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Detection of toxigenic Vibrio cholerae from environmental water samples by an enrichment broth cultivation–pit‐stop semi‐nested PCR procedure
Author(s) -
Theron J.,
Cilliers J.,
Du Preez M.,
Brözel V.S.,
Venter S.N.
Publication year - 2000
Publication title -
journal of applied microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.889
H-Index - 156
eISSN - 1365-2672
pISSN - 1364-5072
DOI - 10.1046/j.1365-2672.2000.01140.x
Subject(s) - vibrio cholerae , microbiology and biotechnology , biology , nested polymerase chain reaction , sewage , cholera , toxin , cholera toxin , vibrionaceae , polymerase chain reaction , bacteria , gene , environmental science , environmental engineering , genetics
A pit‐stop semi‐nested PCR assay for the detection of toxigenic Vibrio cholerae in environmental water samples was developed and its performance evaluated. The PCR technique amplifies sequences within the cholera toxin operon specific for toxigenic V. cholerae . The PCR procedure coupled with an enrichment culture detected as few as four V. cholerae organisms in pure culture. Treated sewage, surface, ground and drinking water samples were seeded with V. cholerae and following enrichment, a detection limit of as few as 1 V. cholerae cfu ml −1 was obtained with amplification reactions from crude bacterial lysates. The proposed method, which includes a combination of enrichment, rapid sample preparation and a pit‐stop semi‐nested PCR, could be applicable in the rapid detection of toxigenic V. cholerae in environmental water samples.