High-throughput isolation and ecogenomics of phages infecting fastidious marine heterotrophic bacteria

Marine Bacteria and their associated viruses are key players in shaping microbial community structures and global biogeochemical cycles. Cell lysis through viral predation is a crucial component for the recycling of carbon compounds and other nutrients, and interaction between viruses and hosts can also alter cellular functions via metabolic ‘hijacking’. The culture-based study of phototrophic cyanobacterial virus-host systems has revealed that these constant co29 evolutionary pressures in virus-host systems are further escalated by virally mediate horizontal gene transfer. Viruses of fastidious, heterotrophic bacteria are among the most abundant and ecologically significant virus-host systems in the oceans, but the dearth of cultured model systems has hampered the progression in testing hypotheses and interpretation of meta’omics data. The central goal of this thesis was to establish efficient and high-throughput methodologies for viral isolation, specifically targeting fastidious heterotrophic microbes. Using a novel isolate of the ubiquitous SAR11 clade as a model for highly abundant hosts, and three novel isolates of the methylotrophic OM43 clade for a lower-abundance host, I devised an optimised workflow based on established Dilution-to-Extinction culturing techniques that resulted in the isolation of over 117 viruses. The results show that among these novel viruses were the first known siphoviruses of the SAR11 clade and the first recorded viruses infecting members of the OM43 clade. Genomic evidence by the Methylophilales phage Melnitz indicated that inter-class host transitioning between streamlined heterotrophs may occur, and highlighted unusual viral features such as curli genes and glutamine riboswitches. Furthermore, utilising the power of metagenomics coupled with culture based host range experiments, the Pelagibacter phage Skadi was revealed as a highly abundant polar virus, and as an example for ecotypic niche specificity among the most abundant viruses on Earth. Culture-based identification of virus-host pairs combined with ecogenomic interpretation of metagenomic data will improve our understanding of ecological patterns and of viral strategies. The results from this thesis illuminated parts of the viral “dark matter” and highlighted possible novel viral strategies. This work can be adapted to many different systems, both of high and low abundance, and should be used to improve future viral isolation campaigns