Development of an assay for the detection and quantification of the measles virus nucleoprotein (N) gene using real-time reverse transcriptase PCR.
We developed a new quantification method for the measles virus (MeV) nucleoprotein (N) gene using real-time reverse transcriptase PCR. This method allowed us to quantify 10(1)-10(7) copies per reaction (corresponding to 5x10(-1)-5x10(5) copies microl(-1)) of the MeV N gene. We also quantified the MeV N gene from the throat swabs of 22 patients with measles as well as the MeV genotypes A, D3, D5, D9 and H1 in viral suspensions derived from MeV-infected cells. As a result, 3.9x10(3)-5.2x10(6) copies ml(-1) and 7.4x10(7)-2.0x10(8) copies ml(-1) of the MeV genomes (N gene) were detected in the throat swabs and viral suspensions, respectively. No other viruses (enteroviruses, respiratory syncytial virus, human metapneumovirus or mumps virus) were detected in the assay. The results suggest that this method is applicable to the detection and quantification of some genotypes of MeV.
Infectious Disease Surveillance Center, National Institute of Infectious Diseases, 4-7-1 Gakuen, Musashimurayama, Tokyo 208-0011, Japan.
Miho
M
Akiyama
Infectious Disease Surveillance Center, National Institute of Infectious Diseases, 4-7-1 Gakuen, Musashimurayama, Tokyo 208-0011, Japan.
Hirokazu
H
Kimura
Gunma Prefectural Institute of Public Health and Environmental Sciences, 378 Kamioki-machi, Maebashi, Gunma 371-0052, Japan.
Hiroyuki
H
Tsukagoshi
Okinawa Prefectural Institute of Public Health and Environmental Sciences, 2085 Ozato, Nanjo, Okinawa 910-1202, Japan.
Katsuya
K
Taira
Yamagata Prefectural Institute of Public Health, 1-6-6 Tokamachi, Yamagata 990-0031, Japan.
Katsumi
K
Mizuta
Gunma Prefectural Institute of Public Health and Environmental Sciences, 378 Kamioki-machi, Maebashi, Gunma 371-0052, Japan.
Mika
M
Saitoh
Molecular and Cellular Biology Division, Applied Biosystems Japan Ltd, 4-5-4 Hatchobori, Chuo-ku, Tokyo 104-0032, Japan.
Manami
M
Nagano
Yamagata Prefectural Institute of Public Health, 1-6-6 Tokamachi, Yamagata 990-0031, Japan.
Asuka
A
Sutoh
Department of Virology III, National Institute of Infectious Diseases, 4-7-1 Gakuen, Musashimurayama, Tokyo 208-0011, Japan.
Masahiro
M
Noda
Gunma Prefectural Institute of Public Health and Environmental Sciences, 378 Kamioki-machi, Maebashi, Gunma 371-0052, Japan.
Yukio
Y
Morita
Molecular and Cellular Biology Division, Applied Biosystems Japan Ltd, 4-5-4 Hatchobori, Chuo-ku, Tokyo 104-0032, Japan.
Osamu
O
Sakatsume
Infectious Disease Surveillance Center, National Institute of Infectious Diseases, 4-7-1 Gakuen, Musashimurayama, Tokyo 208-0011, Japan.
Nobuhiko
N
Okabe
Department of Virology III, National Institute of Infectious Diseases, 4-7-1 Gakuen, Musashimurayama, Tokyo 208-0011, Japan.
Masato
M
Tashiro
D017931:DNA Primers; D005784:Gene Amplification; D005814:Genes, Viral / Q000235:genetics; D005838:Genotype; D006801:Humans; D008457:Measles / Q000821:virology*; D008459:Measles virus / Q000235:genetics* / Q000302:isolation & purification; D008969:Molecular Sequence Data; D019590:Nucleocapsid Proteins / Q000235:genetics*; D010614:Pharynx / Q000821:virology; D010957:Plasmids; D012367:RNA, Viral / Q000235:genetics / Q000302:isolation & purification; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*
D017931:DNA Primers; D019590:Nucleocapsid Proteins; D012367:RNA, Viral