title: A sensitive multiplex, real-time PCR assay for prospective detection of Shiga toxin-producing Escherichia coli from stool samples reveals similar incidences but variable severities of non-O157 and O157 infections in northern California.
abstract: Rapid and accurate detection of Shiga toxin-producing Escherichia coli (STEC) of all serotypes from patients with diarrhea is critical for medical management and for the prevention of ongoing transmission. In this prospective study, we assessed the performance of a multiplex, real-time PCR assay targeting stx1 and stx2 for the detection of O157 and non-O157 STEC in diarrheal stool samples enriched in Gram-negative broth. We show that the assay is 100% sensitive (95% confidence interval [CI], 89.1% to 100%) and 98.5% specific (95% CI, 90.6% to 99.9%) based on a panel of 40 known STEC-positive specimens and 65 known negative specimens. During a 2-year postvalidation period, the assay detected more positive samples from patients in northern California than did culture and PCR testing performed at a public health reference laboratory, with a positive predictive value of 95.6% (95% CI, 87.6% to 99.1%). Serotyping data showed an incidence rate of 51.2% for non-O157 STEC strains, with 5.8% of patients (1/17) with non-O157 strains and 42.9% (6/14) with O157 strains (P = 0.03) developing hemolytic-uremic syndrome. The findings from this study underscore the recommendations of the CDC for laboratories to test all diarrheal stool samples from patients with acute community-acquired diarrhea for non-O157 STEC in addition to the O157 serotype by using a sensitive assay. Additionally, a survey of 17 clinical laboratories in northern California demonstrated that nearly 50% did not screen all stool specimens for the presence of Shiga toxins, indicating that many clinical microbiology laboratories still do not routinely screen all stool specimens for the presence of Shiga toxins as recommended in the 2009 CDC guidelines.
authors.affiliation: Department of Pathology, Stanford University School of Medicine, Stanford, California, USA.
authors.forename: Martina I
authors.initials: MI
authors.lastname: Lefterova
authors.forename: Kathleen A
authors.initials: KA
authors.lastname: Slater
authors.forename: Indre
authors.initials: I
authors.lastname: Budvytiene
authors.forename: Patricia A
authors.initials: PA
authors.lastname: Dadone
authors.forename: Niaz
authors.initials: N
authors.lastname: Banaei
mesh_terms: D000293:Adolescent; D000328:Adult; D000368:Aged; D001431:Bacteriological Techniques / Q000379:methods*; D002140:California / Q000453:epidemiology; D002648:Child; D002675:Child, Preschool; D004927:Escherichia coli Infections / Q000453:epidemiology* / Q000382:microbiology / Q000473:pathology*; D005243:Feces / Q000382:microbiology; D005260:Female; D006801:Humans; D015994:Incidence; D007223:Infant; D008297:Male; D008875:Middle Aged; D060885:Multiplex Polymerase Chain Reaction / Q000379:methods*; D011237:Predictive Value of Tests; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D012703:Serotyping; D012720:Severity of Illness Index; D054323:Shiga-Toxigenic Escherichia coli / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification* / Q000472:pathogenicity; D055815:Young Adult