In vivo gene expression of cold shock and other stress-related genes in Vibrio vulnificus during shellstock temperature control conditions in oysters

Aims: To determine Vibrio vulnificus response to shellstock refrigeration conditions while the bacterium was embedded in oysters (in vivo). Methods and Results: Depurated oysters were artificially inoculated with V. vulnificus. Several cold-shock conditions were examined according to the National Sanitation Shellfish Program guidelines. Culturability of cells along the refrigeration period was measured using specific colony dot-blot. Gene expression of putative cold-shock genes (csp1, csp3, csp4 and csp5) as well as three stress-related genes (rpoS, oxyR and katG) was determined by reverse transcriptase polymerase chain reaction. Vibrio vulnificus exhibited a decline in cell viability after cold shock but a cold adaptation response was observed when oysters were kept at suboptimal (15°C) temperatures. 16SrRNA, and rpoS genes were constitutively expressed while expression of csp genes varied among strains and time points. Conclusions: Vibrio vulnificus culturability was reduced after oysters were subjected to shellstock refrigeration conditions. When V. vulnificus was allowed to acclimate to cold temperatures, its survival after cold shock was higher. None of the cold shock genes analysed behaved as csp type I genes. Significance and Impact of the Study: A model for artificially inoculated specific strains of V. vulnificus into oysters has been established. For the first time, V. vulnficus gene expression was assessed with the pathogen embedded in oysters.