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Journal of Clinical Microbiology, April 2005, p. 1581-1586, Vol. 43, No. 4
0095-1137/05/$08.00+0     doi:10.1128/JCM.43.4.1581-1586.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.

Loop-Mediated Isothermal Amplification Method Targeting the lytA Gene for Detection of Streptococcus pneumoniae

Mitsuko Seki,1,2* Yoshihisa Yamashita,3 Hirotaka Torigoe,1 Hiromasa Tsuda,1 Setsuko Sato,1 and Masao Maeno1,2

Department of Oral Health Sciences,1 Division of Functional Morphology, Dental Research Center, Nihon University School of Dentistry, Tokyo,2 Department of Preventive Dentistry, Faculty of Dental Science, Kyushu University, Fukuoka, Japan3

Received 30 August 2004/ Returned for modification 14 October 2004/ Accepted 24 November 2004

It is difficult to separate Streptococcus pneumoniae from the genotypically similar species Streptococcus mitis and Streptococcus oralis, which are commensals of the human oral cavity. A novel nucleic acid amplification technique, loop-mediated isothermal amplification (LAMP), which amplifies DNA under isothermal conditions (63°C) with high specificity, efficiency, and rapidity, was examined regarding its applicability for detecting S. pneumoniae. An S. pneumoniae-specific LAMP primer targeting the lytA gene was designed. The primer specificity was validated using 10 Streptococcus and 7 non-Streptococcus species. Within 60 min, the assay could detect 10 or more copies of purified S. pneumoniae DNA with a sensitivity 1,000 times that of conventional PCR. Clinical isolates of 21 other strains (3 S. oralis, 17 S. mitis, and 1 Streptococcus species) that harbor virulence-factor-encoding genes (lytA or ply) were tried to differentiate S. pneumoniae. The detection of S. pneumoniae in clinical isolates was more selective using the LAMP method than using conventional PCR. Therefore, LAMP appears to be a sensitive and reliable means of diagnosing S. pneumoniae infection.

* Corresponding author. Mailing address: Department of Oral Health Sciences, Nihon University School of Dentistry, 1-8-13 Kanda-Surugadai, Chiyoda-ku, Tokyo 101-8310, Japan. Phone: 81-3-3219-8128. Fax: 81-3-3219-8138. E-mail: seki-m{at}dent.nihon-u.ac.jp.

Journal of Clinical Microbiology, April 2005, p. 1581-1586, Vol. 43, No. 4
0095-1137/05/$08.00+0     doi:10.1128/JCM.43.4.1581-1586.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.



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