title: Development of a quantitative PCR assay for the quantitation of bovine polyomavirus as a microbial source-tracking tool.
abstract: Adenoviruses and polyomaviruses are two distinct DNA viral families that are excreted in high concentrations and distributed in human and animal populations. Targeting specific virus included in these families has proved to be a promising and useful tool for tracing specifically sources of environmental contamination. In this study, a quantitative PCR assay that is specific for bovine polyomaviruses was developed and used to determine the excretion level and concentration of bovine polyomaviruses in urine and environmental samples, including urban sewage, slaughterhouse sewage, and river water. A set of primers and a TaqMan probe were designed to target a 77-bp region of the bovine polyomavirus VP1 gene, and the conditions of the reaction were optimized. A detection limit was established at 1-10 genome copies per test tube. The assay was specific and produced negative results when samples containing human or porcine fecal contamination were analyzed. This is, to our knowledge, the first description of bovine polyomaviruses excreted in bovine urine samples (mean values of 10(4) GC/l). Bovine polyomaviruses were also detected and quantified in slaughterhouse wastewater and river waters, which shows the spread of these viruses in many environmental samples containing contamination of bovine origin. The procedure described in this paper provides a quantitative source-tracking tool for the analysis of bovine polyomaviruses as indicators of the presence of bovine contamination in environmental samples.
authors.affiliation: Department of Microbiology, University of Barcelona, Avda Diagonal 645, 08028 Barcelona, Spain.
authors.forename: Ayalkibet
authors.initials: A
authors.lastname: Hundesa
authors.forename: Silvia
authors.initials: S
authors.lastname: Bofill-Mas
authors.forename: Carlos
authors.initials: C
authors.lastname: Maluquer de Motes
authors.forename: Jesus
authors.initials: J
authors.lastname: Rodriguez-Manzano
authors.forename: Alex
authors.initials: A
authors.lastname: Bach
authors.forename: Maribel
authors.initials: M
authors.lastname: Casas
authors.forename: Rosina
authors.initials: R
authors.lastname: Girones
mesh_terms: D000818:Animals; D002417:Cattle; D017931:DNA Primers / Q000235:genetics; D006801:Humans; D016133:Polymerase Chain Reaction / Q000379:methods*; D011120:Polyomavirus / Q000235:genetics / Q000302:isolation & purification*; D012680:Sensitivity and Specificity; D012722:Sewage / Q000821:virology*; D014556:Urine / Q000821:virology*; D019562:Viral Load / Q000379:methods*; D015678:Viral Structural Proteins / Q000235:genetics; D014871:Water Microbiology
chemical_list: D017931:DNA Primers; D012722:Sewage; D015678:Viral Structural Proteins