Global ocean’s protist metabarcodingColomban de Vargas, Stéphane Audic, Nicolas Henry, Johan Decelle, Frédéric Mahé, Cédric Berney, Sébastien Colin, Sarah Romac, Daniel Richter, Ian Probert, Raffaele Siano, Gipsi Lima-Mendez, Jeroen Raes, Chris Bowler, Patrick Wincker, Eric Karsenti, & the Tara Oceans Consortium.

43 samples (viral fraction <0.22um) from 26 stations; 2.16 billion Illumina reads (200X larger than the Pacific Ocean Virome).

pelagic upper-ocean viral community sequence space is now well-sampled and approaches a limit of ca. 1 million PCs

Protein Clusters(ORFs)

• 68 stations68 stations 7.2 Tbp DNA data7.2 Tbp DNA data• 3 depths3 depths in the context ofin the context of• 243 samples243 samples the environmentthe environment

A genetic inventory of the ocean:A genetic inventory of the ocean:Ocean Microbial Reference Gene CatalogOcean Microbial Reference Gene Catalog

High sequencing coverageHigh sequencing coverage• only few new genes peradditional sample• 4 x more genes than inhuman gut microbiome

meta-barcoding – V9 rDNA, the microscope of the III millenary

334 plankton communities

4 organismal size fractions

47 stations,sub-surface + DCM layers (photic zone)

~ 2 millions genetic barcodes per sample (total of ~800 million metabarcodes)

13,432 / 24,435 eukaryotic genera / species

all main lineages known from environmental sequences (Sanger clone libraries)

77,500 V9 rDNA reference sequences

V9 PR2* database:

Sub-divided into 97 major morpho-lineages

Annotated for basic ecological functions: auto/heterotrophy; symbioses sensu lato (from parasitism to mutualism, for both host and symbionts)

Stephane AUDIC

Cedric BERNEY

Database available at:http://taraoceans.sb-roscoff.fr/EukDiv/

Reaching the boundary of total eukaryotic plankton diversityin the world sunlit oceans (tropical to temperate):

150 000 genetic types (rDNA OTUs) of eukaryotic planktonFollowing a Preston curve

described species

Tara-Oceans

rDNAOTUs

4,350/12,800 1,350/10,000 5,500/15,300

66X

113X 92X

65X 38X

phototroph (phytoplancton)parasiteosmotroph/saprotrophphagotroph

✓ <1% of the OTUs are strictly identical to reference sequences.

✓ Even in known groups, genetic novelty is massive (e.g. diatoms, dinoflagellates).

✓ ~60 branches of the tree (2/3) are basically ignored from plankton ecology (~25% of assignable OTUs).

✓11 lineages are ‘hyper-diversified (>1,000 OTUs); mostly heterotrophic protists in poorly known eukaryotic supergroups (e.g. diplonemids).

✓ Overall poor diversity of phototrophic lineages (phytoplankton), in comparison to heterotrophic protists)

✓ Hyper-diversification in lineages extending across larger size-fractions, as well as their known parasites.

✓ >85% of eukaryotic OTUs belong to protists (zoo- not that important)

A large proportion of the uncovered diversity represents known or putative parasites / parasitoids.

examples of lineagesinfectingdiatoms

Rhynchopus coscinodiscivorus(Diplonemida) infectingCoscinodiscus concinnus

Cryothecomonaslongipes (Cercozoa)feeding onThalassiosira rotula

plasmodium ofPhagomyxa odontellae(Cercozoa) inside cells

of Odontella sinensis

Pirsonia diadema(Stramenopila) infectingCoscinodiscus wailesii

Dinomyces arenysensis(Chytridiomycota) infectingAlexandrium sp.

examples of lineages infecting

dinoflagellates

Amoebophrya sp.(MALV-II, Alveolata)infecting Alexandrium sp.

zoospores of Parvilucifera sinerae(Perkinsea, Alveolata) released

from Dinophysis caudata

examples of lineages infecting metazoans

Vampyrophryapelagica(Ciliophora,Alveolata)infectinga copepod

Paramikrocytos canceri(Ascetosporea, Cercozoa)infecting Cancer pagurus

Cephaloidophoroids(Apicomplexa, Alveolata),

parasites of copepods

Blastodinium sp.(Dinophyceae, Alveolata)infecting a copepod

Paradinium poucheti(Ascetosporea, Cercozoa)infecting Clausocalanus sp.

Functional metabarcoding

inference of basic trophic and symbiotic ecological modes

227 tree species (out of ±16,000) account for half of all trees in Amazonia

269 OTUs: hyperdominant & cosmopolitan 48%

of all reads

’Hyperdominance and cosmopolitasnism’

25% have poorlydefined identity

(< 95%)

11 are notassignable

at 85%

If 2 species always occur together (co-occurrence) -> mutualism, commensalism, similar niche?

If 2 species never occur together (mutual exclusion)-> competition, amensalism, opposing niches?

127,995 associations92,633 taxon-taxon35,362 taxon-env

co-occurrence muchmore common thanexclusion

•Data processing: sample-size normalization, keeping proportion of unclassified + filtered.• 2 similarity measures: Spearman and KL similarity• P-value calculation by matrix permutation (row shuffling), renormalization and bootstrapping (Faust et al. 2012).•P-value merging (edges supported by >= 2 methods)•Multiple test correction (Benjamini-Hochberg).

Prokaryotes: miTag abundancesPhages: metagenomic contig abundancesProtists: 18S metabarcode abundancesEnvironmental contextual data

68 stations, 2 depths, 7 size fractions

Top-down interactions potentially driving plankton community structure: (1) parasites

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Dinoflagellates infected by syndiniales

Experimental validation of network-predicted interaction (photosymbiosis)

Model for network-driven hypothesis generation

Laser scanning confocal microscopy (LSCM) of acoel flatwormwith endosymbiotic green algae (Tetraselmis)

UniEuk – towards a universal taxonomic frameworkand integrated reference gene databases foreukaryotic biology, ecology, and evolution

A community-based initiative, highly complementary to EukRef

UniEuk organizational scheme

A community-based initiative, highly complementary to EukRef

To address the current deluge of genetic data from environmentalgenetic surveys, meta-barcoding, -transcriptomics, -genomics, single-cell

transcriptomics and genomics, a common taxonomic framework is critical.

Without it, results of different studies using different genesand different reference databases would not be comparable.

common taxonomic framework

gene 1referencedatabase

gene 2referencedatabase

gene 3referencedatabase

gene 4referencedatabase

One taxonomic framework for multiple genetic markers

UniEuk: a pragmatic implementation

gene reference databases

universaltaxonomicframework

existinggenetic datarepositories

UniEuk: a pragmatic implementation

gene reference databases

taxonomytable

sequencetable

markertables

universaltaxonomicframework

existinggenetic datarepositories

UniEuk: a community-based effort

UniEuk webportal

Redmineenvironment

taxonomyimplementer

databaseimplementer

experts / curators

on a web protal, experts/curators have access to the latest version of the framework and databases

experts/curators use a redmine environment to provide feedback, flag issues, make suggestions of changes

the implementers use this feedback to improve the framework/databases and regularly update the web portal

UniEuk Steering Committee

Sina Adl, University of SaskatchewanGuy Cochrane, EMBL-EBIColomban de Vargas, Station Biologique de RoscoffFrank Oliver Glöckner, Max Planck Institute and Jakobs UniversityEunsoo Kim, American Museum of Natural HistoryLaura Wegener-Parfrey, University of British ColumbiaPelin Yilmaz, Max Planck Institute and Jakobs University