Freshwater Products In Pathogenic Vibrio Flora Composition And Main Flora Analysis

Vibrios are very important food-borne pathogen and widely distributed in aquatic products, which are also pathogens in aquatic animals,causing gastroenteritis diseases, wound infection, sepsis and other diseases. Chinese food-borne monitoring network data show that food-borne bacterial disease caused by Vibrio.spp were clearly increased in recent years. At the same time, with the immigration ships and cargo throughput increased year by year, the potential threat to human body caused by Vibrio is also increasing. Therefore, understanding the Vibrio flora distribution in aquatic products, establishing a rapid detection method of Vibrios is of great significance to ensure the safety of food and aquaculture. The methods used in present study and results we obtained are descripted as follows:1、A total of55aquactic samples were collected from different sources of fresh water products in partial regions of Jiangsu Province. After samples processing, we obtained233isolates by double TCBS plate separation and purification. Through the identification of biochemical tests, several kinds of Vibrio species were isolated, e.g. Vibrio parahaemolyticus、Vibrio alginolyticus、Vibrio harveyi、Vibrio aestuarianus、 Vibrio metschnikovii、Vibrio azureus、Vibrio natriegens、Vibrio diazotrophicus and Vibrio owensii, in which, Vibrio parahaemolyticus and Vibrio alginolyticus were major species. Geographically, the detection rates of Vibrios in Lianyungang was higher than Yangzhou、Changzhou and Taizhou.From the sample types, the detection rates of vibrios in fish was higher than shellfish、crabs and shrimp, the detection rates of vibrios in shellfish was lower.2、V.parahaemolyticus and V.alginolyticus which were founded to be the mainly distributed flora were genotyped by RAPD fingerprinting analysis. We obtained18different genotypes in V.parahaemolyticus.These genotypes were clustered into9(from A to I) genetic lineages using UPGMA clustering method, in which A and C were the main genotypes. Analysis of the V.parahaemolyticus molecular types distribution in different regions and samples showed that type A and C were detected in all sampling areas and samples category, Lianyungang originated isolates had a richer fingerprint patterns and the fingerprint patterns of isolates from fish and shrimp were similar. We obtained21different genotypes in V.alginolyticus, these genotypes were clustered into12(from A to L) genetic lineages using UPGMA clustering method, in which A、C and K were the mainly types. The distribution analysis of V.alginolyticus types among regions and samples showed that type A and K were detected in all sampling areas and samples category. Isolates from Lianyungang had a richer fingerprint, and the fingerprint distribution were similar between Yangzhou and Taizhou. There were rich fingerprint pattens in isolates from fish、shrimp and crabs.3、The specific PCR amplification of thermostable direct hemolysin gene (tdh) and thermostable direct hemolysin-related toxin gene (trh) in the84V.parahaemolyticus isolates, although they were all positive in the V.parahaemolyticus species-specific gene(tlh). However, the results of kanagawa test for all stains showed that12strains of V.parahaemolyticus and13strains of V.alginolyticus had significant hemolysis circle, and they were all the main genotypes in strains. The cell adhesion and intestinal fluid accumulation (FA) tests were carried out in8hemolytic Vibrio isolates which were main type, and it was found that8vibrio isolates had certain adhesion ability with spleen cells, the adhesion capacity of the strain92was greater than the positive control strains. All of8vibrio isolates could cause certain level of intestinal fluid accumulation, in which the strain71、92、125and214had higher levels of intestinal fluid accumulation, which suggested these isolates may be pathogenic and the potential biological risk factors to human.