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A method for detecting rash and fever illness‐associated viruses using multiplex reverse transcription polymerase chain reaction
Author(s) -
Matsushima Yuki,
Shimizu Tomomi,
Doi Ikuko,
Mizukoshi Fuminori,
Nagasawa Koo,
Ryo Akihide,
Shimizu Hideaki,
Kobayashi Masae,
Funatogawa Keiji,
Nagata Noriko,
Ishikawa Mariko,
Komane Ayako,
Okabe Nobuhiko,
Mori Yoshio,
Takeda Makoto,
Kimura Hirokazu
Publication year - 2017
Publication title -
microbiology and immunology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.664
H-Index - 70
eISSN - 1348-0421
pISSN - 0385-5600
DOI - 10.1111/1348-0421.12502
Subject(s) - biology , reverse transcriptase , multiplex , polymerase chain reaction , reverse transcription polymerase chain reaction , virology , multiplex polymerase chain reaction , rash , microbiology and biotechnology , genetics , gene , dermatology , medicine , messenger rna
In this study, a new multiplex RT‐PCR method for detecting various viral genes in patients with rash and fever illnesses (RFI s ) was constructed. New primer sets were designed for detection of herpes simplex viruses 1 and 2 (HSV1 and 2), and Epstein–Barr virus (EBV). The newly designed and previously reported primer sets were used to detect 13 types of RFI‐associated viruses by multiplex RT‐PCR assay systems. Moreover, to eliminate non‐specific PCR products, a double‐stranded specific DNase was used to digest double‐stranded DNA derived from the templates in clinical specimens. RFI‐associated viruses were detected in 77.0% of the patients (97/126 cases) by the presented method, multiple viruses being identified in 27.8% of the described cases (35/126 cases). Detected viruses and clinical diagnoses were compatible in 32.5% of the patients (41/126 cases). Sensitivity limits for these viruses were estimated to be 10 1 –10 3 copies/assay. Furthermore, non‐specific PCR products were eliminated by a double‐stranded specific DNase with no influence on sensitivity. These results suggest that this method can detect various RFI‐associated viruses in clinical specimens with high sensitivity and specificity.