IJCRR - 8(23), December, 2016
RAPID DETECTION OF MULTI-DRUG RESISTANT TUBERCULOSIS USING DIRECT DRUG SUSCEPTIBILITY TESTING AND LINE PROBE ASSAY
Author: Pranali Medhekar, Nilma Hirani, Sarala Menon, Ameeta Joshi, Abhay Chowdhary
Introduction: Tuberculosis, particularly multi-drug resistant tuberculosis (MDR TB), is a leading cause of morbidity and mortality in developing countries. Conventional methods of DST have a long turnaround time of around 2-3 months. Rapid detection of MDR TB can be achieved by direct tests, liquid based indirect susceptibility tests and molecular line probe assays. This study aims to evaluate the utility of Direct DST and line probe assay (LPA) for rapid detection of MDR TB.
Methodology: A total of 510 sputum concentrates (smear 1+/ more) were subjected to Direct DST and LPA. Conventional DST was put up from the primary cultures of all 510 strains using economic variant of 1% proportion method, considered the gold standard.
Results: Direct DST showed 100% specificity and sensitivity of 98.7%, 97.3% and 97.5% respectively for detection of Isoniazid (INH), Rifampicin (RIF) resistance and for MDR strains. LPA showed specificity of 98.4%, 96.8%, 97% and sensitivity of 97.5%, 98.9% and 98.9% for detection of INH, RIF resistance and MDR strains respectively. The commonest mutation pattern detected in our region was S531L (MUT3) for rpoB gene locus signifying RIF resistance + S315T1 (MUT 1) for katG gene locus signifying high level INH resistance.
Conclusion: Direct DST can be used as a good alternative for rapid detection of MDR-TB in resource poor settings. LPA is definitely a good tool, but requires requisite laboratory infrastructure and trained man-power. From our study, rifampicin resistance can be considered as a surrogate marker of MDR TB.
Keywords: MDR TB, Direct DST, LPA
Pranali Medhekar, Nilma Hirani, Sarala Menon, Ameeta Joshi, Abhay Chowdhary. RAPID DETECTION OF MULTI-DRUG RESISTANT TUBERCULOSIS USING DIRECT DRUG SUSCEPTIBILITY TESTING AND LINE PROBE ASSAY International Journal of Current Research and Review. 8(23), December, 29-38
1. WHO: Global tuberculosis control 2008: surveillance, planning, financing, Geneva, Switzerland: World Health Organisation; Publication no, WHO/HTM/TB/2008.393, 2008.
2. XDR-TB (Extensively Drug Resistant Tuberculosis). WHO report 2006.
3. WHO: Global Tuberculosis Report, 2015
4. Blumberg HM, Burman WJ. Chaisson RE , Dailey CL, Etkind SC ,Fujivara P, et al: American Thoracic Society/ Centers For Disease Control and Prevention/ Infectious Diseases Society of America; treatment of Tuberculosis . Am. J Resp Crit Care Med 2003, 167(4) :603-662.
5. Manual of Standard Operating Procedures (SOPs). Culture of Mycobacterium tuberculosis and Drug Susceptibility Testing on solid medium. 1-142(2009) available at www.tbcindia.org
6. Mathew S, Nalini SM, Rahman F, Sundaram V and Paramasivan CN. Simple Direct Susceptibility Tests on sputum samples for early detection of resistance in tubercle bacilli: Indian J Tuber 2007; 54: 184-189.
7. Angeby KA , Klintz L, Hoffner SE :Rapid And Inexpensive Drug Susceptibility Testing of Mycobacterium tuberculosis with a nitrate reductase assay, J Clin Microbiol 2008 ,40(2) :553-555
8. Woods GL Susceptibility Testing for Mycobacteria Clin inf Dis 2000, 31 (5);209-1215.
9. Miotto P, PianaF, Cinllo DM, Miglori GB; Genotype MTBDRplus: a further step Toward rapid identification of drug resistant Mycobacterium tuberculosis J Clin Microbiol 2008, 46(1); 393-394.
10. Sharma M, Sethi S, Mishra B, Sengupta C, Sharma SK; Rapid detection of mutations in rpoB gene of rifampicin resistant Mycobacterium tuberculosis strains by line probe assay;Indian J Med Res 2003;pp 76-80.
11. Revised National Tuberculosis Control Programme: Guidelines on Programmatic Management of Drug Resistant TB (PMDT) in India May 2012.
12. Koneman’s Colour Atlas and Textbook of Diagnostic Microbiology. Washington Winn, Jr. Stephen Allen, William Janda, Elmer Koneman Gary Procop, Paul Schreckenberger, Gail Woods. 2006; Sixth Edition.
13. GenoType MTBDRplus,version 1.0 (product insert). (Internet) Feb 2007 (updated 2007 Apr 3; accessed 2007 May 9). Nehren, Germany; Hain Lifescience, GmbH. Available from: http:// www.hain-lifescience.com/pdf/304xx_pbl.pdf
14. Bernard N, Albert H, Coetzee G Rapid Molecular screening of multi drug resistant Tuberculosis in A high volume public health laboratory in South Africa; Am J Resp Cri Care Med, 2008,pp 787-792. 35 Int J Cur Res Rev | Vol 8 • Issue 23 • December 2016 Medhekar et.al.: Rapid detection of multi-drug resistant tuberculosis using direct drug susceptibility testing and line probe assay
15. Farooqui J. Q. Erun Khan, Syed Muhammed Zaheer Alam, Asho Ali, Zahra Hasan, Rumina Hasan, Line Probe Assay for detection of rifampicin and isoniazid resistant tuberculosis in Pakistan; Journal of Pakistan Medical Association; vol 62 No. 8, August 2012.
16. Buyankhishig B, T. Oyuntuya, B. Tserelmaa, J. Sarantuya, Marilla G. Lucero, S. Mitarai. Rapid molecular testing for multiresistant tuberculosis in Mongolia: A diagnostic study; International Journal of Mycobacteriology , vol 1, Issue 1, March 2012i
17. Raveendran R, Wattal C, Oberoi JK, Goel N, Datta S, Prasad KJ. Utility of Geno Type MTBDRplus assay in rapid diagnosis of multidrug resistant tuberculosis at a tertiary care centre in India; Indian Journal of Medical Microbiology vol 30, Issue 1, 2012.
18. Mai NT Huyen, Edine W Tiemersma, Nguyen TN Lan, Frank GJ Cobelens, Nguyen H Dung, Dinh N Sy, et al: Validation of the GenoType® MTBDRplus assay for diagnosis of multidrug resistant tuberculosis in South Vietnam; BMC Infectious Diseases 2010, 10:149.
19. Miotto P, Saleri N, Dembele M, Ouedraogo M, Baooum G, Pinsi G, et al. Molecular detection of rifampicin and isoniazid resistance to guide chronic TB patient management in Burkina Faso. BMC Infect Dis 2009;9:142.
20. Bahrmand AR, Titov LP, Tasbiti AH, Yari S, Graviss EA,. Highlevel rifampin resistance correlates with multiple mutations in the rpo B gene of pulmonary tuberculosis isolates from Afganistan border of Iran. J Clin Microbiol 2009; 47: 2744-50.
21. Van Rie A, Warren R, Mshanga I,Jordaan A, van der Spuy GD, Richardson M, Simpson J, Gie RP, Enarson DA, Beyer’s N et al. Analysis for a limited number of gene codons can predict drug resistance of Mycobacteriun tuberculosis in a high incidence community. J Clin Microbiol 2001;39-636-641.
22. Mokrousov I, Narvaskaya O, Otten T, Limenschenko E, Steklova L, Vyshnevskiy B. High prevalence of katG Ser315Thr substitution among isoniazid resistant Mycobacterium Tuberculosis clinical isolates from Northwestern Russia, 1996-2001. Antimicrob Agents Chemother 2002;46:1417-1424.
23. Kiepiela P, Bishop KS, Smith AN, Roux L, York DF. Genomic mutations in the katG, inhA and ahpC are useful for the prediction of isoniazid resistance in M. Tuberculosis isolates from KWA Zulu Natal, South Africa. Tuber Lung Dis 2000;80:87-56
24. Miotto P, Piana F, Penati V, Canducci F, Migliori GB, Cirillo DM. Use of Genotype MTBDR assay for molecular detection of rifampicin and isonoazid resistance in Mycobacterium tuberculosis in clinical strains isolated in Italy. J Clin Micobiol 2006;44:2485-2491.
25. Hilleman D, Rusch-Gerdes S, Richter E. Application of Genotype MTBDR assay directly on sputum samples. Int J Tuberc Lung Dis 2006;10:1057-1059.
26. Somoskovi A, Dormandy J, Mitsani D, Rivenberg J, Salfinger M. Use of smear-positive samples to assess the PCR-based genotype MTBDR assay for rapid, direct detection of the Mycobacterium tuberculosis complex as well as its resistance to isoniazid and rifampicin. J Clin Microbiol 2006;44:4459-4463.
27. Somoskovi A, Parsons LM, Salfinger M, The molecular basis of resistance to isoniazid, rifampin and pyrizinamide in Mycobacterium tuberculosis. Resspir Res 2001; 2:164-8.
28. Ramaswamy S, Musser JM. Molecular genetic basis of antimicrobial agent resistance in Mycobacterium tuberculosis: 1998 update. Tuber Lung Dis 1998; 79:3-29.
29. Molecular Line Probe Assays for Rapid Screening of Patients at Risk of Multi-Drug Resistant Tuberculosis (MDR-TB): WHO Expert group report.