title: Quantitative PCR and culture evaluation for enterotoxigenic Escherichia coli (ETEC) associated diarrhea in volunteers.
abstract: Recent evidence suggests that the abundance of enteric pathogens in the stool correlates with the presence of clinical diarrhea. We quantified the fecal pathogen after feeding enterotoxigenic Escherichia coli (ETEC) strain H10407 to 30 adult volunteers. Stools were collected daily and examined using qualitative and quantitative (Q) culture. DNA was isolated, and quantitative (Q) PCR targeting the heat-labile toxin (LT) gene was performed. Nine volunteers developed diarrhea. Among 131 stool specimens with complete data, pathogen abundance by QPCR was strongly correlated with Qculture, ρ = 0.61, P < 0.0001. Receiver operating characteristic curve analysis comparing quantitative data against diarrhea status suggested cut-points, based on a maximum Youden Index, of 2.8 × 10(4) LT gene copies and 1.8 × 10(7) CFU. Based on these cut-points, QPCR had a sensitivity and specificity compared with diarrheal status of 0.75 and 0.87, respectively, and an OR of 20.0 (95% CI 5.7-70.2), whereas Qculture had a sensitivity and specificity of 0.73 and 0.91, respectively, and an OR of 28.6 (95% CI 7.7-106.6). Qculture had a sensitivity and specificity of 0.82 and 0.48, respectively and an OR of 4.4 (95% CI 1.2-16.0). The correlation between Qculture and QPCR was highest in diarrheal specimens, and both quantitative methods demonstrated stronger association with diarrhea than qualitative culture.
authors.affiliation: Department of Epidemiology and Public Health, University of Maryland, School of Medicine, Baltimore, MD, USA.
authors.forename: Brianna R
authors.initials: BR
authors.lastname: Lindsay
authors.forename: Subhra
authors.initials: S
authors.lastname: Chakraborty
authors.forename: Clayton
authors.initials: C
authors.lastname: Harro
authors.forename: Shan
authors.initials: S
authors.lastname: Li
authors.forename: James P
authors.initials: JP
authors.lastname: Nataro
authors.forename: Halvor
authors.initials: H
authors.lastname: Sommerfelt
authors.forename: David A
authors.initials: DA
authors.lastname: Sack
authors.forename: Oscar
authors.initials: O
authors.lastname: Colin Stine
mesh_terms: D015169:Colony Count, Microbial / Q000379:methods*; D003967:Diarrhea / Q000175:diagnosis / Q000382:microbiology*; D054307:Enterotoxigenic Escherichia coli / Q000235:genetics / Q000254:growth & development / Q000302:isolation & purification*; D004768:Enterotoxins / Q000235:genetics; D004927:Escherichia coli Infections / Q000175:diagnosis / Q000382:microbiology*; D005243:Feces / Q000382:microbiology; D064368:Healthy Volunteers; D006801:Humans; D016133:Polymerase Chain Reaction / Q000379:methods*
chemical_list: D004768:Enterotoxins
keywords: ETEC; QPCR; diagnostic; quantitative culture; sensitivity; specificity