The prokaryotic and eukaryotic microbiome of Pacific oyster spat is shaped by ocean warming but not acidification.

Pacific oysters (Magallana gigas, a.k.a. Crassostrea gigas), the most widely farmed oysters, are under threat from climate change and emerging pathogens. In part, their resilience may be affected by their microbiome, which, in turn, may be influenced by ocean warming and acidification. To understand these impacts, we exposed early-development Pacific oyster spat to different temperatures (18°C and 24°C) and pCO2 levels (800, 1,600, and 2,800 µatm) in a fully crossed design for 3 weeks. Under all conditions, the microbiome changed over time, with a large decrease in the relative abundance of potentially pathogenic ciliates (Uronema marinum) in all treatments with time. The microbiome composition differed significantly with temperature, but not acidification, indicating that Pacific oyster spat microbiomes can be altered by ocean warming but is resilient to ocean acidification in our experiments. Microbial taxa differed in relative abundance with temperature, implying different adaptive strategies and ecological specializations among microorganisms. Additionally, a small proportion (~0.2% of the total taxa) of the relatively abundant microbial taxa were core constituents (>50% occurrence among samples) across different temperatures, pCO2 levels, or time. Some taxa, including A4b bacteria and members of the family Saprospiraceae in the phyla Chloroflexi (syn. Chloroflexota) and Bacteroidetes (syn. Bacteroidota), respectively, as well as protists in the genera Labyrinthula and Aplanochytrium in the class Labyrinthulomycetes, and Pseudoperkinsus tapetis in the class Ichthyosporea were core constituents across temperatures, pCO2 levels, and time, suggesting that they play an important, albeit unknown, role in maintaining the structural and functional stability of the Pacific oyster spat microbiome in response to ocean warming and acidification. These findings highlight the flexibility of the spat microbiome to environmental changes.IMPORTANCEPacific oysters are the most economically important and widely farmed species of oyster, and their production depends on healthy oyster spat. In turn, spat health and productivity are affected by the associated microbiota; yet, studies have not scrutinized the effects of temperature and pCO2 on the prokaryotic and eukaryotic microbiomes of spat. Here, we show that both the prokaryotic and, for the first time, eukaryotic microbiome of Pacific oyster spat are surprisingly resilient to changes in acidification, but sensitive to ocean warming. The findings have potential implications for oyster survival amid climate change and underscore the need to understand temperature and pCO2 effects on the microbiome and the cascading effects on oyster health and productivity.

Selective cell lysis pressure on rare and abundant prokaryotic taxa across a shelf-to-slope continuum in the Northern South China Sea.

IMPORTANCE: Virus-induced host lysis contributes up to 40% of total prokaryotic mortality and plays crucial roles in shaping microbial composition and diversity in the ocean. Nonetheless, what taxon-specific cell lysis is caused by viruses remains to be studied. The present study, therefore, examined the taxon-specific cell lysis and estimated its contribution to the variations in the rare and abundant microbial taxa. The results demonstrate that taxon-specific mortality differed in surface and bottom of the coastal environment. In addition, active rare taxa are more susceptible to heightened lytic pressure and suggested the importance of viral lysis in regulating the microbial community composition. These results improve our understanding of bottom-up (abiotic environmental variables) and top-down (viral lysis) controls contributing to microbial community assembly in the ocean.

Mortality by ribosomal sequencing (MoRS) provides a window into taxon-specific cell lysis.

Microbes are by far the dominant biomass in the world's oceans and drive biogeochemical cycles that are critical to life on Earth. The composition of marine microbial communities is highly dynamic, spatially and temporally, with consequent effects on their functional roles. In part, these changes in composition result from viral lysis, which is taxon-specific and estimated to account for about half of marine microbial mortality. Here, we show that extracellular ribosomal RNA (rRNAext) is produced by viral lysis, and that specific lysed populations can be identified by sequencing rRNAext recovered from seawater samples. In ten seawater samples collected at five depths between the surface and 265 m during and following a phytoplankton bloom, lysis was detected in about 15% of 16,946 prokaryotic taxa, identified from amplicon sequence variants (ASVs), with lysis occurring in up to 34% of taxa within a water sample. The ratio of rRNAext to cellular rRNA (rRNAcell) was used as an index of taxon-specific lysis, and revealed that higher relative lysis was most commonly associated with copiotrophic bacteria that were in relatively low abundance, such as those in the genera Escherichia and Shigella spp., as well as members of the Bacteriodetes; whereas, relatively low lysis was more common in taxa that are often relatively abundant, such as members of the Pelagibacterales (i.e., SAR11 clade), cyanobacteria in the genus Synechococcus, and members of the phylum Thaumarchaeota (synonym, Nitrososphaerota) that comprised about 13-15% of the 16 S rRNA gene sequences below 30 m. These results provide an explanation for the long-standing conundrum of why highly productive bacteria that are readily isolated from seawater are often in very low abundance. The ability to estimate taxon-specific cell lysis will help explore the distribution and abundance of microbial populations in nature.

Complete Genome Sequence of Vibrio natriegens Strain PWH3a.

Vibrio natriegens strain PWH3a, isolated from the Texas Gulf Coast, is used as a model organism in marine microbiology. Here, we report the complete genome sequence of strain PWH3a, which has two circular chromosomes, 4,650 coding sequences, 34 rRNA, 4 noncoding RNA (ncRNA), 131 tRNA genes, and one Mu-like prophage sequence.

Revealing the composition of the eukaryotic microbiome of oyster spat by CRISPR-Cas Selective Amplicon Sequencing (CCSAS).

BACKGROUND: The microbiome affects the health of plants and animals, including humans, and has many biological, ecological, and evolutionary consequences. Microbiome studies typically rely on sequencing ribosomal 16S RNA gene fragments, which serve as taxonomic markers for prokaryotic communities; however, for eukaryotic microbes this approach is compromised, because 18S rRNA gene sequences from microbial eukaryotes are swamped by contaminating host rRNA gene sequences. RESULTS: To overcome this problem, we developed CRISPR-Cas Selective Amplicon Sequencing (CCSAS), a high-resolution and efficient approach for characterizing eukaryotic microbiomes. CCSAS uses taxon-specific single-guide RNA (sgRNA) to direct Cas9 to cut 18S rRNA gene sequences of the host, while leaving protistan and fungal sequences intact. We validated the specificity of the sgRNA on ten model organisms and an artificially constructed (mock) community of nine protistan and fungal pathogens. The results showed that > 96.5% of host rRNA gene amplicons were cleaved, while 18S rRNA gene sequences from protists and fungi were unaffected. When used to assess the eukaryotic microbiome of oyster spat from a hatchery, CCSAS revealed a diverse community of eukaryotic microbes, typically with much less contamination from oyster 18S rRNA gene sequences than other methods using non-metazoan or blocking primers. However, each method revealed taxonomic groups that were not detected using the other methods, showing that a single approach is unlikely to uncover the entire eukaryotic microbiome in complex communities. To facilitate the application of CCSAS, we designed taxon-specific sgRNA for ~16,000 metazoan and plant taxa, making CCSAS widely available for characterizing eukaryotic microbiomes that have largely been neglected. CONCLUSION: CCSAS provides a high-through-put and cost-effective approach for resolving the eukaryotic microbiome of metazoa and plants with minimal contamination from host 18S rRNA gene sequences. Video Abstract.

A New Freshwater Cyanosiphovirus Harboring Integrase.

Pelagic cyanobacteria are key players in the functioning of aquatic ecosystems, and their viruses (cyanophages) potentially affect the abundance and composition of cyanobacterial communities. Yet, there are few well-described freshwater cyanophages relative to their marine counterparts, and in general, few cyanosiphoviruses (family Siphoviridae) have been characterized, limiting our understanding of the biology and the ecology of this prominent group of viruses. Here, we characterize S-LBS1, a freshwater siphovirus lytic to a phycoerythrin-rich Synechococcus isolate (Strain TCC793). S-LBS1 has a narrow host range, a burst size of ∼400 and a relatively long infecting step before cell lysis occurs. It has a dsDNA 34,641 bp genome with putative genes for structure, DNA packing, lysis, replication, host interactions, DNA repair and metabolism. S-LBS1 is similar in genome size, genome architecture, and gene content, to previously described marine siphoviruses also infecting PE-rich Synechococcus, e.g., S-CBS1 and S-CBS3. However, unlike other Synechococcus phages, S-LBS1 encodes an integrase, suggesting its ability to establish lysogenic relationships with its host. Sequence recruitment from viral metagenomic data showed that S-LBS1-like viruses are diversely present in a wide range of aquatic environments, emphasizing their potential importance in controlling and structuring Synechococcus populations. A comparative analysis with 16 available sequenced cyanosiphoviruses reveals the absence of core genes within the genomes, suggesting high degree of genetic variability in siphoviruses infecting cyanobacteria. It is likely that cyanosiphoviruses have evolved as distinct evolutionary lineages and that adaptive co-evolution occurred between these viruses and their hosts (i.e., Synechococcus, Prochlorococcus, Nodularia, and Acaryochloris), constituting an important driving force for such phage diversification.