Research ArticleArchaeal Community Changes Associated with Cultivation ofAmazon Forest Soil with Oil Palm

Daiva Domenech Tupinambaacute1 Mauriacutecio Egiacutedio Cantatildeo2

Ohana Yonara Assis Costa1 Jessica Carvalho Bergmann1 Ricardo Henrique Kruger3

Cynthia Maria Kyaw3 Cristine Chaves Barreto1 and Betania Ferraz Quirino14

1Genomic Sciences and Biotechnology Program Universidade Catolica de Brasılia 70790-160 Brasılia DF Brazil2Embrapa Swine and Poultry Research Center Embrapa 89700-991 Concordia SC Brazil3Department of Cellular Biology Universidade de Brasılia 70910-900 Brasılia DF Brazil4Embrapa Agroenergy 70770-901 Brasılia DF Brazil

Correspondence should be addressed to Betania Ferraz Quirino bfquwalumnicom

Received 17 September 2015 Revised 14 December 2015 Accepted 10 January 2016

Academic Editor William B Whitman

Copyright copy 2016 Daiva Domenech Tupinamba et al This is an open access article distributed under the Creative CommonsAttribution License which permits unrestricted use distribution and reproduction in any medium provided the original work isproperly cited

This study compared soil archaeal communities of the Amazon forest with that of an adjacent area under oil palm cultivation by16S ribosomal RNA gene pyrosequencing Species richness and diversity were greater in native forest soil than in the oil palm-cultivated area and 130 OTUs (137) were shared between these areas Among the classified sequences Thaumarchaeota werepredominant in the native forest whereas Euryarchaeota were predominant in the oil palm-cultivated area Archaeal speciesdiversity was 17 times higher in the native forest soil according to the Simpson diversity index and the Chao1 index showedthat richness was five times higher in the native forest soil A phylogenetic tree of unclassified Thaumarchaeota sequences showedthat most of the OTUs belong to Miscellaneous Crenarchaeotic Group Several archaeal genera involved in nutrient cycling(eg methanogens and ammonia oxidizers) were identified in both areas but significant differences were found in the relativeabundances ofCandidatusNitrososphaera and unclassified Soil Crenarchaeotic Group (prevalent in the native forest) andCandidatusNitrosotalea and unclassified Terrestrial Group (prevalent in the oil palm-cultivated area) More studies are needed to culture someof these Archaea in the laboratory so that their metabolism and physiology can be studied

1 Introduction

TheAmazon forest area represents 50of theworldrsquos remain-ing rainforests [1] This biome spreads across Brazil BoliviaPeru Ecuador Colombia Venezuela Republic of Guyanaand French Guyana The Amazon is the largest Brazilianbiome and occupies an area of 4196943 km2 correspondingto 67 of the Brazilian territory [2] The Amazon forestprovides important ecosystem services such as hydrologicalcycles and carbon sequestration and storage More impor-tantly it hosts over 20 of all plant and animal species in theworld [3] indicating its high species diversity

Amazonrsquos biodiversity encompasses not only macrofloraandmacrofauna but also its microorganisms which are often

neglectedMineral materials and organic compounds presentin soil create distinct microhabitats populated by differentmicrobial communities Microorganisms are crucial to thebalance of ecosystems with soil microbial communities play-ing important roles in soil fertility plant health and essentialbiogeochemical processes such as nitrification ammoniaoxidation and methanogenesis [4ndash7]

As of 2010 oil palm was cultivated on 112500 hectares ofland in Brazil [8] primarily in the Amazon region Oil palm(Elaeis guineensis Jacq) is a highly productive perennial cropyielding 2000ndash8000 kg oilha [8]The oil which is extractedfrom the fruit has diverse applications in the food and cos-metic industries and can also be used for biodiesel production[9]

Hindawi Publishing CorporationArchaeaVolume 2016 Article ID 3762159 14 pageshttpdxdoiorg10115520163762159

2 Archaea

Although the Amazon is one of the most species-richbiomes on Earth little is known about its archaeal diversityTo date only one published microbial ecology study hasfocused on Amazon soil archaeal diversity using 16S rRNAgene sequencing [10] and only one soil type (ie Amazoniandark earth also called Terra Preta) was studied

The archaeal taxonomy is a matter of constant changesince its proposal in 1977 Initially two phyla were recognizedCrenarchaeota and Euryarchaeota but in the subsequentyears many new phyla were proposed One example is theThaumarchaeota phylum [11] composed predominantly ofmesophilicmembers it encompasses the ammonia-oxidizingarchaea The phylum Korarchaeota was proposed in 1996after the identification of DNA sequences from the ObsidianPool in Yellowstone National Park [12] composed of onecandidate thermophilic species whose genome was com-pletely sequenced [13] Recent works have described putativenew phyla such as Nanoarchaeota Aigarchaeota Aenig-marchaeota Parvarchaeota and Lokiarchaeota but thesephyla are not widely accepted yet due to the low numberof specimens or DNA sequences available In addition fur-ther analyses have positioned sequences belonging to thesephyla in already described phyla such as Euryarchaeota orThaumarchaeota [14ndash17] There are some archaeal groupssuch as MCG (Miscellaneous Crenarchaeotic Group) whichare poorly characterized in terms of phylogenetic affiliationrecent data revealed that this group is probably more closelyrelated to the phylum Thaumarchaeota than to the Crenar-chaeota

Mesophilic archaea seem to play important roles in thecycling of important nutrients such as nitrogen and carbonThe importance of ammonia-oxidizing archaea (AOAs) hasbeen well documented in different ecosystems such as soilsmarine and freshwater environments where they sometimescan be found in higher abundance than the ammonia-oxidizing bacteria (AOBs) [18ndash20] On the other hand theirreal role in nitrification is not yet well understood due to thescarce number of cultured AOAs and the few physiologicalstudies available for this group The methanogens are amongthe nonextremophilic archaea which are widely distributedin anaerobic environments such as flooded soils or marinesoils and vents Methanogens are also found in the gut oftermites in rumenof cows or even in themouth and intestineof humans These organisms play important roles in thecarbon cycle transforming small compounds such as acetateand propionate into methane and removing the hydrogenwhich is potentially hazardous to some bacterial cells On theother hand methane production is one of the major gasesinvolved in the planetrsquos greenhouse effect (reviewed by [21])

There are several studies associating land use withchanges in the structure and abundance of soil microbialcommunities such as the influence of the land use over thediversity of AOA and AOB in grassland soils [22] and theimpacts of edaphic factors on those archaea in tropical soils[23] Therefore the conversion of native forest into palm treeculture can be another example of a potential impact of the oilpalm cultivation in the archaeal communities of Amazoniansoils

This work aimed to improve our understanding of howthe soil archaeal community is impacted by oil palm cultiva-tion To this endmicrobialDNAwas extracted from soil sam-ples from native forest and an adjacent area under oil palmcultivation The archaeal 16S ribosomal RNA (rRNA) genewas amplified and sequenced using high-throughput meth-ods for comparative analysis Here we show for the first timethat soil archaeal diversity is reduced in soil under oil palmcultivation compared to native forest soil

2 Materials and Methods

21 Site Description Sampling and Processing Soil sampleswere collected in the State of Para Brazil in an oil palm-cultivated area and an adjacent area of Amazon native forestnear the city of Moju (Figure 1) The tropical forest in thisregion is dense with trees that are 25ndash35m tall [24] The cli-mate is equatorial hot and humid (Ami type according to theKoppen climate classification) Annual temperatures rangefrom 25∘C to 27∘C and rainfall is 2000ndash3000mm per yearbeing irregularly distributed [25] The soil is predominantlyldquoLatossolo Amarelordquo (a type of Oxisol) [26]

The oil palm cultivation in the sampled farm is not ascontrolled as other crops (Figure 1(b)) There is no irrigationregime natural precipitation of the rainforest is the only waythese plants are irrigated In the Amazon the soil is verymoist due to the high precipitation levels during the yearIn the studied area the annual period of flooding is fromFebruary to April Furthermore the soil around the palmtrees is not fertilized in a homogeneous fashion since onlyone side of the plants is directly fertilized

In October 2010 after plant litter was removed asoil borer was used to obtain four 10 cm deep soilsamples from three points in the oil palm-cultivatedarea (S02∘00101584028910158401015840W048∘37101584057410158401015840 S02∘00101584029210158401015840W048∘37101584056610158401015840 and S02∘00101584031310158401015840W048∘37101584054310158401015840) (Figure 1(b))and four samples from the native Amazon forest area(S02∘00101584027210158401015840W048∘35101584053010158401015840) (Figure 1(a)) The samplescollected in each area were mixed ground and sieved toremove larger particles yielding one composite sample foreach area with approximately 1 kg each The samples werestored in plastic bags on dry ice during transportationA subsample was sent to physicochemical analysis atSoloQuımica Analises de Solo Ltda (Brasılia DF Brazil)The rest of the samples were then stored at minus80∘C until DNAextraction Initially the physicochemical characteristics ofthe soils samples were evaluated individually and a highvariation among replicas was observed This result was dueto the heterogeneous fertilization of the palm trees in thecultivation fields in Amazon therefore composite sampleswere necessary to describe the microbiota in oil palm soil

22 DNA Extraction PCR and Pyrosequencing AnalysisTotal DNA was extracted according to the protocol of Smallaet al [27] using 2 g soil per sample TominimizeDNAextrac-tion bias this procedure was performed in quadruplicatePCR reactions were performed using the following primersspecific for Archaea 340F (51015840-CCC TAY GGG GYG CAS

Archaea 3

(a) (b)

Figure 1 (a) Native forest and (b) oil palm-cultivated sites

CAG-31015840) and 1000R (51015840-GGC CAT GCA CYW CYT CTC-31015840) [28] Archaea 16S rRNA genes were amplified yielding660 bp amplicons Adapters used as priming sites for bothamplification and sequencing (454 Life Sciences BranfordCT USA) were ligated to the 51015840 end of the primer sequencesEach 20120583L PCR reaction contained 10ndash30 ng total DNA 1xreaction buffer 4120583MdNTP 10 120583Mof each primer 200120583gmLbovine serum albumin 05U KAPA2G Robust HotStartpolymerase andMilli-Qwater Amplification was performedin anApplied BiosystemsGeneAmpPCRSystem9700Ther-mal Cycler (Applied Biosystems Foster City CA USA) usingthe following program 2min at 98∘C followed by 30 cyclesof 30 seconds at 95∘C 30 seconds at 57∘C and 1 minute and30 seconds at 72∘C and a final step consisting of 7 minutes at72∘C To minimize PCR bias at least four PCR amplificationreactions per sample were pooled before purification usingthe GeneJET PCR Purification Kit (Thermo Fisher ScientificWaltham MA USA) After quantification using a Qubitfluorometer (Invitrogen Carlsbad CA USA) NanoDrop1000 Spectrophotometer (Thermo Fisher Scientific) and 2agarose gel with ethidium bromide DNA samples were sentfor pyrosequencing (GSFLXTitaniumPlatformatMacrogenSouth Korea) A total of 592145 reads greater than 500 bpwere obtained in three-quarters of a plate 49314 reads fromthe oil palm-cultivated soil and 542831 reads from the nativeforest soil

23 Data Analysis All 16S rRNA gene pyrosequencing readswere analyzed using the original standard flowgram format(SFF) output file from the sequencer in MOTHUR version1333 [29] for the removal of short sequences sequences witherrors low-quality sequences chimeras and possible con-taminants After this removal a total of 39111 sequences foroil palm-cultivated area and 436532 sequences for native for-est were obtained Further analysis was performed after nor-malizing the number of sequences per sample to the lowest

number of high-quality sequences (ie 39111 sequences in theoil palm-cultivated area sample)

All 16S rRNA gene reads were analyzed using MOTHURfor taxonomic classification following the 454 standardoperating procedure available online (httpwwwmothurorgwiki454 SOP) [30] The PyroNoise algorithm [31]was used to denoise the flowgram file (removal of barcodesand quality filtering) The SILVA database (release 119) wasused for sequence alignment and clustering based on 97sequence similarity (003 genetic distance) using the nearestneighbor method [32] The UCHIME algorithm [33] wasused for detection and removal of chimeric reads thisfunction was implemented in MOTHUR and performedwithout a reference database The one-gap method inMOTHURwas used to calculate the pairwise distancematrixArchaeal taxonomic classifications of each representativeOTU were performed in MOTHUR using the SILVA data-base (release 119) clustered at 97 sequence similarity [32]This classificationwas used to estimate the relative abundanceof reads per genus Samples were randomly chosen forsubsampling and normalized to the lowest number ofsequence reads obtained Chao1 [34] abundance-basedcoverage estimator (ACE) [35] Shannon diversity index [36]and Goodrsquos coverage [37] were then used to estimatealpha diversity MOTHUR was also used to perform allstatistical analyses A phylogenetic tree was created usingall Thaumarchaeota OTUs from each sample that wereunclassified at the class level A new tree was created withsequences randomly chosen by MOTHUR that were deemedrepresentative of the entire set Data were analyzed using theneighbor-joining method with the Jukes-Cantor correctionand 1000 bootstrap replicates in MEGA6 [38] In additionOTUs with 3 dissimilarity were selected by MOTHUR toavoid redundancies in the phylogenetic tree The chosensequences were aligned with 16S rRNA gene sequencesfrom GenBank [39] representing the main Archaeagroups Methanosarcina baltica (AB973356) Methanoregula

4 Archaea

Table 1 Number of operational taxonomic units (OTUs) Goodrsquos coverage estimator OTU richness indices (Chao1 ACE) and InverseSimpson and Shannon diversity indices for archaeal communities in native forest soil and soil under oil palm cultivation

Sample OTUslowast Goodrsquos coverage () Chao1 ACE Inverse Simpson ShannonNative forest 821 9871 2124 4645 2645 422Oil palm-cultivated area 253 9972 424 674 1547 331lowastConsidering 97 sequence similarity in MOTHUR

formicica (NR112877) Methanocella paludicola (NR074192)Halococcus hamelinensis (LN651155) and Pyrococcus furiosus(U20163) from the phylum Euryarchaeota Pyrobaculumaerophilum (NR102764) from the phylum CrenarchaeotaCandidatus Nitrososphaera sp (FR773157) and CandidatusNitrososphaera gargensis (GU797786) fromThaumarchaeotaGroup I1b Nitrosopumilus maritimus (JQ346765) andCenarchaeum symbiosum (U51469) from ThaumarchaeotaGroup I1a uncultured acidic red soil AOA (FJ174727)uncultured trembling aspen archaeon (EF021427) and borealforest archaeon (X96688) from Thaumarchaeota GroupI1c and several uncultured organisms classified accordingto the SILVA database as Thaumarchaeota MiscellaneousCrenarchaeotic Group (FR745121 AM910782 JX984848KC510333 FJ485299 KC831395 FJ920714 FJ485307HM051127 HM051130 and HM051125) A 16S rRNA genesequence from Acidobacteria KBS 96 (FJ870384) was usedas an outgroup A second phylogenetic tree was createdusing only Euryarchaeota OTUs from each sample that wereunclassified at the class level with the same parameters andreference sequences

A Venn diagram was generated by grouping OTUs clus-tered at 97 similarity to show the number of OTUs uniqueto each area and those shared between samples

Differences in taxonomic distribution of archaea betweennative forest and the oil palm-cultivated area were evaluatedby Fisherrsquos exact test (119902 le 005) using Statistical Analysisof Metagenomic Profiles (STAMP) software [40] The confi-dence interval was estimated using the asymptotic method[41] and correction of the 119902 value was calculated usingStoreyrsquos false discovery rate (FDR) approach [42] A filterwas applied to show only the differences between proportionsabove 05

3 Results

Goodrsquos coverage estimate of the completeness of sampling(Table 1) was higher for the oil palm-cultivated area than forthe native forest

The number of archaeal OTUs was higher in the nativeforest soil (821 sequences) than in soil under oil palmcultivation (253 sequences) (Figure 2) with 130OTUs (138)shared between the two areas Archaeal species richness asassessed by abundance-based estimators Chao1 and ACEwas higher in the native forest than in the oil palm-cultivatedarea Similarly the Inverse Simpson and Shannon indicesindicated that archaeal species diversity was higher in nativeforest soil compared to soil under oil palm cultivation(Table 1)

Table 2 Physicochemical properties of native forest soil and soilunder oil palm cultivation

Native forest Oil palm-cultivated areapH(in water) 550 483

P (mgsdotdmminus3) 140 7070K (mgsdotdmminus3) 003 011Ca+2 (cmolsdotdmminus3) 140 033Mg+2 (cmolsdotdmminus3) 010 017Al+3 (cmolsdotdmminus3) 040 073H + Al (cmolsdotdmminus3) 620 727Na+ (cmolsdotdmminus3) 001 002Organic matter (gkg) 3100 1163Total C (gkg) 1800 677Total N (mgkg) NA 115NO3

minus1 (mgkg) NA 079NO2

minus1 (mgkg) NA 014NH4

+ (mgkg) NA 021Data not available

Soil physicochemical parameters differed between theareas (Table 2) with higher levels of phosphorous potassiumand total carbon in the oil palm-cultivated soil and higherlevels of calcium and organic matter in the native forest soil

In both areas only two archaeal phyla (Thaumarchaeotaand Euryarchaeota) were detected (Figure 3) Thaumar-chaeota were predominant in the native forest soil (58) andEuryarchaeota were predominant in the oil palm-cultivatedsoil (54)The percentage of unclassified archaea was similarin both soils (native forest 5 oil palm-cultivated area 6)

At the class level a total of 10 groups were detected inboth samples Halobacteria Methanobacteria Methanomi-crobia and Thermoplasmata in the phylum Euryarchaeotaand Marine Group I OPPD003 Soil Crenarchaeotic GroupSouth African Gold Mine Gp 1 Terrestrial Group andMiscellaneous Crenarchaeotic Group in the phylum Thau-marchaeota (Figures 4(a) and 4(b)) The most abundantclass present in native forest soil was the Soil Crenar-chaeotic Group representing 26 of all native forest OTUs(Figure 4(a)) and 45 of theThaumarchaeota sequences (notshown) followed by Methanomicrobia representing 15 ofall native forest OTUs (Figure 4(b)) and 42 of the Eur-yarchaeota sequences (not shown) The most abundant classpresent in the oil palm-cultivated area was Thermoplasmatarepresenting 21 of all cultivated area OTUs (Figure 4(b))and 39 of the Euryarchaeota sequences (not shown) fol-lowed by Methanomicrobia representing 16 of all culti-vated area OTUs (Figure 4(b)) and 30 of all Euryarchaeota

Archaea 5

Venn diagram at distance 003

123Cultivated area

691Native forest

130

Figure 2 Venn diagram depicting the number of shared and unique operational taxonomic units (OTUs) in native forest and oil palm-cultivated area

Thaumarchaeota

Euryarchaeota

Unclassified archaea

Cultivated areaNative forest

10 20 30 40 50 60 700()

Figure 3 Comparison of archaeal phyla present in soil samples from native Amazon forest (orange) and an area under oil palm cultivation(blue) The percentages of unclassified sequences are shown

sequences (not shown) The predominant class from thephylum Thaumarchaeota in the oil palm-cultivated areawas Terrestrial Group representing 14 of the total OTUs(Figure 4(a)) and 35 of all Thaumarchaeota sequences (notshown)

Because it is a newer phylum Thaumarchaeota have fewcultivated representatives compared with Euryarchaeota andfewer sequences in databases such as SILVA To determinehow the Thaumarchaeota sequences that could not be clas-sified at the genus level were related to each other a phylo-genetic tree was constructed with representative sequences(Figure 5(a)) One native forest OTU clustered with GroupI1b (NF6) and one native forest OTU (NF17) and two cul-tivated area OTUs (CA50 CA52) clustered with Group I1cThe remaining OTUs formed different clusters all associatedwith the Miscellaneous Crenarchaeotic Group Most unclas-sified Thaumarchaeota sequences from the native forest andoil palm-cultivated area grouped together although a fewunclassified OTUs from the cultivated area (eg CA51 toCA15 and CA24 to CA33) and native forest (eg NF21 toNF49 and NF5 to NF35) formed separate clusters

The unclassified sequences at genus level in the Euryar-chaeota phylum were all related to the orders Methano-cellales Methanosarcinales Methanomicrobiales Thermo-plasmatales Methanobacteriales and Halobacteriales(Figure 5(b)) The classified sequences at genus level wereMethanocella Methanosarcina Methanoculleus Methano-massiliicoccus Methanomethylophilus MethanobrevibacterMethanobacterium and Candidatus lainarchaeum(Figure 5(b))

The genus-level analysis showed that unclassified SoilCrenarchaeotic Group was predominant in the native forestaccounting for 21 of all native forest OTUs (Figure 6(a))In the oil palm-cultivated area the predominant genus wasUnclassified Terrestrial Group accounting for 14 of allcultivated area OTUs (Figure 6(b))

A total of 31 groups of OTUs were identified (onlymajor groups are shown) most of which are unclassi-fied The classified groups in the phylum Euryarchaeotaare Candidatus lainarchaeum Methanobacterium Methano-brevibacter Methanocella Rice Cluster I Methanoculleus

6 Archaea

Marine Group I

Miscellaneous Crenarchaeotic Group

Soil Crenarchaeotic Group

South African Gold Mine Gp 1

Terrestrial Group

Unclassified Thaumarchaeota

OPPD003

Cultivated areaNative forest

5 10 15 20 25 300()

(a)

Methanobacteria

Methanomicrobia

Halobacteria

Thermoplasmata

Unclassified Euryarchaeota

Cultivated areaNative forest

5 10 15 20 250()

(b)

Figure 4 Comparison of archaeal classes present in soil samples from native Amazon forest (orange) and an area under oil palm cultivation(blue) Classes within the phyla Thaumarchaeota (a) and Euryarchaeota (b) are shown as well as the percentage of unclassified sequences

Methanoregula Methanimicrococcus Methanosarcina Can-didatus Methanomethylophilus and Methanomassiliicoccus(Figure 6(b)) The classified groups in the phylum Thau-marchaeota are Candidatus Nitrososphaera and CandidatusNitrosotalea (Figure 6(a))

The analysis of the genera that differ significantly inrelative abundance between native forest soil and the oilpalm-cultivated area revealed that the greatest difference wasseen for Candidatus Nitrososphaera (Figure 7) This genuswas predominant in the native forest soil accounting for182 of the sequences but represented only 103 of thesequences from the oil palm-cultivated area UnclassifiedSoil Crenarchaeotic Group was also predominant in thenative forest (ie 388 of sequences in the native forest

and 312 of sequences in the oil palm-cultivated area)whereas unclassified South African Gold Mine Gp 1 waspredominant in the oil palm-cultivated area (ie 73 ofsequences in the native forest and 135 of sequences inthe oil palm-cultivated area) Candidatus Nitrosotalea repre-sented 44 of the native forest sequences and 100 of theoil palm-cultivated area sequences The relative abundanceof unclassified Terrestrial Miscellaneous Group unclassifiedThaumarchaeotaunclassifiedThermoplasmatalesunclassifiedMiscellaneous Crenarchaeotic Group Methanosarcina andRice Cluster I also differed significantly between native forestsoil and soils under oil palm cultivation but these differenceswere smaller

Archaea 7

FJ4853071| Uncultured archaeon clone

FJ4852991| Uncultured archaeon clone

FJ9027141| Uncultured archaeon clone

KC8313951| Uncultured archaeon clone

KC5103331| Uncultured archaeon clone

JX9848481| Uncultured archaeon clone

AM9107821| Uncultured archaeon clone

Uncultured archaeon acidic red soil-FJ174727

Ca Nitrososphaera gargensis-GU797786

Ca Nitrososphaeramdash(48 6)

Ca Nitrosotaleamdash(49 12)

Candidatus lainarchaeum (0 2)

Methanobacterium (2 3)

Methanomassillicoccus (6 2)

Methanocella (33 8)

Rice cluster I (43 25)

Methanosarcina baltica-AB973356

Methanocella paludicola-NR074192

Pyrobaculum aerophilum

-NR102764

Acidobacteria bacterium KBS 96-FJ870384

Ca Methanom

ethylophilus (12 0)

Ca Nitrososphaera sp-FR773157

Nitrosopumilus maritimus-JQ346765

Halococcus hamelinensis-LN651155

Pyrococcus furiosus-PFU20163

Cenarchaeum symbiosum-CSU51469

Methanoregula formicica-NR112877

FR7451211| Uncultured soil archaeon

Boreal forest soil archaeon-X96688

Trembling aspen archaeon-EF021427

MCG

MCG005

CA4099

99

7174

63

86

9687

9067

5696

6768 54

57

62

58

65

7559

62

63

65

64

52

64

64

1008654

91

99

999293

53798392

88

7274

5164

5858

CA7CA32

CA50NF17

NF6

CA52

T I1c

T I1b

T I1a

CA24CA6CA12

CA4

CA33

NF16

NF2

NF35

NF9

NF5NF3

6NF4

1NF3

2

NF1

2

NF4

NF1

NF3

0NF2

3

NF7CA3

CA39

CA37

CA46

CA44

CA2

CA42

CA27

Unclassified MCGmdash(16 4)

Unclassified Terrestrial Group (94 36)

Unclassified Marine Group I (2 2)

CA15

CA25

CA30

CA43

CA35

CA14

CA29

CA13

CA20

CA11

CA41

CA36

CA51

NF48

CA9NF4

0NF4

2CA21

CA22

NF38

CA45

CA16

CA47

NF31CA31NF28NF47NF27NF45CA18NF26

NF24NF33NF19

NF18NF37

NF34NF43NF15NF44NF13

NF22NF29NF11

CA49

CA1

CA19CA8

CA10

CA23CA38

CA54HM0511251| Uncultured

HM0511301| Uncultured archaeon clone

HM0511271| Uncultured archaeon clone

archaeon clone

NF46NF14

NF49NF25N

F10N

F39N

F20N

F21

NF8

NF3

CA48

CA34

CA5

CA28CA26CA17CA53

(E)

(B)

(C)

(a)

CA13

CA14

NF1

2

NF1

4M

ethan

ocell

a (3

8)Meth

anosarcina (3

8 4)

Methanoculleus (2 0)

Methanomassilicoccus (6 2)

Methanomassilicoccus (6 2)

Methanomassilicoccus (6 2)

Methanomethylophilus (6 2)

Methanomethylophilus (6 2)

Methanobrevibacter (1 0)

Methanobacterium

(2 3)

Methanobacterium

(2 3)

Methanobacterium

(2 3)

Methanobacterium

(2 3)

Methanobacterium

(2 3)

Methanosarci

na (38 4)

Methanosarcina (38 4)

Methanosarcina (38 4)

Methanosarcina (38 4)

NF16

CA4

CA15CA20

CA19

NF3NF1

Meth

anoc

ella (

33 8

)

Meth

anoc

ella p

aludi

cola-

NR 0741

92 1

Meth

anosa

rcina

baltic

a AB97

3356

1

Methanoregula formicica NR 112877 1

Halococcus hamelinensis LN651155 1

CA6N

F6NF8

NF19 NF5NF7NF20

Pyrobaculum aerophilum NR 102764 2

Ca Nitrososphaera gargensis G

U797786 1

Ca Nitrososphaera gargensis GU797786 1

Nitrosopumilus maritimus JQ346765 1Cenarchaeum symbiosum U51469 1Uncultured archaeon acidic red soil FJ174727 1

Boreal forest soil archaeon X96688 1

Trembling aspen archaeon EF021427 1

HM051125 1 Uncultured archaeon clone

HM051127 1 Uncultured archaeon clone

HM051130 1 Uncultured archaeon clone

FJ485307 1 Uncultured archaeon clone

FJ902714 1 Uncultured archaeon clone

KC831395 1 archaeon clone

FJ485299 1 archaeon clone

KC510333 1 archaeon clone

FR745121 1 archaeon clone

JX984848 1 archaeon clone

AM910782 1 archaeon clone

Acidobacteria bacterium KBS 96 FJ870384 1

Ca Nitrososphaera sp FR773157 1

CA7

Pyrococcus furiosus U20163 1

005

MCG

Halobacteriales

Cand

idat

us la

inar

chae

um (0

2)

Cand

idat

us la

inar

chae

um (0

2)

Thermoplasmatales

Methanomicrobiales

Methanosarcinalles

Methanocellales

T I1c

T I1a

T I1b

(C)

998785

58

100

100

100100

100

71 84 71

53

54

65

86

94

98

59100

100

78

10051

5477

63 73

99

100

8497100

78

100

100

76956970

100 10

010

0

637551

100

9999

9958

94

Methanobacteriales

(b)

Figure 5 Phylogenetic trees of (a) unclassified Thaumarchaeota 16S rRNA gene sequences from native forest soil and soil under oil palmcultivation and (b) 16S rRNA gene sequences classified as Euryarchaeota from native forest soil and soil under oil palm cultivation Theneighbor-joining method and Jukes-Cantor corrections were used with 1000 bootstrap replicates Bootstrap values above 50 are shownin the trees A representative number of operational taxonomic units (OTUs) were selected at 3 dissimilarity by MOTHUR to avoidredundancies in the phylogenetic trees Randomly chosen OTUs from native forest soil and from soil under oil palm cultivation were usedin the construction of the trees along with archaeal reference sequences (from GenBank) from the following groups Acidobacteria (B)Crenarchaeota (C) Euryarchaeota (E) Miscellaneous Crenarchaeotic Group (MCG) Thaumarchaeota Group I1a (T I1a) ThaumarchaeotaGroup I1b (T I1b) andThaumarchaeota Group I1c (T I1c)The AOAs are highlighted with the symbolX Sampling site is indicated by ldquoNFrdquo(native forest) or ldquoCArdquo (oil palm-cultivated area) followed by the number of the OTUs (NAxx or CAxx) OTU sequences classified to at leastthe class level are shown in bold followed by two numbers in parentheses indicating the number of sequences of that OTU detected in nativeforest soil and the number detected in the oil palm-cultivated area The scale bar represents the 5 estimated sequence divergence

8 Archaea

Unclassified Marine Group I

Unclassified Miscellaneous Crenarchaeotic Group

Unclassified Thaumarchaeota

Candidatus Nitrososphaera

Unclassified Soil Crenarchaeotic Group

Candidatus Nitrosotalea

Unclassified South African Gold Mine Gp 1

Unclassified Terrestrial Group

Cultivated areaNative forest

5 10 15 20 250()

(a)

Unclassified Thermoplasmatales

Unclassified Terrestrial Miscellaneous Group

Rice Cluster I

Methanocella

Methanosarcina

Unclassified Euryarchaeota

Unclassified Halobacteriales

Candidatus Methanomethylophilus

Unclassified Methanosarcinaceae

5 10 15 20 250()

Cultivated areaNative forest

(b)

Figure 6 Percentage of archaeal operational taxonomic units within the Thaumarchaeota (a) and Euryarchaeota (b) phyla classified at thegenus level for native forest soil (orange) and soil under oil palm cultivation (blue) The percentage of unclassified sequences at genus level isshown

4 Discussion

The Amazon forest is known for its macrospecies diversityhowever few studies have addressed its microbial diversityAmong the domains of life Archaea are undoubtedly the lesswell known

In this work almost 600000 archaeal 16S rRNA genesequences were obtained from native forest soil and anadjacent area under oil palm cultivation After processing(ie removal of short sequences sequences with errors low-quality sequences chimeras and possible contaminants) andnormalization 39111 sequences per area remained for anal-ysis This number of sequences provided sufficient coverageof archaeal diversity in both areas (Table 1)The primers used

were based on a large number of 16S rRNA sequences fromthe SILVA database [28] however given that little is knownabout this domain it is unlikely that the archaeal diversitywasfully covered [43]

The main physicochemical differences between soils ofthe native forest and the oil palm-cultivated area were thelevels of phosphorous potassium calcium organic matterand total carbon (Table 2) Correlations between phylogenyand physicochemical parameters were evaluated using PCAand NMDS however no clear clusters or correlations wereobserved (results not shown)

In a study of the long-term effects of inorganic fertilizerson microbial communities Zhong and Cai [44] reportedthat phosphorus application indirectly altered microbial

Archaea 9

Candidatus NitrososphaeraUnclassified Soil Crenarchaeotic Group (SCG)

Unclassified South African Gold Mine Gp 1 (SAGMCG-1)Candidatus Nitrosotalea

Unclassified Terrestreial Miscellaneous Gp (TMEG)Unclassified Thaumarchaeota

Unclassified ThermoplasmatalesUnclassified Miscellaneous Crenarchaeotic Group

MethanosarcinaRice Cluster I

95 confidence intervals

q-v

alue

(cor

rect

ed)

Cultivated areaNative forest

531e minus 10

420 6 8minus4minus6minus8 minus2minus10

Difference between proportions ()38800

Proportion ()

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

Figure 7 Archaeal genera that differ significantly (119902 lt 005) in relative abundance between native forest soil (orange) and soil under oil palmcultivation (blue) For the purpose of statistical analysis performed using STAMP software the sum of the percentages of archaea classifiedat the genus level in both soils is 100 Differences between proportions of each genus are shown with 95 confidence intervals Unclassifiedsequences were used to calculate frequency profiles but are not displayed

parameters in soils and increased crop yields by elevating thelevels of organic matter Wessen and collaborators used real-time PCR quantification to show the impacts of fertilizer useon several bacterial phyla and the archaeal phylum Crenar-chaeota (Thaumarchaeota Group I1c after reanalysis usingthe SILVAdatabase 119) [45]They reported that bacterial andarchaeal classes respond differently to fertilization primarilybecause of changes in certain soil parameters such as pHtotal nitrogen and the carbon nitrogen ratio In our studylevels of total carbon and organic matter were higher in thenative forest than in the oil palm-cultivated area which mayhave contributed to higher archaeal diversity Although thereare no studies on the effect of total soil carbon on archaealcommunities there are studies on the bacterial communitiesin Amazonian soils The work of Edgar et al [33] describeddistinct soil bacterial communities under different vegetationtypes The authors found that bacterial diversity correlatedwith total organic carbon and total nitrogen content [46]Furthermore in a study of bacterial diversity and microbialbiomass in forest pasture and fallow soils in the Amazonbasin microbial biomass was highest in pasture soils whichalso had 30ndash47 higher carbon levels than the other sitesover the course of a year [47]

Several studies have also reported alterations in thecomposition of archaeal [48] bacterial [47] and fungal [49]communities associated with land use changes A studyof archaeal communities in Amazonian anthrosols undervarious land uses (agriculture pasture and secondary forest)and cultivation practices (eg eggplant banana citrus andmanioc) [10] reported that agriculture negatively affectedarchaeal community diversity Results of ribosomal inter-genic spacer analysis showed that soil type is also responsiblefor changes in archaeal communities in soils under differenttypes of land use (native grassland native forest Eucalyptus

and Acacia plantations and soybean and watermelon fields)[50]

The Venn diagram showed that the native forest soil has agreater number of unique sequences (Figure 2) This was notsurprising due to the greater archaeal richness and diversityobserved in the native forest soil (Table 2) The number ofunique OTUs found in the oil palm-cultivated area is almostthe same as that of shared OTUs with the native forest areaAlthough oil palm cultivation decreased archaeal diversity inthe soil there is still a core group of archaea OTUs sharedwith the native forest suggesting that these archaea havecertain characteristics that allow them to persist after thechange in land use It cannot be ruled out that these archaeaare metabolically inactive The smaller number of uniquesequences in the oil palm-cultivated area may be related to alower complexity of the vegetation (ie oil palmmonocultureversus great plant diversity in the forest) since plant diversitycould affect these communities [51]

In our study all OTUs identified belong to the domainArchaea and were classified into two phyla Euryarchaeotaand Thaumarchaeota It is important to mention that thedatabases were only recently updated to recognize the phy-lumThaumarchaeota therefore previous studies on Archaeacould not recognize this phylum as proposed by Brochier-Armanet et al [11] The creation of this novel phylum wassupported by a study comparing the genomes of two marineammonia-oxidizing archaea (AOAs) that identified a core setof informational genes specific to this group [52] Thereforeto allow comparisons with other studies we reanalyzed theirdata whenever possible updating the taxonomic classifica-tion to include the phylumThaumarchaeota

Thework ofChao and Shen [35] focused on the impacts ofland use (ie agricultural systems of indigenous people andcattle pasture) on archaeal communities by PCR (amoA gene)

10 Archaea

and denaturing gradient gel electrophoresis analysis of differ-ent Western Amazon soils The databases at the time did notconsiderThaumarchaeota in their classification however ourreanalysis of their dataset using an updated database showedthat Euryarchaeota and Thaumarchaeota were present intheir samples along with Crenarchaeota (Thermoprotei) Inaddition changes were observed in the structure of soilarchaeal communities when comparing the cultivated areaswith forest areas that had been converted to pasture

In our study differences in the percentages of Thau-marchaeota and Euryarchaeota found in the soil sampleswere not striking Thaumarchaeota were predominant in thenative forest (58) whereas Euryarchaeota were predom-inant in the oil palm-cultivated area with 54 (Figure 3)This result is consistent with other comparative studies onarchaeal communities in different types of Brazilian soils (ieAmazonian anthrosols Caatinga and Cerrado) [10 53 54]and a study of different tropical forest soil types that reportedThaumarchaeota as the predominant phylum [44]

The phylumThaumarchaeota comprises all known AOAsand is directly involved in nitrogen metabolism Ammonia isused as an energy source by Group I1a found predominantlyin aquatic systems [52] and Group I1b found in soils [55]Thaumarchaeota Group I1c is primarily found in acidic soilsbut the ecological roles of these archaea are still unknown [5657]

The literature presents conflicting data about the pro-portion of AOAsAOBs soils In some studies AOAs arepredominant [58] while in others AOBs are more abundant[59] However several studies found a predominance of thearchaeal amoA gene over the bacterial amoA gene in soils[60] AOAs are frequently detected in soils their abundancerange from 05 to 10 of the prokaryotic community andtheir abundance is strongly influenced by physicochemicalconditions as well as the organic matter content [61 62] Inacidic soils AOAs appear to have a more important role inammonia oxidation than AOBs [57]

The Soil Crenarchaeotic Group a class within Thaumar-chaeota according to the SILVA database classification wasdetected in both soil samples but a higher representationwas observed in native forest soil This class was discoveredin boreal forest soil [63] Candidatus Nitrososphaera whichbelongs to the Soil Crenarchaeotic Group class is a memberof Thaumarchaeota Group I1b it is an AOA that usesammonia or urea as an energy source [64 65]

On the other hand members of the phylum Eur-yarchaeota are directly involved in carbon metabolismwith several classes of methanogens and methane-oxidizingorganisms [66ndash68] Navarrete et al [48] studied the impactof converting Amazon native forest to pasture and reportedchanges not only to the composition and abundance of soilmicrobial communities but also to a decrease in the diversityof gene function

Within the phylum Euryarchaeota Methanomicrobiaand Thermoplasmata were the main classes detected in oursamples along with a few representatives of Methanobac-teria and Halobacteria Members of Methanobacteria andMethanomicrobia are methanogens [69] Methanomicrobiawere present in both native forest soil and oil palm-cultivated

soil with larger representation in the oil palm-cultivatedarea Within Methanomicrobia the genera Rice Cluster IMethanosarcina andMethanocellawere present in both sam-ples (Figure 6(b)) These genera are closely related and foundin rice fields that produce methane [70] The order Metha-nomicrobiales was detected only in the native forest soilwhich can probably be explained by the relatively high levelsof carbon and organic matter This archaeal order is respon-sible for the final step of methane production in the carboncycle by anaerobic degradation of organic matter [66 71]However the contribution of the Methanomicrobia to thecarbon cycle in the oil palm-cultivated soils could not beassessed since there are no data on the methane productionin the studied areas

Among the Thaumarchaeota sequences used to generatethe phylogenetic tree that were unclassified at the genuslevel several OTUs formed different clusters associated withtheMiscellaneous Crenarchaeotic Group (Figure 5(a))Thesesequences were from both soil samples indicating that theunclassified Thaumarchaeota in the native forest and the oilpalm-cultivated area are phylogenetically related

Most of the sequences remained unclassified at thegenus level for both Thaumarchaeota and Euryarchaeota(Figures 6(a) and 6(b)) This result likely reflects the fact thatmost taxonomic databases do not have many representativesof Archaea especially belonging to the phylum Thaumar-chaeota thus few classifications of new sequences reach thegenus level Several genera were identified as Candidatusaccording to the International Code of Nomenclature ofBacteria indicating that characterization of the culturedprokaryote is incomplete [72]

The proportions of only four groups classified at thegenus level differed significantly between native forest and oilpalm-cultivated soils (Methanosarcina and Rice Cluster I inthe phylum Euryarchaeota and Candidatus NitrososphaeraandCandidatusNitrosotalea in the phylumThaumarchaeota)(Figure 7) With respect to the methanogens sequencesrelated to Methanosarcina were slightly more abundant insoil under oil palm cultivation (Figure 7) whereas sequencesbelonging to the Rice Cluster I were more abundant in nativeforest soil

In the Thaumarchaeota phylum Candidatus Nitrosos-phaera an AOA member of group I1b [65] was detectedin both samples but had a much higher representation innative forest (native forest 182 oil palm-cultivated area103) (Figure 7) In a comparative study of the archaealcommunities in Amazonian dark earth (ldquoTerra Pretardquo)and adjacent soils Candidatus Nitrososphaera was themain archaeal group found in both soils [10] In contrastCandidatus Nitrosotalea another AOA was more abundantin the oil palm-cultivated area (100) than in native forestsoil (44) in our study (Figure 7) Lehtovirta-Morley et al[73] cultivated the obligate acidophilic ammonia oxidizerCandidatus Nitrosotalea devanaterra from a nitrifying acidicsoil Later the same group characterized two strains ofacidophilic AOA belonging to this genus from a Chinesepaddy field and a Scottish agricultural soil [74] In our studythe greater representation of Candidatus Nitrososphaeraand Candidatus Nitrosotalea in native forest and oil

Archaea 11

palm-cultivated soils respectively can be explained by thelow soil pH (native forest pH 550 oil palm-cultivated areapH 483) since in basic soils the activity and growth ofthese archaea are diminished by the exponential decrease innitrite [6 75] In addition soil under oil palm cultivationis fertilized with nitrogen which may explain the higherlevel of Candidatus Nitrosotalea in this area compared withthat of native forest soil Finally although the cultivatedmembers of the genus Ca Nitrososphaera grow betteron neutral pH they have been reported to grow on pHvalues ranging from 60 to 85 In addition a recent workpublished by Wang et al (2014) [76] described active AOAsfrom an acidic soil (pH 492) which are closely related toNitrososphaera known as Nitrososphaera-like organismsby molecular phylogenetic methods Most of our sequenceswere classified as Nitrosotalea-like which are sequencesbelonging to the group I1a This group is typically foundin aquatic environments such as lake sediments or marineenvironments A similar ubiquitous distribution in soils isfound for the group I1b which are not restricted to a specificsoil pH condition

5 Conclusions

Although archaea are known to be ubiquitous there is stillmuch to be learned about their diversity biological rolesand metabolic capabilities This work is the first to comparearchaeal communities associated with the Amazon forest soilwith those of soil under oil palm cultivation

Archaeal diversity was found to be lower in oil palm-cultivated soil than in the native forest soil Only about 30of the OTUs were found in the oil palm soil in comparisonto the native forest However two main phyla were detectedin both samples Thaumarchaeota and Euryarchaeota with apredominance ofThaumarchaeota in native forest soil Theseresults support previous studies in other Brazilian biomenative soils (eg Caatinga Cerrado) as well as soil underdifferent types of land use (eg pasture and agricultural) Itis also important to mention that there are also groups ofuncertain classification associated with the Crenarchaeotaphylum

Although knowledge about Archaea has increased dra-matically in the last few decades little is known aboutthe biology of many archaeal groups and the taxonomyof Archaea is still a work in progress Thus it is notsurprising thatmany sequences identified in thiswork remainunclassified sometimes even at higher taxonomic levels Itis not clear whether the relatively small overall differencebetween soil archaeal communities of the Amazon forest andoil palm-cultivated land is a consequence of the current stateof archaeal taxonomy or whether it indicates that archaealdiversity is lower than bacterial diversity [77]

It is also important to consider that with land usechange due to agricultural practices the microstructure ofthe soil is broken down which can reduce the natural habitatfragmentation allowing potential interactions among theresident microbiota that otherwise would not happen [7879]

A phylogenetic tree of unclassified Thaumarchaeota 16SrRNA gene sequences was useful in exploring relationshipsamong these sequences When taxonomic classification ofsequences was possible it revealed the presence of OTUsfrom genera known to be involved in the metabolism ofcarbon and nitrogen (methanogens Methanocella and Meth-anosarcina AOACandidatusNitrososphaera andCandidatusNitrosotalea) hinting at possible roles for these archaealcommunities in Amazon soils

More studies are needed to elucidate these roles Cultur-ing some of these archaea in the laboratory would allow theirmetabolism and physiology to be studied in detail allowingfor a better understanding of the roles of Archaea in the bio-geochemical cycles

Conflict of Interests

The authors declare that there is no conflict of interestsregarding the publication of this paper

Acknowledgments

This work was supported by grants from FINEP CNPqCAPES and FAP-DF The authors thank Dr Boari for helpwith sample collection logistics Dr Betania Ferraz Quirinowas supported by the Cientista Visitante program fromEmbrapa

References

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[2] IBGE ldquoMapa de Biomas do Brasil primeira aproximacaordquo Riode Janeiro Brazil 2004 httpwwwibgegovbrhome

[3] C Azevedo-Ramos ldquoSustainable development and challengingdeforestation in the Brazilian Amazon the good the badand the uglyrdquo in Proceedings of the Symposium Our CommonGround Innovations in Land Use Decision-Making vol 59 p 5Vancouver Canada May 2007

[4] C A Francis K J Roberts J M Beman A E Santoro and B BOakley ldquoUbiquity and diversity of ammonia-oxidizing archaeainwater columns and sediments of the oceanrdquoProceedings of theNational Academy of Sciences of theUnited States of America vol102 no 41 pp 14683ndash14688 2005

[5] A H Treusch S Leininger A Kietzin S C Schuster H-PKlenk and C Schleper ldquoNovel genes for nitrite reductase andAmo-related proteins indicate a role of uncultivatedmesophiliccrenarchaeota in nitrogen cyclingrdquo Environmental Microbiol-ogy vol 7 no 12 pp 1985ndash1995 2005

[6] G W Nicol S Leininger C Schleper and J I Prosser ldquoTheinfluence of soil pH on the diversity abundance and tran-scriptional activity of ammonia oxidizing archaea and bacteriardquoEnvironmentalMicrobiology vol 10 no 11 pp 2966ndash2978 2008

[7] P Offre A Spang and C Schleper ldquoArchaea in biogeochemicalcyclesrdquo Annual Review of Microbiology vol 67 pp 437ndash4572013

[8] MAPA Anuario Estatıstico da Agroenergia 2012 StatisticalYearbook of Agroenergy 2013

[9] J C Bergmann D D Tupinamba O Y A Costa J R MAlmeida C C Barreto andB FQuirino ldquoBiodiesel production

12 Archaea

in Brazil and alternative biomass feedstocksrdquo Renewable ampSustainable Energy Reviews vol 21 pp 411ndash420 2013

[10] R G Taketani and S M Tsai ldquoThe influence of different landuses on the structure of archaeal communities in Amazoniananthrosols based on 16S rRNA and amoA genesrdquo MicrobialEcology vol 59 no 4 pp 734ndash743 2010

[11] C Brochier-Armanet B Boussau S Gribaldo and P ForterreldquoMesophilic crenarchaeota proposal for a third archaeal phy-lum theThaumarchaeotardquoNature Reviews Microbiology vol 6no 3 pp 245ndash252 2008

[12] S M Barns C F Delwiche J D Palmer and N R PaceldquoPerspectives on archaeal diversity thermophily and mono-phyly from environmental rRNA sequencesrdquo Proceedings of theNational Academy of Sciences of the United States of Americavol 93 no 17 pp 9188ndash9193 1996

[13] J G Elkins M Podar D E Graham et al ldquoA korarchaealgenome reveals insights into the evolution of the Archaeardquo Pro-ceedings of the National Academy of Sciences of the United Statesof America vol 105 no 23 pp 8102ndash8107 2008

[14] H Huber M J Hohn R Rachel T Fuchs V C Wimmer andK O Stetter ldquoA new phylum of Archaea represented by ananosized hyperthermophilic symbiontrdquo Nature vol 417 no6884 pp 63ndash67 2002

[15] T Nunoura H Oida M Nakaseama et al ldquoArchaeal diversityand distribution along thermal and geochemical gradients inhydrothermal sediments at the Yonaguni Knoll IV hydrother-mal field in the SouthernOkinawa TroughrdquoApplied and Enviro-nmental Microbiology vol 76 no 4 pp 1198ndash1211 2010

[16] C Rinke P Schwientek A Sczyrba et al ldquoInsights into thephylogeny and coding potential of microbial dark matterrdquoNature vol 499 no 7459 pp 431ndash437 2013

[17] A Spang J H Saw S L Joslashrgensen et al ldquoComplex archaea thatbridge the gap between prokaryotes and eukaryotesrdquo Naturevol 521 no 7551 pp 173ndash179 2015

[18] S Bagchi S E Vlaeminck L A Sauder et al ldquoTemporal andspatial stability of ammonia-oxidizing archaea and bacteria inaquariumbiofiltersrdquo PLoSONE vol 9 no 12 Article ID e1135152014

[19] W Qin S A Amin W Martens-Habbena et al ldquoMarineammonia-oxidizing archaeal isolates display obligate mixotro-phy and wide ecotypic variationrdquo Proceedings of the NationalAcademy of Sciences of the United States of America vol 111 no34 pp 12504ndash12509 2014

[20] M Coci N Odermatt M M Salcher J Pernthaler and GCorno ldquoEcology and distribution ofThaumarchaea in the deephypolimnion of Lake Maggiorerdquo Archaea vol 2015 Article ID590434 11 pages 2015

[21] J G Ferry ldquoHow tomake a living by exhalingmethanerdquoAnnualReview of Microbiology vol 64 pp 453ndash473 2010

[22] A Meyer A Focks V Radl G Welzl I Schoning and MSchloter ldquoInfluence of land use intensity on the diversity ofammonia oxidizing bacteria and archaea in soils from grasslandecosystemsrdquoMicrobial Ecology vol 67 no 1 pp 161ndash166 2014

[23] V De Gannes G Eudoxie and W J Hickey ldquoImpacts ofedaphic factors on communities of ammonia-oxidizing archaeaammonia-oxidizing bacteria and nitrification in tropical soilsrdquoPLoS ONE vol 9 no 2 Article ID e89568 2014

[24] D S Ribeiro F C J da Silva and TN Carvalho ldquoSobrevivenciade seis especies florestais em uma area explorada seletivamenteno municıpio de Moju Parardquo CERNE vol 9 no 2 pp 153ndash1632003

[25] D HM Costa C A P Ferreira J NM Silva J D C A Lopesand J O P D Carvalho ldquoPotencial madeireiro de floresta densano municıpio de Moju Estado do Parardquo Documentos 1998

[26] C M Almeida S F Lima C V Martins-Da-Silva and JI Gomes ldquoCaracterizacao morfologica e anatomica de dezespecies de leguminosae ocorrentes em uma floresta tropicalumida localizada no municıpio de Moju Estado do Parardquo inA Silvicultura na Amazonia Oriental Contribuicoes do ProjetoEmbrapa-DFID J NMC Silva JO P deCarvalho and J AGYared Eds pp 19ndash54 Embrapa Amazonia Oriental (CPATU)Belem Brazil 2001

[27] K Smalla N Cresswell L C Mendoncahagler A Woltersand J D Vanelsas ldquoRapid extraction protocol from soil forpolymerase chain reaction-mediated amplificationrdquo Journal ofApplied Bacteriology vol 74 pp 78ndash85 1993

[28] S Gantner A F Andersson L Alonso-Saez and S Bertils-son ldquoNovel primers for 16S rRNA-based archaeal communityanalyses in environmental samplesrdquo Journal of MicrobiologicalMethods vol 84 no 1 pp 12ndash18 2011

[29] P D Schloss S L Westcott T Ryabin et al ldquoIntroducingMOTHUR open-source platform-independent community-supported software for describing and comparing microbialcommunitiesrdquoApplied and EnvironmentalMicrobiology vol 75no 23 pp 7537ndash7541 2009

[30] PD SchlossDGevers and S LWestcott ldquoReducing the effectsof PCR amplification and sequencing artifacts on 16S rRNA-based studiesrdquo PLoS ONE vol 6 no 12 Article ID e27310 2011

[31] CQuince A Lanzen T P Curtis et al ldquoAccurate determinationof microbial diversity from 454 pyrosequencing datardquo NatureMethods vol 6 no 9 pp 639ndash641 2009

[32] P Yilmaz L W Parfrey P Yarza et al ldquoThe SILVA and lsquoall-species Living Tree Project (LTP)rsquo taxonomic frameworksrdquoNucleic Acids Research vol 42 no 1 pp D643ndashD648 2014

[33] R C Edgar B J Haas J C Clemente C Quince and RKnight ldquoUCHIME improves sensitivity and speed of chimeradetectionrdquo Bioinformatics vol 27 no 16 pp 2194ndash2200 2011

[34] A Chao and T-J Shen ldquoNonparametric estimation of Shan-nonrsquos index of diversity when there are unseen species insamplerdquo Environmental and Ecological Statistics vol 10 no 4pp 429ndash443 2003

[35] A Chao and T-J Shen Program SPADE (Species Prediction andDiversity Estimation) 2010

[36] C L Shannon andWWeaverTheMathematicalTheory of Com-munication University of Illinois Press Urbana Ill USA 1949

[37] I J Good ldquoThe population frequencies of species and theestimation of population parametersrdquo Biometrika vol 40 pp237ndash264 1953

[38] K Tamura G Stecher D Peterson A Filipski and S KumarldquoMEGA6molecular evolutionary genetics analysis version 60rdquoMolecular Biology and Evolution vol 30 no 12 pp 2725ndash27292013

[39] DA BensonMCavanaugh K Clark et al ldquoGenBankrdquoNucleicAcids Research vol 41 no 1 pp D36ndashD42 2013

[40] D H Parks G W Tyson P Hugenholtz and R G BeikoldquoSTAMP statistical analysis of taxonomic and functional pro-filesrdquo Bioinformatics vol 30 no 21 pp 3123ndash3124 2014

[41] R G Newcombe ldquoInterval estimation for the differencebetween independent proportions comparison of elevenmeth-odsrdquo Statistics in Medicine vol 17 no 8 pp 873ndash890 1998

[42] J D Storey J E Taylor and D Siegmund ldquoStrong controlconservative point estimation and simultaneous conservative

Archaea 13

consistency of false discovery rates a unified approachrdquo Journalof the Royal Statistical Society Series B Statistical Methodologyvol 66 no 1 pp 187ndash205 2004

[43] G C Baker J J Smith and D A Cowan ldquoReview and re-analysis of domain-specific 16S primersrdquo Journal of Microbio-logical Methods vol 55 no 3 pp 541ndash555 2003

[44] W H Zhong and Z C Cai ldquoLong-term effects of inorganicfertilizers on microbial biomass and community functionaldiversity in a paddy soil derived from quaternary red clayrdquoApplied Soil Ecology vol 36 no 2-3 pp 84ndash91 2007

[45] EWessen S Hallin and L Philippot ldquoDifferential responses ofbacterial and archaeal groups at high taxonomical ranks to soilmanagementrdquo Soil Biology and Biochemistry vol 42 no 10 pp1759ndash1765 2010

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[47] K Cenciani M R Lambais C C Cerri L C B De Azevedoand B J Feigl ldquoBacteria diversity and microbial biomass inforest pasture and fallow soils in the southwestern Amazonbasinrdquo Revista Brasileira de Ciencia do Solo vol 33 no 4 pp907ndash916 2009

[48] A A Navarrete R G Taketani L WMendes F de Souza Can-navan F M de Souza Moreira and S M Tsai ldquoLand-use sys-tems affect Archaeal community structure and functional diver-sity in western Amazon soilsrdquo Revista Brasileira de Ciencia doSolo vol 35 no 5 pp 1527ndash1540 2011

[49] E Lumini A Orgiazzi R Borriello P Bonfante and V Bian-ciotto ldquoDisclosing arbuscular mycorrhizal fungal biodiversityin soil through a land-use gradient using a pyrosequencingapproachrdquo Environmental Microbiology vol 12 no 8 pp 2165ndash2179 2010

[50] M Lupatini R J S Jacques Z I Antoniolli A K A SuleimanR R Fulthorpe and L F W Roesch ldquoLand-use change andsoil type are drivers of fungal and archaeal communities in thePampa biomerdquo World Journal of Microbiology and Biotechnol-ogy vol 29 no 2 pp 223ndash233 2013

[51] E G Lamb N Kennedy and S D Siciliano ldquoEffects of plantspecies richness and evenness on soil microbial communitydiversity and functionrdquo Plant and Soil vol 338 no 1 pp 483ndash495 2011

[52] A Spang R Hatzenpichler C Brochier-Armanet et al ldquoDis-tinct gene set in two different lineages of ammonia-oxidizingarchaea supports the phylum Thaumarchaeotardquo Trends inMicrobiology vol 18 no 8 pp 331ndash340 2010

[53] R G Pacchioni F M Carvalho C E Thompson et al ldquoTax-onomic and functional profiles of soil samples from Atlanticforest and Caatinga biomes in northeastern Brazilrdquo Microbio-logyOpen vol 3 no 3 pp 299ndash315 2014

[54] E Catao A P Castro C C Barreto R H Kruger and C MKyaw ldquoDiversity of Archaea in Brazilian savanna soilsrdquoArchivesof Microbiology vol 195 no 7 pp 507ndash512 2013

[55] M Pester T Rattei S Flechl et al ldquoAmoA-based consensusphylogeny of ammonia-oxidizing archaea and deep sequencingof amoA genes from soils of four different geographic regionsrdquoEnvironmental Microbiology vol 14 no 2 pp 525ndash539 2012

[56] J-Z He H-W Hu and L-M Zhang ldquoCurrent insights into theautotrophic thaumarchaeal ammonia oxidation in acidic soilsrdquoSoil Biology amp Biochemistry vol 55 pp 146ndash154 2012

[57] L-M Zhang H-W Hu J-P Shen and J-Z He ldquoAmmonia-oxidizing archaea have more important role than ammonia-oxidizing bacteria in ammonia oxidation of strongly acidicsoilsrdquo ISME Journal vol 6 no 5 pp 1032ndash1045 2012

[58] S Leininger T Urich M Schloter et al ldquoArchaea predominateamong ammonia-oxidizing prokaryotes in soilsrdquo Nature vol442 no 7104 pp 806ndash809 2006

[59] N C Banning L D Maccarone L M Fisk and D VMurphy ldquoAmmonia-oxidising bacteria not archaea dominatenitrification activity in semi-arid agricultural soilrdquo ScientificReports vol 5 Article ID 11146 2015

[60] M Herrmann A Hadrich and K Kusel ldquoPredominance ofthaumarchaeal ammonia oxidizer abundance and transcrip-tional activity in an acidic fenrdquoEnvironmentalMicrobiology vol14 no 11 pp 3013ndash3025 2012

[61] K Zhalnina P Dorr de Quadros F O Camargo and E WTriplett ldquoDrivers of archaeal ammonia-oxidizing communitiesin soilrdquo Frontiers in Microbiology vol 3 article 210 Article IDArticle 210 2012

[62] J Hong J Cho and Y Hong ldquoEnvironmental variables shapingthe ecological niche of Thaumarchaeota in soil direct andindirect causal effectsrdquo PLoS ONE vol 10 no 8 Article IDe0133763 2015

[63] G Jurgens K Lindstrom and A Saano ldquoNovel group withinthe kingdom Crenarchaeota from boreal forest soilrdquo Appliedand Environmental Microbiology vol 63 no 2 pp 803ndash8051997

[64] M Tourna M Stieglmeier A Spang et al ldquoNitrososphaeraviennensis an ammonia oxidizing archaeon from soilrdquoProceed-ings of the National Academy of Sciences of the United States ofAmerica vol 108 no 20 pp 8420ndash8425 2011

[65] K V Zhalnina R Dias M T Leonard et al ldquoGenome sequenceof Candidatus Nitrososphaera evergladensis from group I1benriched from everglades soil reveals novel genomic features ofthe ammonia-oxidizing archaeardquoPLoSONE vol 9 no 7 ArticleID e101648 2014

[66] I Anderson L E Ulrich B Lupa et al ldquoGenomic char-acterization of methanomicrobiales reveals three classes ofmethanogensrdquo PLoS ONE vol 4 no 6 Article ID e5797 2009

[67] J E Galagan C Nusbaum A Roy et al ldquoThe genome of Macetivorans reveals extensive metabolic and physiological diver-sityrdquo Genome Research vol 12 no 4 pp 532ndash542 2002

[68] C Wrede S Kokoschka A Dreier C Heller J Reitner and MHoppert ldquoDeposition of biogenic iron minerals in a methaneoxidizing microbial matrdquo Archaea vol 2013 Article ID 1029728 pages 2013

[69] G Garrity Bergeyrsquo Manual of Systematic Bacteriology 721 2001[70] S Sakai Y Takaki S Shimamura et al ldquoGenome sequence

of a mesophilic hydrogenotrophic methanogen Methanocellapaludicola the first cultivated representative of the orderMethanocellalesrdquo PLoS ONE vol 6 no 7 Article ID e228982011

[71] E Bapteste C Brochier and Y Boucher ldquoHigher-level clas-sification of the Archaea evolution of methanogenesis andmethanogensrdquo Archaea vol 1 no 5 pp 353ndash363 2005

[72] R G E Murray and E Stackebrandt ldquoTaxonomic note imple-mentation of the provisional status Candidatus for incompletelydescribed procaryotesrdquo International Journal of Systematic Bac-teriology vol 45 no 1 pp 186ndash187 1995

[73] L E Lehtovirta-Morley K Stoecker A Vilcinskas J I Prosserand G W Nicol ldquoCultivation of an obligate acidophilic ammo-nia oxidizer from a nitrifying acid soilrdquo Proceedings of the

14 Archaea

National Academy of Sciences of the United States of Americavol 108 no 38 pp 15892ndash15897 2011

[74] L E Lehtovirta-Morley C Ge J Ross H Yao G W Nicol andJ I Prosser ldquoCharacterisation of terrestrial acidophilic archaealammonia oxidisers and their inhibition and stimulation byorganic compoundsrdquo FEMSMicrobiology Ecology vol 89 no 3pp 542ndash552 2015

[75] M J Frijlink T Abee H J Laanbroek W de Boer and W NKonings ldquoThe bioenergetics of ammonia and hydroxylamineoxidation in Nitrosomonas europaea at acid and alkaline pHrdquoArchives of Microbiology vol 157 no 2 pp 194ndash199 1992

[76] B Wang Y Zheng R Huang et al ldquoActive ammonia oxidizersin an acidic soil are phylogenetically closely related to neu-trophilic archaeonrdquo Applied and Environmental Microbiologyvol 80 no 5 pp 1684ndash1691 2014

[77] J Y Aller and P F Kemp ldquoAre Archaea inherently less diversethan Bacteria in the same environmentsrdquo FEMS MicrobiologyEcology vol 65 no 1 pp 74ndash87 2008

[78] K Potthast U Hamer and FMakeschin ldquoLand-use change in atropical mountain rainforest region of southern Ecuador affectssoilmicroorganisms and nutrient cyclingrdquoBiogeochemistry vol111 no 1ndash3 pp 151ndash167 2012

[79] J I Prosser and G W Nicol ldquoArchaeal and bacterial ammonia-oxidisers in soil the quest for niche specialisation and differen-tiationrdquoTrends inMicrobiology vol 20 no 11 pp 523ndash531 2012

Submit your manuscripts athttpwwwhindawicom

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Anatomy Research International

PeptidesInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporation httpwwwhindawicom

International Journal of

Volume 2014

Zoology

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Molecular Biology International

GenomicsInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioinformaticsAdvances in

Marine BiologyJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Signal TransductionJournal of

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BioMed Research International

Evolutionary BiologyInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

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Biochemistry Research International

ArchaeaHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

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Advances in

Virolog y

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Nucleic AcidsJournal of

Volume 2014

Stem CellsInternational

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

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Enzyme Research

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

International Journal of

Microbiology

2 Archaea

Although the Amazon is one of the most species-richbiomes on Earth little is known about its archaeal diversityTo date only one published microbial ecology study hasfocused on Amazon soil archaeal diversity using 16S rRNAgene sequencing [10] and only one soil type (ie Amazoniandark earth also called Terra Preta) was studied

The archaeal taxonomy is a matter of constant changesince its proposal in 1977 Initially two phyla were recognizedCrenarchaeota and Euryarchaeota but in the subsequentyears many new phyla were proposed One example is theThaumarchaeota phylum [11] composed predominantly ofmesophilicmembers it encompasses the ammonia-oxidizingarchaea The phylum Korarchaeota was proposed in 1996after the identification of DNA sequences from the ObsidianPool in Yellowstone National Park [12] composed of onecandidate thermophilic species whose genome was com-pletely sequenced [13] Recent works have described putativenew phyla such as Nanoarchaeota Aigarchaeota Aenig-marchaeota Parvarchaeota and Lokiarchaeota but thesephyla are not widely accepted yet due to the low numberof specimens or DNA sequences available In addition fur-ther analyses have positioned sequences belonging to thesephyla in already described phyla such as Euryarchaeota orThaumarchaeota [14ndash17] There are some archaeal groupssuch as MCG (Miscellaneous Crenarchaeotic Group) whichare poorly characterized in terms of phylogenetic affiliationrecent data revealed that this group is probably more closelyrelated to the phylum Thaumarchaeota than to the Crenar-chaeota

Mesophilic archaea seem to play important roles in thecycling of important nutrients such as nitrogen and carbonThe importance of ammonia-oxidizing archaea (AOAs) hasbeen well documented in different ecosystems such as soilsmarine and freshwater environments where they sometimescan be found in higher abundance than the ammonia-oxidizing bacteria (AOBs) [18ndash20] On the other hand theirreal role in nitrification is not yet well understood due to thescarce number of cultured AOAs and the few physiologicalstudies available for this group The methanogens are amongthe nonextremophilic archaea which are widely distributedin anaerobic environments such as flooded soils or marinesoils and vents Methanogens are also found in the gut oftermites in rumenof cows or even in themouth and intestineof humans These organisms play important roles in thecarbon cycle transforming small compounds such as acetateand propionate into methane and removing the hydrogenwhich is potentially hazardous to some bacterial cells On theother hand methane production is one of the major gasesinvolved in the planetrsquos greenhouse effect (reviewed by [21])

There are several studies associating land use withchanges in the structure and abundance of soil microbialcommunities such as the influence of the land use over thediversity of AOA and AOB in grassland soils [22] and theimpacts of edaphic factors on those archaea in tropical soils[23] Therefore the conversion of native forest into palm treeculture can be another example of a potential impact of the oilpalm cultivation in the archaeal communities of Amazoniansoils

This work aimed to improve our understanding of howthe soil archaeal community is impacted by oil palm cultiva-tion To this endmicrobialDNAwas extracted from soil sam-ples from native forest and an adjacent area under oil palmcultivation The archaeal 16S ribosomal RNA (rRNA) genewas amplified and sequenced using high-throughput meth-ods for comparative analysis Here we show for the first timethat soil archaeal diversity is reduced in soil under oil palmcultivation compared to native forest soil

2 Materials and Methods

21 Site Description Sampling and Processing Soil sampleswere collected in the State of Para Brazil in an oil palm-cultivated area and an adjacent area of Amazon native forestnear the city of Moju (Figure 1) The tropical forest in thisregion is dense with trees that are 25ndash35m tall [24] The cli-mate is equatorial hot and humid (Ami type according to theKoppen climate classification) Annual temperatures rangefrom 25∘C to 27∘C and rainfall is 2000ndash3000mm per yearbeing irregularly distributed [25] The soil is predominantlyldquoLatossolo Amarelordquo (a type of Oxisol) [26]

The oil palm cultivation in the sampled farm is not ascontrolled as other crops (Figure 1(b)) There is no irrigationregime natural precipitation of the rainforest is the only waythese plants are irrigated In the Amazon the soil is verymoist due to the high precipitation levels during the yearIn the studied area the annual period of flooding is fromFebruary to April Furthermore the soil around the palmtrees is not fertilized in a homogeneous fashion since onlyone side of the plants is directly fertilized

In October 2010 after plant litter was removed asoil borer was used to obtain four 10 cm deep soilsamples from three points in the oil palm-cultivatedarea (S02∘00101584028910158401015840W048∘37101584057410158401015840 S02∘00101584029210158401015840W048∘37101584056610158401015840 and S02∘00101584031310158401015840W048∘37101584054310158401015840) (Figure 1(b))and four samples from the native Amazon forest area(S02∘00101584027210158401015840W048∘35101584053010158401015840) (Figure 1(a)) The samplescollected in each area were mixed ground and sieved toremove larger particles yielding one composite sample foreach area with approximately 1 kg each The samples werestored in plastic bags on dry ice during transportationA subsample was sent to physicochemical analysis atSoloQuımica Analises de Solo Ltda (Brasılia DF Brazil)The rest of the samples were then stored at minus80∘C until DNAextraction Initially the physicochemical characteristics ofthe soils samples were evaluated individually and a highvariation among replicas was observed This result was dueto the heterogeneous fertilization of the palm trees in thecultivation fields in Amazon therefore composite sampleswere necessary to describe the microbiota in oil palm soil

22 DNA Extraction PCR and Pyrosequencing AnalysisTotal DNA was extracted according to the protocol of Smallaet al [27] using 2 g soil per sample TominimizeDNAextrac-tion bias this procedure was performed in quadruplicatePCR reactions were performed using the following primersspecific for Archaea 340F (51015840-CCC TAY GGG GYG CAS

Archaea 3

(a) (b)

Figure 1 (a) Native forest and (b) oil palm-cultivated sites

CAG-31015840) and 1000R (51015840-GGC CAT GCA CYW CYT CTC-31015840) [28] Archaea 16S rRNA genes were amplified yielding660 bp amplicons Adapters used as priming sites for bothamplification and sequencing (454 Life Sciences BranfordCT USA) were ligated to the 51015840 end of the primer sequencesEach 20120583L PCR reaction contained 10ndash30 ng total DNA 1xreaction buffer 4120583MdNTP 10 120583Mof each primer 200120583gmLbovine serum albumin 05U KAPA2G Robust HotStartpolymerase andMilli-Qwater Amplification was performedin anApplied BiosystemsGeneAmpPCRSystem9700Ther-mal Cycler (Applied Biosystems Foster City CA USA) usingthe following program 2min at 98∘C followed by 30 cyclesof 30 seconds at 95∘C 30 seconds at 57∘C and 1 minute and30 seconds at 72∘C and a final step consisting of 7 minutes at72∘C To minimize PCR bias at least four PCR amplificationreactions per sample were pooled before purification usingthe GeneJET PCR Purification Kit (Thermo Fisher ScientificWaltham MA USA) After quantification using a Qubitfluorometer (Invitrogen Carlsbad CA USA) NanoDrop1000 Spectrophotometer (Thermo Fisher Scientific) and 2agarose gel with ethidium bromide DNA samples were sentfor pyrosequencing (GSFLXTitaniumPlatformatMacrogenSouth Korea) A total of 592145 reads greater than 500 bpwere obtained in three-quarters of a plate 49314 reads fromthe oil palm-cultivated soil and 542831 reads from the nativeforest soil

23 Data Analysis All 16S rRNA gene pyrosequencing readswere analyzed using the original standard flowgram format(SFF) output file from the sequencer in MOTHUR version1333 [29] for the removal of short sequences sequences witherrors low-quality sequences chimeras and possible con-taminants After this removal a total of 39111 sequences foroil palm-cultivated area and 436532 sequences for native for-est were obtained Further analysis was performed after nor-malizing the number of sequences per sample to the lowest

number of high-quality sequences (ie 39111 sequences in theoil palm-cultivated area sample)

All 16S rRNA gene reads were analyzed using MOTHURfor taxonomic classification following the 454 standardoperating procedure available online (httpwwwmothurorgwiki454 SOP) [30] The PyroNoise algorithm [31]was used to denoise the flowgram file (removal of barcodesand quality filtering) The SILVA database (release 119) wasused for sequence alignment and clustering based on 97sequence similarity (003 genetic distance) using the nearestneighbor method [32] The UCHIME algorithm [33] wasused for detection and removal of chimeric reads thisfunction was implemented in MOTHUR and performedwithout a reference database The one-gap method inMOTHURwas used to calculate the pairwise distancematrixArchaeal taxonomic classifications of each representativeOTU were performed in MOTHUR using the SILVA data-base (release 119) clustered at 97 sequence similarity [32]This classificationwas used to estimate the relative abundanceof reads per genus Samples were randomly chosen forsubsampling and normalized to the lowest number ofsequence reads obtained Chao1 [34] abundance-basedcoverage estimator (ACE) [35] Shannon diversity index [36]and Goodrsquos coverage [37] were then used to estimatealpha diversity MOTHUR was also used to perform allstatistical analyses A phylogenetic tree was created usingall Thaumarchaeota OTUs from each sample that wereunclassified at the class level A new tree was created withsequences randomly chosen by MOTHUR that were deemedrepresentative of the entire set Data were analyzed using theneighbor-joining method with the Jukes-Cantor correctionand 1000 bootstrap replicates in MEGA6 [38] In additionOTUs with 3 dissimilarity were selected by MOTHUR toavoid redundancies in the phylogenetic tree The chosensequences were aligned with 16S rRNA gene sequencesfrom GenBank [39] representing the main Archaeagroups Methanosarcina baltica (AB973356) Methanoregula

4 Archaea

Table 1 Number of operational taxonomic units (OTUs) Goodrsquos coverage estimator OTU richness indices (Chao1 ACE) and InverseSimpson and Shannon diversity indices for archaeal communities in native forest soil and soil under oil palm cultivation

Sample OTUslowast Goodrsquos coverage () Chao1 ACE Inverse Simpson ShannonNative forest 821 9871 2124 4645 2645 422Oil palm-cultivated area 253 9972 424 674 1547 331lowastConsidering 97 sequence similarity in MOTHUR

formicica (NR112877) Methanocella paludicola (NR074192)Halococcus hamelinensis (LN651155) and Pyrococcus furiosus(U20163) from the phylum Euryarchaeota Pyrobaculumaerophilum (NR102764) from the phylum CrenarchaeotaCandidatus Nitrososphaera sp (FR773157) and CandidatusNitrososphaera gargensis (GU797786) fromThaumarchaeotaGroup I1b Nitrosopumilus maritimus (JQ346765) andCenarchaeum symbiosum (U51469) from ThaumarchaeotaGroup I1a uncultured acidic red soil AOA (FJ174727)uncultured trembling aspen archaeon (EF021427) and borealforest archaeon (X96688) from Thaumarchaeota GroupI1c and several uncultured organisms classified accordingto the SILVA database as Thaumarchaeota MiscellaneousCrenarchaeotic Group (FR745121 AM910782 JX984848KC510333 FJ485299 KC831395 FJ920714 FJ485307HM051127 HM051130 and HM051125) A 16S rRNA genesequence from Acidobacteria KBS 96 (FJ870384) was usedas an outgroup A second phylogenetic tree was createdusing only Euryarchaeota OTUs from each sample that wereunclassified at the class level with the same parameters andreference sequences

A Venn diagram was generated by grouping OTUs clus-tered at 97 similarity to show the number of OTUs uniqueto each area and those shared between samples

Differences in taxonomic distribution of archaea betweennative forest and the oil palm-cultivated area were evaluatedby Fisherrsquos exact test (119902 le 005) using Statistical Analysisof Metagenomic Profiles (STAMP) software [40] The confi-dence interval was estimated using the asymptotic method[41] and correction of the 119902 value was calculated usingStoreyrsquos false discovery rate (FDR) approach [42] A filterwas applied to show only the differences between proportionsabove 05

3 Results

Goodrsquos coverage estimate of the completeness of sampling(Table 1) was higher for the oil palm-cultivated area than forthe native forest

The number of archaeal OTUs was higher in the nativeforest soil (821 sequences) than in soil under oil palmcultivation (253 sequences) (Figure 2) with 130OTUs (138)shared between the two areas Archaeal species richness asassessed by abundance-based estimators Chao1 and ACEwas higher in the native forest than in the oil palm-cultivatedarea Similarly the Inverse Simpson and Shannon indicesindicated that archaeal species diversity was higher in nativeforest soil compared to soil under oil palm cultivation(Table 1)

Table 2 Physicochemical properties of native forest soil and soilunder oil palm cultivation

Native forest Oil palm-cultivated areapH(in water) 550 483

P (mgsdotdmminus3) 140 7070K (mgsdotdmminus3) 003 011Ca+2 (cmolsdotdmminus3) 140 033Mg+2 (cmolsdotdmminus3) 010 017Al+3 (cmolsdotdmminus3) 040 073H + Al (cmolsdotdmminus3) 620 727Na+ (cmolsdotdmminus3) 001 002Organic matter (gkg) 3100 1163Total C (gkg) 1800 677Total N (mgkg) NA 115NO3

minus1 (mgkg) NA 079NO2

minus1 (mgkg) NA 014NH4

+ (mgkg) NA 021Data not available

Soil physicochemical parameters differed between theareas (Table 2) with higher levels of phosphorous potassiumand total carbon in the oil palm-cultivated soil and higherlevels of calcium and organic matter in the native forest soil

In both areas only two archaeal phyla (Thaumarchaeotaand Euryarchaeota) were detected (Figure 3) Thaumar-chaeota were predominant in the native forest soil (58) andEuryarchaeota were predominant in the oil palm-cultivatedsoil (54)The percentage of unclassified archaea was similarin both soils (native forest 5 oil palm-cultivated area 6)

At the class level a total of 10 groups were detected inboth samples Halobacteria Methanobacteria Methanomi-crobia and Thermoplasmata in the phylum Euryarchaeotaand Marine Group I OPPD003 Soil Crenarchaeotic GroupSouth African Gold Mine Gp 1 Terrestrial Group andMiscellaneous Crenarchaeotic Group in the phylum Thau-marchaeota (Figures 4(a) and 4(b)) The most abundantclass present in native forest soil was the Soil Crenar-chaeotic Group representing 26 of all native forest OTUs(Figure 4(a)) and 45 of theThaumarchaeota sequences (notshown) followed by Methanomicrobia representing 15 ofall native forest OTUs (Figure 4(b)) and 42 of the Eur-yarchaeota sequences (not shown) The most abundant classpresent in the oil palm-cultivated area was Thermoplasmatarepresenting 21 of all cultivated area OTUs (Figure 4(b))and 39 of the Euryarchaeota sequences (not shown) fol-lowed by Methanomicrobia representing 16 of all culti-vated area OTUs (Figure 4(b)) and 30 of all Euryarchaeota

Archaea 5

Venn diagram at distance 003

123Cultivated area

691Native forest

130

Figure 2 Venn diagram depicting the number of shared and unique operational taxonomic units (OTUs) in native forest and oil palm-cultivated area

Thaumarchaeota

Euryarchaeota

Unclassified archaea

Cultivated areaNative forest

10 20 30 40 50 60 700()

Figure 3 Comparison of archaeal phyla present in soil samples from native Amazon forest (orange) and an area under oil palm cultivation(blue) The percentages of unclassified sequences are shown

sequences (not shown) The predominant class from thephylum Thaumarchaeota in the oil palm-cultivated areawas Terrestrial Group representing 14 of the total OTUs(Figure 4(a)) and 35 of all Thaumarchaeota sequences (notshown)

Because it is a newer phylum Thaumarchaeota have fewcultivated representatives compared with Euryarchaeota andfewer sequences in databases such as SILVA To determinehow the Thaumarchaeota sequences that could not be clas-sified at the genus level were related to each other a phylo-genetic tree was constructed with representative sequences(Figure 5(a)) One native forest OTU clustered with GroupI1b (NF6) and one native forest OTU (NF17) and two cul-tivated area OTUs (CA50 CA52) clustered with Group I1cThe remaining OTUs formed different clusters all associatedwith the Miscellaneous Crenarchaeotic Group Most unclas-sified Thaumarchaeota sequences from the native forest andoil palm-cultivated area grouped together although a fewunclassified OTUs from the cultivated area (eg CA51 toCA15 and CA24 to CA33) and native forest (eg NF21 toNF49 and NF5 to NF35) formed separate clusters

The unclassified sequences at genus level in the Euryar-chaeota phylum were all related to the orders Methano-cellales Methanosarcinales Methanomicrobiales Thermo-plasmatales Methanobacteriales and Halobacteriales(Figure 5(b)) The classified sequences at genus level wereMethanocella Methanosarcina Methanoculleus Methano-massiliicoccus Methanomethylophilus MethanobrevibacterMethanobacterium and Candidatus lainarchaeum(Figure 5(b))

The genus-level analysis showed that unclassified SoilCrenarchaeotic Group was predominant in the native forestaccounting for 21 of all native forest OTUs (Figure 6(a))In the oil palm-cultivated area the predominant genus wasUnclassified Terrestrial Group accounting for 14 of allcultivated area OTUs (Figure 6(b))

A total of 31 groups of OTUs were identified (onlymajor groups are shown) most of which are unclassi-fied The classified groups in the phylum Euryarchaeotaare Candidatus lainarchaeum Methanobacterium Methano-brevibacter Methanocella Rice Cluster I Methanoculleus

6 Archaea

Marine Group I

Miscellaneous Crenarchaeotic Group

Soil Crenarchaeotic Group

South African Gold Mine Gp 1

Terrestrial Group

Unclassified Thaumarchaeota

OPPD003

Cultivated areaNative forest

5 10 15 20 25 300()

(a)

Methanobacteria

Methanomicrobia

Halobacteria

Thermoplasmata

Unclassified Euryarchaeota

Cultivated areaNative forest

5 10 15 20 250()

(b)

Figure 4 Comparison of archaeal classes present in soil samples from native Amazon forest (orange) and an area under oil palm cultivation(blue) Classes within the phyla Thaumarchaeota (a) and Euryarchaeota (b) are shown as well as the percentage of unclassified sequences

Methanoregula Methanimicrococcus Methanosarcina Can-didatus Methanomethylophilus and Methanomassiliicoccus(Figure 6(b)) The classified groups in the phylum Thau-marchaeota are Candidatus Nitrososphaera and CandidatusNitrosotalea (Figure 6(a))

The analysis of the genera that differ significantly inrelative abundance between native forest soil and the oilpalm-cultivated area revealed that the greatest difference wasseen for Candidatus Nitrososphaera (Figure 7) This genuswas predominant in the native forest soil accounting for182 of the sequences but represented only 103 of thesequences from the oil palm-cultivated area UnclassifiedSoil Crenarchaeotic Group was also predominant in thenative forest (ie 388 of sequences in the native forest

and 312 of sequences in the oil palm-cultivated area)whereas unclassified South African Gold Mine Gp 1 waspredominant in the oil palm-cultivated area (ie 73 ofsequences in the native forest and 135 of sequences inthe oil palm-cultivated area) Candidatus Nitrosotalea repre-sented 44 of the native forest sequences and 100 of theoil palm-cultivated area sequences The relative abundanceof unclassified Terrestrial Miscellaneous Group unclassifiedThaumarchaeotaunclassifiedThermoplasmatalesunclassifiedMiscellaneous Crenarchaeotic Group Methanosarcina andRice Cluster I also differed significantly between native forestsoil and soils under oil palm cultivation but these differenceswere smaller

Archaea 7

FJ4853071| Uncultured archaeon clone

FJ4852991| Uncultured archaeon clone

FJ9027141| Uncultured archaeon clone

KC8313951| Uncultured archaeon clone

KC5103331| Uncultured archaeon clone

JX9848481| Uncultured archaeon clone

AM9107821| Uncultured archaeon clone

Uncultured archaeon acidic red soil-FJ174727

Ca Nitrososphaera gargensis-GU797786

Ca Nitrososphaeramdash(48 6)

Ca Nitrosotaleamdash(49 12)

Candidatus lainarchaeum (0 2)

Methanobacterium (2 3)

Methanomassillicoccus (6 2)

Methanocella (33 8)

Rice cluster I (43 25)

Methanosarcina baltica-AB973356

Methanocella paludicola-NR074192

Pyrobaculum aerophilum

-NR102764

Acidobacteria bacterium KBS 96-FJ870384

Ca Methanom

ethylophilus (12 0)

Ca Nitrososphaera sp-FR773157

Nitrosopumilus maritimus-JQ346765

Halococcus hamelinensis-LN651155

Pyrococcus furiosus-PFU20163

Cenarchaeum symbiosum-CSU51469

Methanoregula formicica-NR112877

FR7451211| Uncultured soil archaeon

Boreal forest soil archaeon-X96688

Trembling aspen archaeon-EF021427

MCG

MCG005

CA4099

99

7174

63

86

9687

9067

5696

6768 54

57

62

58

65

7559

62

63

65

64

52

64

64

1008654

91

99

999293

53798392

88

7274

5164

5858

CA7CA32

CA50NF17

NF6

CA52

T I1c

T I1b

T I1a

CA24CA6CA12

CA4

CA33

NF16

NF2

NF35

NF9

NF5NF3

6NF4

1NF3

2

NF1

2

NF4

NF1

NF3

0NF2

3

NF7CA3

CA39

CA37

CA46

CA44

CA2

CA42

CA27

Unclassified MCGmdash(16 4)

Unclassified Terrestrial Group (94 36)

Unclassified Marine Group I (2 2)

CA15

CA25

CA30

CA43

CA35

CA14

CA29

CA13

CA20

CA11

CA41

CA36

CA51

NF48

CA9NF4

0NF4

2CA21

CA22

NF38

CA45

CA16

CA47

NF31CA31NF28NF47NF27NF45CA18NF26

NF24NF33NF19

NF18NF37

NF34NF43NF15NF44NF13

NF22NF29NF11

CA49

CA1

CA19CA8

CA10

CA23CA38

CA54HM0511251| Uncultured

HM0511301| Uncultured archaeon clone

HM0511271| Uncultured archaeon clone

archaeon clone

NF46NF14

NF49NF25N

F10N

F39N

F20N

F21

NF8

NF3

CA48

CA34

CA5

CA28CA26CA17CA53

(E)

(B)

(C)

(a)

CA13

CA14

NF1

2

NF1

4M

ethan

ocell

a (3

8)Meth

anosarcina (3

8 4)

Methanoculleus (2 0)

Methanomassilicoccus (6 2)

Methanomassilicoccus (6 2)

Methanomassilicoccus (6 2)

Methanomethylophilus (6 2)

Methanomethylophilus (6 2)

Methanobrevibacter (1 0)

Methanobacterium

(2 3)

Methanobacterium

(2 3)

Methanobacterium

(2 3)

Methanobacterium

(2 3)

Methanobacterium

(2 3)

Methanosarci

na (38 4)

Methanosarcina (38 4)

Methanosarcina (38 4)

Methanosarcina (38 4)

NF16

CA4

CA15CA20

CA19

NF3NF1

Meth

anoc

ella (

33 8

)

Meth

anoc

ella p

aludi

cola-

NR 0741

92 1

Meth

anosa

rcina

baltic

a AB97

3356

1

Methanoregula formicica NR 112877 1

Halococcus hamelinensis LN651155 1

CA6N

F6NF8

NF19 NF5NF7NF20

Pyrobaculum aerophilum NR 102764 2

Ca Nitrososphaera gargensis G

U797786 1

Ca Nitrososphaera gargensis GU797786 1

Nitrosopumilus maritimus JQ346765 1Cenarchaeum symbiosum U51469 1Uncultured archaeon acidic red soil FJ174727 1

Boreal forest soil archaeon X96688 1

Trembling aspen archaeon EF021427 1

HM051125 1 Uncultured archaeon clone

HM051127 1 Uncultured archaeon clone

HM051130 1 Uncultured archaeon clone

FJ485307 1 Uncultured archaeon clone

FJ902714 1 Uncultured archaeon clone

KC831395 1 archaeon clone

FJ485299 1 archaeon clone

KC510333 1 archaeon clone

FR745121 1 archaeon clone

JX984848 1 archaeon clone

AM910782 1 archaeon clone

Acidobacteria bacterium KBS 96 FJ870384 1

Ca Nitrososphaera sp FR773157 1

CA7

Pyrococcus furiosus U20163 1

005

MCG

Halobacteriales

Cand

idat

us la

inar

chae

um (0

2)

Cand

idat

us la

inar

chae

um (0

2)

Thermoplasmatales

Methanomicrobiales

Methanosarcinalles

Methanocellales

T I1c

T I1a

T I1b

(C)

998785

58

100

100

100100

100

71 84 71

53

54

65

86

94

98

59100

100

78

10051

5477

63 73

99

100

8497100

78

100

100

76956970

100 10

010

0

637551

100

9999

9958

94

Methanobacteriales

(b)

Figure 5 Phylogenetic trees of (a) unclassified Thaumarchaeota 16S rRNA gene sequences from native forest soil and soil under oil palmcultivation and (b) 16S rRNA gene sequences classified as Euryarchaeota from native forest soil and soil under oil palm cultivation Theneighbor-joining method and Jukes-Cantor corrections were used with 1000 bootstrap replicates Bootstrap values above 50 are shownin the trees A representative number of operational taxonomic units (OTUs) were selected at 3 dissimilarity by MOTHUR to avoidredundancies in the phylogenetic trees Randomly chosen OTUs from native forest soil and from soil under oil palm cultivation were usedin the construction of the trees along with archaeal reference sequences (from GenBank) from the following groups Acidobacteria (B)Crenarchaeota (C) Euryarchaeota (E) Miscellaneous Crenarchaeotic Group (MCG) Thaumarchaeota Group I1a (T I1a) ThaumarchaeotaGroup I1b (T I1b) andThaumarchaeota Group I1c (T I1c)The AOAs are highlighted with the symbolX Sampling site is indicated by ldquoNFrdquo(native forest) or ldquoCArdquo (oil palm-cultivated area) followed by the number of the OTUs (NAxx or CAxx) OTU sequences classified to at leastthe class level are shown in bold followed by two numbers in parentheses indicating the number of sequences of that OTU detected in nativeforest soil and the number detected in the oil palm-cultivated area The scale bar represents the 5 estimated sequence divergence

8 Archaea

Unclassified Marine Group I

Unclassified Miscellaneous Crenarchaeotic Group

Unclassified Thaumarchaeota

Candidatus Nitrososphaera

Unclassified Soil Crenarchaeotic Group

Candidatus Nitrosotalea

Unclassified South African Gold Mine Gp 1

Unclassified Terrestrial Group

Cultivated areaNative forest

5 10 15 20 250()

(a)

Unclassified Thermoplasmatales

Unclassified Terrestrial Miscellaneous Group

Rice Cluster I

Methanocella

Methanosarcina

Unclassified Euryarchaeota

Unclassified Halobacteriales

Candidatus Methanomethylophilus

Unclassified Methanosarcinaceae

5 10 15 20 250()

Cultivated areaNative forest

(b)

Figure 6 Percentage of archaeal operational taxonomic units within the Thaumarchaeota (a) and Euryarchaeota (b) phyla classified at thegenus level for native forest soil (orange) and soil under oil palm cultivation (blue) The percentage of unclassified sequences at genus level isshown

4 Discussion

The Amazon forest is known for its macrospecies diversityhowever few studies have addressed its microbial diversityAmong the domains of life Archaea are undoubtedly the lesswell known

In this work almost 600000 archaeal 16S rRNA genesequences were obtained from native forest soil and anadjacent area under oil palm cultivation After processing(ie removal of short sequences sequences with errors low-quality sequences chimeras and possible contaminants) andnormalization 39111 sequences per area remained for anal-ysis This number of sequences provided sufficient coverageof archaeal diversity in both areas (Table 1)The primers used

were based on a large number of 16S rRNA sequences fromthe SILVA database [28] however given that little is knownabout this domain it is unlikely that the archaeal diversitywasfully covered [43]

The main physicochemical differences between soils ofthe native forest and the oil palm-cultivated area were thelevels of phosphorous potassium calcium organic matterand total carbon (Table 2) Correlations between phylogenyand physicochemical parameters were evaluated using PCAand NMDS however no clear clusters or correlations wereobserved (results not shown)

In a study of the long-term effects of inorganic fertilizerson microbial communities Zhong and Cai [44] reportedthat phosphorus application indirectly altered microbial

Archaea 9

Candidatus NitrososphaeraUnclassified Soil Crenarchaeotic Group (SCG)

Unclassified South African Gold Mine Gp 1 (SAGMCG-1)Candidatus Nitrosotalea

Unclassified Terrestreial Miscellaneous Gp (TMEG)Unclassified Thaumarchaeota

Unclassified ThermoplasmatalesUnclassified Miscellaneous Crenarchaeotic Group

MethanosarcinaRice Cluster I

95 confidence intervals

q-v

alue

(cor

rect

ed)

Cultivated areaNative forest

531e minus 10

420 6 8minus4minus6minus8 minus2minus10

Difference between proportions ()38800

Proportion ()

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

Figure 7 Archaeal genera that differ significantly (119902 lt 005) in relative abundance between native forest soil (orange) and soil under oil palmcultivation (blue) For the purpose of statistical analysis performed using STAMP software the sum of the percentages of archaea classifiedat the genus level in both soils is 100 Differences between proportions of each genus are shown with 95 confidence intervals Unclassifiedsequences were used to calculate frequency profiles but are not displayed

parameters in soils and increased crop yields by elevating thelevels of organic matter Wessen and collaborators used real-time PCR quantification to show the impacts of fertilizer useon several bacterial phyla and the archaeal phylum Crenar-chaeota (Thaumarchaeota Group I1c after reanalysis usingthe SILVAdatabase 119) [45]They reported that bacterial andarchaeal classes respond differently to fertilization primarilybecause of changes in certain soil parameters such as pHtotal nitrogen and the carbon nitrogen ratio In our studylevels of total carbon and organic matter were higher in thenative forest than in the oil palm-cultivated area which mayhave contributed to higher archaeal diversity Although thereare no studies on the effect of total soil carbon on archaealcommunities there are studies on the bacterial communitiesin Amazonian soils The work of Edgar et al [33] describeddistinct soil bacterial communities under different vegetationtypes The authors found that bacterial diversity correlatedwith total organic carbon and total nitrogen content [46]Furthermore in a study of bacterial diversity and microbialbiomass in forest pasture and fallow soils in the Amazonbasin microbial biomass was highest in pasture soils whichalso had 30ndash47 higher carbon levels than the other sitesover the course of a year [47]

Several studies have also reported alterations in thecomposition of archaeal [48] bacterial [47] and fungal [49]communities associated with land use changes A studyof archaeal communities in Amazonian anthrosols undervarious land uses (agriculture pasture and secondary forest)and cultivation practices (eg eggplant banana citrus andmanioc) [10] reported that agriculture negatively affectedarchaeal community diversity Results of ribosomal inter-genic spacer analysis showed that soil type is also responsiblefor changes in archaeal communities in soils under differenttypes of land use (native grassland native forest Eucalyptus

and Acacia plantations and soybean and watermelon fields)[50]

The Venn diagram showed that the native forest soil has agreater number of unique sequences (Figure 2) This was notsurprising due to the greater archaeal richness and diversityobserved in the native forest soil (Table 2) The number ofunique OTUs found in the oil palm-cultivated area is almostthe same as that of shared OTUs with the native forest areaAlthough oil palm cultivation decreased archaeal diversity inthe soil there is still a core group of archaea OTUs sharedwith the native forest suggesting that these archaea havecertain characteristics that allow them to persist after thechange in land use It cannot be ruled out that these archaeaare metabolically inactive The smaller number of uniquesequences in the oil palm-cultivated area may be related to alower complexity of the vegetation (ie oil palmmonocultureversus great plant diversity in the forest) since plant diversitycould affect these communities [51]

In our study all OTUs identified belong to the domainArchaea and were classified into two phyla Euryarchaeotaand Thaumarchaeota It is important to mention that thedatabases were only recently updated to recognize the phy-lumThaumarchaeota therefore previous studies on Archaeacould not recognize this phylum as proposed by Brochier-Armanet et al [11] The creation of this novel phylum wassupported by a study comparing the genomes of two marineammonia-oxidizing archaea (AOAs) that identified a core setof informational genes specific to this group [52] Thereforeto allow comparisons with other studies we reanalyzed theirdata whenever possible updating the taxonomic classifica-tion to include the phylumThaumarchaeota

Thework ofChao and Shen [35] focused on the impacts ofland use (ie agricultural systems of indigenous people andcattle pasture) on archaeal communities by PCR (amoA gene)

10 Archaea

and denaturing gradient gel electrophoresis analysis of differ-ent Western Amazon soils The databases at the time did notconsiderThaumarchaeota in their classification however ourreanalysis of their dataset using an updated database showedthat Euryarchaeota and Thaumarchaeota were present intheir samples along with Crenarchaeota (Thermoprotei) Inaddition changes were observed in the structure of soilarchaeal communities when comparing the cultivated areaswith forest areas that had been converted to pasture

In our study differences in the percentages of Thau-marchaeota and Euryarchaeota found in the soil sampleswere not striking Thaumarchaeota were predominant in thenative forest (58) whereas Euryarchaeota were predom-inant in the oil palm-cultivated area with 54 (Figure 3)This result is consistent with other comparative studies onarchaeal communities in different types of Brazilian soils (ieAmazonian anthrosols Caatinga and Cerrado) [10 53 54]and a study of different tropical forest soil types that reportedThaumarchaeota as the predominant phylum [44]

The phylumThaumarchaeota comprises all known AOAsand is directly involved in nitrogen metabolism Ammonia isused as an energy source by Group I1a found predominantlyin aquatic systems [52] and Group I1b found in soils [55]Thaumarchaeota Group I1c is primarily found in acidic soilsbut the ecological roles of these archaea are still unknown [5657]

The literature presents conflicting data about the pro-portion of AOAsAOBs soils In some studies AOAs arepredominant [58] while in others AOBs are more abundant[59] However several studies found a predominance of thearchaeal amoA gene over the bacterial amoA gene in soils[60] AOAs are frequently detected in soils their abundancerange from 05 to 10 of the prokaryotic community andtheir abundance is strongly influenced by physicochemicalconditions as well as the organic matter content [61 62] Inacidic soils AOAs appear to have a more important role inammonia oxidation than AOBs [57]

The Soil Crenarchaeotic Group a class within Thaumar-chaeota according to the SILVA database classification wasdetected in both soil samples but a higher representationwas observed in native forest soil This class was discoveredin boreal forest soil [63] Candidatus Nitrososphaera whichbelongs to the Soil Crenarchaeotic Group class is a memberof Thaumarchaeota Group I1b it is an AOA that usesammonia or urea as an energy source [64 65]

On the other hand members of the phylum Eur-yarchaeota are directly involved in carbon metabolismwith several classes of methanogens and methane-oxidizingorganisms [66ndash68] Navarrete et al [48] studied the impactof converting Amazon native forest to pasture and reportedchanges not only to the composition and abundance of soilmicrobial communities but also to a decrease in the diversityof gene function

Within the phylum Euryarchaeota Methanomicrobiaand Thermoplasmata were the main classes detected in oursamples along with a few representatives of Methanobac-teria and Halobacteria Members of Methanobacteria andMethanomicrobia are methanogens [69] Methanomicrobiawere present in both native forest soil and oil palm-cultivated

soil with larger representation in the oil palm-cultivatedarea Within Methanomicrobia the genera Rice Cluster IMethanosarcina andMethanocellawere present in both sam-ples (Figure 6(b)) These genera are closely related and foundin rice fields that produce methane [70] The order Metha-nomicrobiales was detected only in the native forest soilwhich can probably be explained by the relatively high levelsof carbon and organic matter This archaeal order is respon-sible for the final step of methane production in the carboncycle by anaerobic degradation of organic matter [66 71]However the contribution of the Methanomicrobia to thecarbon cycle in the oil palm-cultivated soils could not beassessed since there are no data on the methane productionin the studied areas

Among the Thaumarchaeota sequences used to generatethe phylogenetic tree that were unclassified at the genuslevel several OTUs formed different clusters associated withtheMiscellaneous Crenarchaeotic Group (Figure 5(a))Thesesequences were from both soil samples indicating that theunclassified Thaumarchaeota in the native forest and the oilpalm-cultivated area are phylogenetically related

Most of the sequences remained unclassified at thegenus level for both Thaumarchaeota and Euryarchaeota(Figures 6(a) and 6(b)) This result likely reflects the fact thatmost taxonomic databases do not have many representativesof Archaea especially belonging to the phylum Thaumar-chaeota thus few classifications of new sequences reach thegenus level Several genera were identified as Candidatusaccording to the International Code of Nomenclature ofBacteria indicating that characterization of the culturedprokaryote is incomplete [72]

The proportions of only four groups classified at thegenus level differed significantly between native forest and oilpalm-cultivated soils (Methanosarcina and Rice Cluster I inthe phylum Euryarchaeota and Candidatus NitrososphaeraandCandidatusNitrosotalea in the phylumThaumarchaeota)(Figure 7) With respect to the methanogens sequencesrelated to Methanosarcina were slightly more abundant insoil under oil palm cultivation (Figure 7) whereas sequencesbelonging to the Rice Cluster I were more abundant in nativeforest soil

In the Thaumarchaeota phylum Candidatus Nitrosos-phaera an AOA member of group I1b [65] was detectedin both samples but had a much higher representation innative forest (native forest 182 oil palm-cultivated area103) (Figure 7) In a comparative study of the archaealcommunities in Amazonian dark earth (ldquoTerra Pretardquo)and adjacent soils Candidatus Nitrososphaera was themain archaeal group found in both soils [10] In contrastCandidatus Nitrosotalea another AOA was more abundantin the oil palm-cultivated area (100) than in native forestsoil (44) in our study (Figure 7) Lehtovirta-Morley et al[73] cultivated the obligate acidophilic ammonia oxidizerCandidatus Nitrosotalea devanaterra from a nitrifying acidicsoil Later the same group characterized two strains ofacidophilic AOA belonging to this genus from a Chinesepaddy field and a Scottish agricultural soil [74] In our studythe greater representation of Candidatus Nitrososphaeraand Candidatus Nitrosotalea in native forest and oil

Archaea 11

palm-cultivated soils respectively can be explained by thelow soil pH (native forest pH 550 oil palm-cultivated areapH 483) since in basic soils the activity and growth ofthese archaea are diminished by the exponential decrease innitrite [6 75] In addition soil under oil palm cultivationis fertilized with nitrogen which may explain the higherlevel of Candidatus Nitrosotalea in this area compared withthat of native forest soil Finally although the cultivatedmembers of the genus Ca Nitrososphaera grow betteron neutral pH they have been reported to grow on pHvalues ranging from 60 to 85 In addition a recent workpublished by Wang et al (2014) [76] described active AOAsfrom an acidic soil (pH 492) which are closely related toNitrososphaera known as Nitrososphaera-like organismsby molecular phylogenetic methods Most of our sequenceswere classified as Nitrosotalea-like which are sequencesbelonging to the group I1a This group is typically foundin aquatic environments such as lake sediments or marineenvironments A similar ubiquitous distribution in soils isfound for the group I1b which are not restricted to a specificsoil pH condition

5 Conclusions

Although archaea are known to be ubiquitous there is stillmuch to be learned about their diversity biological rolesand metabolic capabilities This work is the first to comparearchaeal communities associated with the Amazon forest soilwith those of soil under oil palm cultivation

Archaeal diversity was found to be lower in oil palm-cultivated soil than in the native forest soil Only about 30of the OTUs were found in the oil palm soil in comparisonto the native forest However two main phyla were detectedin both samples Thaumarchaeota and Euryarchaeota with apredominance ofThaumarchaeota in native forest soil Theseresults support previous studies in other Brazilian biomenative soils (eg Caatinga Cerrado) as well as soil underdifferent types of land use (eg pasture and agricultural) Itis also important to mention that there are also groups ofuncertain classification associated with the Crenarchaeotaphylum

Although knowledge about Archaea has increased dra-matically in the last few decades little is known aboutthe biology of many archaeal groups and the taxonomyof Archaea is still a work in progress Thus it is notsurprising thatmany sequences identified in thiswork remainunclassified sometimes even at higher taxonomic levels Itis not clear whether the relatively small overall differencebetween soil archaeal communities of the Amazon forest andoil palm-cultivated land is a consequence of the current stateof archaeal taxonomy or whether it indicates that archaealdiversity is lower than bacterial diversity [77]

It is also important to consider that with land usechange due to agricultural practices the microstructure ofthe soil is broken down which can reduce the natural habitatfragmentation allowing potential interactions among theresident microbiota that otherwise would not happen [7879]

A phylogenetic tree of unclassified Thaumarchaeota 16SrRNA gene sequences was useful in exploring relationshipsamong these sequences When taxonomic classification ofsequences was possible it revealed the presence of OTUsfrom genera known to be involved in the metabolism ofcarbon and nitrogen (methanogens Methanocella and Meth-anosarcina AOACandidatusNitrososphaera andCandidatusNitrosotalea) hinting at possible roles for these archaealcommunities in Amazon soils

More studies are needed to elucidate these roles Cultur-ing some of these archaea in the laboratory would allow theirmetabolism and physiology to be studied in detail allowingfor a better understanding of the roles of Archaea in the bio-geochemical cycles

Conflict of Interests

The authors declare that there is no conflict of interestsregarding the publication of this paper

Acknowledgments

This work was supported by grants from FINEP CNPqCAPES and FAP-DF The authors thank Dr Boari for helpwith sample collection logistics Dr Betania Ferraz Quirinowas supported by the Cientista Visitante program fromEmbrapa

References

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[2] IBGE ldquoMapa de Biomas do Brasil primeira aproximacaordquo Riode Janeiro Brazil 2004 httpwwwibgegovbrhome

[3] C Azevedo-Ramos ldquoSustainable development and challengingdeforestation in the Brazilian Amazon the good the badand the uglyrdquo in Proceedings of the Symposium Our CommonGround Innovations in Land Use Decision-Making vol 59 p 5Vancouver Canada May 2007

[4] C A Francis K J Roberts J M Beman A E Santoro and B BOakley ldquoUbiquity and diversity of ammonia-oxidizing archaeainwater columns and sediments of the oceanrdquoProceedings of theNational Academy of Sciences of theUnited States of America vol102 no 41 pp 14683ndash14688 2005

[5] A H Treusch S Leininger A Kietzin S C Schuster H-PKlenk and C Schleper ldquoNovel genes for nitrite reductase andAmo-related proteins indicate a role of uncultivatedmesophiliccrenarchaeota in nitrogen cyclingrdquo Environmental Microbiol-ogy vol 7 no 12 pp 1985ndash1995 2005

[6] G W Nicol S Leininger C Schleper and J I Prosser ldquoTheinfluence of soil pH on the diversity abundance and tran-scriptional activity of ammonia oxidizing archaea and bacteriardquoEnvironmentalMicrobiology vol 10 no 11 pp 2966ndash2978 2008

[7] P Offre A Spang and C Schleper ldquoArchaea in biogeochemicalcyclesrdquo Annual Review of Microbiology vol 67 pp 437ndash4572013

[8] MAPA Anuario Estatıstico da Agroenergia 2012 StatisticalYearbook of Agroenergy 2013

[9] J C Bergmann D D Tupinamba O Y A Costa J R MAlmeida C C Barreto andB FQuirino ldquoBiodiesel production

12 Archaea

in Brazil and alternative biomass feedstocksrdquo Renewable ampSustainable Energy Reviews vol 21 pp 411ndash420 2013

[10] R G Taketani and S M Tsai ldquoThe influence of different landuses on the structure of archaeal communities in Amazoniananthrosols based on 16S rRNA and amoA genesrdquo MicrobialEcology vol 59 no 4 pp 734ndash743 2010

[11] C Brochier-Armanet B Boussau S Gribaldo and P ForterreldquoMesophilic crenarchaeota proposal for a third archaeal phy-lum theThaumarchaeotardquoNature Reviews Microbiology vol 6no 3 pp 245ndash252 2008

[12] S M Barns C F Delwiche J D Palmer and N R PaceldquoPerspectives on archaeal diversity thermophily and mono-phyly from environmental rRNA sequencesrdquo Proceedings of theNational Academy of Sciences of the United States of Americavol 93 no 17 pp 9188ndash9193 1996

[13] J G Elkins M Podar D E Graham et al ldquoA korarchaealgenome reveals insights into the evolution of the Archaeardquo Pro-ceedings of the National Academy of Sciences of the United Statesof America vol 105 no 23 pp 8102ndash8107 2008

[14] H Huber M J Hohn R Rachel T Fuchs V C Wimmer andK O Stetter ldquoA new phylum of Archaea represented by ananosized hyperthermophilic symbiontrdquo Nature vol 417 no6884 pp 63ndash67 2002

[15] T Nunoura H Oida M Nakaseama et al ldquoArchaeal diversityand distribution along thermal and geochemical gradients inhydrothermal sediments at the Yonaguni Knoll IV hydrother-mal field in the SouthernOkinawa TroughrdquoApplied and Enviro-nmental Microbiology vol 76 no 4 pp 1198ndash1211 2010

[16] C Rinke P Schwientek A Sczyrba et al ldquoInsights into thephylogeny and coding potential of microbial dark matterrdquoNature vol 499 no 7459 pp 431ndash437 2013

[17] A Spang J H Saw S L Joslashrgensen et al ldquoComplex archaea thatbridge the gap between prokaryotes and eukaryotesrdquo Naturevol 521 no 7551 pp 173ndash179 2015

[18] S Bagchi S E Vlaeminck L A Sauder et al ldquoTemporal andspatial stability of ammonia-oxidizing archaea and bacteria inaquariumbiofiltersrdquo PLoSONE vol 9 no 12 Article ID e1135152014

[19] W Qin S A Amin W Martens-Habbena et al ldquoMarineammonia-oxidizing archaeal isolates display obligate mixotro-phy and wide ecotypic variationrdquo Proceedings of the NationalAcademy of Sciences of the United States of America vol 111 no34 pp 12504ndash12509 2014

[20] M Coci N Odermatt M M Salcher J Pernthaler and GCorno ldquoEcology and distribution ofThaumarchaea in the deephypolimnion of Lake Maggiorerdquo Archaea vol 2015 Article ID590434 11 pages 2015

[21] J G Ferry ldquoHow tomake a living by exhalingmethanerdquoAnnualReview of Microbiology vol 64 pp 453ndash473 2010

[22] A Meyer A Focks V Radl G Welzl I Schoning and MSchloter ldquoInfluence of land use intensity on the diversity ofammonia oxidizing bacteria and archaea in soils from grasslandecosystemsrdquoMicrobial Ecology vol 67 no 1 pp 161ndash166 2014

[23] V De Gannes G Eudoxie and W J Hickey ldquoImpacts ofedaphic factors on communities of ammonia-oxidizing archaeaammonia-oxidizing bacteria and nitrification in tropical soilsrdquoPLoS ONE vol 9 no 2 Article ID e89568 2014

[24] D S Ribeiro F C J da Silva and TN Carvalho ldquoSobrevivenciade seis especies florestais em uma area explorada seletivamenteno municıpio de Moju Parardquo CERNE vol 9 no 2 pp 153ndash1632003

[25] D HM Costa C A P Ferreira J NM Silva J D C A Lopesand J O P D Carvalho ldquoPotencial madeireiro de floresta densano municıpio de Moju Estado do Parardquo Documentos 1998

[26] C M Almeida S F Lima C V Martins-Da-Silva and JI Gomes ldquoCaracterizacao morfologica e anatomica de dezespecies de leguminosae ocorrentes em uma floresta tropicalumida localizada no municıpio de Moju Estado do Parardquo inA Silvicultura na Amazonia Oriental Contribuicoes do ProjetoEmbrapa-DFID J NMC Silva JO P deCarvalho and J AGYared Eds pp 19ndash54 Embrapa Amazonia Oriental (CPATU)Belem Brazil 2001

[27] K Smalla N Cresswell L C Mendoncahagler A Woltersand J D Vanelsas ldquoRapid extraction protocol from soil forpolymerase chain reaction-mediated amplificationrdquo Journal ofApplied Bacteriology vol 74 pp 78ndash85 1993

[28] S Gantner A F Andersson L Alonso-Saez and S Bertils-son ldquoNovel primers for 16S rRNA-based archaeal communityanalyses in environmental samplesrdquo Journal of MicrobiologicalMethods vol 84 no 1 pp 12ndash18 2011

[29] P D Schloss S L Westcott T Ryabin et al ldquoIntroducingMOTHUR open-source platform-independent community-supported software for describing and comparing microbialcommunitiesrdquoApplied and EnvironmentalMicrobiology vol 75no 23 pp 7537ndash7541 2009

[30] PD SchlossDGevers and S LWestcott ldquoReducing the effectsof PCR amplification and sequencing artifacts on 16S rRNA-based studiesrdquo PLoS ONE vol 6 no 12 Article ID e27310 2011

[31] CQuince A Lanzen T P Curtis et al ldquoAccurate determinationof microbial diversity from 454 pyrosequencing datardquo NatureMethods vol 6 no 9 pp 639ndash641 2009

[32] P Yilmaz L W Parfrey P Yarza et al ldquoThe SILVA and lsquoall-species Living Tree Project (LTP)rsquo taxonomic frameworksrdquoNucleic Acids Research vol 42 no 1 pp D643ndashD648 2014

[33] R C Edgar B J Haas J C Clemente C Quince and RKnight ldquoUCHIME improves sensitivity and speed of chimeradetectionrdquo Bioinformatics vol 27 no 16 pp 2194ndash2200 2011

[34] A Chao and T-J Shen ldquoNonparametric estimation of Shan-nonrsquos index of diversity when there are unseen species insamplerdquo Environmental and Ecological Statistics vol 10 no 4pp 429ndash443 2003

[35] A Chao and T-J Shen Program SPADE (Species Prediction andDiversity Estimation) 2010

[36] C L Shannon andWWeaverTheMathematicalTheory of Com-munication University of Illinois Press Urbana Ill USA 1949

[37] I J Good ldquoThe population frequencies of species and theestimation of population parametersrdquo Biometrika vol 40 pp237ndash264 1953

[38] K Tamura G Stecher D Peterson A Filipski and S KumarldquoMEGA6molecular evolutionary genetics analysis version 60rdquoMolecular Biology and Evolution vol 30 no 12 pp 2725ndash27292013

[39] DA BensonMCavanaugh K Clark et al ldquoGenBankrdquoNucleicAcids Research vol 41 no 1 pp D36ndashD42 2013

[40] D H Parks G W Tyson P Hugenholtz and R G BeikoldquoSTAMP statistical analysis of taxonomic and functional pro-filesrdquo Bioinformatics vol 30 no 21 pp 3123ndash3124 2014

[41] R G Newcombe ldquoInterval estimation for the differencebetween independent proportions comparison of elevenmeth-odsrdquo Statistics in Medicine vol 17 no 8 pp 873ndash890 1998

[42] J D Storey J E Taylor and D Siegmund ldquoStrong controlconservative point estimation and simultaneous conservative

Archaea 13

consistency of false discovery rates a unified approachrdquo Journalof the Royal Statistical Society Series B Statistical Methodologyvol 66 no 1 pp 187ndash205 2004

[43] G C Baker J J Smith and D A Cowan ldquoReview and re-analysis of domain-specific 16S primersrdquo Journal of Microbio-logical Methods vol 55 no 3 pp 541ndash555 2003

[44] W H Zhong and Z C Cai ldquoLong-term effects of inorganicfertilizers on microbial biomass and community functionaldiversity in a paddy soil derived from quaternary red clayrdquoApplied Soil Ecology vol 36 no 2-3 pp 84ndash91 2007

[45] EWessen S Hallin and L Philippot ldquoDifferential responses ofbacterial and archaeal groups at high taxonomical ranks to soilmanagementrdquo Soil Biology and Biochemistry vol 42 no 10 pp1759ndash1765 2010

[46] ZWuW Lin B Li et al ldquoTerminal restriction fragment lengthpolymorphism analysis of soil bacterial communities underdifferent vegetation types in subtropical area rdquo PLOS ONE vol10 no 6 Article ID e0129397 2015

[47] K Cenciani M R Lambais C C Cerri L C B De Azevedoand B J Feigl ldquoBacteria diversity and microbial biomass inforest pasture and fallow soils in the southwestern Amazonbasinrdquo Revista Brasileira de Ciencia do Solo vol 33 no 4 pp907ndash916 2009

[48] A A Navarrete R G Taketani L WMendes F de Souza Can-navan F M de Souza Moreira and S M Tsai ldquoLand-use sys-tems affect Archaeal community structure and functional diver-sity in western Amazon soilsrdquo Revista Brasileira de Ciencia doSolo vol 35 no 5 pp 1527ndash1540 2011

[49] E Lumini A Orgiazzi R Borriello P Bonfante and V Bian-ciotto ldquoDisclosing arbuscular mycorrhizal fungal biodiversityin soil through a land-use gradient using a pyrosequencingapproachrdquo Environmental Microbiology vol 12 no 8 pp 2165ndash2179 2010

[50] M Lupatini R J S Jacques Z I Antoniolli A K A SuleimanR R Fulthorpe and L F W Roesch ldquoLand-use change andsoil type are drivers of fungal and archaeal communities in thePampa biomerdquo World Journal of Microbiology and Biotechnol-ogy vol 29 no 2 pp 223ndash233 2013

[51] E G Lamb N Kennedy and S D Siciliano ldquoEffects of plantspecies richness and evenness on soil microbial communitydiversity and functionrdquo Plant and Soil vol 338 no 1 pp 483ndash495 2011

[52] A Spang R Hatzenpichler C Brochier-Armanet et al ldquoDis-tinct gene set in two different lineages of ammonia-oxidizingarchaea supports the phylum Thaumarchaeotardquo Trends inMicrobiology vol 18 no 8 pp 331ndash340 2010

[53] R G Pacchioni F M Carvalho C E Thompson et al ldquoTax-onomic and functional profiles of soil samples from Atlanticforest and Caatinga biomes in northeastern Brazilrdquo Microbio-logyOpen vol 3 no 3 pp 299ndash315 2014

[54] E Catao A P Castro C C Barreto R H Kruger and C MKyaw ldquoDiversity of Archaea in Brazilian savanna soilsrdquoArchivesof Microbiology vol 195 no 7 pp 507ndash512 2013

[55] M Pester T Rattei S Flechl et al ldquoAmoA-based consensusphylogeny of ammonia-oxidizing archaea and deep sequencingof amoA genes from soils of four different geographic regionsrdquoEnvironmental Microbiology vol 14 no 2 pp 525ndash539 2012

[56] J-Z He H-W Hu and L-M Zhang ldquoCurrent insights into theautotrophic thaumarchaeal ammonia oxidation in acidic soilsrdquoSoil Biology amp Biochemistry vol 55 pp 146ndash154 2012

[57] L-M Zhang H-W Hu J-P Shen and J-Z He ldquoAmmonia-oxidizing archaea have more important role than ammonia-oxidizing bacteria in ammonia oxidation of strongly acidicsoilsrdquo ISME Journal vol 6 no 5 pp 1032ndash1045 2012

[58] S Leininger T Urich M Schloter et al ldquoArchaea predominateamong ammonia-oxidizing prokaryotes in soilsrdquo Nature vol442 no 7104 pp 806ndash809 2006

[59] N C Banning L D Maccarone L M Fisk and D VMurphy ldquoAmmonia-oxidising bacteria not archaea dominatenitrification activity in semi-arid agricultural soilrdquo ScientificReports vol 5 Article ID 11146 2015

[60] M Herrmann A Hadrich and K Kusel ldquoPredominance ofthaumarchaeal ammonia oxidizer abundance and transcrip-tional activity in an acidic fenrdquoEnvironmentalMicrobiology vol14 no 11 pp 3013ndash3025 2012

[61] K Zhalnina P Dorr de Quadros F O Camargo and E WTriplett ldquoDrivers of archaeal ammonia-oxidizing communitiesin soilrdquo Frontiers in Microbiology vol 3 article 210 Article IDArticle 210 2012

[62] J Hong J Cho and Y Hong ldquoEnvironmental variables shapingthe ecological niche of Thaumarchaeota in soil direct andindirect causal effectsrdquo PLoS ONE vol 10 no 8 Article IDe0133763 2015

[63] G Jurgens K Lindstrom and A Saano ldquoNovel group withinthe kingdom Crenarchaeota from boreal forest soilrdquo Appliedand Environmental Microbiology vol 63 no 2 pp 803ndash8051997

[64] M Tourna M Stieglmeier A Spang et al ldquoNitrososphaeraviennensis an ammonia oxidizing archaeon from soilrdquoProceed-ings of the National Academy of Sciences of the United States ofAmerica vol 108 no 20 pp 8420ndash8425 2011

[65] K V Zhalnina R Dias M T Leonard et al ldquoGenome sequenceof Candidatus Nitrososphaera evergladensis from group I1benriched from everglades soil reveals novel genomic features ofthe ammonia-oxidizing archaeardquoPLoSONE vol 9 no 7 ArticleID e101648 2014

[66] I Anderson L E Ulrich B Lupa et al ldquoGenomic char-acterization of methanomicrobiales reveals three classes ofmethanogensrdquo PLoS ONE vol 4 no 6 Article ID e5797 2009

[67] J E Galagan C Nusbaum A Roy et al ldquoThe genome of Macetivorans reveals extensive metabolic and physiological diver-sityrdquo Genome Research vol 12 no 4 pp 532ndash542 2002

[68] C Wrede S Kokoschka A Dreier C Heller J Reitner and MHoppert ldquoDeposition of biogenic iron minerals in a methaneoxidizing microbial matrdquo Archaea vol 2013 Article ID 1029728 pages 2013

[69] G Garrity Bergeyrsquo Manual of Systematic Bacteriology 721 2001[70] S Sakai Y Takaki S Shimamura et al ldquoGenome sequence

of a mesophilic hydrogenotrophic methanogen Methanocellapaludicola the first cultivated representative of the orderMethanocellalesrdquo PLoS ONE vol 6 no 7 Article ID e228982011

[71] E Bapteste C Brochier and Y Boucher ldquoHigher-level clas-sification of the Archaea evolution of methanogenesis andmethanogensrdquo Archaea vol 1 no 5 pp 353ndash363 2005

[72] R G E Murray and E Stackebrandt ldquoTaxonomic note imple-mentation of the provisional status Candidatus for incompletelydescribed procaryotesrdquo International Journal of Systematic Bac-teriology vol 45 no 1 pp 186ndash187 1995

[73] L E Lehtovirta-Morley K Stoecker A Vilcinskas J I Prosserand G W Nicol ldquoCultivation of an obligate acidophilic ammo-nia oxidizer from a nitrifying acid soilrdquo Proceedings of the

14 Archaea

National Academy of Sciences of the United States of Americavol 108 no 38 pp 15892ndash15897 2011

[74] L E Lehtovirta-Morley C Ge J Ross H Yao G W Nicol andJ I Prosser ldquoCharacterisation of terrestrial acidophilic archaealammonia oxidisers and their inhibition and stimulation byorganic compoundsrdquo FEMSMicrobiology Ecology vol 89 no 3pp 542ndash552 2015

[75] M J Frijlink T Abee H J Laanbroek W de Boer and W NKonings ldquoThe bioenergetics of ammonia and hydroxylamineoxidation in Nitrosomonas europaea at acid and alkaline pHrdquoArchives of Microbiology vol 157 no 2 pp 194ndash199 1992

[76] B Wang Y Zheng R Huang et al ldquoActive ammonia oxidizersin an acidic soil are phylogenetically closely related to neu-trophilic archaeonrdquo Applied and Environmental Microbiologyvol 80 no 5 pp 1684ndash1691 2014

[77] J Y Aller and P F Kemp ldquoAre Archaea inherently less diversethan Bacteria in the same environmentsrdquo FEMS MicrobiologyEcology vol 65 no 1 pp 74ndash87 2008

[78] K Potthast U Hamer and FMakeschin ldquoLand-use change in atropical mountain rainforest region of southern Ecuador affectssoilmicroorganisms and nutrient cyclingrdquoBiogeochemistry vol111 no 1ndash3 pp 151ndash167 2012

[79] J I Prosser and G W Nicol ldquoArchaeal and bacterial ammonia-oxidisers in soil the quest for niche specialisation and differen-tiationrdquoTrends inMicrobiology vol 20 no 11 pp 523ndash531 2012

Submit your manuscripts athttpwwwhindawicom

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Anatomy Research International

PeptidesInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporation httpwwwhindawicom

International Journal of

Volume 2014

Zoology

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Molecular Biology International

GenomicsInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioinformaticsAdvances in

Marine BiologyJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Signal TransductionJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioMed Research International

Evolutionary BiologyInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Biochemistry Research International

ArchaeaHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Genetics Research International

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Advances in

Virolog y

Hindawi Publishing Corporationhttpwwwhindawicom

Nucleic AcidsJournal of

Volume 2014

Stem CellsInternational

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

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Enzyme Research

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

International Journal of

Microbiology

Archaea 3

(a) (b)

Figure 1 (a) Native forest and (b) oil palm-cultivated sites

CAG-31015840) and 1000R (51015840-GGC CAT GCA CYW CYT CTC-31015840) [28] Archaea 16S rRNA genes were amplified yielding660 bp amplicons Adapters used as priming sites for bothamplification and sequencing (454 Life Sciences BranfordCT USA) were ligated to the 51015840 end of the primer sequencesEach 20120583L PCR reaction contained 10ndash30 ng total DNA 1xreaction buffer 4120583MdNTP 10 120583Mof each primer 200120583gmLbovine serum albumin 05U KAPA2G Robust HotStartpolymerase andMilli-Qwater Amplification was performedin anApplied BiosystemsGeneAmpPCRSystem9700Ther-mal Cycler (Applied Biosystems Foster City CA USA) usingthe following program 2min at 98∘C followed by 30 cyclesof 30 seconds at 95∘C 30 seconds at 57∘C and 1 minute and30 seconds at 72∘C and a final step consisting of 7 minutes at72∘C To minimize PCR bias at least four PCR amplificationreactions per sample were pooled before purification usingthe GeneJET PCR Purification Kit (Thermo Fisher ScientificWaltham MA USA) After quantification using a Qubitfluorometer (Invitrogen Carlsbad CA USA) NanoDrop1000 Spectrophotometer (Thermo Fisher Scientific) and 2agarose gel with ethidium bromide DNA samples were sentfor pyrosequencing (GSFLXTitaniumPlatformatMacrogenSouth Korea) A total of 592145 reads greater than 500 bpwere obtained in three-quarters of a plate 49314 reads fromthe oil palm-cultivated soil and 542831 reads from the nativeforest soil

23 Data Analysis All 16S rRNA gene pyrosequencing readswere analyzed using the original standard flowgram format(SFF) output file from the sequencer in MOTHUR version1333 [29] for the removal of short sequences sequences witherrors low-quality sequences chimeras and possible con-taminants After this removal a total of 39111 sequences foroil palm-cultivated area and 436532 sequences for native for-est were obtained Further analysis was performed after nor-malizing the number of sequences per sample to the lowest

number of high-quality sequences (ie 39111 sequences in theoil palm-cultivated area sample)

All 16S rRNA gene reads were analyzed using MOTHURfor taxonomic classification following the 454 standardoperating procedure available online (httpwwwmothurorgwiki454 SOP) [30] The PyroNoise algorithm [31]was used to denoise the flowgram file (removal of barcodesand quality filtering) The SILVA database (release 119) wasused for sequence alignment and clustering based on 97sequence similarity (003 genetic distance) using the nearestneighbor method [32] The UCHIME algorithm [33] wasused for detection and removal of chimeric reads thisfunction was implemented in MOTHUR and performedwithout a reference database The one-gap method inMOTHURwas used to calculate the pairwise distancematrixArchaeal taxonomic classifications of each representativeOTU were performed in MOTHUR using the SILVA data-base (release 119) clustered at 97 sequence similarity [32]This classificationwas used to estimate the relative abundanceof reads per genus Samples were randomly chosen forsubsampling and normalized to the lowest number ofsequence reads obtained Chao1 [34] abundance-basedcoverage estimator (ACE) [35] Shannon diversity index [36]and Goodrsquos coverage [37] were then used to estimatealpha diversity MOTHUR was also used to perform allstatistical analyses A phylogenetic tree was created usingall Thaumarchaeota OTUs from each sample that wereunclassified at the class level A new tree was created withsequences randomly chosen by MOTHUR that were deemedrepresentative of the entire set Data were analyzed using theneighbor-joining method with the Jukes-Cantor correctionand 1000 bootstrap replicates in MEGA6 [38] In additionOTUs with 3 dissimilarity were selected by MOTHUR toavoid redundancies in the phylogenetic tree The chosensequences were aligned with 16S rRNA gene sequencesfrom GenBank [39] representing the main Archaeagroups Methanosarcina baltica (AB973356) Methanoregula

4 Archaea

Table 1 Number of operational taxonomic units (OTUs) Goodrsquos coverage estimator OTU richness indices (Chao1 ACE) and InverseSimpson and Shannon diversity indices for archaeal communities in native forest soil and soil under oil palm cultivation

Sample OTUslowast Goodrsquos coverage () Chao1 ACE Inverse Simpson ShannonNative forest 821 9871 2124 4645 2645 422Oil palm-cultivated area 253 9972 424 674 1547 331lowastConsidering 97 sequence similarity in MOTHUR

formicica (NR112877) Methanocella paludicola (NR074192)Halococcus hamelinensis (LN651155) and Pyrococcus furiosus(U20163) from the phylum Euryarchaeota Pyrobaculumaerophilum (NR102764) from the phylum CrenarchaeotaCandidatus Nitrososphaera sp (FR773157) and CandidatusNitrososphaera gargensis (GU797786) fromThaumarchaeotaGroup I1b Nitrosopumilus maritimus (JQ346765) andCenarchaeum symbiosum (U51469) from ThaumarchaeotaGroup I1a uncultured acidic red soil AOA (FJ174727)uncultured trembling aspen archaeon (EF021427) and borealforest archaeon (X96688) from Thaumarchaeota GroupI1c and several uncultured organisms classified accordingto the SILVA database as Thaumarchaeota MiscellaneousCrenarchaeotic Group (FR745121 AM910782 JX984848KC510333 FJ485299 KC831395 FJ920714 FJ485307HM051127 HM051130 and HM051125) A 16S rRNA genesequence from Acidobacteria KBS 96 (FJ870384) was usedas an outgroup A second phylogenetic tree was createdusing only Euryarchaeota OTUs from each sample that wereunclassified at the class level with the same parameters andreference sequences

A Venn diagram was generated by grouping OTUs clus-tered at 97 similarity to show the number of OTUs uniqueto each area and those shared between samples

Differences in taxonomic distribution of archaea betweennative forest and the oil palm-cultivated area were evaluatedby Fisherrsquos exact test (119902 le 005) using Statistical Analysisof Metagenomic Profiles (STAMP) software [40] The confi-dence interval was estimated using the asymptotic method[41] and correction of the 119902 value was calculated usingStoreyrsquos false discovery rate (FDR) approach [42] A filterwas applied to show only the differences between proportionsabove 05

3 Results

Goodrsquos coverage estimate of the completeness of sampling(Table 1) was higher for the oil palm-cultivated area than forthe native forest

The number of archaeal OTUs was higher in the nativeforest soil (821 sequences) than in soil under oil palmcultivation (253 sequences) (Figure 2) with 130OTUs (138)shared between the two areas Archaeal species richness asassessed by abundance-based estimators Chao1 and ACEwas higher in the native forest than in the oil palm-cultivatedarea Similarly the Inverse Simpson and Shannon indicesindicated that archaeal species diversity was higher in nativeforest soil compared to soil under oil palm cultivation(Table 1)

Table 2 Physicochemical properties of native forest soil and soilunder oil palm cultivation

Native forest Oil palm-cultivated areapH(in water) 550 483

P (mgsdotdmminus3) 140 7070K (mgsdotdmminus3) 003 011Ca+2 (cmolsdotdmminus3) 140 033Mg+2 (cmolsdotdmminus3) 010 017Al+3 (cmolsdotdmminus3) 040 073H + Al (cmolsdotdmminus3) 620 727Na+ (cmolsdotdmminus3) 001 002Organic matter (gkg) 3100 1163Total C (gkg) 1800 677Total N (mgkg) NA 115NO3

minus1 (mgkg) NA 079NO2

minus1 (mgkg) NA 014NH4

+ (mgkg) NA 021Data not available

Soil physicochemical parameters differed between theareas (Table 2) with higher levels of phosphorous potassiumand total carbon in the oil palm-cultivated soil and higherlevels of calcium and organic matter in the native forest soil

In both areas only two archaeal phyla (Thaumarchaeotaand Euryarchaeota) were detected (Figure 3) Thaumar-chaeota were predominant in the native forest soil (58) andEuryarchaeota were predominant in the oil palm-cultivatedsoil (54)The percentage of unclassified archaea was similarin both soils (native forest 5 oil palm-cultivated area 6)

At the class level a total of 10 groups were detected inboth samples Halobacteria Methanobacteria Methanomi-crobia and Thermoplasmata in the phylum Euryarchaeotaand Marine Group I OPPD003 Soil Crenarchaeotic GroupSouth African Gold Mine Gp 1 Terrestrial Group andMiscellaneous Crenarchaeotic Group in the phylum Thau-marchaeota (Figures 4(a) and 4(b)) The most abundantclass present in native forest soil was the Soil Crenar-chaeotic Group representing 26 of all native forest OTUs(Figure 4(a)) and 45 of theThaumarchaeota sequences (notshown) followed by Methanomicrobia representing 15 ofall native forest OTUs (Figure 4(b)) and 42 of the Eur-yarchaeota sequences (not shown) The most abundant classpresent in the oil palm-cultivated area was Thermoplasmatarepresenting 21 of all cultivated area OTUs (Figure 4(b))and 39 of the Euryarchaeota sequences (not shown) fol-lowed by Methanomicrobia representing 16 of all culti-vated area OTUs (Figure 4(b)) and 30 of all Euryarchaeota

Archaea 5

Venn diagram at distance 003

123Cultivated area

691Native forest

130

Figure 2 Venn diagram depicting the number of shared and unique operational taxonomic units (OTUs) in native forest and oil palm-cultivated area

Thaumarchaeota

Euryarchaeota

Unclassified archaea

Cultivated areaNative forest

10 20 30 40 50 60 700()

Figure 3 Comparison of archaeal phyla present in soil samples from native Amazon forest (orange) and an area under oil palm cultivation(blue) The percentages of unclassified sequences are shown

sequences (not shown) The predominant class from thephylum Thaumarchaeota in the oil palm-cultivated areawas Terrestrial Group representing 14 of the total OTUs(Figure 4(a)) and 35 of all Thaumarchaeota sequences (notshown)

Because it is a newer phylum Thaumarchaeota have fewcultivated representatives compared with Euryarchaeota andfewer sequences in databases such as SILVA To determinehow the Thaumarchaeota sequences that could not be clas-sified at the genus level were related to each other a phylo-genetic tree was constructed with representative sequences(Figure 5(a)) One native forest OTU clustered with GroupI1b (NF6) and one native forest OTU (NF17) and two cul-tivated area OTUs (CA50 CA52) clustered with Group I1cThe remaining OTUs formed different clusters all associatedwith the Miscellaneous Crenarchaeotic Group Most unclas-sified Thaumarchaeota sequences from the native forest andoil palm-cultivated area grouped together although a fewunclassified OTUs from the cultivated area (eg CA51 toCA15 and CA24 to CA33) and native forest (eg NF21 toNF49 and NF5 to NF35) formed separate clusters

The unclassified sequences at genus level in the Euryar-chaeota phylum were all related to the orders Methano-cellales Methanosarcinales Methanomicrobiales Thermo-plasmatales Methanobacteriales and Halobacteriales(Figure 5(b)) The classified sequences at genus level wereMethanocella Methanosarcina Methanoculleus Methano-massiliicoccus Methanomethylophilus MethanobrevibacterMethanobacterium and Candidatus lainarchaeum(Figure 5(b))

The genus-level analysis showed that unclassified SoilCrenarchaeotic Group was predominant in the native forestaccounting for 21 of all native forest OTUs (Figure 6(a))In the oil palm-cultivated area the predominant genus wasUnclassified Terrestrial Group accounting for 14 of allcultivated area OTUs (Figure 6(b))

A total of 31 groups of OTUs were identified (onlymajor groups are shown) most of which are unclassi-fied The classified groups in the phylum Euryarchaeotaare Candidatus lainarchaeum Methanobacterium Methano-brevibacter Methanocella Rice Cluster I Methanoculleus

6 Archaea

Marine Group I

Miscellaneous Crenarchaeotic Group

Soil Crenarchaeotic Group

South African Gold Mine Gp 1

Terrestrial Group

Unclassified Thaumarchaeota

OPPD003

Cultivated areaNative forest

5 10 15 20 25 300()

(a)

Methanobacteria

Methanomicrobia

Halobacteria

Thermoplasmata

Unclassified Euryarchaeota

Cultivated areaNative forest

5 10 15 20 250()

(b)

Figure 4 Comparison of archaeal classes present in soil samples from native Amazon forest (orange) and an area under oil palm cultivation(blue) Classes within the phyla Thaumarchaeota (a) and Euryarchaeota (b) are shown as well as the percentage of unclassified sequences

Methanoregula Methanimicrococcus Methanosarcina Can-didatus Methanomethylophilus and Methanomassiliicoccus(Figure 6(b)) The classified groups in the phylum Thau-marchaeota are Candidatus Nitrososphaera and CandidatusNitrosotalea (Figure 6(a))

The analysis of the genera that differ significantly inrelative abundance between native forest soil and the oilpalm-cultivated area revealed that the greatest difference wasseen for Candidatus Nitrososphaera (Figure 7) This genuswas predominant in the native forest soil accounting for182 of the sequences but represented only 103 of thesequences from the oil palm-cultivated area UnclassifiedSoil Crenarchaeotic Group was also predominant in thenative forest (ie 388 of sequences in the native forest

and 312 of sequences in the oil palm-cultivated area)whereas unclassified South African Gold Mine Gp 1 waspredominant in the oil palm-cultivated area (ie 73 ofsequences in the native forest and 135 of sequences inthe oil palm-cultivated area) Candidatus Nitrosotalea repre-sented 44 of the native forest sequences and 100 of theoil palm-cultivated area sequences The relative abundanceof unclassified Terrestrial Miscellaneous Group unclassifiedThaumarchaeotaunclassifiedThermoplasmatalesunclassifiedMiscellaneous Crenarchaeotic Group Methanosarcina andRice Cluster I also differed significantly between native forestsoil and soils under oil palm cultivation but these differenceswere smaller

Archaea 7

FJ4853071| Uncultured archaeon clone

FJ4852991| Uncultured archaeon clone

FJ9027141| Uncultured archaeon clone

KC8313951| Uncultured archaeon clone

KC5103331| Uncultured archaeon clone

JX9848481| Uncultured archaeon clone

AM9107821| Uncultured archaeon clone

Uncultured archaeon acidic red soil-FJ174727

Ca Nitrososphaera gargensis-GU797786

Ca Nitrososphaeramdash(48 6)

Ca Nitrosotaleamdash(49 12)

Candidatus lainarchaeum (0 2)

Methanobacterium (2 3)

Methanomassillicoccus (6 2)

Methanocella (33 8)

Rice cluster I (43 25)

Methanosarcina baltica-AB973356

Methanocella paludicola-NR074192

Pyrobaculum aerophilum

-NR102764

Acidobacteria bacterium KBS 96-FJ870384

Ca Methanom

ethylophilus (12 0)

Ca Nitrososphaera sp-FR773157

Nitrosopumilus maritimus-JQ346765

Halococcus hamelinensis-LN651155

Pyrococcus furiosus-PFU20163

Cenarchaeum symbiosum-CSU51469

Methanoregula formicica-NR112877

FR7451211| Uncultured soil archaeon

Boreal forest soil archaeon-X96688

Trembling aspen archaeon-EF021427

MCG

MCG005

CA4099

99

7174

63

86

9687

9067

5696

6768 54

57

62

58

65

7559

62

63

65

64

52

64

64

1008654

91

99

999293

53798392

88

7274

5164

5858

CA7CA32

CA50NF17

NF6

CA52

T I1c

T I1b

T I1a

CA24CA6CA12

CA4

CA33

NF16

NF2

NF35

NF9

NF5NF3

6NF4

1NF3

2

NF1

2

NF4

NF1

NF3

0NF2

3

NF7CA3

CA39

CA37

CA46

CA44

CA2

CA42

CA27

Unclassified MCGmdash(16 4)

Unclassified Terrestrial Group (94 36)

Unclassified Marine Group I (2 2)

CA15

CA25

CA30

CA43

CA35

CA14

CA29

CA13

CA20

CA11

CA41

CA36

CA51

NF48

CA9NF4

0NF4

2CA21

CA22

NF38

CA45

CA16

CA47

NF31CA31NF28NF47NF27NF45CA18NF26

NF24NF33NF19

NF18NF37

NF34NF43NF15NF44NF13

NF22NF29NF11

CA49

CA1

CA19CA8

CA10

CA23CA38

CA54HM0511251| Uncultured

HM0511301| Uncultured archaeon clone

HM0511271| Uncultured archaeon clone

archaeon clone

NF46NF14

NF49NF25N

F10N

F39N

F20N

F21

NF8

NF3

CA48

CA34

CA5

CA28CA26CA17CA53

(E)

(B)

(C)

(a)

CA13

CA14

NF1

2

NF1

4M

ethan

ocell

a (3

8)Meth

anosarcina (3

8 4)

Methanoculleus (2 0)

Methanomassilicoccus (6 2)

Methanomassilicoccus (6 2)

Methanomassilicoccus (6 2)

Methanomethylophilus (6 2)

Methanomethylophilus (6 2)

Methanobrevibacter (1 0)

Methanobacterium

(2 3)

Methanobacterium

(2 3)

Methanobacterium

(2 3)

Methanobacterium

(2 3)

Methanobacterium

(2 3)

Methanosarci

na (38 4)

Methanosarcina (38 4)

Methanosarcina (38 4)

Methanosarcina (38 4)

NF16

CA4

CA15CA20

CA19

NF3NF1

Meth

anoc

ella (

33 8

)

Meth

anoc

ella p

aludi

cola-

NR 0741

92 1

Meth

anosa

rcina

baltic

a AB97

3356

1

Methanoregula formicica NR 112877 1

Halococcus hamelinensis LN651155 1

CA6N

F6NF8

NF19 NF5NF7NF20

Pyrobaculum aerophilum NR 102764 2

Ca Nitrososphaera gargensis G

U797786 1

Ca Nitrososphaera gargensis GU797786 1

Nitrosopumilus maritimus JQ346765 1Cenarchaeum symbiosum U51469 1Uncultured archaeon acidic red soil FJ174727 1

Boreal forest soil archaeon X96688 1

Trembling aspen archaeon EF021427 1

HM051125 1 Uncultured archaeon clone

HM051127 1 Uncultured archaeon clone

HM051130 1 Uncultured archaeon clone

FJ485307 1 Uncultured archaeon clone

FJ902714 1 Uncultured archaeon clone

KC831395 1 archaeon clone

FJ485299 1 archaeon clone

KC510333 1 archaeon clone

FR745121 1 archaeon clone

JX984848 1 archaeon clone

AM910782 1 archaeon clone

Acidobacteria bacterium KBS 96 FJ870384 1

Ca Nitrososphaera sp FR773157 1

CA7

Pyrococcus furiosus U20163 1

005

MCG

Halobacteriales

Cand

idat

us la

inar

chae

um (0

2)

Cand

idat

us la

inar

chae

um (0

2)

Thermoplasmatales

Methanomicrobiales

Methanosarcinalles

Methanocellales

T I1c

T I1a

T I1b

(C)

998785

58

100

100

100100

100

71 84 71

53

54

65

86

94

98

59100

100

78

10051

5477

63 73

99

100

8497100

78

100

100

76956970

100 10

010

0

637551

100

9999

9958

94

Methanobacteriales

(b)

Figure 5 Phylogenetic trees of (a) unclassified Thaumarchaeota 16S rRNA gene sequences from native forest soil and soil under oil palmcultivation and (b) 16S rRNA gene sequences classified as Euryarchaeota from native forest soil and soil under oil palm cultivation Theneighbor-joining method and Jukes-Cantor corrections were used with 1000 bootstrap replicates Bootstrap values above 50 are shownin the trees A representative number of operational taxonomic units (OTUs) were selected at 3 dissimilarity by MOTHUR to avoidredundancies in the phylogenetic trees Randomly chosen OTUs from native forest soil and from soil under oil palm cultivation were usedin the construction of the trees along with archaeal reference sequences (from GenBank) from the following groups Acidobacteria (B)Crenarchaeota (C) Euryarchaeota (E) Miscellaneous Crenarchaeotic Group (MCG) Thaumarchaeota Group I1a (T I1a) ThaumarchaeotaGroup I1b (T I1b) andThaumarchaeota Group I1c (T I1c)The AOAs are highlighted with the symbolX Sampling site is indicated by ldquoNFrdquo(native forest) or ldquoCArdquo (oil palm-cultivated area) followed by the number of the OTUs (NAxx or CAxx) OTU sequences classified to at leastthe class level are shown in bold followed by two numbers in parentheses indicating the number of sequences of that OTU detected in nativeforest soil and the number detected in the oil palm-cultivated area The scale bar represents the 5 estimated sequence divergence

8 Archaea

Unclassified Marine Group I

Unclassified Miscellaneous Crenarchaeotic Group

Unclassified Thaumarchaeota

Candidatus Nitrososphaera

Unclassified Soil Crenarchaeotic Group

Candidatus Nitrosotalea

Unclassified South African Gold Mine Gp 1

Unclassified Terrestrial Group

Cultivated areaNative forest

5 10 15 20 250()

(a)

Unclassified Thermoplasmatales

Unclassified Terrestrial Miscellaneous Group

Rice Cluster I

Methanocella

Methanosarcina

Unclassified Euryarchaeota

Unclassified Halobacteriales

Candidatus Methanomethylophilus

Unclassified Methanosarcinaceae

5 10 15 20 250()

Cultivated areaNative forest

(b)

Figure 6 Percentage of archaeal operational taxonomic units within the Thaumarchaeota (a) and Euryarchaeota (b) phyla classified at thegenus level for native forest soil (orange) and soil under oil palm cultivation (blue) The percentage of unclassified sequences at genus level isshown

4 Discussion

The Amazon forest is known for its macrospecies diversityhowever few studies have addressed its microbial diversityAmong the domains of life Archaea are undoubtedly the lesswell known

In this work almost 600000 archaeal 16S rRNA genesequences were obtained from native forest soil and anadjacent area under oil palm cultivation After processing(ie removal of short sequences sequences with errors low-quality sequences chimeras and possible contaminants) andnormalization 39111 sequences per area remained for anal-ysis This number of sequences provided sufficient coverageof archaeal diversity in both areas (Table 1)The primers used

were based on a large number of 16S rRNA sequences fromthe SILVA database [28] however given that little is knownabout this domain it is unlikely that the archaeal diversitywasfully covered [43]

The main physicochemical differences between soils ofthe native forest and the oil palm-cultivated area were thelevels of phosphorous potassium calcium organic matterand total carbon (Table 2) Correlations between phylogenyand physicochemical parameters were evaluated using PCAand NMDS however no clear clusters or correlations wereobserved (results not shown)

In a study of the long-term effects of inorganic fertilizerson microbial communities Zhong and Cai [44] reportedthat phosphorus application indirectly altered microbial

Archaea 9

Candidatus NitrososphaeraUnclassified Soil Crenarchaeotic Group (SCG)

Unclassified South African Gold Mine Gp 1 (SAGMCG-1)Candidatus Nitrosotalea

Unclassified Terrestreial Miscellaneous Gp (TMEG)Unclassified Thaumarchaeota

Unclassified ThermoplasmatalesUnclassified Miscellaneous Crenarchaeotic Group

MethanosarcinaRice Cluster I

95 confidence intervals

q-v

alue

(cor

rect

ed)

Cultivated areaNative forest

531e minus 10

420 6 8minus4minus6minus8 minus2minus10

Difference between proportions ()38800

Proportion ()

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

Figure 7 Archaeal genera that differ significantly (119902 lt 005) in relative abundance between native forest soil (orange) and soil under oil palmcultivation (blue) For the purpose of statistical analysis performed using STAMP software the sum of the percentages of archaea classifiedat the genus level in both soils is 100 Differences between proportions of each genus are shown with 95 confidence intervals Unclassifiedsequences were used to calculate frequency profiles but are not displayed

parameters in soils and increased crop yields by elevating thelevels of organic matter Wessen and collaborators used real-time PCR quantification to show the impacts of fertilizer useon several bacterial phyla and the archaeal phylum Crenar-chaeota (Thaumarchaeota Group I1c after reanalysis usingthe SILVAdatabase 119) [45]They reported that bacterial andarchaeal classes respond differently to fertilization primarilybecause of changes in certain soil parameters such as pHtotal nitrogen and the carbon nitrogen ratio In our studylevels of total carbon and organic matter were higher in thenative forest than in the oil palm-cultivated area which mayhave contributed to higher archaeal diversity Although thereare no studies on the effect of total soil carbon on archaealcommunities there are studies on the bacterial communitiesin Amazonian soils The work of Edgar et al [33] describeddistinct soil bacterial communities under different vegetationtypes The authors found that bacterial diversity correlatedwith total organic carbon and total nitrogen content [46]Furthermore in a study of bacterial diversity and microbialbiomass in forest pasture and fallow soils in the Amazonbasin microbial biomass was highest in pasture soils whichalso had 30ndash47 higher carbon levels than the other sitesover the course of a year [47]

Several studies have also reported alterations in thecomposition of archaeal [48] bacterial [47] and fungal [49]communities associated with land use changes A studyof archaeal communities in Amazonian anthrosols undervarious land uses (agriculture pasture and secondary forest)and cultivation practices (eg eggplant banana citrus andmanioc) [10] reported that agriculture negatively affectedarchaeal community diversity Results of ribosomal inter-genic spacer analysis showed that soil type is also responsiblefor changes in archaeal communities in soils under differenttypes of land use (native grassland native forest Eucalyptus

and Acacia plantations and soybean and watermelon fields)[50]

The Venn diagram showed that the native forest soil has agreater number of unique sequences (Figure 2) This was notsurprising due to the greater archaeal richness and diversityobserved in the native forest soil (Table 2) The number ofunique OTUs found in the oil palm-cultivated area is almostthe same as that of shared OTUs with the native forest areaAlthough oil palm cultivation decreased archaeal diversity inthe soil there is still a core group of archaea OTUs sharedwith the native forest suggesting that these archaea havecertain characteristics that allow them to persist after thechange in land use It cannot be ruled out that these archaeaare metabolically inactive The smaller number of uniquesequences in the oil palm-cultivated area may be related to alower complexity of the vegetation (ie oil palmmonocultureversus great plant diversity in the forest) since plant diversitycould affect these communities [51]

In our study all OTUs identified belong to the domainArchaea and were classified into two phyla Euryarchaeotaand Thaumarchaeota It is important to mention that thedatabases were only recently updated to recognize the phy-lumThaumarchaeota therefore previous studies on Archaeacould not recognize this phylum as proposed by Brochier-Armanet et al [11] The creation of this novel phylum wassupported by a study comparing the genomes of two marineammonia-oxidizing archaea (AOAs) that identified a core setof informational genes specific to this group [52] Thereforeto allow comparisons with other studies we reanalyzed theirdata whenever possible updating the taxonomic classifica-tion to include the phylumThaumarchaeota

Thework ofChao and Shen [35] focused on the impacts ofland use (ie agricultural systems of indigenous people andcattle pasture) on archaeal communities by PCR (amoA gene)

10 Archaea

and denaturing gradient gel electrophoresis analysis of differ-ent Western Amazon soils The databases at the time did notconsiderThaumarchaeota in their classification however ourreanalysis of their dataset using an updated database showedthat Euryarchaeota and Thaumarchaeota were present intheir samples along with Crenarchaeota (Thermoprotei) Inaddition changes were observed in the structure of soilarchaeal communities when comparing the cultivated areaswith forest areas that had been converted to pasture

In our study differences in the percentages of Thau-marchaeota and Euryarchaeota found in the soil sampleswere not striking Thaumarchaeota were predominant in thenative forest (58) whereas Euryarchaeota were predom-inant in the oil palm-cultivated area with 54 (Figure 3)This result is consistent with other comparative studies onarchaeal communities in different types of Brazilian soils (ieAmazonian anthrosols Caatinga and Cerrado) [10 53 54]and a study of different tropical forest soil types that reportedThaumarchaeota as the predominant phylum [44]

The phylumThaumarchaeota comprises all known AOAsand is directly involved in nitrogen metabolism Ammonia isused as an energy source by Group I1a found predominantlyin aquatic systems [52] and Group I1b found in soils [55]Thaumarchaeota Group I1c is primarily found in acidic soilsbut the ecological roles of these archaea are still unknown [5657]

The literature presents conflicting data about the pro-portion of AOAsAOBs soils In some studies AOAs arepredominant [58] while in others AOBs are more abundant[59] However several studies found a predominance of thearchaeal amoA gene over the bacterial amoA gene in soils[60] AOAs are frequently detected in soils their abundancerange from 05 to 10 of the prokaryotic community andtheir abundance is strongly influenced by physicochemicalconditions as well as the organic matter content [61 62] Inacidic soils AOAs appear to have a more important role inammonia oxidation than AOBs [57]

The Soil Crenarchaeotic Group a class within Thaumar-chaeota according to the SILVA database classification wasdetected in both soil samples but a higher representationwas observed in native forest soil This class was discoveredin boreal forest soil [63] Candidatus Nitrososphaera whichbelongs to the Soil Crenarchaeotic Group class is a memberof Thaumarchaeota Group I1b it is an AOA that usesammonia or urea as an energy source [64 65]

On the other hand members of the phylum Eur-yarchaeota are directly involved in carbon metabolismwith several classes of methanogens and methane-oxidizingorganisms [66ndash68] Navarrete et al [48] studied the impactof converting Amazon native forest to pasture and reportedchanges not only to the composition and abundance of soilmicrobial communities but also to a decrease in the diversityof gene function

Within the phylum Euryarchaeota Methanomicrobiaand Thermoplasmata were the main classes detected in oursamples along with a few representatives of Methanobac-teria and Halobacteria Members of Methanobacteria andMethanomicrobia are methanogens [69] Methanomicrobiawere present in both native forest soil and oil palm-cultivated

soil with larger representation in the oil palm-cultivatedarea Within Methanomicrobia the genera Rice Cluster IMethanosarcina andMethanocellawere present in both sam-ples (Figure 6(b)) These genera are closely related and foundin rice fields that produce methane [70] The order Metha-nomicrobiales was detected only in the native forest soilwhich can probably be explained by the relatively high levelsof carbon and organic matter This archaeal order is respon-sible for the final step of methane production in the carboncycle by anaerobic degradation of organic matter [66 71]However the contribution of the Methanomicrobia to thecarbon cycle in the oil palm-cultivated soils could not beassessed since there are no data on the methane productionin the studied areas

Among the Thaumarchaeota sequences used to generatethe phylogenetic tree that were unclassified at the genuslevel several OTUs formed different clusters associated withtheMiscellaneous Crenarchaeotic Group (Figure 5(a))Thesesequences were from both soil samples indicating that theunclassified Thaumarchaeota in the native forest and the oilpalm-cultivated area are phylogenetically related

Most of the sequences remained unclassified at thegenus level for both Thaumarchaeota and Euryarchaeota(Figures 6(a) and 6(b)) This result likely reflects the fact thatmost taxonomic databases do not have many representativesof Archaea especially belonging to the phylum Thaumar-chaeota thus few classifications of new sequences reach thegenus level Several genera were identified as Candidatusaccording to the International Code of Nomenclature ofBacteria indicating that characterization of the culturedprokaryote is incomplete [72]

The proportions of only four groups classified at thegenus level differed significantly between native forest and oilpalm-cultivated soils (Methanosarcina and Rice Cluster I inthe phylum Euryarchaeota and Candidatus NitrososphaeraandCandidatusNitrosotalea in the phylumThaumarchaeota)(Figure 7) With respect to the methanogens sequencesrelated to Methanosarcina were slightly more abundant insoil under oil palm cultivation (Figure 7) whereas sequencesbelonging to the Rice Cluster I were more abundant in nativeforest soil

In the Thaumarchaeota phylum Candidatus Nitrosos-phaera an AOA member of group I1b [65] was detectedin both samples but had a much higher representation innative forest (native forest 182 oil palm-cultivated area103) (Figure 7) In a comparative study of the archaealcommunities in Amazonian dark earth (ldquoTerra Pretardquo)and adjacent soils Candidatus Nitrososphaera was themain archaeal group found in both soils [10] In contrastCandidatus Nitrosotalea another AOA was more abundantin the oil palm-cultivated area (100) than in native forestsoil (44) in our study (Figure 7) Lehtovirta-Morley et al[73] cultivated the obligate acidophilic ammonia oxidizerCandidatus Nitrosotalea devanaterra from a nitrifying acidicsoil Later the same group characterized two strains ofacidophilic AOA belonging to this genus from a Chinesepaddy field and a Scottish agricultural soil [74] In our studythe greater representation of Candidatus Nitrososphaeraand Candidatus Nitrosotalea in native forest and oil

Archaea 11

palm-cultivated soils respectively can be explained by thelow soil pH (native forest pH 550 oil palm-cultivated areapH 483) since in basic soils the activity and growth ofthese archaea are diminished by the exponential decrease innitrite [6 75] In addition soil under oil palm cultivationis fertilized with nitrogen which may explain the higherlevel of Candidatus Nitrosotalea in this area compared withthat of native forest soil Finally although the cultivatedmembers of the genus Ca Nitrososphaera grow betteron neutral pH they have been reported to grow on pHvalues ranging from 60 to 85 In addition a recent workpublished by Wang et al (2014) [76] described active AOAsfrom an acidic soil (pH 492) which are closely related toNitrososphaera known as Nitrososphaera-like organismsby molecular phylogenetic methods Most of our sequenceswere classified as Nitrosotalea-like which are sequencesbelonging to the group I1a This group is typically foundin aquatic environments such as lake sediments or marineenvironments A similar ubiquitous distribution in soils isfound for the group I1b which are not restricted to a specificsoil pH condition

5 Conclusions

Although archaea are known to be ubiquitous there is stillmuch to be learned about their diversity biological rolesand metabolic capabilities This work is the first to comparearchaeal communities associated with the Amazon forest soilwith those of soil under oil palm cultivation

Archaeal diversity was found to be lower in oil palm-cultivated soil than in the native forest soil Only about 30of the OTUs were found in the oil palm soil in comparisonto the native forest However two main phyla were detectedin both samples Thaumarchaeota and Euryarchaeota with apredominance ofThaumarchaeota in native forest soil Theseresults support previous studies in other Brazilian biomenative soils (eg Caatinga Cerrado) as well as soil underdifferent types of land use (eg pasture and agricultural) Itis also important to mention that there are also groups ofuncertain classification associated with the Crenarchaeotaphylum

Although knowledge about Archaea has increased dra-matically in the last few decades little is known aboutthe biology of many archaeal groups and the taxonomyof Archaea is still a work in progress Thus it is notsurprising thatmany sequences identified in thiswork remainunclassified sometimes even at higher taxonomic levels Itis not clear whether the relatively small overall differencebetween soil archaeal communities of the Amazon forest andoil palm-cultivated land is a consequence of the current stateof archaeal taxonomy or whether it indicates that archaealdiversity is lower than bacterial diversity [77]

It is also important to consider that with land usechange due to agricultural practices the microstructure ofthe soil is broken down which can reduce the natural habitatfragmentation allowing potential interactions among theresident microbiota that otherwise would not happen [7879]

A phylogenetic tree of unclassified Thaumarchaeota 16SrRNA gene sequences was useful in exploring relationshipsamong these sequences When taxonomic classification ofsequences was possible it revealed the presence of OTUsfrom genera known to be involved in the metabolism ofcarbon and nitrogen (methanogens Methanocella and Meth-anosarcina AOACandidatusNitrososphaera andCandidatusNitrosotalea) hinting at possible roles for these archaealcommunities in Amazon soils

More studies are needed to elucidate these roles Cultur-ing some of these archaea in the laboratory would allow theirmetabolism and physiology to be studied in detail allowingfor a better understanding of the roles of Archaea in the bio-geochemical cycles

Conflict of Interests

The authors declare that there is no conflict of interestsregarding the publication of this paper

Acknowledgments

This work was supported by grants from FINEP CNPqCAPES and FAP-DF The authors thank Dr Boari for helpwith sample collection logistics Dr Betania Ferraz Quirinowas supported by the Cientista Visitante program fromEmbrapa

References

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[2] IBGE ldquoMapa de Biomas do Brasil primeira aproximacaordquo Riode Janeiro Brazil 2004 httpwwwibgegovbrhome

[3] C Azevedo-Ramos ldquoSustainable development and challengingdeforestation in the Brazilian Amazon the good the badand the uglyrdquo in Proceedings of the Symposium Our CommonGround Innovations in Land Use Decision-Making vol 59 p 5Vancouver Canada May 2007

[4] C A Francis K J Roberts J M Beman A E Santoro and B BOakley ldquoUbiquity and diversity of ammonia-oxidizing archaeainwater columns and sediments of the oceanrdquoProceedings of theNational Academy of Sciences of theUnited States of America vol102 no 41 pp 14683ndash14688 2005

[5] A H Treusch S Leininger A Kietzin S C Schuster H-PKlenk and C Schleper ldquoNovel genes for nitrite reductase andAmo-related proteins indicate a role of uncultivatedmesophiliccrenarchaeota in nitrogen cyclingrdquo Environmental Microbiol-ogy vol 7 no 12 pp 1985ndash1995 2005

[6] G W Nicol S Leininger C Schleper and J I Prosser ldquoTheinfluence of soil pH on the diversity abundance and tran-scriptional activity of ammonia oxidizing archaea and bacteriardquoEnvironmentalMicrobiology vol 10 no 11 pp 2966ndash2978 2008

[7] P Offre A Spang and C Schleper ldquoArchaea in biogeochemicalcyclesrdquo Annual Review of Microbiology vol 67 pp 437ndash4572013

[8] MAPA Anuario Estatıstico da Agroenergia 2012 StatisticalYearbook of Agroenergy 2013

[9] J C Bergmann D D Tupinamba O Y A Costa J R MAlmeida C C Barreto andB FQuirino ldquoBiodiesel production

12 Archaea

in Brazil and alternative biomass feedstocksrdquo Renewable ampSustainable Energy Reviews vol 21 pp 411ndash420 2013

[10] R G Taketani and S M Tsai ldquoThe influence of different landuses on the structure of archaeal communities in Amazoniananthrosols based on 16S rRNA and amoA genesrdquo MicrobialEcology vol 59 no 4 pp 734ndash743 2010

[11] C Brochier-Armanet B Boussau S Gribaldo and P ForterreldquoMesophilic crenarchaeota proposal for a third archaeal phy-lum theThaumarchaeotardquoNature Reviews Microbiology vol 6no 3 pp 245ndash252 2008

[12] S M Barns C F Delwiche J D Palmer and N R PaceldquoPerspectives on archaeal diversity thermophily and mono-phyly from environmental rRNA sequencesrdquo Proceedings of theNational Academy of Sciences of the United States of Americavol 93 no 17 pp 9188ndash9193 1996

[13] J G Elkins M Podar D E Graham et al ldquoA korarchaealgenome reveals insights into the evolution of the Archaeardquo Pro-ceedings of the National Academy of Sciences of the United Statesof America vol 105 no 23 pp 8102ndash8107 2008

[14] H Huber M J Hohn R Rachel T Fuchs V C Wimmer andK O Stetter ldquoA new phylum of Archaea represented by ananosized hyperthermophilic symbiontrdquo Nature vol 417 no6884 pp 63ndash67 2002

[15] T Nunoura H Oida M Nakaseama et al ldquoArchaeal diversityand distribution along thermal and geochemical gradients inhydrothermal sediments at the Yonaguni Knoll IV hydrother-mal field in the SouthernOkinawa TroughrdquoApplied and Enviro-nmental Microbiology vol 76 no 4 pp 1198ndash1211 2010

[16] C Rinke P Schwientek A Sczyrba et al ldquoInsights into thephylogeny and coding potential of microbial dark matterrdquoNature vol 499 no 7459 pp 431ndash437 2013

[17] A Spang J H Saw S L Joslashrgensen et al ldquoComplex archaea thatbridge the gap between prokaryotes and eukaryotesrdquo Naturevol 521 no 7551 pp 173ndash179 2015

[18] S Bagchi S E Vlaeminck L A Sauder et al ldquoTemporal andspatial stability of ammonia-oxidizing archaea and bacteria inaquariumbiofiltersrdquo PLoSONE vol 9 no 12 Article ID e1135152014

[19] W Qin S A Amin W Martens-Habbena et al ldquoMarineammonia-oxidizing archaeal isolates display obligate mixotro-phy and wide ecotypic variationrdquo Proceedings of the NationalAcademy of Sciences of the United States of America vol 111 no34 pp 12504ndash12509 2014

[20] M Coci N Odermatt M M Salcher J Pernthaler and GCorno ldquoEcology and distribution ofThaumarchaea in the deephypolimnion of Lake Maggiorerdquo Archaea vol 2015 Article ID590434 11 pages 2015

[21] J G Ferry ldquoHow tomake a living by exhalingmethanerdquoAnnualReview of Microbiology vol 64 pp 453ndash473 2010

[22] A Meyer A Focks V Radl G Welzl I Schoning and MSchloter ldquoInfluence of land use intensity on the diversity ofammonia oxidizing bacteria and archaea in soils from grasslandecosystemsrdquoMicrobial Ecology vol 67 no 1 pp 161ndash166 2014

[23] V De Gannes G Eudoxie and W J Hickey ldquoImpacts ofedaphic factors on communities of ammonia-oxidizing archaeaammonia-oxidizing bacteria and nitrification in tropical soilsrdquoPLoS ONE vol 9 no 2 Article ID e89568 2014

[24] D S Ribeiro F C J da Silva and TN Carvalho ldquoSobrevivenciade seis especies florestais em uma area explorada seletivamenteno municıpio de Moju Parardquo CERNE vol 9 no 2 pp 153ndash1632003

[25] D HM Costa C A P Ferreira J NM Silva J D C A Lopesand J O P D Carvalho ldquoPotencial madeireiro de floresta densano municıpio de Moju Estado do Parardquo Documentos 1998

[26] C M Almeida S F Lima C V Martins-Da-Silva and JI Gomes ldquoCaracterizacao morfologica e anatomica de dezespecies de leguminosae ocorrentes em uma floresta tropicalumida localizada no municıpio de Moju Estado do Parardquo inA Silvicultura na Amazonia Oriental Contribuicoes do ProjetoEmbrapa-DFID J NMC Silva JO P deCarvalho and J AGYared Eds pp 19ndash54 Embrapa Amazonia Oriental (CPATU)Belem Brazil 2001

[27] K Smalla N Cresswell L C Mendoncahagler A Woltersand J D Vanelsas ldquoRapid extraction protocol from soil forpolymerase chain reaction-mediated amplificationrdquo Journal ofApplied Bacteriology vol 74 pp 78ndash85 1993

[28] S Gantner A F Andersson L Alonso-Saez and S Bertils-son ldquoNovel primers for 16S rRNA-based archaeal communityanalyses in environmental samplesrdquo Journal of MicrobiologicalMethods vol 84 no 1 pp 12ndash18 2011

[29] P D Schloss S L Westcott T Ryabin et al ldquoIntroducingMOTHUR open-source platform-independent community-supported software for describing and comparing microbialcommunitiesrdquoApplied and EnvironmentalMicrobiology vol 75no 23 pp 7537ndash7541 2009

[30] PD SchlossDGevers and S LWestcott ldquoReducing the effectsof PCR amplification and sequencing artifacts on 16S rRNA-based studiesrdquo PLoS ONE vol 6 no 12 Article ID e27310 2011

[31] CQuince A Lanzen T P Curtis et al ldquoAccurate determinationof microbial diversity from 454 pyrosequencing datardquo NatureMethods vol 6 no 9 pp 639ndash641 2009

[32] P Yilmaz L W Parfrey P Yarza et al ldquoThe SILVA and lsquoall-species Living Tree Project (LTP)rsquo taxonomic frameworksrdquoNucleic Acids Research vol 42 no 1 pp D643ndashD648 2014

[33] R C Edgar B J Haas J C Clemente C Quince and RKnight ldquoUCHIME improves sensitivity and speed of chimeradetectionrdquo Bioinformatics vol 27 no 16 pp 2194ndash2200 2011

[34] A Chao and T-J Shen ldquoNonparametric estimation of Shan-nonrsquos index of diversity when there are unseen species insamplerdquo Environmental and Ecological Statistics vol 10 no 4pp 429ndash443 2003

[35] A Chao and T-J Shen Program SPADE (Species Prediction andDiversity Estimation) 2010

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[37] I J Good ldquoThe population frequencies of species and theestimation of population parametersrdquo Biometrika vol 40 pp237ndash264 1953

[38] K Tamura G Stecher D Peterson A Filipski and S KumarldquoMEGA6molecular evolutionary genetics analysis version 60rdquoMolecular Biology and Evolution vol 30 no 12 pp 2725ndash27292013

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[40] D H Parks G W Tyson P Hugenholtz and R G BeikoldquoSTAMP statistical analysis of taxonomic and functional pro-filesrdquo Bioinformatics vol 30 no 21 pp 3123ndash3124 2014

[41] R G Newcombe ldquoInterval estimation for the differencebetween independent proportions comparison of elevenmeth-odsrdquo Statistics in Medicine vol 17 no 8 pp 873ndash890 1998

[42] J D Storey J E Taylor and D Siegmund ldquoStrong controlconservative point estimation and simultaneous conservative

Archaea 13

consistency of false discovery rates a unified approachrdquo Journalof the Royal Statistical Society Series B Statistical Methodologyvol 66 no 1 pp 187ndash205 2004

[43] G C Baker J J Smith and D A Cowan ldquoReview and re-analysis of domain-specific 16S primersrdquo Journal of Microbio-logical Methods vol 55 no 3 pp 541ndash555 2003

[44] W H Zhong and Z C Cai ldquoLong-term effects of inorganicfertilizers on microbial biomass and community functionaldiversity in a paddy soil derived from quaternary red clayrdquoApplied Soil Ecology vol 36 no 2-3 pp 84ndash91 2007

[45] EWessen S Hallin and L Philippot ldquoDifferential responses ofbacterial and archaeal groups at high taxonomical ranks to soilmanagementrdquo Soil Biology and Biochemistry vol 42 no 10 pp1759ndash1765 2010

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[47] K Cenciani M R Lambais C C Cerri L C B De Azevedoand B J Feigl ldquoBacteria diversity and microbial biomass inforest pasture and fallow soils in the southwestern Amazonbasinrdquo Revista Brasileira de Ciencia do Solo vol 33 no 4 pp907ndash916 2009

[48] A A Navarrete R G Taketani L WMendes F de Souza Can-navan F M de Souza Moreira and S M Tsai ldquoLand-use sys-tems affect Archaeal community structure and functional diver-sity in western Amazon soilsrdquo Revista Brasileira de Ciencia doSolo vol 35 no 5 pp 1527ndash1540 2011

[49] E Lumini A Orgiazzi R Borriello P Bonfante and V Bian-ciotto ldquoDisclosing arbuscular mycorrhizal fungal biodiversityin soil through a land-use gradient using a pyrosequencingapproachrdquo Environmental Microbiology vol 12 no 8 pp 2165ndash2179 2010

[50] M Lupatini R J S Jacques Z I Antoniolli A K A SuleimanR R Fulthorpe and L F W Roesch ldquoLand-use change andsoil type are drivers of fungal and archaeal communities in thePampa biomerdquo World Journal of Microbiology and Biotechnol-ogy vol 29 no 2 pp 223ndash233 2013

[51] E G Lamb N Kennedy and S D Siciliano ldquoEffects of plantspecies richness and evenness on soil microbial communitydiversity and functionrdquo Plant and Soil vol 338 no 1 pp 483ndash495 2011

[52] A Spang R Hatzenpichler C Brochier-Armanet et al ldquoDis-tinct gene set in two different lineages of ammonia-oxidizingarchaea supports the phylum Thaumarchaeotardquo Trends inMicrobiology vol 18 no 8 pp 331ndash340 2010

[53] R G Pacchioni F M Carvalho C E Thompson et al ldquoTax-onomic and functional profiles of soil samples from Atlanticforest and Caatinga biomes in northeastern Brazilrdquo Microbio-logyOpen vol 3 no 3 pp 299ndash315 2014

[54] E Catao A P Castro C C Barreto R H Kruger and C MKyaw ldquoDiversity of Archaea in Brazilian savanna soilsrdquoArchivesof Microbiology vol 195 no 7 pp 507ndash512 2013

[55] M Pester T Rattei S Flechl et al ldquoAmoA-based consensusphylogeny of ammonia-oxidizing archaea and deep sequencingof amoA genes from soils of four different geographic regionsrdquoEnvironmental Microbiology vol 14 no 2 pp 525ndash539 2012

[56] J-Z He H-W Hu and L-M Zhang ldquoCurrent insights into theautotrophic thaumarchaeal ammonia oxidation in acidic soilsrdquoSoil Biology amp Biochemistry vol 55 pp 146ndash154 2012

[57] L-M Zhang H-W Hu J-P Shen and J-Z He ldquoAmmonia-oxidizing archaea have more important role than ammonia-oxidizing bacteria in ammonia oxidation of strongly acidicsoilsrdquo ISME Journal vol 6 no 5 pp 1032ndash1045 2012

[58] S Leininger T Urich M Schloter et al ldquoArchaea predominateamong ammonia-oxidizing prokaryotes in soilsrdquo Nature vol442 no 7104 pp 806ndash809 2006

[59] N C Banning L D Maccarone L M Fisk and D VMurphy ldquoAmmonia-oxidising bacteria not archaea dominatenitrification activity in semi-arid agricultural soilrdquo ScientificReports vol 5 Article ID 11146 2015

[60] M Herrmann A Hadrich and K Kusel ldquoPredominance ofthaumarchaeal ammonia oxidizer abundance and transcrip-tional activity in an acidic fenrdquoEnvironmentalMicrobiology vol14 no 11 pp 3013ndash3025 2012

[61] K Zhalnina P Dorr de Quadros F O Camargo and E WTriplett ldquoDrivers of archaeal ammonia-oxidizing communitiesin soilrdquo Frontiers in Microbiology vol 3 article 210 Article IDArticle 210 2012

[62] J Hong J Cho and Y Hong ldquoEnvironmental variables shapingthe ecological niche of Thaumarchaeota in soil direct andindirect causal effectsrdquo PLoS ONE vol 10 no 8 Article IDe0133763 2015

[63] G Jurgens K Lindstrom and A Saano ldquoNovel group withinthe kingdom Crenarchaeota from boreal forest soilrdquo Appliedand Environmental Microbiology vol 63 no 2 pp 803ndash8051997

[64] M Tourna M Stieglmeier A Spang et al ldquoNitrososphaeraviennensis an ammonia oxidizing archaeon from soilrdquoProceed-ings of the National Academy of Sciences of the United States ofAmerica vol 108 no 20 pp 8420ndash8425 2011

[65] K V Zhalnina R Dias M T Leonard et al ldquoGenome sequenceof Candidatus Nitrososphaera evergladensis from group I1benriched from everglades soil reveals novel genomic features ofthe ammonia-oxidizing archaeardquoPLoSONE vol 9 no 7 ArticleID e101648 2014

[66] I Anderson L E Ulrich B Lupa et al ldquoGenomic char-acterization of methanomicrobiales reveals three classes ofmethanogensrdquo PLoS ONE vol 4 no 6 Article ID e5797 2009

[67] J E Galagan C Nusbaum A Roy et al ldquoThe genome of Macetivorans reveals extensive metabolic and physiological diver-sityrdquo Genome Research vol 12 no 4 pp 532ndash542 2002

[68] C Wrede S Kokoschka A Dreier C Heller J Reitner and MHoppert ldquoDeposition of biogenic iron minerals in a methaneoxidizing microbial matrdquo Archaea vol 2013 Article ID 1029728 pages 2013

[69] G Garrity Bergeyrsquo Manual of Systematic Bacteriology 721 2001[70] S Sakai Y Takaki S Shimamura et al ldquoGenome sequence

of a mesophilic hydrogenotrophic methanogen Methanocellapaludicola the first cultivated representative of the orderMethanocellalesrdquo PLoS ONE vol 6 no 7 Article ID e228982011

[71] E Bapteste C Brochier and Y Boucher ldquoHigher-level clas-sification of the Archaea evolution of methanogenesis andmethanogensrdquo Archaea vol 1 no 5 pp 353ndash363 2005

[72] R G E Murray and E Stackebrandt ldquoTaxonomic note imple-mentation of the provisional status Candidatus for incompletelydescribed procaryotesrdquo International Journal of Systematic Bac-teriology vol 45 no 1 pp 186ndash187 1995

[73] L E Lehtovirta-Morley K Stoecker A Vilcinskas J I Prosserand G W Nicol ldquoCultivation of an obligate acidophilic ammo-nia oxidizer from a nitrifying acid soilrdquo Proceedings of the

14 Archaea

National Academy of Sciences of the United States of Americavol 108 no 38 pp 15892ndash15897 2011

[74] L E Lehtovirta-Morley C Ge J Ross H Yao G W Nicol andJ I Prosser ldquoCharacterisation of terrestrial acidophilic archaealammonia oxidisers and their inhibition and stimulation byorganic compoundsrdquo FEMSMicrobiology Ecology vol 89 no 3pp 542ndash552 2015

[75] M J Frijlink T Abee H J Laanbroek W de Boer and W NKonings ldquoThe bioenergetics of ammonia and hydroxylamineoxidation in Nitrosomonas europaea at acid and alkaline pHrdquoArchives of Microbiology vol 157 no 2 pp 194ndash199 1992

[76] B Wang Y Zheng R Huang et al ldquoActive ammonia oxidizersin an acidic soil are phylogenetically closely related to neu-trophilic archaeonrdquo Applied and Environmental Microbiologyvol 80 no 5 pp 1684ndash1691 2014

[77] J Y Aller and P F Kemp ldquoAre Archaea inherently less diversethan Bacteria in the same environmentsrdquo FEMS MicrobiologyEcology vol 65 no 1 pp 74ndash87 2008

[78] K Potthast U Hamer and FMakeschin ldquoLand-use change in atropical mountain rainforest region of southern Ecuador affectssoilmicroorganisms and nutrient cyclingrdquoBiogeochemistry vol111 no 1ndash3 pp 151ndash167 2012

[79] J I Prosser and G W Nicol ldquoArchaeal and bacterial ammonia-oxidisers in soil the quest for niche specialisation and differen-tiationrdquoTrends inMicrobiology vol 20 no 11 pp 523ndash531 2012

Submit your manuscripts athttpwwwhindawicom

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International Journal of

Volume 2014

Zoology

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4 Archaea

Table 1 Number of operational taxonomic units (OTUs) Goodrsquos coverage estimator OTU richness indices (Chao1 ACE) and InverseSimpson and Shannon diversity indices for archaeal communities in native forest soil and soil under oil palm cultivation

Sample OTUslowast Goodrsquos coverage () Chao1 ACE Inverse Simpson ShannonNative forest 821 9871 2124 4645 2645 422Oil palm-cultivated area 253 9972 424 674 1547 331lowastConsidering 97 sequence similarity in MOTHUR

formicica (NR112877) Methanocella paludicola (NR074192)Halococcus hamelinensis (LN651155) and Pyrococcus furiosus(U20163) from the phylum Euryarchaeota Pyrobaculumaerophilum (NR102764) from the phylum CrenarchaeotaCandidatus Nitrososphaera sp (FR773157) and CandidatusNitrososphaera gargensis (GU797786) fromThaumarchaeotaGroup I1b Nitrosopumilus maritimus (JQ346765) andCenarchaeum symbiosum (U51469) from ThaumarchaeotaGroup I1a uncultured acidic red soil AOA (FJ174727)uncultured trembling aspen archaeon (EF021427) and borealforest archaeon (X96688) from Thaumarchaeota GroupI1c and several uncultured organisms classified accordingto the SILVA database as Thaumarchaeota MiscellaneousCrenarchaeotic Group (FR745121 AM910782 JX984848KC510333 FJ485299 KC831395 FJ920714 FJ485307HM051127 HM051130 and HM051125) A 16S rRNA genesequence from Acidobacteria KBS 96 (FJ870384) was usedas an outgroup A second phylogenetic tree was createdusing only Euryarchaeota OTUs from each sample that wereunclassified at the class level with the same parameters andreference sequences

A Venn diagram was generated by grouping OTUs clus-tered at 97 similarity to show the number of OTUs uniqueto each area and those shared between samples

Differences in taxonomic distribution of archaea betweennative forest and the oil palm-cultivated area were evaluatedby Fisherrsquos exact test (119902 le 005) using Statistical Analysisof Metagenomic Profiles (STAMP) software [40] The confi-dence interval was estimated using the asymptotic method[41] and correction of the 119902 value was calculated usingStoreyrsquos false discovery rate (FDR) approach [42] A filterwas applied to show only the differences between proportionsabove 05

3 Results

Goodrsquos coverage estimate of the completeness of sampling(Table 1) was higher for the oil palm-cultivated area than forthe native forest

The number of archaeal OTUs was higher in the nativeforest soil (821 sequences) than in soil under oil palmcultivation (253 sequences) (Figure 2) with 130OTUs (138)shared between the two areas Archaeal species richness asassessed by abundance-based estimators Chao1 and ACEwas higher in the native forest than in the oil palm-cultivatedarea Similarly the Inverse Simpson and Shannon indicesindicated that archaeal species diversity was higher in nativeforest soil compared to soil under oil palm cultivation(Table 1)

Table 2 Physicochemical properties of native forest soil and soilunder oil palm cultivation

Native forest Oil palm-cultivated areapH(in water) 550 483

P (mgsdotdmminus3) 140 7070K (mgsdotdmminus3) 003 011Ca+2 (cmolsdotdmminus3) 140 033Mg+2 (cmolsdotdmminus3) 010 017Al+3 (cmolsdotdmminus3) 040 073H + Al (cmolsdotdmminus3) 620 727Na+ (cmolsdotdmminus3) 001 002Organic matter (gkg) 3100 1163Total C (gkg) 1800 677Total N (mgkg) NA 115NO3

minus1 (mgkg) NA 079NO2

minus1 (mgkg) NA 014NH4

+ (mgkg) NA 021Data not available

Soil physicochemical parameters differed between theareas (Table 2) with higher levels of phosphorous potassiumand total carbon in the oil palm-cultivated soil and higherlevels of calcium and organic matter in the native forest soil

In both areas only two archaeal phyla (Thaumarchaeotaand Euryarchaeota) were detected (Figure 3) Thaumar-chaeota were predominant in the native forest soil (58) andEuryarchaeota were predominant in the oil palm-cultivatedsoil (54)The percentage of unclassified archaea was similarin both soils (native forest 5 oil palm-cultivated area 6)

At the class level a total of 10 groups were detected inboth samples Halobacteria Methanobacteria Methanomi-crobia and Thermoplasmata in the phylum Euryarchaeotaand Marine Group I OPPD003 Soil Crenarchaeotic GroupSouth African Gold Mine Gp 1 Terrestrial Group andMiscellaneous Crenarchaeotic Group in the phylum Thau-marchaeota (Figures 4(a) and 4(b)) The most abundantclass present in native forest soil was the Soil Crenar-chaeotic Group representing 26 of all native forest OTUs(Figure 4(a)) and 45 of theThaumarchaeota sequences (notshown) followed by Methanomicrobia representing 15 ofall native forest OTUs (Figure 4(b)) and 42 of the Eur-yarchaeota sequences (not shown) The most abundant classpresent in the oil palm-cultivated area was Thermoplasmatarepresenting 21 of all cultivated area OTUs (Figure 4(b))and 39 of the Euryarchaeota sequences (not shown) fol-lowed by Methanomicrobia representing 16 of all culti-vated area OTUs (Figure 4(b)) and 30 of all Euryarchaeota

Archaea 5

Venn diagram at distance 003

123Cultivated area

691Native forest

130

Figure 2 Venn diagram depicting the number of shared and unique operational taxonomic units (OTUs) in native forest and oil palm-cultivated area

Thaumarchaeota

Euryarchaeota

Unclassified archaea

Cultivated areaNative forest

10 20 30 40 50 60 700()

Figure 3 Comparison of archaeal phyla present in soil samples from native Amazon forest (orange) and an area under oil palm cultivation(blue) The percentages of unclassified sequences are shown

sequences (not shown) The predominant class from thephylum Thaumarchaeota in the oil palm-cultivated areawas Terrestrial Group representing 14 of the total OTUs(Figure 4(a)) and 35 of all Thaumarchaeota sequences (notshown)

Because it is a newer phylum Thaumarchaeota have fewcultivated representatives compared with Euryarchaeota andfewer sequences in databases such as SILVA To determinehow the Thaumarchaeota sequences that could not be clas-sified at the genus level were related to each other a phylo-genetic tree was constructed with representative sequences(Figure 5(a)) One native forest OTU clustered with GroupI1b (NF6) and one native forest OTU (NF17) and two cul-tivated area OTUs (CA50 CA52) clustered with Group I1cThe remaining OTUs formed different clusters all associatedwith the Miscellaneous Crenarchaeotic Group Most unclas-sified Thaumarchaeota sequences from the native forest andoil palm-cultivated area grouped together although a fewunclassified OTUs from the cultivated area (eg CA51 toCA15 and CA24 to CA33) and native forest (eg NF21 toNF49 and NF5 to NF35) formed separate clusters

The unclassified sequences at genus level in the Euryar-chaeota phylum were all related to the orders Methano-cellales Methanosarcinales Methanomicrobiales Thermo-plasmatales Methanobacteriales and Halobacteriales(Figure 5(b)) The classified sequences at genus level wereMethanocella Methanosarcina Methanoculleus Methano-massiliicoccus Methanomethylophilus MethanobrevibacterMethanobacterium and Candidatus lainarchaeum(Figure 5(b))

The genus-level analysis showed that unclassified SoilCrenarchaeotic Group was predominant in the native forestaccounting for 21 of all native forest OTUs (Figure 6(a))In the oil palm-cultivated area the predominant genus wasUnclassified Terrestrial Group accounting for 14 of allcultivated area OTUs (Figure 6(b))

A total of 31 groups of OTUs were identified (onlymajor groups are shown) most of which are unclassi-fied The classified groups in the phylum Euryarchaeotaare Candidatus lainarchaeum Methanobacterium Methano-brevibacter Methanocella Rice Cluster I Methanoculleus

6 Archaea

Marine Group I

Miscellaneous Crenarchaeotic Group

Soil Crenarchaeotic Group

South African Gold Mine Gp 1

Terrestrial Group

Unclassified Thaumarchaeota

OPPD003

Cultivated areaNative forest

5 10 15 20 25 300()

(a)

Methanobacteria

Methanomicrobia

Halobacteria

Thermoplasmata

Unclassified Euryarchaeota

Cultivated areaNative forest

5 10 15 20 250()

(b)

Figure 4 Comparison of archaeal classes present in soil samples from native Amazon forest (orange) and an area under oil palm cultivation(blue) Classes within the phyla Thaumarchaeota (a) and Euryarchaeota (b) are shown as well as the percentage of unclassified sequences

Methanoregula Methanimicrococcus Methanosarcina Can-didatus Methanomethylophilus and Methanomassiliicoccus(Figure 6(b)) The classified groups in the phylum Thau-marchaeota are Candidatus Nitrososphaera and CandidatusNitrosotalea (Figure 6(a))

The analysis of the genera that differ significantly inrelative abundance between native forest soil and the oilpalm-cultivated area revealed that the greatest difference wasseen for Candidatus Nitrososphaera (Figure 7) This genuswas predominant in the native forest soil accounting for182 of the sequences but represented only 103 of thesequences from the oil palm-cultivated area UnclassifiedSoil Crenarchaeotic Group was also predominant in thenative forest (ie 388 of sequences in the native forest

and 312 of sequences in the oil palm-cultivated area)whereas unclassified South African Gold Mine Gp 1 waspredominant in the oil palm-cultivated area (ie 73 ofsequences in the native forest and 135 of sequences inthe oil palm-cultivated area) Candidatus Nitrosotalea repre-sented 44 of the native forest sequences and 100 of theoil palm-cultivated area sequences The relative abundanceof unclassified Terrestrial Miscellaneous Group unclassifiedThaumarchaeotaunclassifiedThermoplasmatalesunclassifiedMiscellaneous Crenarchaeotic Group Methanosarcina andRice Cluster I also differed significantly between native forestsoil and soils under oil palm cultivation but these differenceswere smaller

Archaea 7

FJ4853071| Uncultured archaeon clone

FJ4852991| Uncultured archaeon clone

FJ9027141| Uncultured archaeon clone

KC8313951| Uncultured archaeon clone

KC5103331| Uncultured archaeon clone

JX9848481| Uncultured archaeon clone

AM9107821| Uncultured archaeon clone

Uncultured archaeon acidic red soil-FJ174727

Ca Nitrososphaera gargensis-GU797786

Ca Nitrososphaeramdash(48 6)

Ca Nitrosotaleamdash(49 12)

Candidatus lainarchaeum (0 2)

Methanobacterium (2 3)

Methanomassillicoccus (6 2)

Methanocella (33 8)

Rice cluster I (43 25)

Methanosarcina baltica-AB973356

Methanocella paludicola-NR074192

Pyrobaculum aerophilum

-NR102764

Acidobacteria bacterium KBS 96-FJ870384

Ca Methanom

ethylophilus (12 0)

Ca Nitrososphaera sp-FR773157

Nitrosopumilus maritimus-JQ346765

Halococcus hamelinensis-LN651155

Pyrococcus furiosus-PFU20163

Cenarchaeum symbiosum-CSU51469

Methanoregula formicica-NR112877

FR7451211| Uncultured soil archaeon

Boreal forest soil archaeon-X96688

Trembling aspen archaeon-EF021427

MCG

MCG005

CA4099

99

7174

63

86

9687

9067

5696

6768 54

57

62

58

65

7559

62

63

65

64

52

64

64

1008654

91

99

999293

53798392

88

7274

5164

5858

CA7CA32

CA50NF17

NF6

CA52

T I1c

T I1b

T I1a

CA24CA6CA12

CA4

CA33

NF16

NF2

NF35

NF9

NF5NF3

6NF4

1NF3

2

NF1

2

NF4

NF1

NF3

0NF2

3

NF7CA3

CA39

CA37

CA46

CA44

CA2

CA42

CA27

Unclassified MCGmdash(16 4)

Unclassified Terrestrial Group (94 36)

Unclassified Marine Group I (2 2)

CA15

CA25

CA30

CA43

CA35

CA14

CA29

CA13

CA20

CA11

CA41

CA36

CA51

NF48

CA9NF4

0NF4

2CA21

CA22

NF38

CA45

CA16

CA47

NF31CA31NF28NF47NF27NF45CA18NF26

NF24NF33NF19

NF18NF37

NF34NF43NF15NF44NF13

NF22NF29NF11

CA49

CA1

CA19CA8

CA10

CA23CA38

CA54HM0511251| Uncultured

HM0511301| Uncultured archaeon clone

HM0511271| Uncultured archaeon clone

archaeon clone

NF46NF14

NF49NF25N

F10N

F39N

F20N

F21

NF8

NF3

CA48

CA34

CA5

CA28CA26CA17CA53

(E)

(B)

(C)

(a)

CA13

CA14

NF1

2

NF1

4M

ethan

ocell

a (3

8)Meth

anosarcina (3

8 4)

Methanoculleus (2 0)

Methanomassilicoccus (6 2)

Methanomassilicoccus (6 2)

Methanomassilicoccus (6 2)

Methanomethylophilus (6 2)

Methanomethylophilus (6 2)

Methanobrevibacter (1 0)

Methanobacterium

(2 3)

Methanobacterium

(2 3)

Methanobacterium

(2 3)

Methanobacterium

(2 3)

Methanobacterium

(2 3)

Methanosarci

na (38 4)

Methanosarcina (38 4)

Methanosarcina (38 4)

Methanosarcina (38 4)

NF16

CA4

CA15CA20

CA19

NF3NF1

Meth

anoc

ella (

33 8

)

Meth

anoc

ella p

aludi

cola-

NR 0741

92 1

Meth

anosa

rcina

baltic

a AB97

3356

1

Methanoregula formicica NR 112877 1

Halococcus hamelinensis LN651155 1

CA6N

F6NF8

NF19 NF5NF7NF20

Pyrobaculum aerophilum NR 102764 2

Ca Nitrososphaera gargensis G

U797786 1

Ca Nitrososphaera gargensis GU797786 1

Nitrosopumilus maritimus JQ346765 1Cenarchaeum symbiosum U51469 1Uncultured archaeon acidic red soil FJ174727 1

Boreal forest soil archaeon X96688 1

Trembling aspen archaeon EF021427 1

HM051125 1 Uncultured archaeon clone

HM051127 1 Uncultured archaeon clone

HM051130 1 Uncultured archaeon clone

FJ485307 1 Uncultured archaeon clone

FJ902714 1 Uncultured archaeon clone

KC831395 1 archaeon clone

FJ485299 1 archaeon clone

KC510333 1 archaeon clone

FR745121 1 archaeon clone

JX984848 1 archaeon clone

AM910782 1 archaeon clone

Acidobacteria bacterium KBS 96 FJ870384 1

Ca Nitrososphaera sp FR773157 1

CA7

Pyrococcus furiosus U20163 1

005

MCG

Halobacteriales

Cand

idat

us la

inar

chae

um (0

2)

Cand

idat

us la

inar

chae

um (0

2)

Thermoplasmatales

Methanomicrobiales

Methanosarcinalles

Methanocellales

T I1c

T I1a

T I1b

(C)

998785

58

100

100

100100

100

71 84 71

53

54

65

86

94

98

59100

100

78

10051

5477

63 73

99

100

8497100

78

100

100

76956970

100 10

010

0

637551

100

9999

9958

94

Methanobacteriales

(b)

Figure 5 Phylogenetic trees of (a) unclassified Thaumarchaeota 16S rRNA gene sequences from native forest soil and soil under oil palmcultivation and (b) 16S rRNA gene sequences classified as Euryarchaeota from native forest soil and soil under oil palm cultivation Theneighbor-joining method and Jukes-Cantor corrections were used with 1000 bootstrap replicates Bootstrap values above 50 are shownin the trees A representative number of operational taxonomic units (OTUs) were selected at 3 dissimilarity by MOTHUR to avoidredundancies in the phylogenetic trees Randomly chosen OTUs from native forest soil and from soil under oil palm cultivation were usedin the construction of the trees along with archaeal reference sequences (from GenBank) from the following groups Acidobacteria (B)Crenarchaeota (C) Euryarchaeota (E) Miscellaneous Crenarchaeotic Group (MCG) Thaumarchaeota Group I1a (T I1a) ThaumarchaeotaGroup I1b (T I1b) andThaumarchaeota Group I1c (T I1c)The AOAs are highlighted with the symbolX Sampling site is indicated by ldquoNFrdquo(native forest) or ldquoCArdquo (oil palm-cultivated area) followed by the number of the OTUs (NAxx or CAxx) OTU sequences classified to at leastthe class level are shown in bold followed by two numbers in parentheses indicating the number of sequences of that OTU detected in nativeforest soil and the number detected in the oil palm-cultivated area The scale bar represents the 5 estimated sequence divergence

8 Archaea

Unclassified Marine Group I

Unclassified Miscellaneous Crenarchaeotic Group

Unclassified Thaumarchaeota

Candidatus Nitrososphaera

Unclassified Soil Crenarchaeotic Group

Candidatus Nitrosotalea

Unclassified South African Gold Mine Gp 1

Unclassified Terrestrial Group

Cultivated areaNative forest

5 10 15 20 250()

(a)

Unclassified Thermoplasmatales

Unclassified Terrestrial Miscellaneous Group

Rice Cluster I

Methanocella

Methanosarcina

Unclassified Euryarchaeota

Unclassified Halobacteriales

Candidatus Methanomethylophilus

Unclassified Methanosarcinaceae

5 10 15 20 250()

Cultivated areaNative forest

(b)

Figure 6 Percentage of archaeal operational taxonomic units within the Thaumarchaeota (a) and Euryarchaeota (b) phyla classified at thegenus level for native forest soil (orange) and soil under oil palm cultivation (blue) The percentage of unclassified sequences at genus level isshown

4 Discussion

The Amazon forest is known for its macrospecies diversityhowever few studies have addressed its microbial diversityAmong the domains of life Archaea are undoubtedly the lesswell known

In this work almost 600000 archaeal 16S rRNA genesequences were obtained from native forest soil and anadjacent area under oil palm cultivation After processing(ie removal of short sequences sequences with errors low-quality sequences chimeras and possible contaminants) andnormalization 39111 sequences per area remained for anal-ysis This number of sequences provided sufficient coverageof archaeal diversity in both areas (Table 1)The primers used

were based on a large number of 16S rRNA sequences fromthe SILVA database [28] however given that little is knownabout this domain it is unlikely that the archaeal diversitywasfully covered [43]

The main physicochemical differences between soils ofthe native forest and the oil palm-cultivated area were thelevels of phosphorous potassium calcium organic matterand total carbon (Table 2) Correlations between phylogenyand physicochemical parameters were evaluated using PCAand NMDS however no clear clusters or correlations wereobserved (results not shown)

In a study of the long-term effects of inorganic fertilizerson microbial communities Zhong and Cai [44] reportedthat phosphorus application indirectly altered microbial

Archaea 9

Candidatus NitrososphaeraUnclassified Soil Crenarchaeotic Group (SCG)

Unclassified South African Gold Mine Gp 1 (SAGMCG-1)Candidatus Nitrosotalea

Unclassified Terrestreial Miscellaneous Gp (TMEG)Unclassified Thaumarchaeota

Unclassified ThermoplasmatalesUnclassified Miscellaneous Crenarchaeotic Group

MethanosarcinaRice Cluster I

95 confidence intervals

q-v

alue

(cor

rect

ed)

Cultivated areaNative forest

531e minus 10

420 6 8minus4minus6minus8 minus2minus10

Difference between proportions ()38800

Proportion ()

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

Figure 7 Archaeal genera that differ significantly (119902 lt 005) in relative abundance between native forest soil (orange) and soil under oil palmcultivation (blue) For the purpose of statistical analysis performed using STAMP software the sum of the percentages of archaea classifiedat the genus level in both soils is 100 Differences between proportions of each genus are shown with 95 confidence intervals Unclassifiedsequences were used to calculate frequency profiles but are not displayed

parameters in soils and increased crop yields by elevating thelevels of organic matter Wessen and collaborators used real-time PCR quantification to show the impacts of fertilizer useon several bacterial phyla and the archaeal phylum Crenar-chaeota (Thaumarchaeota Group I1c after reanalysis usingthe SILVAdatabase 119) [45]They reported that bacterial andarchaeal classes respond differently to fertilization primarilybecause of changes in certain soil parameters such as pHtotal nitrogen and the carbon nitrogen ratio In our studylevels of total carbon and organic matter were higher in thenative forest than in the oil palm-cultivated area which mayhave contributed to higher archaeal diversity Although thereare no studies on the effect of total soil carbon on archaealcommunities there are studies on the bacterial communitiesin Amazonian soils The work of Edgar et al [33] describeddistinct soil bacterial communities under different vegetationtypes The authors found that bacterial diversity correlatedwith total organic carbon and total nitrogen content [46]Furthermore in a study of bacterial diversity and microbialbiomass in forest pasture and fallow soils in the Amazonbasin microbial biomass was highest in pasture soils whichalso had 30ndash47 higher carbon levels than the other sitesover the course of a year [47]

Several studies have also reported alterations in thecomposition of archaeal [48] bacterial [47] and fungal [49]communities associated with land use changes A studyof archaeal communities in Amazonian anthrosols undervarious land uses (agriculture pasture and secondary forest)and cultivation practices (eg eggplant banana citrus andmanioc) [10] reported that agriculture negatively affectedarchaeal community diversity Results of ribosomal inter-genic spacer analysis showed that soil type is also responsiblefor changes in archaeal communities in soils under differenttypes of land use (native grassland native forest Eucalyptus

and Acacia plantations and soybean and watermelon fields)[50]

The Venn diagram showed that the native forest soil has agreater number of unique sequences (Figure 2) This was notsurprising due to the greater archaeal richness and diversityobserved in the native forest soil (Table 2) The number ofunique OTUs found in the oil palm-cultivated area is almostthe same as that of shared OTUs with the native forest areaAlthough oil palm cultivation decreased archaeal diversity inthe soil there is still a core group of archaea OTUs sharedwith the native forest suggesting that these archaea havecertain characteristics that allow them to persist after thechange in land use It cannot be ruled out that these archaeaare metabolically inactive The smaller number of uniquesequences in the oil palm-cultivated area may be related to alower complexity of the vegetation (ie oil palmmonocultureversus great plant diversity in the forest) since plant diversitycould affect these communities [51]

In our study all OTUs identified belong to the domainArchaea and were classified into two phyla Euryarchaeotaand Thaumarchaeota It is important to mention that thedatabases were only recently updated to recognize the phy-lumThaumarchaeota therefore previous studies on Archaeacould not recognize this phylum as proposed by Brochier-Armanet et al [11] The creation of this novel phylum wassupported by a study comparing the genomes of two marineammonia-oxidizing archaea (AOAs) that identified a core setof informational genes specific to this group [52] Thereforeto allow comparisons with other studies we reanalyzed theirdata whenever possible updating the taxonomic classifica-tion to include the phylumThaumarchaeota

Thework ofChao and Shen [35] focused on the impacts ofland use (ie agricultural systems of indigenous people andcattle pasture) on archaeal communities by PCR (amoA gene)

10 Archaea

and denaturing gradient gel electrophoresis analysis of differ-ent Western Amazon soils The databases at the time did notconsiderThaumarchaeota in their classification however ourreanalysis of their dataset using an updated database showedthat Euryarchaeota and Thaumarchaeota were present intheir samples along with Crenarchaeota (Thermoprotei) Inaddition changes were observed in the structure of soilarchaeal communities when comparing the cultivated areaswith forest areas that had been converted to pasture

In our study differences in the percentages of Thau-marchaeota and Euryarchaeota found in the soil sampleswere not striking Thaumarchaeota were predominant in thenative forest (58) whereas Euryarchaeota were predom-inant in the oil palm-cultivated area with 54 (Figure 3)This result is consistent with other comparative studies onarchaeal communities in different types of Brazilian soils (ieAmazonian anthrosols Caatinga and Cerrado) [10 53 54]and a study of different tropical forest soil types that reportedThaumarchaeota as the predominant phylum [44]

The phylumThaumarchaeota comprises all known AOAsand is directly involved in nitrogen metabolism Ammonia isused as an energy source by Group I1a found predominantlyin aquatic systems [52] and Group I1b found in soils [55]Thaumarchaeota Group I1c is primarily found in acidic soilsbut the ecological roles of these archaea are still unknown [5657]

The literature presents conflicting data about the pro-portion of AOAsAOBs soils In some studies AOAs arepredominant [58] while in others AOBs are more abundant[59] However several studies found a predominance of thearchaeal amoA gene over the bacterial amoA gene in soils[60] AOAs are frequently detected in soils their abundancerange from 05 to 10 of the prokaryotic community andtheir abundance is strongly influenced by physicochemicalconditions as well as the organic matter content [61 62] Inacidic soils AOAs appear to have a more important role inammonia oxidation than AOBs [57]

The Soil Crenarchaeotic Group a class within Thaumar-chaeota according to the SILVA database classification wasdetected in both soil samples but a higher representationwas observed in native forest soil This class was discoveredin boreal forest soil [63] Candidatus Nitrososphaera whichbelongs to the Soil Crenarchaeotic Group class is a memberof Thaumarchaeota Group I1b it is an AOA that usesammonia or urea as an energy source [64 65]

On the other hand members of the phylum Eur-yarchaeota are directly involved in carbon metabolismwith several classes of methanogens and methane-oxidizingorganisms [66ndash68] Navarrete et al [48] studied the impactof converting Amazon native forest to pasture and reportedchanges not only to the composition and abundance of soilmicrobial communities but also to a decrease in the diversityof gene function

Within the phylum Euryarchaeota Methanomicrobiaand Thermoplasmata were the main classes detected in oursamples along with a few representatives of Methanobac-teria and Halobacteria Members of Methanobacteria andMethanomicrobia are methanogens [69] Methanomicrobiawere present in both native forest soil and oil palm-cultivated

soil with larger representation in the oil palm-cultivatedarea Within Methanomicrobia the genera Rice Cluster IMethanosarcina andMethanocellawere present in both sam-ples (Figure 6(b)) These genera are closely related and foundin rice fields that produce methane [70] The order Metha-nomicrobiales was detected only in the native forest soilwhich can probably be explained by the relatively high levelsof carbon and organic matter This archaeal order is respon-sible for the final step of methane production in the carboncycle by anaerobic degradation of organic matter [66 71]However the contribution of the Methanomicrobia to thecarbon cycle in the oil palm-cultivated soils could not beassessed since there are no data on the methane productionin the studied areas

Among the Thaumarchaeota sequences used to generatethe phylogenetic tree that were unclassified at the genuslevel several OTUs formed different clusters associated withtheMiscellaneous Crenarchaeotic Group (Figure 5(a))Thesesequences were from both soil samples indicating that theunclassified Thaumarchaeota in the native forest and the oilpalm-cultivated area are phylogenetically related

Most of the sequences remained unclassified at thegenus level for both Thaumarchaeota and Euryarchaeota(Figures 6(a) and 6(b)) This result likely reflects the fact thatmost taxonomic databases do not have many representativesof Archaea especially belonging to the phylum Thaumar-chaeota thus few classifications of new sequences reach thegenus level Several genera were identified as Candidatusaccording to the International Code of Nomenclature ofBacteria indicating that characterization of the culturedprokaryote is incomplete [72]

The proportions of only four groups classified at thegenus level differed significantly between native forest and oilpalm-cultivated soils (Methanosarcina and Rice Cluster I inthe phylum Euryarchaeota and Candidatus NitrososphaeraandCandidatusNitrosotalea in the phylumThaumarchaeota)(Figure 7) With respect to the methanogens sequencesrelated to Methanosarcina were slightly more abundant insoil under oil palm cultivation (Figure 7) whereas sequencesbelonging to the Rice Cluster I were more abundant in nativeforest soil

In the Thaumarchaeota phylum Candidatus Nitrosos-phaera an AOA member of group I1b [65] was detectedin both samples but had a much higher representation innative forest (native forest 182 oil palm-cultivated area103) (Figure 7) In a comparative study of the archaealcommunities in Amazonian dark earth (ldquoTerra Pretardquo)and adjacent soils Candidatus Nitrososphaera was themain archaeal group found in both soils [10] In contrastCandidatus Nitrosotalea another AOA was more abundantin the oil palm-cultivated area (100) than in native forestsoil (44) in our study (Figure 7) Lehtovirta-Morley et al[73] cultivated the obligate acidophilic ammonia oxidizerCandidatus Nitrosotalea devanaterra from a nitrifying acidicsoil Later the same group characterized two strains ofacidophilic AOA belonging to this genus from a Chinesepaddy field and a Scottish agricultural soil [74] In our studythe greater representation of Candidatus Nitrososphaeraand Candidatus Nitrosotalea in native forest and oil

Archaea 11

palm-cultivated soils respectively can be explained by thelow soil pH (native forest pH 550 oil palm-cultivated areapH 483) since in basic soils the activity and growth ofthese archaea are diminished by the exponential decrease innitrite [6 75] In addition soil under oil palm cultivationis fertilized with nitrogen which may explain the higherlevel of Candidatus Nitrosotalea in this area compared withthat of native forest soil Finally although the cultivatedmembers of the genus Ca Nitrososphaera grow betteron neutral pH they have been reported to grow on pHvalues ranging from 60 to 85 In addition a recent workpublished by Wang et al (2014) [76] described active AOAsfrom an acidic soil (pH 492) which are closely related toNitrososphaera known as Nitrososphaera-like organismsby molecular phylogenetic methods Most of our sequenceswere classified as Nitrosotalea-like which are sequencesbelonging to the group I1a This group is typically foundin aquatic environments such as lake sediments or marineenvironments A similar ubiquitous distribution in soils isfound for the group I1b which are not restricted to a specificsoil pH condition

5 Conclusions

Although archaea are known to be ubiquitous there is stillmuch to be learned about their diversity biological rolesand metabolic capabilities This work is the first to comparearchaeal communities associated with the Amazon forest soilwith those of soil under oil palm cultivation

Archaeal diversity was found to be lower in oil palm-cultivated soil than in the native forest soil Only about 30of the OTUs were found in the oil palm soil in comparisonto the native forest However two main phyla were detectedin both samples Thaumarchaeota and Euryarchaeota with apredominance ofThaumarchaeota in native forest soil Theseresults support previous studies in other Brazilian biomenative soils (eg Caatinga Cerrado) as well as soil underdifferent types of land use (eg pasture and agricultural) Itis also important to mention that there are also groups ofuncertain classification associated with the Crenarchaeotaphylum

Although knowledge about Archaea has increased dra-matically in the last few decades little is known aboutthe biology of many archaeal groups and the taxonomyof Archaea is still a work in progress Thus it is notsurprising thatmany sequences identified in thiswork remainunclassified sometimes even at higher taxonomic levels Itis not clear whether the relatively small overall differencebetween soil archaeal communities of the Amazon forest andoil palm-cultivated land is a consequence of the current stateof archaeal taxonomy or whether it indicates that archaealdiversity is lower than bacterial diversity [77]

It is also important to consider that with land usechange due to agricultural practices the microstructure ofthe soil is broken down which can reduce the natural habitatfragmentation allowing potential interactions among theresident microbiota that otherwise would not happen [7879]

A phylogenetic tree of unclassified Thaumarchaeota 16SrRNA gene sequences was useful in exploring relationshipsamong these sequences When taxonomic classification ofsequences was possible it revealed the presence of OTUsfrom genera known to be involved in the metabolism ofcarbon and nitrogen (methanogens Methanocella and Meth-anosarcina AOACandidatusNitrososphaera andCandidatusNitrosotalea) hinting at possible roles for these archaealcommunities in Amazon soils

More studies are needed to elucidate these roles Cultur-ing some of these archaea in the laboratory would allow theirmetabolism and physiology to be studied in detail allowingfor a better understanding of the roles of Archaea in the bio-geochemical cycles

Conflict of Interests

The authors declare that there is no conflict of interestsregarding the publication of this paper

Acknowledgments

This work was supported by grants from FINEP CNPqCAPES and FAP-DF The authors thank Dr Boari for helpwith sample collection logistics Dr Betania Ferraz Quirinowas supported by the Cientista Visitante program fromEmbrapa

References

[1] FAO Global Forest Resources Assessment Food and AgricultureOrganization of the United Nations 2010

[2] IBGE ldquoMapa de Biomas do Brasil primeira aproximacaordquo Riode Janeiro Brazil 2004 httpwwwibgegovbrhome

[3] C Azevedo-Ramos ldquoSustainable development and challengingdeforestation in the Brazilian Amazon the good the badand the uglyrdquo in Proceedings of the Symposium Our CommonGround Innovations in Land Use Decision-Making vol 59 p 5Vancouver Canada May 2007

[4] C A Francis K J Roberts J M Beman A E Santoro and B BOakley ldquoUbiquity and diversity of ammonia-oxidizing archaeainwater columns and sediments of the oceanrdquoProceedings of theNational Academy of Sciences of theUnited States of America vol102 no 41 pp 14683ndash14688 2005

[5] A H Treusch S Leininger A Kietzin S C Schuster H-PKlenk and C Schleper ldquoNovel genes for nitrite reductase andAmo-related proteins indicate a role of uncultivatedmesophiliccrenarchaeota in nitrogen cyclingrdquo Environmental Microbiol-ogy vol 7 no 12 pp 1985ndash1995 2005

[6] G W Nicol S Leininger C Schleper and J I Prosser ldquoTheinfluence of soil pH on the diversity abundance and tran-scriptional activity of ammonia oxidizing archaea and bacteriardquoEnvironmentalMicrobiology vol 10 no 11 pp 2966ndash2978 2008

[7] P Offre A Spang and C Schleper ldquoArchaea in biogeochemicalcyclesrdquo Annual Review of Microbiology vol 67 pp 437ndash4572013

[8] MAPA Anuario Estatıstico da Agroenergia 2012 StatisticalYearbook of Agroenergy 2013

[9] J C Bergmann D D Tupinamba O Y A Costa J R MAlmeida C C Barreto andB FQuirino ldquoBiodiesel production

12 Archaea

in Brazil and alternative biomass feedstocksrdquo Renewable ampSustainable Energy Reviews vol 21 pp 411ndash420 2013

[10] R G Taketani and S M Tsai ldquoThe influence of different landuses on the structure of archaeal communities in Amazoniananthrosols based on 16S rRNA and amoA genesrdquo MicrobialEcology vol 59 no 4 pp 734ndash743 2010

[11] C Brochier-Armanet B Boussau S Gribaldo and P ForterreldquoMesophilic crenarchaeota proposal for a third archaeal phy-lum theThaumarchaeotardquoNature Reviews Microbiology vol 6no 3 pp 245ndash252 2008

[12] S M Barns C F Delwiche J D Palmer and N R PaceldquoPerspectives on archaeal diversity thermophily and mono-phyly from environmental rRNA sequencesrdquo Proceedings of theNational Academy of Sciences of the United States of Americavol 93 no 17 pp 9188ndash9193 1996

[13] J G Elkins M Podar D E Graham et al ldquoA korarchaealgenome reveals insights into the evolution of the Archaeardquo Pro-ceedings of the National Academy of Sciences of the United Statesof America vol 105 no 23 pp 8102ndash8107 2008

[14] H Huber M J Hohn R Rachel T Fuchs V C Wimmer andK O Stetter ldquoA new phylum of Archaea represented by ananosized hyperthermophilic symbiontrdquo Nature vol 417 no6884 pp 63ndash67 2002

[15] T Nunoura H Oida M Nakaseama et al ldquoArchaeal diversityand distribution along thermal and geochemical gradients inhydrothermal sediments at the Yonaguni Knoll IV hydrother-mal field in the SouthernOkinawa TroughrdquoApplied and Enviro-nmental Microbiology vol 76 no 4 pp 1198ndash1211 2010

[16] C Rinke P Schwientek A Sczyrba et al ldquoInsights into thephylogeny and coding potential of microbial dark matterrdquoNature vol 499 no 7459 pp 431ndash437 2013

[17] A Spang J H Saw S L Joslashrgensen et al ldquoComplex archaea thatbridge the gap between prokaryotes and eukaryotesrdquo Naturevol 521 no 7551 pp 173ndash179 2015

[18] S Bagchi S E Vlaeminck L A Sauder et al ldquoTemporal andspatial stability of ammonia-oxidizing archaea and bacteria inaquariumbiofiltersrdquo PLoSONE vol 9 no 12 Article ID e1135152014

[19] W Qin S A Amin W Martens-Habbena et al ldquoMarineammonia-oxidizing archaeal isolates display obligate mixotro-phy and wide ecotypic variationrdquo Proceedings of the NationalAcademy of Sciences of the United States of America vol 111 no34 pp 12504ndash12509 2014

[20] M Coci N Odermatt M M Salcher J Pernthaler and GCorno ldquoEcology and distribution ofThaumarchaea in the deephypolimnion of Lake Maggiorerdquo Archaea vol 2015 Article ID590434 11 pages 2015

[21] J G Ferry ldquoHow tomake a living by exhalingmethanerdquoAnnualReview of Microbiology vol 64 pp 453ndash473 2010

[22] A Meyer A Focks V Radl G Welzl I Schoning and MSchloter ldquoInfluence of land use intensity on the diversity ofammonia oxidizing bacteria and archaea in soils from grasslandecosystemsrdquoMicrobial Ecology vol 67 no 1 pp 161ndash166 2014

[23] V De Gannes G Eudoxie and W J Hickey ldquoImpacts ofedaphic factors on communities of ammonia-oxidizing archaeaammonia-oxidizing bacteria and nitrification in tropical soilsrdquoPLoS ONE vol 9 no 2 Article ID e89568 2014

[24] D S Ribeiro F C J da Silva and TN Carvalho ldquoSobrevivenciade seis especies florestais em uma area explorada seletivamenteno municıpio de Moju Parardquo CERNE vol 9 no 2 pp 153ndash1632003

[25] D HM Costa C A P Ferreira J NM Silva J D C A Lopesand J O P D Carvalho ldquoPotencial madeireiro de floresta densano municıpio de Moju Estado do Parardquo Documentos 1998

[26] C M Almeida S F Lima C V Martins-Da-Silva and JI Gomes ldquoCaracterizacao morfologica e anatomica de dezespecies de leguminosae ocorrentes em uma floresta tropicalumida localizada no municıpio de Moju Estado do Parardquo inA Silvicultura na Amazonia Oriental Contribuicoes do ProjetoEmbrapa-DFID J NMC Silva JO P deCarvalho and J AGYared Eds pp 19ndash54 Embrapa Amazonia Oriental (CPATU)Belem Brazil 2001

[27] K Smalla N Cresswell L C Mendoncahagler A Woltersand J D Vanelsas ldquoRapid extraction protocol from soil forpolymerase chain reaction-mediated amplificationrdquo Journal ofApplied Bacteriology vol 74 pp 78ndash85 1993

[28] S Gantner A F Andersson L Alonso-Saez and S Bertils-son ldquoNovel primers for 16S rRNA-based archaeal communityanalyses in environmental samplesrdquo Journal of MicrobiologicalMethods vol 84 no 1 pp 12ndash18 2011

[29] P D Schloss S L Westcott T Ryabin et al ldquoIntroducingMOTHUR open-source platform-independent community-supported software for describing and comparing microbialcommunitiesrdquoApplied and EnvironmentalMicrobiology vol 75no 23 pp 7537ndash7541 2009

[30] PD SchlossDGevers and S LWestcott ldquoReducing the effectsof PCR amplification and sequencing artifacts on 16S rRNA-based studiesrdquo PLoS ONE vol 6 no 12 Article ID e27310 2011

[31] CQuince A Lanzen T P Curtis et al ldquoAccurate determinationof microbial diversity from 454 pyrosequencing datardquo NatureMethods vol 6 no 9 pp 639ndash641 2009

[32] P Yilmaz L W Parfrey P Yarza et al ldquoThe SILVA and lsquoall-species Living Tree Project (LTP)rsquo taxonomic frameworksrdquoNucleic Acids Research vol 42 no 1 pp D643ndashD648 2014

[33] R C Edgar B J Haas J C Clemente C Quince and RKnight ldquoUCHIME improves sensitivity and speed of chimeradetectionrdquo Bioinformatics vol 27 no 16 pp 2194ndash2200 2011

[34] A Chao and T-J Shen ldquoNonparametric estimation of Shan-nonrsquos index of diversity when there are unseen species insamplerdquo Environmental and Ecological Statistics vol 10 no 4pp 429ndash443 2003

[35] A Chao and T-J Shen Program SPADE (Species Prediction andDiversity Estimation) 2010

[36] C L Shannon andWWeaverTheMathematicalTheory of Com-munication University of Illinois Press Urbana Ill USA 1949

[37] I J Good ldquoThe population frequencies of species and theestimation of population parametersrdquo Biometrika vol 40 pp237ndash264 1953

[38] K Tamura G Stecher D Peterson A Filipski and S KumarldquoMEGA6molecular evolutionary genetics analysis version 60rdquoMolecular Biology and Evolution vol 30 no 12 pp 2725ndash27292013

[39] DA BensonMCavanaugh K Clark et al ldquoGenBankrdquoNucleicAcids Research vol 41 no 1 pp D36ndashD42 2013

[40] D H Parks G W Tyson P Hugenholtz and R G BeikoldquoSTAMP statistical analysis of taxonomic and functional pro-filesrdquo Bioinformatics vol 30 no 21 pp 3123ndash3124 2014

[41] R G Newcombe ldquoInterval estimation for the differencebetween independent proportions comparison of elevenmeth-odsrdquo Statistics in Medicine vol 17 no 8 pp 873ndash890 1998

[42] J D Storey J E Taylor and D Siegmund ldquoStrong controlconservative point estimation and simultaneous conservative

Archaea 13

consistency of false discovery rates a unified approachrdquo Journalof the Royal Statistical Society Series B Statistical Methodologyvol 66 no 1 pp 187ndash205 2004

[43] G C Baker J J Smith and D A Cowan ldquoReview and re-analysis of domain-specific 16S primersrdquo Journal of Microbio-logical Methods vol 55 no 3 pp 541ndash555 2003

[44] W H Zhong and Z C Cai ldquoLong-term effects of inorganicfertilizers on microbial biomass and community functionaldiversity in a paddy soil derived from quaternary red clayrdquoApplied Soil Ecology vol 36 no 2-3 pp 84ndash91 2007

[45] EWessen S Hallin and L Philippot ldquoDifferential responses ofbacterial and archaeal groups at high taxonomical ranks to soilmanagementrdquo Soil Biology and Biochemistry vol 42 no 10 pp1759ndash1765 2010

[46] ZWuW Lin B Li et al ldquoTerminal restriction fragment lengthpolymorphism analysis of soil bacterial communities underdifferent vegetation types in subtropical area rdquo PLOS ONE vol10 no 6 Article ID e0129397 2015

[47] K Cenciani M R Lambais C C Cerri L C B De Azevedoand B J Feigl ldquoBacteria diversity and microbial biomass inforest pasture and fallow soils in the southwestern Amazonbasinrdquo Revista Brasileira de Ciencia do Solo vol 33 no 4 pp907ndash916 2009

[48] A A Navarrete R G Taketani L WMendes F de Souza Can-navan F M de Souza Moreira and S M Tsai ldquoLand-use sys-tems affect Archaeal community structure and functional diver-sity in western Amazon soilsrdquo Revista Brasileira de Ciencia doSolo vol 35 no 5 pp 1527ndash1540 2011

[49] E Lumini A Orgiazzi R Borriello P Bonfante and V Bian-ciotto ldquoDisclosing arbuscular mycorrhizal fungal biodiversityin soil through a land-use gradient using a pyrosequencingapproachrdquo Environmental Microbiology vol 12 no 8 pp 2165ndash2179 2010

[50] M Lupatini R J S Jacques Z I Antoniolli A K A SuleimanR R Fulthorpe and L F W Roesch ldquoLand-use change andsoil type are drivers of fungal and archaeal communities in thePampa biomerdquo World Journal of Microbiology and Biotechnol-ogy vol 29 no 2 pp 223ndash233 2013

[51] E G Lamb N Kennedy and S D Siciliano ldquoEffects of plantspecies richness and evenness on soil microbial communitydiversity and functionrdquo Plant and Soil vol 338 no 1 pp 483ndash495 2011

[52] A Spang R Hatzenpichler C Brochier-Armanet et al ldquoDis-tinct gene set in two different lineages of ammonia-oxidizingarchaea supports the phylum Thaumarchaeotardquo Trends inMicrobiology vol 18 no 8 pp 331ndash340 2010

[53] R G Pacchioni F M Carvalho C E Thompson et al ldquoTax-onomic and functional profiles of soil samples from Atlanticforest and Caatinga biomes in northeastern Brazilrdquo Microbio-logyOpen vol 3 no 3 pp 299ndash315 2014

[54] E Catao A P Castro C C Barreto R H Kruger and C MKyaw ldquoDiversity of Archaea in Brazilian savanna soilsrdquoArchivesof Microbiology vol 195 no 7 pp 507ndash512 2013

[55] M Pester T Rattei S Flechl et al ldquoAmoA-based consensusphylogeny of ammonia-oxidizing archaea and deep sequencingof amoA genes from soils of four different geographic regionsrdquoEnvironmental Microbiology vol 14 no 2 pp 525ndash539 2012

[56] J-Z He H-W Hu and L-M Zhang ldquoCurrent insights into theautotrophic thaumarchaeal ammonia oxidation in acidic soilsrdquoSoil Biology amp Biochemistry vol 55 pp 146ndash154 2012

[57] L-M Zhang H-W Hu J-P Shen and J-Z He ldquoAmmonia-oxidizing archaea have more important role than ammonia-oxidizing bacteria in ammonia oxidation of strongly acidicsoilsrdquo ISME Journal vol 6 no 5 pp 1032ndash1045 2012

[58] S Leininger T Urich M Schloter et al ldquoArchaea predominateamong ammonia-oxidizing prokaryotes in soilsrdquo Nature vol442 no 7104 pp 806ndash809 2006

[59] N C Banning L D Maccarone L M Fisk and D VMurphy ldquoAmmonia-oxidising bacteria not archaea dominatenitrification activity in semi-arid agricultural soilrdquo ScientificReports vol 5 Article ID 11146 2015

[60] M Herrmann A Hadrich and K Kusel ldquoPredominance ofthaumarchaeal ammonia oxidizer abundance and transcrip-tional activity in an acidic fenrdquoEnvironmentalMicrobiology vol14 no 11 pp 3013ndash3025 2012

[61] K Zhalnina P Dorr de Quadros F O Camargo and E WTriplett ldquoDrivers of archaeal ammonia-oxidizing communitiesin soilrdquo Frontiers in Microbiology vol 3 article 210 Article IDArticle 210 2012

[62] J Hong J Cho and Y Hong ldquoEnvironmental variables shapingthe ecological niche of Thaumarchaeota in soil direct andindirect causal effectsrdquo PLoS ONE vol 10 no 8 Article IDe0133763 2015

[63] G Jurgens K Lindstrom and A Saano ldquoNovel group withinthe kingdom Crenarchaeota from boreal forest soilrdquo Appliedand Environmental Microbiology vol 63 no 2 pp 803ndash8051997

[64] M Tourna M Stieglmeier A Spang et al ldquoNitrososphaeraviennensis an ammonia oxidizing archaeon from soilrdquoProceed-ings of the National Academy of Sciences of the United States ofAmerica vol 108 no 20 pp 8420ndash8425 2011

[65] K V Zhalnina R Dias M T Leonard et al ldquoGenome sequenceof Candidatus Nitrososphaera evergladensis from group I1benriched from everglades soil reveals novel genomic features ofthe ammonia-oxidizing archaeardquoPLoSONE vol 9 no 7 ArticleID e101648 2014

[66] I Anderson L E Ulrich B Lupa et al ldquoGenomic char-acterization of methanomicrobiales reveals three classes ofmethanogensrdquo PLoS ONE vol 4 no 6 Article ID e5797 2009

[67] J E Galagan C Nusbaum A Roy et al ldquoThe genome of Macetivorans reveals extensive metabolic and physiological diver-sityrdquo Genome Research vol 12 no 4 pp 532ndash542 2002

[68] C Wrede S Kokoschka A Dreier C Heller J Reitner and MHoppert ldquoDeposition of biogenic iron minerals in a methaneoxidizing microbial matrdquo Archaea vol 2013 Article ID 1029728 pages 2013

[69] G Garrity Bergeyrsquo Manual of Systematic Bacteriology 721 2001[70] S Sakai Y Takaki S Shimamura et al ldquoGenome sequence

of a mesophilic hydrogenotrophic methanogen Methanocellapaludicola the first cultivated representative of the orderMethanocellalesrdquo PLoS ONE vol 6 no 7 Article ID e228982011

[71] E Bapteste C Brochier and Y Boucher ldquoHigher-level clas-sification of the Archaea evolution of methanogenesis andmethanogensrdquo Archaea vol 1 no 5 pp 353ndash363 2005

[72] R G E Murray and E Stackebrandt ldquoTaxonomic note imple-mentation of the provisional status Candidatus for incompletelydescribed procaryotesrdquo International Journal of Systematic Bac-teriology vol 45 no 1 pp 186ndash187 1995

[73] L E Lehtovirta-Morley K Stoecker A Vilcinskas J I Prosserand G W Nicol ldquoCultivation of an obligate acidophilic ammo-nia oxidizer from a nitrifying acid soilrdquo Proceedings of the

14 Archaea

National Academy of Sciences of the United States of Americavol 108 no 38 pp 15892ndash15897 2011

[74] L E Lehtovirta-Morley C Ge J Ross H Yao G W Nicol andJ I Prosser ldquoCharacterisation of terrestrial acidophilic archaealammonia oxidisers and their inhibition and stimulation byorganic compoundsrdquo FEMSMicrobiology Ecology vol 89 no 3pp 542ndash552 2015

[75] M J Frijlink T Abee H J Laanbroek W de Boer and W NKonings ldquoThe bioenergetics of ammonia and hydroxylamineoxidation in Nitrosomonas europaea at acid and alkaline pHrdquoArchives of Microbiology vol 157 no 2 pp 194ndash199 1992

[76] B Wang Y Zheng R Huang et al ldquoActive ammonia oxidizersin an acidic soil are phylogenetically closely related to neu-trophilic archaeonrdquo Applied and Environmental Microbiologyvol 80 no 5 pp 1684ndash1691 2014

[77] J Y Aller and P F Kemp ldquoAre Archaea inherently less diversethan Bacteria in the same environmentsrdquo FEMS MicrobiologyEcology vol 65 no 1 pp 74ndash87 2008

[78] K Potthast U Hamer and FMakeschin ldquoLand-use change in atropical mountain rainforest region of southern Ecuador affectssoilmicroorganisms and nutrient cyclingrdquoBiogeochemistry vol111 no 1ndash3 pp 151ndash167 2012

[79] J I Prosser and G W Nicol ldquoArchaeal and bacterial ammonia-oxidisers in soil the quest for niche specialisation and differen-tiationrdquoTrends inMicrobiology vol 20 no 11 pp 523ndash531 2012

Submit your manuscripts athttpwwwhindawicom

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

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Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporation httpwwwhindawicom

International Journal of

Volume 2014

Zoology

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Molecular Biology International

GenomicsInternational Journal of

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The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioinformaticsAdvances in

Marine BiologyJournal of

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Signal TransductionJournal of

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Evolutionary BiologyInternational Journal of

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ArchaeaHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

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Enzyme Research

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International Journal of

Microbiology

Archaea 5

Venn diagram at distance 003

123Cultivated area

691Native forest

130

Figure 2 Venn diagram depicting the number of shared and unique operational taxonomic units (OTUs) in native forest and oil palm-cultivated area

Thaumarchaeota

Euryarchaeota

Unclassified archaea

Cultivated areaNative forest

10 20 30 40 50 60 700()

Figure 3 Comparison of archaeal phyla present in soil samples from native Amazon forest (orange) and an area under oil palm cultivation(blue) The percentages of unclassified sequences are shown

sequences (not shown) The predominant class from thephylum Thaumarchaeota in the oil palm-cultivated areawas Terrestrial Group representing 14 of the total OTUs(Figure 4(a)) and 35 of all Thaumarchaeota sequences (notshown)

Because it is a newer phylum Thaumarchaeota have fewcultivated representatives compared with Euryarchaeota andfewer sequences in databases such as SILVA To determinehow the Thaumarchaeota sequences that could not be clas-sified at the genus level were related to each other a phylo-genetic tree was constructed with representative sequences(Figure 5(a)) One native forest OTU clustered with GroupI1b (NF6) and one native forest OTU (NF17) and two cul-tivated area OTUs (CA50 CA52) clustered with Group I1cThe remaining OTUs formed different clusters all associatedwith the Miscellaneous Crenarchaeotic Group Most unclas-sified Thaumarchaeota sequences from the native forest andoil palm-cultivated area grouped together although a fewunclassified OTUs from the cultivated area (eg CA51 toCA15 and CA24 to CA33) and native forest (eg NF21 toNF49 and NF5 to NF35) formed separate clusters

The unclassified sequences at genus level in the Euryar-chaeota phylum were all related to the orders Methano-cellales Methanosarcinales Methanomicrobiales Thermo-plasmatales Methanobacteriales and Halobacteriales(Figure 5(b)) The classified sequences at genus level wereMethanocella Methanosarcina Methanoculleus Methano-massiliicoccus Methanomethylophilus MethanobrevibacterMethanobacterium and Candidatus lainarchaeum(Figure 5(b))

The genus-level analysis showed that unclassified SoilCrenarchaeotic Group was predominant in the native forestaccounting for 21 of all native forest OTUs (Figure 6(a))In the oil palm-cultivated area the predominant genus wasUnclassified Terrestrial Group accounting for 14 of allcultivated area OTUs (Figure 6(b))

A total of 31 groups of OTUs were identified (onlymajor groups are shown) most of which are unclassi-fied The classified groups in the phylum Euryarchaeotaare Candidatus lainarchaeum Methanobacterium Methano-brevibacter Methanocella Rice Cluster I Methanoculleus

6 Archaea

Marine Group I

Miscellaneous Crenarchaeotic Group

Soil Crenarchaeotic Group

South African Gold Mine Gp 1

Terrestrial Group

Unclassified Thaumarchaeota

OPPD003

Cultivated areaNative forest

5 10 15 20 25 300()

(a)

Methanobacteria

Methanomicrobia

Halobacteria

Thermoplasmata

Unclassified Euryarchaeota

Cultivated areaNative forest

5 10 15 20 250()

(b)

Figure 4 Comparison of archaeal classes present in soil samples from native Amazon forest (orange) and an area under oil palm cultivation(blue) Classes within the phyla Thaumarchaeota (a) and Euryarchaeota (b) are shown as well as the percentage of unclassified sequences

Methanoregula Methanimicrococcus Methanosarcina Can-didatus Methanomethylophilus and Methanomassiliicoccus(Figure 6(b)) The classified groups in the phylum Thau-marchaeota are Candidatus Nitrososphaera and CandidatusNitrosotalea (Figure 6(a))

The analysis of the genera that differ significantly inrelative abundance between native forest soil and the oilpalm-cultivated area revealed that the greatest difference wasseen for Candidatus Nitrososphaera (Figure 7) This genuswas predominant in the native forest soil accounting for182 of the sequences but represented only 103 of thesequences from the oil palm-cultivated area UnclassifiedSoil Crenarchaeotic Group was also predominant in thenative forest (ie 388 of sequences in the native forest

and 312 of sequences in the oil palm-cultivated area)whereas unclassified South African Gold Mine Gp 1 waspredominant in the oil palm-cultivated area (ie 73 ofsequences in the native forest and 135 of sequences inthe oil palm-cultivated area) Candidatus Nitrosotalea repre-sented 44 of the native forest sequences and 100 of theoil palm-cultivated area sequences The relative abundanceof unclassified Terrestrial Miscellaneous Group unclassifiedThaumarchaeotaunclassifiedThermoplasmatalesunclassifiedMiscellaneous Crenarchaeotic Group Methanosarcina andRice Cluster I also differed significantly between native forestsoil and soils under oil palm cultivation but these differenceswere smaller

Archaea 7

FJ4853071| Uncultured archaeon clone

FJ4852991| Uncultured archaeon clone

FJ9027141| Uncultured archaeon clone

KC8313951| Uncultured archaeon clone

KC5103331| Uncultured archaeon clone

JX9848481| Uncultured archaeon clone

AM9107821| Uncultured archaeon clone

Uncultured archaeon acidic red soil-FJ174727

Ca Nitrososphaera gargensis-GU797786

Ca Nitrososphaeramdash(48 6)

Ca Nitrosotaleamdash(49 12)

Candidatus lainarchaeum (0 2)

Methanobacterium (2 3)

Methanomassillicoccus (6 2)

Methanocella (33 8)

Rice cluster I (43 25)

Methanosarcina baltica-AB973356

Methanocella paludicola-NR074192

Pyrobaculum aerophilum

-NR102764

Acidobacteria bacterium KBS 96-FJ870384

Ca Methanom

ethylophilus (12 0)

Ca Nitrososphaera sp-FR773157

Nitrosopumilus maritimus-JQ346765

Halococcus hamelinensis-LN651155

Pyrococcus furiosus-PFU20163

Cenarchaeum symbiosum-CSU51469

Methanoregula formicica-NR112877

FR7451211| Uncultured soil archaeon

Boreal forest soil archaeon-X96688

Trembling aspen archaeon-EF021427

MCG

MCG005

CA4099

99

7174

63

86

9687

9067

5696

6768 54

57

62

58

65

7559

62

63

65

64

52

64

64

1008654

91

99

999293

53798392

88

7274

5164

5858

CA7CA32

CA50NF17

NF6

CA52

T I1c

T I1b

T I1a

CA24CA6CA12

CA4

CA33

NF16

NF2

NF35

NF9

NF5NF3

6NF4

1NF3

2

NF1

2

NF4

NF1

NF3

0NF2

3

NF7CA3

CA39

CA37

CA46

CA44

CA2

CA42

CA27

Unclassified MCGmdash(16 4)

Unclassified Terrestrial Group (94 36)

Unclassified Marine Group I (2 2)

CA15

CA25

CA30

CA43

CA35

CA14

CA29

CA13

CA20

CA11

CA41

CA36

CA51

NF48

CA9NF4

0NF4

2CA21

CA22

NF38

CA45

CA16

CA47

NF31CA31NF28NF47NF27NF45CA18NF26

NF24NF33NF19

NF18NF37

NF34NF43NF15NF44NF13

NF22NF29NF11

CA49

CA1

CA19CA8

CA10

CA23CA38

CA54HM0511251| Uncultured

HM0511301| Uncultured archaeon clone

HM0511271| Uncultured archaeon clone

archaeon clone

NF46NF14

NF49NF25N

F10N

F39N

F20N

F21

NF8

NF3

CA48

CA34

CA5

CA28CA26CA17CA53

(E)

(B)

(C)

(a)

CA13

CA14

NF1

2

NF1

4M

ethan

ocell

a (3

8)Meth

anosarcina (3

8 4)

Methanoculleus (2 0)

Methanomassilicoccus (6 2)

Methanomassilicoccus (6 2)

Methanomassilicoccus (6 2)

Methanomethylophilus (6 2)

Methanomethylophilus (6 2)

Methanobrevibacter (1 0)

Methanobacterium

(2 3)

Methanobacterium

(2 3)

Methanobacterium

(2 3)

Methanobacterium

(2 3)

Methanobacterium

(2 3)

Methanosarci

na (38 4)

Methanosarcina (38 4)

Methanosarcina (38 4)

Methanosarcina (38 4)

NF16

CA4

CA15CA20

CA19

NF3NF1

Meth

anoc

ella (

33 8

)

Meth

anoc

ella p

aludi

cola-

NR 0741

92 1

Meth

anosa

rcina

baltic

a AB97

3356

1

Methanoregula formicica NR 112877 1

Halococcus hamelinensis LN651155 1

CA6N

F6NF8

NF19 NF5NF7NF20

Pyrobaculum aerophilum NR 102764 2

Ca Nitrososphaera gargensis G

U797786 1

Ca Nitrososphaera gargensis GU797786 1

Nitrosopumilus maritimus JQ346765 1Cenarchaeum symbiosum U51469 1Uncultured archaeon acidic red soil FJ174727 1

Boreal forest soil archaeon X96688 1

Trembling aspen archaeon EF021427 1

HM051125 1 Uncultured archaeon clone

HM051127 1 Uncultured archaeon clone

HM051130 1 Uncultured archaeon clone

FJ485307 1 Uncultured archaeon clone

FJ902714 1 Uncultured archaeon clone

KC831395 1 archaeon clone

FJ485299 1 archaeon clone

KC510333 1 archaeon clone

FR745121 1 archaeon clone

JX984848 1 archaeon clone

AM910782 1 archaeon clone

Acidobacteria bacterium KBS 96 FJ870384 1

Ca Nitrososphaera sp FR773157 1

CA7

Pyrococcus furiosus U20163 1

005

MCG

Halobacteriales

Cand

idat

us la

inar

chae

um (0

2)

Cand

idat

us la

inar

chae

um (0

2)

Thermoplasmatales

Methanomicrobiales

Methanosarcinalles

Methanocellales

T I1c

T I1a

T I1b

(C)

998785

58

100

100

100100

100

71 84 71

53

54

65

86

94

98

59100

100

78

10051

5477

63 73

99

100

8497100

78

100

100

76956970

100 10

010

0

637551

100

9999

9958

94

Methanobacteriales

(b)

Figure 5 Phylogenetic trees of (a) unclassified Thaumarchaeota 16S rRNA gene sequences from native forest soil and soil under oil palmcultivation and (b) 16S rRNA gene sequences classified as Euryarchaeota from native forest soil and soil under oil palm cultivation Theneighbor-joining method and Jukes-Cantor corrections were used with 1000 bootstrap replicates Bootstrap values above 50 are shownin the trees A representative number of operational taxonomic units (OTUs) were selected at 3 dissimilarity by MOTHUR to avoidredundancies in the phylogenetic trees Randomly chosen OTUs from native forest soil and from soil under oil palm cultivation were usedin the construction of the trees along with archaeal reference sequences (from GenBank) from the following groups Acidobacteria (B)Crenarchaeota (C) Euryarchaeota (E) Miscellaneous Crenarchaeotic Group (MCG) Thaumarchaeota Group I1a (T I1a) ThaumarchaeotaGroup I1b (T I1b) andThaumarchaeota Group I1c (T I1c)The AOAs are highlighted with the symbolX Sampling site is indicated by ldquoNFrdquo(native forest) or ldquoCArdquo (oil palm-cultivated area) followed by the number of the OTUs (NAxx or CAxx) OTU sequences classified to at leastthe class level are shown in bold followed by two numbers in parentheses indicating the number of sequences of that OTU detected in nativeforest soil and the number detected in the oil palm-cultivated area The scale bar represents the 5 estimated sequence divergence

8 Archaea

Unclassified Marine Group I

Unclassified Miscellaneous Crenarchaeotic Group

Unclassified Thaumarchaeota

Candidatus Nitrososphaera

Unclassified Soil Crenarchaeotic Group

Candidatus Nitrosotalea

Unclassified South African Gold Mine Gp 1

Unclassified Terrestrial Group

Cultivated areaNative forest

5 10 15 20 250()

(a)

Unclassified Thermoplasmatales

Unclassified Terrestrial Miscellaneous Group

Rice Cluster I

Methanocella

Methanosarcina

Unclassified Euryarchaeota

Unclassified Halobacteriales

Candidatus Methanomethylophilus

Unclassified Methanosarcinaceae

5 10 15 20 250()

Cultivated areaNative forest

(b)

Figure 6 Percentage of archaeal operational taxonomic units within the Thaumarchaeota (a) and Euryarchaeota (b) phyla classified at thegenus level for native forest soil (orange) and soil under oil palm cultivation (blue) The percentage of unclassified sequences at genus level isshown

4 Discussion

The Amazon forest is known for its macrospecies diversityhowever few studies have addressed its microbial diversityAmong the domains of life Archaea are undoubtedly the lesswell known

In this work almost 600000 archaeal 16S rRNA genesequences were obtained from native forest soil and anadjacent area under oil palm cultivation After processing(ie removal of short sequences sequences with errors low-quality sequences chimeras and possible contaminants) andnormalization 39111 sequences per area remained for anal-ysis This number of sequences provided sufficient coverageof archaeal diversity in both areas (Table 1)The primers used

were based on a large number of 16S rRNA sequences fromthe SILVA database [28] however given that little is knownabout this domain it is unlikely that the archaeal diversitywasfully covered [43]

The main physicochemical differences between soils ofthe native forest and the oil palm-cultivated area were thelevels of phosphorous potassium calcium organic matterand total carbon (Table 2) Correlations between phylogenyand physicochemical parameters were evaluated using PCAand NMDS however no clear clusters or correlations wereobserved (results not shown)

In a study of the long-term effects of inorganic fertilizerson microbial communities Zhong and Cai [44] reportedthat phosphorus application indirectly altered microbial

Archaea 9

Candidatus NitrososphaeraUnclassified Soil Crenarchaeotic Group (SCG)

Unclassified South African Gold Mine Gp 1 (SAGMCG-1)Candidatus Nitrosotalea

Unclassified Terrestreial Miscellaneous Gp (TMEG)Unclassified Thaumarchaeota

Unclassified ThermoplasmatalesUnclassified Miscellaneous Crenarchaeotic Group

MethanosarcinaRice Cluster I

95 confidence intervals

q-v

alue

(cor

rect

ed)

Cultivated areaNative forest

531e minus 10

420 6 8minus4minus6minus8 minus2minus10

Difference between proportions ()38800

Proportion ()

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

Figure 7 Archaeal genera that differ significantly (119902 lt 005) in relative abundance between native forest soil (orange) and soil under oil palmcultivation (blue) For the purpose of statistical analysis performed using STAMP software the sum of the percentages of archaea classifiedat the genus level in both soils is 100 Differences between proportions of each genus are shown with 95 confidence intervals Unclassifiedsequences were used to calculate frequency profiles but are not displayed

parameters in soils and increased crop yields by elevating thelevels of organic matter Wessen and collaborators used real-time PCR quantification to show the impacts of fertilizer useon several bacterial phyla and the archaeal phylum Crenar-chaeota (Thaumarchaeota Group I1c after reanalysis usingthe SILVAdatabase 119) [45]They reported that bacterial andarchaeal classes respond differently to fertilization primarilybecause of changes in certain soil parameters such as pHtotal nitrogen and the carbon nitrogen ratio In our studylevels of total carbon and organic matter were higher in thenative forest than in the oil palm-cultivated area which mayhave contributed to higher archaeal diversity Although thereare no studies on the effect of total soil carbon on archaealcommunities there are studies on the bacterial communitiesin Amazonian soils The work of Edgar et al [33] describeddistinct soil bacterial communities under different vegetationtypes The authors found that bacterial diversity correlatedwith total organic carbon and total nitrogen content [46]Furthermore in a study of bacterial diversity and microbialbiomass in forest pasture and fallow soils in the Amazonbasin microbial biomass was highest in pasture soils whichalso had 30ndash47 higher carbon levels than the other sitesover the course of a year [47]

Several studies have also reported alterations in thecomposition of archaeal [48] bacterial [47] and fungal [49]communities associated with land use changes A studyof archaeal communities in Amazonian anthrosols undervarious land uses (agriculture pasture and secondary forest)and cultivation practices (eg eggplant banana citrus andmanioc) [10] reported that agriculture negatively affectedarchaeal community diversity Results of ribosomal inter-genic spacer analysis showed that soil type is also responsiblefor changes in archaeal communities in soils under differenttypes of land use (native grassland native forest Eucalyptus

and Acacia plantations and soybean and watermelon fields)[50]

The Venn diagram showed that the native forest soil has agreater number of unique sequences (Figure 2) This was notsurprising due to the greater archaeal richness and diversityobserved in the native forest soil (Table 2) The number ofunique OTUs found in the oil palm-cultivated area is almostthe same as that of shared OTUs with the native forest areaAlthough oil palm cultivation decreased archaeal diversity inthe soil there is still a core group of archaea OTUs sharedwith the native forest suggesting that these archaea havecertain characteristics that allow them to persist after thechange in land use It cannot be ruled out that these archaeaare metabolically inactive The smaller number of uniquesequences in the oil palm-cultivated area may be related to alower complexity of the vegetation (ie oil palmmonocultureversus great plant diversity in the forest) since plant diversitycould affect these communities [51]

In our study all OTUs identified belong to the domainArchaea and were classified into two phyla Euryarchaeotaand Thaumarchaeota It is important to mention that thedatabases were only recently updated to recognize the phy-lumThaumarchaeota therefore previous studies on Archaeacould not recognize this phylum as proposed by Brochier-Armanet et al [11] The creation of this novel phylum wassupported by a study comparing the genomes of two marineammonia-oxidizing archaea (AOAs) that identified a core setof informational genes specific to this group [52] Thereforeto allow comparisons with other studies we reanalyzed theirdata whenever possible updating the taxonomic classifica-tion to include the phylumThaumarchaeota

Thework ofChao and Shen [35] focused on the impacts ofland use (ie agricultural systems of indigenous people andcattle pasture) on archaeal communities by PCR (amoA gene)

10 Archaea

and denaturing gradient gel electrophoresis analysis of differ-ent Western Amazon soils The databases at the time did notconsiderThaumarchaeota in their classification however ourreanalysis of their dataset using an updated database showedthat Euryarchaeota and Thaumarchaeota were present intheir samples along with Crenarchaeota (Thermoprotei) Inaddition changes were observed in the structure of soilarchaeal communities when comparing the cultivated areaswith forest areas that had been converted to pasture

In our study differences in the percentages of Thau-marchaeota and Euryarchaeota found in the soil sampleswere not striking Thaumarchaeota were predominant in thenative forest (58) whereas Euryarchaeota were predom-inant in the oil palm-cultivated area with 54 (Figure 3)This result is consistent with other comparative studies onarchaeal communities in different types of Brazilian soils (ieAmazonian anthrosols Caatinga and Cerrado) [10 53 54]and a study of different tropical forest soil types that reportedThaumarchaeota as the predominant phylum [44]

The phylumThaumarchaeota comprises all known AOAsand is directly involved in nitrogen metabolism Ammonia isused as an energy source by Group I1a found predominantlyin aquatic systems [52] and Group I1b found in soils [55]Thaumarchaeota Group I1c is primarily found in acidic soilsbut the ecological roles of these archaea are still unknown [5657]

The literature presents conflicting data about the pro-portion of AOAsAOBs soils In some studies AOAs arepredominant [58] while in others AOBs are more abundant[59] However several studies found a predominance of thearchaeal amoA gene over the bacterial amoA gene in soils[60] AOAs are frequently detected in soils their abundancerange from 05 to 10 of the prokaryotic community andtheir abundance is strongly influenced by physicochemicalconditions as well as the organic matter content [61 62] Inacidic soils AOAs appear to have a more important role inammonia oxidation than AOBs [57]

The Soil Crenarchaeotic Group a class within Thaumar-chaeota according to the SILVA database classification wasdetected in both soil samples but a higher representationwas observed in native forest soil This class was discoveredin boreal forest soil [63] Candidatus Nitrososphaera whichbelongs to the Soil Crenarchaeotic Group class is a memberof Thaumarchaeota Group I1b it is an AOA that usesammonia or urea as an energy source [64 65]

On the other hand members of the phylum Eur-yarchaeota are directly involved in carbon metabolismwith several classes of methanogens and methane-oxidizingorganisms [66ndash68] Navarrete et al [48] studied the impactof converting Amazon native forest to pasture and reportedchanges not only to the composition and abundance of soilmicrobial communities but also to a decrease in the diversityof gene function

Within the phylum Euryarchaeota Methanomicrobiaand Thermoplasmata were the main classes detected in oursamples along with a few representatives of Methanobac-teria and Halobacteria Members of Methanobacteria andMethanomicrobia are methanogens [69] Methanomicrobiawere present in both native forest soil and oil palm-cultivated

soil with larger representation in the oil palm-cultivatedarea Within Methanomicrobia the genera Rice Cluster IMethanosarcina andMethanocellawere present in both sam-ples (Figure 6(b)) These genera are closely related and foundin rice fields that produce methane [70] The order Metha-nomicrobiales was detected only in the native forest soilwhich can probably be explained by the relatively high levelsof carbon and organic matter This archaeal order is respon-sible for the final step of methane production in the carboncycle by anaerobic degradation of organic matter [66 71]However the contribution of the Methanomicrobia to thecarbon cycle in the oil palm-cultivated soils could not beassessed since there are no data on the methane productionin the studied areas

Among the Thaumarchaeota sequences used to generatethe phylogenetic tree that were unclassified at the genuslevel several OTUs formed different clusters associated withtheMiscellaneous Crenarchaeotic Group (Figure 5(a))Thesesequences were from both soil samples indicating that theunclassified Thaumarchaeota in the native forest and the oilpalm-cultivated area are phylogenetically related

Most of the sequences remained unclassified at thegenus level for both Thaumarchaeota and Euryarchaeota(Figures 6(a) and 6(b)) This result likely reflects the fact thatmost taxonomic databases do not have many representativesof Archaea especially belonging to the phylum Thaumar-chaeota thus few classifications of new sequences reach thegenus level Several genera were identified as Candidatusaccording to the International Code of Nomenclature ofBacteria indicating that characterization of the culturedprokaryote is incomplete [72]

The proportions of only four groups classified at thegenus level differed significantly between native forest and oilpalm-cultivated soils (Methanosarcina and Rice Cluster I inthe phylum Euryarchaeota and Candidatus NitrososphaeraandCandidatusNitrosotalea in the phylumThaumarchaeota)(Figure 7) With respect to the methanogens sequencesrelated to Methanosarcina were slightly more abundant insoil under oil palm cultivation (Figure 7) whereas sequencesbelonging to the Rice Cluster I were more abundant in nativeforest soil

In the Thaumarchaeota phylum Candidatus Nitrosos-phaera an AOA member of group I1b [65] was detectedin both samples but had a much higher representation innative forest (native forest 182 oil palm-cultivated area103) (Figure 7) In a comparative study of the archaealcommunities in Amazonian dark earth (ldquoTerra Pretardquo)and adjacent soils Candidatus Nitrososphaera was themain archaeal group found in both soils [10] In contrastCandidatus Nitrosotalea another AOA was more abundantin the oil palm-cultivated area (100) than in native forestsoil (44) in our study (Figure 7) Lehtovirta-Morley et al[73] cultivated the obligate acidophilic ammonia oxidizerCandidatus Nitrosotalea devanaterra from a nitrifying acidicsoil Later the same group characterized two strains ofacidophilic AOA belonging to this genus from a Chinesepaddy field and a Scottish agricultural soil [74] In our studythe greater representation of Candidatus Nitrososphaeraand Candidatus Nitrosotalea in native forest and oil

Archaea 11

palm-cultivated soils respectively can be explained by thelow soil pH (native forest pH 550 oil palm-cultivated areapH 483) since in basic soils the activity and growth ofthese archaea are diminished by the exponential decrease innitrite [6 75] In addition soil under oil palm cultivationis fertilized with nitrogen which may explain the higherlevel of Candidatus Nitrosotalea in this area compared withthat of native forest soil Finally although the cultivatedmembers of the genus Ca Nitrososphaera grow betteron neutral pH they have been reported to grow on pHvalues ranging from 60 to 85 In addition a recent workpublished by Wang et al (2014) [76] described active AOAsfrom an acidic soil (pH 492) which are closely related toNitrososphaera known as Nitrososphaera-like organismsby molecular phylogenetic methods Most of our sequenceswere classified as Nitrosotalea-like which are sequencesbelonging to the group I1a This group is typically foundin aquatic environments such as lake sediments or marineenvironments A similar ubiquitous distribution in soils isfound for the group I1b which are not restricted to a specificsoil pH condition

5 Conclusions

Although archaea are known to be ubiquitous there is stillmuch to be learned about their diversity biological rolesand metabolic capabilities This work is the first to comparearchaeal communities associated with the Amazon forest soilwith those of soil under oil palm cultivation

Archaeal diversity was found to be lower in oil palm-cultivated soil than in the native forest soil Only about 30of the OTUs were found in the oil palm soil in comparisonto the native forest However two main phyla were detectedin both samples Thaumarchaeota and Euryarchaeota with apredominance ofThaumarchaeota in native forest soil Theseresults support previous studies in other Brazilian biomenative soils (eg Caatinga Cerrado) as well as soil underdifferent types of land use (eg pasture and agricultural) Itis also important to mention that there are also groups ofuncertain classification associated with the Crenarchaeotaphylum

Although knowledge about Archaea has increased dra-matically in the last few decades little is known aboutthe biology of many archaeal groups and the taxonomyof Archaea is still a work in progress Thus it is notsurprising thatmany sequences identified in thiswork remainunclassified sometimes even at higher taxonomic levels Itis not clear whether the relatively small overall differencebetween soil archaeal communities of the Amazon forest andoil palm-cultivated land is a consequence of the current stateof archaeal taxonomy or whether it indicates that archaealdiversity is lower than bacterial diversity [77]

It is also important to consider that with land usechange due to agricultural practices the microstructure ofthe soil is broken down which can reduce the natural habitatfragmentation allowing potential interactions among theresident microbiota that otherwise would not happen [7879]

A phylogenetic tree of unclassified Thaumarchaeota 16SrRNA gene sequences was useful in exploring relationshipsamong these sequences When taxonomic classification ofsequences was possible it revealed the presence of OTUsfrom genera known to be involved in the metabolism ofcarbon and nitrogen (methanogens Methanocella and Meth-anosarcina AOACandidatusNitrososphaera andCandidatusNitrosotalea) hinting at possible roles for these archaealcommunities in Amazon soils

More studies are needed to elucidate these roles Cultur-ing some of these archaea in the laboratory would allow theirmetabolism and physiology to be studied in detail allowingfor a better understanding of the roles of Archaea in the bio-geochemical cycles

Conflict of Interests

The authors declare that there is no conflict of interestsregarding the publication of this paper

Acknowledgments

This work was supported by grants from FINEP CNPqCAPES and FAP-DF The authors thank Dr Boari for helpwith sample collection logistics Dr Betania Ferraz Quirinowas supported by the Cientista Visitante program fromEmbrapa

References

[1] FAO Global Forest Resources Assessment Food and AgricultureOrganization of the United Nations 2010

[2] IBGE ldquoMapa de Biomas do Brasil primeira aproximacaordquo Riode Janeiro Brazil 2004 httpwwwibgegovbrhome

[3] C Azevedo-Ramos ldquoSustainable development and challengingdeforestation in the Brazilian Amazon the good the badand the uglyrdquo in Proceedings of the Symposium Our CommonGround Innovations in Land Use Decision-Making vol 59 p 5Vancouver Canada May 2007

[4] C A Francis K J Roberts J M Beman A E Santoro and B BOakley ldquoUbiquity and diversity of ammonia-oxidizing archaeainwater columns and sediments of the oceanrdquoProceedings of theNational Academy of Sciences of theUnited States of America vol102 no 41 pp 14683ndash14688 2005

[5] A H Treusch S Leininger A Kietzin S C Schuster H-PKlenk and C Schleper ldquoNovel genes for nitrite reductase andAmo-related proteins indicate a role of uncultivatedmesophiliccrenarchaeota in nitrogen cyclingrdquo Environmental Microbiol-ogy vol 7 no 12 pp 1985ndash1995 2005

[6] G W Nicol S Leininger C Schleper and J I Prosser ldquoTheinfluence of soil pH on the diversity abundance and tran-scriptional activity of ammonia oxidizing archaea and bacteriardquoEnvironmentalMicrobiology vol 10 no 11 pp 2966ndash2978 2008

[7] P Offre A Spang and C Schleper ldquoArchaea in biogeochemicalcyclesrdquo Annual Review of Microbiology vol 67 pp 437ndash4572013

[8] MAPA Anuario Estatıstico da Agroenergia 2012 StatisticalYearbook of Agroenergy 2013

[9] J C Bergmann D D Tupinamba O Y A Costa J R MAlmeida C C Barreto andB FQuirino ldquoBiodiesel production

12 Archaea

in Brazil and alternative biomass feedstocksrdquo Renewable ampSustainable Energy Reviews vol 21 pp 411ndash420 2013

[10] R G Taketani and S M Tsai ldquoThe influence of different landuses on the structure of archaeal communities in Amazoniananthrosols based on 16S rRNA and amoA genesrdquo MicrobialEcology vol 59 no 4 pp 734ndash743 2010

[11] C Brochier-Armanet B Boussau S Gribaldo and P ForterreldquoMesophilic crenarchaeota proposal for a third archaeal phy-lum theThaumarchaeotardquoNature Reviews Microbiology vol 6no 3 pp 245ndash252 2008

[12] S M Barns C F Delwiche J D Palmer and N R PaceldquoPerspectives on archaeal diversity thermophily and mono-phyly from environmental rRNA sequencesrdquo Proceedings of theNational Academy of Sciences of the United States of Americavol 93 no 17 pp 9188ndash9193 1996

[13] J G Elkins M Podar D E Graham et al ldquoA korarchaealgenome reveals insights into the evolution of the Archaeardquo Pro-ceedings of the National Academy of Sciences of the United Statesof America vol 105 no 23 pp 8102ndash8107 2008

[14] H Huber M J Hohn R Rachel T Fuchs V C Wimmer andK O Stetter ldquoA new phylum of Archaea represented by ananosized hyperthermophilic symbiontrdquo Nature vol 417 no6884 pp 63ndash67 2002

[15] T Nunoura H Oida M Nakaseama et al ldquoArchaeal diversityand distribution along thermal and geochemical gradients inhydrothermal sediments at the Yonaguni Knoll IV hydrother-mal field in the SouthernOkinawa TroughrdquoApplied and Enviro-nmental Microbiology vol 76 no 4 pp 1198ndash1211 2010

[16] C Rinke P Schwientek A Sczyrba et al ldquoInsights into thephylogeny and coding potential of microbial dark matterrdquoNature vol 499 no 7459 pp 431ndash437 2013

[17] A Spang J H Saw S L Joslashrgensen et al ldquoComplex archaea thatbridge the gap between prokaryotes and eukaryotesrdquo Naturevol 521 no 7551 pp 173ndash179 2015

[18] S Bagchi S E Vlaeminck L A Sauder et al ldquoTemporal andspatial stability of ammonia-oxidizing archaea and bacteria inaquariumbiofiltersrdquo PLoSONE vol 9 no 12 Article ID e1135152014

[19] W Qin S A Amin W Martens-Habbena et al ldquoMarineammonia-oxidizing archaeal isolates display obligate mixotro-phy and wide ecotypic variationrdquo Proceedings of the NationalAcademy of Sciences of the United States of America vol 111 no34 pp 12504ndash12509 2014

[20] M Coci N Odermatt M M Salcher J Pernthaler and GCorno ldquoEcology and distribution ofThaumarchaea in the deephypolimnion of Lake Maggiorerdquo Archaea vol 2015 Article ID590434 11 pages 2015

[21] J G Ferry ldquoHow tomake a living by exhalingmethanerdquoAnnualReview of Microbiology vol 64 pp 453ndash473 2010

[22] A Meyer A Focks V Radl G Welzl I Schoning and MSchloter ldquoInfluence of land use intensity on the diversity ofammonia oxidizing bacteria and archaea in soils from grasslandecosystemsrdquoMicrobial Ecology vol 67 no 1 pp 161ndash166 2014

[23] V De Gannes G Eudoxie and W J Hickey ldquoImpacts ofedaphic factors on communities of ammonia-oxidizing archaeaammonia-oxidizing bacteria and nitrification in tropical soilsrdquoPLoS ONE vol 9 no 2 Article ID e89568 2014

[24] D S Ribeiro F C J da Silva and TN Carvalho ldquoSobrevivenciade seis especies florestais em uma area explorada seletivamenteno municıpio de Moju Parardquo CERNE vol 9 no 2 pp 153ndash1632003

[25] D HM Costa C A P Ferreira J NM Silva J D C A Lopesand J O P D Carvalho ldquoPotencial madeireiro de floresta densano municıpio de Moju Estado do Parardquo Documentos 1998

[26] C M Almeida S F Lima C V Martins-Da-Silva and JI Gomes ldquoCaracterizacao morfologica e anatomica de dezespecies de leguminosae ocorrentes em uma floresta tropicalumida localizada no municıpio de Moju Estado do Parardquo inA Silvicultura na Amazonia Oriental Contribuicoes do ProjetoEmbrapa-DFID J NMC Silva JO P deCarvalho and J AGYared Eds pp 19ndash54 Embrapa Amazonia Oriental (CPATU)Belem Brazil 2001

[27] K Smalla N Cresswell L C Mendoncahagler A Woltersand J D Vanelsas ldquoRapid extraction protocol from soil forpolymerase chain reaction-mediated amplificationrdquo Journal ofApplied Bacteriology vol 74 pp 78ndash85 1993

[28] S Gantner A F Andersson L Alonso-Saez and S Bertils-son ldquoNovel primers for 16S rRNA-based archaeal communityanalyses in environmental samplesrdquo Journal of MicrobiologicalMethods vol 84 no 1 pp 12ndash18 2011

[29] P D Schloss S L Westcott T Ryabin et al ldquoIntroducingMOTHUR open-source platform-independent community-supported software for describing and comparing microbialcommunitiesrdquoApplied and EnvironmentalMicrobiology vol 75no 23 pp 7537ndash7541 2009

[30] PD SchlossDGevers and S LWestcott ldquoReducing the effectsof PCR amplification and sequencing artifacts on 16S rRNA-based studiesrdquo PLoS ONE vol 6 no 12 Article ID e27310 2011

[31] CQuince A Lanzen T P Curtis et al ldquoAccurate determinationof microbial diversity from 454 pyrosequencing datardquo NatureMethods vol 6 no 9 pp 639ndash641 2009

[32] P Yilmaz L W Parfrey P Yarza et al ldquoThe SILVA and lsquoall-species Living Tree Project (LTP)rsquo taxonomic frameworksrdquoNucleic Acids Research vol 42 no 1 pp D643ndashD648 2014

[33] R C Edgar B J Haas J C Clemente C Quince and RKnight ldquoUCHIME improves sensitivity and speed of chimeradetectionrdquo Bioinformatics vol 27 no 16 pp 2194ndash2200 2011

[34] A Chao and T-J Shen ldquoNonparametric estimation of Shan-nonrsquos index of diversity when there are unseen species insamplerdquo Environmental and Ecological Statistics vol 10 no 4pp 429ndash443 2003

[35] A Chao and T-J Shen Program SPADE (Species Prediction andDiversity Estimation) 2010

[36] C L Shannon andWWeaverTheMathematicalTheory of Com-munication University of Illinois Press Urbana Ill USA 1949

[37] I J Good ldquoThe population frequencies of species and theestimation of population parametersrdquo Biometrika vol 40 pp237ndash264 1953

[38] K Tamura G Stecher D Peterson A Filipski and S KumarldquoMEGA6molecular evolutionary genetics analysis version 60rdquoMolecular Biology and Evolution vol 30 no 12 pp 2725ndash27292013

[39] DA BensonMCavanaugh K Clark et al ldquoGenBankrdquoNucleicAcids Research vol 41 no 1 pp D36ndashD42 2013

[40] D H Parks G W Tyson P Hugenholtz and R G BeikoldquoSTAMP statistical analysis of taxonomic and functional pro-filesrdquo Bioinformatics vol 30 no 21 pp 3123ndash3124 2014

[41] R G Newcombe ldquoInterval estimation for the differencebetween independent proportions comparison of elevenmeth-odsrdquo Statistics in Medicine vol 17 no 8 pp 873ndash890 1998

[42] J D Storey J E Taylor and D Siegmund ldquoStrong controlconservative point estimation and simultaneous conservative

Archaea 13

consistency of false discovery rates a unified approachrdquo Journalof the Royal Statistical Society Series B Statistical Methodologyvol 66 no 1 pp 187ndash205 2004

[43] G C Baker J J Smith and D A Cowan ldquoReview and re-analysis of domain-specific 16S primersrdquo Journal of Microbio-logical Methods vol 55 no 3 pp 541ndash555 2003

[44] W H Zhong and Z C Cai ldquoLong-term effects of inorganicfertilizers on microbial biomass and community functionaldiversity in a paddy soil derived from quaternary red clayrdquoApplied Soil Ecology vol 36 no 2-3 pp 84ndash91 2007

[45] EWessen S Hallin and L Philippot ldquoDifferential responses ofbacterial and archaeal groups at high taxonomical ranks to soilmanagementrdquo Soil Biology and Biochemistry vol 42 no 10 pp1759ndash1765 2010

[46] ZWuW Lin B Li et al ldquoTerminal restriction fragment lengthpolymorphism analysis of soil bacterial communities underdifferent vegetation types in subtropical area rdquo PLOS ONE vol10 no 6 Article ID e0129397 2015

[47] K Cenciani M R Lambais C C Cerri L C B De Azevedoand B J Feigl ldquoBacteria diversity and microbial biomass inforest pasture and fallow soils in the southwestern Amazonbasinrdquo Revista Brasileira de Ciencia do Solo vol 33 no 4 pp907ndash916 2009

[48] A A Navarrete R G Taketani L WMendes F de Souza Can-navan F M de Souza Moreira and S M Tsai ldquoLand-use sys-tems affect Archaeal community structure and functional diver-sity in western Amazon soilsrdquo Revista Brasileira de Ciencia doSolo vol 35 no 5 pp 1527ndash1540 2011

[49] E Lumini A Orgiazzi R Borriello P Bonfante and V Bian-ciotto ldquoDisclosing arbuscular mycorrhizal fungal biodiversityin soil through a land-use gradient using a pyrosequencingapproachrdquo Environmental Microbiology vol 12 no 8 pp 2165ndash2179 2010

[50] M Lupatini R J S Jacques Z I Antoniolli A K A SuleimanR R Fulthorpe and L F W Roesch ldquoLand-use change andsoil type are drivers of fungal and archaeal communities in thePampa biomerdquo World Journal of Microbiology and Biotechnol-ogy vol 29 no 2 pp 223ndash233 2013

[51] E G Lamb N Kennedy and S D Siciliano ldquoEffects of plantspecies richness and evenness on soil microbial communitydiversity and functionrdquo Plant and Soil vol 338 no 1 pp 483ndash495 2011

[52] A Spang R Hatzenpichler C Brochier-Armanet et al ldquoDis-tinct gene set in two different lineages of ammonia-oxidizingarchaea supports the phylum Thaumarchaeotardquo Trends inMicrobiology vol 18 no 8 pp 331ndash340 2010

[53] R G Pacchioni F M Carvalho C E Thompson et al ldquoTax-onomic and functional profiles of soil samples from Atlanticforest and Caatinga biomes in northeastern Brazilrdquo Microbio-logyOpen vol 3 no 3 pp 299ndash315 2014

[54] E Catao A P Castro C C Barreto R H Kruger and C MKyaw ldquoDiversity of Archaea in Brazilian savanna soilsrdquoArchivesof Microbiology vol 195 no 7 pp 507ndash512 2013

[55] M Pester T Rattei S Flechl et al ldquoAmoA-based consensusphylogeny of ammonia-oxidizing archaea and deep sequencingof amoA genes from soils of four different geographic regionsrdquoEnvironmental Microbiology vol 14 no 2 pp 525ndash539 2012

[56] J-Z He H-W Hu and L-M Zhang ldquoCurrent insights into theautotrophic thaumarchaeal ammonia oxidation in acidic soilsrdquoSoil Biology amp Biochemistry vol 55 pp 146ndash154 2012

[57] L-M Zhang H-W Hu J-P Shen and J-Z He ldquoAmmonia-oxidizing archaea have more important role than ammonia-oxidizing bacteria in ammonia oxidation of strongly acidicsoilsrdquo ISME Journal vol 6 no 5 pp 1032ndash1045 2012

[58] S Leininger T Urich M Schloter et al ldquoArchaea predominateamong ammonia-oxidizing prokaryotes in soilsrdquo Nature vol442 no 7104 pp 806ndash809 2006

[59] N C Banning L D Maccarone L M Fisk and D VMurphy ldquoAmmonia-oxidising bacteria not archaea dominatenitrification activity in semi-arid agricultural soilrdquo ScientificReports vol 5 Article ID 11146 2015

[60] M Herrmann A Hadrich and K Kusel ldquoPredominance ofthaumarchaeal ammonia oxidizer abundance and transcrip-tional activity in an acidic fenrdquoEnvironmentalMicrobiology vol14 no 11 pp 3013ndash3025 2012

[61] K Zhalnina P Dorr de Quadros F O Camargo and E WTriplett ldquoDrivers of archaeal ammonia-oxidizing communitiesin soilrdquo Frontiers in Microbiology vol 3 article 210 Article IDArticle 210 2012

[62] J Hong J Cho and Y Hong ldquoEnvironmental variables shapingthe ecological niche of Thaumarchaeota in soil direct andindirect causal effectsrdquo PLoS ONE vol 10 no 8 Article IDe0133763 2015

[63] G Jurgens K Lindstrom and A Saano ldquoNovel group withinthe kingdom Crenarchaeota from boreal forest soilrdquo Appliedand Environmental Microbiology vol 63 no 2 pp 803ndash8051997

[64] M Tourna M Stieglmeier A Spang et al ldquoNitrososphaeraviennensis an ammonia oxidizing archaeon from soilrdquoProceed-ings of the National Academy of Sciences of the United States ofAmerica vol 108 no 20 pp 8420ndash8425 2011

[65] K V Zhalnina R Dias M T Leonard et al ldquoGenome sequenceof Candidatus Nitrososphaera evergladensis from group I1benriched from everglades soil reveals novel genomic features ofthe ammonia-oxidizing archaeardquoPLoSONE vol 9 no 7 ArticleID e101648 2014

[66] I Anderson L E Ulrich B Lupa et al ldquoGenomic char-acterization of methanomicrobiales reveals three classes ofmethanogensrdquo PLoS ONE vol 4 no 6 Article ID e5797 2009

[67] J E Galagan C Nusbaum A Roy et al ldquoThe genome of Macetivorans reveals extensive metabolic and physiological diver-sityrdquo Genome Research vol 12 no 4 pp 532ndash542 2002

[68] C Wrede S Kokoschka A Dreier C Heller J Reitner and MHoppert ldquoDeposition of biogenic iron minerals in a methaneoxidizing microbial matrdquo Archaea vol 2013 Article ID 1029728 pages 2013

[69] G Garrity Bergeyrsquo Manual of Systematic Bacteriology 721 2001[70] S Sakai Y Takaki S Shimamura et al ldquoGenome sequence

of a mesophilic hydrogenotrophic methanogen Methanocellapaludicola the first cultivated representative of the orderMethanocellalesrdquo PLoS ONE vol 6 no 7 Article ID e228982011

[71] E Bapteste C Brochier and Y Boucher ldquoHigher-level clas-sification of the Archaea evolution of methanogenesis andmethanogensrdquo Archaea vol 1 no 5 pp 353ndash363 2005

[72] R G E Murray and E Stackebrandt ldquoTaxonomic note imple-mentation of the provisional status Candidatus for incompletelydescribed procaryotesrdquo International Journal of Systematic Bac-teriology vol 45 no 1 pp 186ndash187 1995

[73] L E Lehtovirta-Morley K Stoecker A Vilcinskas J I Prosserand G W Nicol ldquoCultivation of an obligate acidophilic ammo-nia oxidizer from a nitrifying acid soilrdquo Proceedings of the

14 Archaea

National Academy of Sciences of the United States of Americavol 108 no 38 pp 15892ndash15897 2011

[74] L E Lehtovirta-Morley C Ge J Ross H Yao G W Nicol andJ I Prosser ldquoCharacterisation of terrestrial acidophilic archaealammonia oxidisers and their inhibition and stimulation byorganic compoundsrdquo FEMSMicrobiology Ecology vol 89 no 3pp 542ndash552 2015

[75] M J Frijlink T Abee H J Laanbroek W de Boer and W NKonings ldquoThe bioenergetics of ammonia and hydroxylamineoxidation in Nitrosomonas europaea at acid and alkaline pHrdquoArchives of Microbiology vol 157 no 2 pp 194ndash199 1992

[76] B Wang Y Zheng R Huang et al ldquoActive ammonia oxidizersin an acidic soil are phylogenetically closely related to neu-trophilic archaeonrdquo Applied and Environmental Microbiologyvol 80 no 5 pp 1684ndash1691 2014

[77] J Y Aller and P F Kemp ldquoAre Archaea inherently less diversethan Bacteria in the same environmentsrdquo FEMS MicrobiologyEcology vol 65 no 1 pp 74ndash87 2008

[78] K Potthast U Hamer and FMakeschin ldquoLand-use change in atropical mountain rainforest region of southern Ecuador affectssoilmicroorganisms and nutrient cyclingrdquoBiogeochemistry vol111 no 1ndash3 pp 151ndash167 2012

[79] J I Prosser and G W Nicol ldquoArchaeal and bacterial ammonia-oxidisers in soil the quest for niche specialisation and differen-tiationrdquoTrends inMicrobiology vol 20 no 11 pp 523ndash531 2012

Submit your manuscripts athttpwwwhindawicom

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Anatomy Research International

PeptidesInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporation httpwwwhindawicom

International Journal of

Volume 2014

Zoology

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Molecular Biology International

GenomicsInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioinformaticsAdvances in

Marine BiologyJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Signal TransductionJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioMed Research International

Evolutionary BiologyInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Biochemistry Research International

ArchaeaHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Genetics Research International

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Advances in

Virolog y

Hindawi Publishing Corporationhttpwwwhindawicom

Nucleic AcidsJournal of

Volume 2014

Stem CellsInternational

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Enzyme Research

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

International Journal of

Microbiology

6 Archaea

Marine Group I

Miscellaneous Crenarchaeotic Group

Soil Crenarchaeotic Group

South African Gold Mine Gp 1

Terrestrial Group

Unclassified Thaumarchaeota

OPPD003

Cultivated areaNative forest

5 10 15 20 25 300()

(a)

Methanobacteria

Methanomicrobia

Halobacteria

Thermoplasmata

Unclassified Euryarchaeota

Cultivated areaNative forest

5 10 15 20 250()

(b)

Figure 4 Comparison of archaeal classes present in soil samples from native Amazon forest (orange) and an area under oil palm cultivation(blue) Classes within the phyla Thaumarchaeota (a) and Euryarchaeota (b) are shown as well as the percentage of unclassified sequences

Methanoregula Methanimicrococcus Methanosarcina Can-didatus Methanomethylophilus and Methanomassiliicoccus(Figure 6(b)) The classified groups in the phylum Thau-marchaeota are Candidatus Nitrososphaera and CandidatusNitrosotalea (Figure 6(a))

The analysis of the genera that differ significantly inrelative abundance between native forest soil and the oilpalm-cultivated area revealed that the greatest difference wasseen for Candidatus Nitrososphaera (Figure 7) This genuswas predominant in the native forest soil accounting for182 of the sequences but represented only 103 of thesequences from the oil palm-cultivated area UnclassifiedSoil Crenarchaeotic Group was also predominant in thenative forest (ie 388 of sequences in the native forest

and 312 of sequences in the oil palm-cultivated area)whereas unclassified South African Gold Mine Gp 1 waspredominant in the oil palm-cultivated area (ie 73 ofsequences in the native forest and 135 of sequences inthe oil palm-cultivated area) Candidatus Nitrosotalea repre-sented 44 of the native forest sequences and 100 of theoil palm-cultivated area sequences The relative abundanceof unclassified Terrestrial Miscellaneous Group unclassifiedThaumarchaeotaunclassifiedThermoplasmatalesunclassifiedMiscellaneous Crenarchaeotic Group Methanosarcina andRice Cluster I also differed significantly between native forestsoil and soils under oil palm cultivation but these differenceswere smaller

Archaea 7

FJ4853071| Uncultured archaeon clone

FJ4852991| Uncultured archaeon clone

FJ9027141| Uncultured archaeon clone

KC8313951| Uncultured archaeon clone

KC5103331| Uncultured archaeon clone

JX9848481| Uncultured archaeon clone

AM9107821| Uncultured archaeon clone

Uncultured archaeon acidic red soil-FJ174727

Ca Nitrososphaera gargensis-GU797786

Ca Nitrososphaeramdash(48 6)

Ca Nitrosotaleamdash(49 12)

Candidatus lainarchaeum (0 2)

Methanobacterium (2 3)

Methanomassillicoccus (6 2)

Methanocella (33 8)

Rice cluster I (43 25)

Methanosarcina baltica-AB973356

Methanocella paludicola-NR074192

Pyrobaculum aerophilum

-NR102764

Acidobacteria bacterium KBS 96-FJ870384

Ca Methanom

ethylophilus (12 0)

Ca Nitrososphaera sp-FR773157

Nitrosopumilus maritimus-JQ346765

Halococcus hamelinensis-LN651155

Pyrococcus furiosus-PFU20163

Cenarchaeum symbiosum-CSU51469

Methanoregula formicica-NR112877

FR7451211| Uncultured soil archaeon

Boreal forest soil archaeon-X96688

Trembling aspen archaeon-EF021427

MCG

MCG005

CA4099

99

7174

63

86

9687

9067

5696

6768 54

57

62

58

65

7559

62

63

65

64

52

64

64

1008654

91

99

999293

53798392

88

7274

5164

5858

CA7CA32

CA50NF17

NF6

CA52

T I1c

T I1b

T I1a

CA24CA6CA12

CA4

CA33

NF16

NF2

NF35

NF9

NF5NF3

6NF4

1NF3

2

NF1

2

NF4

NF1

NF3

0NF2

3

NF7CA3

CA39

CA37

CA46

CA44

CA2

CA42

CA27

Unclassified MCGmdash(16 4)

Unclassified Terrestrial Group (94 36)

Unclassified Marine Group I (2 2)

CA15

CA25

CA30

CA43

CA35

CA14

CA29

CA13

CA20

CA11

CA41

CA36

CA51

NF48

CA9NF4

0NF4

2CA21

CA22

NF38

CA45

CA16

CA47

NF31CA31NF28NF47NF27NF45CA18NF26

NF24NF33NF19

NF18NF37

NF34NF43NF15NF44NF13

NF22NF29NF11

CA49

CA1

CA19CA8

CA10

CA23CA38

CA54HM0511251| Uncultured

HM0511301| Uncultured archaeon clone

HM0511271| Uncultured archaeon clone

archaeon clone

NF46NF14

NF49NF25N

F10N

F39N

F20N

F21

NF8

NF3

CA48

CA34

CA5

CA28CA26CA17CA53

(E)

(B)

(C)

(a)

CA13

CA14

NF1

2

NF1

4M

ethan

ocell

a (3

8)Meth

anosarcina (3

8 4)

Methanoculleus (2 0)

Methanomassilicoccus (6 2)

Methanomassilicoccus (6 2)

Methanomassilicoccus (6 2)

Methanomethylophilus (6 2)

Methanomethylophilus (6 2)

Methanobrevibacter (1 0)

Methanobacterium

(2 3)

Methanobacterium

(2 3)

Methanobacterium

(2 3)

Methanobacterium

(2 3)

Methanobacterium

(2 3)

Methanosarci

na (38 4)

Methanosarcina (38 4)

Methanosarcina (38 4)

Methanosarcina (38 4)

NF16

CA4

CA15CA20

CA19

NF3NF1

Meth

anoc

ella (

33 8

)

Meth

anoc

ella p

aludi

cola-

NR 0741

92 1

Meth

anosa

rcina

baltic

a AB97

3356

1

Methanoregula formicica NR 112877 1

Halococcus hamelinensis LN651155 1

CA6N

F6NF8

NF19 NF5NF7NF20

Pyrobaculum aerophilum NR 102764 2

Ca Nitrososphaera gargensis G

U797786 1

Ca Nitrososphaera gargensis GU797786 1

Nitrosopumilus maritimus JQ346765 1Cenarchaeum symbiosum U51469 1Uncultured archaeon acidic red soil FJ174727 1

Boreal forest soil archaeon X96688 1

Trembling aspen archaeon EF021427 1

HM051125 1 Uncultured archaeon clone

HM051127 1 Uncultured archaeon clone

HM051130 1 Uncultured archaeon clone

FJ485307 1 Uncultured archaeon clone

FJ902714 1 Uncultured archaeon clone

KC831395 1 archaeon clone

FJ485299 1 archaeon clone

KC510333 1 archaeon clone

FR745121 1 archaeon clone

JX984848 1 archaeon clone

AM910782 1 archaeon clone

Acidobacteria bacterium KBS 96 FJ870384 1

Ca Nitrososphaera sp FR773157 1

CA7

Pyrococcus furiosus U20163 1

005

MCG

Halobacteriales

Cand

idat

us la

inar

chae

um (0

2)

Cand

idat

us la

inar

chae

um (0

2)

Thermoplasmatales

Methanomicrobiales

Methanosarcinalles

Methanocellales

T I1c

T I1a

T I1b

(C)

998785

58

100

100

100100

100

71 84 71

53

54

65

86

94

98

59100

100

78

10051

5477

63 73

99

100

8497100

78

100

100

76956970

100 10

010

0

637551

100

9999

9958

94

Methanobacteriales

(b)

Figure 5 Phylogenetic trees of (a) unclassified Thaumarchaeota 16S rRNA gene sequences from native forest soil and soil under oil palmcultivation and (b) 16S rRNA gene sequences classified as Euryarchaeota from native forest soil and soil under oil palm cultivation Theneighbor-joining method and Jukes-Cantor corrections were used with 1000 bootstrap replicates Bootstrap values above 50 are shownin the trees A representative number of operational taxonomic units (OTUs) were selected at 3 dissimilarity by MOTHUR to avoidredundancies in the phylogenetic trees Randomly chosen OTUs from native forest soil and from soil under oil palm cultivation were usedin the construction of the trees along with archaeal reference sequences (from GenBank) from the following groups Acidobacteria (B)Crenarchaeota (C) Euryarchaeota (E) Miscellaneous Crenarchaeotic Group (MCG) Thaumarchaeota Group I1a (T I1a) ThaumarchaeotaGroup I1b (T I1b) andThaumarchaeota Group I1c (T I1c)The AOAs are highlighted with the symbolX Sampling site is indicated by ldquoNFrdquo(native forest) or ldquoCArdquo (oil palm-cultivated area) followed by the number of the OTUs (NAxx or CAxx) OTU sequences classified to at leastthe class level are shown in bold followed by two numbers in parentheses indicating the number of sequences of that OTU detected in nativeforest soil and the number detected in the oil palm-cultivated area The scale bar represents the 5 estimated sequence divergence

8 Archaea

Unclassified Marine Group I

Unclassified Miscellaneous Crenarchaeotic Group

Unclassified Thaumarchaeota

Candidatus Nitrososphaera

Unclassified Soil Crenarchaeotic Group

Candidatus Nitrosotalea

Unclassified South African Gold Mine Gp 1

Unclassified Terrestrial Group

Cultivated areaNative forest

5 10 15 20 250()

(a)

Unclassified Thermoplasmatales

Unclassified Terrestrial Miscellaneous Group

Rice Cluster I

Methanocella

Methanosarcina

Unclassified Euryarchaeota

Unclassified Halobacteriales

Candidatus Methanomethylophilus

Unclassified Methanosarcinaceae

5 10 15 20 250()

Cultivated areaNative forest

(b)

Figure 6 Percentage of archaeal operational taxonomic units within the Thaumarchaeota (a) and Euryarchaeota (b) phyla classified at thegenus level for native forest soil (orange) and soil under oil palm cultivation (blue) The percentage of unclassified sequences at genus level isshown

4 Discussion

The Amazon forest is known for its macrospecies diversityhowever few studies have addressed its microbial diversityAmong the domains of life Archaea are undoubtedly the lesswell known

In this work almost 600000 archaeal 16S rRNA genesequences were obtained from native forest soil and anadjacent area under oil palm cultivation After processing(ie removal of short sequences sequences with errors low-quality sequences chimeras and possible contaminants) andnormalization 39111 sequences per area remained for anal-ysis This number of sequences provided sufficient coverageof archaeal diversity in both areas (Table 1)The primers used

were based on a large number of 16S rRNA sequences fromthe SILVA database [28] however given that little is knownabout this domain it is unlikely that the archaeal diversitywasfully covered [43]

The main physicochemical differences between soils ofthe native forest and the oil palm-cultivated area were thelevels of phosphorous potassium calcium organic matterand total carbon (Table 2) Correlations between phylogenyand physicochemical parameters were evaluated using PCAand NMDS however no clear clusters or correlations wereobserved (results not shown)

In a study of the long-term effects of inorganic fertilizerson microbial communities Zhong and Cai [44] reportedthat phosphorus application indirectly altered microbial

Archaea 9

Candidatus NitrososphaeraUnclassified Soil Crenarchaeotic Group (SCG)

Unclassified South African Gold Mine Gp 1 (SAGMCG-1)Candidatus Nitrosotalea

Unclassified Terrestreial Miscellaneous Gp (TMEG)Unclassified Thaumarchaeota

Unclassified ThermoplasmatalesUnclassified Miscellaneous Crenarchaeotic Group

MethanosarcinaRice Cluster I

95 confidence intervals

q-v

alue

(cor

rect

ed)

Cultivated areaNative forest

531e minus 10

420 6 8minus4minus6minus8 minus2minus10

Difference between proportions ()38800

Proportion ()

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

Figure 7 Archaeal genera that differ significantly (119902 lt 005) in relative abundance between native forest soil (orange) and soil under oil palmcultivation (blue) For the purpose of statistical analysis performed using STAMP software the sum of the percentages of archaea classifiedat the genus level in both soils is 100 Differences between proportions of each genus are shown with 95 confidence intervals Unclassifiedsequences were used to calculate frequency profiles but are not displayed

parameters in soils and increased crop yields by elevating thelevels of organic matter Wessen and collaborators used real-time PCR quantification to show the impacts of fertilizer useon several bacterial phyla and the archaeal phylum Crenar-chaeota (Thaumarchaeota Group I1c after reanalysis usingthe SILVAdatabase 119) [45]They reported that bacterial andarchaeal classes respond differently to fertilization primarilybecause of changes in certain soil parameters such as pHtotal nitrogen and the carbon nitrogen ratio In our studylevels of total carbon and organic matter were higher in thenative forest than in the oil palm-cultivated area which mayhave contributed to higher archaeal diversity Although thereare no studies on the effect of total soil carbon on archaealcommunities there are studies on the bacterial communitiesin Amazonian soils The work of Edgar et al [33] describeddistinct soil bacterial communities under different vegetationtypes The authors found that bacterial diversity correlatedwith total organic carbon and total nitrogen content [46]Furthermore in a study of bacterial diversity and microbialbiomass in forest pasture and fallow soils in the Amazonbasin microbial biomass was highest in pasture soils whichalso had 30ndash47 higher carbon levels than the other sitesover the course of a year [47]

Several studies have also reported alterations in thecomposition of archaeal [48] bacterial [47] and fungal [49]communities associated with land use changes A studyof archaeal communities in Amazonian anthrosols undervarious land uses (agriculture pasture and secondary forest)and cultivation practices (eg eggplant banana citrus andmanioc) [10] reported that agriculture negatively affectedarchaeal community diversity Results of ribosomal inter-genic spacer analysis showed that soil type is also responsiblefor changes in archaeal communities in soils under differenttypes of land use (native grassland native forest Eucalyptus

and Acacia plantations and soybean and watermelon fields)[50]

The Venn diagram showed that the native forest soil has agreater number of unique sequences (Figure 2) This was notsurprising due to the greater archaeal richness and diversityobserved in the native forest soil (Table 2) The number ofunique OTUs found in the oil palm-cultivated area is almostthe same as that of shared OTUs with the native forest areaAlthough oil palm cultivation decreased archaeal diversity inthe soil there is still a core group of archaea OTUs sharedwith the native forest suggesting that these archaea havecertain characteristics that allow them to persist after thechange in land use It cannot be ruled out that these archaeaare metabolically inactive The smaller number of uniquesequences in the oil palm-cultivated area may be related to alower complexity of the vegetation (ie oil palmmonocultureversus great plant diversity in the forest) since plant diversitycould affect these communities [51]

In our study all OTUs identified belong to the domainArchaea and were classified into two phyla Euryarchaeotaand Thaumarchaeota It is important to mention that thedatabases were only recently updated to recognize the phy-lumThaumarchaeota therefore previous studies on Archaeacould not recognize this phylum as proposed by Brochier-Armanet et al [11] The creation of this novel phylum wassupported by a study comparing the genomes of two marineammonia-oxidizing archaea (AOAs) that identified a core setof informational genes specific to this group [52] Thereforeto allow comparisons with other studies we reanalyzed theirdata whenever possible updating the taxonomic classifica-tion to include the phylumThaumarchaeota

Thework ofChao and Shen [35] focused on the impacts ofland use (ie agricultural systems of indigenous people andcattle pasture) on archaeal communities by PCR (amoA gene)

10 Archaea

and denaturing gradient gel electrophoresis analysis of differ-ent Western Amazon soils The databases at the time did notconsiderThaumarchaeota in their classification however ourreanalysis of their dataset using an updated database showedthat Euryarchaeota and Thaumarchaeota were present intheir samples along with Crenarchaeota (Thermoprotei) Inaddition changes were observed in the structure of soilarchaeal communities when comparing the cultivated areaswith forest areas that had been converted to pasture

In our study differences in the percentages of Thau-marchaeota and Euryarchaeota found in the soil sampleswere not striking Thaumarchaeota were predominant in thenative forest (58) whereas Euryarchaeota were predom-inant in the oil palm-cultivated area with 54 (Figure 3)This result is consistent with other comparative studies onarchaeal communities in different types of Brazilian soils (ieAmazonian anthrosols Caatinga and Cerrado) [10 53 54]and a study of different tropical forest soil types that reportedThaumarchaeota as the predominant phylum [44]

The phylumThaumarchaeota comprises all known AOAsand is directly involved in nitrogen metabolism Ammonia isused as an energy source by Group I1a found predominantlyin aquatic systems [52] and Group I1b found in soils [55]Thaumarchaeota Group I1c is primarily found in acidic soilsbut the ecological roles of these archaea are still unknown [5657]

The literature presents conflicting data about the pro-portion of AOAsAOBs soils In some studies AOAs arepredominant [58] while in others AOBs are more abundant[59] However several studies found a predominance of thearchaeal amoA gene over the bacterial amoA gene in soils[60] AOAs are frequently detected in soils their abundancerange from 05 to 10 of the prokaryotic community andtheir abundance is strongly influenced by physicochemicalconditions as well as the organic matter content [61 62] Inacidic soils AOAs appear to have a more important role inammonia oxidation than AOBs [57]

The Soil Crenarchaeotic Group a class within Thaumar-chaeota according to the SILVA database classification wasdetected in both soil samples but a higher representationwas observed in native forest soil This class was discoveredin boreal forest soil [63] Candidatus Nitrososphaera whichbelongs to the Soil Crenarchaeotic Group class is a memberof Thaumarchaeota Group I1b it is an AOA that usesammonia or urea as an energy source [64 65]

On the other hand members of the phylum Eur-yarchaeota are directly involved in carbon metabolismwith several classes of methanogens and methane-oxidizingorganisms [66ndash68] Navarrete et al [48] studied the impactof converting Amazon native forest to pasture and reportedchanges not only to the composition and abundance of soilmicrobial communities but also to a decrease in the diversityof gene function

Within the phylum Euryarchaeota Methanomicrobiaand Thermoplasmata were the main classes detected in oursamples along with a few representatives of Methanobac-teria and Halobacteria Members of Methanobacteria andMethanomicrobia are methanogens [69] Methanomicrobiawere present in both native forest soil and oil palm-cultivated

soil with larger representation in the oil palm-cultivatedarea Within Methanomicrobia the genera Rice Cluster IMethanosarcina andMethanocellawere present in both sam-ples (Figure 6(b)) These genera are closely related and foundin rice fields that produce methane [70] The order Metha-nomicrobiales was detected only in the native forest soilwhich can probably be explained by the relatively high levelsof carbon and organic matter This archaeal order is respon-sible for the final step of methane production in the carboncycle by anaerobic degradation of organic matter [66 71]However the contribution of the Methanomicrobia to thecarbon cycle in the oil palm-cultivated soils could not beassessed since there are no data on the methane productionin the studied areas

Among the Thaumarchaeota sequences used to generatethe phylogenetic tree that were unclassified at the genuslevel several OTUs formed different clusters associated withtheMiscellaneous Crenarchaeotic Group (Figure 5(a))Thesesequences were from both soil samples indicating that theunclassified Thaumarchaeota in the native forest and the oilpalm-cultivated area are phylogenetically related

Most of the sequences remained unclassified at thegenus level for both Thaumarchaeota and Euryarchaeota(Figures 6(a) and 6(b)) This result likely reflects the fact thatmost taxonomic databases do not have many representativesof Archaea especially belonging to the phylum Thaumar-chaeota thus few classifications of new sequences reach thegenus level Several genera were identified as Candidatusaccording to the International Code of Nomenclature ofBacteria indicating that characterization of the culturedprokaryote is incomplete [72]

The proportions of only four groups classified at thegenus level differed significantly between native forest and oilpalm-cultivated soils (Methanosarcina and Rice Cluster I inthe phylum Euryarchaeota and Candidatus NitrososphaeraandCandidatusNitrosotalea in the phylumThaumarchaeota)(Figure 7) With respect to the methanogens sequencesrelated to Methanosarcina were slightly more abundant insoil under oil palm cultivation (Figure 7) whereas sequencesbelonging to the Rice Cluster I were more abundant in nativeforest soil

In the Thaumarchaeota phylum Candidatus Nitrosos-phaera an AOA member of group I1b [65] was detectedin both samples but had a much higher representation innative forest (native forest 182 oil palm-cultivated area103) (Figure 7) In a comparative study of the archaealcommunities in Amazonian dark earth (ldquoTerra Pretardquo)and adjacent soils Candidatus Nitrososphaera was themain archaeal group found in both soils [10] In contrastCandidatus Nitrosotalea another AOA was more abundantin the oil palm-cultivated area (100) than in native forestsoil (44) in our study (Figure 7) Lehtovirta-Morley et al[73] cultivated the obligate acidophilic ammonia oxidizerCandidatus Nitrosotalea devanaterra from a nitrifying acidicsoil Later the same group characterized two strains ofacidophilic AOA belonging to this genus from a Chinesepaddy field and a Scottish agricultural soil [74] In our studythe greater representation of Candidatus Nitrososphaeraand Candidatus Nitrosotalea in native forest and oil

Archaea 11

palm-cultivated soils respectively can be explained by thelow soil pH (native forest pH 550 oil palm-cultivated areapH 483) since in basic soils the activity and growth ofthese archaea are diminished by the exponential decrease innitrite [6 75] In addition soil under oil palm cultivationis fertilized with nitrogen which may explain the higherlevel of Candidatus Nitrosotalea in this area compared withthat of native forest soil Finally although the cultivatedmembers of the genus Ca Nitrososphaera grow betteron neutral pH they have been reported to grow on pHvalues ranging from 60 to 85 In addition a recent workpublished by Wang et al (2014) [76] described active AOAsfrom an acidic soil (pH 492) which are closely related toNitrososphaera known as Nitrososphaera-like organismsby molecular phylogenetic methods Most of our sequenceswere classified as Nitrosotalea-like which are sequencesbelonging to the group I1a This group is typically foundin aquatic environments such as lake sediments or marineenvironments A similar ubiquitous distribution in soils isfound for the group I1b which are not restricted to a specificsoil pH condition

5 Conclusions

Although archaea are known to be ubiquitous there is stillmuch to be learned about their diversity biological rolesand metabolic capabilities This work is the first to comparearchaeal communities associated with the Amazon forest soilwith those of soil under oil palm cultivation

Archaeal diversity was found to be lower in oil palm-cultivated soil than in the native forest soil Only about 30of the OTUs were found in the oil palm soil in comparisonto the native forest However two main phyla were detectedin both samples Thaumarchaeota and Euryarchaeota with apredominance ofThaumarchaeota in native forest soil Theseresults support previous studies in other Brazilian biomenative soils (eg Caatinga Cerrado) as well as soil underdifferent types of land use (eg pasture and agricultural) Itis also important to mention that there are also groups ofuncertain classification associated with the Crenarchaeotaphylum

Although knowledge about Archaea has increased dra-matically in the last few decades little is known aboutthe biology of many archaeal groups and the taxonomyof Archaea is still a work in progress Thus it is notsurprising thatmany sequences identified in thiswork remainunclassified sometimes even at higher taxonomic levels Itis not clear whether the relatively small overall differencebetween soil archaeal communities of the Amazon forest andoil palm-cultivated land is a consequence of the current stateof archaeal taxonomy or whether it indicates that archaealdiversity is lower than bacterial diversity [77]

It is also important to consider that with land usechange due to agricultural practices the microstructure ofthe soil is broken down which can reduce the natural habitatfragmentation allowing potential interactions among theresident microbiota that otherwise would not happen [7879]

A phylogenetic tree of unclassified Thaumarchaeota 16SrRNA gene sequences was useful in exploring relationshipsamong these sequences When taxonomic classification ofsequences was possible it revealed the presence of OTUsfrom genera known to be involved in the metabolism ofcarbon and nitrogen (methanogens Methanocella and Meth-anosarcina AOACandidatusNitrososphaera andCandidatusNitrosotalea) hinting at possible roles for these archaealcommunities in Amazon soils

More studies are needed to elucidate these roles Cultur-ing some of these archaea in the laboratory would allow theirmetabolism and physiology to be studied in detail allowingfor a better understanding of the roles of Archaea in the bio-geochemical cycles

Conflict of Interests

The authors declare that there is no conflict of interestsregarding the publication of this paper

Acknowledgments

This work was supported by grants from FINEP CNPqCAPES and FAP-DF The authors thank Dr Boari for helpwith sample collection logistics Dr Betania Ferraz Quirinowas supported by the Cientista Visitante program fromEmbrapa

References

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[2] IBGE ldquoMapa de Biomas do Brasil primeira aproximacaordquo Riode Janeiro Brazil 2004 httpwwwibgegovbrhome

[3] C Azevedo-Ramos ldquoSustainable development and challengingdeforestation in the Brazilian Amazon the good the badand the uglyrdquo in Proceedings of the Symposium Our CommonGround Innovations in Land Use Decision-Making vol 59 p 5Vancouver Canada May 2007

[4] C A Francis K J Roberts J M Beman A E Santoro and B BOakley ldquoUbiquity and diversity of ammonia-oxidizing archaeainwater columns and sediments of the oceanrdquoProceedings of theNational Academy of Sciences of theUnited States of America vol102 no 41 pp 14683ndash14688 2005

[5] A H Treusch S Leininger A Kietzin S C Schuster H-PKlenk and C Schleper ldquoNovel genes for nitrite reductase andAmo-related proteins indicate a role of uncultivatedmesophiliccrenarchaeota in nitrogen cyclingrdquo Environmental Microbiol-ogy vol 7 no 12 pp 1985ndash1995 2005

[6] G W Nicol S Leininger C Schleper and J I Prosser ldquoTheinfluence of soil pH on the diversity abundance and tran-scriptional activity of ammonia oxidizing archaea and bacteriardquoEnvironmentalMicrobiology vol 10 no 11 pp 2966ndash2978 2008

[7] P Offre A Spang and C Schleper ldquoArchaea in biogeochemicalcyclesrdquo Annual Review of Microbiology vol 67 pp 437ndash4572013

[8] MAPA Anuario Estatıstico da Agroenergia 2012 StatisticalYearbook of Agroenergy 2013

[9] J C Bergmann D D Tupinamba O Y A Costa J R MAlmeida C C Barreto andB FQuirino ldquoBiodiesel production

12 Archaea

in Brazil and alternative biomass feedstocksrdquo Renewable ampSustainable Energy Reviews vol 21 pp 411ndash420 2013

[10] R G Taketani and S M Tsai ldquoThe influence of different landuses on the structure of archaeal communities in Amazoniananthrosols based on 16S rRNA and amoA genesrdquo MicrobialEcology vol 59 no 4 pp 734ndash743 2010

[11] C Brochier-Armanet B Boussau S Gribaldo and P ForterreldquoMesophilic crenarchaeota proposal for a third archaeal phy-lum theThaumarchaeotardquoNature Reviews Microbiology vol 6no 3 pp 245ndash252 2008

[12] S M Barns C F Delwiche J D Palmer and N R PaceldquoPerspectives on archaeal diversity thermophily and mono-phyly from environmental rRNA sequencesrdquo Proceedings of theNational Academy of Sciences of the United States of Americavol 93 no 17 pp 9188ndash9193 1996

[13] J G Elkins M Podar D E Graham et al ldquoA korarchaealgenome reveals insights into the evolution of the Archaeardquo Pro-ceedings of the National Academy of Sciences of the United Statesof America vol 105 no 23 pp 8102ndash8107 2008

[14] H Huber M J Hohn R Rachel T Fuchs V C Wimmer andK O Stetter ldquoA new phylum of Archaea represented by ananosized hyperthermophilic symbiontrdquo Nature vol 417 no6884 pp 63ndash67 2002

[15] T Nunoura H Oida M Nakaseama et al ldquoArchaeal diversityand distribution along thermal and geochemical gradients inhydrothermal sediments at the Yonaguni Knoll IV hydrother-mal field in the SouthernOkinawa TroughrdquoApplied and Enviro-nmental Microbiology vol 76 no 4 pp 1198ndash1211 2010

[16] C Rinke P Schwientek A Sczyrba et al ldquoInsights into thephylogeny and coding potential of microbial dark matterrdquoNature vol 499 no 7459 pp 431ndash437 2013

[17] A Spang J H Saw S L Joslashrgensen et al ldquoComplex archaea thatbridge the gap between prokaryotes and eukaryotesrdquo Naturevol 521 no 7551 pp 173ndash179 2015

[18] S Bagchi S E Vlaeminck L A Sauder et al ldquoTemporal andspatial stability of ammonia-oxidizing archaea and bacteria inaquariumbiofiltersrdquo PLoSONE vol 9 no 12 Article ID e1135152014

[19] W Qin S A Amin W Martens-Habbena et al ldquoMarineammonia-oxidizing archaeal isolates display obligate mixotro-phy and wide ecotypic variationrdquo Proceedings of the NationalAcademy of Sciences of the United States of America vol 111 no34 pp 12504ndash12509 2014

[20] M Coci N Odermatt M M Salcher J Pernthaler and GCorno ldquoEcology and distribution ofThaumarchaea in the deephypolimnion of Lake Maggiorerdquo Archaea vol 2015 Article ID590434 11 pages 2015

[21] J G Ferry ldquoHow tomake a living by exhalingmethanerdquoAnnualReview of Microbiology vol 64 pp 453ndash473 2010

[22] A Meyer A Focks V Radl G Welzl I Schoning and MSchloter ldquoInfluence of land use intensity on the diversity ofammonia oxidizing bacteria and archaea in soils from grasslandecosystemsrdquoMicrobial Ecology vol 67 no 1 pp 161ndash166 2014

[23] V De Gannes G Eudoxie and W J Hickey ldquoImpacts ofedaphic factors on communities of ammonia-oxidizing archaeaammonia-oxidizing bacteria and nitrification in tropical soilsrdquoPLoS ONE vol 9 no 2 Article ID e89568 2014

[24] D S Ribeiro F C J da Silva and TN Carvalho ldquoSobrevivenciade seis especies florestais em uma area explorada seletivamenteno municıpio de Moju Parardquo CERNE vol 9 no 2 pp 153ndash1632003

[25] D HM Costa C A P Ferreira J NM Silva J D C A Lopesand J O P D Carvalho ldquoPotencial madeireiro de floresta densano municıpio de Moju Estado do Parardquo Documentos 1998

[26] C M Almeida S F Lima C V Martins-Da-Silva and JI Gomes ldquoCaracterizacao morfologica e anatomica de dezespecies de leguminosae ocorrentes em uma floresta tropicalumida localizada no municıpio de Moju Estado do Parardquo inA Silvicultura na Amazonia Oriental Contribuicoes do ProjetoEmbrapa-DFID J NMC Silva JO P deCarvalho and J AGYared Eds pp 19ndash54 Embrapa Amazonia Oriental (CPATU)Belem Brazil 2001

[27] K Smalla N Cresswell L C Mendoncahagler A Woltersand J D Vanelsas ldquoRapid extraction protocol from soil forpolymerase chain reaction-mediated amplificationrdquo Journal ofApplied Bacteriology vol 74 pp 78ndash85 1993

[28] S Gantner A F Andersson L Alonso-Saez and S Bertils-son ldquoNovel primers for 16S rRNA-based archaeal communityanalyses in environmental samplesrdquo Journal of MicrobiologicalMethods vol 84 no 1 pp 12ndash18 2011

[29] P D Schloss S L Westcott T Ryabin et al ldquoIntroducingMOTHUR open-source platform-independent community-supported software for describing and comparing microbialcommunitiesrdquoApplied and EnvironmentalMicrobiology vol 75no 23 pp 7537ndash7541 2009

[30] PD SchlossDGevers and S LWestcott ldquoReducing the effectsof PCR amplification and sequencing artifacts on 16S rRNA-based studiesrdquo PLoS ONE vol 6 no 12 Article ID e27310 2011

[31] CQuince A Lanzen T P Curtis et al ldquoAccurate determinationof microbial diversity from 454 pyrosequencing datardquo NatureMethods vol 6 no 9 pp 639ndash641 2009

[32] P Yilmaz L W Parfrey P Yarza et al ldquoThe SILVA and lsquoall-species Living Tree Project (LTP)rsquo taxonomic frameworksrdquoNucleic Acids Research vol 42 no 1 pp D643ndashD648 2014

[33] R C Edgar B J Haas J C Clemente C Quince and RKnight ldquoUCHIME improves sensitivity and speed of chimeradetectionrdquo Bioinformatics vol 27 no 16 pp 2194ndash2200 2011

[34] A Chao and T-J Shen ldquoNonparametric estimation of Shan-nonrsquos index of diversity when there are unseen species insamplerdquo Environmental and Ecological Statistics vol 10 no 4pp 429ndash443 2003

[35] A Chao and T-J Shen Program SPADE (Species Prediction andDiversity Estimation) 2010

[36] C L Shannon andWWeaverTheMathematicalTheory of Com-munication University of Illinois Press Urbana Ill USA 1949

[37] I J Good ldquoThe population frequencies of species and theestimation of population parametersrdquo Biometrika vol 40 pp237ndash264 1953

[38] K Tamura G Stecher D Peterson A Filipski and S KumarldquoMEGA6molecular evolutionary genetics analysis version 60rdquoMolecular Biology and Evolution vol 30 no 12 pp 2725ndash27292013

[39] DA BensonMCavanaugh K Clark et al ldquoGenBankrdquoNucleicAcids Research vol 41 no 1 pp D36ndashD42 2013

[40] D H Parks G W Tyson P Hugenholtz and R G BeikoldquoSTAMP statistical analysis of taxonomic and functional pro-filesrdquo Bioinformatics vol 30 no 21 pp 3123ndash3124 2014

[41] R G Newcombe ldquoInterval estimation for the differencebetween independent proportions comparison of elevenmeth-odsrdquo Statistics in Medicine vol 17 no 8 pp 873ndash890 1998

[42] J D Storey J E Taylor and D Siegmund ldquoStrong controlconservative point estimation and simultaneous conservative

Archaea 13

consistency of false discovery rates a unified approachrdquo Journalof the Royal Statistical Society Series B Statistical Methodologyvol 66 no 1 pp 187ndash205 2004

[43] G C Baker J J Smith and D A Cowan ldquoReview and re-analysis of domain-specific 16S primersrdquo Journal of Microbio-logical Methods vol 55 no 3 pp 541ndash555 2003

[44] W H Zhong and Z C Cai ldquoLong-term effects of inorganicfertilizers on microbial biomass and community functionaldiversity in a paddy soil derived from quaternary red clayrdquoApplied Soil Ecology vol 36 no 2-3 pp 84ndash91 2007

[45] EWessen S Hallin and L Philippot ldquoDifferential responses ofbacterial and archaeal groups at high taxonomical ranks to soilmanagementrdquo Soil Biology and Biochemistry vol 42 no 10 pp1759ndash1765 2010

[46] ZWuW Lin B Li et al ldquoTerminal restriction fragment lengthpolymorphism analysis of soil bacterial communities underdifferent vegetation types in subtropical area rdquo PLOS ONE vol10 no 6 Article ID e0129397 2015

[47] K Cenciani M R Lambais C C Cerri L C B De Azevedoand B J Feigl ldquoBacteria diversity and microbial biomass inforest pasture and fallow soils in the southwestern Amazonbasinrdquo Revista Brasileira de Ciencia do Solo vol 33 no 4 pp907ndash916 2009

[48] A A Navarrete R G Taketani L WMendes F de Souza Can-navan F M de Souza Moreira and S M Tsai ldquoLand-use sys-tems affect Archaeal community structure and functional diver-sity in western Amazon soilsrdquo Revista Brasileira de Ciencia doSolo vol 35 no 5 pp 1527ndash1540 2011

[49] E Lumini A Orgiazzi R Borriello P Bonfante and V Bian-ciotto ldquoDisclosing arbuscular mycorrhizal fungal biodiversityin soil through a land-use gradient using a pyrosequencingapproachrdquo Environmental Microbiology vol 12 no 8 pp 2165ndash2179 2010

[50] M Lupatini R J S Jacques Z I Antoniolli A K A SuleimanR R Fulthorpe and L F W Roesch ldquoLand-use change andsoil type are drivers of fungal and archaeal communities in thePampa biomerdquo World Journal of Microbiology and Biotechnol-ogy vol 29 no 2 pp 223ndash233 2013

[51] E G Lamb N Kennedy and S D Siciliano ldquoEffects of plantspecies richness and evenness on soil microbial communitydiversity and functionrdquo Plant and Soil vol 338 no 1 pp 483ndash495 2011

[52] A Spang R Hatzenpichler C Brochier-Armanet et al ldquoDis-tinct gene set in two different lineages of ammonia-oxidizingarchaea supports the phylum Thaumarchaeotardquo Trends inMicrobiology vol 18 no 8 pp 331ndash340 2010

[53] R G Pacchioni F M Carvalho C E Thompson et al ldquoTax-onomic and functional profiles of soil samples from Atlanticforest and Caatinga biomes in northeastern Brazilrdquo Microbio-logyOpen vol 3 no 3 pp 299ndash315 2014

[54] E Catao A P Castro C C Barreto R H Kruger and C MKyaw ldquoDiversity of Archaea in Brazilian savanna soilsrdquoArchivesof Microbiology vol 195 no 7 pp 507ndash512 2013

[55] M Pester T Rattei S Flechl et al ldquoAmoA-based consensusphylogeny of ammonia-oxidizing archaea and deep sequencingof amoA genes from soils of four different geographic regionsrdquoEnvironmental Microbiology vol 14 no 2 pp 525ndash539 2012

[56] J-Z He H-W Hu and L-M Zhang ldquoCurrent insights into theautotrophic thaumarchaeal ammonia oxidation in acidic soilsrdquoSoil Biology amp Biochemistry vol 55 pp 146ndash154 2012

[57] L-M Zhang H-W Hu J-P Shen and J-Z He ldquoAmmonia-oxidizing archaea have more important role than ammonia-oxidizing bacteria in ammonia oxidation of strongly acidicsoilsrdquo ISME Journal vol 6 no 5 pp 1032ndash1045 2012

[58] S Leininger T Urich M Schloter et al ldquoArchaea predominateamong ammonia-oxidizing prokaryotes in soilsrdquo Nature vol442 no 7104 pp 806ndash809 2006

[59] N C Banning L D Maccarone L M Fisk and D VMurphy ldquoAmmonia-oxidising bacteria not archaea dominatenitrification activity in semi-arid agricultural soilrdquo ScientificReports vol 5 Article ID 11146 2015

[60] M Herrmann A Hadrich and K Kusel ldquoPredominance ofthaumarchaeal ammonia oxidizer abundance and transcrip-tional activity in an acidic fenrdquoEnvironmentalMicrobiology vol14 no 11 pp 3013ndash3025 2012

[61] K Zhalnina P Dorr de Quadros F O Camargo and E WTriplett ldquoDrivers of archaeal ammonia-oxidizing communitiesin soilrdquo Frontiers in Microbiology vol 3 article 210 Article IDArticle 210 2012

[62] J Hong J Cho and Y Hong ldquoEnvironmental variables shapingthe ecological niche of Thaumarchaeota in soil direct andindirect causal effectsrdquo PLoS ONE vol 10 no 8 Article IDe0133763 2015

[63] G Jurgens K Lindstrom and A Saano ldquoNovel group withinthe kingdom Crenarchaeota from boreal forest soilrdquo Appliedand Environmental Microbiology vol 63 no 2 pp 803ndash8051997

[64] M Tourna M Stieglmeier A Spang et al ldquoNitrososphaeraviennensis an ammonia oxidizing archaeon from soilrdquoProceed-ings of the National Academy of Sciences of the United States ofAmerica vol 108 no 20 pp 8420ndash8425 2011

[65] K V Zhalnina R Dias M T Leonard et al ldquoGenome sequenceof Candidatus Nitrososphaera evergladensis from group I1benriched from everglades soil reveals novel genomic features ofthe ammonia-oxidizing archaeardquoPLoSONE vol 9 no 7 ArticleID e101648 2014

[66] I Anderson L E Ulrich B Lupa et al ldquoGenomic char-acterization of methanomicrobiales reveals three classes ofmethanogensrdquo PLoS ONE vol 4 no 6 Article ID e5797 2009

[67] J E Galagan C Nusbaum A Roy et al ldquoThe genome of Macetivorans reveals extensive metabolic and physiological diver-sityrdquo Genome Research vol 12 no 4 pp 532ndash542 2002

[68] C Wrede S Kokoschka A Dreier C Heller J Reitner and MHoppert ldquoDeposition of biogenic iron minerals in a methaneoxidizing microbial matrdquo Archaea vol 2013 Article ID 1029728 pages 2013

[69] G Garrity Bergeyrsquo Manual of Systematic Bacteriology 721 2001[70] S Sakai Y Takaki S Shimamura et al ldquoGenome sequence

of a mesophilic hydrogenotrophic methanogen Methanocellapaludicola the first cultivated representative of the orderMethanocellalesrdquo PLoS ONE vol 6 no 7 Article ID e228982011

[71] E Bapteste C Brochier and Y Boucher ldquoHigher-level clas-sification of the Archaea evolution of methanogenesis andmethanogensrdquo Archaea vol 1 no 5 pp 353ndash363 2005

[72] R G E Murray and E Stackebrandt ldquoTaxonomic note imple-mentation of the provisional status Candidatus for incompletelydescribed procaryotesrdquo International Journal of Systematic Bac-teriology vol 45 no 1 pp 186ndash187 1995

[73] L E Lehtovirta-Morley K Stoecker A Vilcinskas J I Prosserand G W Nicol ldquoCultivation of an obligate acidophilic ammo-nia oxidizer from a nitrifying acid soilrdquo Proceedings of the

14 Archaea

National Academy of Sciences of the United States of Americavol 108 no 38 pp 15892ndash15897 2011

[74] L E Lehtovirta-Morley C Ge J Ross H Yao G W Nicol andJ I Prosser ldquoCharacterisation of terrestrial acidophilic archaealammonia oxidisers and their inhibition and stimulation byorganic compoundsrdquo FEMSMicrobiology Ecology vol 89 no 3pp 542ndash552 2015

[75] M J Frijlink T Abee H J Laanbroek W de Boer and W NKonings ldquoThe bioenergetics of ammonia and hydroxylamineoxidation in Nitrosomonas europaea at acid and alkaline pHrdquoArchives of Microbiology vol 157 no 2 pp 194ndash199 1992

[76] B Wang Y Zheng R Huang et al ldquoActive ammonia oxidizersin an acidic soil are phylogenetically closely related to neu-trophilic archaeonrdquo Applied and Environmental Microbiologyvol 80 no 5 pp 1684ndash1691 2014

[77] J Y Aller and P F Kemp ldquoAre Archaea inherently less diversethan Bacteria in the same environmentsrdquo FEMS MicrobiologyEcology vol 65 no 1 pp 74ndash87 2008

[78] K Potthast U Hamer and FMakeschin ldquoLand-use change in atropical mountain rainforest region of southern Ecuador affectssoilmicroorganisms and nutrient cyclingrdquoBiogeochemistry vol111 no 1ndash3 pp 151ndash167 2012

[79] J I Prosser and G W Nicol ldquoArchaeal and bacterial ammonia-oxidisers in soil the quest for niche specialisation and differen-tiationrdquoTrends inMicrobiology vol 20 no 11 pp 523ndash531 2012

Submit your manuscripts athttpwwwhindawicom

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

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PeptidesInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

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International Journal of

Volume 2014

Zoology

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Molecular Biology International

GenomicsInternational Journal of

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The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014

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BioinformaticsAdvances in

Marine BiologyJournal of

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Signal TransductionJournal of

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Evolutionary BiologyInternational Journal of

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Biochemistry Research International

ArchaeaHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

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Microbiology

Archaea 7

FJ4853071| Uncultured archaeon clone

FJ4852991| Uncultured archaeon clone

FJ9027141| Uncultured archaeon clone

KC8313951| Uncultured archaeon clone

KC5103331| Uncultured archaeon clone

JX9848481| Uncultured archaeon clone

AM9107821| Uncultured archaeon clone

Uncultured archaeon acidic red soil-FJ174727

Ca Nitrososphaera gargensis-GU797786

Ca Nitrososphaeramdash(48 6)

Ca Nitrosotaleamdash(49 12)

Candidatus lainarchaeum (0 2)

Methanobacterium (2 3)

Methanomassillicoccus (6 2)

Methanocella (33 8)

Rice cluster I (43 25)

Methanosarcina baltica-AB973356

Methanocella paludicola-NR074192

Pyrobaculum aerophilum

-NR102764

Acidobacteria bacterium KBS 96-FJ870384

Ca Methanom

ethylophilus (12 0)

Ca Nitrososphaera sp-FR773157

Nitrosopumilus maritimus-JQ346765

Halococcus hamelinensis-LN651155

Pyrococcus furiosus-PFU20163

Cenarchaeum symbiosum-CSU51469

Methanoregula formicica-NR112877

FR7451211| Uncultured soil archaeon

Boreal forest soil archaeon-X96688

Trembling aspen archaeon-EF021427

MCG

MCG005

CA4099

99

7174

63

86

9687

9067

5696

6768 54

57

62

58

65

7559

62

63

65

64

52

64

64

1008654

91

99

999293

53798392

88

7274

5164

5858

CA7CA32

CA50NF17

NF6

CA52

T I1c

T I1b

T I1a

CA24CA6CA12

CA4

CA33

NF16

NF2

NF35

NF9

NF5NF3

6NF4

1NF3

2

NF1

2

NF4

NF1

NF3

0NF2

3

NF7CA3

CA39

CA37

CA46

CA44

CA2

CA42

CA27

Unclassified MCGmdash(16 4)

Unclassified Terrestrial Group (94 36)

Unclassified Marine Group I (2 2)

CA15

CA25

CA30

CA43

CA35

CA14

CA29

CA13

CA20

CA11

CA41

CA36

CA51

NF48

CA9NF4

0NF4

2CA21

CA22

NF38

CA45

CA16

CA47

NF31CA31NF28NF47NF27NF45CA18NF26

NF24NF33NF19

NF18NF37

NF34NF43NF15NF44NF13

NF22NF29NF11

CA49

CA1

CA19CA8

CA10

CA23CA38

CA54HM0511251| Uncultured

HM0511301| Uncultured archaeon clone

HM0511271| Uncultured archaeon clone

archaeon clone

NF46NF14

NF49NF25N

F10N

F39N

F20N

F21

NF8

NF3

CA48

CA34

CA5

CA28CA26CA17CA53

(E)

(B)

(C)

(a)

CA13

CA14

NF1

2

NF1

4M

ethan

ocell

a (3

8)Meth

anosarcina (3

8 4)

Methanoculleus (2 0)

Methanomassilicoccus (6 2)

Methanomassilicoccus (6 2)

Methanomassilicoccus (6 2)

Methanomethylophilus (6 2)

Methanomethylophilus (6 2)

Methanobrevibacter (1 0)

Methanobacterium

(2 3)

Methanobacterium

(2 3)

Methanobacterium

(2 3)

Methanobacterium

(2 3)

Methanobacterium

(2 3)

Methanosarci

na (38 4)

Methanosarcina (38 4)

Methanosarcina (38 4)

Methanosarcina (38 4)

NF16

CA4

CA15CA20

CA19

NF3NF1

Meth

anoc

ella (

33 8

)

Meth

anoc

ella p

aludi

cola-

NR 0741

92 1

Meth

anosa

rcina

baltic

a AB97

3356

1

Methanoregula formicica NR 112877 1

Halococcus hamelinensis LN651155 1

CA6N

F6NF8

NF19 NF5NF7NF20

Pyrobaculum aerophilum NR 102764 2

Ca Nitrososphaera gargensis G

U797786 1

Ca Nitrososphaera gargensis GU797786 1

Nitrosopumilus maritimus JQ346765 1Cenarchaeum symbiosum U51469 1Uncultured archaeon acidic red soil FJ174727 1

Boreal forest soil archaeon X96688 1

Trembling aspen archaeon EF021427 1

HM051125 1 Uncultured archaeon clone

HM051127 1 Uncultured archaeon clone

HM051130 1 Uncultured archaeon clone

FJ485307 1 Uncultured archaeon clone

FJ902714 1 Uncultured archaeon clone

KC831395 1 archaeon clone

FJ485299 1 archaeon clone

KC510333 1 archaeon clone

FR745121 1 archaeon clone

JX984848 1 archaeon clone

AM910782 1 archaeon clone

Acidobacteria bacterium KBS 96 FJ870384 1

Ca Nitrososphaera sp FR773157 1

CA7

Pyrococcus furiosus U20163 1

005

MCG

Halobacteriales

Cand

idat

us la

inar

chae

um (0

2)

Cand

idat

us la

inar

chae

um (0

2)

Thermoplasmatales

Methanomicrobiales

Methanosarcinalles

Methanocellales

T I1c

T I1a

T I1b

(C)

998785

58

100

100

100100

100

71 84 71

53

54

65

86

94

98

59100

100

78

10051

5477

63 73

99

100

8497100

78

100

100

76956970

100 10

010

0

637551

100

9999

9958

94

Methanobacteriales

(b)

Figure 5 Phylogenetic trees of (a) unclassified Thaumarchaeota 16S rRNA gene sequences from native forest soil and soil under oil palmcultivation and (b) 16S rRNA gene sequences classified as Euryarchaeota from native forest soil and soil under oil palm cultivation Theneighbor-joining method and Jukes-Cantor corrections were used with 1000 bootstrap replicates Bootstrap values above 50 are shownin the trees A representative number of operational taxonomic units (OTUs) were selected at 3 dissimilarity by MOTHUR to avoidredundancies in the phylogenetic trees Randomly chosen OTUs from native forest soil and from soil under oil palm cultivation were usedin the construction of the trees along with archaeal reference sequences (from GenBank) from the following groups Acidobacteria (B)Crenarchaeota (C) Euryarchaeota (E) Miscellaneous Crenarchaeotic Group (MCG) Thaumarchaeota Group I1a (T I1a) ThaumarchaeotaGroup I1b (T I1b) andThaumarchaeota Group I1c (T I1c)The AOAs are highlighted with the symbolX Sampling site is indicated by ldquoNFrdquo(native forest) or ldquoCArdquo (oil palm-cultivated area) followed by the number of the OTUs (NAxx or CAxx) OTU sequences classified to at leastthe class level are shown in bold followed by two numbers in parentheses indicating the number of sequences of that OTU detected in nativeforest soil and the number detected in the oil palm-cultivated area The scale bar represents the 5 estimated sequence divergence

8 Archaea

Unclassified Marine Group I

Unclassified Miscellaneous Crenarchaeotic Group

Unclassified Thaumarchaeota

Candidatus Nitrososphaera

Unclassified Soil Crenarchaeotic Group

Candidatus Nitrosotalea

Unclassified South African Gold Mine Gp 1

Unclassified Terrestrial Group

Cultivated areaNative forest

5 10 15 20 250()

(a)

Unclassified Thermoplasmatales

Unclassified Terrestrial Miscellaneous Group

Rice Cluster I

Methanocella

Methanosarcina

Unclassified Euryarchaeota

Unclassified Halobacteriales

Candidatus Methanomethylophilus

Unclassified Methanosarcinaceae

5 10 15 20 250()

Cultivated areaNative forest

(b)

Figure 6 Percentage of archaeal operational taxonomic units within the Thaumarchaeota (a) and Euryarchaeota (b) phyla classified at thegenus level for native forest soil (orange) and soil under oil palm cultivation (blue) The percentage of unclassified sequences at genus level isshown

4 Discussion

The Amazon forest is known for its macrospecies diversityhowever few studies have addressed its microbial diversityAmong the domains of life Archaea are undoubtedly the lesswell known

In this work almost 600000 archaeal 16S rRNA genesequences were obtained from native forest soil and anadjacent area under oil palm cultivation After processing(ie removal of short sequences sequences with errors low-quality sequences chimeras and possible contaminants) andnormalization 39111 sequences per area remained for anal-ysis This number of sequences provided sufficient coverageof archaeal diversity in both areas (Table 1)The primers used

were based on a large number of 16S rRNA sequences fromthe SILVA database [28] however given that little is knownabout this domain it is unlikely that the archaeal diversitywasfully covered [43]

The main physicochemical differences between soils ofthe native forest and the oil palm-cultivated area were thelevels of phosphorous potassium calcium organic matterand total carbon (Table 2) Correlations between phylogenyand physicochemical parameters were evaluated using PCAand NMDS however no clear clusters or correlations wereobserved (results not shown)

In a study of the long-term effects of inorganic fertilizerson microbial communities Zhong and Cai [44] reportedthat phosphorus application indirectly altered microbial

Archaea 9

Candidatus NitrososphaeraUnclassified Soil Crenarchaeotic Group (SCG)

Unclassified South African Gold Mine Gp 1 (SAGMCG-1)Candidatus Nitrosotalea

Unclassified Terrestreial Miscellaneous Gp (TMEG)Unclassified Thaumarchaeota

Unclassified ThermoplasmatalesUnclassified Miscellaneous Crenarchaeotic Group

MethanosarcinaRice Cluster I

95 confidence intervals

q-v

alue

(cor

rect

ed)

Cultivated areaNative forest

531e minus 10

420 6 8minus4minus6minus8 minus2minus10

Difference between proportions ()38800

Proportion ()

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

Figure 7 Archaeal genera that differ significantly (119902 lt 005) in relative abundance between native forest soil (orange) and soil under oil palmcultivation (blue) For the purpose of statistical analysis performed using STAMP software the sum of the percentages of archaea classifiedat the genus level in both soils is 100 Differences between proportions of each genus are shown with 95 confidence intervals Unclassifiedsequences were used to calculate frequency profiles but are not displayed

parameters in soils and increased crop yields by elevating thelevels of organic matter Wessen and collaborators used real-time PCR quantification to show the impacts of fertilizer useon several bacterial phyla and the archaeal phylum Crenar-chaeota (Thaumarchaeota Group I1c after reanalysis usingthe SILVAdatabase 119) [45]They reported that bacterial andarchaeal classes respond differently to fertilization primarilybecause of changes in certain soil parameters such as pHtotal nitrogen and the carbon nitrogen ratio In our studylevels of total carbon and organic matter were higher in thenative forest than in the oil palm-cultivated area which mayhave contributed to higher archaeal diversity Although thereare no studies on the effect of total soil carbon on archaealcommunities there are studies on the bacterial communitiesin Amazonian soils The work of Edgar et al [33] describeddistinct soil bacterial communities under different vegetationtypes The authors found that bacterial diversity correlatedwith total organic carbon and total nitrogen content [46]Furthermore in a study of bacterial diversity and microbialbiomass in forest pasture and fallow soils in the Amazonbasin microbial biomass was highest in pasture soils whichalso had 30ndash47 higher carbon levels than the other sitesover the course of a year [47]

Several studies have also reported alterations in thecomposition of archaeal [48] bacterial [47] and fungal [49]communities associated with land use changes A studyof archaeal communities in Amazonian anthrosols undervarious land uses (agriculture pasture and secondary forest)and cultivation practices (eg eggplant banana citrus andmanioc) [10] reported that agriculture negatively affectedarchaeal community diversity Results of ribosomal inter-genic spacer analysis showed that soil type is also responsiblefor changes in archaeal communities in soils under differenttypes of land use (native grassland native forest Eucalyptus

and Acacia plantations and soybean and watermelon fields)[50]

The Venn diagram showed that the native forest soil has agreater number of unique sequences (Figure 2) This was notsurprising due to the greater archaeal richness and diversityobserved in the native forest soil (Table 2) The number ofunique OTUs found in the oil palm-cultivated area is almostthe same as that of shared OTUs with the native forest areaAlthough oil palm cultivation decreased archaeal diversity inthe soil there is still a core group of archaea OTUs sharedwith the native forest suggesting that these archaea havecertain characteristics that allow them to persist after thechange in land use It cannot be ruled out that these archaeaare metabolically inactive The smaller number of uniquesequences in the oil palm-cultivated area may be related to alower complexity of the vegetation (ie oil palmmonocultureversus great plant diversity in the forest) since plant diversitycould affect these communities [51]

In our study all OTUs identified belong to the domainArchaea and were classified into two phyla Euryarchaeotaand Thaumarchaeota It is important to mention that thedatabases were only recently updated to recognize the phy-lumThaumarchaeota therefore previous studies on Archaeacould not recognize this phylum as proposed by Brochier-Armanet et al [11] The creation of this novel phylum wassupported by a study comparing the genomes of two marineammonia-oxidizing archaea (AOAs) that identified a core setof informational genes specific to this group [52] Thereforeto allow comparisons with other studies we reanalyzed theirdata whenever possible updating the taxonomic classifica-tion to include the phylumThaumarchaeota

Thework ofChao and Shen [35] focused on the impacts ofland use (ie agricultural systems of indigenous people andcattle pasture) on archaeal communities by PCR (amoA gene)

10 Archaea

and denaturing gradient gel electrophoresis analysis of differ-ent Western Amazon soils The databases at the time did notconsiderThaumarchaeota in their classification however ourreanalysis of their dataset using an updated database showedthat Euryarchaeota and Thaumarchaeota were present intheir samples along with Crenarchaeota (Thermoprotei) Inaddition changes were observed in the structure of soilarchaeal communities when comparing the cultivated areaswith forest areas that had been converted to pasture

In our study differences in the percentages of Thau-marchaeota and Euryarchaeota found in the soil sampleswere not striking Thaumarchaeota were predominant in thenative forest (58) whereas Euryarchaeota were predom-inant in the oil palm-cultivated area with 54 (Figure 3)This result is consistent with other comparative studies onarchaeal communities in different types of Brazilian soils (ieAmazonian anthrosols Caatinga and Cerrado) [10 53 54]and a study of different tropical forest soil types that reportedThaumarchaeota as the predominant phylum [44]

The phylumThaumarchaeota comprises all known AOAsand is directly involved in nitrogen metabolism Ammonia isused as an energy source by Group I1a found predominantlyin aquatic systems [52] and Group I1b found in soils [55]Thaumarchaeota Group I1c is primarily found in acidic soilsbut the ecological roles of these archaea are still unknown [5657]

The literature presents conflicting data about the pro-portion of AOAsAOBs soils In some studies AOAs arepredominant [58] while in others AOBs are more abundant[59] However several studies found a predominance of thearchaeal amoA gene over the bacterial amoA gene in soils[60] AOAs are frequently detected in soils their abundancerange from 05 to 10 of the prokaryotic community andtheir abundance is strongly influenced by physicochemicalconditions as well as the organic matter content [61 62] Inacidic soils AOAs appear to have a more important role inammonia oxidation than AOBs [57]

The Soil Crenarchaeotic Group a class within Thaumar-chaeota according to the SILVA database classification wasdetected in both soil samples but a higher representationwas observed in native forest soil This class was discoveredin boreal forest soil [63] Candidatus Nitrososphaera whichbelongs to the Soil Crenarchaeotic Group class is a memberof Thaumarchaeota Group I1b it is an AOA that usesammonia or urea as an energy source [64 65]

On the other hand members of the phylum Eur-yarchaeota are directly involved in carbon metabolismwith several classes of methanogens and methane-oxidizingorganisms [66ndash68] Navarrete et al [48] studied the impactof converting Amazon native forest to pasture and reportedchanges not only to the composition and abundance of soilmicrobial communities but also to a decrease in the diversityof gene function

Within the phylum Euryarchaeota Methanomicrobiaand Thermoplasmata were the main classes detected in oursamples along with a few representatives of Methanobac-teria and Halobacteria Members of Methanobacteria andMethanomicrobia are methanogens [69] Methanomicrobiawere present in both native forest soil and oil palm-cultivated

soil with larger representation in the oil palm-cultivatedarea Within Methanomicrobia the genera Rice Cluster IMethanosarcina andMethanocellawere present in both sam-ples (Figure 6(b)) These genera are closely related and foundin rice fields that produce methane [70] The order Metha-nomicrobiales was detected only in the native forest soilwhich can probably be explained by the relatively high levelsof carbon and organic matter This archaeal order is respon-sible for the final step of methane production in the carboncycle by anaerobic degradation of organic matter [66 71]However the contribution of the Methanomicrobia to thecarbon cycle in the oil palm-cultivated soils could not beassessed since there are no data on the methane productionin the studied areas

Among the Thaumarchaeota sequences used to generatethe phylogenetic tree that were unclassified at the genuslevel several OTUs formed different clusters associated withtheMiscellaneous Crenarchaeotic Group (Figure 5(a))Thesesequences were from both soil samples indicating that theunclassified Thaumarchaeota in the native forest and the oilpalm-cultivated area are phylogenetically related

Most of the sequences remained unclassified at thegenus level for both Thaumarchaeota and Euryarchaeota(Figures 6(a) and 6(b)) This result likely reflects the fact thatmost taxonomic databases do not have many representativesof Archaea especially belonging to the phylum Thaumar-chaeota thus few classifications of new sequences reach thegenus level Several genera were identified as Candidatusaccording to the International Code of Nomenclature ofBacteria indicating that characterization of the culturedprokaryote is incomplete [72]

The proportions of only four groups classified at thegenus level differed significantly between native forest and oilpalm-cultivated soils (Methanosarcina and Rice Cluster I inthe phylum Euryarchaeota and Candidatus NitrososphaeraandCandidatusNitrosotalea in the phylumThaumarchaeota)(Figure 7) With respect to the methanogens sequencesrelated to Methanosarcina were slightly more abundant insoil under oil palm cultivation (Figure 7) whereas sequencesbelonging to the Rice Cluster I were more abundant in nativeforest soil

In the Thaumarchaeota phylum Candidatus Nitrosos-phaera an AOA member of group I1b [65] was detectedin both samples but had a much higher representation innative forest (native forest 182 oil palm-cultivated area103) (Figure 7) In a comparative study of the archaealcommunities in Amazonian dark earth (ldquoTerra Pretardquo)and adjacent soils Candidatus Nitrososphaera was themain archaeal group found in both soils [10] In contrastCandidatus Nitrosotalea another AOA was more abundantin the oil palm-cultivated area (100) than in native forestsoil (44) in our study (Figure 7) Lehtovirta-Morley et al[73] cultivated the obligate acidophilic ammonia oxidizerCandidatus Nitrosotalea devanaterra from a nitrifying acidicsoil Later the same group characterized two strains ofacidophilic AOA belonging to this genus from a Chinesepaddy field and a Scottish agricultural soil [74] In our studythe greater representation of Candidatus Nitrososphaeraand Candidatus Nitrosotalea in native forest and oil

Archaea 11

palm-cultivated soils respectively can be explained by thelow soil pH (native forest pH 550 oil palm-cultivated areapH 483) since in basic soils the activity and growth ofthese archaea are diminished by the exponential decrease innitrite [6 75] In addition soil under oil palm cultivationis fertilized with nitrogen which may explain the higherlevel of Candidatus Nitrosotalea in this area compared withthat of native forest soil Finally although the cultivatedmembers of the genus Ca Nitrososphaera grow betteron neutral pH they have been reported to grow on pHvalues ranging from 60 to 85 In addition a recent workpublished by Wang et al (2014) [76] described active AOAsfrom an acidic soil (pH 492) which are closely related toNitrososphaera known as Nitrososphaera-like organismsby molecular phylogenetic methods Most of our sequenceswere classified as Nitrosotalea-like which are sequencesbelonging to the group I1a This group is typically foundin aquatic environments such as lake sediments or marineenvironments A similar ubiquitous distribution in soils isfound for the group I1b which are not restricted to a specificsoil pH condition

5 Conclusions

Although archaea are known to be ubiquitous there is stillmuch to be learned about their diversity biological rolesand metabolic capabilities This work is the first to comparearchaeal communities associated with the Amazon forest soilwith those of soil under oil palm cultivation

Archaeal diversity was found to be lower in oil palm-cultivated soil than in the native forest soil Only about 30of the OTUs were found in the oil palm soil in comparisonto the native forest However two main phyla were detectedin both samples Thaumarchaeota and Euryarchaeota with apredominance ofThaumarchaeota in native forest soil Theseresults support previous studies in other Brazilian biomenative soils (eg Caatinga Cerrado) as well as soil underdifferent types of land use (eg pasture and agricultural) Itis also important to mention that there are also groups ofuncertain classification associated with the Crenarchaeotaphylum

Although knowledge about Archaea has increased dra-matically in the last few decades little is known aboutthe biology of many archaeal groups and the taxonomyof Archaea is still a work in progress Thus it is notsurprising thatmany sequences identified in thiswork remainunclassified sometimes even at higher taxonomic levels Itis not clear whether the relatively small overall differencebetween soil archaeal communities of the Amazon forest andoil palm-cultivated land is a consequence of the current stateof archaeal taxonomy or whether it indicates that archaealdiversity is lower than bacterial diversity [77]

It is also important to consider that with land usechange due to agricultural practices the microstructure ofthe soil is broken down which can reduce the natural habitatfragmentation allowing potential interactions among theresident microbiota that otherwise would not happen [7879]

A phylogenetic tree of unclassified Thaumarchaeota 16SrRNA gene sequences was useful in exploring relationshipsamong these sequences When taxonomic classification ofsequences was possible it revealed the presence of OTUsfrom genera known to be involved in the metabolism ofcarbon and nitrogen (methanogens Methanocella and Meth-anosarcina AOACandidatusNitrososphaera andCandidatusNitrosotalea) hinting at possible roles for these archaealcommunities in Amazon soils

More studies are needed to elucidate these roles Cultur-ing some of these archaea in the laboratory would allow theirmetabolism and physiology to be studied in detail allowingfor a better understanding of the roles of Archaea in the bio-geochemical cycles

Conflict of Interests

The authors declare that there is no conflict of interestsregarding the publication of this paper

Acknowledgments

This work was supported by grants from FINEP CNPqCAPES and FAP-DF The authors thank Dr Boari for helpwith sample collection logistics Dr Betania Ferraz Quirinowas supported by the Cientista Visitante program fromEmbrapa

References

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[2] IBGE ldquoMapa de Biomas do Brasil primeira aproximacaordquo Riode Janeiro Brazil 2004 httpwwwibgegovbrhome

[3] C Azevedo-Ramos ldquoSustainable development and challengingdeforestation in the Brazilian Amazon the good the badand the uglyrdquo in Proceedings of the Symposium Our CommonGround Innovations in Land Use Decision-Making vol 59 p 5Vancouver Canada May 2007

[4] C A Francis K J Roberts J M Beman A E Santoro and B BOakley ldquoUbiquity and diversity of ammonia-oxidizing archaeainwater columns and sediments of the oceanrdquoProceedings of theNational Academy of Sciences of theUnited States of America vol102 no 41 pp 14683ndash14688 2005

[5] A H Treusch S Leininger A Kietzin S C Schuster H-PKlenk and C Schleper ldquoNovel genes for nitrite reductase andAmo-related proteins indicate a role of uncultivatedmesophiliccrenarchaeota in nitrogen cyclingrdquo Environmental Microbiol-ogy vol 7 no 12 pp 1985ndash1995 2005

[6] G W Nicol S Leininger C Schleper and J I Prosser ldquoTheinfluence of soil pH on the diversity abundance and tran-scriptional activity of ammonia oxidizing archaea and bacteriardquoEnvironmentalMicrobiology vol 10 no 11 pp 2966ndash2978 2008

[7] P Offre A Spang and C Schleper ldquoArchaea in biogeochemicalcyclesrdquo Annual Review of Microbiology vol 67 pp 437ndash4572013

[8] MAPA Anuario Estatıstico da Agroenergia 2012 StatisticalYearbook of Agroenergy 2013

[9] J C Bergmann D D Tupinamba O Y A Costa J R MAlmeida C C Barreto andB FQuirino ldquoBiodiesel production

12 Archaea

in Brazil and alternative biomass feedstocksrdquo Renewable ampSustainable Energy Reviews vol 21 pp 411ndash420 2013

[10] R G Taketani and S M Tsai ldquoThe influence of different landuses on the structure of archaeal communities in Amazoniananthrosols based on 16S rRNA and amoA genesrdquo MicrobialEcology vol 59 no 4 pp 734ndash743 2010

[11] C Brochier-Armanet B Boussau S Gribaldo and P ForterreldquoMesophilic crenarchaeota proposal for a third archaeal phy-lum theThaumarchaeotardquoNature Reviews Microbiology vol 6no 3 pp 245ndash252 2008

[12] S M Barns C F Delwiche J D Palmer and N R PaceldquoPerspectives on archaeal diversity thermophily and mono-phyly from environmental rRNA sequencesrdquo Proceedings of theNational Academy of Sciences of the United States of Americavol 93 no 17 pp 9188ndash9193 1996

[13] J G Elkins M Podar D E Graham et al ldquoA korarchaealgenome reveals insights into the evolution of the Archaeardquo Pro-ceedings of the National Academy of Sciences of the United Statesof America vol 105 no 23 pp 8102ndash8107 2008

[14] H Huber M J Hohn R Rachel T Fuchs V C Wimmer andK O Stetter ldquoA new phylum of Archaea represented by ananosized hyperthermophilic symbiontrdquo Nature vol 417 no6884 pp 63ndash67 2002

[15] T Nunoura H Oida M Nakaseama et al ldquoArchaeal diversityand distribution along thermal and geochemical gradients inhydrothermal sediments at the Yonaguni Knoll IV hydrother-mal field in the SouthernOkinawa TroughrdquoApplied and Enviro-nmental Microbiology vol 76 no 4 pp 1198ndash1211 2010

[16] C Rinke P Schwientek A Sczyrba et al ldquoInsights into thephylogeny and coding potential of microbial dark matterrdquoNature vol 499 no 7459 pp 431ndash437 2013

[17] A Spang J H Saw S L Joslashrgensen et al ldquoComplex archaea thatbridge the gap between prokaryotes and eukaryotesrdquo Naturevol 521 no 7551 pp 173ndash179 2015

[18] S Bagchi S E Vlaeminck L A Sauder et al ldquoTemporal andspatial stability of ammonia-oxidizing archaea and bacteria inaquariumbiofiltersrdquo PLoSONE vol 9 no 12 Article ID e1135152014

[19] W Qin S A Amin W Martens-Habbena et al ldquoMarineammonia-oxidizing archaeal isolates display obligate mixotro-phy and wide ecotypic variationrdquo Proceedings of the NationalAcademy of Sciences of the United States of America vol 111 no34 pp 12504ndash12509 2014

[20] M Coci N Odermatt M M Salcher J Pernthaler and GCorno ldquoEcology and distribution ofThaumarchaea in the deephypolimnion of Lake Maggiorerdquo Archaea vol 2015 Article ID590434 11 pages 2015

[21] J G Ferry ldquoHow tomake a living by exhalingmethanerdquoAnnualReview of Microbiology vol 64 pp 453ndash473 2010

[22] A Meyer A Focks V Radl G Welzl I Schoning and MSchloter ldquoInfluence of land use intensity on the diversity ofammonia oxidizing bacteria and archaea in soils from grasslandecosystemsrdquoMicrobial Ecology vol 67 no 1 pp 161ndash166 2014

[23] V De Gannes G Eudoxie and W J Hickey ldquoImpacts ofedaphic factors on communities of ammonia-oxidizing archaeaammonia-oxidizing bacteria and nitrification in tropical soilsrdquoPLoS ONE vol 9 no 2 Article ID e89568 2014

[24] D S Ribeiro F C J da Silva and TN Carvalho ldquoSobrevivenciade seis especies florestais em uma area explorada seletivamenteno municıpio de Moju Parardquo CERNE vol 9 no 2 pp 153ndash1632003

[25] D HM Costa C A P Ferreira J NM Silva J D C A Lopesand J O P D Carvalho ldquoPotencial madeireiro de floresta densano municıpio de Moju Estado do Parardquo Documentos 1998

[26] C M Almeida S F Lima C V Martins-Da-Silva and JI Gomes ldquoCaracterizacao morfologica e anatomica de dezespecies de leguminosae ocorrentes em uma floresta tropicalumida localizada no municıpio de Moju Estado do Parardquo inA Silvicultura na Amazonia Oriental Contribuicoes do ProjetoEmbrapa-DFID J NMC Silva JO P deCarvalho and J AGYared Eds pp 19ndash54 Embrapa Amazonia Oriental (CPATU)Belem Brazil 2001

[27] K Smalla N Cresswell L C Mendoncahagler A Woltersand J D Vanelsas ldquoRapid extraction protocol from soil forpolymerase chain reaction-mediated amplificationrdquo Journal ofApplied Bacteriology vol 74 pp 78ndash85 1993

[28] S Gantner A F Andersson L Alonso-Saez and S Bertils-son ldquoNovel primers for 16S rRNA-based archaeal communityanalyses in environmental samplesrdquo Journal of MicrobiologicalMethods vol 84 no 1 pp 12ndash18 2011

[29] P D Schloss S L Westcott T Ryabin et al ldquoIntroducingMOTHUR open-source platform-independent community-supported software for describing and comparing microbialcommunitiesrdquoApplied and EnvironmentalMicrobiology vol 75no 23 pp 7537ndash7541 2009

[30] PD SchlossDGevers and S LWestcott ldquoReducing the effectsof PCR amplification and sequencing artifacts on 16S rRNA-based studiesrdquo PLoS ONE vol 6 no 12 Article ID e27310 2011

[31] CQuince A Lanzen T P Curtis et al ldquoAccurate determinationof microbial diversity from 454 pyrosequencing datardquo NatureMethods vol 6 no 9 pp 639ndash641 2009

[32] P Yilmaz L W Parfrey P Yarza et al ldquoThe SILVA and lsquoall-species Living Tree Project (LTP)rsquo taxonomic frameworksrdquoNucleic Acids Research vol 42 no 1 pp D643ndashD648 2014

[33] R C Edgar B J Haas J C Clemente C Quince and RKnight ldquoUCHIME improves sensitivity and speed of chimeradetectionrdquo Bioinformatics vol 27 no 16 pp 2194ndash2200 2011

[34] A Chao and T-J Shen ldquoNonparametric estimation of Shan-nonrsquos index of diversity when there are unseen species insamplerdquo Environmental and Ecological Statistics vol 10 no 4pp 429ndash443 2003

[35] A Chao and T-J Shen Program SPADE (Species Prediction andDiversity Estimation) 2010

[36] C L Shannon andWWeaverTheMathematicalTheory of Com-munication University of Illinois Press Urbana Ill USA 1949

[37] I J Good ldquoThe population frequencies of species and theestimation of population parametersrdquo Biometrika vol 40 pp237ndash264 1953

[38] K Tamura G Stecher D Peterson A Filipski and S KumarldquoMEGA6molecular evolutionary genetics analysis version 60rdquoMolecular Biology and Evolution vol 30 no 12 pp 2725ndash27292013

[39] DA BensonMCavanaugh K Clark et al ldquoGenBankrdquoNucleicAcids Research vol 41 no 1 pp D36ndashD42 2013

[40] D H Parks G W Tyson P Hugenholtz and R G BeikoldquoSTAMP statistical analysis of taxonomic and functional pro-filesrdquo Bioinformatics vol 30 no 21 pp 3123ndash3124 2014

[41] R G Newcombe ldquoInterval estimation for the differencebetween independent proportions comparison of elevenmeth-odsrdquo Statistics in Medicine vol 17 no 8 pp 873ndash890 1998

[42] J D Storey J E Taylor and D Siegmund ldquoStrong controlconservative point estimation and simultaneous conservative

Archaea 13

consistency of false discovery rates a unified approachrdquo Journalof the Royal Statistical Society Series B Statistical Methodologyvol 66 no 1 pp 187ndash205 2004

[43] G C Baker J J Smith and D A Cowan ldquoReview and re-analysis of domain-specific 16S primersrdquo Journal of Microbio-logical Methods vol 55 no 3 pp 541ndash555 2003

[44] W H Zhong and Z C Cai ldquoLong-term effects of inorganicfertilizers on microbial biomass and community functionaldiversity in a paddy soil derived from quaternary red clayrdquoApplied Soil Ecology vol 36 no 2-3 pp 84ndash91 2007

[45] EWessen S Hallin and L Philippot ldquoDifferential responses ofbacterial and archaeal groups at high taxonomical ranks to soilmanagementrdquo Soil Biology and Biochemistry vol 42 no 10 pp1759ndash1765 2010

[46] ZWuW Lin B Li et al ldquoTerminal restriction fragment lengthpolymorphism analysis of soil bacterial communities underdifferent vegetation types in subtropical area rdquo PLOS ONE vol10 no 6 Article ID e0129397 2015

[47] K Cenciani M R Lambais C C Cerri L C B De Azevedoand B J Feigl ldquoBacteria diversity and microbial biomass inforest pasture and fallow soils in the southwestern Amazonbasinrdquo Revista Brasileira de Ciencia do Solo vol 33 no 4 pp907ndash916 2009

[48] A A Navarrete R G Taketani L WMendes F de Souza Can-navan F M de Souza Moreira and S M Tsai ldquoLand-use sys-tems affect Archaeal community structure and functional diver-sity in western Amazon soilsrdquo Revista Brasileira de Ciencia doSolo vol 35 no 5 pp 1527ndash1540 2011

[49] E Lumini A Orgiazzi R Borriello P Bonfante and V Bian-ciotto ldquoDisclosing arbuscular mycorrhizal fungal biodiversityin soil through a land-use gradient using a pyrosequencingapproachrdquo Environmental Microbiology vol 12 no 8 pp 2165ndash2179 2010

[50] M Lupatini R J S Jacques Z I Antoniolli A K A SuleimanR R Fulthorpe and L F W Roesch ldquoLand-use change andsoil type are drivers of fungal and archaeal communities in thePampa biomerdquo World Journal of Microbiology and Biotechnol-ogy vol 29 no 2 pp 223ndash233 2013

[51] E G Lamb N Kennedy and S D Siciliano ldquoEffects of plantspecies richness and evenness on soil microbial communitydiversity and functionrdquo Plant and Soil vol 338 no 1 pp 483ndash495 2011

[52] A Spang R Hatzenpichler C Brochier-Armanet et al ldquoDis-tinct gene set in two different lineages of ammonia-oxidizingarchaea supports the phylum Thaumarchaeotardquo Trends inMicrobiology vol 18 no 8 pp 331ndash340 2010

[53] R G Pacchioni F M Carvalho C E Thompson et al ldquoTax-onomic and functional profiles of soil samples from Atlanticforest and Caatinga biomes in northeastern Brazilrdquo Microbio-logyOpen vol 3 no 3 pp 299ndash315 2014

[54] E Catao A P Castro C C Barreto R H Kruger and C MKyaw ldquoDiversity of Archaea in Brazilian savanna soilsrdquoArchivesof Microbiology vol 195 no 7 pp 507ndash512 2013

[55] M Pester T Rattei S Flechl et al ldquoAmoA-based consensusphylogeny of ammonia-oxidizing archaea and deep sequencingof amoA genes from soils of four different geographic regionsrdquoEnvironmental Microbiology vol 14 no 2 pp 525ndash539 2012

[56] J-Z He H-W Hu and L-M Zhang ldquoCurrent insights into theautotrophic thaumarchaeal ammonia oxidation in acidic soilsrdquoSoil Biology amp Biochemistry vol 55 pp 146ndash154 2012

[57] L-M Zhang H-W Hu J-P Shen and J-Z He ldquoAmmonia-oxidizing archaea have more important role than ammonia-oxidizing bacteria in ammonia oxidation of strongly acidicsoilsrdquo ISME Journal vol 6 no 5 pp 1032ndash1045 2012

[58] S Leininger T Urich M Schloter et al ldquoArchaea predominateamong ammonia-oxidizing prokaryotes in soilsrdquo Nature vol442 no 7104 pp 806ndash809 2006

[59] N C Banning L D Maccarone L M Fisk and D VMurphy ldquoAmmonia-oxidising bacteria not archaea dominatenitrification activity in semi-arid agricultural soilrdquo ScientificReports vol 5 Article ID 11146 2015

[60] M Herrmann A Hadrich and K Kusel ldquoPredominance ofthaumarchaeal ammonia oxidizer abundance and transcrip-tional activity in an acidic fenrdquoEnvironmentalMicrobiology vol14 no 11 pp 3013ndash3025 2012

[61] K Zhalnina P Dorr de Quadros F O Camargo and E WTriplett ldquoDrivers of archaeal ammonia-oxidizing communitiesin soilrdquo Frontiers in Microbiology vol 3 article 210 Article IDArticle 210 2012

[62] J Hong J Cho and Y Hong ldquoEnvironmental variables shapingthe ecological niche of Thaumarchaeota in soil direct andindirect causal effectsrdquo PLoS ONE vol 10 no 8 Article IDe0133763 2015

[63] G Jurgens K Lindstrom and A Saano ldquoNovel group withinthe kingdom Crenarchaeota from boreal forest soilrdquo Appliedand Environmental Microbiology vol 63 no 2 pp 803ndash8051997

[64] M Tourna M Stieglmeier A Spang et al ldquoNitrososphaeraviennensis an ammonia oxidizing archaeon from soilrdquoProceed-ings of the National Academy of Sciences of the United States ofAmerica vol 108 no 20 pp 8420ndash8425 2011

[65] K V Zhalnina R Dias M T Leonard et al ldquoGenome sequenceof Candidatus Nitrososphaera evergladensis from group I1benriched from everglades soil reveals novel genomic features ofthe ammonia-oxidizing archaeardquoPLoSONE vol 9 no 7 ArticleID e101648 2014

[66] I Anderson L E Ulrich B Lupa et al ldquoGenomic char-acterization of methanomicrobiales reveals three classes ofmethanogensrdquo PLoS ONE vol 4 no 6 Article ID e5797 2009

[67] J E Galagan C Nusbaum A Roy et al ldquoThe genome of Macetivorans reveals extensive metabolic and physiological diver-sityrdquo Genome Research vol 12 no 4 pp 532ndash542 2002

[68] C Wrede S Kokoschka A Dreier C Heller J Reitner and MHoppert ldquoDeposition of biogenic iron minerals in a methaneoxidizing microbial matrdquo Archaea vol 2013 Article ID 1029728 pages 2013

[69] G Garrity Bergeyrsquo Manual of Systematic Bacteriology 721 2001[70] S Sakai Y Takaki S Shimamura et al ldquoGenome sequence

of a mesophilic hydrogenotrophic methanogen Methanocellapaludicola the first cultivated representative of the orderMethanocellalesrdquo PLoS ONE vol 6 no 7 Article ID e228982011

[71] E Bapteste C Brochier and Y Boucher ldquoHigher-level clas-sification of the Archaea evolution of methanogenesis andmethanogensrdquo Archaea vol 1 no 5 pp 353ndash363 2005

[72] R G E Murray and E Stackebrandt ldquoTaxonomic note imple-mentation of the provisional status Candidatus for incompletelydescribed procaryotesrdquo International Journal of Systematic Bac-teriology vol 45 no 1 pp 186ndash187 1995

[73] L E Lehtovirta-Morley K Stoecker A Vilcinskas J I Prosserand G W Nicol ldquoCultivation of an obligate acidophilic ammo-nia oxidizer from a nitrifying acid soilrdquo Proceedings of the

14 Archaea

National Academy of Sciences of the United States of Americavol 108 no 38 pp 15892ndash15897 2011

[74] L E Lehtovirta-Morley C Ge J Ross H Yao G W Nicol andJ I Prosser ldquoCharacterisation of terrestrial acidophilic archaealammonia oxidisers and their inhibition and stimulation byorganic compoundsrdquo FEMSMicrobiology Ecology vol 89 no 3pp 542ndash552 2015

[75] M J Frijlink T Abee H J Laanbroek W de Boer and W NKonings ldquoThe bioenergetics of ammonia and hydroxylamineoxidation in Nitrosomonas europaea at acid and alkaline pHrdquoArchives of Microbiology vol 157 no 2 pp 194ndash199 1992

[76] B Wang Y Zheng R Huang et al ldquoActive ammonia oxidizersin an acidic soil are phylogenetically closely related to neu-trophilic archaeonrdquo Applied and Environmental Microbiologyvol 80 no 5 pp 1684ndash1691 2014

[77] J Y Aller and P F Kemp ldquoAre Archaea inherently less diversethan Bacteria in the same environmentsrdquo FEMS MicrobiologyEcology vol 65 no 1 pp 74ndash87 2008

[78] K Potthast U Hamer and FMakeschin ldquoLand-use change in atropical mountain rainforest region of southern Ecuador affectssoilmicroorganisms and nutrient cyclingrdquoBiogeochemistry vol111 no 1ndash3 pp 151ndash167 2012

[79] J I Prosser and G W Nicol ldquoArchaeal and bacterial ammonia-oxidisers in soil the quest for niche specialisation and differen-tiationrdquoTrends inMicrobiology vol 20 no 11 pp 523ndash531 2012

Submit your manuscripts athttpwwwhindawicom

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Anatomy Research International

PeptidesInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporation httpwwwhindawicom

International Journal of

Volume 2014

Zoology

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Molecular Biology International

GenomicsInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioinformaticsAdvances in

Marine BiologyJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Signal TransductionJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioMed Research International

Evolutionary BiologyInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Biochemistry Research International

ArchaeaHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Genetics Research International

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Advances in

Virolog y

Hindawi Publishing Corporationhttpwwwhindawicom

Nucleic AcidsJournal of

Volume 2014

Stem CellsInternational

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Enzyme Research

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

International Journal of

Microbiology

8 Archaea

Unclassified Marine Group I

Unclassified Miscellaneous Crenarchaeotic Group

Unclassified Thaumarchaeota

Candidatus Nitrososphaera

Unclassified Soil Crenarchaeotic Group

Candidatus Nitrosotalea

Unclassified South African Gold Mine Gp 1

Unclassified Terrestrial Group

Cultivated areaNative forest

5 10 15 20 250()

(a)

Unclassified Thermoplasmatales

Unclassified Terrestrial Miscellaneous Group

Rice Cluster I

Methanocella

Methanosarcina

Unclassified Euryarchaeota

Unclassified Halobacteriales

Candidatus Methanomethylophilus

Unclassified Methanosarcinaceae

5 10 15 20 250()

Cultivated areaNative forest

(b)

Figure 6 Percentage of archaeal operational taxonomic units within the Thaumarchaeota (a) and Euryarchaeota (b) phyla classified at thegenus level for native forest soil (orange) and soil under oil palm cultivation (blue) The percentage of unclassified sequences at genus level isshown

4 Discussion

The Amazon forest is known for its macrospecies diversityhowever few studies have addressed its microbial diversityAmong the domains of life Archaea are undoubtedly the lesswell known

In this work almost 600000 archaeal 16S rRNA genesequences were obtained from native forest soil and anadjacent area under oil palm cultivation After processing(ie removal of short sequences sequences with errors low-quality sequences chimeras and possible contaminants) andnormalization 39111 sequences per area remained for anal-ysis This number of sequences provided sufficient coverageof archaeal diversity in both areas (Table 1)The primers used

were based on a large number of 16S rRNA sequences fromthe SILVA database [28] however given that little is knownabout this domain it is unlikely that the archaeal diversitywasfully covered [43]

The main physicochemical differences between soils ofthe native forest and the oil palm-cultivated area were thelevels of phosphorous potassium calcium organic matterand total carbon (Table 2) Correlations between phylogenyand physicochemical parameters were evaluated using PCAand NMDS however no clear clusters or correlations wereobserved (results not shown)

In a study of the long-term effects of inorganic fertilizerson microbial communities Zhong and Cai [44] reportedthat phosphorus application indirectly altered microbial

Archaea 9

Candidatus NitrososphaeraUnclassified Soil Crenarchaeotic Group (SCG)

Unclassified South African Gold Mine Gp 1 (SAGMCG-1)Candidatus Nitrosotalea

Unclassified Terrestreial Miscellaneous Gp (TMEG)Unclassified Thaumarchaeota

Unclassified ThermoplasmatalesUnclassified Miscellaneous Crenarchaeotic Group

MethanosarcinaRice Cluster I

95 confidence intervals

q-v

alue

(cor

rect

ed)

Cultivated areaNative forest

531e minus 10

420 6 8minus4minus6minus8 minus2minus10

Difference between proportions ()38800

Proportion ()

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

Figure 7 Archaeal genera that differ significantly (119902 lt 005) in relative abundance between native forest soil (orange) and soil under oil palmcultivation (blue) For the purpose of statistical analysis performed using STAMP software the sum of the percentages of archaea classifiedat the genus level in both soils is 100 Differences between proportions of each genus are shown with 95 confidence intervals Unclassifiedsequences were used to calculate frequency profiles but are not displayed

parameters in soils and increased crop yields by elevating thelevels of organic matter Wessen and collaborators used real-time PCR quantification to show the impacts of fertilizer useon several bacterial phyla and the archaeal phylum Crenar-chaeota (Thaumarchaeota Group I1c after reanalysis usingthe SILVAdatabase 119) [45]They reported that bacterial andarchaeal classes respond differently to fertilization primarilybecause of changes in certain soil parameters such as pHtotal nitrogen and the carbon nitrogen ratio In our studylevels of total carbon and organic matter were higher in thenative forest than in the oil palm-cultivated area which mayhave contributed to higher archaeal diversity Although thereare no studies on the effect of total soil carbon on archaealcommunities there are studies on the bacterial communitiesin Amazonian soils The work of Edgar et al [33] describeddistinct soil bacterial communities under different vegetationtypes The authors found that bacterial diversity correlatedwith total organic carbon and total nitrogen content [46]Furthermore in a study of bacterial diversity and microbialbiomass in forest pasture and fallow soils in the Amazonbasin microbial biomass was highest in pasture soils whichalso had 30ndash47 higher carbon levels than the other sitesover the course of a year [47]

Several studies have also reported alterations in thecomposition of archaeal [48] bacterial [47] and fungal [49]communities associated with land use changes A studyof archaeal communities in Amazonian anthrosols undervarious land uses (agriculture pasture and secondary forest)and cultivation practices (eg eggplant banana citrus andmanioc) [10] reported that agriculture negatively affectedarchaeal community diversity Results of ribosomal inter-genic spacer analysis showed that soil type is also responsiblefor changes in archaeal communities in soils under differenttypes of land use (native grassland native forest Eucalyptus

and Acacia plantations and soybean and watermelon fields)[50]

The Venn diagram showed that the native forest soil has agreater number of unique sequences (Figure 2) This was notsurprising due to the greater archaeal richness and diversityobserved in the native forest soil (Table 2) The number ofunique OTUs found in the oil palm-cultivated area is almostthe same as that of shared OTUs with the native forest areaAlthough oil palm cultivation decreased archaeal diversity inthe soil there is still a core group of archaea OTUs sharedwith the native forest suggesting that these archaea havecertain characteristics that allow them to persist after thechange in land use It cannot be ruled out that these archaeaare metabolically inactive The smaller number of uniquesequences in the oil palm-cultivated area may be related to alower complexity of the vegetation (ie oil palmmonocultureversus great plant diversity in the forest) since plant diversitycould affect these communities [51]

In our study all OTUs identified belong to the domainArchaea and were classified into two phyla Euryarchaeotaand Thaumarchaeota It is important to mention that thedatabases were only recently updated to recognize the phy-lumThaumarchaeota therefore previous studies on Archaeacould not recognize this phylum as proposed by Brochier-Armanet et al [11] The creation of this novel phylum wassupported by a study comparing the genomes of two marineammonia-oxidizing archaea (AOAs) that identified a core setof informational genes specific to this group [52] Thereforeto allow comparisons with other studies we reanalyzed theirdata whenever possible updating the taxonomic classifica-tion to include the phylumThaumarchaeota

Thework ofChao and Shen [35] focused on the impacts ofland use (ie agricultural systems of indigenous people andcattle pasture) on archaeal communities by PCR (amoA gene)

10 Archaea

and denaturing gradient gel electrophoresis analysis of differ-ent Western Amazon soils The databases at the time did notconsiderThaumarchaeota in their classification however ourreanalysis of their dataset using an updated database showedthat Euryarchaeota and Thaumarchaeota were present intheir samples along with Crenarchaeota (Thermoprotei) Inaddition changes were observed in the structure of soilarchaeal communities when comparing the cultivated areaswith forest areas that had been converted to pasture

In our study differences in the percentages of Thau-marchaeota and Euryarchaeota found in the soil sampleswere not striking Thaumarchaeota were predominant in thenative forest (58) whereas Euryarchaeota were predom-inant in the oil palm-cultivated area with 54 (Figure 3)This result is consistent with other comparative studies onarchaeal communities in different types of Brazilian soils (ieAmazonian anthrosols Caatinga and Cerrado) [10 53 54]and a study of different tropical forest soil types that reportedThaumarchaeota as the predominant phylum [44]

The phylumThaumarchaeota comprises all known AOAsand is directly involved in nitrogen metabolism Ammonia isused as an energy source by Group I1a found predominantlyin aquatic systems [52] and Group I1b found in soils [55]Thaumarchaeota Group I1c is primarily found in acidic soilsbut the ecological roles of these archaea are still unknown [5657]

The literature presents conflicting data about the pro-portion of AOAsAOBs soils In some studies AOAs arepredominant [58] while in others AOBs are more abundant[59] However several studies found a predominance of thearchaeal amoA gene over the bacterial amoA gene in soils[60] AOAs are frequently detected in soils their abundancerange from 05 to 10 of the prokaryotic community andtheir abundance is strongly influenced by physicochemicalconditions as well as the organic matter content [61 62] Inacidic soils AOAs appear to have a more important role inammonia oxidation than AOBs [57]

The Soil Crenarchaeotic Group a class within Thaumar-chaeota according to the SILVA database classification wasdetected in both soil samples but a higher representationwas observed in native forest soil This class was discoveredin boreal forest soil [63] Candidatus Nitrososphaera whichbelongs to the Soil Crenarchaeotic Group class is a memberof Thaumarchaeota Group I1b it is an AOA that usesammonia or urea as an energy source [64 65]

On the other hand members of the phylum Eur-yarchaeota are directly involved in carbon metabolismwith several classes of methanogens and methane-oxidizingorganisms [66ndash68] Navarrete et al [48] studied the impactof converting Amazon native forest to pasture and reportedchanges not only to the composition and abundance of soilmicrobial communities but also to a decrease in the diversityof gene function

Within the phylum Euryarchaeota Methanomicrobiaand Thermoplasmata were the main classes detected in oursamples along with a few representatives of Methanobac-teria and Halobacteria Members of Methanobacteria andMethanomicrobia are methanogens [69] Methanomicrobiawere present in both native forest soil and oil palm-cultivated

soil with larger representation in the oil palm-cultivatedarea Within Methanomicrobia the genera Rice Cluster IMethanosarcina andMethanocellawere present in both sam-ples (Figure 6(b)) These genera are closely related and foundin rice fields that produce methane [70] The order Metha-nomicrobiales was detected only in the native forest soilwhich can probably be explained by the relatively high levelsof carbon and organic matter This archaeal order is respon-sible for the final step of methane production in the carboncycle by anaerobic degradation of organic matter [66 71]However the contribution of the Methanomicrobia to thecarbon cycle in the oil palm-cultivated soils could not beassessed since there are no data on the methane productionin the studied areas

Among the Thaumarchaeota sequences used to generatethe phylogenetic tree that were unclassified at the genuslevel several OTUs formed different clusters associated withtheMiscellaneous Crenarchaeotic Group (Figure 5(a))Thesesequences were from both soil samples indicating that theunclassified Thaumarchaeota in the native forest and the oilpalm-cultivated area are phylogenetically related

Most of the sequences remained unclassified at thegenus level for both Thaumarchaeota and Euryarchaeota(Figures 6(a) and 6(b)) This result likely reflects the fact thatmost taxonomic databases do not have many representativesof Archaea especially belonging to the phylum Thaumar-chaeota thus few classifications of new sequences reach thegenus level Several genera were identified as Candidatusaccording to the International Code of Nomenclature ofBacteria indicating that characterization of the culturedprokaryote is incomplete [72]

The proportions of only four groups classified at thegenus level differed significantly between native forest and oilpalm-cultivated soils (Methanosarcina and Rice Cluster I inthe phylum Euryarchaeota and Candidatus NitrososphaeraandCandidatusNitrosotalea in the phylumThaumarchaeota)(Figure 7) With respect to the methanogens sequencesrelated to Methanosarcina were slightly more abundant insoil under oil palm cultivation (Figure 7) whereas sequencesbelonging to the Rice Cluster I were more abundant in nativeforest soil

In the Thaumarchaeota phylum Candidatus Nitrosos-phaera an AOA member of group I1b [65] was detectedin both samples but had a much higher representation innative forest (native forest 182 oil palm-cultivated area103) (Figure 7) In a comparative study of the archaealcommunities in Amazonian dark earth (ldquoTerra Pretardquo)and adjacent soils Candidatus Nitrososphaera was themain archaeal group found in both soils [10] In contrastCandidatus Nitrosotalea another AOA was more abundantin the oil palm-cultivated area (100) than in native forestsoil (44) in our study (Figure 7) Lehtovirta-Morley et al[73] cultivated the obligate acidophilic ammonia oxidizerCandidatus Nitrosotalea devanaterra from a nitrifying acidicsoil Later the same group characterized two strains ofacidophilic AOA belonging to this genus from a Chinesepaddy field and a Scottish agricultural soil [74] In our studythe greater representation of Candidatus Nitrososphaeraand Candidatus Nitrosotalea in native forest and oil

Archaea 11

palm-cultivated soils respectively can be explained by thelow soil pH (native forest pH 550 oil palm-cultivated areapH 483) since in basic soils the activity and growth ofthese archaea are diminished by the exponential decrease innitrite [6 75] In addition soil under oil palm cultivationis fertilized with nitrogen which may explain the higherlevel of Candidatus Nitrosotalea in this area compared withthat of native forest soil Finally although the cultivatedmembers of the genus Ca Nitrososphaera grow betteron neutral pH they have been reported to grow on pHvalues ranging from 60 to 85 In addition a recent workpublished by Wang et al (2014) [76] described active AOAsfrom an acidic soil (pH 492) which are closely related toNitrososphaera known as Nitrososphaera-like organismsby molecular phylogenetic methods Most of our sequenceswere classified as Nitrosotalea-like which are sequencesbelonging to the group I1a This group is typically foundin aquatic environments such as lake sediments or marineenvironments A similar ubiquitous distribution in soils isfound for the group I1b which are not restricted to a specificsoil pH condition

5 Conclusions

Although archaea are known to be ubiquitous there is stillmuch to be learned about their diversity biological rolesand metabolic capabilities This work is the first to comparearchaeal communities associated with the Amazon forest soilwith those of soil under oil palm cultivation

Archaeal diversity was found to be lower in oil palm-cultivated soil than in the native forest soil Only about 30of the OTUs were found in the oil palm soil in comparisonto the native forest However two main phyla were detectedin both samples Thaumarchaeota and Euryarchaeota with apredominance ofThaumarchaeota in native forest soil Theseresults support previous studies in other Brazilian biomenative soils (eg Caatinga Cerrado) as well as soil underdifferent types of land use (eg pasture and agricultural) Itis also important to mention that there are also groups ofuncertain classification associated with the Crenarchaeotaphylum

Although knowledge about Archaea has increased dra-matically in the last few decades little is known aboutthe biology of many archaeal groups and the taxonomyof Archaea is still a work in progress Thus it is notsurprising thatmany sequences identified in thiswork remainunclassified sometimes even at higher taxonomic levels Itis not clear whether the relatively small overall differencebetween soil archaeal communities of the Amazon forest andoil palm-cultivated land is a consequence of the current stateof archaeal taxonomy or whether it indicates that archaealdiversity is lower than bacterial diversity [77]

It is also important to consider that with land usechange due to agricultural practices the microstructure ofthe soil is broken down which can reduce the natural habitatfragmentation allowing potential interactions among theresident microbiota that otherwise would not happen [7879]

A phylogenetic tree of unclassified Thaumarchaeota 16SrRNA gene sequences was useful in exploring relationshipsamong these sequences When taxonomic classification ofsequences was possible it revealed the presence of OTUsfrom genera known to be involved in the metabolism ofcarbon and nitrogen (methanogens Methanocella and Meth-anosarcina AOACandidatusNitrososphaera andCandidatusNitrosotalea) hinting at possible roles for these archaealcommunities in Amazon soils

More studies are needed to elucidate these roles Cultur-ing some of these archaea in the laboratory would allow theirmetabolism and physiology to be studied in detail allowingfor a better understanding of the roles of Archaea in the bio-geochemical cycles

Conflict of Interests

The authors declare that there is no conflict of interestsregarding the publication of this paper

Acknowledgments

This work was supported by grants from FINEP CNPqCAPES and FAP-DF The authors thank Dr Boari for helpwith sample collection logistics Dr Betania Ferraz Quirinowas supported by the Cientista Visitante program fromEmbrapa

References

[1] FAO Global Forest Resources Assessment Food and AgricultureOrganization of the United Nations 2010

[2] IBGE ldquoMapa de Biomas do Brasil primeira aproximacaordquo Riode Janeiro Brazil 2004 httpwwwibgegovbrhome

[3] C Azevedo-Ramos ldquoSustainable development and challengingdeforestation in the Brazilian Amazon the good the badand the uglyrdquo in Proceedings of the Symposium Our CommonGround Innovations in Land Use Decision-Making vol 59 p 5Vancouver Canada May 2007

[4] C A Francis K J Roberts J M Beman A E Santoro and B BOakley ldquoUbiquity and diversity of ammonia-oxidizing archaeainwater columns and sediments of the oceanrdquoProceedings of theNational Academy of Sciences of theUnited States of America vol102 no 41 pp 14683ndash14688 2005

[5] A H Treusch S Leininger A Kietzin S C Schuster H-PKlenk and C Schleper ldquoNovel genes for nitrite reductase andAmo-related proteins indicate a role of uncultivatedmesophiliccrenarchaeota in nitrogen cyclingrdquo Environmental Microbiol-ogy vol 7 no 12 pp 1985ndash1995 2005

[6] G W Nicol S Leininger C Schleper and J I Prosser ldquoTheinfluence of soil pH on the diversity abundance and tran-scriptional activity of ammonia oxidizing archaea and bacteriardquoEnvironmentalMicrobiology vol 10 no 11 pp 2966ndash2978 2008

[7] P Offre A Spang and C Schleper ldquoArchaea in biogeochemicalcyclesrdquo Annual Review of Microbiology vol 67 pp 437ndash4572013

[8] MAPA Anuario Estatıstico da Agroenergia 2012 StatisticalYearbook of Agroenergy 2013

[9] J C Bergmann D D Tupinamba O Y A Costa J R MAlmeida C C Barreto andB FQuirino ldquoBiodiesel production

12 Archaea

in Brazil and alternative biomass feedstocksrdquo Renewable ampSustainable Energy Reviews vol 21 pp 411ndash420 2013

[10] R G Taketani and S M Tsai ldquoThe influence of different landuses on the structure of archaeal communities in Amazoniananthrosols based on 16S rRNA and amoA genesrdquo MicrobialEcology vol 59 no 4 pp 734ndash743 2010

[11] C Brochier-Armanet B Boussau S Gribaldo and P ForterreldquoMesophilic crenarchaeota proposal for a third archaeal phy-lum theThaumarchaeotardquoNature Reviews Microbiology vol 6no 3 pp 245ndash252 2008

[12] S M Barns C F Delwiche J D Palmer and N R PaceldquoPerspectives on archaeal diversity thermophily and mono-phyly from environmental rRNA sequencesrdquo Proceedings of theNational Academy of Sciences of the United States of Americavol 93 no 17 pp 9188ndash9193 1996

[13] J G Elkins M Podar D E Graham et al ldquoA korarchaealgenome reveals insights into the evolution of the Archaeardquo Pro-ceedings of the National Academy of Sciences of the United Statesof America vol 105 no 23 pp 8102ndash8107 2008

[14] H Huber M J Hohn R Rachel T Fuchs V C Wimmer andK O Stetter ldquoA new phylum of Archaea represented by ananosized hyperthermophilic symbiontrdquo Nature vol 417 no6884 pp 63ndash67 2002

[15] T Nunoura H Oida M Nakaseama et al ldquoArchaeal diversityand distribution along thermal and geochemical gradients inhydrothermal sediments at the Yonaguni Knoll IV hydrother-mal field in the SouthernOkinawa TroughrdquoApplied and Enviro-nmental Microbiology vol 76 no 4 pp 1198ndash1211 2010

[16] C Rinke P Schwientek A Sczyrba et al ldquoInsights into thephylogeny and coding potential of microbial dark matterrdquoNature vol 499 no 7459 pp 431ndash437 2013

[17] A Spang J H Saw S L Joslashrgensen et al ldquoComplex archaea thatbridge the gap between prokaryotes and eukaryotesrdquo Naturevol 521 no 7551 pp 173ndash179 2015

[18] S Bagchi S E Vlaeminck L A Sauder et al ldquoTemporal andspatial stability of ammonia-oxidizing archaea and bacteria inaquariumbiofiltersrdquo PLoSONE vol 9 no 12 Article ID e1135152014

[19] W Qin S A Amin W Martens-Habbena et al ldquoMarineammonia-oxidizing archaeal isolates display obligate mixotro-phy and wide ecotypic variationrdquo Proceedings of the NationalAcademy of Sciences of the United States of America vol 111 no34 pp 12504ndash12509 2014

[20] M Coci N Odermatt M M Salcher J Pernthaler and GCorno ldquoEcology and distribution ofThaumarchaea in the deephypolimnion of Lake Maggiorerdquo Archaea vol 2015 Article ID590434 11 pages 2015

[21] J G Ferry ldquoHow tomake a living by exhalingmethanerdquoAnnualReview of Microbiology vol 64 pp 453ndash473 2010

[22] A Meyer A Focks V Radl G Welzl I Schoning and MSchloter ldquoInfluence of land use intensity on the diversity ofammonia oxidizing bacteria and archaea in soils from grasslandecosystemsrdquoMicrobial Ecology vol 67 no 1 pp 161ndash166 2014

[23] V De Gannes G Eudoxie and W J Hickey ldquoImpacts ofedaphic factors on communities of ammonia-oxidizing archaeaammonia-oxidizing bacteria and nitrification in tropical soilsrdquoPLoS ONE vol 9 no 2 Article ID e89568 2014

[24] D S Ribeiro F C J da Silva and TN Carvalho ldquoSobrevivenciade seis especies florestais em uma area explorada seletivamenteno municıpio de Moju Parardquo CERNE vol 9 no 2 pp 153ndash1632003

[25] D HM Costa C A P Ferreira J NM Silva J D C A Lopesand J O P D Carvalho ldquoPotencial madeireiro de floresta densano municıpio de Moju Estado do Parardquo Documentos 1998

[26] C M Almeida S F Lima C V Martins-Da-Silva and JI Gomes ldquoCaracterizacao morfologica e anatomica de dezespecies de leguminosae ocorrentes em uma floresta tropicalumida localizada no municıpio de Moju Estado do Parardquo inA Silvicultura na Amazonia Oriental Contribuicoes do ProjetoEmbrapa-DFID J NMC Silva JO P deCarvalho and J AGYared Eds pp 19ndash54 Embrapa Amazonia Oriental (CPATU)Belem Brazil 2001

[27] K Smalla N Cresswell L C Mendoncahagler A Woltersand J D Vanelsas ldquoRapid extraction protocol from soil forpolymerase chain reaction-mediated amplificationrdquo Journal ofApplied Bacteriology vol 74 pp 78ndash85 1993

[28] S Gantner A F Andersson L Alonso-Saez and S Bertils-son ldquoNovel primers for 16S rRNA-based archaeal communityanalyses in environmental samplesrdquo Journal of MicrobiologicalMethods vol 84 no 1 pp 12ndash18 2011

[29] P D Schloss S L Westcott T Ryabin et al ldquoIntroducingMOTHUR open-source platform-independent community-supported software for describing and comparing microbialcommunitiesrdquoApplied and EnvironmentalMicrobiology vol 75no 23 pp 7537ndash7541 2009

[30] PD SchlossDGevers and S LWestcott ldquoReducing the effectsof PCR amplification and sequencing artifacts on 16S rRNA-based studiesrdquo PLoS ONE vol 6 no 12 Article ID e27310 2011

[31] CQuince A Lanzen T P Curtis et al ldquoAccurate determinationof microbial diversity from 454 pyrosequencing datardquo NatureMethods vol 6 no 9 pp 639ndash641 2009

[32] P Yilmaz L W Parfrey P Yarza et al ldquoThe SILVA and lsquoall-species Living Tree Project (LTP)rsquo taxonomic frameworksrdquoNucleic Acids Research vol 42 no 1 pp D643ndashD648 2014

[33] R C Edgar B J Haas J C Clemente C Quince and RKnight ldquoUCHIME improves sensitivity and speed of chimeradetectionrdquo Bioinformatics vol 27 no 16 pp 2194ndash2200 2011

[34] A Chao and T-J Shen ldquoNonparametric estimation of Shan-nonrsquos index of diversity when there are unseen species insamplerdquo Environmental and Ecological Statistics vol 10 no 4pp 429ndash443 2003

[35] A Chao and T-J Shen Program SPADE (Species Prediction andDiversity Estimation) 2010

[36] C L Shannon andWWeaverTheMathematicalTheory of Com-munication University of Illinois Press Urbana Ill USA 1949

[37] I J Good ldquoThe population frequencies of species and theestimation of population parametersrdquo Biometrika vol 40 pp237ndash264 1953

[38] K Tamura G Stecher D Peterson A Filipski and S KumarldquoMEGA6molecular evolutionary genetics analysis version 60rdquoMolecular Biology and Evolution vol 30 no 12 pp 2725ndash27292013

[39] DA BensonMCavanaugh K Clark et al ldquoGenBankrdquoNucleicAcids Research vol 41 no 1 pp D36ndashD42 2013

[40] D H Parks G W Tyson P Hugenholtz and R G BeikoldquoSTAMP statistical analysis of taxonomic and functional pro-filesrdquo Bioinformatics vol 30 no 21 pp 3123ndash3124 2014

[41] R G Newcombe ldquoInterval estimation for the differencebetween independent proportions comparison of elevenmeth-odsrdquo Statistics in Medicine vol 17 no 8 pp 873ndash890 1998

[42] J D Storey J E Taylor and D Siegmund ldquoStrong controlconservative point estimation and simultaneous conservative

Archaea 13

consistency of false discovery rates a unified approachrdquo Journalof the Royal Statistical Society Series B Statistical Methodologyvol 66 no 1 pp 187ndash205 2004

[43] G C Baker J J Smith and D A Cowan ldquoReview and re-analysis of domain-specific 16S primersrdquo Journal of Microbio-logical Methods vol 55 no 3 pp 541ndash555 2003

[44] W H Zhong and Z C Cai ldquoLong-term effects of inorganicfertilizers on microbial biomass and community functionaldiversity in a paddy soil derived from quaternary red clayrdquoApplied Soil Ecology vol 36 no 2-3 pp 84ndash91 2007

[45] EWessen S Hallin and L Philippot ldquoDifferential responses ofbacterial and archaeal groups at high taxonomical ranks to soilmanagementrdquo Soil Biology and Biochemistry vol 42 no 10 pp1759ndash1765 2010

[46] ZWuW Lin B Li et al ldquoTerminal restriction fragment lengthpolymorphism analysis of soil bacterial communities underdifferent vegetation types in subtropical area rdquo PLOS ONE vol10 no 6 Article ID e0129397 2015

[47] K Cenciani M R Lambais C C Cerri L C B De Azevedoand B J Feigl ldquoBacteria diversity and microbial biomass inforest pasture and fallow soils in the southwestern Amazonbasinrdquo Revista Brasileira de Ciencia do Solo vol 33 no 4 pp907ndash916 2009

[48] A A Navarrete R G Taketani L WMendes F de Souza Can-navan F M de Souza Moreira and S M Tsai ldquoLand-use sys-tems affect Archaeal community structure and functional diver-sity in western Amazon soilsrdquo Revista Brasileira de Ciencia doSolo vol 35 no 5 pp 1527ndash1540 2011

[49] E Lumini A Orgiazzi R Borriello P Bonfante and V Bian-ciotto ldquoDisclosing arbuscular mycorrhizal fungal biodiversityin soil through a land-use gradient using a pyrosequencingapproachrdquo Environmental Microbiology vol 12 no 8 pp 2165ndash2179 2010

[50] M Lupatini R J S Jacques Z I Antoniolli A K A SuleimanR R Fulthorpe and L F W Roesch ldquoLand-use change andsoil type are drivers of fungal and archaeal communities in thePampa biomerdquo World Journal of Microbiology and Biotechnol-ogy vol 29 no 2 pp 223ndash233 2013

[51] E G Lamb N Kennedy and S D Siciliano ldquoEffects of plantspecies richness and evenness on soil microbial communitydiversity and functionrdquo Plant and Soil vol 338 no 1 pp 483ndash495 2011

[52] A Spang R Hatzenpichler C Brochier-Armanet et al ldquoDis-tinct gene set in two different lineages of ammonia-oxidizingarchaea supports the phylum Thaumarchaeotardquo Trends inMicrobiology vol 18 no 8 pp 331ndash340 2010

[53] R G Pacchioni F M Carvalho C E Thompson et al ldquoTax-onomic and functional profiles of soil samples from Atlanticforest and Caatinga biomes in northeastern Brazilrdquo Microbio-logyOpen vol 3 no 3 pp 299ndash315 2014

[54] E Catao A P Castro C C Barreto R H Kruger and C MKyaw ldquoDiversity of Archaea in Brazilian savanna soilsrdquoArchivesof Microbiology vol 195 no 7 pp 507ndash512 2013

[55] M Pester T Rattei S Flechl et al ldquoAmoA-based consensusphylogeny of ammonia-oxidizing archaea and deep sequencingof amoA genes from soils of four different geographic regionsrdquoEnvironmental Microbiology vol 14 no 2 pp 525ndash539 2012

[56] J-Z He H-W Hu and L-M Zhang ldquoCurrent insights into theautotrophic thaumarchaeal ammonia oxidation in acidic soilsrdquoSoil Biology amp Biochemistry vol 55 pp 146ndash154 2012

[57] L-M Zhang H-W Hu J-P Shen and J-Z He ldquoAmmonia-oxidizing archaea have more important role than ammonia-oxidizing bacteria in ammonia oxidation of strongly acidicsoilsrdquo ISME Journal vol 6 no 5 pp 1032ndash1045 2012

[58] S Leininger T Urich M Schloter et al ldquoArchaea predominateamong ammonia-oxidizing prokaryotes in soilsrdquo Nature vol442 no 7104 pp 806ndash809 2006

[59] N C Banning L D Maccarone L M Fisk and D VMurphy ldquoAmmonia-oxidising bacteria not archaea dominatenitrification activity in semi-arid agricultural soilrdquo ScientificReports vol 5 Article ID 11146 2015

[60] M Herrmann A Hadrich and K Kusel ldquoPredominance ofthaumarchaeal ammonia oxidizer abundance and transcrip-tional activity in an acidic fenrdquoEnvironmentalMicrobiology vol14 no 11 pp 3013ndash3025 2012

[61] K Zhalnina P Dorr de Quadros F O Camargo and E WTriplett ldquoDrivers of archaeal ammonia-oxidizing communitiesin soilrdquo Frontiers in Microbiology vol 3 article 210 Article IDArticle 210 2012

[62] J Hong J Cho and Y Hong ldquoEnvironmental variables shapingthe ecological niche of Thaumarchaeota in soil direct andindirect causal effectsrdquo PLoS ONE vol 10 no 8 Article IDe0133763 2015

[63] G Jurgens K Lindstrom and A Saano ldquoNovel group withinthe kingdom Crenarchaeota from boreal forest soilrdquo Appliedand Environmental Microbiology vol 63 no 2 pp 803ndash8051997

[64] M Tourna M Stieglmeier A Spang et al ldquoNitrososphaeraviennensis an ammonia oxidizing archaeon from soilrdquoProceed-ings of the National Academy of Sciences of the United States ofAmerica vol 108 no 20 pp 8420ndash8425 2011

[65] K V Zhalnina R Dias M T Leonard et al ldquoGenome sequenceof Candidatus Nitrososphaera evergladensis from group I1benriched from everglades soil reveals novel genomic features ofthe ammonia-oxidizing archaeardquoPLoSONE vol 9 no 7 ArticleID e101648 2014

[66] I Anderson L E Ulrich B Lupa et al ldquoGenomic char-acterization of methanomicrobiales reveals three classes ofmethanogensrdquo PLoS ONE vol 4 no 6 Article ID e5797 2009

[67] J E Galagan C Nusbaum A Roy et al ldquoThe genome of Macetivorans reveals extensive metabolic and physiological diver-sityrdquo Genome Research vol 12 no 4 pp 532ndash542 2002

[68] C Wrede S Kokoschka A Dreier C Heller J Reitner and MHoppert ldquoDeposition of biogenic iron minerals in a methaneoxidizing microbial matrdquo Archaea vol 2013 Article ID 1029728 pages 2013

[69] G Garrity Bergeyrsquo Manual of Systematic Bacteriology 721 2001[70] S Sakai Y Takaki S Shimamura et al ldquoGenome sequence

of a mesophilic hydrogenotrophic methanogen Methanocellapaludicola the first cultivated representative of the orderMethanocellalesrdquo PLoS ONE vol 6 no 7 Article ID e228982011

[71] E Bapteste C Brochier and Y Boucher ldquoHigher-level clas-sification of the Archaea evolution of methanogenesis andmethanogensrdquo Archaea vol 1 no 5 pp 353ndash363 2005

[72] R G E Murray and E Stackebrandt ldquoTaxonomic note imple-mentation of the provisional status Candidatus for incompletelydescribed procaryotesrdquo International Journal of Systematic Bac-teriology vol 45 no 1 pp 186ndash187 1995

[73] L E Lehtovirta-Morley K Stoecker A Vilcinskas J I Prosserand G W Nicol ldquoCultivation of an obligate acidophilic ammo-nia oxidizer from a nitrifying acid soilrdquo Proceedings of the

14 Archaea

National Academy of Sciences of the United States of Americavol 108 no 38 pp 15892ndash15897 2011

[74] L E Lehtovirta-Morley C Ge J Ross H Yao G W Nicol andJ I Prosser ldquoCharacterisation of terrestrial acidophilic archaealammonia oxidisers and their inhibition and stimulation byorganic compoundsrdquo FEMSMicrobiology Ecology vol 89 no 3pp 542ndash552 2015

[75] M J Frijlink T Abee H J Laanbroek W de Boer and W NKonings ldquoThe bioenergetics of ammonia and hydroxylamineoxidation in Nitrosomonas europaea at acid and alkaline pHrdquoArchives of Microbiology vol 157 no 2 pp 194ndash199 1992

[76] B Wang Y Zheng R Huang et al ldquoActive ammonia oxidizersin an acidic soil are phylogenetically closely related to neu-trophilic archaeonrdquo Applied and Environmental Microbiologyvol 80 no 5 pp 1684ndash1691 2014

[77] J Y Aller and P F Kemp ldquoAre Archaea inherently less diversethan Bacteria in the same environmentsrdquo FEMS MicrobiologyEcology vol 65 no 1 pp 74ndash87 2008

[78] K Potthast U Hamer and FMakeschin ldquoLand-use change in atropical mountain rainforest region of southern Ecuador affectssoilmicroorganisms and nutrient cyclingrdquoBiogeochemistry vol111 no 1ndash3 pp 151ndash167 2012

[79] J I Prosser and G W Nicol ldquoArchaeal and bacterial ammonia-oxidisers in soil the quest for niche specialisation and differen-tiationrdquoTrends inMicrobiology vol 20 no 11 pp 523ndash531 2012

Submit your manuscripts athttpwwwhindawicom

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Anatomy Research International

PeptidesInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporation httpwwwhindawicom

International Journal of

Volume 2014

Zoology

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Molecular Biology International

GenomicsInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioinformaticsAdvances in

Marine BiologyJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Signal TransductionJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioMed Research International

Evolutionary BiologyInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Biochemistry Research International

ArchaeaHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Genetics Research International

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Advances in

Virolog y

Hindawi Publishing Corporationhttpwwwhindawicom

Nucleic AcidsJournal of

Volume 2014

Stem CellsInternational

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

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Enzyme Research

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

International Journal of

Microbiology

Archaea 9

Candidatus NitrososphaeraUnclassified Soil Crenarchaeotic Group (SCG)

Unclassified South African Gold Mine Gp 1 (SAGMCG-1)Candidatus Nitrosotalea

Unclassified Terrestreial Miscellaneous Gp (TMEG)Unclassified Thaumarchaeota

Unclassified ThermoplasmatalesUnclassified Miscellaneous Crenarchaeotic Group

MethanosarcinaRice Cluster I

95 confidence intervals

q-v

alue

(cor

rect

ed)

Cultivated areaNative forest

531e minus 10

420 6 8minus4minus6minus8 minus2minus10

Difference between proportions ()38800

Proportion ()

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

lt1e minus 15

Figure 7 Archaeal genera that differ significantly (119902 lt 005) in relative abundance between native forest soil (orange) and soil under oil palmcultivation (blue) For the purpose of statistical analysis performed using STAMP software the sum of the percentages of archaea classifiedat the genus level in both soils is 100 Differences between proportions of each genus are shown with 95 confidence intervals Unclassifiedsequences were used to calculate frequency profiles but are not displayed

parameters in soils and increased crop yields by elevating thelevels of organic matter Wessen and collaborators used real-time PCR quantification to show the impacts of fertilizer useon several bacterial phyla and the archaeal phylum Crenar-chaeota (Thaumarchaeota Group I1c after reanalysis usingthe SILVAdatabase 119) [45]They reported that bacterial andarchaeal classes respond differently to fertilization primarilybecause of changes in certain soil parameters such as pHtotal nitrogen and the carbon nitrogen ratio In our studylevels of total carbon and organic matter were higher in thenative forest than in the oil palm-cultivated area which mayhave contributed to higher archaeal diversity Although thereare no studies on the effect of total soil carbon on archaealcommunities there are studies on the bacterial communitiesin Amazonian soils The work of Edgar et al [33] describeddistinct soil bacterial communities under different vegetationtypes The authors found that bacterial diversity correlatedwith total organic carbon and total nitrogen content [46]Furthermore in a study of bacterial diversity and microbialbiomass in forest pasture and fallow soils in the Amazonbasin microbial biomass was highest in pasture soils whichalso had 30ndash47 higher carbon levels than the other sitesover the course of a year [47]

Several studies have also reported alterations in thecomposition of archaeal [48] bacterial [47] and fungal [49]communities associated with land use changes A studyof archaeal communities in Amazonian anthrosols undervarious land uses (agriculture pasture and secondary forest)and cultivation practices (eg eggplant banana citrus andmanioc) [10] reported that agriculture negatively affectedarchaeal community diversity Results of ribosomal inter-genic spacer analysis showed that soil type is also responsiblefor changes in archaeal communities in soils under differenttypes of land use (native grassland native forest Eucalyptus

and Acacia plantations and soybean and watermelon fields)[50]

The Venn diagram showed that the native forest soil has agreater number of unique sequences (Figure 2) This was notsurprising due to the greater archaeal richness and diversityobserved in the native forest soil (Table 2) The number ofunique OTUs found in the oil palm-cultivated area is almostthe same as that of shared OTUs with the native forest areaAlthough oil palm cultivation decreased archaeal diversity inthe soil there is still a core group of archaea OTUs sharedwith the native forest suggesting that these archaea havecertain characteristics that allow them to persist after thechange in land use It cannot be ruled out that these archaeaare metabolically inactive The smaller number of uniquesequences in the oil palm-cultivated area may be related to alower complexity of the vegetation (ie oil palmmonocultureversus great plant diversity in the forest) since plant diversitycould affect these communities [51]

In our study all OTUs identified belong to the domainArchaea and were classified into two phyla Euryarchaeotaand Thaumarchaeota It is important to mention that thedatabases were only recently updated to recognize the phy-lumThaumarchaeota therefore previous studies on Archaeacould not recognize this phylum as proposed by Brochier-Armanet et al [11] The creation of this novel phylum wassupported by a study comparing the genomes of two marineammonia-oxidizing archaea (AOAs) that identified a core setof informational genes specific to this group [52] Thereforeto allow comparisons with other studies we reanalyzed theirdata whenever possible updating the taxonomic classifica-tion to include the phylumThaumarchaeota

Thework ofChao and Shen [35] focused on the impacts ofland use (ie agricultural systems of indigenous people andcattle pasture) on archaeal communities by PCR (amoA gene)

10 Archaea

and denaturing gradient gel electrophoresis analysis of differ-ent Western Amazon soils The databases at the time did notconsiderThaumarchaeota in their classification however ourreanalysis of their dataset using an updated database showedthat Euryarchaeota and Thaumarchaeota were present intheir samples along with Crenarchaeota (Thermoprotei) Inaddition changes were observed in the structure of soilarchaeal communities when comparing the cultivated areaswith forest areas that had been converted to pasture

In our study differences in the percentages of Thau-marchaeota and Euryarchaeota found in the soil sampleswere not striking Thaumarchaeota were predominant in thenative forest (58) whereas Euryarchaeota were predom-inant in the oil palm-cultivated area with 54 (Figure 3)This result is consistent with other comparative studies onarchaeal communities in different types of Brazilian soils (ieAmazonian anthrosols Caatinga and Cerrado) [10 53 54]and a study of different tropical forest soil types that reportedThaumarchaeota as the predominant phylum [44]

The phylumThaumarchaeota comprises all known AOAsand is directly involved in nitrogen metabolism Ammonia isused as an energy source by Group I1a found predominantlyin aquatic systems [52] and Group I1b found in soils [55]Thaumarchaeota Group I1c is primarily found in acidic soilsbut the ecological roles of these archaea are still unknown [5657]

The literature presents conflicting data about the pro-portion of AOAsAOBs soils In some studies AOAs arepredominant [58] while in others AOBs are more abundant[59] However several studies found a predominance of thearchaeal amoA gene over the bacterial amoA gene in soils[60] AOAs are frequently detected in soils their abundancerange from 05 to 10 of the prokaryotic community andtheir abundance is strongly influenced by physicochemicalconditions as well as the organic matter content [61 62] Inacidic soils AOAs appear to have a more important role inammonia oxidation than AOBs [57]

The Soil Crenarchaeotic Group a class within Thaumar-chaeota according to the SILVA database classification wasdetected in both soil samples but a higher representationwas observed in native forest soil This class was discoveredin boreal forest soil [63] Candidatus Nitrososphaera whichbelongs to the Soil Crenarchaeotic Group class is a memberof Thaumarchaeota Group I1b it is an AOA that usesammonia or urea as an energy source [64 65]

On the other hand members of the phylum Eur-yarchaeota are directly involved in carbon metabolismwith several classes of methanogens and methane-oxidizingorganisms [66ndash68] Navarrete et al [48] studied the impactof converting Amazon native forest to pasture and reportedchanges not only to the composition and abundance of soilmicrobial communities but also to a decrease in the diversityof gene function

Within the phylum Euryarchaeota Methanomicrobiaand Thermoplasmata were the main classes detected in oursamples along with a few representatives of Methanobac-teria and Halobacteria Members of Methanobacteria andMethanomicrobia are methanogens [69] Methanomicrobiawere present in both native forest soil and oil palm-cultivated

soil with larger representation in the oil palm-cultivatedarea Within Methanomicrobia the genera Rice Cluster IMethanosarcina andMethanocellawere present in both sam-ples (Figure 6(b)) These genera are closely related and foundin rice fields that produce methane [70] The order Metha-nomicrobiales was detected only in the native forest soilwhich can probably be explained by the relatively high levelsof carbon and organic matter This archaeal order is respon-sible for the final step of methane production in the carboncycle by anaerobic degradation of organic matter [66 71]However the contribution of the Methanomicrobia to thecarbon cycle in the oil palm-cultivated soils could not beassessed since there are no data on the methane productionin the studied areas

Among the Thaumarchaeota sequences used to generatethe phylogenetic tree that were unclassified at the genuslevel several OTUs formed different clusters associated withtheMiscellaneous Crenarchaeotic Group (Figure 5(a))Thesesequences were from both soil samples indicating that theunclassified Thaumarchaeota in the native forest and the oilpalm-cultivated area are phylogenetically related

Most of the sequences remained unclassified at thegenus level for both Thaumarchaeota and Euryarchaeota(Figures 6(a) and 6(b)) This result likely reflects the fact thatmost taxonomic databases do not have many representativesof Archaea especially belonging to the phylum Thaumar-chaeota thus few classifications of new sequences reach thegenus level Several genera were identified as Candidatusaccording to the International Code of Nomenclature ofBacteria indicating that characterization of the culturedprokaryote is incomplete [72]

The proportions of only four groups classified at thegenus level differed significantly between native forest and oilpalm-cultivated soils (Methanosarcina and Rice Cluster I inthe phylum Euryarchaeota and Candidatus NitrososphaeraandCandidatusNitrosotalea in the phylumThaumarchaeota)(Figure 7) With respect to the methanogens sequencesrelated to Methanosarcina were slightly more abundant insoil under oil palm cultivation (Figure 7) whereas sequencesbelonging to the Rice Cluster I were more abundant in nativeforest soil

In the Thaumarchaeota phylum Candidatus Nitrosos-phaera an AOA member of group I1b [65] was detectedin both samples but had a much higher representation innative forest (native forest 182 oil palm-cultivated area103) (Figure 7) In a comparative study of the archaealcommunities in Amazonian dark earth (ldquoTerra Pretardquo)and adjacent soils Candidatus Nitrososphaera was themain archaeal group found in both soils [10] In contrastCandidatus Nitrosotalea another AOA was more abundantin the oil palm-cultivated area (100) than in native forestsoil (44) in our study (Figure 7) Lehtovirta-Morley et al[73] cultivated the obligate acidophilic ammonia oxidizerCandidatus Nitrosotalea devanaterra from a nitrifying acidicsoil Later the same group characterized two strains ofacidophilic AOA belonging to this genus from a Chinesepaddy field and a Scottish agricultural soil [74] In our studythe greater representation of Candidatus Nitrososphaeraand Candidatus Nitrosotalea in native forest and oil

Archaea 11

palm-cultivated soils respectively can be explained by thelow soil pH (native forest pH 550 oil palm-cultivated areapH 483) since in basic soils the activity and growth ofthese archaea are diminished by the exponential decrease innitrite [6 75] In addition soil under oil palm cultivationis fertilized with nitrogen which may explain the higherlevel of Candidatus Nitrosotalea in this area compared withthat of native forest soil Finally although the cultivatedmembers of the genus Ca Nitrososphaera grow betteron neutral pH they have been reported to grow on pHvalues ranging from 60 to 85 In addition a recent workpublished by Wang et al (2014) [76] described active AOAsfrom an acidic soil (pH 492) which are closely related toNitrososphaera known as Nitrososphaera-like organismsby molecular phylogenetic methods Most of our sequenceswere classified as Nitrosotalea-like which are sequencesbelonging to the group I1a This group is typically foundin aquatic environments such as lake sediments or marineenvironments A similar ubiquitous distribution in soils isfound for the group I1b which are not restricted to a specificsoil pH condition

5 Conclusions

Although archaea are known to be ubiquitous there is stillmuch to be learned about their diversity biological rolesand metabolic capabilities This work is the first to comparearchaeal communities associated with the Amazon forest soilwith those of soil under oil palm cultivation

Archaeal diversity was found to be lower in oil palm-cultivated soil than in the native forest soil Only about 30of the OTUs were found in the oil palm soil in comparisonto the native forest However two main phyla were detectedin both samples Thaumarchaeota and Euryarchaeota with apredominance ofThaumarchaeota in native forest soil Theseresults support previous studies in other Brazilian biomenative soils (eg Caatinga Cerrado) as well as soil underdifferent types of land use (eg pasture and agricultural) Itis also important to mention that there are also groups ofuncertain classification associated with the Crenarchaeotaphylum

Although knowledge about Archaea has increased dra-matically in the last few decades little is known aboutthe biology of many archaeal groups and the taxonomyof Archaea is still a work in progress Thus it is notsurprising thatmany sequences identified in thiswork remainunclassified sometimes even at higher taxonomic levels Itis not clear whether the relatively small overall differencebetween soil archaeal communities of the Amazon forest andoil palm-cultivated land is a consequence of the current stateof archaeal taxonomy or whether it indicates that archaealdiversity is lower than bacterial diversity [77]

It is also important to consider that with land usechange due to agricultural practices the microstructure ofthe soil is broken down which can reduce the natural habitatfragmentation allowing potential interactions among theresident microbiota that otherwise would not happen [7879]

A phylogenetic tree of unclassified Thaumarchaeota 16SrRNA gene sequences was useful in exploring relationshipsamong these sequences When taxonomic classification ofsequences was possible it revealed the presence of OTUsfrom genera known to be involved in the metabolism ofcarbon and nitrogen (methanogens Methanocella and Meth-anosarcina AOACandidatusNitrososphaera andCandidatusNitrosotalea) hinting at possible roles for these archaealcommunities in Amazon soils

More studies are needed to elucidate these roles Cultur-ing some of these archaea in the laboratory would allow theirmetabolism and physiology to be studied in detail allowingfor a better understanding of the roles of Archaea in the bio-geochemical cycles

Conflict of Interests

The authors declare that there is no conflict of interestsregarding the publication of this paper

Acknowledgments

This work was supported by grants from FINEP CNPqCAPES and FAP-DF The authors thank Dr Boari for helpwith sample collection logistics Dr Betania Ferraz Quirinowas supported by the Cientista Visitante program fromEmbrapa

References

[1] FAO Global Forest Resources Assessment Food and AgricultureOrganization of the United Nations 2010

[2] IBGE ldquoMapa de Biomas do Brasil primeira aproximacaordquo Riode Janeiro Brazil 2004 httpwwwibgegovbrhome

[3] C Azevedo-Ramos ldquoSustainable development and challengingdeforestation in the Brazilian Amazon the good the badand the uglyrdquo in Proceedings of the Symposium Our CommonGround Innovations in Land Use Decision-Making vol 59 p 5Vancouver Canada May 2007

[4] C A Francis K J Roberts J M Beman A E Santoro and B BOakley ldquoUbiquity and diversity of ammonia-oxidizing archaeainwater columns and sediments of the oceanrdquoProceedings of theNational Academy of Sciences of theUnited States of America vol102 no 41 pp 14683ndash14688 2005

[5] A H Treusch S Leininger A Kietzin S C Schuster H-PKlenk and C Schleper ldquoNovel genes for nitrite reductase andAmo-related proteins indicate a role of uncultivatedmesophiliccrenarchaeota in nitrogen cyclingrdquo Environmental Microbiol-ogy vol 7 no 12 pp 1985ndash1995 2005

[6] G W Nicol S Leininger C Schleper and J I Prosser ldquoTheinfluence of soil pH on the diversity abundance and tran-scriptional activity of ammonia oxidizing archaea and bacteriardquoEnvironmentalMicrobiology vol 10 no 11 pp 2966ndash2978 2008

[7] P Offre A Spang and C Schleper ldquoArchaea in biogeochemicalcyclesrdquo Annual Review of Microbiology vol 67 pp 437ndash4572013

[8] MAPA Anuario Estatıstico da Agroenergia 2012 StatisticalYearbook of Agroenergy 2013

[9] J C Bergmann D D Tupinamba O Y A Costa J R MAlmeida C C Barreto andB FQuirino ldquoBiodiesel production

12 Archaea

in Brazil and alternative biomass feedstocksrdquo Renewable ampSustainable Energy Reviews vol 21 pp 411ndash420 2013

[10] R G Taketani and S M Tsai ldquoThe influence of different landuses on the structure of archaeal communities in Amazoniananthrosols based on 16S rRNA and amoA genesrdquo MicrobialEcology vol 59 no 4 pp 734ndash743 2010

[11] C Brochier-Armanet B Boussau S Gribaldo and P ForterreldquoMesophilic crenarchaeota proposal for a third archaeal phy-lum theThaumarchaeotardquoNature Reviews Microbiology vol 6no 3 pp 245ndash252 2008

[12] S M Barns C F Delwiche J D Palmer and N R PaceldquoPerspectives on archaeal diversity thermophily and mono-phyly from environmental rRNA sequencesrdquo Proceedings of theNational Academy of Sciences of the United States of Americavol 93 no 17 pp 9188ndash9193 1996

[13] J G Elkins M Podar D E Graham et al ldquoA korarchaealgenome reveals insights into the evolution of the Archaeardquo Pro-ceedings of the National Academy of Sciences of the United Statesof America vol 105 no 23 pp 8102ndash8107 2008

[14] H Huber M J Hohn R Rachel T Fuchs V C Wimmer andK O Stetter ldquoA new phylum of Archaea represented by ananosized hyperthermophilic symbiontrdquo Nature vol 417 no6884 pp 63ndash67 2002

[15] T Nunoura H Oida M Nakaseama et al ldquoArchaeal diversityand distribution along thermal and geochemical gradients inhydrothermal sediments at the Yonaguni Knoll IV hydrother-mal field in the SouthernOkinawa TroughrdquoApplied and Enviro-nmental Microbiology vol 76 no 4 pp 1198ndash1211 2010

[16] C Rinke P Schwientek A Sczyrba et al ldquoInsights into thephylogeny and coding potential of microbial dark matterrdquoNature vol 499 no 7459 pp 431ndash437 2013

[17] A Spang J H Saw S L Joslashrgensen et al ldquoComplex archaea thatbridge the gap between prokaryotes and eukaryotesrdquo Naturevol 521 no 7551 pp 173ndash179 2015

[18] S Bagchi S E Vlaeminck L A Sauder et al ldquoTemporal andspatial stability of ammonia-oxidizing archaea and bacteria inaquariumbiofiltersrdquo PLoSONE vol 9 no 12 Article ID e1135152014

[19] W Qin S A Amin W Martens-Habbena et al ldquoMarineammonia-oxidizing archaeal isolates display obligate mixotro-phy and wide ecotypic variationrdquo Proceedings of the NationalAcademy of Sciences of the United States of America vol 111 no34 pp 12504ndash12509 2014

[20] M Coci N Odermatt M M Salcher J Pernthaler and GCorno ldquoEcology and distribution ofThaumarchaea in the deephypolimnion of Lake Maggiorerdquo Archaea vol 2015 Article ID590434 11 pages 2015

[21] J G Ferry ldquoHow tomake a living by exhalingmethanerdquoAnnualReview of Microbiology vol 64 pp 453ndash473 2010

[22] A Meyer A Focks V Radl G Welzl I Schoning and MSchloter ldquoInfluence of land use intensity on the diversity ofammonia oxidizing bacteria and archaea in soils from grasslandecosystemsrdquoMicrobial Ecology vol 67 no 1 pp 161ndash166 2014

[23] V De Gannes G Eudoxie and W J Hickey ldquoImpacts ofedaphic factors on communities of ammonia-oxidizing archaeaammonia-oxidizing bacteria and nitrification in tropical soilsrdquoPLoS ONE vol 9 no 2 Article ID e89568 2014

[24] D S Ribeiro F C J da Silva and TN Carvalho ldquoSobrevivenciade seis especies florestais em uma area explorada seletivamenteno municıpio de Moju Parardquo CERNE vol 9 no 2 pp 153ndash1632003

[25] D HM Costa C A P Ferreira J NM Silva J D C A Lopesand J O P D Carvalho ldquoPotencial madeireiro de floresta densano municıpio de Moju Estado do Parardquo Documentos 1998

[26] C M Almeida S F Lima C V Martins-Da-Silva and JI Gomes ldquoCaracterizacao morfologica e anatomica de dezespecies de leguminosae ocorrentes em uma floresta tropicalumida localizada no municıpio de Moju Estado do Parardquo inA Silvicultura na Amazonia Oriental Contribuicoes do ProjetoEmbrapa-DFID J NMC Silva JO P deCarvalho and J AGYared Eds pp 19ndash54 Embrapa Amazonia Oriental (CPATU)Belem Brazil 2001

[27] K Smalla N Cresswell L C Mendoncahagler A Woltersand J D Vanelsas ldquoRapid extraction protocol from soil forpolymerase chain reaction-mediated amplificationrdquo Journal ofApplied Bacteriology vol 74 pp 78ndash85 1993

[28] S Gantner A F Andersson L Alonso-Saez and S Bertils-son ldquoNovel primers for 16S rRNA-based archaeal communityanalyses in environmental samplesrdquo Journal of MicrobiologicalMethods vol 84 no 1 pp 12ndash18 2011

[29] P D Schloss S L Westcott T Ryabin et al ldquoIntroducingMOTHUR open-source platform-independent community-supported software for describing and comparing microbialcommunitiesrdquoApplied and EnvironmentalMicrobiology vol 75no 23 pp 7537ndash7541 2009

[30] PD SchlossDGevers and S LWestcott ldquoReducing the effectsof PCR amplification and sequencing artifacts on 16S rRNA-based studiesrdquo PLoS ONE vol 6 no 12 Article ID e27310 2011

[31] CQuince A Lanzen T P Curtis et al ldquoAccurate determinationof microbial diversity from 454 pyrosequencing datardquo NatureMethods vol 6 no 9 pp 639ndash641 2009

[32] P Yilmaz L W Parfrey P Yarza et al ldquoThe SILVA and lsquoall-species Living Tree Project (LTP)rsquo taxonomic frameworksrdquoNucleic Acids Research vol 42 no 1 pp D643ndashD648 2014

[33] R C Edgar B J Haas J C Clemente C Quince and RKnight ldquoUCHIME improves sensitivity and speed of chimeradetectionrdquo Bioinformatics vol 27 no 16 pp 2194ndash2200 2011

[34] A Chao and T-J Shen ldquoNonparametric estimation of Shan-nonrsquos index of diversity when there are unseen species insamplerdquo Environmental and Ecological Statistics vol 10 no 4pp 429ndash443 2003

[35] A Chao and T-J Shen Program SPADE (Species Prediction andDiversity Estimation) 2010

[36] C L Shannon andWWeaverTheMathematicalTheory of Com-munication University of Illinois Press Urbana Ill USA 1949

[37] I J Good ldquoThe population frequencies of species and theestimation of population parametersrdquo Biometrika vol 40 pp237ndash264 1953

[38] K Tamura G Stecher D Peterson A Filipski and S KumarldquoMEGA6molecular evolutionary genetics analysis version 60rdquoMolecular Biology and Evolution vol 30 no 12 pp 2725ndash27292013

[39] DA BensonMCavanaugh K Clark et al ldquoGenBankrdquoNucleicAcids Research vol 41 no 1 pp D36ndashD42 2013

[40] D H Parks G W Tyson P Hugenholtz and R G BeikoldquoSTAMP statistical analysis of taxonomic and functional pro-filesrdquo Bioinformatics vol 30 no 21 pp 3123ndash3124 2014

[41] R G Newcombe ldquoInterval estimation for the differencebetween independent proportions comparison of elevenmeth-odsrdquo Statistics in Medicine vol 17 no 8 pp 873ndash890 1998

[42] J D Storey J E Taylor and D Siegmund ldquoStrong controlconservative point estimation and simultaneous conservative

Archaea 13

consistency of false discovery rates a unified approachrdquo Journalof the Royal Statistical Society Series B Statistical Methodologyvol 66 no 1 pp 187ndash205 2004

[43] G C Baker J J Smith and D A Cowan ldquoReview and re-analysis of domain-specific 16S primersrdquo Journal of Microbio-logical Methods vol 55 no 3 pp 541ndash555 2003

[44] W H Zhong and Z C Cai ldquoLong-term effects of inorganicfertilizers on microbial biomass and community functionaldiversity in a paddy soil derived from quaternary red clayrdquoApplied Soil Ecology vol 36 no 2-3 pp 84ndash91 2007

[45] EWessen S Hallin and L Philippot ldquoDifferential responses ofbacterial and archaeal groups at high taxonomical ranks to soilmanagementrdquo Soil Biology and Biochemistry vol 42 no 10 pp1759ndash1765 2010

[46] ZWuW Lin B Li et al ldquoTerminal restriction fragment lengthpolymorphism analysis of soil bacterial communities underdifferent vegetation types in subtropical area rdquo PLOS ONE vol10 no 6 Article ID e0129397 2015

[47] K Cenciani M R Lambais C C Cerri L C B De Azevedoand B J Feigl ldquoBacteria diversity and microbial biomass inforest pasture and fallow soils in the southwestern Amazonbasinrdquo Revista Brasileira de Ciencia do Solo vol 33 no 4 pp907ndash916 2009

[48] A A Navarrete R G Taketani L WMendes F de Souza Can-navan F M de Souza Moreira and S M Tsai ldquoLand-use sys-tems affect Archaeal community structure and functional diver-sity in western Amazon soilsrdquo Revista Brasileira de Ciencia doSolo vol 35 no 5 pp 1527ndash1540 2011

[49] E Lumini A Orgiazzi R Borriello P Bonfante and V Bian-ciotto ldquoDisclosing arbuscular mycorrhizal fungal biodiversityin soil through a land-use gradient using a pyrosequencingapproachrdquo Environmental Microbiology vol 12 no 8 pp 2165ndash2179 2010

[50] M Lupatini R J S Jacques Z I Antoniolli A K A SuleimanR R Fulthorpe and L F W Roesch ldquoLand-use change andsoil type are drivers of fungal and archaeal communities in thePampa biomerdquo World Journal of Microbiology and Biotechnol-ogy vol 29 no 2 pp 223ndash233 2013

[51] E G Lamb N Kennedy and S D Siciliano ldquoEffects of plantspecies richness and evenness on soil microbial communitydiversity and functionrdquo Plant and Soil vol 338 no 1 pp 483ndash495 2011

[52] A Spang R Hatzenpichler C Brochier-Armanet et al ldquoDis-tinct gene set in two different lineages of ammonia-oxidizingarchaea supports the phylum Thaumarchaeotardquo Trends inMicrobiology vol 18 no 8 pp 331ndash340 2010

[53] R G Pacchioni F M Carvalho C E Thompson et al ldquoTax-onomic and functional profiles of soil samples from Atlanticforest and Caatinga biomes in northeastern Brazilrdquo Microbio-logyOpen vol 3 no 3 pp 299ndash315 2014

[54] E Catao A P Castro C C Barreto R H Kruger and C MKyaw ldquoDiversity of Archaea in Brazilian savanna soilsrdquoArchivesof Microbiology vol 195 no 7 pp 507ndash512 2013

[55] M Pester T Rattei S Flechl et al ldquoAmoA-based consensusphylogeny of ammonia-oxidizing archaea and deep sequencingof amoA genes from soils of four different geographic regionsrdquoEnvironmental Microbiology vol 14 no 2 pp 525ndash539 2012

[56] J-Z He H-W Hu and L-M Zhang ldquoCurrent insights into theautotrophic thaumarchaeal ammonia oxidation in acidic soilsrdquoSoil Biology amp Biochemistry vol 55 pp 146ndash154 2012

[57] L-M Zhang H-W Hu J-P Shen and J-Z He ldquoAmmonia-oxidizing archaea have more important role than ammonia-oxidizing bacteria in ammonia oxidation of strongly acidicsoilsrdquo ISME Journal vol 6 no 5 pp 1032ndash1045 2012

[58] S Leininger T Urich M Schloter et al ldquoArchaea predominateamong ammonia-oxidizing prokaryotes in soilsrdquo Nature vol442 no 7104 pp 806ndash809 2006

[59] N C Banning L D Maccarone L M Fisk and D VMurphy ldquoAmmonia-oxidising bacteria not archaea dominatenitrification activity in semi-arid agricultural soilrdquo ScientificReports vol 5 Article ID 11146 2015

[60] M Herrmann A Hadrich and K Kusel ldquoPredominance ofthaumarchaeal ammonia oxidizer abundance and transcrip-tional activity in an acidic fenrdquoEnvironmentalMicrobiology vol14 no 11 pp 3013ndash3025 2012

[61] K Zhalnina P Dorr de Quadros F O Camargo and E WTriplett ldquoDrivers of archaeal ammonia-oxidizing communitiesin soilrdquo Frontiers in Microbiology vol 3 article 210 Article IDArticle 210 2012

[62] J Hong J Cho and Y Hong ldquoEnvironmental variables shapingthe ecological niche of Thaumarchaeota in soil direct andindirect causal effectsrdquo PLoS ONE vol 10 no 8 Article IDe0133763 2015

[63] G Jurgens K Lindstrom and A Saano ldquoNovel group withinthe kingdom Crenarchaeota from boreal forest soilrdquo Appliedand Environmental Microbiology vol 63 no 2 pp 803ndash8051997

[64] M Tourna M Stieglmeier A Spang et al ldquoNitrososphaeraviennensis an ammonia oxidizing archaeon from soilrdquoProceed-ings of the National Academy of Sciences of the United States ofAmerica vol 108 no 20 pp 8420ndash8425 2011

[65] K V Zhalnina R Dias M T Leonard et al ldquoGenome sequenceof Candidatus Nitrososphaera evergladensis from group I1benriched from everglades soil reveals novel genomic features ofthe ammonia-oxidizing archaeardquoPLoSONE vol 9 no 7 ArticleID e101648 2014

[66] I Anderson L E Ulrich B Lupa et al ldquoGenomic char-acterization of methanomicrobiales reveals three classes ofmethanogensrdquo PLoS ONE vol 4 no 6 Article ID e5797 2009

[67] J E Galagan C Nusbaum A Roy et al ldquoThe genome of Macetivorans reveals extensive metabolic and physiological diver-sityrdquo Genome Research vol 12 no 4 pp 532ndash542 2002

[68] C Wrede S Kokoschka A Dreier C Heller J Reitner and MHoppert ldquoDeposition of biogenic iron minerals in a methaneoxidizing microbial matrdquo Archaea vol 2013 Article ID 1029728 pages 2013

[69] G Garrity Bergeyrsquo Manual of Systematic Bacteriology 721 2001[70] S Sakai Y Takaki S Shimamura et al ldquoGenome sequence

of a mesophilic hydrogenotrophic methanogen Methanocellapaludicola the first cultivated representative of the orderMethanocellalesrdquo PLoS ONE vol 6 no 7 Article ID e228982011

[71] E Bapteste C Brochier and Y Boucher ldquoHigher-level clas-sification of the Archaea evolution of methanogenesis andmethanogensrdquo Archaea vol 1 no 5 pp 353ndash363 2005

[72] R G E Murray and E Stackebrandt ldquoTaxonomic note imple-mentation of the provisional status Candidatus for incompletelydescribed procaryotesrdquo International Journal of Systematic Bac-teriology vol 45 no 1 pp 186ndash187 1995

[73] L E Lehtovirta-Morley K Stoecker A Vilcinskas J I Prosserand G W Nicol ldquoCultivation of an obligate acidophilic ammo-nia oxidizer from a nitrifying acid soilrdquo Proceedings of the

14 Archaea

National Academy of Sciences of the United States of Americavol 108 no 38 pp 15892ndash15897 2011

[74] L E Lehtovirta-Morley C Ge J Ross H Yao G W Nicol andJ I Prosser ldquoCharacterisation of terrestrial acidophilic archaealammonia oxidisers and their inhibition and stimulation byorganic compoundsrdquo FEMSMicrobiology Ecology vol 89 no 3pp 542ndash552 2015

[75] M J Frijlink T Abee H J Laanbroek W de Boer and W NKonings ldquoThe bioenergetics of ammonia and hydroxylamineoxidation in Nitrosomonas europaea at acid and alkaline pHrdquoArchives of Microbiology vol 157 no 2 pp 194ndash199 1992

[76] B Wang Y Zheng R Huang et al ldquoActive ammonia oxidizersin an acidic soil are phylogenetically closely related to neu-trophilic archaeonrdquo Applied and Environmental Microbiologyvol 80 no 5 pp 1684ndash1691 2014

[77] J Y Aller and P F Kemp ldquoAre Archaea inherently less diversethan Bacteria in the same environmentsrdquo FEMS MicrobiologyEcology vol 65 no 1 pp 74ndash87 2008

[78] K Potthast U Hamer and FMakeschin ldquoLand-use change in atropical mountain rainforest region of southern Ecuador affectssoilmicroorganisms and nutrient cyclingrdquoBiogeochemistry vol111 no 1ndash3 pp 151ndash167 2012

[79] J I Prosser and G W Nicol ldquoArchaeal and bacterial ammonia-oxidisers in soil the quest for niche specialisation and differen-tiationrdquoTrends inMicrobiology vol 20 no 11 pp 523ndash531 2012

Submit your manuscripts athttpwwwhindawicom

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Anatomy Research International

PeptidesInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporation httpwwwhindawicom

International Journal of

Volume 2014

Zoology

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Molecular Biology International

GenomicsInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioinformaticsAdvances in

Marine BiologyJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Signal TransductionJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioMed Research International

Evolutionary BiologyInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Biochemistry Research International

ArchaeaHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Genetics Research International

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Advances in

Virolog y

Hindawi Publishing Corporationhttpwwwhindawicom

Nucleic AcidsJournal of

Volume 2014

Stem CellsInternational

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Enzyme Research

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

International Journal of

Microbiology

10 Archaea

and denaturing gradient gel electrophoresis analysis of differ-ent Western Amazon soils The databases at the time did notconsiderThaumarchaeota in their classification however ourreanalysis of their dataset using an updated database showedthat Euryarchaeota and Thaumarchaeota were present intheir samples along with Crenarchaeota (Thermoprotei) Inaddition changes were observed in the structure of soilarchaeal communities when comparing the cultivated areaswith forest areas that had been converted to pasture

In our study differences in the percentages of Thau-marchaeota and Euryarchaeota found in the soil sampleswere not striking Thaumarchaeota were predominant in thenative forest (58) whereas Euryarchaeota were predom-inant in the oil palm-cultivated area with 54 (Figure 3)This result is consistent with other comparative studies onarchaeal communities in different types of Brazilian soils (ieAmazonian anthrosols Caatinga and Cerrado) [10 53 54]and a study of different tropical forest soil types that reportedThaumarchaeota as the predominant phylum [44]

The phylumThaumarchaeota comprises all known AOAsand is directly involved in nitrogen metabolism Ammonia isused as an energy source by Group I1a found predominantlyin aquatic systems [52] and Group I1b found in soils [55]Thaumarchaeota Group I1c is primarily found in acidic soilsbut the ecological roles of these archaea are still unknown [5657]

The literature presents conflicting data about the pro-portion of AOAsAOBs soils In some studies AOAs arepredominant [58] while in others AOBs are more abundant[59] However several studies found a predominance of thearchaeal amoA gene over the bacterial amoA gene in soils[60] AOAs are frequently detected in soils their abundancerange from 05 to 10 of the prokaryotic community andtheir abundance is strongly influenced by physicochemicalconditions as well as the organic matter content [61 62] Inacidic soils AOAs appear to have a more important role inammonia oxidation than AOBs [57]

The Soil Crenarchaeotic Group a class within Thaumar-chaeota according to the SILVA database classification wasdetected in both soil samples but a higher representationwas observed in native forest soil This class was discoveredin boreal forest soil [63] Candidatus Nitrososphaera whichbelongs to the Soil Crenarchaeotic Group class is a memberof Thaumarchaeota Group I1b it is an AOA that usesammonia or urea as an energy source [64 65]

On the other hand members of the phylum Eur-yarchaeota are directly involved in carbon metabolismwith several classes of methanogens and methane-oxidizingorganisms [66ndash68] Navarrete et al [48] studied the impactof converting Amazon native forest to pasture and reportedchanges not only to the composition and abundance of soilmicrobial communities but also to a decrease in the diversityof gene function

Within the phylum Euryarchaeota Methanomicrobiaand Thermoplasmata were the main classes detected in oursamples along with a few representatives of Methanobac-teria and Halobacteria Members of Methanobacteria andMethanomicrobia are methanogens [69] Methanomicrobiawere present in both native forest soil and oil palm-cultivated

soil with larger representation in the oil palm-cultivatedarea Within Methanomicrobia the genera Rice Cluster IMethanosarcina andMethanocellawere present in both sam-ples (Figure 6(b)) These genera are closely related and foundin rice fields that produce methane [70] The order Metha-nomicrobiales was detected only in the native forest soilwhich can probably be explained by the relatively high levelsof carbon and organic matter This archaeal order is respon-sible for the final step of methane production in the carboncycle by anaerobic degradation of organic matter [66 71]However the contribution of the Methanomicrobia to thecarbon cycle in the oil palm-cultivated soils could not beassessed since there are no data on the methane productionin the studied areas

Among the Thaumarchaeota sequences used to generatethe phylogenetic tree that were unclassified at the genuslevel several OTUs formed different clusters associated withtheMiscellaneous Crenarchaeotic Group (Figure 5(a))Thesesequences were from both soil samples indicating that theunclassified Thaumarchaeota in the native forest and the oilpalm-cultivated area are phylogenetically related

Most of the sequences remained unclassified at thegenus level for both Thaumarchaeota and Euryarchaeota(Figures 6(a) and 6(b)) This result likely reflects the fact thatmost taxonomic databases do not have many representativesof Archaea especially belonging to the phylum Thaumar-chaeota thus few classifications of new sequences reach thegenus level Several genera were identified as Candidatusaccording to the International Code of Nomenclature ofBacteria indicating that characterization of the culturedprokaryote is incomplete [72]

The proportions of only four groups classified at thegenus level differed significantly between native forest and oilpalm-cultivated soils (Methanosarcina and Rice Cluster I inthe phylum Euryarchaeota and Candidatus NitrososphaeraandCandidatusNitrosotalea in the phylumThaumarchaeota)(Figure 7) With respect to the methanogens sequencesrelated to Methanosarcina were slightly more abundant insoil under oil palm cultivation (Figure 7) whereas sequencesbelonging to the Rice Cluster I were more abundant in nativeforest soil

In the Thaumarchaeota phylum Candidatus Nitrosos-phaera an AOA member of group I1b [65] was detectedin both samples but had a much higher representation innative forest (native forest 182 oil palm-cultivated area103) (Figure 7) In a comparative study of the archaealcommunities in Amazonian dark earth (ldquoTerra Pretardquo)and adjacent soils Candidatus Nitrososphaera was themain archaeal group found in both soils [10] In contrastCandidatus Nitrosotalea another AOA was more abundantin the oil palm-cultivated area (100) than in native forestsoil (44) in our study (Figure 7) Lehtovirta-Morley et al[73] cultivated the obligate acidophilic ammonia oxidizerCandidatus Nitrosotalea devanaterra from a nitrifying acidicsoil Later the same group characterized two strains ofacidophilic AOA belonging to this genus from a Chinesepaddy field and a Scottish agricultural soil [74] In our studythe greater representation of Candidatus Nitrososphaeraand Candidatus Nitrosotalea in native forest and oil

Archaea 11

palm-cultivated soils respectively can be explained by thelow soil pH (native forest pH 550 oil palm-cultivated areapH 483) since in basic soils the activity and growth ofthese archaea are diminished by the exponential decrease innitrite [6 75] In addition soil under oil palm cultivationis fertilized with nitrogen which may explain the higherlevel of Candidatus Nitrosotalea in this area compared withthat of native forest soil Finally although the cultivatedmembers of the genus Ca Nitrososphaera grow betteron neutral pH they have been reported to grow on pHvalues ranging from 60 to 85 In addition a recent workpublished by Wang et al (2014) [76] described active AOAsfrom an acidic soil (pH 492) which are closely related toNitrososphaera known as Nitrososphaera-like organismsby molecular phylogenetic methods Most of our sequenceswere classified as Nitrosotalea-like which are sequencesbelonging to the group I1a This group is typically foundin aquatic environments such as lake sediments or marineenvironments A similar ubiquitous distribution in soils isfound for the group I1b which are not restricted to a specificsoil pH condition

5 Conclusions

Although archaea are known to be ubiquitous there is stillmuch to be learned about their diversity biological rolesand metabolic capabilities This work is the first to comparearchaeal communities associated with the Amazon forest soilwith those of soil under oil palm cultivation

Archaeal diversity was found to be lower in oil palm-cultivated soil than in the native forest soil Only about 30of the OTUs were found in the oil palm soil in comparisonto the native forest However two main phyla were detectedin both samples Thaumarchaeota and Euryarchaeota with apredominance ofThaumarchaeota in native forest soil Theseresults support previous studies in other Brazilian biomenative soils (eg Caatinga Cerrado) as well as soil underdifferent types of land use (eg pasture and agricultural) Itis also important to mention that there are also groups ofuncertain classification associated with the Crenarchaeotaphylum

Although knowledge about Archaea has increased dra-matically in the last few decades little is known aboutthe biology of many archaeal groups and the taxonomyof Archaea is still a work in progress Thus it is notsurprising thatmany sequences identified in thiswork remainunclassified sometimes even at higher taxonomic levels Itis not clear whether the relatively small overall differencebetween soil archaeal communities of the Amazon forest andoil palm-cultivated land is a consequence of the current stateof archaeal taxonomy or whether it indicates that archaealdiversity is lower than bacterial diversity [77]

It is also important to consider that with land usechange due to agricultural practices the microstructure ofthe soil is broken down which can reduce the natural habitatfragmentation allowing potential interactions among theresident microbiota that otherwise would not happen [7879]

A phylogenetic tree of unclassified Thaumarchaeota 16SrRNA gene sequences was useful in exploring relationshipsamong these sequences When taxonomic classification ofsequences was possible it revealed the presence of OTUsfrom genera known to be involved in the metabolism ofcarbon and nitrogen (methanogens Methanocella and Meth-anosarcina AOACandidatusNitrososphaera andCandidatusNitrosotalea) hinting at possible roles for these archaealcommunities in Amazon soils

More studies are needed to elucidate these roles Cultur-ing some of these archaea in the laboratory would allow theirmetabolism and physiology to be studied in detail allowingfor a better understanding of the roles of Archaea in the bio-geochemical cycles

Conflict of Interests

The authors declare that there is no conflict of interestsregarding the publication of this paper

Acknowledgments

This work was supported by grants from FINEP CNPqCAPES and FAP-DF The authors thank Dr Boari for helpwith sample collection logistics Dr Betania Ferraz Quirinowas supported by the Cientista Visitante program fromEmbrapa

References

[1] FAO Global Forest Resources Assessment Food and AgricultureOrganization of the United Nations 2010

[2] IBGE ldquoMapa de Biomas do Brasil primeira aproximacaordquo Riode Janeiro Brazil 2004 httpwwwibgegovbrhome

[3] C Azevedo-Ramos ldquoSustainable development and challengingdeforestation in the Brazilian Amazon the good the badand the uglyrdquo in Proceedings of the Symposium Our CommonGround Innovations in Land Use Decision-Making vol 59 p 5Vancouver Canada May 2007

[4] C A Francis K J Roberts J M Beman A E Santoro and B BOakley ldquoUbiquity and diversity of ammonia-oxidizing archaeainwater columns and sediments of the oceanrdquoProceedings of theNational Academy of Sciences of theUnited States of America vol102 no 41 pp 14683ndash14688 2005

[5] A H Treusch S Leininger A Kietzin S C Schuster H-PKlenk and C Schleper ldquoNovel genes for nitrite reductase andAmo-related proteins indicate a role of uncultivatedmesophiliccrenarchaeota in nitrogen cyclingrdquo Environmental Microbiol-ogy vol 7 no 12 pp 1985ndash1995 2005

[6] G W Nicol S Leininger C Schleper and J I Prosser ldquoTheinfluence of soil pH on the diversity abundance and tran-scriptional activity of ammonia oxidizing archaea and bacteriardquoEnvironmentalMicrobiology vol 10 no 11 pp 2966ndash2978 2008

[7] P Offre A Spang and C Schleper ldquoArchaea in biogeochemicalcyclesrdquo Annual Review of Microbiology vol 67 pp 437ndash4572013

[8] MAPA Anuario Estatıstico da Agroenergia 2012 StatisticalYearbook of Agroenergy 2013

[9] J C Bergmann D D Tupinamba O Y A Costa J R MAlmeida C C Barreto andB FQuirino ldquoBiodiesel production

12 Archaea

in Brazil and alternative biomass feedstocksrdquo Renewable ampSustainable Energy Reviews vol 21 pp 411ndash420 2013

[10] R G Taketani and S M Tsai ldquoThe influence of different landuses on the structure of archaeal communities in Amazoniananthrosols based on 16S rRNA and amoA genesrdquo MicrobialEcology vol 59 no 4 pp 734ndash743 2010

[11] C Brochier-Armanet B Boussau S Gribaldo and P ForterreldquoMesophilic crenarchaeota proposal for a third archaeal phy-lum theThaumarchaeotardquoNature Reviews Microbiology vol 6no 3 pp 245ndash252 2008

[12] S M Barns C F Delwiche J D Palmer and N R PaceldquoPerspectives on archaeal diversity thermophily and mono-phyly from environmental rRNA sequencesrdquo Proceedings of theNational Academy of Sciences of the United States of Americavol 93 no 17 pp 9188ndash9193 1996

[13] J G Elkins M Podar D E Graham et al ldquoA korarchaealgenome reveals insights into the evolution of the Archaeardquo Pro-ceedings of the National Academy of Sciences of the United Statesof America vol 105 no 23 pp 8102ndash8107 2008

[14] H Huber M J Hohn R Rachel T Fuchs V C Wimmer andK O Stetter ldquoA new phylum of Archaea represented by ananosized hyperthermophilic symbiontrdquo Nature vol 417 no6884 pp 63ndash67 2002

[15] T Nunoura H Oida M Nakaseama et al ldquoArchaeal diversityand distribution along thermal and geochemical gradients inhydrothermal sediments at the Yonaguni Knoll IV hydrother-mal field in the SouthernOkinawa TroughrdquoApplied and Enviro-nmental Microbiology vol 76 no 4 pp 1198ndash1211 2010

[16] C Rinke P Schwientek A Sczyrba et al ldquoInsights into thephylogeny and coding potential of microbial dark matterrdquoNature vol 499 no 7459 pp 431ndash437 2013

[17] A Spang J H Saw S L Joslashrgensen et al ldquoComplex archaea thatbridge the gap between prokaryotes and eukaryotesrdquo Naturevol 521 no 7551 pp 173ndash179 2015

[18] S Bagchi S E Vlaeminck L A Sauder et al ldquoTemporal andspatial stability of ammonia-oxidizing archaea and bacteria inaquariumbiofiltersrdquo PLoSONE vol 9 no 12 Article ID e1135152014

[19] W Qin S A Amin W Martens-Habbena et al ldquoMarineammonia-oxidizing archaeal isolates display obligate mixotro-phy and wide ecotypic variationrdquo Proceedings of the NationalAcademy of Sciences of the United States of America vol 111 no34 pp 12504ndash12509 2014

[20] M Coci N Odermatt M M Salcher J Pernthaler and GCorno ldquoEcology and distribution ofThaumarchaea in the deephypolimnion of Lake Maggiorerdquo Archaea vol 2015 Article ID590434 11 pages 2015

[21] J G Ferry ldquoHow tomake a living by exhalingmethanerdquoAnnualReview of Microbiology vol 64 pp 453ndash473 2010

[22] A Meyer A Focks V Radl G Welzl I Schoning and MSchloter ldquoInfluence of land use intensity on the diversity ofammonia oxidizing bacteria and archaea in soils from grasslandecosystemsrdquoMicrobial Ecology vol 67 no 1 pp 161ndash166 2014

[23] V De Gannes G Eudoxie and W J Hickey ldquoImpacts ofedaphic factors on communities of ammonia-oxidizing archaeaammonia-oxidizing bacteria and nitrification in tropical soilsrdquoPLoS ONE vol 9 no 2 Article ID e89568 2014

[24] D S Ribeiro F C J da Silva and TN Carvalho ldquoSobrevivenciade seis especies florestais em uma area explorada seletivamenteno municıpio de Moju Parardquo CERNE vol 9 no 2 pp 153ndash1632003

[25] D HM Costa C A P Ferreira J NM Silva J D C A Lopesand J O P D Carvalho ldquoPotencial madeireiro de floresta densano municıpio de Moju Estado do Parardquo Documentos 1998

[26] C M Almeida S F Lima C V Martins-Da-Silva and JI Gomes ldquoCaracterizacao morfologica e anatomica de dezespecies de leguminosae ocorrentes em uma floresta tropicalumida localizada no municıpio de Moju Estado do Parardquo inA Silvicultura na Amazonia Oriental Contribuicoes do ProjetoEmbrapa-DFID J NMC Silva JO P deCarvalho and J AGYared Eds pp 19ndash54 Embrapa Amazonia Oriental (CPATU)Belem Brazil 2001

[27] K Smalla N Cresswell L C Mendoncahagler A Woltersand J D Vanelsas ldquoRapid extraction protocol from soil forpolymerase chain reaction-mediated amplificationrdquo Journal ofApplied Bacteriology vol 74 pp 78ndash85 1993

[28] S Gantner A F Andersson L Alonso-Saez and S Bertils-son ldquoNovel primers for 16S rRNA-based archaeal communityanalyses in environmental samplesrdquo Journal of MicrobiologicalMethods vol 84 no 1 pp 12ndash18 2011

[29] P D Schloss S L Westcott T Ryabin et al ldquoIntroducingMOTHUR open-source platform-independent community-supported software for describing and comparing microbialcommunitiesrdquoApplied and EnvironmentalMicrobiology vol 75no 23 pp 7537ndash7541 2009

[30] PD SchlossDGevers and S LWestcott ldquoReducing the effectsof PCR amplification and sequencing artifacts on 16S rRNA-based studiesrdquo PLoS ONE vol 6 no 12 Article ID e27310 2011

[31] CQuince A Lanzen T P Curtis et al ldquoAccurate determinationof microbial diversity from 454 pyrosequencing datardquo NatureMethods vol 6 no 9 pp 639ndash641 2009

[32] P Yilmaz L W Parfrey P Yarza et al ldquoThe SILVA and lsquoall-species Living Tree Project (LTP)rsquo taxonomic frameworksrdquoNucleic Acids Research vol 42 no 1 pp D643ndashD648 2014

[33] R C Edgar B J Haas J C Clemente C Quince and RKnight ldquoUCHIME improves sensitivity and speed of chimeradetectionrdquo Bioinformatics vol 27 no 16 pp 2194ndash2200 2011

[34] A Chao and T-J Shen ldquoNonparametric estimation of Shan-nonrsquos index of diversity when there are unseen species insamplerdquo Environmental and Ecological Statistics vol 10 no 4pp 429ndash443 2003

[35] A Chao and T-J Shen Program SPADE (Species Prediction andDiversity Estimation) 2010

[36] C L Shannon andWWeaverTheMathematicalTheory of Com-munication University of Illinois Press Urbana Ill USA 1949

[37] I J Good ldquoThe population frequencies of species and theestimation of population parametersrdquo Biometrika vol 40 pp237ndash264 1953

[38] K Tamura G Stecher D Peterson A Filipski and S KumarldquoMEGA6molecular evolutionary genetics analysis version 60rdquoMolecular Biology and Evolution vol 30 no 12 pp 2725ndash27292013

[39] DA BensonMCavanaugh K Clark et al ldquoGenBankrdquoNucleicAcids Research vol 41 no 1 pp D36ndashD42 2013

[40] D H Parks G W Tyson P Hugenholtz and R G BeikoldquoSTAMP statistical analysis of taxonomic and functional pro-filesrdquo Bioinformatics vol 30 no 21 pp 3123ndash3124 2014

[41] R G Newcombe ldquoInterval estimation for the differencebetween independent proportions comparison of elevenmeth-odsrdquo Statistics in Medicine vol 17 no 8 pp 873ndash890 1998

[42] J D Storey J E Taylor and D Siegmund ldquoStrong controlconservative point estimation and simultaneous conservative

Archaea 13

consistency of false discovery rates a unified approachrdquo Journalof the Royal Statistical Society Series B Statistical Methodologyvol 66 no 1 pp 187ndash205 2004

[43] G C Baker J J Smith and D A Cowan ldquoReview and re-analysis of domain-specific 16S primersrdquo Journal of Microbio-logical Methods vol 55 no 3 pp 541ndash555 2003

[44] W H Zhong and Z C Cai ldquoLong-term effects of inorganicfertilizers on microbial biomass and community functionaldiversity in a paddy soil derived from quaternary red clayrdquoApplied Soil Ecology vol 36 no 2-3 pp 84ndash91 2007

[45] EWessen S Hallin and L Philippot ldquoDifferential responses ofbacterial and archaeal groups at high taxonomical ranks to soilmanagementrdquo Soil Biology and Biochemistry vol 42 no 10 pp1759ndash1765 2010

[46] ZWuW Lin B Li et al ldquoTerminal restriction fragment lengthpolymorphism analysis of soil bacterial communities underdifferent vegetation types in subtropical area rdquo PLOS ONE vol10 no 6 Article ID e0129397 2015

[47] K Cenciani M R Lambais C C Cerri L C B De Azevedoand B J Feigl ldquoBacteria diversity and microbial biomass inforest pasture and fallow soils in the southwestern Amazonbasinrdquo Revista Brasileira de Ciencia do Solo vol 33 no 4 pp907ndash916 2009

[48] A A Navarrete R G Taketani L WMendes F de Souza Can-navan F M de Souza Moreira and S M Tsai ldquoLand-use sys-tems affect Archaeal community structure and functional diver-sity in western Amazon soilsrdquo Revista Brasileira de Ciencia doSolo vol 35 no 5 pp 1527ndash1540 2011

[49] E Lumini A Orgiazzi R Borriello P Bonfante and V Bian-ciotto ldquoDisclosing arbuscular mycorrhizal fungal biodiversityin soil through a land-use gradient using a pyrosequencingapproachrdquo Environmental Microbiology vol 12 no 8 pp 2165ndash2179 2010

[50] M Lupatini R J S Jacques Z I Antoniolli A K A SuleimanR R Fulthorpe and L F W Roesch ldquoLand-use change andsoil type are drivers of fungal and archaeal communities in thePampa biomerdquo World Journal of Microbiology and Biotechnol-ogy vol 29 no 2 pp 223ndash233 2013

[51] E G Lamb N Kennedy and S D Siciliano ldquoEffects of plantspecies richness and evenness on soil microbial communitydiversity and functionrdquo Plant and Soil vol 338 no 1 pp 483ndash495 2011

[52] A Spang R Hatzenpichler C Brochier-Armanet et al ldquoDis-tinct gene set in two different lineages of ammonia-oxidizingarchaea supports the phylum Thaumarchaeotardquo Trends inMicrobiology vol 18 no 8 pp 331ndash340 2010

[53] R G Pacchioni F M Carvalho C E Thompson et al ldquoTax-onomic and functional profiles of soil samples from Atlanticforest and Caatinga biomes in northeastern Brazilrdquo Microbio-logyOpen vol 3 no 3 pp 299ndash315 2014

[54] E Catao A P Castro C C Barreto R H Kruger and C MKyaw ldquoDiversity of Archaea in Brazilian savanna soilsrdquoArchivesof Microbiology vol 195 no 7 pp 507ndash512 2013

[55] M Pester T Rattei S Flechl et al ldquoAmoA-based consensusphylogeny of ammonia-oxidizing archaea and deep sequencingof amoA genes from soils of four different geographic regionsrdquoEnvironmental Microbiology vol 14 no 2 pp 525ndash539 2012

[56] J-Z He H-W Hu and L-M Zhang ldquoCurrent insights into theautotrophic thaumarchaeal ammonia oxidation in acidic soilsrdquoSoil Biology amp Biochemistry vol 55 pp 146ndash154 2012

[57] L-M Zhang H-W Hu J-P Shen and J-Z He ldquoAmmonia-oxidizing archaea have more important role than ammonia-oxidizing bacteria in ammonia oxidation of strongly acidicsoilsrdquo ISME Journal vol 6 no 5 pp 1032ndash1045 2012

[58] S Leininger T Urich M Schloter et al ldquoArchaea predominateamong ammonia-oxidizing prokaryotes in soilsrdquo Nature vol442 no 7104 pp 806ndash809 2006

[59] N C Banning L D Maccarone L M Fisk and D VMurphy ldquoAmmonia-oxidising bacteria not archaea dominatenitrification activity in semi-arid agricultural soilrdquo ScientificReports vol 5 Article ID 11146 2015

[60] M Herrmann A Hadrich and K Kusel ldquoPredominance ofthaumarchaeal ammonia oxidizer abundance and transcrip-tional activity in an acidic fenrdquoEnvironmentalMicrobiology vol14 no 11 pp 3013ndash3025 2012

[61] K Zhalnina P Dorr de Quadros F O Camargo and E WTriplett ldquoDrivers of archaeal ammonia-oxidizing communitiesin soilrdquo Frontiers in Microbiology vol 3 article 210 Article IDArticle 210 2012

[62] J Hong J Cho and Y Hong ldquoEnvironmental variables shapingthe ecological niche of Thaumarchaeota in soil direct andindirect causal effectsrdquo PLoS ONE vol 10 no 8 Article IDe0133763 2015

[63] G Jurgens K Lindstrom and A Saano ldquoNovel group withinthe kingdom Crenarchaeota from boreal forest soilrdquo Appliedand Environmental Microbiology vol 63 no 2 pp 803ndash8051997

[64] M Tourna M Stieglmeier A Spang et al ldquoNitrososphaeraviennensis an ammonia oxidizing archaeon from soilrdquoProceed-ings of the National Academy of Sciences of the United States ofAmerica vol 108 no 20 pp 8420ndash8425 2011

[65] K V Zhalnina R Dias M T Leonard et al ldquoGenome sequenceof Candidatus Nitrososphaera evergladensis from group I1benriched from everglades soil reveals novel genomic features ofthe ammonia-oxidizing archaeardquoPLoSONE vol 9 no 7 ArticleID e101648 2014

[66] I Anderson L E Ulrich B Lupa et al ldquoGenomic char-acterization of methanomicrobiales reveals three classes ofmethanogensrdquo PLoS ONE vol 4 no 6 Article ID e5797 2009

[67] J E Galagan C Nusbaum A Roy et al ldquoThe genome of Macetivorans reveals extensive metabolic and physiological diver-sityrdquo Genome Research vol 12 no 4 pp 532ndash542 2002

[68] C Wrede S Kokoschka A Dreier C Heller J Reitner and MHoppert ldquoDeposition of biogenic iron minerals in a methaneoxidizing microbial matrdquo Archaea vol 2013 Article ID 1029728 pages 2013

[69] G Garrity Bergeyrsquo Manual of Systematic Bacteriology 721 2001[70] S Sakai Y Takaki S Shimamura et al ldquoGenome sequence

of a mesophilic hydrogenotrophic methanogen Methanocellapaludicola the first cultivated representative of the orderMethanocellalesrdquo PLoS ONE vol 6 no 7 Article ID e228982011

[71] E Bapteste C Brochier and Y Boucher ldquoHigher-level clas-sification of the Archaea evolution of methanogenesis andmethanogensrdquo Archaea vol 1 no 5 pp 353ndash363 2005

[72] R G E Murray and E Stackebrandt ldquoTaxonomic note imple-mentation of the provisional status Candidatus for incompletelydescribed procaryotesrdquo International Journal of Systematic Bac-teriology vol 45 no 1 pp 186ndash187 1995

[73] L E Lehtovirta-Morley K Stoecker A Vilcinskas J I Prosserand G W Nicol ldquoCultivation of an obligate acidophilic ammo-nia oxidizer from a nitrifying acid soilrdquo Proceedings of the

14 Archaea

National Academy of Sciences of the United States of Americavol 108 no 38 pp 15892ndash15897 2011

[74] L E Lehtovirta-Morley C Ge J Ross H Yao G W Nicol andJ I Prosser ldquoCharacterisation of terrestrial acidophilic archaealammonia oxidisers and their inhibition and stimulation byorganic compoundsrdquo FEMSMicrobiology Ecology vol 89 no 3pp 542ndash552 2015

[75] M J Frijlink T Abee H J Laanbroek W de Boer and W NKonings ldquoThe bioenergetics of ammonia and hydroxylamineoxidation in Nitrosomonas europaea at acid and alkaline pHrdquoArchives of Microbiology vol 157 no 2 pp 194ndash199 1992

[76] B Wang Y Zheng R Huang et al ldquoActive ammonia oxidizersin an acidic soil are phylogenetically closely related to neu-trophilic archaeonrdquo Applied and Environmental Microbiologyvol 80 no 5 pp 1684ndash1691 2014

[77] J Y Aller and P F Kemp ldquoAre Archaea inherently less diversethan Bacteria in the same environmentsrdquo FEMS MicrobiologyEcology vol 65 no 1 pp 74ndash87 2008

[78] K Potthast U Hamer and FMakeschin ldquoLand-use change in atropical mountain rainforest region of southern Ecuador affectssoilmicroorganisms and nutrient cyclingrdquoBiogeochemistry vol111 no 1ndash3 pp 151ndash167 2012

[79] J I Prosser and G W Nicol ldquoArchaeal and bacterial ammonia-oxidisers in soil the quest for niche specialisation and differen-tiationrdquoTrends inMicrobiology vol 20 no 11 pp 523ndash531 2012

Submit your manuscripts athttpwwwhindawicom

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Anatomy Research International

PeptidesInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporation httpwwwhindawicom

International Journal of

Volume 2014

Zoology

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Molecular Biology International

GenomicsInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioinformaticsAdvances in

Marine BiologyJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Signal TransductionJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioMed Research International

Evolutionary BiologyInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Biochemistry Research International

ArchaeaHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Genetics Research International

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Advances in

Virolog y

Hindawi Publishing Corporationhttpwwwhindawicom

Nucleic AcidsJournal of

Volume 2014

Stem CellsInternational

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Enzyme Research

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

International Journal of

Microbiology

Archaea 11

palm-cultivated soils respectively can be explained by thelow soil pH (native forest pH 550 oil palm-cultivated areapH 483) since in basic soils the activity and growth ofthese archaea are diminished by the exponential decrease innitrite [6 75] In addition soil under oil palm cultivationis fertilized with nitrogen which may explain the higherlevel of Candidatus Nitrosotalea in this area compared withthat of native forest soil Finally although the cultivatedmembers of the genus Ca Nitrososphaera grow betteron neutral pH they have been reported to grow on pHvalues ranging from 60 to 85 In addition a recent workpublished by Wang et al (2014) [76] described active AOAsfrom an acidic soil (pH 492) which are closely related toNitrososphaera known as Nitrososphaera-like organismsby molecular phylogenetic methods Most of our sequenceswere classified as Nitrosotalea-like which are sequencesbelonging to the group I1a This group is typically foundin aquatic environments such as lake sediments or marineenvironments A similar ubiquitous distribution in soils isfound for the group I1b which are not restricted to a specificsoil pH condition

5 Conclusions

Although archaea are known to be ubiquitous there is stillmuch to be learned about their diversity biological rolesand metabolic capabilities This work is the first to comparearchaeal communities associated with the Amazon forest soilwith those of soil under oil palm cultivation

Archaeal diversity was found to be lower in oil palm-cultivated soil than in the native forest soil Only about 30of the OTUs were found in the oil palm soil in comparisonto the native forest However two main phyla were detectedin both samples Thaumarchaeota and Euryarchaeota with apredominance ofThaumarchaeota in native forest soil Theseresults support previous studies in other Brazilian biomenative soils (eg Caatinga Cerrado) as well as soil underdifferent types of land use (eg pasture and agricultural) Itis also important to mention that there are also groups ofuncertain classification associated with the Crenarchaeotaphylum

Although knowledge about Archaea has increased dra-matically in the last few decades little is known aboutthe biology of many archaeal groups and the taxonomyof Archaea is still a work in progress Thus it is notsurprising thatmany sequences identified in thiswork remainunclassified sometimes even at higher taxonomic levels Itis not clear whether the relatively small overall differencebetween soil archaeal communities of the Amazon forest andoil palm-cultivated land is a consequence of the current stateof archaeal taxonomy or whether it indicates that archaealdiversity is lower than bacterial diversity [77]

It is also important to consider that with land usechange due to agricultural practices the microstructure ofthe soil is broken down which can reduce the natural habitatfragmentation allowing potential interactions among theresident microbiota that otherwise would not happen [7879]

A phylogenetic tree of unclassified Thaumarchaeota 16SrRNA gene sequences was useful in exploring relationshipsamong these sequences When taxonomic classification ofsequences was possible it revealed the presence of OTUsfrom genera known to be involved in the metabolism ofcarbon and nitrogen (methanogens Methanocella and Meth-anosarcina AOACandidatusNitrososphaera andCandidatusNitrosotalea) hinting at possible roles for these archaealcommunities in Amazon soils

More studies are needed to elucidate these roles Cultur-ing some of these archaea in the laboratory would allow theirmetabolism and physiology to be studied in detail allowingfor a better understanding of the roles of Archaea in the bio-geochemical cycles

Conflict of Interests

The authors declare that there is no conflict of interestsregarding the publication of this paper

Acknowledgments

This work was supported by grants from FINEP CNPqCAPES and FAP-DF The authors thank Dr Boari for helpwith sample collection logistics Dr Betania Ferraz Quirinowas supported by the Cientista Visitante program fromEmbrapa

References

[1] FAO Global Forest Resources Assessment Food and AgricultureOrganization of the United Nations 2010

[2] IBGE ldquoMapa de Biomas do Brasil primeira aproximacaordquo Riode Janeiro Brazil 2004 httpwwwibgegovbrhome

[3] C Azevedo-Ramos ldquoSustainable development and challengingdeforestation in the Brazilian Amazon the good the badand the uglyrdquo in Proceedings of the Symposium Our CommonGround Innovations in Land Use Decision-Making vol 59 p 5Vancouver Canada May 2007

[4] C A Francis K J Roberts J M Beman A E Santoro and B BOakley ldquoUbiquity and diversity of ammonia-oxidizing archaeainwater columns and sediments of the oceanrdquoProceedings of theNational Academy of Sciences of theUnited States of America vol102 no 41 pp 14683ndash14688 2005

[5] A H Treusch S Leininger A Kietzin S C Schuster H-PKlenk and C Schleper ldquoNovel genes for nitrite reductase andAmo-related proteins indicate a role of uncultivatedmesophiliccrenarchaeota in nitrogen cyclingrdquo Environmental Microbiol-ogy vol 7 no 12 pp 1985ndash1995 2005

[6] G W Nicol S Leininger C Schleper and J I Prosser ldquoTheinfluence of soil pH on the diversity abundance and tran-scriptional activity of ammonia oxidizing archaea and bacteriardquoEnvironmentalMicrobiology vol 10 no 11 pp 2966ndash2978 2008

[7] P Offre A Spang and C Schleper ldquoArchaea in biogeochemicalcyclesrdquo Annual Review of Microbiology vol 67 pp 437ndash4572013

[8] MAPA Anuario Estatıstico da Agroenergia 2012 StatisticalYearbook of Agroenergy 2013

[9] J C Bergmann D D Tupinamba O Y A Costa J R MAlmeida C C Barreto andB FQuirino ldquoBiodiesel production

12 Archaea

in Brazil and alternative biomass feedstocksrdquo Renewable ampSustainable Energy Reviews vol 21 pp 411ndash420 2013

[10] R G Taketani and S M Tsai ldquoThe influence of different landuses on the structure of archaeal communities in Amazoniananthrosols based on 16S rRNA and amoA genesrdquo MicrobialEcology vol 59 no 4 pp 734ndash743 2010

[11] C Brochier-Armanet B Boussau S Gribaldo and P ForterreldquoMesophilic crenarchaeota proposal for a third archaeal phy-lum theThaumarchaeotardquoNature Reviews Microbiology vol 6no 3 pp 245ndash252 2008

[12] S M Barns C F Delwiche J D Palmer and N R PaceldquoPerspectives on archaeal diversity thermophily and mono-phyly from environmental rRNA sequencesrdquo Proceedings of theNational Academy of Sciences of the United States of Americavol 93 no 17 pp 9188ndash9193 1996

[13] J G Elkins M Podar D E Graham et al ldquoA korarchaealgenome reveals insights into the evolution of the Archaeardquo Pro-ceedings of the National Academy of Sciences of the United Statesof America vol 105 no 23 pp 8102ndash8107 2008

[14] H Huber M J Hohn R Rachel T Fuchs V C Wimmer andK O Stetter ldquoA new phylum of Archaea represented by ananosized hyperthermophilic symbiontrdquo Nature vol 417 no6884 pp 63ndash67 2002

[15] T Nunoura H Oida M Nakaseama et al ldquoArchaeal diversityand distribution along thermal and geochemical gradients inhydrothermal sediments at the Yonaguni Knoll IV hydrother-mal field in the SouthernOkinawa TroughrdquoApplied and Enviro-nmental Microbiology vol 76 no 4 pp 1198ndash1211 2010

[16] C Rinke P Schwientek A Sczyrba et al ldquoInsights into thephylogeny and coding potential of microbial dark matterrdquoNature vol 499 no 7459 pp 431ndash437 2013

[17] A Spang J H Saw S L Joslashrgensen et al ldquoComplex archaea thatbridge the gap between prokaryotes and eukaryotesrdquo Naturevol 521 no 7551 pp 173ndash179 2015

[18] S Bagchi S E Vlaeminck L A Sauder et al ldquoTemporal andspatial stability of ammonia-oxidizing archaea and bacteria inaquariumbiofiltersrdquo PLoSONE vol 9 no 12 Article ID e1135152014

[19] W Qin S A Amin W Martens-Habbena et al ldquoMarineammonia-oxidizing archaeal isolates display obligate mixotro-phy and wide ecotypic variationrdquo Proceedings of the NationalAcademy of Sciences of the United States of America vol 111 no34 pp 12504ndash12509 2014

[20] M Coci N Odermatt M M Salcher J Pernthaler and GCorno ldquoEcology and distribution ofThaumarchaea in the deephypolimnion of Lake Maggiorerdquo Archaea vol 2015 Article ID590434 11 pages 2015

[21] J G Ferry ldquoHow tomake a living by exhalingmethanerdquoAnnualReview of Microbiology vol 64 pp 453ndash473 2010

[22] A Meyer A Focks V Radl G Welzl I Schoning and MSchloter ldquoInfluence of land use intensity on the diversity ofammonia oxidizing bacteria and archaea in soils from grasslandecosystemsrdquoMicrobial Ecology vol 67 no 1 pp 161ndash166 2014

[23] V De Gannes G Eudoxie and W J Hickey ldquoImpacts ofedaphic factors on communities of ammonia-oxidizing archaeaammonia-oxidizing bacteria and nitrification in tropical soilsrdquoPLoS ONE vol 9 no 2 Article ID e89568 2014

[24] D S Ribeiro F C J da Silva and TN Carvalho ldquoSobrevivenciade seis especies florestais em uma area explorada seletivamenteno municıpio de Moju Parardquo CERNE vol 9 no 2 pp 153ndash1632003

[25] D HM Costa C A P Ferreira J NM Silva J D C A Lopesand J O P D Carvalho ldquoPotencial madeireiro de floresta densano municıpio de Moju Estado do Parardquo Documentos 1998

[26] C M Almeida S F Lima C V Martins-Da-Silva and JI Gomes ldquoCaracterizacao morfologica e anatomica de dezespecies de leguminosae ocorrentes em uma floresta tropicalumida localizada no municıpio de Moju Estado do Parardquo inA Silvicultura na Amazonia Oriental Contribuicoes do ProjetoEmbrapa-DFID J NMC Silva JO P deCarvalho and J AGYared Eds pp 19ndash54 Embrapa Amazonia Oriental (CPATU)Belem Brazil 2001

[27] K Smalla N Cresswell L C Mendoncahagler A Woltersand J D Vanelsas ldquoRapid extraction protocol from soil forpolymerase chain reaction-mediated amplificationrdquo Journal ofApplied Bacteriology vol 74 pp 78ndash85 1993

[28] S Gantner A F Andersson L Alonso-Saez and S Bertils-son ldquoNovel primers for 16S rRNA-based archaeal communityanalyses in environmental samplesrdquo Journal of MicrobiologicalMethods vol 84 no 1 pp 12ndash18 2011

[29] P D Schloss S L Westcott T Ryabin et al ldquoIntroducingMOTHUR open-source platform-independent community-supported software for describing and comparing microbialcommunitiesrdquoApplied and EnvironmentalMicrobiology vol 75no 23 pp 7537ndash7541 2009

[30] PD SchlossDGevers and S LWestcott ldquoReducing the effectsof PCR amplification and sequencing artifacts on 16S rRNA-based studiesrdquo PLoS ONE vol 6 no 12 Article ID e27310 2011

[31] CQuince A Lanzen T P Curtis et al ldquoAccurate determinationof microbial diversity from 454 pyrosequencing datardquo NatureMethods vol 6 no 9 pp 639ndash641 2009

[32] P Yilmaz L W Parfrey P Yarza et al ldquoThe SILVA and lsquoall-species Living Tree Project (LTP)rsquo taxonomic frameworksrdquoNucleic Acids Research vol 42 no 1 pp D643ndashD648 2014

[33] R C Edgar B J Haas J C Clemente C Quince and RKnight ldquoUCHIME improves sensitivity and speed of chimeradetectionrdquo Bioinformatics vol 27 no 16 pp 2194ndash2200 2011

[34] A Chao and T-J Shen ldquoNonparametric estimation of Shan-nonrsquos index of diversity when there are unseen species insamplerdquo Environmental and Ecological Statistics vol 10 no 4pp 429ndash443 2003

[35] A Chao and T-J Shen Program SPADE (Species Prediction andDiversity Estimation) 2010

[36] C L Shannon andWWeaverTheMathematicalTheory of Com-munication University of Illinois Press Urbana Ill USA 1949

[37] I J Good ldquoThe population frequencies of species and theestimation of population parametersrdquo Biometrika vol 40 pp237ndash264 1953

[38] K Tamura G Stecher D Peterson A Filipski and S KumarldquoMEGA6molecular evolutionary genetics analysis version 60rdquoMolecular Biology and Evolution vol 30 no 12 pp 2725ndash27292013

[39] DA BensonMCavanaugh K Clark et al ldquoGenBankrdquoNucleicAcids Research vol 41 no 1 pp D36ndashD42 2013

[40] D H Parks G W Tyson P Hugenholtz and R G BeikoldquoSTAMP statistical analysis of taxonomic and functional pro-filesrdquo Bioinformatics vol 30 no 21 pp 3123ndash3124 2014

[41] R G Newcombe ldquoInterval estimation for the differencebetween independent proportions comparison of elevenmeth-odsrdquo Statistics in Medicine vol 17 no 8 pp 873ndash890 1998

[42] J D Storey J E Taylor and D Siegmund ldquoStrong controlconservative point estimation and simultaneous conservative

Archaea 13

consistency of false discovery rates a unified approachrdquo Journalof the Royal Statistical Society Series B Statistical Methodologyvol 66 no 1 pp 187ndash205 2004

[43] G C Baker J J Smith and D A Cowan ldquoReview and re-analysis of domain-specific 16S primersrdquo Journal of Microbio-logical Methods vol 55 no 3 pp 541ndash555 2003

[44] W H Zhong and Z C Cai ldquoLong-term effects of inorganicfertilizers on microbial biomass and community functionaldiversity in a paddy soil derived from quaternary red clayrdquoApplied Soil Ecology vol 36 no 2-3 pp 84ndash91 2007

[45] EWessen S Hallin and L Philippot ldquoDifferential responses ofbacterial and archaeal groups at high taxonomical ranks to soilmanagementrdquo Soil Biology and Biochemistry vol 42 no 10 pp1759ndash1765 2010

[46] ZWuW Lin B Li et al ldquoTerminal restriction fragment lengthpolymorphism analysis of soil bacterial communities underdifferent vegetation types in subtropical area rdquo PLOS ONE vol10 no 6 Article ID e0129397 2015

[47] K Cenciani M R Lambais C C Cerri L C B De Azevedoand B J Feigl ldquoBacteria diversity and microbial biomass inforest pasture and fallow soils in the southwestern Amazonbasinrdquo Revista Brasileira de Ciencia do Solo vol 33 no 4 pp907ndash916 2009

[48] A A Navarrete R G Taketani L WMendes F de Souza Can-navan F M de Souza Moreira and S M Tsai ldquoLand-use sys-tems affect Archaeal community structure and functional diver-sity in western Amazon soilsrdquo Revista Brasileira de Ciencia doSolo vol 35 no 5 pp 1527ndash1540 2011

[49] E Lumini A Orgiazzi R Borriello P Bonfante and V Bian-ciotto ldquoDisclosing arbuscular mycorrhizal fungal biodiversityin soil through a land-use gradient using a pyrosequencingapproachrdquo Environmental Microbiology vol 12 no 8 pp 2165ndash2179 2010

[50] M Lupatini R J S Jacques Z I Antoniolli A K A SuleimanR R Fulthorpe and L F W Roesch ldquoLand-use change andsoil type are drivers of fungal and archaeal communities in thePampa biomerdquo World Journal of Microbiology and Biotechnol-ogy vol 29 no 2 pp 223ndash233 2013

[51] E G Lamb N Kennedy and S D Siciliano ldquoEffects of plantspecies richness and evenness on soil microbial communitydiversity and functionrdquo Plant and Soil vol 338 no 1 pp 483ndash495 2011

[52] A Spang R Hatzenpichler C Brochier-Armanet et al ldquoDis-tinct gene set in two different lineages of ammonia-oxidizingarchaea supports the phylum Thaumarchaeotardquo Trends inMicrobiology vol 18 no 8 pp 331ndash340 2010

[53] R G Pacchioni F M Carvalho C E Thompson et al ldquoTax-onomic and functional profiles of soil samples from Atlanticforest and Caatinga biomes in northeastern Brazilrdquo Microbio-logyOpen vol 3 no 3 pp 299ndash315 2014

[54] E Catao A P Castro C C Barreto R H Kruger and C MKyaw ldquoDiversity of Archaea in Brazilian savanna soilsrdquoArchivesof Microbiology vol 195 no 7 pp 507ndash512 2013

[55] M Pester T Rattei S Flechl et al ldquoAmoA-based consensusphylogeny of ammonia-oxidizing archaea and deep sequencingof amoA genes from soils of four different geographic regionsrdquoEnvironmental Microbiology vol 14 no 2 pp 525ndash539 2012

[56] J-Z He H-W Hu and L-M Zhang ldquoCurrent insights into theautotrophic thaumarchaeal ammonia oxidation in acidic soilsrdquoSoil Biology amp Biochemistry vol 55 pp 146ndash154 2012

[57] L-M Zhang H-W Hu J-P Shen and J-Z He ldquoAmmonia-oxidizing archaea have more important role than ammonia-oxidizing bacteria in ammonia oxidation of strongly acidicsoilsrdquo ISME Journal vol 6 no 5 pp 1032ndash1045 2012

[58] S Leininger T Urich M Schloter et al ldquoArchaea predominateamong ammonia-oxidizing prokaryotes in soilsrdquo Nature vol442 no 7104 pp 806ndash809 2006

[59] N C Banning L D Maccarone L M Fisk and D VMurphy ldquoAmmonia-oxidising bacteria not archaea dominatenitrification activity in semi-arid agricultural soilrdquo ScientificReports vol 5 Article ID 11146 2015

[60] M Herrmann A Hadrich and K Kusel ldquoPredominance ofthaumarchaeal ammonia oxidizer abundance and transcrip-tional activity in an acidic fenrdquoEnvironmentalMicrobiology vol14 no 11 pp 3013ndash3025 2012

[61] K Zhalnina P Dorr de Quadros F O Camargo and E WTriplett ldquoDrivers of archaeal ammonia-oxidizing communitiesin soilrdquo Frontiers in Microbiology vol 3 article 210 Article IDArticle 210 2012

[62] J Hong J Cho and Y Hong ldquoEnvironmental variables shapingthe ecological niche of Thaumarchaeota in soil direct andindirect causal effectsrdquo PLoS ONE vol 10 no 8 Article IDe0133763 2015

[63] G Jurgens K Lindstrom and A Saano ldquoNovel group withinthe kingdom Crenarchaeota from boreal forest soilrdquo Appliedand Environmental Microbiology vol 63 no 2 pp 803ndash8051997

[64] M Tourna M Stieglmeier A Spang et al ldquoNitrososphaeraviennensis an ammonia oxidizing archaeon from soilrdquoProceed-ings of the National Academy of Sciences of the United States ofAmerica vol 108 no 20 pp 8420ndash8425 2011

[65] K V Zhalnina R Dias M T Leonard et al ldquoGenome sequenceof Candidatus Nitrososphaera evergladensis from group I1benriched from everglades soil reveals novel genomic features ofthe ammonia-oxidizing archaeardquoPLoSONE vol 9 no 7 ArticleID e101648 2014

[66] I Anderson L E Ulrich B Lupa et al ldquoGenomic char-acterization of methanomicrobiales reveals three classes ofmethanogensrdquo PLoS ONE vol 4 no 6 Article ID e5797 2009

[67] J E Galagan C Nusbaum A Roy et al ldquoThe genome of Macetivorans reveals extensive metabolic and physiological diver-sityrdquo Genome Research vol 12 no 4 pp 532ndash542 2002

[68] C Wrede S Kokoschka A Dreier C Heller J Reitner and MHoppert ldquoDeposition of biogenic iron minerals in a methaneoxidizing microbial matrdquo Archaea vol 2013 Article ID 1029728 pages 2013

[69] G Garrity Bergeyrsquo Manual of Systematic Bacteriology 721 2001[70] S Sakai Y Takaki S Shimamura et al ldquoGenome sequence

of a mesophilic hydrogenotrophic methanogen Methanocellapaludicola the first cultivated representative of the orderMethanocellalesrdquo PLoS ONE vol 6 no 7 Article ID e228982011

[71] E Bapteste C Brochier and Y Boucher ldquoHigher-level clas-sification of the Archaea evolution of methanogenesis andmethanogensrdquo Archaea vol 1 no 5 pp 353ndash363 2005

[72] R G E Murray and E Stackebrandt ldquoTaxonomic note imple-mentation of the provisional status Candidatus for incompletelydescribed procaryotesrdquo International Journal of Systematic Bac-teriology vol 45 no 1 pp 186ndash187 1995

[73] L E Lehtovirta-Morley K Stoecker A Vilcinskas J I Prosserand G W Nicol ldquoCultivation of an obligate acidophilic ammo-nia oxidizer from a nitrifying acid soilrdquo Proceedings of the

14 Archaea

National Academy of Sciences of the United States of Americavol 108 no 38 pp 15892ndash15897 2011

[74] L E Lehtovirta-Morley C Ge J Ross H Yao G W Nicol andJ I Prosser ldquoCharacterisation of terrestrial acidophilic archaealammonia oxidisers and their inhibition and stimulation byorganic compoundsrdquo FEMSMicrobiology Ecology vol 89 no 3pp 542ndash552 2015

[75] M J Frijlink T Abee H J Laanbroek W de Boer and W NKonings ldquoThe bioenergetics of ammonia and hydroxylamineoxidation in Nitrosomonas europaea at acid and alkaline pHrdquoArchives of Microbiology vol 157 no 2 pp 194ndash199 1992

[76] B Wang Y Zheng R Huang et al ldquoActive ammonia oxidizersin an acidic soil are phylogenetically closely related to neu-trophilic archaeonrdquo Applied and Environmental Microbiologyvol 80 no 5 pp 1684ndash1691 2014

[77] J Y Aller and P F Kemp ldquoAre Archaea inherently less diversethan Bacteria in the same environmentsrdquo FEMS MicrobiologyEcology vol 65 no 1 pp 74ndash87 2008

[78] K Potthast U Hamer and FMakeschin ldquoLand-use change in atropical mountain rainforest region of southern Ecuador affectssoilmicroorganisms and nutrient cyclingrdquoBiogeochemistry vol111 no 1ndash3 pp 151ndash167 2012

[79] J I Prosser and G W Nicol ldquoArchaeal and bacterial ammonia-oxidisers in soil the quest for niche specialisation and differen-tiationrdquoTrends inMicrobiology vol 20 no 11 pp 523ndash531 2012

Submit your manuscripts athttpwwwhindawicom

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Anatomy Research International

PeptidesInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporation httpwwwhindawicom

International Journal of

Volume 2014

Zoology

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Molecular Biology International

GenomicsInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioinformaticsAdvances in

Marine BiologyJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Signal TransductionJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioMed Research International

Evolutionary BiologyInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Biochemistry Research International

ArchaeaHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Genetics Research International

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Advances in

Virolog y

Hindawi Publishing Corporationhttpwwwhindawicom

Nucleic AcidsJournal of

Volume 2014

Stem CellsInternational

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Enzyme Research

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

International Journal of

Microbiology

12 Archaea

in Brazil and alternative biomass feedstocksrdquo Renewable ampSustainable Energy Reviews vol 21 pp 411ndash420 2013

[10] R G Taketani and S M Tsai ldquoThe influence of different landuses on the structure of archaeal communities in Amazoniananthrosols based on 16S rRNA and amoA genesrdquo MicrobialEcology vol 59 no 4 pp 734ndash743 2010

[11] C Brochier-Armanet B Boussau S Gribaldo and P ForterreldquoMesophilic crenarchaeota proposal for a third archaeal phy-lum theThaumarchaeotardquoNature Reviews Microbiology vol 6no 3 pp 245ndash252 2008

[12] S M Barns C F Delwiche J D Palmer and N R PaceldquoPerspectives on archaeal diversity thermophily and mono-phyly from environmental rRNA sequencesrdquo Proceedings of theNational Academy of Sciences of the United States of Americavol 93 no 17 pp 9188ndash9193 1996

[13] J G Elkins M Podar D E Graham et al ldquoA korarchaealgenome reveals insights into the evolution of the Archaeardquo Pro-ceedings of the National Academy of Sciences of the United Statesof America vol 105 no 23 pp 8102ndash8107 2008

[14] H Huber M J Hohn R Rachel T Fuchs V C Wimmer andK O Stetter ldquoA new phylum of Archaea represented by ananosized hyperthermophilic symbiontrdquo Nature vol 417 no6884 pp 63ndash67 2002

[15] T Nunoura H Oida M Nakaseama et al ldquoArchaeal diversityand distribution along thermal and geochemical gradients inhydrothermal sediments at the Yonaguni Knoll IV hydrother-mal field in the SouthernOkinawa TroughrdquoApplied and Enviro-nmental Microbiology vol 76 no 4 pp 1198ndash1211 2010

[16] C Rinke P Schwientek A Sczyrba et al ldquoInsights into thephylogeny and coding potential of microbial dark matterrdquoNature vol 499 no 7459 pp 431ndash437 2013

[17] A Spang J H Saw S L Joslashrgensen et al ldquoComplex archaea thatbridge the gap between prokaryotes and eukaryotesrdquo Naturevol 521 no 7551 pp 173ndash179 2015

[18] S Bagchi S E Vlaeminck L A Sauder et al ldquoTemporal andspatial stability of ammonia-oxidizing archaea and bacteria inaquariumbiofiltersrdquo PLoSONE vol 9 no 12 Article ID e1135152014

[19] W Qin S A Amin W Martens-Habbena et al ldquoMarineammonia-oxidizing archaeal isolates display obligate mixotro-phy and wide ecotypic variationrdquo Proceedings of the NationalAcademy of Sciences of the United States of America vol 111 no34 pp 12504ndash12509 2014

[20] M Coci N Odermatt M M Salcher J Pernthaler and GCorno ldquoEcology and distribution ofThaumarchaea in the deephypolimnion of Lake Maggiorerdquo Archaea vol 2015 Article ID590434 11 pages 2015

[21] J G Ferry ldquoHow tomake a living by exhalingmethanerdquoAnnualReview of Microbiology vol 64 pp 453ndash473 2010

[22] A Meyer A Focks V Radl G Welzl I Schoning and MSchloter ldquoInfluence of land use intensity on the diversity ofammonia oxidizing bacteria and archaea in soils from grasslandecosystemsrdquoMicrobial Ecology vol 67 no 1 pp 161ndash166 2014

[23] V De Gannes G Eudoxie and W J Hickey ldquoImpacts ofedaphic factors on communities of ammonia-oxidizing archaeaammonia-oxidizing bacteria and nitrification in tropical soilsrdquoPLoS ONE vol 9 no 2 Article ID e89568 2014

[24] D S Ribeiro F C J da Silva and TN Carvalho ldquoSobrevivenciade seis especies florestais em uma area explorada seletivamenteno municıpio de Moju Parardquo CERNE vol 9 no 2 pp 153ndash1632003

[25] D HM Costa C A P Ferreira J NM Silva J D C A Lopesand J O P D Carvalho ldquoPotencial madeireiro de floresta densano municıpio de Moju Estado do Parardquo Documentos 1998

[26] C M Almeida S F Lima C V Martins-Da-Silva and JI Gomes ldquoCaracterizacao morfologica e anatomica de dezespecies de leguminosae ocorrentes em uma floresta tropicalumida localizada no municıpio de Moju Estado do Parardquo inA Silvicultura na Amazonia Oriental Contribuicoes do ProjetoEmbrapa-DFID J NMC Silva JO P deCarvalho and J AGYared Eds pp 19ndash54 Embrapa Amazonia Oriental (CPATU)Belem Brazil 2001

[27] K Smalla N Cresswell L C Mendoncahagler A Woltersand J D Vanelsas ldquoRapid extraction protocol from soil forpolymerase chain reaction-mediated amplificationrdquo Journal ofApplied Bacteriology vol 74 pp 78ndash85 1993

[28] S Gantner A F Andersson L Alonso-Saez and S Bertils-son ldquoNovel primers for 16S rRNA-based archaeal communityanalyses in environmental samplesrdquo Journal of MicrobiologicalMethods vol 84 no 1 pp 12ndash18 2011

[29] P D Schloss S L Westcott T Ryabin et al ldquoIntroducingMOTHUR open-source platform-independent community-supported software for describing and comparing microbialcommunitiesrdquoApplied and EnvironmentalMicrobiology vol 75no 23 pp 7537ndash7541 2009

[30] PD SchlossDGevers and S LWestcott ldquoReducing the effectsof PCR amplification and sequencing artifacts on 16S rRNA-based studiesrdquo PLoS ONE vol 6 no 12 Article ID e27310 2011

[31] CQuince A Lanzen T P Curtis et al ldquoAccurate determinationof microbial diversity from 454 pyrosequencing datardquo NatureMethods vol 6 no 9 pp 639ndash641 2009

[32] P Yilmaz L W Parfrey P Yarza et al ldquoThe SILVA and lsquoall-species Living Tree Project (LTP)rsquo taxonomic frameworksrdquoNucleic Acids Research vol 42 no 1 pp D643ndashD648 2014

[33] R C Edgar B J Haas J C Clemente C Quince and RKnight ldquoUCHIME improves sensitivity and speed of chimeradetectionrdquo Bioinformatics vol 27 no 16 pp 2194ndash2200 2011

[34] A Chao and T-J Shen ldquoNonparametric estimation of Shan-nonrsquos index of diversity when there are unseen species insamplerdquo Environmental and Ecological Statistics vol 10 no 4pp 429ndash443 2003

[35] A Chao and T-J Shen Program SPADE (Species Prediction andDiversity Estimation) 2010

[36] C L Shannon andWWeaverTheMathematicalTheory of Com-munication University of Illinois Press Urbana Ill USA 1949

[37] I J Good ldquoThe population frequencies of species and theestimation of population parametersrdquo Biometrika vol 40 pp237ndash264 1953

[38] K Tamura G Stecher D Peterson A Filipski and S KumarldquoMEGA6molecular evolutionary genetics analysis version 60rdquoMolecular Biology and Evolution vol 30 no 12 pp 2725ndash27292013

[39] DA BensonMCavanaugh K Clark et al ldquoGenBankrdquoNucleicAcids Research vol 41 no 1 pp D36ndashD42 2013

[40] D H Parks G W Tyson P Hugenholtz and R G BeikoldquoSTAMP statistical analysis of taxonomic and functional pro-filesrdquo Bioinformatics vol 30 no 21 pp 3123ndash3124 2014

[41] R G Newcombe ldquoInterval estimation for the differencebetween independent proportions comparison of elevenmeth-odsrdquo Statistics in Medicine vol 17 no 8 pp 873ndash890 1998

[42] J D Storey J E Taylor and D Siegmund ldquoStrong controlconservative point estimation and simultaneous conservative

Archaea 13

consistency of false discovery rates a unified approachrdquo Journalof the Royal Statistical Society Series B Statistical Methodologyvol 66 no 1 pp 187ndash205 2004

[43] G C Baker J J Smith and D A Cowan ldquoReview and re-analysis of domain-specific 16S primersrdquo Journal of Microbio-logical Methods vol 55 no 3 pp 541ndash555 2003

[44] W H Zhong and Z C Cai ldquoLong-term effects of inorganicfertilizers on microbial biomass and community functionaldiversity in a paddy soil derived from quaternary red clayrdquoApplied Soil Ecology vol 36 no 2-3 pp 84ndash91 2007

[45] EWessen S Hallin and L Philippot ldquoDifferential responses ofbacterial and archaeal groups at high taxonomical ranks to soilmanagementrdquo Soil Biology and Biochemistry vol 42 no 10 pp1759ndash1765 2010

[46] ZWuW Lin B Li et al ldquoTerminal restriction fragment lengthpolymorphism analysis of soil bacterial communities underdifferent vegetation types in subtropical area rdquo PLOS ONE vol10 no 6 Article ID e0129397 2015

[47] K Cenciani M R Lambais C C Cerri L C B De Azevedoand B J Feigl ldquoBacteria diversity and microbial biomass inforest pasture and fallow soils in the southwestern Amazonbasinrdquo Revista Brasileira de Ciencia do Solo vol 33 no 4 pp907ndash916 2009

[48] A A Navarrete R G Taketani L WMendes F de Souza Can-navan F M de Souza Moreira and S M Tsai ldquoLand-use sys-tems affect Archaeal community structure and functional diver-sity in western Amazon soilsrdquo Revista Brasileira de Ciencia doSolo vol 35 no 5 pp 1527ndash1540 2011

[49] E Lumini A Orgiazzi R Borriello P Bonfante and V Bian-ciotto ldquoDisclosing arbuscular mycorrhizal fungal biodiversityin soil through a land-use gradient using a pyrosequencingapproachrdquo Environmental Microbiology vol 12 no 8 pp 2165ndash2179 2010

[50] M Lupatini R J S Jacques Z I Antoniolli A K A SuleimanR R Fulthorpe and L F W Roesch ldquoLand-use change andsoil type are drivers of fungal and archaeal communities in thePampa biomerdquo World Journal of Microbiology and Biotechnol-ogy vol 29 no 2 pp 223ndash233 2013

[51] E G Lamb N Kennedy and S D Siciliano ldquoEffects of plantspecies richness and evenness on soil microbial communitydiversity and functionrdquo Plant and Soil vol 338 no 1 pp 483ndash495 2011

[52] A Spang R Hatzenpichler C Brochier-Armanet et al ldquoDis-tinct gene set in two different lineages of ammonia-oxidizingarchaea supports the phylum Thaumarchaeotardquo Trends inMicrobiology vol 18 no 8 pp 331ndash340 2010

[53] R G Pacchioni F M Carvalho C E Thompson et al ldquoTax-onomic and functional profiles of soil samples from Atlanticforest and Caatinga biomes in northeastern Brazilrdquo Microbio-logyOpen vol 3 no 3 pp 299ndash315 2014

[54] E Catao A P Castro C C Barreto R H Kruger and C MKyaw ldquoDiversity of Archaea in Brazilian savanna soilsrdquoArchivesof Microbiology vol 195 no 7 pp 507ndash512 2013

[55] M Pester T Rattei S Flechl et al ldquoAmoA-based consensusphylogeny of ammonia-oxidizing archaea and deep sequencingof amoA genes from soils of four different geographic regionsrdquoEnvironmental Microbiology vol 14 no 2 pp 525ndash539 2012

[56] J-Z He H-W Hu and L-M Zhang ldquoCurrent insights into theautotrophic thaumarchaeal ammonia oxidation in acidic soilsrdquoSoil Biology amp Biochemistry vol 55 pp 146ndash154 2012

[57] L-M Zhang H-W Hu J-P Shen and J-Z He ldquoAmmonia-oxidizing archaea have more important role than ammonia-oxidizing bacteria in ammonia oxidation of strongly acidicsoilsrdquo ISME Journal vol 6 no 5 pp 1032ndash1045 2012

[58] S Leininger T Urich M Schloter et al ldquoArchaea predominateamong ammonia-oxidizing prokaryotes in soilsrdquo Nature vol442 no 7104 pp 806ndash809 2006

[59] N C Banning L D Maccarone L M Fisk and D VMurphy ldquoAmmonia-oxidising bacteria not archaea dominatenitrification activity in semi-arid agricultural soilrdquo ScientificReports vol 5 Article ID 11146 2015

[60] M Herrmann A Hadrich and K Kusel ldquoPredominance ofthaumarchaeal ammonia oxidizer abundance and transcrip-tional activity in an acidic fenrdquoEnvironmentalMicrobiology vol14 no 11 pp 3013ndash3025 2012

[61] K Zhalnina P Dorr de Quadros F O Camargo and E WTriplett ldquoDrivers of archaeal ammonia-oxidizing communitiesin soilrdquo Frontiers in Microbiology vol 3 article 210 Article IDArticle 210 2012

[62] J Hong J Cho and Y Hong ldquoEnvironmental variables shapingthe ecological niche of Thaumarchaeota in soil direct andindirect causal effectsrdquo PLoS ONE vol 10 no 8 Article IDe0133763 2015

[63] G Jurgens K Lindstrom and A Saano ldquoNovel group withinthe kingdom Crenarchaeota from boreal forest soilrdquo Appliedand Environmental Microbiology vol 63 no 2 pp 803ndash8051997

[64] M Tourna M Stieglmeier A Spang et al ldquoNitrososphaeraviennensis an ammonia oxidizing archaeon from soilrdquoProceed-ings of the National Academy of Sciences of the United States ofAmerica vol 108 no 20 pp 8420ndash8425 2011

[65] K V Zhalnina R Dias M T Leonard et al ldquoGenome sequenceof Candidatus Nitrososphaera evergladensis from group I1benriched from everglades soil reveals novel genomic features ofthe ammonia-oxidizing archaeardquoPLoSONE vol 9 no 7 ArticleID e101648 2014

[66] I Anderson L E Ulrich B Lupa et al ldquoGenomic char-acterization of methanomicrobiales reveals three classes ofmethanogensrdquo PLoS ONE vol 4 no 6 Article ID e5797 2009

[67] J E Galagan C Nusbaum A Roy et al ldquoThe genome of Macetivorans reveals extensive metabolic and physiological diver-sityrdquo Genome Research vol 12 no 4 pp 532ndash542 2002

[68] C Wrede S Kokoschka A Dreier C Heller J Reitner and MHoppert ldquoDeposition of biogenic iron minerals in a methaneoxidizing microbial matrdquo Archaea vol 2013 Article ID 1029728 pages 2013

[69] G Garrity Bergeyrsquo Manual of Systematic Bacteriology 721 2001[70] S Sakai Y Takaki S Shimamura et al ldquoGenome sequence

of a mesophilic hydrogenotrophic methanogen Methanocellapaludicola the first cultivated representative of the orderMethanocellalesrdquo PLoS ONE vol 6 no 7 Article ID e228982011

[71] E Bapteste C Brochier and Y Boucher ldquoHigher-level clas-sification of the Archaea evolution of methanogenesis andmethanogensrdquo Archaea vol 1 no 5 pp 353ndash363 2005

[72] R G E Murray and E Stackebrandt ldquoTaxonomic note imple-mentation of the provisional status Candidatus for incompletelydescribed procaryotesrdquo International Journal of Systematic Bac-teriology vol 45 no 1 pp 186ndash187 1995

[73] L E Lehtovirta-Morley K Stoecker A Vilcinskas J I Prosserand G W Nicol ldquoCultivation of an obligate acidophilic ammo-nia oxidizer from a nitrifying acid soilrdquo Proceedings of the

14 Archaea

National Academy of Sciences of the United States of Americavol 108 no 38 pp 15892ndash15897 2011

[74] L E Lehtovirta-Morley C Ge J Ross H Yao G W Nicol andJ I Prosser ldquoCharacterisation of terrestrial acidophilic archaealammonia oxidisers and their inhibition and stimulation byorganic compoundsrdquo FEMSMicrobiology Ecology vol 89 no 3pp 542ndash552 2015

[75] M J Frijlink T Abee H J Laanbroek W de Boer and W NKonings ldquoThe bioenergetics of ammonia and hydroxylamineoxidation in Nitrosomonas europaea at acid and alkaline pHrdquoArchives of Microbiology vol 157 no 2 pp 194ndash199 1992

[76] B Wang Y Zheng R Huang et al ldquoActive ammonia oxidizersin an acidic soil are phylogenetically closely related to neu-trophilic archaeonrdquo Applied and Environmental Microbiologyvol 80 no 5 pp 1684ndash1691 2014

[77] J Y Aller and P F Kemp ldquoAre Archaea inherently less diversethan Bacteria in the same environmentsrdquo FEMS MicrobiologyEcology vol 65 no 1 pp 74ndash87 2008

[78] K Potthast U Hamer and FMakeschin ldquoLand-use change in atropical mountain rainforest region of southern Ecuador affectssoilmicroorganisms and nutrient cyclingrdquoBiogeochemistry vol111 no 1ndash3 pp 151ndash167 2012

[79] J I Prosser and G W Nicol ldquoArchaeal and bacterial ammonia-oxidisers in soil the quest for niche specialisation and differen-tiationrdquoTrends inMicrobiology vol 20 no 11 pp 523ndash531 2012

Submit your manuscripts athttpwwwhindawicom

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Anatomy Research International

PeptidesInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporation httpwwwhindawicom

International Journal of

Volume 2014

Zoology

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Molecular Biology International

GenomicsInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioinformaticsAdvances in

Marine BiologyJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Signal TransductionJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioMed Research International

Evolutionary BiologyInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Biochemistry Research International

ArchaeaHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Genetics Research International

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Advances in

Virolog y

Hindawi Publishing Corporationhttpwwwhindawicom

Nucleic AcidsJournal of

Volume 2014

Stem CellsInternational

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Enzyme Research

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

International Journal of

Microbiology

Archaea 13

consistency of false discovery rates a unified approachrdquo Journalof the Royal Statistical Society Series B Statistical Methodologyvol 66 no 1 pp 187ndash205 2004

[43] G C Baker J J Smith and D A Cowan ldquoReview and re-analysis of domain-specific 16S primersrdquo Journal of Microbio-logical Methods vol 55 no 3 pp 541ndash555 2003

[44] W H Zhong and Z C Cai ldquoLong-term effects of inorganicfertilizers on microbial biomass and community functionaldiversity in a paddy soil derived from quaternary red clayrdquoApplied Soil Ecology vol 36 no 2-3 pp 84ndash91 2007

[45] EWessen S Hallin and L Philippot ldquoDifferential responses ofbacterial and archaeal groups at high taxonomical ranks to soilmanagementrdquo Soil Biology and Biochemistry vol 42 no 10 pp1759ndash1765 2010

[46] ZWuW Lin B Li et al ldquoTerminal restriction fragment lengthpolymorphism analysis of soil bacterial communities underdifferent vegetation types in subtropical area rdquo PLOS ONE vol10 no 6 Article ID e0129397 2015

[47] K Cenciani M R Lambais C C Cerri L C B De Azevedoand B J Feigl ldquoBacteria diversity and microbial biomass inforest pasture and fallow soils in the southwestern Amazonbasinrdquo Revista Brasileira de Ciencia do Solo vol 33 no 4 pp907ndash916 2009

[48] A A Navarrete R G Taketani L WMendes F de Souza Can-navan F M de Souza Moreira and S M Tsai ldquoLand-use sys-tems affect Archaeal community structure and functional diver-sity in western Amazon soilsrdquo Revista Brasileira de Ciencia doSolo vol 35 no 5 pp 1527ndash1540 2011

[49] E Lumini A Orgiazzi R Borriello P Bonfante and V Bian-ciotto ldquoDisclosing arbuscular mycorrhizal fungal biodiversityin soil through a land-use gradient using a pyrosequencingapproachrdquo Environmental Microbiology vol 12 no 8 pp 2165ndash2179 2010

[50] M Lupatini R J S Jacques Z I Antoniolli A K A SuleimanR R Fulthorpe and L F W Roesch ldquoLand-use change andsoil type are drivers of fungal and archaeal communities in thePampa biomerdquo World Journal of Microbiology and Biotechnol-ogy vol 29 no 2 pp 223ndash233 2013

[51] E G Lamb N Kennedy and S D Siciliano ldquoEffects of plantspecies richness and evenness on soil microbial communitydiversity and functionrdquo Plant and Soil vol 338 no 1 pp 483ndash495 2011

[52] A Spang R Hatzenpichler C Brochier-Armanet et al ldquoDis-tinct gene set in two different lineages of ammonia-oxidizingarchaea supports the phylum Thaumarchaeotardquo Trends inMicrobiology vol 18 no 8 pp 331ndash340 2010

[53] R G Pacchioni F M Carvalho C E Thompson et al ldquoTax-onomic and functional profiles of soil samples from Atlanticforest and Caatinga biomes in northeastern Brazilrdquo Microbio-logyOpen vol 3 no 3 pp 299ndash315 2014

[54] E Catao A P Castro C C Barreto R H Kruger and C MKyaw ldquoDiversity of Archaea in Brazilian savanna soilsrdquoArchivesof Microbiology vol 195 no 7 pp 507ndash512 2013

[55] M Pester T Rattei S Flechl et al ldquoAmoA-based consensusphylogeny of ammonia-oxidizing archaea and deep sequencingof amoA genes from soils of four different geographic regionsrdquoEnvironmental Microbiology vol 14 no 2 pp 525ndash539 2012

[56] J-Z He H-W Hu and L-M Zhang ldquoCurrent insights into theautotrophic thaumarchaeal ammonia oxidation in acidic soilsrdquoSoil Biology amp Biochemistry vol 55 pp 146ndash154 2012

[57] L-M Zhang H-W Hu J-P Shen and J-Z He ldquoAmmonia-oxidizing archaea have more important role than ammonia-oxidizing bacteria in ammonia oxidation of strongly acidicsoilsrdquo ISME Journal vol 6 no 5 pp 1032ndash1045 2012

[58] S Leininger T Urich M Schloter et al ldquoArchaea predominateamong ammonia-oxidizing prokaryotes in soilsrdquo Nature vol442 no 7104 pp 806ndash809 2006

[59] N C Banning L D Maccarone L M Fisk and D VMurphy ldquoAmmonia-oxidising bacteria not archaea dominatenitrification activity in semi-arid agricultural soilrdquo ScientificReports vol 5 Article ID 11146 2015

[60] M Herrmann A Hadrich and K Kusel ldquoPredominance ofthaumarchaeal ammonia oxidizer abundance and transcrip-tional activity in an acidic fenrdquoEnvironmentalMicrobiology vol14 no 11 pp 3013ndash3025 2012

[61] K Zhalnina P Dorr de Quadros F O Camargo and E WTriplett ldquoDrivers of archaeal ammonia-oxidizing communitiesin soilrdquo Frontiers in Microbiology vol 3 article 210 Article IDArticle 210 2012

[62] J Hong J Cho and Y Hong ldquoEnvironmental variables shapingthe ecological niche of Thaumarchaeota in soil direct andindirect causal effectsrdquo PLoS ONE vol 10 no 8 Article IDe0133763 2015

[63] G Jurgens K Lindstrom and A Saano ldquoNovel group withinthe kingdom Crenarchaeota from boreal forest soilrdquo Appliedand Environmental Microbiology vol 63 no 2 pp 803ndash8051997

[64] M Tourna M Stieglmeier A Spang et al ldquoNitrososphaeraviennensis an ammonia oxidizing archaeon from soilrdquoProceed-ings of the National Academy of Sciences of the United States ofAmerica vol 108 no 20 pp 8420ndash8425 2011

[65] K V Zhalnina R Dias M T Leonard et al ldquoGenome sequenceof Candidatus Nitrososphaera evergladensis from group I1benriched from everglades soil reveals novel genomic features ofthe ammonia-oxidizing archaeardquoPLoSONE vol 9 no 7 ArticleID e101648 2014

[66] I Anderson L E Ulrich B Lupa et al ldquoGenomic char-acterization of methanomicrobiales reveals three classes ofmethanogensrdquo PLoS ONE vol 4 no 6 Article ID e5797 2009

[67] J E Galagan C Nusbaum A Roy et al ldquoThe genome of Macetivorans reveals extensive metabolic and physiological diver-sityrdquo Genome Research vol 12 no 4 pp 532ndash542 2002

[68] C Wrede S Kokoschka A Dreier C Heller J Reitner and MHoppert ldquoDeposition of biogenic iron minerals in a methaneoxidizing microbial matrdquo Archaea vol 2013 Article ID 1029728 pages 2013

[69] G Garrity Bergeyrsquo Manual of Systematic Bacteriology 721 2001[70] S Sakai Y Takaki S Shimamura et al ldquoGenome sequence

of a mesophilic hydrogenotrophic methanogen Methanocellapaludicola the first cultivated representative of the orderMethanocellalesrdquo PLoS ONE vol 6 no 7 Article ID e228982011

[71] E Bapteste C Brochier and Y Boucher ldquoHigher-level clas-sification of the Archaea evolution of methanogenesis andmethanogensrdquo Archaea vol 1 no 5 pp 353ndash363 2005

[72] R G E Murray and E Stackebrandt ldquoTaxonomic note imple-mentation of the provisional status Candidatus for incompletelydescribed procaryotesrdquo International Journal of Systematic Bac-teriology vol 45 no 1 pp 186ndash187 1995

[73] L E Lehtovirta-Morley K Stoecker A Vilcinskas J I Prosserand G W Nicol ldquoCultivation of an obligate acidophilic ammo-nia oxidizer from a nitrifying acid soilrdquo Proceedings of the

14 Archaea

National Academy of Sciences of the United States of Americavol 108 no 38 pp 15892ndash15897 2011

[74] L E Lehtovirta-Morley C Ge J Ross H Yao G W Nicol andJ I Prosser ldquoCharacterisation of terrestrial acidophilic archaealammonia oxidisers and their inhibition and stimulation byorganic compoundsrdquo FEMSMicrobiology Ecology vol 89 no 3pp 542ndash552 2015

[75] M J Frijlink T Abee H J Laanbroek W de Boer and W NKonings ldquoThe bioenergetics of ammonia and hydroxylamineoxidation in Nitrosomonas europaea at acid and alkaline pHrdquoArchives of Microbiology vol 157 no 2 pp 194ndash199 1992

[76] B Wang Y Zheng R Huang et al ldquoActive ammonia oxidizersin an acidic soil are phylogenetically closely related to neu-trophilic archaeonrdquo Applied and Environmental Microbiologyvol 80 no 5 pp 1684ndash1691 2014

[77] J Y Aller and P F Kemp ldquoAre Archaea inherently less diversethan Bacteria in the same environmentsrdquo FEMS MicrobiologyEcology vol 65 no 1 pp 74ndash87 2008

[78] K Potthast U Hamer and FMakeschin ldquoLand-use change in atropical mountain rainforest region of southern Ecuador affectssoilmicroorganisms and nutrient cyclingrdquoBiogeochemistry vol111 no 1ndash3 pp 151ndash167 2012

[79] J I Prosser and G W Nicol ldquoArchaeal and bacterial ammonia-oxidisers in soil the quest for niche specialisation and differen-tiationrdquoTrends inMicrobiology vol 20 no 11 pp 523ndash531 2012

Submit your manuscripts athttpwwwhindawicom

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Anatomy Research International

PeptidesInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporation httpwwwhindawicom

International Journal of

Volume 2014

Zoology

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Molecular Biology International

GenomicsInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioinformaticsAdvances in

Marine BiologyJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Signal TransductionJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioMed Research International

Evolutionary BiologyInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Biochemistry Research International

ArchaeaHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Genetics Research International

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Advances in

Virolog y

Hindawi Publishing Corporationhttpwwwhindawicom

Nucleic AcidsJournal of

Volume 2014

Stem CellsInternational

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Enzyme Research

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

International Journal of

Microbiology

14 Archaea

National Academy of Sciences of the United States of Americavol 108 no 38 pp 15892ndash15897 2011

[74] L E Lehtovirta-Morley C Ge J Ross H Yao G W Nicol andJ I Prosser ldquoCharacterisation of terrestrial acidophilic archaealammonia oxidisers and their inhibition and stimulation byorganic compoundsrdquo FEMSMicrobiology Ecology vol 89 no 3pp 542ndash552 2015

[75] M J Frijlink T Abee H J Laanbroek W de Boer and W NKonings ldquoThe bioenergetics of ammonia and hydroxylamineoxidation in Nitrosomonas europaea at acid and alkaline pHrdquoArchives of Microbiology vol 157 no 2 pp 194ndash199 1992

[76] B Wang Y Zheng R Huang et al ldquoActive ammonia oxidizersin an acidic soil are phylogenetically closely related to neu-trophilic archaeonrdquo Applied and Environmental Microbiologyvol 80 no 5 pp 1684ndash1691 2014

[77] J Y Aller and P F Kemp ldquoAre Archaea inherently less diversethan Bacteria in the same environmentsrdquo FEMS MicrobiologyEcology vol 65 no 1 pp 74ndash87 2008

[78] K Potthast U Hamer and FMakeschin ldquoLand-use change in atropical mountain rainforest region of southern Ecuador affectssoilmicroorganisms and nutrient cyclingrdquoBiogeochemistry vol111 no 1ndash3 pp 151ndash167 2012

[79] J I Prosser and G W Nicol ldquoArchaeal and bacterial ammonia-oxidisers in soil the quest for niche specialisation and differen-tiationrdquoTrends inMicrobiology vol 20 no 11 pp 523ndash531 2012

Submit your manuscripts athttpwwwhindawicom

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Anatomy Research International

PeptidesInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporation httpwwwhindawicom

International Journal of

Volume 2014

Zoology

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Molecular Biology International

GenomicsInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioinformaticsAdvances in

Marine BiologyJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Signal TransductionJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioMed Research International

Evolutionary BiologyInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Biochemistry Research International

ArchaeaHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Genetics Research International

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Advances in

Virolog y

Hindawi Publishing Corporationhttpwwwhindawicom

Nucleic AcidsJournal of

Volume 2014

Stem CellsInternational

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Enzyme Research

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

International Journal of

Microbiology

Submit your manuscripts athttpwwwhindawicom

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Anatomy Research International

PeptidesInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporation httpwwwhindawicom

International Journal of

Volume 2014

Zoology

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Molecular Biology International

GenomicsInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioinformaticsAdvances in

Marine BiologyJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Signal TransductionJournal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

BioMed Research International

Evolutionary BiologyInternational Journal of

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Biochemistry Research International

ArchaeaHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Genetics Research International

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Advances in

Virolog y

Hindawi Publishing Corporationhttpwwwhindawicom

Nucleic AcidsJournal of

Volume 2014

Stem CellsInternational

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Enzyme Research

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

International Journal of

Microbiology