Evaluation of alkaline phosphatase- and digoxigenin-labelled probes for detection of the thermolabile hemolysin (tlh) gene of Vibrio parahaemolyticus.
The biochemical identification and enumeration of Vibrio parahaemolyticus as described in the FDA Bacteriological Analytical Manual is expensive and labour-intensive. To reduce the time and effort necessary to verify the identity of V. parahaemolyticus, the use of a thermolabile haemolysin (tlh) gene probe is proposed. An alkaline phosphatase (AP)-labelled probe was evaluated for specificity against 26 strains of V. parahaemolyticus, 88 strains of other Vibrio species and 10 strains of non-vibrio species. Of the 124 isolates tested, the probe hybridized only with the 26 strains of V. parahaemolyticus, indicating species specificity. Two hundred and six suspect V. parahaemolyticus isolates from oysters were tested by this probe and API-20E diagnostic strips; there was 97% agreement between results. A digoxigenin (DIG)-labelled probe for detection of the tlh gene fragment was prepared by PCR and compared with the AP-labelled probe. When tested on 584 suspect V. parahaemolyticus isolates, results obtained with the AP- and DIG-labelled probes were in 98% agreement. These results suggest that the probes are equivalent for detection of the V. parahaemolyticus tlh gene.
US Food and Drug Administration, Dauphin Island, AL 36528, USA.
S A
SA
McCarthy
A
A
DePaola
D W
DW
Cook
C A
CA
Kaysner
W E
WE
Hill
D000469:Alkaline Phosphatase; D000818:Animals; D015373:Bacterial Typing Techniques; D004076:Digoxigenin; D004196:Disease Outbreaks; D004783:Environmental Microbiology; D005069:Evaluation Studies as Topic; D005517:Foodborne Diseases / Q000382:microbiology; D005798:Genes, Bacterial; D006460:Hemolysin Proteins / Q000235:genetics*; D006801:Humans; D015345:Oligonucleotide Probes; D010125:Ostreidae / Q000382:microbiology; D012680:Sensitivity and Specificity; D013045:Species Specificity; D013696:Temperature; D014735:Vibrio Infections / Q000382:microbiology; D014736:Vibrio parahaemolyticus / Q000145:classification / Q000235:genetics* / Q000302:isolation & purification*
D006460:Hemolysin Proteins; D015345:Oligonucleotide Probes; D000469:Alkaline Phosphatase; D004076:Digoxigenin