title: Comparison of two poultry litter qPCR assays targeting the 16S rRNA gene of Brevibacterium sp.
abstract: Chicken feces commonly contain human pathogens and are also important sources of fecal pollution in environmental waters. Consequently, methods that can detect chicken fecal pollution are needed in public health and environmental monitoring studies. In this study, we compared a previously developed SYBR green qPCR assay (LA35) to a novel TaqMan qPCR assay (CL) for the environmental detection of poultry-associated fecal pollution. We tested both assays against chicken litter (n = 40), chicken fecal samples (n = 186), non-chicken fecal sources (n = 484), and environmental water samples (n = 323). Most chicken litter samples (i.e., ≥ 98%) were positive for both assays with relatively high signal intensities, whereas only 23% and 12% of poultry fecal samples (n = 186) were positive with the LA35 and the CL assays, respectively. Data using fecal samples from non-target animal species showed that the assays are highly host-associated (≥ 95%). Bayesian statistical models showed that the two assays are associated with relatively low probability of false-positive and false-negative signals in water samples. The CL marker had a lower prevalence than the LA35 assay when tested against environmental water samples (i.e., 21% vs. 31% positive signals). However, by combining the results from the two assays the detection levels increased to 41%, suggesting that using multiple assays can improve the detection of chicken-fecal pollution in environmental waters.
authors.affiliation: National Risk Management Research Laboratory, Cincinnati, OH, USA.
authors.forename: Hodon
authors.initials: H
authors.lastname: Ryu
authors.forename: Michael
authors.initials: M
authors.lastname: Elk
authors.forename: Izhar U H
authors.initials: IU
authors.lastname: Khan
authors.forename: Valerie J
authors.initials: VJ
authors.lastname: Harwood
authors.forename: Marirosa
authors.initials: M
authors.lastname: Molina
authors.forename: Thomas A
authors.initials: TA
authors.lastname: Edge
authors.forename: Jorge Santo
authors.initials: JS
authors.lastname: Domingo
mesh_terms: D000818:Animals; D001483:Base Sequence; D001499:Bayes Theorem; D001951:Brevibacterium / Q000235:genetics*; D017931:DNA Primers; D005798:Genes, Bacterial; D008372:Manure; D016133:Polymerase Chain Reaction / Q000379:methods*; D011200:Poultry; D012336:RNA, Ribosomal, 16S / Q000235:genetics*
chemical_list: D017931:DNA Primers; D008372:Manure; D012336:RNA, Ribosomal, 16S
keywords: Fecal pollution; Microbial source tracking; Poultry-associated assays; qPCR