This Article Full Text Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Reprints and Permissions Copyright Information Books from ASM Press MicrobeWorld Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Li, F. Articles by Zhang, D. Y. Search for Related Content PubMed PubMed Citation Articles by Li, F. Articles by Zhang, D. Y.

 Previous Article  |  Next Article 

Journal of Clinical Microbiology, December 2005, p. 6086-6090, Vol. 43, No. 12
0095-1137/05/$08.00+0     doi:10.1128/JCM.43.12.6086-6090.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.

Use of Ramification Amplification Assay for Detection of Escherichia coli O157:H7 and Other E. coli Shiga Toxin-Producing Strains

Department of Pathogenobiology, School of Basic Medical Sciences, Jilin University, Changchun, China,¹ Department of Pathology, Mount Sinai School of Medicine, New York, New York,² Department of Nutrition and Food Science, University of Maryland, College Park, Maryland³

Received 10 February 2005/ Returned for modification 28 April 2005/ Accepted 17 August 2005

Escherichia coli O157:H7 and other Shiga toxin-producing E. coli (STEC) strains are important human pathogens that are mainly transmitted through the food chain. These pathogens have a low infectious dose and may cause life-threatening illnesses. However, detection of this microorganism in contaminated food or a patient's stool specimens presents a diagnostic challenge because of the low copy number in the sample. Often, a more sensitive nucleic acid amplification method, such as PCR, is required for rapid detection of this microorganism. Ramification amplification (RAM) is a recently introduced isothermal DNA amplification technique that utilizes a circular probe for target detection and achieves exponential amplification through the mechanism of primer extension, strand displacement, and ramification. In this study, we synthesized a circular probe specific for the Shiga toxin 2 gene (stx₂). Our results showed that as few as 10 copies of stx₂ could be detected, indicating that the RAM assay was as sensitive as conventional PCR. We further tested 33 isolates of E coli O157:H7, STEC, Shigella dysenteriae, and nonpathogenic E. coli by RAM assay. Results showed that all 27 STEC isolates containing the stx₂ gene were identified by RAM assay, while S. dysenteriae and nonpathogenic E. coli isolates were undetected. The RAM results were 100% in concordance with those of PCR. Because of its simplicity and isothermal amplification, the RAM assay could be a useful method for detecting STEC in food and human specimens.

This article has been cited by other articles:

• Hussein, H. S. (2007). Prevalence and pathogenicity of Shiga toxin-producing Escherichia coli in beef cattle and their products. J ANIM SCI 85: E63-E72 [Abstract] [Full Text]