Verocytotoxin-producing Escherichia coli (VTEC) O26 is an emergent pathotype that has caused an increasing number of sporadic cases and outbreaks of gastroenteritis, hemorrhagic colitis, and hemolytic uremic syndrome in the United States and Europe. Many cases are associated with the consumption of milk and undercooked or fermented meats. The stx2 strains of VTEC O26 seem to be more likely to cause human infections than isolates expressing only stx1. The isolation and identification of VTEC O26 from foods is labor intensive and time-consuming. We developed a multiplex PCR (M-PCR) assay for the identification and characterization of E. coli O26 VTEC and its detection in raw milk and ground beef. The method is based on the amplification of the wzx, stx1, and stx2 genes for the simultaneous detection of the O26 antigen and verocytotoxin types 1 and 2. This M-PCR assay had a sensitivity of 108 CFU/ml when applied to a bacterial suspension and of 106 CFU/ml or g when applied to both inoculated milk and minced beef samples. This M-PCR assay also was highly specific, and results were consistently negative for negative controls (nonpathogenic E. coli strains, uninoculated milk and beef samples, and samples inoculated with the nontarget microorganisms). This method could be used for the rapid detection of E. coli O26 VTEC from foods and for the rapid identification and characterization of clinical and environmental isolates.
MULTIPLEX PCR AND LATEX AGGLUTINATION FOR IDENTIFICATION AND CHARACTERIZATION OF TOXIGENIC CLOSTRIDIUM DIFFICILE FROM RAW MILK
