Direct plating procedure for enumerating Vibrio vulnificus in oysters (Crassostrea virginica).
A procedure for enumerating and identifying Vibrio vulnificus in oysters was developed and evaluated. This method consists of growth on a direct plating medium (VVE medium) for isolating the organism from shellfish tissues, followed by biochemical tests for differentiating and identifying presumptively positive isolates. Densities of V. vulnificus are reliably obtained in 2 to 4 days, and as few as 10 culturable cells per 100 g can be identified. The procedure was evaluated by using a DNA probe technique specific for the cytotoxin-hemolysin gene of V. vulnificus and gas chromatographic analysis of the fatty acid contents of positive isolates. Only 3.2 and 0.4% of the isolates gave false-positive and false-negative results, respectively. The average level of recovery on VVE medium for 33 strains, including both clinical and environmental isolates, was 92% of the level of recovery obtained with brain heart infusion agar supplemented with 1% NaCl. The densities of V. vulnificus in oyster homogenates and individual oysters harvested from gulf and Atlantic coastal waters revealed that seasonally high levels occurred. The VVE medium procedure facilitated enumeration of this pathogen in molluscan shellfish and had a distinct advantage over the widely used most-probable-number procedure for V. vulnificus enumeration, which requires 5 to 7 days and often gives improbable and imprecise results.
Miceli
G A
GA
Department of Microbiology, University of Rhode Island, Kingston 02881.
Watkins
W D
WD
Rippey
S R
SR
D000818:Animals; D001431:Bacteriological Techniques; D015169:Colony Count, Microbial; D003470:Culture Media; D015342:DNA Probes; D005069:Evaluation Studies as Topic; D005227:Fatty Acids / Q000032:analysis; D005516:Food Microbiology; D006801:Humans; D010125:Ostreidae / Q000382:microbiology*; D012680:Sensitivity and Specificity; D014733:Vibrio / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification*
D003470:Culture Media; D015342:DNA Probes; D005227:Fatty Acids