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Degenerate polymerase chain reaction strategy with DNA microarray for detection of multiple and various subtypes of virus during blood screening
Author(s) -
Takizawa Kazuya,
Nakashima Tatsuo,
Mizukami Takuo,
Kuramitsu Madoka,
Endoh Daiji,
Kawauchi Shigeto,
Sasaki Kohsuke,
Momose Haruka,
Kiba Yoshiharu,
Mizutani Tetsuya,
Furuta Rika A.,
Yamaguchi Kazunari,
Hamaguchi Isao
Publication year - 2013
Publication title -
transfusion
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.045
H-Index - 132
eISSN - 1537-2995
pISSN - 0041-1132
DOI - 10.1111/trf.12193
Subject(s) - polymerase chain reaction , virus , virology , real time polymerase chain reaction , dna , microbiology and biotechnology , biology , computational biology , gene , genetics
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. Study Design and 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 B 19 ( PVB 19), 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. Conclusion 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 , PVB 19, 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.