title: Degenerate polymerase chain reaction strategy with DNA microarray for detection of multiple and various subtypes of virus during blood screening.
abstract: BACKGROUND
  The risk of transferring blood-borne infections during transfusion is continually increasing because of newly emerging and reemerging viruses. Development of a rapid screening method for emerging viruses that might be transmitted by transfusion is required to eliminate such pathogens during blood donor screening. Owing to increased use of human materials in organ transplants and cell therapy, the risk of donor-transmitted viral infections is also increasing. Although nucleic acid amplification technology (NAT) is dedicated to blood screening, a small, convenient detection system is needed at the laboratory and hospital level.
  
  
  METHODS
  We developed a new pathogen detection system that can detect multiple viruses simultaneously, using originally designed degenerate polymerase chain reaction primers to amplify a wide range of viral genotypes. Amplified samples were identified using a DNA microarray of pathogen-specific probes.
  
  
  RESULTS
  We detected very low copy numbers of multiple subtypes of viruses, such as human hepatitis C virus (HCV), human hepatitis B virus (HBV), human parvovirus B19 (PVB19), and West Nile virus (WNV), using a single plate. We also detected all genotypes of human immunodeficiency virus (HIV) but sensitivity was less than for the other viruses.
  
  
  CONCLUSIONS
  We developed a microarray assay using novel primers for detection of a wide range of multiple pathogens and subtypes. Our NAT system was accurate and reliable for detection of HIV, HBV, HCV, PVB19, and WNV, with respect to specificity, sensitivity, and genotype inclusivity. Our system could be customized and extended for emerging pathogens and is suitable as a future NAT system.
authors.affiliation: Department of Safety Research on Blood and Biologics, Department of Virology, Medicine, National Institute of Infectious Diseases, Tokyo, Japan; Department of Pathology, Yamaguchi University Graduate School of Medicine, Yamaguchi, Japan; Department of Veterinary Medicine, Rakuno Gakuen University, Hokkaido, Japan; Osaka Red Cross Blood Center, Osaka, Japan; Nihon Parkerizing Hiroshima Works Co. Ltd, Hiroshima, Japan.
authors.forename: Kazuya
authors.initials: K
authors.lastname: Takizawa
authors.forename: Tatsuo
authors.initials: T
authors.lastname: Nakashima
authors.forename: Takuo
authors.initials: T
authors.lastname: Mizukami
authors.forename: Madoka
authors.initials: M
authors.lastname: Kuramitsu
authors.forename: Daiji
authors.initials: D
authors.lastname: Endoh
authors.forename: Shigeto
authors.initials: S
authors.lastname: Kawauchi
authors.forename: Kohsuke
authors.initials: K
authors.lastname: Sasaki
authors.forename: Haruka
authors.initials: H
authors.lastname: Momose
authors.forename: Yoshiharu
authors.initials: Y
authors.lastname: Kiba
authors.forename: Tetsuya
authors.initials: T
authors.lastname: Mizutani
authors.forename: Rika A
authors.initials: RA
authors.lastname: Furuta
authors.forename: Kazunari
authors.initials: K
authors.lastname: Yamaguchi
authors.forename: Isao
authors.initials: I
authors.lastname: Hamaguchi
mesh_terms: D001782:Blood Donors; D017848:Blood-Borne Pathogens / Q000302:isolation & purification; D017931:DNA Primers / Q000235:genetics; D004279:DNA, Viral / Q000235:genetics / Q000302:isolation & purification; D005838:Genotype; D006801:Humans; D008403:Mass Screening / Q000379:methods*; D020411:Oligonucleotide Array Sequence Analysis / Q000295:instrumentation / Q000379:methods*; D016133:Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D014780:Viruses / Q000235:genetics / Q000302:isolation & purification*; D014944:World Health Organization
chemical_list: D017931:DNA Primers; D004279:DNA, Viral