Multiplex PCR assay and simple preparation method for stool specimens detect enterotoxigenic Escherichia coli DNA during course of infection

Abstract

Infection with enterotoxigenic Escherichia coli (ETEC) is a common cause of diarrhea among travelers and residents of developing countries. ETEC produces either a heat-stable toxin or a heat-labile toxin, or both, encoded by plasmid-borne ST and LT genes, respectively. Diagnosis of infection with this subclass of E. coli can be performed with oligonucleotide hybridization probes; however, the sensitivity and specificity of this method are insufficient. A nonradioactive multiplex PCR assay that provides a sensitive and specific method for detecting the presence of either or both toxin genes has been developed. A simple procedure that removed inhibitors of the PCR while efficiently releasing ETEC DNA from stool specimens for subsequent amplification was used. The results for samples from a human volunteer study of ETEC infection indicated that this method of sample preparation results in greater clinical sensitivity than conventional total nucleic acid extraction and ethanol precipitation. Detection of ETEC by a multiplex PCR assay in stool specimens directly processed with a glass matrix and chaotropic solution had greater sensitivity than culture.

EnteroaggregativeEscherichia colipathotype: a genetically heterogeneous emerging foodborne enteropathogen

“…Molecular biology assays based on virulence genes have been extensively used to identify most diarrheagenic E. coli (Gunzburg et al, 1995;Stacy-Phipps et al, 1995;Paton & Paton, 1998;Lopez-Saucedo et al, 2003). However, for a long time, two main limitations have hampered the development of molecular identification methods for EAEC.…”

The structures ofEscherichia coliO-polysaccharide antigens

“…The assays are based on nucleic acid probes and PCR and have been extensively used. The advantages of PCR include its high sensitivity in detection of target templates and both rapid and reliable results due to its high specificity (Schultsz et al, 1994;Ramotar et al, 1995;Stacy-Phipps et al, 1995;Kai et al, 2000;Dutta et al, 2001;Pulz et al, 2003;Gioffre et al, 2004).…”

Genotyping of fimbrial adhesins inEscherichia coli strains isolated from slovak piglets suffering from diarrhea

“…The effectiveness of serological tests is affected by the level of expression of antigen and the quality of the antiserum used. In the recent period diagnosis of these strains has been carried out increasingly with the help of the PCR method as a quite sensitive and specific method with primers for detection of genes coding for enterotoxin (Stacy-Philips et al 1995) or adherence antigens (Ojeniyi et al 1994;Osek 1999;Alexa et al 2001;Holoda et al 2002;Vu-Khac et al 2003).…”