Comparative 16SrDNA Gene-Based Microbiota Profiles of the Pacific Oyster (Crassostrea gigas) and the Mediterranean Mussel (Mytilus galloprovincialis) from a Shellfish Farm (Ligurian Sea, Italy).

The pacific oyster Crassostrea gigas and the Mediterranean mussel Mytilus galloprovincialis are two widely farmed bivalve species which show contrasting behaviour in relation to microbial diseases, with C. gigas being more susceptible and M. galloprovincialis being generally resistant. In a recent study, we showed that different susceptibility to infection exhibited by these two bivalve species may depend on their different capability to kill invading pathogens (e.g., Vibrio spp.) through the action of haemolymph components. Specific microbial-host interactions may also impact bivalve microbiome structure and further influence susceptibility/resistance to microbial diseases. To further investigate this concept, a comparative study of haemolymph and digestive gland 16SrDNA gene-based bacterial microbiota profiles in C. gigas and M. galloprovincialis co-cultivated at the same aquaculture site was carried out using pyrosequencing. Bacterial communities associated with bivalve tissues (hemolymph and digestive gland) were significantly different from those of seawater, and were dominated by relatively few genera such as Vibrio and Pseudoalteromonas. In general, Vibrio accounted for a larger fraction of the microbiota in C. gigas (on average 1.7-fold in the haemolymph) compared to M. galloprovincialis, suggesting that C. gigas may provide better conditions for survival for these bacteria, including potential pathogenic species such as V. aestuarianus. Vibrios appeared to be important members of C. gigas and M. galloprovincialis microbiota and might play a contrasting role in health and disease of bivalve species. Accordingly, microbiome analyses performed on bivalve specimens subjected to commercial depuration highlighted the ineffectiveness of such practice in removing Vibrio species from bivalve tissues.

Whole-Genome Enrichment Provides Deep Insights into Vibrio cholerae Metagenome from an African River.

The detection and typing of Vibrio cholerae in natural aquatic environments encounter major methodological challenges related to the fact that the bacterium is often present in environmental matrices at very low abundance in nonculturable state. This study applied, for the first time to our knowledge, a whole-genome enrichment (WGE) and next-generation sequencing (NGS) approach for direct genotyping and metagenomic analysis of low abundant V. cholerae DNA (<50 genome unit/L) from natural water collected in the Morogoro river (Tanzania). The protocol is based on the use of biotinylated RNA baits for target enrichment of V. cholerae metagenomic DNA via hybridization. An enriched V. cholerae metagenome library was generated and sequenced on an Illumina MiSeq platform. Up to 1.8 × 107 bp (4.5× mean read depth) were found to map against V. cholerae reference genome sequences representing an increase of about 2500 times in target DNA coverage compared to theoretical calculations of performance for shotgun metagenomics. Analysis of metagenomic data revealed the presence of several V. cholerae virulence and virulence associated genes in river water including major virulence regions (e.g. CTX prophage and Vibrio pathogenicity island-1) and genetic markers of epidemic strains (e.g. O1-antigen biosynthesis gene cluster) that were not detectable by standard culture and molecular techniques. Overall, besides providing a powerful tool for direct genotyping of V. cholerae in complex environmental matrices, this study provides a 'proof of concept' on the methodological gap that might currently preclude a more comprehensive understanding of toxigenic V. cholerae emergence from natural aquatic environments.