The official method for screening Escherichia coli O157 and O26 in food involves the detection of the VT genes by PCR. In this study, we evaluated the efficacy of the application of an internal control (EC3 DNA is coamplified with the VT gene) in preventing false-negative results of VT gene screening. A modular approach was applied to consider the effect of each parameter: (1) 4 food samples were tested by using 3 types of cultures artificially inoculated with E. coli O157 before or after enrichment; (2) 2 commercial kits were used; and (3) 2 DNA extraction methods in the official method were employed. We found that the PCR performance was greatly affected by each experimental parameter; in particular, the DNA extraction method that influenced the concentration of PCR inhibitors in each food sample. Hence, the use of internal control was found to be critical for avoiding false-negative results. Isolation of bacteria is the final assessment in the official method. If it is true-negative, however, any steps for isolating the E. coli will not be required. In conclusion, the results of our study may be useful in establishing an overall rapid and efficient test.
