Detection of small numbers of Campylobacter jejuni and Campylobacter coli cells in environmental water, sewage, and food samples by a seminested PCR assay

A rapid and sensitive assay was developed for detection of small numbers of Campylobacter jejuni and Campylobacter coli cells in environmental water, sewage, and food samples. Water and sewage samples were filtered, and the f ilters were enriched overnight in a nonselective medium. The enrichment cul tures were prepared for PCR by a rapid and simple procedure consisting of c entrifugation, proteinase K treatment, and boiling. A seminested PCR based on specific amplification of the intergenic sequence between the two Campyl obacter flagellin genes, flaA and flaB, was performed, and the PCR products were visualized by agarose gel electrophoresis, The assay allowed us to de tect 3 to 15 CFU of C. jejuni per 100 ml in water samples containing a back ground flora consisting of up to 8,700 heterotrophic organisms per mi and 1 0,000 CFU of coliform bacteria per 100 mi, Dilution of the enriched culture s 1:10 with sterile broth prior to the PCR was sometimes necessary to obtai n positive results. The assay was also conducted with food samples analyzed with or without overnight enrichment, As few as less than or equal to 3 CF U per g of food could be detected with samples subjected to overnight enric hment, while variable results were obtained for samples analyzed without pr ior enrichment. This rapid and sensitive nested PCR assay provides a useful tool for specific detection of C. jejuni or C. coli in drinking water, as well as environmental water, sewage, and food samples containing high level s of background organisms.