Performance Evaluation of Xpert MTB/RIF Assay and Prevalence of rpoB Gene Mutation in <em> Mycobacterium Tuberculosis</em> in Clinical Isolates Quetta, Pakistan: A Cross Sectional Retrospective Cohort Study
Keywords:RRDR, rpoB, MDR-TB, 81 bp, XpertÂ® MTB/RIF.
With the use of Xpert assay, the detection mechanism of tuberculosis has completely transformed. Concurrently detecting mycobacterium and its resistance to rifampin (rif) which is also one of the primary marker for MDR-TB in addition to its surrogate activity against MTB in first line drugs. At large, resistance of mycobacterium strains against rifampicin is caused by rpoB gene mutations about 96.1 % globally while these mutations are generally situated at a site of 507-533rd residuals of amino acid in the rpoB gene of Mycobacterium, this region is also termed as Rifampicin resistance determining region (RRDR). In this study, we determined the mutations of various rpoB genes in different probes of 81 bp of RRD Region. A total of 6353 different specimens obtained at provincial reference laboratory of Fatima Jinnah general and chest hospital, Quetta (FJHQ), between Jan 2016 and Apr 2018 were examined by Xpert MTB/RIF assay. Altogether positive and negative samples for MTB were further examined with the light emitting diode (LED) fluorescence microscope. Pearson Chi square test was used to find out the associations between probe type and gender, treatment history and age group of the patients. Out of 6353 specimens 1297 MTB positive cases were detected by Xpert assay including 184 samples having both mycobacterium and RIF-resistance advised by mutations in 81 bp-RRD Region of rpoB gene which is further divided into five different probes. Further LED fluorescence microscope detected and confirmed 327 positive MTB samples out of 1297 specimens of Xpert assay. The associated probes mutation for rifampicin resistance were as follow: E (98/184), D (23/184), B (15/184), C (5/184), A (3/184), two probe mutation (5/184) in D&E (3/184) and C&D (2/184) and all probe mutation (35/184). The percentage of rpoB gene mutation in the studied population was 14.187 %. The commonest mutation associated with rpoB gene was in Probe E (53.2608%) also termed as 531 and 533 codons, additionally two probe mutation (2.7173%) in D&E (1.6304%) and C&D (1.0869%) and all probe mutation (19.0217%) were also detected that has never been detected before globally.
Helb D, Jones M, Story E, Boehme C, Wallace E, Ho K, Kop J, Owens MR, Rodgers R, Banada P, Safi H, Blakemore R, Lan NT, Jones-López EC, Levi M, Burday M, Ayakaka I, Mugerwa RD, McMillan B, Winn-Deen E, Christel L, Dailey P, Perkins MD, Persing DH, Alland D: Rapid detection of Mycobacterium tuberculosis and rifampin resistance by use of on-demand, near-patient technology. J Clin Microbiol 2010, 48:229–37.
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, Rodriques C, Alland D, Perkins MD: Rapid molecular detection of tuberculosis and rifampin resistance. N Engl J Med 2010, 363:1005–15.
Drobniewski FA, Wilson SM: The rapid diagnosis of isoniazid and rifampin resistance in Mycobacterium tuberculosis: a molecular story. J. Med. Microbiol 1998, 47:189–96.
Ramaswamy S, Musser JM: Molecular genetic basis of antimicrobial agent resistance in Mycobacterium tuberculosis. Tuberc Lung Dis 1998, 79:3–29.
Organization WH. World Health Organization Global Tuberculosis Report 2013. Geneva, Switzerland: World Health Organization, WHO Press; 2013.
Li W, Wang S, Li C, Liu Y, Shen G, Zhang X, Niu T, Gao Q, Van Soolingen D, Kremer K. Molecular epidemiology of Mycobacterium tuberculosis in China: a nationwide random survey in 2000. Int J Tuberc Lung Dis. 2005;9(12):1314–9.
Brossier F, Veziris N, Truffot-Pernot C, Jarlier V, Sougakoff W. Performance of the genotype MTBDR line probe assay for detection of resistance to rifampin and isoniazid in strains of Mycobacterium tuberculosis with lowand high-level resistance. J Clin Microbiol. 2006;44(10):3659–64.
Ocheretina O, Escuyer VE, Mabou M-M, Royal-Mardi G, Collins S, Vilbrun SC, Pape JW, Fitzgerald DW. Correlation between genotypic and phenotypic testing for resistance to rifampin in Mycobacterium tuberculosis clinical isolates in Haiti: investigation of cases with discrepant susceptibility results. PLoS One. 2014;9(3):e90569.
Jones-López E, Manabe YC, Palaci M, Kayiza C, Armstrong D, Nakiyingi L, Ssengooba W, Gaeddert M, Kubiak R, Almeida JP, Alland D, Dietze R, Joloba LM, Ellner JJ, Dorman SE: Prospective cross-sectional evaluation of the small membrane filtration method for diagnosis of pulmonary tuberculosis. J Clin Microbiol 2014, 52(7):2513–20.
Ioerger TR, Koo S, No E-G, Chen X, Larsen MH, Jacobs Jr WR, Pillay M, Sturm AW, Sacchettini JC. Genome analysis of multi-and extensively-drug-resistant tuberculosis from KwaZulu-Natal, South Africa. PloS One. 2009;4(11):e7778.
Yue J, Shi W, Xie J, Li Y, Zeng E: Mutations in the rpoB Gene of MultidrugResistant Mycobacterium tuberculosis Isolates from China. J Clin Microbiol 2003, 41(5):2209–12.
Ullah I, Shah AA, Basit A, Ali M, Khan A, Ullah U, Ihtesham M, Mehreen S, Mughal A, Javaid A. Rifampicin resistance mutations in the 81 bp RRDR of rpoB gene in Mycobacterium tuberculosis clinical isolates using Xpert MTB/RIF in Khyber Pakhtunkhwa, Pakistan: a retrospective study. BMC Infect Dis. 2016; 16:413.
Mboowa G, Namaganda C, Ssengooba W. Rifampicin resistance mutations in the 81 bp RRDR of rpoB gene in Mycobacterium tuberculosis clinical isolates using Xpert MTB/RIF in Kampala, Uganda: a retrospective study. BMC Infect Dis. 2014; 14(1):481.
Khan SN, Niemann S, Gulfraz M, Qayyum M, Siddiqi S, Mirza ZS, Tahsin S, Ebrahimi-Rad M, Khanum A. Molecular characterization of multidrug resistant isolates of Mycobacterium tuberculosis from patients in Punjab, Pakistan. Pakistan J Zool. 2013;45(1):93–100.
Boehme CC, Nicol MP, Nabeta P, Michael JS, Gotuzzo E, Tahirli R, Gler MT, Blakemore R, Worodria W, Gray C. Feasibility, diagnostic accuracy, and effectiveness of decentralised use of the Xpert MTB/RIF test for diagnosis of tuberculosis and multidrug resistance: a multicentre implementation study. The lancet. 2011;377(9776):1495–505.
Moure R, Muñoz L, Torres M, Santin M, Martín R, Alcaide F. Rapid detection of Mycobacterium tuberculosis complex and rifampin resistance in smearnegative clinical samples by use of an integrated real-time PCR method. J Clin Microbiol. 2011;49(3):1137–9.
Liu Q, Zhu L, Shao Y, Song H, Li G, Zhou Y, Shi J, Zhong C, Chen C, Lu W. Rates and risk factors for drug resistance tuberculosis in Northeastern China. BMC Public Health. 2013;13(1):1171.
Luiz RSS, Suffys P, Barroso EC, Kerr LRFS, Duarte CR, Freitas MVC, Mota RMS, Frota CC. Genotyping and drug resistance patterns of Mycobacterium tuberculosis strains observed in a tuberculosis high-burden municipality in Northeast, Brazil. Braz J Infect Dis. 2013;17(3):338–45.
Javaid A, Hasan R, Zafar A, Ghafoor A, Pathan A, Rab A, Sadiq A, Akram C, Burki I, Shah K. Prevalence of primary multidrug resistance to antituberculosis drugs in Pakistan. Int J Tuberc Lung Dis. 2008;12(3):326–31.
Ullah I, Javaid A, Tahir Z, Ullah O, Shah AA, Hasan F, Ayub N. Pattern of Drug Resistance and Risk Factors Associated with Development of Drug Resistant Mycobacterium tuberculosis in Pakistan. PLoS One. 2016;11(1):e0147529.
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Copyright (c) 2021 Ashraf Ullah Khan, Shereen Khan, Hidayatullah, Zabi Ullah
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