Microbial biomass and community composition of a tallgrass prairie soil subjected to simulated global warming and clipping

A. Belay-Tedla, M. Elshahed, L. Krumholz, Y. LuoDepartment of Botany and Microbiology, University of Oklahoma, Norman, OK 73019, USA

Abstract

Introduction

Soil microorganisms are the driving force behind most belowground processes and may considerably influence ecosystem responses to global warming. However, the consequences of warming on the microbial community, both in terms of total biomass and species composition, are not known for certain. The present study assessed the effects of experimental warming and clipping on soil microbial biomass contents and examined warming-induced alterations in the microbial community structure and diversity using the 16S ribosomal ribonucleic acid (16S rRNA) sequence analysis.

Materials and methods

Site descriptions and experimental design: The study was conducted in the Great Plain Apiaries of Central Oklahoma (34058’54’’ N, 97031’14’’ W). The design of the experiment was a paired factorial with warming as the main factor and clipping as the secondary factor, replicated six times. The plot size was 2mX2m each divided into four 1mX1m subplots. Two diagonal subplots in each main plot were clipped 10 cm above the ground once every year. The other two diagonal subplots were the unclipped control. An infrared radiator (Kalglo Electronics, Bethlehem, Pennsylvania) suspended at 1.5 m above the ground in each warmed plot was used as the heating device. The plots have been warmed continuously since November 21, 1999. The four treatments in the experiment were clipped control (CC), unclipped control (UC), clipped warmed (CW) and unclipped warmed (UW).

Sampling and analysis: soil samples were collected at 0-20cm depth in the Winter and Spring seasons between 2002 and 2004.

- Soil microbial biomass: Chloroform-fumigation

- Community composition: 16S rRNA

The present study examined the effects of simulated global warming and clipping on soil microbial biomass and community composition of tallgrass prairie soil in Central Oklahoma. Soil microbial biomass C and N contents were significantly increased (>50%) by the warming but remained unaffected by the clipping treatment. Warming also enhanced the microbes’ C and N use efficiency by more than 48%. The 16S ribosomal RNA analysis revealed seasonal and differential effects of warming on bacterial diversity and community composition. Assays conducted in the present study have provided both metabolic and molecular evidence that soil microorganisms may adjust their activity rate, physiology, and/or community composition in response to global warming.

Fig. 2. Soil microbial biomass C (SMB-C) and N (SMB-N) on two different seasons. SMB-C in unclipped plots increased by >50% as a result of the warming treatment. SMB-N was also higher (with average increase of more than 20 mg/kg over other treatments) in the UW plots but differed significantly only from CC plots.

Results

Infrared heater“dummy” heater

clip

clip

Unclip Unclip

Soil microbial biomass C and N

SMB-C

0

200

400

600

800

1000

19/12/02 27/6/03

Date

mg

C k

g-1

dry

soi

l CC

UC

CW

UW

b bb aab

b b

a

a SMB-N

0

20

40

60

80

100

120

19/12/02 27/6/03

Date

mg

N k

g-1

dry

soi

l CC

UC

CW

UW

aaab

b

abab

b

a

b

SMB:LP ratio

0

0.1

0.2

0.3

0.4

0.5

0.6

SMBC/LPC SMBN/LPN

SM

B:L

P r

ati

o CC

UC

CW

UW

abb

aba

bab

aab

Fig. 3. Soil microbial biomass C (SMB-C) and N (SMB-N) to labile C (LPC) and N (LPN) ratio. SMB-C:LP-C rose by 48, 31, and 22% in UW, CW, and CC. SMB-N:LPN ratio also increased by warming

Soil sample(0.5 g)

Bulk DNA

Amplified16S rRNA

16S rRNAclones

16S rRNAsequences

FastDNA® SPIN Kit for Soil (Qbiogene, Inc.)

Gene Amp PCR system 9700 thermocycler

TOPO-TA

Advanced Center forGenome Technology

DNA extraction

PCR

Cloning

Sequencing

Fig. 1. Overview of the 16S rRNA procedure

Library Season Clone # OTU #

Control

Warmed

Spring ’03

Winter ’04

Spring ’03

Winter ‘04

152

102

137

93

105

81

107

81

Bacterial community diversity and composition

Table 1. Total number of clones sequenced and OTUs identified.Clone number in UC higher by 10% ; OTU richness was similar for both.

Shannon-Weiner index (H') and species eveness (J')

0

0.5

1

1.5

2

2.5

3

H' J'

ind

ex Control

Warmed

Shannon-Weiner index (H') and species evenness (J')

0

0.5

1

1.5

2

2.5

H' J'

Ind

ex Control

Warmed

Fig. 4. Shannon-Weinerindex and species even-ness (Sp03; Win04).

Expected number of OTUs vs clones

0

20

40

60

80

100

120

1 10 19 28 37 46 55 64 73 82 91 100 109 118 127 136 145

Number of clones

Exp

ecte

d O

TU

s Control

Upper95

Lower95

Warmed

Upper95

Lower95

Expected number of OTUs vs. clones

0

10

20

30

40

50

60

70

80

90

100

1 7 13 19 25 31 37 43 49 55 61 67 73 79 85 91

Number of clones

Exp

ecte

d O

TU

s

Control

Warmed

Fig. 5. Collectors’ curveAnalysis (Sp03; Win04)

Distribution of clones

05

10152025303540

Bacterial division

% c

lone

s

Control

Warmed

Clone distribution

05

101520253035

Bacterial division

% C

lone

s

Conrol

Warmed

Fig. 7. Bacterial clone distribution. There was a persistent relatively higherproportion of bacterial groups belonging to Actinobacteria, CFB, beta-Proteobacteria, Chloroflexi and comparative decline of those belonging toAlpha-proteobacteria, Gamma-proteobacteria, and Gametoides.

ConclusionThe study has demonstrated consistent and significant increases of soil microbial biomass indicating enhanced microbial activity in response to warming. An alteration in C and N use efficiency of the microbes was also observed, suggesting warming-induced physiological changes. The observed differences in the relative proportions of different bacterial groups indicate that the microbial community structure or specific components of the population could have been changed as a result of the imposed warming.