Revision of new diagnostics for TB

1. Revision of new diagnostics for TB Churchyard GJ

2. Overview • Introduction • Xpert MTB/RIF • Line probe assays • Urine LAM • Diagnostics pipeline • Conclusion

3. Introduction • The global burden of TB is declining slowly • The lack of a rapid & accurate diagnostics is compromising progress towards TB elimination • Sputum microscopy remains the mainstay of TB diagnosis in many resource poor countries • Globally <10% of MDR TB patients are diagnosed & treated • HIV associated TB • Is more difficult to diagnose • If undiagnosed, is associated with a high mortality

4. Overview • Introduction • Xpert MTB/RIF • Line probe assays • Urine LAM • Diagnostics pipeline • Conclusion

5. Xpert MTB/RIF • Detects M. tuberculosis and common mutations that confer resistance to rifampin • Is a hemi-nested real-time PCR of MTB-specific region of rpoB gene, which is then probed with molecular beacons for mutations • Fully automated • Uses GeneXpert platform (Cepheid, CA)

6. Assay Procedure for the MTB/RIF Test

7. Evaluation Study of Xpert MTB/RIF • Cross-sectional study of diagnostic test accuracy • Population and Procedures • Peru, Azerbaijan, South Africa x 2, India • Adults with pulmonary TB symptoms • 3 sputa obtained (2 spot, 1 morning) Results • 1730 eligible participants • 976 with HIV status known; 40.2% HIV-positive (C. Boehme et al. NEJM 2010;363:1005)

8. Evaluation Study of Xpert MTB/RIF (C. Boehme et al. NEJM 2010;363:1005)

9. Evaluation Study of Xpert MTB/RIF (C. Boehme et al. NEJM 2010;363:1005)

10. Xpert MTB/RIF: false positives • WHO estimates Positive Predictive value at • >90% if greater than 15% of isolates are Rif resistant • <70% if less than 5% of isolates are Rif resistant • Further culture based DST to first and second line drugs recommended • Patients should receive MDR TB treatment pending further results • The software & cartridge have been redesigned to address these limitations

11. Xpert MTB/RIF: Operational performanceDetection of MTB (Lawn, Nicol. Future Micobiol. 2012; Dorman et al. PLoS ONE. 2012)

12. Xpert MTB/RIF Attributes & Advantages • Simple to perform, minimal training required • Not prone to cross-contamination • Requires minimal biosafety facilities • “Near-care” Shortcomings & Disadvantages • Complex instrument (calibration, power supply) • Cost for instrument • Cost of cartridges reduced to ~$10 • Single supplier

13. Xpert MTB/RIF • WHO expert group recommendations: • “Xpert should be used as the initial diagnostic test in individuals suspected of having MDR-TB or HIV-associated TB” (strong recommendation) • “Xpert may be used as a follow-on test to microscopy where MDR and/or HIV is of lesser concern, especially in smear-negative specimens” (conditional recommendation, recognizing resource implications)

14. Overview • Introduction • Xpert MTB/RIF • Line probe assays • Urine LAM • Diagnostics pipeline • Conclusion

15. GenoTypeMTBDRplus (Hain) Is a molecular line probe assay with probes for • MTB • Almost all of the common rifampicin resistance-conferring mutations • A subset of the mutations conferring resistance to isoniazid

16. GenoTypeMTBDRplus (Hain) • Validation study* showed that performance on • Smear positive specimens was good • Smear negative specimens was reasonable (* Barnard M. Am J RespirCrit Care Med 2008;177:787-792)

17. Genotype MTBDRplusvs MGIT for detection of MTB, by smear status • Missed 15% of RIF resistant and 37% of INH resistant TB • (Dorman S. PLoS One. 2012. In press)

18. Genotype MTBDRplus 2.0vsMGIT & clinical TB

19. GenoTypeMTBDRsl

20. GenoTypeMTBDRsl

21. GenoTypeMTBDRsl

22. GenoTypeMTBDRsl

23. Overview • Introduction • Xpert MTB/RIF • Line probe assays • Urine LAM • Diagnostics pipeline • Conclusion

24. Urine Assays for MycobacterialLipoarabinomannan (LAM) • LAM • A lipopolysaccharide component of MTB cell wall • Released from metabolically active or degraded MTB • Urine-based test • Urine easy to obtain • Lacks infection control issues of blood, sputum • Inverness: ELISA format • Alere: lateral flow

25. Urine Assays for MycobacterialLipoarabinomannan (LAM) • LAM • A lipopolysaccharide component of MTB cell wall • Released from metabolically active or degraded MTB • Urine-based test • Urine easy to obtain • Lacks infection control issues of blood, sputum • Inverness: ELISA format • Alere: lateral flow

26. Determine LAM lateral flow assay (Alere) • uses Determine testing platform • No sample processing; results in 25 minutes • Analytical sensitivity reported to be 0.25 ng/ml • Reporting scale: no band (neg), 1+ to 5+ (pos) sample application pad patient result window control window

27. Determine TB-LAM (* Dorman S, Interim unpublished data)

28. Overview • Introduction • Xpert MTB/RIF • Line probe assays • Urine LAM • Diagnostics pipeline • Conclusion

29. Global TB diagnostics pipeline

30. Conclusion • In many high burden countries sputum microscopy remains the mainstay of TB diagnosis • New diagnostics can substantially reduce the time to diagnosis of TB and drug resistant TB • A point of care test is urgently required

31. Acknowledgements • Susan Dorman