title: Development of a direct blood-based PCR system to detect BLV provirus using CoCoMo primers.
abstract: Bovine leukemia virus (BLV), the etiologic agent of enzootic bovine leucosis, has caused pandemic outbreaks worldwide. Because transcription of the BLV is quickly blocked after infection, detecting integrated provirus at host genome is an important method of identifying whether an animal is infected. The aim of the present study was to develop a novel direct blood-based PCR system to detect the BLV provirus with high specificity and at low cost. The assay was based on the BLV-CoCoMo degenerate primers, which amplify all known BLV strains. Cattle blood samples (n = 182) were collected from the same BLV-positive farm and subjected to BLV-CoCoMo-direct-PCR to detect the BLV provirus. The proviral load was then estimated. This novel PCR method showed 100 % specificity. The BLV-CoCoMo-direct-PCR can be used in a variety of laboratory situations because it does not require expensive equipment/reagents, DNA purification, or a second round of PCR. Therefore, the method is extremely cost-effective and the risk of a false-positive result due to DNA contamination is very low.
authors.affiliation: Viral Infectious Diseases Unit, RIKEN, 2-1 Hirosawa, Wako, Saitama, 351-0198, Japan.
authors.forename: Shin-Nosuke
authors.initials: SN
authors.lastname: Takeshima
authors.affiliation: Viral Infectious Diseases Unit, RIKEN, 2-1 Hirosawa, Wako, Saitama, 351-0198, Japan.
authors.forename: Sonoko
authors.initials: S
authors.lastname: Watanuki
authors.affiliation: NARO Institute of Livestock and Grassland Sciences, 768 Senbonmatsu, Nasushiobara, Tochigi, 329-2793, Japan.
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Ishizaki
authors.affiliation: NARO Institute of Livestock and Grassland Sciences, 768 Senbonmatsu, Nasushiobara, Tochigi, 329-2793, Japan.
authors.forename: Kazuhiro
authors.initials: K
authors.lastname: Matoba
authors.affiliation: Viral Infectious Diseases Unit, RIKEN, 2-1 Hirosawa, Wako, Saitama, 351-0198, Japan. aida@riken.jp.
authors.forename: Yoko
authors.initials: Y
authors.lastname: Aida
mesh_terms: D000818:Animals; D000914:Antibodies, Viral / Q000097:blood; D002417:Cattle; D017931:DNA Primers / Q000235:genetics; D004279:DNA, Viral / Q000097:blood / Q000235:genetics; D016583:Enzootic Bovine Leukosis / Q000097:blood* / Q000175:diagnosis / Q000821:virology*; D004797:Enzyme-Linked Immunosorbent Assay; D001909:Leukemia Virus, Bovine / Q000235:genetics* / Q000302:isolation & purification*; D016133:Polymerase Chain Reaction / Q000379:methods*; D011533:Proviruses / Q000235:genetics* / Q000302:isolation & purification*; D012680:Sensitivity and Specificity
chemical_list: D000914:Antibodies, Viral; D017931:DNA Primers; D004279:DNA, Viral