The diagnostic accuracy of the GenoType(®) MTBDRsl assay for the detection of resistance to second-line anti-tuberculosis drugs

Abstract

Background: Accurate and rapid tests for tuberculosis (TB) drug resistance are critical for improving patient care and decreasing the transmission of drug-resistant TB. Genotype(®)MTBDRsl (MTBDRsl) is the only commercially-available molecular test for detecting resistance in TB to the fluoroquinolones (FQs; ofloxacin, moxifloxacin and levofloxacin) and the second-line injectable drugs (SLIDs; amikacin, kanamycin and capreomycin), which are used to treat patients with multidrug-resistant (MDR-)TB.

Objectives: To obtain summary estimates of the diagnostic accuracy of MTBDRsl for FQ resistance, SLID resistance and extensively drug-resistant TB (XDR-TB; defined as MDR-TB plus resistance to a FQ and a SLID) when performed (1) indirectly (ie on culture isolates confirmed as TB positive) and (2) directly (ie on smear-positive sputum specimens).To compare summary estimates of the diagnostic accuracy of MTBDRsl for FQ resistance, SLID resistance and XDR-TB by type of testing (indirect versus direct testing).The populations of interest were adults with drug-susceptible TB or drug-resistant TB. The settings of interest were intermediate and central laboratories.

Search methods: We searched the following databases without any language restriction up to 30 January 2014: Cochrane Infectious Diseases Group Specialized Register; MEDLINE; EMBASE; ISI Web of Knowledge; MEDION; LILACS; BIOSIS; SCOPUS; the metaRegister of Controlled Trials; the search portal of the World Health Organization International Clinical Trials Registry Platform; and ProQuest Dissertations & Theses A&I.

Selection criteria: We included all studies that determined MTBDRsl accuracy against a defined reference standard (culture-based drug susceptibility testing (DST), genetic testing or both). We included cross-sectional and diagnostic case-control studies. We excluded unpublished data and conference proceedings.

Data collection and analysis: For each study, two review authors independently extracted data using a standardized form and assessed study quality using the Quality Assessment of Diagnostic Accuracy Studies (QUADAS-2) tool. We performed meta-analyses to estimate the pooled sensitivity and specificity of MTBDRsl for FQ resistance, SLID resistance, and XDR-TB. We explored the influence of different reference standards. We performed the majority of analyses using a bivariate random-effects model against culture-based DST as the reference standard.

Main results: We included 21 unique studies: 14 studies reported the accuracy of MTBDRsl when done directly, five studies when done indirectly and two studies that did both. Of the 21 studies, 15 studies (71%) were cross-sectional and 11 studies (58%) were located in low-income or middle-income countries. All studies but two were written in English. Nine (43%) of the 21 included studies had a high risk of bias for patient selection. At least half of the studies had low risk of bias for the other QUADAS-2 domains.As a test for FQ resistance measured against culture-based DST, the pooled sensitivity of MTBDRsl when performed indirectly was 83.1% (95% confidence interval (CI) 78.7% to 86.7%) and the pooled specificity was 97.7% (95% CI 94.3% to 99.1%), respectively (16 studies, 1766 participants; 610 confirmed cases of FQ-resistant TB; moderate quality evidence). When performed directly, the pooled sensitivity was 85.1% (95% CI 71.9% to 92.7%) and the pooled specificity was 98.2% (95% CI 96.8% to 99.0%), respectively (seven studies, 1033 participants; 230 confirmed cases of FQ-resistant TB; moderate quality evidence). For indirect testing for FQ resistance, four (0.2%) of 1766 MTBDRsl results were indeterminate, whereas for direct testing 20 (1.9%) of 1033 were MTBDRsl indeterminate (P < 0.001).As a test for SLID resistance measured against culture-based DST, the pooled sensitivity of MTBDRsl when performed indirectly was 76.9% (95% CI 61.1% to 87.6%) and the pooled specificity was 99.5% (95% CI 97.1% to 99.9%), respectively (14 studies, 1637 participants; 414 confirmed cases of SLID-resistant TB; moderate quality evidence). For amikacin resistance, the pooled sensitivity and specificity were 87.9% (95% CI 82.1% to 92.0%) and 99.5% (95% CI 97.5% to 99.9%), respectively. For kanamycin resistance, the pooled sensitivity and specificity were 66.9% (95% CI 44.1% to 83.8%) and 98.6% (95% CI 96.1% to 99.5%), respectively. For capreomycin resistance, the pooled sensitivity and specificity were 79.5% (95% CI 58.3% to 91.4%) and 95.8% (95% CI 93.4% to 97.3%), respectively. When performed directly, the pooled sensitivity for SLID resistance was 94.4% (95% CI 25.2% to 99.9%) and the pooled specificity was 98.2% (95% CI 88.9% to 99.7%), respectively (six studies, 947 participants; 207 confirmed cases of SLID-resistant TB, 740 SLID susceptible cases of TB; very low quality evidence). For indirect testing for SLID resistance, three (0.4%) of 774 MTBDRsl results were indeterminate, whereas for direct testing 53 (6.1%) of 873 were MTBDRsl indeterminate (P < 0.001).As a test for XDR-TB measured against culture-based DST, the pooled sensitivity of MTBDRsl when performed indirectly was 70.9% (95% CI 42.9% to 88.8%) and the pooled specificity was 98.8% (95% CI 96.1% to 99.6%), respectively (eight studies, 880 participants; 173 confirmed cases of XDR-TB; low quality evidence).

Authors' conclusions: In adults with TB, a positive MTBDRsl result for FQ resistance, SLID resistance, or XDR-TB can be treated with confidence. However, MTBDRsl does not detect approximately one in five cases of FQ-resistant TB, and does not detect approximately one in four cases of SLID-resistant TB. Of the three SLIDs, MTBDRsl has the poorest sensitivity for kanamycin resistance. MTBDRsl will miss between one in four and one in three cases of XDR-TB. The diagnostic accuracy of MTBDRsl is similar when done using either culture isolates or smear-positive sputum. As the location of the resistance causing mutations can vary on a strain-by-strain basis, further research is required on test accuracy in different settings and, if genetic sequencing is used as a reference standard, it should examine all resistance-determining regions. Given the confidence one can have in a positive result, and the ability of the test to provide results within a matter of days, MTBDRsl may be used as an initial test for second-line drug resistance. However, when the test reports a negative result, clinicians may still wish to carry out conventional testing.

Figure 1

Clinical pathway diagram showing how molecular drug susceptibility testing (DST), which may use the MTBDRsl assay, is applied. A patient with suspected TB or suspected drug-resistant TB supplies a biological specimen (usually sputum), which is examined by smear microscopy and cultured. If acid-fast bacilli are observed under the microscope (smear-positive), a molecular DST can be performed directly on the specimen. If acid-fast bacilli are not observed (smear-negative), molecular DST can only be performed with acceptable accuracy on the culture isolate grown from the specimen. A molecular test for first-line drug resistance (for example, the MTBDRplus assay) is performed first and, only if resistance to the first-line drugs is indicated, the specimen is tested further for resistance to the second-line drugs using the MTBDRsl assay. Where molecular testing is not available, phenotypic testing for drug resistance may be performed on culture-positive isolates. Although phenotypic testing is being replaced by molecular-based methods in some settings, it is still usually performed in research studies seeking to measure the accuracy of the molecular test. Furthermore, some research studies also use gene sequencing as a reference standard or any specimens with discordant molecular DST-culture results.

Figure 2

Examples of different GenoType® MTBDRsl strip readouts (Hain Life Sciences 2012b).

Figure 3

Study flow diagram.

Figure 4

Risk of bias and applicability concerns graph: review authors' judgements about each domain presented as percentages across included studies.

Figure 5

Risk of bias and applicability concerns summary: review authors' judgements about each domain for each included study.

Figure 6

Forest plots of MTBDRsl sensitivity and specificity when performed indirectly or directly for FQ resistance detection and using phenotypic culture-based DST as a reference standard. The individual studies are ordered by decreasing sensitivity. TP = true positive; FP = false positive; FN = false negative; TN = true negative. Values between brackets are the 95% CIs of sensitivity and specificity. The figure shows the estimated sensitivity and specificity of the study (blue square) and its 95% CI (black horizontal line).

Figure 7

Summary plots of MTBDRsl sensitivity and specificity comparing detection of fluoroquinolone resistance by indirect and direct testing. The solid circles correspond to the summary estimates of sensitivity and specificity and are shown with 95% confidence regions (dotted lines) and 95% prediction regions (dashed lines).

Figure 8

Forest plots of MTBDRsl sensitivity and specificity for SLID resistance detection when performed indirectly or directly and using phenotypic culture-based DST as a reference standard. The individual studies are ordered by decreasing sensitivity. TP = true positive; FP = false positive; FN = false negative; TN = true negative. Values between brackets are the 95% CIs of sensitivity and specificity. The figure shows the estimated sensitivity and specificity of the study (blue square) and its 95% CI (black horizontal line)

Figure 9

Summary plots of MTBDRsl sensitivity and specificity comparing detection of resistance for second-line injectable drugs by indirect and direct testing. The solid circles correspond to the summary estimates of sensitivity and specificity and are shown with 95% confidence regions (dotted lines) and 95% prediction regions (dashed lines).

Figure 10

Forest plots of MTBDRsl sensitivity and specificity when performed indirectly for the detection of resistance to amikacin (Ak), kanamycin (Kn) and capreomycin (Cm) using culture as a reference standard. The individual studies are ordered by decreasing sensitivity. TP = true positive; FP = false positive; FN = false negative; TN = true negative. Values between brackets are the 95% CIs of sensitivity and specificity. The figure shows the estimated sensitivity and specificity of the study (blue square) and its 95% CI (black horizontal line).

Figure 11

Summary plots of MTBDRsl sensitivity and specificity comparing indirect detection of resistance for amikacin (Ak), kanamycin (Kn) and capreomycin (Cm) using culture as a reference standard. The solid circles correspond to the summary estimates of sensitivity and specificity and are shown with 95% confidence regions (dotted lines) and 95% prediction regions (dashed lines).

Figure 12

Forest plots of MTBDRsl sensitivity and specificity when performed directly for the detection of resistance to amikacin (Ak), kanamycin (Kn) and capreomycin (Cm) using culture as a reference standard. The individual studies are ordered by decreasing sensitivity. TP = true positive; FP = false positive; FN = false negative; TN = true negative. Between brackets are the 95% CI of sensitivity and specificity. The figure shows the estimated sensitivity and specificity of the study (blue square) and its 95% CI (black horizontal line).

Figure 13

Summary plots of MTBDRsl sensitivity and specificity comparing direct detection of resistance for amikacin (Ak), kanamycin (Kn) and capreomycin (Cm) using culture as a reference standard. The solid circles correspond to the summary estimates of sensitivity and specificity and are shown with 95% confidence regions (dotted lines) and 95% prediction regions (dashed lines).

Figure 14

Forest plots of MTBDRsl sensitivity and specificity when performed indirectly and directly for the detection of XDR-TB using phenotypic culture-based DST as a reference standard. The individual studies are ordered by decreasing sensitivity. TP = true positive; FP = false positive; FN = false negative; TN = true negative. Between brackets are the 95% CI of sensitivity and specificity. The figure shows the estimated sensitivity and specificity of the study (blue square) and its 95% CI (black horizontal line).

Figure 15

An example of the manufacturer-supplied result template.

Figure 16

An example of a readout from an automated strip reader. The results are generated automatically and validated manually by a technician.

Figure 17

Forest plots of MTBDRsl sensitivity and specificity for fluoroquinolone (FQ) resistance detection when performed indirectly using different reference standards. The individual studies are ordered by decreasing sensitivity. TP = true positive; FP = false positive; FN = false negative; TN = true negative. Values between brackets are the 95% CIs of sensitivity and specificity. The figure shows the estimated sensitivity and specificity of the study (blue square) and its 95% CI (black horizontal line).

Figure 18

Forest plots of MTBDRsl sensitivity and specificity for ofloxacin (Ofl) and moxifloxacin (Mx) resistance detection when performed indirectly and using phenotypic culture-based DST as a reference standard. The individual studies are ordered by decreasing sensitivity. TP = true positive; FP = false positive; FN = false negative; TN = true negative. Values between brackets are the 95% CIs of sensitivity and specificity. The figure shows the estimated sensitivity and specificity of the study (blue square) and its 95% CI (black horizontal line).

Figure 19

Summary plots of MTBDRsl sensitivity and specificity comparing direct detection of resistance for ofloxacin (Ofl) and moxifloxacin (Mx) using culture as a reference standard. The solid circles correspond to the summary estimates of sensitivity and specificity and are shown with 95% confidence regions (dotted lines) and 95% prediction regions (dashed lines).

Figure 20

Forest plots of MTBDRsl sensitivity and specificity when performed indirectly for second-line injectable drug (SLID) resistance detection and using three different reference standards. The individual studies are ordered by decreasing sensitivity. TP = true positive; FP = false positive; FN = false negative; TN = true negative. Values between brackets are the 95% CIs of sensitivity and specificity. The figure shows the estimated sensitivity and specificity of the study (blue square) and its 95% CI (black horizontal line).

Test 1.

Indirect, FQ, culture.

Test 2.

Indirect, Ofl, culture.

Test 3.

Indirect, Mx, culture.

Test 4.

Indirect, SLID, culture.

Test 5.

Indirect, Ak, culture.

Test 6.

Indirect, Kn, culture.

Test 7.

Indirect, Cm, culture.

Test 8.

Indirect, XDR, culture.

Test 9.

Indirect, FQ, sequencing.

Test 10.

Indirect, SLID, sequencing.

Test 11.

Indirect, XDR, sequencing.

Test 12.

Indirect, FQ, sequencing and culture.

Test 13.

Indirect, SLID, sequencing and culture.

Test 14.

Indirect, XDR, sequencing and culture.

Test 15.

Indirect, FQ, culture followed by sequencing of discrepants.

Test 16.

Indirect, SLID, culture followed by sequencing of discrepants.

Test 17.

Direct, FQ, culture.

Test 18.

Direct, Ofl, culture.

Test 19.

Direct, SLID, culture.

Test 20.

Direct, Ak, culture.

Test 21.

Direct, Kn, culture.

Test 22.

Direct, Cm, culture.

Test 23.

Direct, XDR, culture.

Test 24.

Direct, FQ, culture followed by sequencing of discrepants.

Test 25.

Direct, SLID, culture followed by sequencing of discrepants.

Test 26.

Direct, XDR, culture followed by sequencing of discrepants.