title: Detection and typing of BKV, JCV, and SV40 by multiplex nested polymerase chain reaction.
abstract: A multiplex nested polymerase chain reaction (PCR) method was developed for detecting and differentiating simultaneously the DNA of polyomaviruses JC, BK, and SV40 in a single tube. In the first amplification step the same set of primers was used to amplify a conserved DNA region of the large T antigen gene of JCV, BKV, and SV40. The second round was carried out using a set of primers designed to obtain products of different size for each related virus. Subsequently, the sensitivity of the multiplex nested PCR was maximized by optimizing parameters such as primer, magnesium, and dNTP concentrations. The sensitivity of the method ranged between 1 and 10 copies of the polyomavirus genome. The assay was then used for detecting polyomavirus DNA in urine, serum, and biopsy specimens from renal transplant recipients. Based on the results obtained, the multiplex nested PCR developed in our study represents a useful tool for supporting the diagnosis of polyomavirus infection and could be used for epidemiological purposes and to better define the role of polyomaviruses in human pathology.
authors.affiliation: Department of Public Health and Microbiology, Virology Unit, University of Turin, Via Santena 9 - 10126, Turin, Italy.
authors.forename: Massimiliano
authors.initials: M
authors.lastname: Bergallo
authors.forename: Cristina
authors.initials: C
authors.lastname: Costa
authors.forename: Samuela
authors.initials: S
authors.lastname: Margio
authors.forename: Francesca
authors.initials: F
authors.lastname: Sidoti
authors.forename: Giuseppe P
authors.initials: GP
authors.lastname: Segoloni
authors.forename: Alessandro Negro
authors.initials: AN
authors.lastname: Ponzi
authors.forename: Rossana
authors.initials: R
authors.lastname: Cavallo
mesh_terms: D000368:Aged; D000956:Antigens, Viral / Q000276:immunology; D001483:Base Sequence; D017931:DNA Primers; D004279:DNA, Viral / Q000235:genetics; D005260:Female; D006801:Humans; D008297:Male; D008875:Middle Aged; D016133:Polymerase Chain Reaction / Q000379:methods*; D011120:Polyomavirus / Q000145:classification* / Q000235:genetics / Q000276:immunology; D012680:Sensitivity and Specificity
chemical_list: D000956:Antigens, Viral; D017931:DNA Primers; D004279:DNA, Viral