Isolation of Vibrio cholerae 0139 from freshwater fish and its characterization vis-à-vis virulence genes, genetic heterogeneity and response to antimicrobials

Abstract

Vibrio cholerae belonging to the pathogenic O139 serogroup was isolated from freshwater fish, Catla catla procured from fish market in Visakhapatnam, Andhra Pradesh. The V. cholerae O139 isolate was identified by employing V. cholerae species specific PCR, PCR targeting O139-rfb specific primers and serologically using O139 antiserum. The V. cholerae O139 isolate was positive for the presence of virulence genes viz., cholera toxin, toxin co-regulated pilus and accessory cholera enterotoxin genes but was negative for zonula occludens toxin gene. The heterogeneity of the V. cholerae O139 isolate was compared with other cholera toxin gene positive V. cholerae isolates from fish and fishery environment. Biochemically, the O139 isolate was similar to other ctx positive V. cholerae but differed with some isolates in acetoin production. The V. cholerae O139 isolate haemolysed human RBC, was positive for gelatinase, caseinase, DNAse, lipase, phosphatase, amylase and chitinase activities. The lipase activity of V. cholerae O139 isolate was relatively higher than other ctx positive V. cholerae. The V. cholerae O139 isolate was resistant to nitrofurantoin, cefotaxime and ampicillin but was sensitive to 23 other antibiotics. Plant essential oils inhibited the growth of V. cholerae including O139 isolate in the order thyme oil greater than clove bud oil greater than clove leaf oil greater than rosemary oil. ERIC-PCR results showed that V. cholerae O139 isolate showed extensive dissimilarity with other ctx gene positive V. cholerae isolates that belonged to serogroup-O1 and serogroup non-O1/non-O139 but exhibited more similarity with V. cholerae O139 type culture.