General Patterns of Diversity in Major Marine Microeukaryote Lineages

Pernice, Massimo C.; Logares, Ramiro; Guillou, Laure; Massana, Ramón

doi:10.1371/journal.pone.0057170

Cited by 57 publications

(60 citation statements)

References 48 publications

(52 reference statements)

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“…The table was manually curated by removing OTUs represented by short sequences (o250 bp) and OTUs that occurred in only one sample, regardless of their abundance (Supplementary Figure S1). OTUs were then taxonomically assigned using three reference databases: SILVA 108 (Quast et al, 2013), PR 2 and MAS9013, an in-house database (Pernice et al, 2013; available on request). SILVA 108 was used to exclude prokaryotic 16S rDNA OTUs, whereas PR 2 and MAS9013 were used to classify eukaryotic OTUs mostly to the class level.…”

Section: Methodsmentioning

confidence: 99%

Large variability of bathypelagic microbial eukaryotic communities across the world’s oceans

et al. 2015

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In this work, we study the diversity of bathypelagic microbial eukaryotes (0.8-20 μm) in the global ocean. Seawater samples from 3000 to 4000 m depth from 27 stations in the Atlantic, Pacific and Indian Oceans were analyzed by pyrosequencing the V4 region of the 18S ribosomal DNA. The relative abundance of the most abundant operational taxonomic units agreed with the results of a parallel metagenomic analysis, suggesting limited PCR biases in the tag approach. Although rarefaction curves for single stations were seldom saturated, the global analysis of all sequences together suggested an adequate recovery of bathypelagic diversity. Community composition presented a large variability among samples, which was poorly explained by linear geographic distance. In fact, the similarity between communities was better explained by water mass composition (26% of the variability) and the ratio in cell abundance between prokaryotes and microbial eukaryotes (21%). Deep diversity appeared dominated by four taxonomic groups (Collodaria, Chrysophytes, Basidiomycota and MALV-II) appearing in different proportions in each sample. Novel diversity amounted to 1% of the pyrotags and was lower than expected. Our study represents an essential step in the investigation of bathypelagic microbial eukaryotes, indicating dominating taxonomic groups and suggesting idiosyncratic assemblages in distinct oceanic regions.

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Section: Methodsmentioning

confidence: 99%

Large variability of bathypelagic microbial eukaryotic communities across the world’s oceans

et al. 2015

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“…The final curated reads were clustered into operational taxonomic units (OTUs) by using UCLUST 1.2.22 (45), with similarity thresholds of 97% for V4 reads and 95% for V9 reads. Representative reads of each OTU were taxonomically classified by using BLAST against SILVA108, PR2, and a marine microeukaryote database (46). After the taxonomic assignment, metazoan OTUs were removed.…”

Section: Methodsmentioning

confidence: 99%

Environmental Sequencing Provides Reasonable Estimates of the Relative Abundance of Specific Picoeukaryotes

Giner

Forn

Romac

et al. 2016

Appl Environ Microbiol

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High-throughput sequencing (HTS) is revolutionizing environmental surveys of microbial diversity in the three domains of life by providing detailed information on which taxa are present in microbial assemblages. However, it is still unclear how the relative abundance of specific taxa gathered by HTS correlates with cell abundances. Here, we quantified the relative cell abundance of 6 picoeukaryotic taxa in 13 planktonic samples from 6 European coastal sites using epifluorescence microscopy on tyramide signal amplification-fluorescence in situ hybridization preparations. These relative abundance values were then compared with HTS data obtained in three separate molecular surveys: 454 sequencing of the V4 region of the 18S ribosomal DNA (rDNA) using DNA and RNA extracts (DNA-V4 and cDNA-V4) and Illumina sequencing of the V9 region (cDNA-V9). The microscopic and molecular signals were generally correlated, indicating that a relative increase in specific 18S rDNA was the result of a large proportion of cells in the given taxa. Despite these positive correlations, the slopes often deviated from 1, precluding a direct translation of sequences to cells. Our data highlighted clear differences depending on the nucleic acid template or the 18S rDNA region targeted. Thus, the molecular signal obtained using cDNA templates was always closer to relative cell abundances, while the V4 and V9 regions gave better results depending on the taxa. Our data support the quantitative use of HTS data but warn about considering it as a direct proxy of cell abundances. IMPORTANCEDirect studies on marine picoeukaryotes by epifluorescence microscopy are problematic due to the lack of morphological features and due to the limited number and poor resolution of specific phylogenetic probes used in fluorescence in situ hybridization (FISH) routines. As a consequence, there is an increasing use of molecular methods, including high-throughput sequencing (HTS), to study marine microbial diversity. HTS can provide a detailed picture of the taxa present in a community and can reveal diversity not evident using other methods, but it is still unclear what the meaning of the sequence abundance in a given taxon is. Our aim is to investigate the correspondence between the relative HTS signal and relative cell abundances in selected picoeukaryotic taxa. Environmental sequencing provides reasonable estimates of the relative abundance of specific taxa. Better results are obtained when using RNA extracts as the templates, while the region of 18S ribosomal DNA had different influences depending on the taxa assayed. Protists are key components of marine ecosystems, being major players in the global respiration and production budgets (1, 2) and playing central roles in marine food webs (3). Despite their importance and ubiquity, it was only during the past decade that environmental studies, based on molecular (i.e., culture-independent) techniques, revealed an unsuspected protist diversity in a large variety of marine ecosystems (4-13). These studies we...

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“…On the other hand, these high numbers may be due to missing intraspecific sequence variation in the PR 2 reference database, which usually contains only one accession per species. In natural communities, intraspecific genetic variation of microbial organisms may be much more diverse than just a few base pair differences, especially in hypervariable gene regions (Brown et al, 2015;Decelle et al, 2014;Dunthorn et al, 2012;Pernice et al, 2013 (2014) and Gimmler and Stoeck (2015).…”

Section: Contrasting Otu Results From Three Approachesmentioning

confidence: 99%

Comparison of three clustering approaches for detecting novel environmental microbial diversity

Forster

Dunthorn

Stoeck

et al. 2015

Preprint

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Discovery of novel diversity in high-throughput sequencing (HTS) studies is a central task in environmental microbial ecology. To evaluate the effects that amplicon clustering methods have on novel diversity discovery, we clustered an environmental marine protist HTS dataset of protist reads together with accessions from the taxonomically curated PR 2 reference database using three de novo approaches: sequence similarity networks, USEARCH, and Swarm. The novel diversity uncovered by each clustering approach differed drastically in the number of operational taxonomic units (OTUs) and the number of environmental amplicons in these novel diversity OTUs. Global pairwise alignment comparisons revealed that numerous amplicons classified as novel by USEARCH and Swarm were actually highly similar to reference accessions. Using graph theory we found additional novel diversity within OTUs that would have gone unnoticed without further using their underlying network topologies. Our results suggest that novel diversity inferred from clustering approaches requires further validation, whereas graph theory provides a powerful tool for microbial ecology and the analyses of environmental HTS datasets.

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General Patterns of Diversity in Major Marine Microeukaryote Lineages

Cited by 57 publications

References 48 publications

Large variability of bathypelagic microbial eukaryotic communities across the world’s oceans

Large variability of bathypelagic microbial eukaryotic communities across the world’s oceans

Environmental Sequencing Provides Reasonable Estimates of the Relative Abundance of Specific Picoeukaryotes

Comparison of three clustering approaches for detecting novel environmental microbial diversity

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