A robust approach to estimate relative phytoplankton cell abundance from metagenomic data

Phytoplankton account for >45% of primary production on Earth, and hence have an enormous impact on aquatic food webs and on the entire Earth System. Their members are found across many of the major clades of the tree of life, including prokaryotes (cyanobacteria) and multiple eukaryotic lineages. Phytoplankton communities are generally studied by PCR amplification of rRNA marker genes from the DNA extracted from environmental samples and high-throughput sequencing of the amplicons. However, our appreciation of phytoplankton abundance or biomass is limited by PCR-amplification biases, rRNA gene copy number variations across taxa and the fact that rRNA do not provide insights into metabolic traits such as photosynthesis (i.e., it is sometimes difficult to distinguish an rRNA sequence of a phytoplanktonic species from a phylogenetically close non-photosynthetic species). In addition, rRNA marker genes fail to capture both cyanobacteria and photosynthetic eukaryotes simultaneously, thus hampering assessment of the entire phytoplankton community. Here, we propose to use the core single-copy photosynthetic gene psbO extracted from metagenomes to circumvent these limitations: the method is PCR-free, and the gene is present in both prokaryotes and eukaryotes, mainly in one copy per genome. We applied and validated this new strategy with the Tara Oceans datasets, and showed in most cases improved correlations with flow cytometry and microscopy than when based on rRNA genes. Furthermore, we revealed unexpected features of the ecology of these organisms, such as the high abundance of picocyanobacterial aggregates and symbionts in the ocean, and the decrease in relative abundance of phototrophs towards the larger size classes of marine communities of dinoflagellates. To facilitate the incorporation of psbO in future molecular-based surveys, we released a curated database including >20,000 unique sequences. Overall, psbO appears to be a promising new gene marker for improving molecular-based evaluations of phytoplankton communities.