Development and evaluation of a loop-mediated isothermal amplification assay for rapid detection of Mycoplasma pneumoniae

1 Department of Infection Control and Prevention, University of Tokyo Hospital, Bunkyo-ku, Tokyo 113-8655, Japan 2 Department of Microbiology and Immunology, Graduate School of Allied Health Sciences, Tokyo Medical and Dental University, Bunkyo-ku, Tokyo 113-8510, Japan 3 Kitasato Institute for Life Sciences and Graduate School of Infection Control Sciences, Kitasato University, Minato-ku, Tokyo 108-8641, Japan Correspondence Ryoichi Saito saito-lab{at}h.u-tokyo.ac.jp Received March 3, 2005 Accepted July 29, 2005 A loop-mediated isothermal amplification (LAMP) assay for the rapid detection of Mycoplasma pneumoniae was developed and evaluated. The assay specifically amplified only M. pneumoniae sequences, and no cross-reactivity was observed for other Mycoplasma species or respiratory bacterial species. The detection limit for this assay was found to be 2 x 10 2 copies, corresponding to 2–20 colour changing units of M. pneumoniae in 1 h, as observed in a real-time turbidimeter and electrophoretic analysis. The accuracy of the LAMP reaction was confirmed by restriction endonuclease analysis as well as direct sequencing of the amplified product. The assay was applied to 95 nasopharyngeal swab samples collected from patients or from healthy individuals, and compared to a real-time PCR assay in-house. A concordance of 100 % was observed between the two assays. The LAMP assay is easy to perform, shows a rapid reaction and is inexpensive. It may therefore be applied in the routine diagnosis of M. pneumoniae infection in the clinical laboratory. Abbreviation: LAMP, loop-mediated isothermal amplification.