Research On Profile And Diversity Of Vibrio Parahaemolyticus Isolated From Seafood In Shanghai

Vibrio parahaemolyticus is a intracellular pathogen responsible for seafood-borne outbreak in coastal cities.Many study reported that vibrio parahaemolyticus are most associated with consumption of raw seafood through across contamination。This study was to investigate the distribution of Vibrio parahaemolyticus isolated from seafood in Shanghai,effect on Caco-2 of non-pathogenic Vibrio parahaemolyticus and pathogenic Vibrio parahaemolyticus,and molecular typing technique to analyze the genetic diversity of pathogenic vibrio parahaemolyticus using REP-PCR and ERIC-PCR.1.Detection of Vibrio parahaemolyticus and virulence genes isolated from seafood in wharf and market of ShanghaiThe aim of this study was to investigate the profiles of V.parahaemolyticus in seafood from Shanghai in 2015-2016.186 suspected V.parahaemolyticus strains were isolated from 206 aquatic samples by rapid separable and detectable assays,including GB/T4789.2013,polymerase chain reaction(PCR)and loop-mediated isothermal amplification(LAMP).Suspected colonies were streaked onto CHROMagar Vibrio agar plate to determine the most probable number(MPN).DNA from the purple colonies was extracted for use as a template for PCR and LAMP detection.A total of 71 samples were collected from the sea wharf,59 of these samples were tested positive for V.parahaemolyticus with levels as high as 88.36 MPN/100 g.In contrast,the prevalence rate was 91.11% with levels as high as 1361.76MPN/100 g for samples collected in the marketplace.The prevalence rates of tdh+ and trh+ were 1.6%,1.0% respectively.These results showed that shrimp was a particular concern for total V.parahaemolyticus and V.parahaemolyticus(tdh+/trh+)contamination.Furthermore,the seafood marketplace has a higher contamination rate than natural environment.As shown in this study,the post-harvest practices increase the prevalence rate and level of V.parahaemolyticus in seafood.These results can be used to control outbreak of foodborne diseases in Shanghai.2.Effect on Caco-2 of Vibrio parahaemolyticus isolated from different sourceThe aim of this study was to investigated the pathogenic potential of Vibrio parahaemolyticus isolated from different source by studying their invasion ability and their cytotoxicity through infected Caco-2 cells.40 strains of V.parahaemolyticus isolated from seafood and patient were tested by two different experimental method.The invasion ability of pathogenic Vibrio parahaemolyticus was higher than non pathogenic Vibrio parahaemolyticus.The cytotoxicity of pathogenic Vibrio parahaemolyticushave was higher than non-pathogenic Vibrio parahaemolyticus too.Also data corresponding invasion ability and cytotoxicity of Vibrio parahaemolyticus isolated from shrimp and Shanghai Luchao wharf were highest.On the basis of data show that distinct virulence profiles and environmental Vibrio parahaemolyticus may represent a risk for human health.3.Prevalence,Molecular typing of Vibrio parahaemolyticus by PCR-based MethodsThe aim of this study was to use two PCR-based techniques for the analysis of genetic variability among V.parahaemolyticus virulence strains isolated from seafood in Shanghai from 2009 to 2016 and hospital patients.All isolates were analyzed for the presence of tdh and trh gene by PCR.40 strains(tdh+/trh+)of V.parahaemolyticus isolated from seafood and clinical patient were analyzed by repetitive extragenic palindromic PCR(REP-PCR)and renterobacterial repetitive intergenic consensus sequence PCR(ERIC-PCR).Results indicated that 40 strains of V.parahaemolyticus,representing a wide range of patterns,were grouped into 6 and 8 clusters at a similarity coefficient of 0.8 by REP-PCR,and ERIC-PCR,respectively.ERIC-PCR is preferable to REP-PCR because of the greater discriminative abilities of its fingerprints,while REP-PCR may be a practical method because it generates fewer amplification bands and patterns.Vibrio parahaemolyticus isolated from shrimp has genetic diversity.Shanghai Hengda aquaculture market has not only rich diversity but also dominant molecular typing.Thus,this study demonstrated the genetic variability of V.parahaemolyticus(tdh+ /trh+)in seafood,provided insights into the dissemination of virulence strains,and improved our knowledge of methods of microbiological risk assessment in seafood from market.