A direct polymerase chain reaction method to EC cultured broth (named EC-PCR) for screening detection of enteropathogenic E. coli (ETEC, VTEC and EIEC) in foods and feces is described.
Eleven E. coli strains belonging to three groups (4ETEC: 06, 025, 0128, 0148; 4VTEC: 026, 0111, 0145, 0157; and 3EIEC: 028ac, 0124, 0164) were used for this study. Four pairs of oligonucleotide primers homologous to LT, ST, VT and EIEC genes were used in combination. The culture condition at 37-43°C for 16-20 h in EC broth was most suitable for the EC-PCR method, and no cross readings were observed with other bacteria, substances in meats or feces.
After a 20-h enrichment step, it was possible to detect fewer than 10 to 102 bacteria per g of the aritificially inoculated meat. E. coli was easily detected because of low contamination with other bacteria which disturb the detection of E. coli. However, in feces, it ranged from 103 to 104 cfu per g. The large number of bacteria with feces are the main limiting factor of the EC-PCR detection assay. All E. coli strains examined were detected from all the enrichment cultures on both the EC-PCR and the culture methods. In two sporadic cases and two food poisoning cases, the enteropathogenicity of E. coli isolates from patients was rapidly judged by the EC-PCR method.
These findings were consistent with those of the culture method. Thus, the findings suggest that the EC-PCR method is a suitable, sensitive and rapid method for detection of the potentially enteropathogenic E. coli.
