Detection of Virulence-Associated Genes andGene Transfer Fromsp. Isolated From Aquatic Environments of Sub-himalayan West Bengal.

Abstract

is omnipresent in aquatic environments and cause disease within a wide host range. A total of thirty-four isolates from water samples of small fish farms were identified asbased on biochemical characteristics and 16S rRNA gene sequence. A total of six virulent factors were analyzed which indicated 100% of isolates as beta-haemolytic and proteolytic, whereas 44.1, 38.2, and 70.6% of isolates produced DNAse, siderophore, and amylase, respectively. Studies on the occurrence of four genetic determinants of virulence factors revealed that(haemolytic toxin) and(polar flagella) genes were present in 44.1% of strains whereas(type 3 secretion system) and(serine protease) genes were detected in 21.5 and 8.82% of strains, respectively. Fish () challenge studies showed that the isolate GP3 () bearing five virulent factors with the combination ofgenes induced severe lesions leading to 100% of mortality. In contrast, RB7 possessing four virulence factors and three genes () could not produce severe lesions and any mortality indicating the absence of correlation between the virulence factors, its genes, and the pathogenicity in fishes. GP3 was cytotoxic to human liver cell line (WRL-68) in trypan blue dye exclusion assay. The 431 bpgene of GP3 was transferable toDh5α with a conjugational efficiency of 0.394 × 10transconjugants per recipient cell. The transfer was confirmed by PCR and by the presence of 23-kb plasmids in both donor and transconjugants. Therefore, the occurrence of mobile genetic elements bearing virulence-associated genes inindicates the need for periodic monitoring of the aquatic habitat to prevent disease outbreaks.