Single cell genomics yields a wide diversity of small planktonic protists across major ocean ecosystems

M E Sieracki,R Massana,R Stepanauskas,N J Poulton,R Logares,O Jaillon,L Rubinat-Ripoll,C De Vargas,P Wincker

doi:10.1038/s41598-019-42487-1

Abstract

Marine planktonic protists are critical components of ocean ecosystems and are highly diverse. Molecular sequencing methods are being used to describe this diversity and reveal new associations and metabolisms that are important to how these ecosystems function. We describe here the use of the single cell genomics approach to sample and interrogate the diversity of the smaller (pico- and nano-sized) protists from a range of oceanic samples. We created over 900 single amplified genomes (SAGs) from 8 Tara Ocean samples across the Indian Ocean and the Mediterranean Sea. We show that flow cytometric sorting of single cells effectively distinguishes plastidic and aplastidic cell types that agree with our understanding of protist phylogeny. Yields of genomic DNA with PCR-identifiable 18S rRNA gene sequence from single cells was low (15% of aplastidic cell sorts, and 7% of plastidic sorts) and tests with alternate primers and comparisons to metabarcoding did not reveal phylogenetic bias in the major protist groups. There was little evidence of significant bias against or in favor of any phylogenetic group expected or known to be present. The four open ocean stations in the Indian Ocean had similar communities, despite ranging from 14°N to 20°S latitude, and they differed from the Mediterranean station. Single cell genomics of protists suggests that the taxonomic diversity of the dominant taxa found in only several hundreds of microliters of surface seawater is similar to that found in molecular surveys where liters of sample are filtered.

Highlights

Planktonic protists in the surface ocean are ubiquitous, abundant and highly diverse
The single cell approach has proven its power in the discovery of new potential metabolisms in uncultured prokaryotes[13], and has the advantage of yielding large amounts of genomic DNA from individual microorganisms for further sequencing and investigation
Chlorophyll fluorescence was preserved in the plastidic cells for discrimination by flow cytometry (Supplementary Fig. S2)

Summary

Introduction

Planktonic protists in the surface ocean are ubiquitous, abundant and highly diverse. Genetic methods have revealed remarkably diverse ocean planktonic protist communities[4] These methods include direct cloning of environmental DNA, fingerprinting methods, tag sequencing, and metagenomics of filtered or sorted fractions of the community. For assessing the diversity of the dominant forms present in seawater, clone libraries and tag sequencing have been the favored approaches These methods have the disadvantage of being biased in favor of particular, often larger, cell types, which can have 10’s to 100’s www.nature.com/scientificreports/. Fine plankton nets and filter fractionation is often used to characterize plankton communities, but these can break up fragile animals, colonies, and individual cells, sending their DNA into small size fractions[8] It has been known for some time that many marine protists are mixotrophic and are not assigned to photo- or heterotrophic categories[9,10]. Sequencing of three “picobiliphyte” ( Picozoa) SAGs from that sample showed how this approach can reveal microbial interactions between eukaryotes, prokaryotes and viruses[14]

Methods

Results

Conclusion