THE GENETIC DIVERSITY AND VIRULENCE CHARACTERISTICS OF AEROMONAS HYDROPHILA ISOLATED FROM FISHPONDS WITH DISEASE OUTBREAKS IN HUBEI PROVINCE

to investigate the genetic diversity and virulence characteristics, and to clarify the epidemic regularity of Aeromonas hydrophila which caused motile Aeromonas septicaemia outbreak (commonly known as disease outbreaks) in fish, 30 strains were isolated from 6 diseased fishponds which located in three different regions of Hubei province from 2006 to 2009. Among these strains, 20 were clinical strains (isolated from blood, liver, kidney or ascites fluid), 6 were intestinal strains and 4 were pond-water strains. A phylogenetic tree was constructed on the basis of the gyrB gene sequence of all 30 strains; the genetic typing of these strains were done through ERIC (enterobacterial repetitive intergenic consensus)-fingerprinting; the distribution patterns of seven virulence genes in these strains were tested by PCR method, including aerolysin (aerA), haemolysin (hlyA), heat-labile cytotonic enterotoxin (alt), heat-stable cytotonic enterotoxin (ast), elastase (ahpB), lipase (lip), and flagella (fla). In addition, the virulence of 15 representative strains was assayed in zebrafish by intraperitoneal injection. The results showed that regardless of the origin, all the 20 clinical strains, 1 intestinal strain and 3 pond-water strains had the same genetic characteristics. These included that clustered together in the phylogenetic tree, with 100% sequence similarities among these strains; have the same ERIC-finger-printing and the same virulence-gene pattern: aerA+hlyA+alt+ast+ahpB+lip+fla+; all of them were high virulent strains (LD50 9.74104　cfu/fish). Compared with clinical strains, the rest 5 intestinal strains and 1 pond-water strain either had different ERIC-fingerprintings or have different virulence-gene patterns, showing higher genetic diversity and lower virulence (LD501.01106　cfu/fish). These results indicated that as the pathogen of disease outbreaks, a clonal lineage of A. hydrophila was in popular within a certain time and areas, and no obvious variation or genetic diversity was found among these strains. This result would help in the clarification of the epidemic regularity of A. hydrophila underlying outbreaks, and help to make the right defense measures to prevent this infection. Meanwhile, the combined prevalence of multiple virulence genes in pathogenic strains of A. hydrophila laid the foundation for clarify the pathogenic mechanism underlie this infection. Moreover, the correlation of virulence and virulence-gene patterns of A. hydrophila showed that virulence genes can be chosen as gene-marker to detect pathogenic strains of this bacterium.