DETECTION OF ENTEROTOXIGENIC CLOSTRIDIUM-PERFRINGENS IN RAW BEEF BY POLYMERASE CHAIN-REACTION

A polymerase chain reaction (PCR) procedure was developed for direct d etection of Clostridium perfringens strains with potential for food po isoning in raw beef samples. An oligonucleotide primer pair was used t o amplify a 364 base pair sequence internal to the C. perfingens enter otoxin gene. One milliliter portions of the meat homogenates Were inoc ulated into cooked meat medium (CMM) or reduced Fluid Thioglycollate ( FTG) medium and incubated at 37 degrees C. Portions sampled at 2, 4, 6 , 8 and 24 h of enrichment were assayed for detection of the enterotox in sequence by PCR. Amplification of the 364 bp sequence could be dete cted in 6 h by agarose gel electrophoresis and as early as ? h by hybr idization to a 150 bp digoxigenin (DIG)-labeled probe. To increase the sensitivity of the detection assay a commercial chromosomal deoxyribo nucleic acid (DNA) extraction assay was Compared with a nested PCR app roach. Both methods allowed detection of less than 1 log(10) colony fo rming units (CFU)/g of C. perfringens strains harboring the enterotoxi n gene, with no interference with the background microflora present in the raw ground beef.