Rapid detection of Vibrio parahaemolyticus in oysters by immunofluorescence microscopy

Abstract

A method of indirect immunofluorescence microscopy was developed for the rapid detection of Vibrio parahaemolyticus in oysters. Affinity-purified antibodies against two outer membrane proteins (molecular mass 36 and 34 kDa, respectively) of V. parahaemolyticus were used as the primary antibodies to label the vibrio, and fluorescein isothiocyanate-conjugated goat anti-rabbit immunoglobulin G was used to reveal the antigen-antibody reaction. Of 85 strains of V. parahaemolyticus tested, all produced strong fluorescence under the fluorescence microscope. However, for 63 strains (including 27 vibrios) of other bacteria tested, 6 produced weak to moderate fluorescence. The sensitivity and specificity of the immunostaining technique were 100 and 90.5%, respectively. In the inoculation experiments, as low as 1.7 CFU g of V. parahaemolyticus spiked in oysters could be detected by the immunostaining method after 18-h enrichment in alkaline peptone water containing 3% NaCl. The immunofluorescence microscopy is recommended for rapid screening of V. parahaemolyticus in oysters, a presumptive positive result could be obtained within 24 h.