Chain Reaction PCR. PCR Transcription-Reverse Transcription Concerted reaction TRC. Gene Xpert Cepheid, USA. Satoshi MITARAI

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1 Satoshi MITARAI 2 1, ml 10, Polymerase Chain Reaction PCR 5, 6 PCR Realtime PCR Transcription-Reverse Transcription Concerted reaction TRC Gene Xpert Cepheid, USA TB-LAMP Gene Xpert Professor/Head, Department of Mycobacterium Reference and Research, Research Institute of Tuberculosis, Japan Anti-Tuberculosis Association (JATA) (3-1-24, Matsuyama, Kiyose-shi, Tokyo)

2 DNA FR -MTB TMA Hybridization protection assay HPA 5 TaqMan MTB Real-time PCR TaqMan probe 3 TRC Rapid M. TB TRC INAF probe, M. avium complex, M. kansasii 1.5 GENECUBE PCR QProbe, M. avium complex 50 Loopamp LAMP 50 Xpert MTB Rif 2 RFP rpob Real-time PCR Molecular Beacon CI, CI, RFP CI, CI, TB-LAMP PURE Procedure for Ultra Rapid Extraction, 1 Isothermal 9 DNA-DNA hybridization DDH, mg 16S rrna SPEED-OLIGO MYCOBACT- ERIA Vircell, Spain PCR GenoType Mycobacterium CM AS Hain Lifescience, Germany M. tuberculosis M. bovis M. africanum M. canetti M. caprae M. microti M. pinipedii M. tuberculosis BCG 15

3 RFP Xpert MTB Rif RFP DNA RNA RNA 2 α 1 1 -subunit rpob - subunit rpob RFP RNA RFP RRDR Refampicin Resistance Determining Region, 81bp RFP 95 rpob RFP Rifabutin 18 rpob Rif TB PCR rpob RRDR DNA Line Probe Assay ; LPA , 20 INH fura-katg, fabg1-inha, 21 European-American katg 315 inha -15 MTBDRplus Hain Lifescience, Germany LPA 22 embc-emba-embb / EMB R gid / SM R gyrb-gyra / FQs R rpob / RIF R rpsl / SM R M. tuberculosis H37Rv 4.4 Mbp rrs / KM R, AMK R, KM R, CPM R fabg1-inha / INH R oxyr ahpc / INH R pnca / PZA R fura-katg / INH R

4 PZA PZA ph 10 MGIT AST PZA PZA 24 PZA PZA pnca Pyrazinoic acid pnca PZA pnca PZA TB LPA 308 PZA PCR LPA Genotyping Restriction Fragment Length Polymorphisms : RFLP Gold standard Variable Number Tandem Repeat : VNTR VNTR PCR VNTR Whole Genome Sequence : WGS pnca 5 pnca 7 pnca 1-47 pnca 20 pnca 21 pnca 20 pnca 21 pnca : WT pnca : D12A pnca : 59 deletion c g232a pnca : deletion PZA TB PZA

5 1,, MGIT : , Tortoli E, Cichero P, Piersimoni C, Simonetti MT, Gesu G, Nista D. Use of BACTEC MGIT 960 for recovery of mycobacteria from clinical specimens : multicenter study. J Clin Microbiol. : , MAC ,,,,,. PCR. : , Bergmann JS, Woods GL. Clinical evaluation of the Roche AMPLICOR PCR Mycobacterium tuberculosis test for detection of M. tuberculosis in respiratory specimens. J Clin Microbiol. : , Bergmann JS, Keating WE, Woods GL. Clinical evaluation of the BDProbeTec ET system for rapid detection of Mycobacterium tuberculosis. J Clin Microbiol. : , Boehme C C, Nabeta P, Henostroza G, Raqib R, Rahim Z, Gerhardt M, Sanga E, Hoelscher M, Notomi T, Hase T, Perkins M D. Operational feasibility of using loop-mediated isothermal amplification for diagnosis of pulmonary tuberculosis in microscopy centers of developing countries. J Clin Microbiol. : , Mitarai S, Okumura M, Toyota E, Yoshiyama T, Aono A, Sejimo A, Azuma Y, Sugahara K, Nagasawa T, Nagayama N, Yamane A, Yano R, Kokuto H, Morimoto K, Ueyama M, Kubota M, Yi R, Ogata H, Kudoh S, Mori T. Evaluation of a simple loop-mediated isothermal amplification test kit for the diagnosis of tuberculosis. Int J Tuber Lung Dis. : , Boehme CC, Nabeta P, Hillemann D, Nicol MP, Shenai S, Krapp F, Allen J, Tahirli R, Blakemore R, Rustomjee R, Milovic A, Jones M, O Brien SM, Persing DH, Ruesch- Gerdes S, Gotuzzo E, Rodrigues C, Alland D, Perkins MD. Rapid molecular detection of tuberculosis and rifampin resistance. N Engl J Med. : , Peter JG, Theron G, Muchinga TE, Govender U, Dheda K. The diagnostic accuracy of urine-based Xpert MTB RIF in HIV-infected hospitalized patients who are smear-negative or sputum scarce. PLoS One. : e39966, Kazumi Y, Mitarai S. The evaluation of an identification algorithm for Myconacterium species using the 16S rrna coding gene and rpob. Int J Myco. : 21-28, Quezel-Guerraz NM, Arriaza MM, Avila JA, Sánchez- Yebra Romera WE, Martínez-Lirola MJ ; Indal-TB Group. Evaluation of the Speed-oligo Mycobacteria assay for identification of Mycobacterium spp. from fresh liquid and solid cultures of human clinical samples. Diagn Microbiol Infect Dis. : , Lee AS, Jelfs P, Sintchenko V, Gilbert GL. Identification of non-tuberculous mycobacteria : utility of the GenoType Mycobacterium CM AS assay compared with HPLC and 16S rrna gene sequencing. J Med Microbiol. : , ,,,,,. M. bovis Zhang Y, Yew WW. Mechanisms of drug resistance in Mycobacterium tuberculosis. Int J Tuberc Lung Dis. : , ,,,. Rifampicin Rifabutin. : , Hirano K, Abe C, Takahashi M. Mutations in the rpob gene of rifampin-resistant Mycobacterium tuberculosis strains isolated mostly in Asian countries and their rapid detection by line probe assay. J Clin Microbiol. : , Rossau R, Traore H, De Beenhouwer H, Mijs W, Jannes G, De Rijk P, Portaels F. Evaluation of the INNO-LiPA Rif. TB assay, a reverse hybridization assay for the simultaneous detection of Mycobacterium tuberculosis complex and its resistance to rifampin. Antimicrob Agents Chemother. : , Ando H, Kondo Y, Suetake T, Toyota E, Kato S, Mori T, Kirikae T. Identification of katg mutations associated with high-level isoniazid resistance in Mycobacterium tuberculosis. Antimicrob Agents Chemother. : , ,,,,,,. GenoType MTBDR plus : , Chedore P, Bertucci L, Wolfe J, Sharma M, Jamieson F. Potential for erroneous results indicating resistance when using the Bactec MGIT 960 system for testing susceptibility of Mycobacterium tuberculosis to pyrazinamide. J clin

6 microbiol. : , Werngren J, Sturegard E, Jureen P, Angebyc K, Hoffner S, Schon T. Reevaluation of the critical concentration for drug susceptibility testing of Mycobacterium tuberculosis against pyrazinamide using wild-type MIC distributions and pnca gene sequencing. Antimicrob Agent Chemother. : , Mitarai S, Kato S, Ogata H, Aono A, Chikamatsu K, Mizuno K, Toyota E, Sejimo A, Suzuki K, Yoshida S, Saito T, Moriya A, Fujita A, Sato S, Matsumoto T, Ano H, Suetake T, Kondo Y, Kirikae T, Mori T. Comprehensive multicenter evaluation of a new line probe assay kit for identification of Mycobacterium species and detection of drugresistant Mycobacterium tuberculosis. J Clin Microbiol. : , 2012.

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