Transcript This is the title slide

Revision of new diagnostics for TB
Churchyard GJ
Overview
 Introduction
 Xpert MTB/RIF
 Line probe assays
 Urine LAM
 Diagnostics pipeline
 Conclusion
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
Overview
 Introduction
 Xpert MTB/RIF
 Line probe assays
 Urine LAM
 Diagnostics pipeline
 Conclusion
Xpert MTB/RIF
 Detects M. tuberculosis and common
mutations that confer resistance to rifampin
 Is a hemi-nested real-time PCR of MTBspecific region of rpoB gene, which is then
probed with molecular beacons for
mutations
 Fully automated
 Uses GeneXpert platform (Cepheid, CA)
Assay Procedure for the MTB/RIF Test
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)
Evaluation Study of Xpert MTB/RIF
# Xpert
tests per
participant
Sensitivity
Specificity
All CX POS
SM POS,
CX POS
SM NEG,
CX POS
1 sputum
675/732
92.2%
551/561
98.2%
124/171
72.5%
604/609
99.2%
3 sputa
723/741
97.6%
566/567
99.8%
157/174
90.2%
604/616
98.1%
(C. Boehme et al. NEJM 2010;363:1005)
Evaluation Study of Xpert MTB/RIF
Xpert SENSITIVITY for Xpert SPECIFICITY for
Rifampin Resistance Rifampin Resistance
# correct/# total
%
# correct/# total
%
Phenotypic DST
200/205
97.6%
505/515
98.1%
Phenotypic DST
and Discrepant
Resolution by
Sequencing
209/211
99.1%
506/506
100.0%
(C. Boehme et al. NEJM 2010;363:1005)
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
Xpert MTB/RIF: Operational performance
Detection of MTB
Sensitivity
Specificity
90.3%
99%
58%-73%
99.2%-93.9%
Extrapulmonary TB 53%-95%
98.2%-100%
Active case finding 62.6%
99.6%
Decentralised
implementation
HIV-associated TB
(Lawn, Nicol. Future Micobiol. 2012; Dorman et al. PLoS ONE. 2012)
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
Xpert MTB/RIF
 WHO expert group recommendations:
 “Xpert should be used as the initial diagnostic test
in individuals suspected of having MDR-TB or HIVassociated 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)
Overview
 Introduction
 Xpert MTB/RIF
 Line probe assays
 Urine LAM
 Diagnostics pipeline
 Conclusion
GenoType MTBDRplus (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
GenoType MTBDRplus (Hain)
Ling D. 2008
Rifampicin
Isoniazid
Sensitivity
98.4%
88.7%
Specificity
98.9%
99.2%
Validation study* showed that performance on
• Smear positive specimens was good
• Smear negative specimens was reasonable
(* Barnard M. Am J Respir Crit Care Med 2008;177:787-792)
Genotype MTBDRplus vs 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)
Genotype MTBDRplus 2.0 vs
MGIT & clinical TB
Crudu. JCM, 2012
MTB
Overall
Smear negative
Rifampicin resistance
Overall
Smear negative
Isoniazid resistance
Overall
Smear negative
Sensitivity
Specificity
87.6%
79.8%
99.2%
99.2%
94.3%
90.7%
96.0%
96.0%
95.8%
93.5%
88.9%
82.3%
GenoType MTBDRsl
Drug
Kontsevaya,I. JCM. 2011
Fluoroquinolone
Kiet,V.S. JCM. 2010
Fluoroquinolone
kanamycin
Ethambutol
Hillemann,D. JCM. 2009
Fluoroquinolone
Amikacin
Ethambutol
Sensitivity
Specificity
86.2%
100%
75.6%
100%
100%
64.2%
100%
100%
88.9%
75.0%
38.5%
GenoType MTBDRsl
Drug
Kontsevaya,I. JCM. 2011
Fluoroquinolone
Kiet,V.S. JCM. 2010
Fluoroquinolone
kanamycin
Ethambutol
Hillemann,D. JCM. 2009
Fluoroquinolone
Amikacin
Ethambutol
Sensitivity
Specificity
86.2%
100%
75.6%
100%
100%
64.2%
100%
100%
88.9%
75.0%
38.5%
GenoType MTBDRsl
Drug
Kontsevaya,I. JCM. 2011
Fluoroquinolone
Kiet,V.S. JCM. 2010
Fluoroquinolone
kanamycin
Ethambutol
Hillemann,D. JCM. 2009
Fluoroquinolone
Amikacin
Ethambutol
Sensitivity
Specificity
86.2%
100%
75.6%
100%
100%
64.2%
100%
100%
88.9%
75.0%
38.5%
GenoType MTBDRsl
Drug
Kontsevaya,I. JCM. 2011
Fluoroquinolone
Kiet,V.S. JCM. 2010
Fluoroquinolone
kanamycin
Ethambutol
Hillemann,D. JCM. 2009
Fluoroquinolone
Amikacin
Ethambutol
Sensitivity
Specificity
86.2%
100%
75.6%
100%
100%
64.2%
100%
100%
88.9%
75.0%
38.5%
Overview
 Introduction
 Xpert MTB/RIF
 Line probe assays
 Urine LAM
 Diagnostics pipeline
 Conclusion
Urine Assays for Mycobacterial
Lipoarabinomannan (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
Urine Assays for Mycobacterial
Lipoarabinomannan (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
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
Determine TB-LAM
Author/
Year
N
Setting
Sensitivity
Overall CD4<100
45%
96%
Peter, 2012
335
Inpatients
Lawn, 2012
Dorman S,
2012 *
516
561
ART clinic
Outpatients
Inpatients
28%
45%
52%
80%
(* Dorman S, Interim unpublished data)
Specificity
99%
90%
Overview
 Introduction
 Xpert MTB/RIF
 Line probe assays
 Urine LAM
 Diagnostics pipeline
 Conclusion
Global TB diagnostics pipeline
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
Acknowledgements
 Susan Dorman