Shiga toxin-producing E. coli (STEC) are a serious cause of human diseases. The infections are mainly associated with strains belonging to serogroups O157, O26, O103, O111 and O113, while the main source of infection is contaminated food of animal origin (especially beef). In this study the molecular identification methods for STEC were used which enabled detection of STEC in raw beef using multiplex PCR (mPCR) assays, isolation of individual bacterial colonies with digoxigenin (DIG)-labeled DNA probe and characterization of the virulence markers by the use of mPCR. The molecular method was used to examine 272 raw beef samples obtained from slaughterhouses. After the mPCR-1 analysis 16 stx-positive samples (5.88%) were detected. The STEC isolates were then tested using the mPCR and PCR assays. Eight of them belonged to O157:H7 serotype by the presence of the rfbO157 and fliCн₇ genes. The stx₁ marker was observed in all E. coli O157:H7 and in four of the non-O157:H7 isolates tested. Moreover, the stx₂ gene was detected in all E. coli O157:H7 and in seven of the non-O157:H7, three of them also possessed the stx₁ marker. Eight of the rfbO157-negative STEC were examined with the mPCR-3 assay to detect the rfbO111, rfbO113 and O26wzx markers, three of them produced the characteristic amplicon for the O26 group. These isolates also carried the stx₁ (one strain) or stx₂ genes (two isolates). Among the STEC tested, none belonged to the O111 or O113 groups. The amplification with the mPCR-4 assay showed that all isolates harbored the enterohemolysin (ehlyA) gene whereas the intimin marker (eaeA) was observed in all E. coli O157:H7 and O26 isolates.
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