Multiplex PCR for Simultaneous Detection of Virulence Genes in Escherichia coli

Thong, K.L.; Ke, X.Y. (2009) Multiplex PCR for Simultaneous Detection of Virulence Genes in Escherichia coli. Malaysian Journal of Science, 28 (1). pp. 1-14. ISSN 1394-3065

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    Abstract

    Diarrhea caused by Escherichia coli infection is a major cause of public health problems in developing countries. In view of the deficits and limitations of conventional methods of detecting the virulence determinants, a multiplex Polymerase Chain Reaction (PCR) assay was optimized and developed to provide an effective, rapid and specific diagnostic tool to simultaneously detect virulence genes such as heatstable toxin 1 (ST1), heat-labile toxin 1 (LT1), heat-labile toxin 2 (LT2), verotoxin1 (VT1), verotoxin 2 (VT2) and attachment and effacement (eaeA) in pathogenic E. coli. Five sets of primers targeting these six virulence genes were optimized by using postitive control E. coli strains. The optimized conditions consisted of 3.0 mM of MgCl2, 0.2 mM of dNTPs, 1.5 U of Taq DNA polymerase (Promega), 0.70 µM of VT primer, 0.60 µM of LT2 primer and 0.07 µM each of LT1 primer, ST primer and AE primer. The mPCR assay was then applied to a panel of 87 E. coli isolates from different sources. One food isolate (EC 375) was positive for eaeA gene while another environmental isolate had ST, LT1, eaeA and VT genes. The study shows that the mPCR assay is a useful tool to differentiate the pathogenic potential (pathotypes) of E. coli by presence of known virulence genes.

    Item Type: Article
    Creators:
    1. Thong, K.L.(Institute of Biological Sciences, Faculty of Science Building, University of Malaya, 50603 Kuala Lumpur, MALAYSIA)
    2. Ke, X.Y.
    Journal or Publication Title: Malaysian Journal of Science
    Uncontrolled Keywords: Escherichia coli, multiplex Polymerase Chain Reaction, virulence genes
    Subjects: Q Science > Q Science (General)
    Q Science > QR Microbiology
    Divisions: Faculty of Science > Institute of Biological Sciences
    Depositing User: miss munirah saadom
    Date Deposited: 05 Jan 2013 00:35
    Last Modified: 15 Dec 2014 14:13
    URI: http://eprints.um.edu.my/id/eprint/5657

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