Phase variation, capsular polysaccharide, pilus and flagella contribute to uptake ofVibrio vulnificusby the Eastern oyster (Crassostrea virginica)

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SummaryVibrio vulnificusinfections are associated with raw oyster consumption, and disease reservoirs are determined by the ability of this bacterium to infect and persist in oysters. Surface structures, such as capsular polysaccharide (CPS), pili and flagella, function as virulence factors in mouse infection models. Furthermore, virulence is related to phase variation in colony morphology, which reflects CPS expression and includes opaque (encapsulated, virulent), translucent (reduced encapsulation, avirulent) and rugose (wrinkled, biofilm-enhanced) colony types. The role of these factors in environmental survival is unknown; therefore, mutational analysis and phase variation ofV. vulnificuswere examined in an oyster infection model. Oysters (Crassostrea virginica) were pre-treated with tetracycline to reduce background bacteria and subsequently inoculated via filter feeding with 106 colony-forming units (cfu) ml−1 ofV. vulnificuswild-type strains and phase variants, as well as strains with deletion mutations in genes related to CPS (Δwza), pili (ΔpilA), flagella (ΔflaCDE/ΔflaFBA) and motility (ΔmotAB). All mutants were significantly reduced in their dissemination to oyster haemolymph as compared with wild type; however, recovery of mutants from gills and intestinal tissue was generally similar to wild type. Translucent and rugose inocula showed induction of high-frequency phase variation to the opaque encapsulated phenotype (100% and 72% respectively) during oyster infections that did not occur in strains recovered from seawater. Thus, multiple bacterial factors determine uptake ofV. vulnificusin oysters, and phase variation during oyster infection is a likely mechanism for environmental survival and for induction of the more virulent phenotype.

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