From Aug 11, 2009, until June 26, 2010, we enrolled 6648 eligible adults (, ). One-off MTB/RIF testing correctly detected tuberculosis in more than 90% of patients with positive cultures, with 99% specificity for non-tuberculosis (). Performance was much the same during validation and implementation phases (webappendix p 2
). A one-off MTB/RIF test identified significantly (p<0·0001) more cases of tuberculosis than did 2–3 smear microscopy examinations per patient, which detected 699 of 1041 culture-positive patients (sensitivity of 67·1%) and 3700 of 3718 patients without tuberculosis (specificity of 99·5%). Although HIV co-infection significantly decreased the sensitivity of smear microscopy (p<0·0001), the sensitivity of MTB/RIF was not significantly affected by HIV co-infection status (p=0·0849; ). MTB/RIF test sensitivity and specificity were much the same between basic health centres and sites with increased capacity both between countries (p=0·895 and p=0·097, respectively; webappendix p 2
), and within countries with more than one site (webappendix p 2
Characteristics of patients and study sites
HIV statuses in patients with suspected cases of tuberculosis and multidrug-resistant tuberculosis
Sensitivity, specificity, and predictive values of a one-off direct MTB/RIF test
Sensitivity and specificity of smear microscopy (two to three microscopy examinations as per routine practice) and a one-off direct MTB/RIF test, stratified by HIV status of patients
Sensitivity of MTB/RIF testing for smear-negative tuberculosis varied between countries (p<0·0001). It was lower at sites that used a reference standard of solid and liquid cultures (Azerbaijan, Uganda, and the Philippines) and slightly higher at sites that tested morning sputum samples rather than spot sputum collections (Peru and India).
MTB/RIF testing correctly identified 242 of 250 cases of rifampicin-resistant tuberculosis (sensitivity of 96·8%) and 779 of 810 rifampicin-sensitive cases (specificity of 96·2%). However, because of concern over false-positive results, especially for settings with a low-prevalence of multidrug-resistant disease, we changed the software cutoff defining drug resistance during the study on May 12, 2010. With modified software definitions, our post-hoc analysis showed that sensitivity decreased to 94·4% and specificity increased to 98·3% (). 17 (6·8%) of 250 cases of rifampicin-resistant tuberculosis were sensitive to isoniazid.
MTB/RIF test sensitivity and specificity for detection of rifampicin resistance after change to software cutoff
24 (16%) of 153 patients with clinically diagnosed tuberculosis, but negative culture had positive results on MTB/RIF testing. 20 (83%) of these 24 patients had clinical and radiological follow-up, and all 20 improved on tuberculosis treatment. For the 118 (91%) of 129 patients who tested negative on MTB/RIF but were treated for tuberculosis on the basis of a clinical diagnosis and had clinical and radiological follow-up, only 67 (57%) showed improvement (p<0·0001).
Median time to detection of tuberculosis for the MTB/RIF test was 0 days (IQR 0–1), compared with 1 day (0–1) for smear microscopy, 30 days (23–43) for solid culture, and 16 days (13–21) for liquid culture (). Median time to detection of rifampicin resistance was 1 day (0–1) for the MTB/RIF test, 20 days (10–26) for line-probe assay (done directly from sputum pellet for smear-positive specimens and from culture isolates for smear-negative specimens) and 106 days (30–124) for phenotypic drug-susceptibility testing (). Although MTB/RIF testing and microscopy were done near the clinics and results were rapidly received by clinicians (median 1 day [IQR 0–2] for MTB/RIF testing and 2 days [2–3] for microscopy), there were significant delays in receiving results from cultures (median 58 days [42–62]), line-probe assays (40 days [27–53]), and conventional drug-susceptibility testing (63 days [38–102]). Some results were lost or unreported ().
Proportion of tuberculosis cases detected by each method in culture-positive patients
Proportion of results reported to the clinics for each method from date of first sputum sample
Time between sputum collection and treatment initiation was very dependent on the testing method (). In the baseline group in South Africa and the validation phase at other sites (ie, when MTB/RIF test results were not used to direct therapy), patients with smear-negative, culture-positive tuberculosis started treatment after a median of 56 days (IQR 39–81). Once MTB/RIF test results were used to direct therapy, the median time-to-treatment for smear-negative tuberculosis reduced to 5 days (2–8). Rates of untreated smear-negative, culture-positive tuberculosis reduced from 39·3% (95% CI 32·6–46·6) at baseline to 14·7% (9·9–21·2) after implementation of the MTB/RIF test.
Figure 4 Time to treatment during validation phase (treatment based on conventional methods only) and implementation phase (treatment based on MTB/RIF test and conventional methods) for patients with smear-positive, culture-positive tuberculosis, smear-negative, (more ...)
GeneXpert provides an indeterminate result if unexpected results occur with any of the internal control measures. The MTB/RIF test was indeterminate in 126 (2%) of 5321 samples tested. 112 repeat tests were successful when adequate sputum remained, with the indeterminate rate reduced to less than 1% (14/5321 samples). In 1449 samples that were positive on MTB/RIF testing, 17 (1%) had indeterminate results for rifampicin resistance. These tests were not repeated. By comparison, the contamination rate was 441 (5%) of 9690 cultures, including repeated cultures from re-decontaminated pellets from all countries apart from South Africa and the Philippines.
Operators without previous molecular biology experience or computer skills passed proficiency testing after 1–3 days of training on MTB/RIF tests, including three hands-on runs. A 1 day online training was successfully used at two sites (Peru and Azerbaijan). Monthly variation in MTB/RIF test performance did not differ between sites (psensitivity=0·52 on Cochran-Mantel-Haenszel test stratified by smear status and pspecificity=0·46 on χ2).
In one of the high HIV-prevalence sites, microscopy was introduced at the same time as MTB/RIF testing. Although MTB/RIF sensitivity for culture-positive tuberculosis at this site was much the same as in other centres (85·9%; 116 of 135 cases), the sensitivity of microscopy with one smear per patient was only 17·8% (21 of 118 smears) compared with 46·6% (55 of 118 smears) with a second smear at the reference laboratory. These findings support the laboratory managers' perception, expressed in user appraisal questionnaires, that MTB/RIF test performance might be less dependent on user skills, motivation, or workload than is microscopy.
We did not detect any DNA contamination events during monthly negative control runs, and test specificity was high across sites. The four-module GeneXpert device was used for 1–24 tests a day with only two incidents needing product support (one network-card failure requiring device replacement and one module replacement). At four sites, the recorded operating temperatures exceeded the maximum recommended operating temperature (15–30°C) during more than 10% of runs. Test performance and frequency of indeterminate results did not show seasonal variation in these sites. In one case, the operating temperature exceeded 40°C and an error message appeared as described in the manual. Several sites had daily temperatures higher than the 2–28°C recommended for cartridge storage temperature; cartridges were stored centrally and distributed twice every month. All sites had power cuts, but used uninterruptible power supplies to support the device during short power cuts and one site used an inverter and serial car batteries during a longer power outage.