Abstract
Bacteriophages have immense potential as antibiotic therapies and in genetic engineering. Understanding the mechanisms that bacteriophages implement to infect their hosts will allow researchers to manipulate these systems and adapt them to specific bacterial targets. In this study, we isolated a bacteriophage capable of infecting the marine alphaproteobacterium Phaeobacter inhibens and determined its mechanism of infection. Phaeobacter virus MD18, a novel species of bacteriophage isolated in Woods Hole, MA, exhibits potent lytic ability against P. inhibens and appears to be of the Siphoviridae morphotype. The genomic sequence of MD18 displayed significant similarity to another siphophage, the recently discovered Roseobacter phage DSS3P8, but genomic and phylogenetic analyses, assessing host range and a search of available metagenomes are all consistent with the conclusion that Phaeobacter phage MD18 is a novel lytic phage. We incubated MD18 with a library of barcoded P. inhibens transposon insertion mutants and identified 22 genes that appear to be required for phage predation of this host. Network analysis of these genes using genomic position, Gene Ontology (GO) term enrichment, and protein associations revealed that these genes are enriched for roles in assembly of a type IV pilus (T4P) and regulators of cellular morphology. Our results suggest that T4P serve as receptors for a novel marine virus that targets P. inhibens.IMPORTANCE Bacteriophages are useful nonantibiotic therapeutics for bacterial infections as well as threats to industries utilizing bacterial agents. This study identified Phaeobacter virus MD18, a phage antagonist of Phaeobacter inhibens, a bacterium with promising use as a probiotic for aquatic farming industries. Genomic analysis suggested that Phaeobacter phage MD18 has evolved to enhance its replication in P. inhibens by adopting favorable tRNA genes as well as through genomic sequence adaptation to resemble host codon usage. Lastly, a high-throughput analysis of P. inhibens transposon insertion mutants identified genes that modulate host susceptibility to phage MD18 and implicated the type IV pilus as the likely receptor recognized for adsorption. This study marks the first characterization of the relationship between P. inhibens and an environmentally sampled phage, which informs our understanding of natural threats to the bacterium and may promote the development of novel phage technologies for genetic manipulation of this host.
Highlights
Viruses are the largest source of genetic diversity known
We identified a novel lytic bacteriophage, Phaeobacter phage MD18, that infects P. inhibens
We identified genetic determinants to MD18 infection in P. inhibens using BarSeq
Summary
Viruses are the largest source of genetic diversity known As such, they have given rise to an incredible assortment of potential therapeutic and genetic tools used as sustainable substitutes for antibiotics[1, 2], targeted bacterial delivery systems[3], and for bacterial engineering[4]. P. inhibens is most known for the production of the antibiotic tropodithietic acid (TDA), which protects its natural algal symbiont from marine pathogens[8, 9]. Due to this property, P. inhibens serves as a useful probiotic in oyster and other aquatic farms to prevent colonization by pathogenic Vibrio species[10, 11]. Characterizing phage-host interactions within this industrially relevant host will enable researchers to implement phage engineering approaches for this species and provide insight into potential natural threats of P. inhibens
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