Rapid microimmunodiffusion method with species-specific antiserum raised to purified antigen for identification of Vibrio vulnificus

Abstract

An antigen common to Vibrio vulnificus strains, designated VVA, was purified by ammonium sulfate precipitation, gel filtration, ion-exchange column chromatography, and preparative gel electrophoresis. The molecular weight of VVA was 64,000 when estimated by gel filtration and 40,000 when measured by denaturing polyacrylamide gel electrophoresis. Antiserum prepared against purified VVA (anti-VVA serum) did not agglutinate whole cells of V. vulnificus. Therefore, VVA was considered a possible internal antigen. By using anti-VVA serum, a microimmunodiffusion method was designed to detect the antigen VVA in bacterial cell lysates prepared from a single colony. This simple method allowed the specific identification of V. vulnificus as soon as 10 h after antigen preparation and therefore can be a useful tool in the identification of V. vulnificus from environmental or clinical specimens. VVA was not detected as a line of complete identity in some 20 other Vibrio species or in 7 other bacterial genera. VVA was present in all 63 isolates of V. vulnificus obtained from clinical and nonclinical sources.