Rapid and specific identification of Shiga toxin-producing Escherichia coli in faeces by multiplex PCR.
OBJECTIVE
The object of this study was to develop a multiplex PCR system for rapid and specific identification of Shiga toxin-producing Escherichia coli (STEC) in faeces.


RESULTS
A multiplex PCR (mPCR) protocol was developed using a primer pair specific for genes that are involved in the biosynthesis of the O157 E. coli antigen, and primers that identify the sequences of Shiga toxin 1 and 2 (stx 1 and stx1) and the intimin protein (eaeA). The mPCR assay was used for amplification of STEC genes in bacteria directly (after enrichment) in faeces. The test was very sensitive and could detect between 9 and 1 bacterial cells per gram of faeces. The mPCR was used for the examination of 69 bovine faecal samples derived from healthy cattle. The results indicated that 62 x 3% of the samples were positive, generating at least one PCR amplicon of the expected size.


CONCLUSIONS
The method can be applied for rapid and specific identification of STEC bacteria in faecal samples, and for differentiation of their main virulence marker genes.


CONCLUSIONS
The ability to sensitively detect Shiga toxin-producing E. coli directly in faeces within a short time represents a considerable advancement over more time-consuming and less sensitive methods for identification and characterization of STEC bacteria.
Osek
J
J
Department of Microbiology, National Veterinary Research Institute, Pulawy, Poland. josek@piwet.pulawy.pl
D000818:Animals; D001483:Base Sequence; D002417:Cattle / Q000382:microbiology*; D004269:DNA, Bacterial / Q000032:analysis; D019453:Escherichia coli O157 / Q000235:genetics / Q000302:isolation & purification* / Q000472:pathogenicity; D005243:Feces / Q000382:microbiology*; D006801:Humans; D016133:Polymerase Chain Reaction / Q000379:methods / Q000662:veterinary*; D012680:Sensitivity and Specificity; D022603:Shiga Toxins / Q000096:biosynthesis / Q000235:genetics*; D014774:Virulence
D004269:DNA, Bacterial; D022603:Shiga Toxins