Rapid detection of V. cholerae 01

Abstract

Monoclonal antibodies directed against group specific antigen A of Vibio cholerae 01 were obtained through hybridoma technology which involved fusions of Sp 2/0 myeloma cells with splenocytes of Balb/c mice immunized with whole cell lysates of V. cholerae 01. Specificity and crossreactivity of the monoclonal antibodies were determined against whole cell lysates prepared from 38 strains of V. cholerae 01, six strains of V. cholerae non-01, five strains of other vibrios, 45 strains of enterobacteria and Entamoeba histolytica by indirect and dot-blot enzyme-linked immunosorbent assay (ELISA). The monoclonal antibodies (MAb) reacted specifically to the whole cell lysates of the V. cholerae serogroup 01 and did not react to the antigens prepared from other organisms. The MAb were used in a dot-blot ELISA for detecting V. cholerae 01 antigen in 335 seafood specimens and in stools and rectal swabs of patients with acute watery diarrhoea. The dotblot ELISA performed on rectal swab specimens enriched in alkaline peptone water gave 96.0% sensitivity, 99.3% specificity, 94.7% positive predictive value, 99.5% negative predictive value and 98.9% efficacy, respectively, when compared to the conventional culture method. The two methods showed excellent agreement beyond chance, on the basis of a k coefficient value of 94.7%. No V. cholerae 01 was isolated from the 335 seafood samples and the dot-blot ELISA for V. cholerae 01 antigen was also negative, consistent with a high specificity of the assay. The dot-blot ELISA is easy to perform, relatively inexpensive, highly sensitive and specific. It permits multisample analysis at a single time, requires no special equipment and does not pose any disposal problem (compared with the culture method). Most of all, diagnosis of cholera cases could be made accurately within 1–3 h (the dot-blot ELISA takes 1 h, while the culture method takes 2 days). The method is recommended for rapid detection of V. cholerae 01 in contaminated foods, in environmental samples and in stools of diarrhoeic patients.