A Quantitative PCR Assay for Rapid Detection of 9 Pathogens Directly From Stools of Travelers with Diarrhea.
Clin Gastroenterol Hepatol. 2013 Apr 29;
Authors: Antikainen J, Kantele A, Pakkanen SH, Lääveri T, Riutta J, Vaara M, Kirveskari J
Abstract
BACKGROUND: & Aims: Every year 80 million tourists travelling to tropical and subtropical areas contract traveler's diarrhea (TD). Forty percent to 80% of cases are caused by bacteria, yet clinical diagnostic tests are available to identify only a few of the strains that cause TD. We aimed to develop a quantitative PCR (qPCR) assay to identify all major pathogens in stool samples. METHODS: We developed a low-cost, high-throughput, multiplex qPCR assay for simultaneous detection of 9 bacterial pathogens in stool samples: Salmonella, Yersinia, Campylobacter, and Vibrio cholerae, as well as Shigella or enteroinvasive Escherichia coli (EIEC), enterohemorrhagic E coli (EHEC), enterotoxigenic E coli (ETEC), enteroaggregative E coli (EAEC), and enteropathogenic E coli (EPEC). The assay was validated using positive (n=245) and negative (n=243) control strains as well as preselected positive and negative stool samples. In addition, stool samples were collected from 96 returning travelers with TD. The findings were compared with those from routine diagnostic tests. RESULTS: The assay detected the bacterial strains with 100% sensitivity and specificity, compared with results from the reference tests. Of all stool samples collected from travelers with TD, EPEC was found in 47%, EAEC in 46%, ETEC in 22%, EHEC in 7%, Campylobacter in 6%, Shigella or EIEC in 2%, and Salmonella in 2%. Multiple pathogens were found in 37% of all samples. CONCLUSIONS: We developed a low-cost, high throughput qPCR assay for use in routine diagnostic analysis and research. It detects the pathogenic bacteria most commonly associated with TD in stool samples with 100% sensitivity and specificity, compared to reference methods. The assay requires 4 hours, whereas current detection methods require 1-7 days. At least 1 TD pathogen was identified in stool samples from 76% of returning travelers, whereas conventional methods found a pathogen in only 17%. The most commonly detected bacteria were EPEC, EAEC, and ETEC.
PMID: 23639597 [PubMed - as supplied by publisher]