Choanoflagellates alongside diverse uncultured predatory protists consume the abundant open-ocean cyanobacterium Prochlorococcus.

Prochlorococcus is a key member of open-ocean primary producer communities. Despite its importance, little is known about the predators that consume this cyanobacterium and make its biomass available to higher trophic levels. We identify potential predators along a gradient wherein Prochlorococcus abundance increased from near detection limits (coastal California) to >200,000 cells mL-1 (subtropical North Pacific Gyre). A replicated RNA-Stable Isotope Probing experiment involving the in situ community, and labeled Prochlorococcus as prey, revealed choanoflagellates as the most active predators of Prochlorococcus, alongside a radiolarian, chrysophytes, dictyochophytes, and specific MAST lineages. These predators were not appropriately highlighted in multiyear conventional 18S rRNA gene amplicon surveys where dinoflagellates and other taxa had highest relative amplicon abundances across the gradient. In identifying direct consumers of Prochlorococcus, we reveal food-web linkages of individual protistan taxa and resolve routes of carbon transfer from the base of marine food webs.

Quantification of Polyphosphate in Environmental Planktonic Samples Using a Novel Fluorescence Dye JC-D7.

Polyphosphate (polyP) is found in plankton of diverse aquatic ecosystems and is important for plankton ecology and biogeochemical cycling. However, our knowledge of polyP in aquatic environments is hindered by a lack of data due to the limitations of quantification methods. The estimate of polyP in model organisms using phenol-chloroform extraction followed by enzymatic hydrolysis is complicated and fails for environmental samples. The commonly used 4',6-diamidino-2-phenylindole (DAPI) fluorescence method for environmental studies, on the contrary, severely overestimates polyP due to interference. In this paper, we develop a plankton lysis buffer to extract polyP and a quantification method using a novel polyP-specific fluorescence dye JC-D7. We test the methods using cultured algae and bacteria, as well as natural samples from marine and freshwater environments. We show that our plankton lysis extracts polyP with high recovery while requiring substantially less time and effort. Subsequent polyP quantification using JC-D7 fluorescence overcomes the interference encountered by the DAPI method and provides an accurate measurement of polyP down to <0.5 µmol L-1. This novel method enables more accurate quantification of polyP in aquatic environments and will profoundly enhance our knowledge of polyP, plankton ecology, and biogeochemistry.

Characterization of chasmoendolithic community in Miers Valley, McMurdo Dry Valleys, Antarctica.

The Antarctic Dry Valleys are unable to support higher plant and animal life and so microbial communities dominate biotic ecosystem processes. Soil communities are well characterized, but rocky surfaces have also emerged as a significant microbial habitat. Here, we identify extensive colonization of weathered granite on a landscape scale by chasmoendolithic microbial communities. A transect across north-facing and south-facing slopes plus valley floor moraines revealed 30-100 % of available substrate was colonized up to an altitude of 800 m. Communities were assessed at a multidomain level and were clearly distinct from those in surrounding soils and other rock-inhabiting cryptoendolithic and hypolithic communities. All colonized rocks were dominated by the cyanobacterial genus Leptolyngbya (Oscillatoriales), with heterotrophic bacteria, archaea, algae, and fungi also identified. Striking patterns in community distribution were evident with regard to microclimate as determined by aspect. Notably, a shift in cyanobacterial assemblages from Chroococcidiopsis-like phylotypes (Pleurocapsales) on colder-drier slopes, to Synechococcus-like phylotypes (Chroococcales) on warmer-wetter slopes. Greater relative abundance of known desiccation-tolerant bacterial taxa occurred on colder-drier slopes. Archaeal phylotypes indicated halotolerant taxa and also taxa possibly derived from nearby volcanic sources. Among the eukaryotes, the lichen photobiont Trebouxia (Chlorophyta) was ubiquitous, but known lichen-forming fungi were not recovered. Instead, fungal assemblages were dominated by ascomycetous yeasts. We conclude that chasmoendoliths likely constitute a significant geobiological phenomenon at lower elevations in granite-dominated Antarctic Dry Valley systems.

Hidden genomic diversity drives niche partitioning in a cosmopolitan eukaryotic picophytoplankton.

Marine eukaryotic phytoplankton are fundamental to the marine food web, yet the lack of reference genomes or just a single genome representing a taxon has led to an underestimation of their taxonomic, adaptive, and functional diversity. Here, we integrated strain isolation with metagenomic binning to recover genomes from the cosmopolitan picophytoplankton genus Bathycoccus, traditionally considered monospecific. Our recovery and analysis of 37 Bathycoccus genomes delineated their global genomic diversity and established four evolutionary clades (BI, BII, BIII, BIV). Our metagenomic abundance survey revealed well-differentiated ecological niches and distinct biogeographic distributions for each clade, predominantly shaped by temperature, salinity, and nutrient availability. Comparative genomics analyses further revealed clade-specific genomic traits that underpin niche adaptation and contribute to the global prevalence of Bathycoccus. Our findings underscore temperature as a major driver of genome diversification in this genus, with clade divergences coinciding with major paleoclimatic events that influenced their contemporary thermal niches. Moreover, the unique enrichment of C2H2 zinc finger and ankyrin repeat gene families in polar-adapted clades suggests previously unrecognized cold-adaptation mechanisms in marine eukaryotic phytoplankton. Our study offers a comprehensive genomic landscape of this crucial eukaryotic picophytoplankton, providing insights into their microdiversity and adaptive evolution in response to changing environments.

Gradients of bacteria in the oceanic water column reveal finely-resolved vertical distributions.

Bacterial communities directly influence ecological processes in the ocean, and depth has a major influence due to the changeover in primary energy sources between the sunlit photic zone and dark ocean. Here, we examine the abundance and diversity of bacteria in Monterey Bay depth profiles collected from the surface to just above the sediments (e.g., 2000 m). Bacterial abundance in these Pacific Ocean samples decreased by >1 order of magnitude, from 1.22 ±0.69 ×106 cells ml-1 in the variable photic zone to 1.44 ± 0.25 ×105 and 6.71 ± 1.23 ×104 cells ml-1 in the mesopelagic and bathypelagic, respectively. V1-V2 16S rRNA gene profiling showed diversity increased sharply between the photic and mesopelagic zones. Weighted Gene Correlation Network Analysis clustered co-occurring bacterial amplicon sequence variants (ASVs) into seven subnetwork modules, of which five strongly correlated with depth-related factors. Within surface-associated modules there was a clear distinction between a 'copiotrophic' module, correlating with chlorophyll and dominated by e.g., Flavobacteriales and Rhodobacteraceae, and an 'oligotrophic' module dominated by diverse Oceanospirillales (such as uncultured JL-ETNP-Y6, SAR86) and Pelagibacterales. Phylogenetic reconstructions of Pelagibacterales and SAR324 using full-length 16S rRNA gene data revealed several additional subclades, expanding known microdiversity within these abundant lineages, including new Pelagibacterales subclades Ia.B, Id, and IIc, which comprised 4-10% of amplicons depending on the subclade and depth zone. SAR324 and Oceanospirillales dominated in the mesopelagic, with SAR324 clade II exhibiting its highest relative abundances (17±4%) in the lower mesopelagic (300-750 m). The two newly-identified SAR324 clades showed highest relative abundances in the photic zone (clade III), while clade IV was extremely low in relative abundance, but present across dark ocean depths. Hierarchical clustering placed microbial communities from 900 m samples with those from the bathypelagic, where Marinimicrobia was distinctively relatively abundant. The patterns resolved herein, through high resolution and statistical replication, establish baselines for marine bacterial abundance and taxonomic distributions across the Monterey Bay water column, against which future change can be assessed.

Revealing the intricate temporal dynamics and adaptive responses of prokaryotic and eukaryotic microbes in the coastal South China Sea.

This comprehensive two-year investigation in the coastal South China Sea has advanced our understanding of marine microbes at both community and genomic levels. By combining metagenomics and metatranscriptomics, we have revealed the intricate temporal dynamics and remarkable adaptability of microbial communities and phytoplankton metagenome-assembled genomes (MAGs) in response to environmental fluctuations. We observed distinct seasonal shifts in microbial community composition and function: cyanobacteria were predominant during warmer months, whereas photosynthetic protists were more abundant during colder seasons. Notably, metabolic marker KOs of photosynthesis were consistently active throughout the year, underscoring the persistent role of these processes irrespective of seasonal changes. Our analysis reveals that environmental parameters such as temperature, salinity, and nitrate concentrations profoundly influence microbial community composition, while temperature and silicate have emerged as crucial factors shaping their functional traits. Through the recovery and analysis of 37 phytoplankton MAGs, encompassing nine prokaryotic cyanobacteria and 28 eukaryotic protists from diverse phyla, we have gained insights into their genetic diversity and metabolic capabilities. Distinct profiles of photosynthesis-related pathways including carbon fixation, carotenoid biosynthesis, photosynthesis-antenna proteins, and photosynthesis among the MAGs indicated their genetic adaptations to changing environmental conditions. This study not only enhances our understanding of microbial dynamics in coastal marine ecosystems but also sheds light on the ecological roles and adaptive responses of different microbial groups to environmental changes.

Unveiling Prasinovirus diversity and host specificity through targeted enrichment in the South China Sea.

Unicellular green picophytoplankton from the Mamiellales order are pervasive in marine ecosystems and susceptible to infections by prasinoviruses, large double-stranded DNA viruses within the Nucleocytoviricota phylum. We developed a double-stranded DNA virus enrichment and shotgun sequencing method, and successfully assembled 80 prasinovirus genomes from 43 samples in the South China Sea. Our research delivered the first direct estimation of 94% accuracy in correlating genome similarity to host range. Stirkingly, our analyses uncovered unexpected host-switching across diverse algal lineages, challenging the existing paradigms of host-virus co-speciation and revealing the dynamic nature of viral evolution. We also detected six instances of horizontal gene transfer between prasinoviruses and their hosts, including a novel alternative oxidase. Additionally, diversifying selection on a major capsid protein suggests an ongoing co-evolutionary arms race. These insights not only expand our understanding of prasinovirus genomic diversity but also highlight the intricate evolutionary mechanisms driving their ecological success and shaping broader virus-host interactions in marine environments.

The microbiome of a bacterivorous marine choanoflagellate contains a resource-demanding obligate bacterial associate.

Microbial predators such as choanoflagellates are key players in ocean food webs. Choanoflagellates, which are the closest unicellular relatives of animals, consume bacteria and also exhibit marked biological transitions triggered by bacterial compounds, yet their native microbiomes remain uncharacterized. Here we report the discovery of a ubiquitous, uncultured bacterial lineage we name Candidatus Comchoanobacterales ord. nov., related to the human pathogen Coxiella and physically associated with the uncultured marine choanoflagellate Bicosta minor. We analyse complete 'Comchoano' genomes acquired after sorting single Bicosta cells, finding signatures of obligate host-dependence, including reduction of pathways encoding glycolysis, membrane components, amino acids and B-vitamins. Comchoano encode the necessary apparatus to import energy and other compounds from the host, proteins for host-cell associations and a type IV secretion system closest to Coxiella's that is expressed in Pacific Ocean metatranscriptomes. Interactions between choanoflagellates and their microbiota could reshape the direction of energy and resource flow attributed to microbial predators, adding complexity and nuance to marine food webs.

Viruses infecting a warm water picoeukaryote shed light on spatial co-occurrence dynamics of marine viruses and their hosts.

The marine picoeukaryote Bathycoccus prasinos has been considered a cosmopolitan alga, although recent studies indicate two ecotypes exist, Clade BI (B. prasinos) and Clade BII. Viruses that infect Bathycoccus Clade BI are known (BpVs), but not that infect BII. We isolated three dsDNA prasinoviruses from the Sargasso Sea against Clade BII isolate RCC716. The BII-Vs do not infect BI, and two (BII-V2 and BII-V3) have larger genomes (~210 kb) than BI-Viruses and BII-V1. BII-Vs share ~90% of their proteins, and between 65% to 83% of their proteins with sequenced BpVs. Phylogenomic reconstructions and PolB analyses establish close-relatedness of BII-V2 and BII-V3, yet BII-V2 has 10-fold higher infectivity and induces greater mortality on host isolate RCC716. BII-V1 is more distant, has a shorter latent period, and infects both available BII isolates, RCC716 and RCC715, while BII-V2 and BII-V3 do not exhibit productive infection of the latter in our experiments. Global metagenome analyses show Clade BI and BII algal relative abundances correlate positively with their respective viruses. The distributions delineate BI/BpVs as occupying lower temperature mesotrophic and coastal systems, whereas BII/BII-Vs occupy warmer temperature, higher salinity ecosystems. Accordingly, with molecular diagnostic support, we name Clade BII Bathycoccus calidus sp. nov. and propose that molecular diversity within this new species likely connects to the differentiated host-virus dynamics observed in our time course experiments. Overall, the tightly linked biogeography of Bathycoccus host and virus clades observed herein supports species-level host specificity, with strain-level variations in infection parameters.

The need to account for cell biology in characterizing predatory mixotrophs in aquatic environments.

Photosynthesis in eukaryotes first arose through phagocytotic processes wherein an engulfed cyanobacterium was not digested, but instead became a permanent organelle. Other photosynthetic lineages then arose when eukaryotic cells engulfed other already photosynthetic eukaryotic cells. Some of the resulting lineages subsequently lost their ability for phagocytosis, while many others maintained the ability to do both processes. These mixotrophic taxa have more complicated ecological roles, in that they are both primary producers and consumers that can shift more towards producing the organic matter that forms the base of aquatic food chains, or towards respiring and releasing CO2. We still have much to learn about which taxa are predatory mixotrophs as well as about the physiological consequences of this lifestyle, in part, because much of the diversity of unicellular eukaryotes in aquatic ecosystems remains uncultured. Here, we discuss existing methods for studying predatory mixotrophs, their individual biases, and how single-cell approaches can enhance knowledge of these important taxa. The question remains what the gold standard should be for assigning a mixotrophic status to ill-characterized or uncultured taxa-a status that dictates how organisms are incorporated into carbon cycle models and how their ecosystem roles may shift in future lakes and oceans. This article is part of a discussion meeting issue 'Single cell ecology'.