Rapid and simple detection of Escherichia coli O157: H7 using multiplex PCR
Keywords:
Escherichia coli O157:H7, Detection, Specificity, Multiplex PCR, SensitivityAbstract
Background: Escherichia coli O157:H7 is a major foodborne pathogen and can be found globally. Traditional microbiological methods to detect E.coli O157:H7 are expensive and time consuming. However, detection assay using polymerase chain reaction (PCR) can be very specific and may be a suitable alternative.
Objective: The present study aimed to evaluate a simple and rapid diagnostic multiplex PCR assay for the selective identification of attaching and effacing genes of E.coli O157:H7.
Methods: One-step multiplex PCR using specific primers for eae (Intimin) and rfbE (O157 antigen) genes was carried out. PCR products were analyzed by agarose gel electrophoresis. The specificity of this method was determined using E. coli O157:H7, Salmonella typhi, Pseudomonas aeruginosa, and Vibrio cholera. Sensitivity was determined from serial dilutions of E. coli O157:H7 genomic DNA.
Results: Specific PCR products of 206 and 497 base pairs were obtained from the amplification of eae and rfbE, respectively. No cross-amplification of other bacterial genes was observed. Assay sensitivity was 380 femtogram.
Conclusion: This multiplex PCR assay can be used as a simple diagnostic test in clinical laboratories for the specific and rapid identification of E. coli O157:H7.
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