Performance of the TB-LAMP diagnostic assay in reference laboratories: Results from a multicentre study.

OBJECTIVE: To evaluate the diagnostic performance of TB-LAMP, a manual molecular tuberculosis (TB) detection method, and provide comparison to the Xpert MTB/RIF assay. METHODS: In a large multicentre study, two sputum samples were collected from participants with TB symptoms in reference laboratories in Peru, South Africa, Brazil, and Vietnam. Each sample was tested with TB-LAMP. The reference standard consisted of four direct smears, four cultures, and clinical and radiological findings. Individuals negative on conventional tests were followed up after 8 weeks. The Xpert MTB/RIF assay was performed on fresh or frozen samples as a molecular test comparison. RESULTS: A total of 1036 adults with suspected TB were enrolled. Among 375 culture-confirmed TB cases with 750 sputum samples, TB-LAMP detected 75.6% (95% confidence interval (CI) 71.8-79.4%), including 97.9% (95% CI 96.4-99.4%) of smear-positive TB samples and 46.6% (95% CI 40.6-52.7%) of smear-negative TB samples. Specificity in 477 culture-negative participants not treated for TB (954 sputum samples) was 98.7% (95% CI 97.9-99.6%). TB-LAMP test results were indeterminate in 0.3% of cases. CONCLUSIONS: TB-LAMP detects nearly all smear-positive and half of smear-negative TB cases and has a high specificity when performed in reference laboratories. Performance was similar to the Xpert MTB/RIF assay.

Diagnostic Performance of Tuberculosis-Specific IgG Antibody Profiles in Patients with Presumptive Tuberculosis from Two Continents.

BACKGROUND: Development of rapid diagnostic tests for tuberculosis is a global priority. A whole proteome screen identified Mycobacterium tuberculosis antigens associated with serological responses in tuberculosis patients. We used World Health Organization (WHO) target product profile (TPP) criteria for a detection test and triage test to evaluate these antigens. METHODS: Consecutive patients presenting to microscopy centers and district hospitals in Peru and to outpatient clinics at a tuberculosis reference center in Vietnam were recruited. We tested blood samples from 755 HIV-uninfected adults with presumptive pulmonary tuberculosis to measure IgG antibody responses to 57 M. tuberculosis antigens using a field-based multiplexed serological assay and a 132-antigen bead-based reference assay. We evaluated single antigen performance and models of all possible 3-antigen combinations and multiantigen combinations. RESULTS: Three-antigen and multiantigen models performed similarly and were superior to single antigens. With specificity set at 90% for a detection test, the best sensitivity of a 3-antigen model was 35% (95% confidence interval [CI], 31-40). With sensitivity set at 85% for a triage test, the specificity of the best 3-antigen model was 34% (95% CI, 29-40). The reference assay also did not meet study targets. Antigen performance differed significantly between the study sites for 7/22 of the best-performing antigens. CONCLUSIONS: Although M. tuberculosis antigens were recognized by the IgG response during tuberculosis, no single antigen or multiantigen set performance approached WHO TPP criteria for clinical utility among HIV-uninfected adults with presumed tuberculosis in high-volume, urban settings in tuberculosis-endemic countries.

Cost-effectiveness analysis of introduction of rapid, alternative methods to identify multidrug-resistant tuberculosis in middle-income countries.

BACKGROUND: Resistance to commonly used antituberculosis drugs is emerging worldwide. Conventional drug-susceptibility testing (DST) methods are slow and demanding. Alternative, rapid DST methods would permit the early detection of drug resistance and, in turn, arrest tuberculosis transmission. METHODS: A cost-effectiveness analysis of 5 DST methods was performed in the context of a clinical trial that compared rapid with conventional DST methods. The methods under investigation were direct phage-replication assay (FASTPlaque-Response; Biotech), direct amplification and reverse hybridization of the rpoB gene (INNO-LiPA; Innogenetics), indirect colorimetric minimum inhibitory concentration assay (MTT; ICN Biomedicals), and direct proportion method on Löwenstein-Jensen medium. These were compared with the widely used indirect proportion method on Löwenstein-Jensen medium. RESULTS: All alternative DST methods were found to be cost-effective, compared with other health care interventions. DST methods also generate substantial cost savings in settings of high prevalence of multidrug-resistant tuberculosis. Excluding the effects of transmission, the direct proportion method on Löwenstein-Jensen medium was the most cost-effective alternative DST method for patient groups with prevalences of multidrug-resistant tuberculosis of 2%, 5%, 20%, and 50% (cost in US$2004, $94, $36, $8, and $2 per disability-adjusted life year, respectively). CONCLUSION: Alternative, rapid methods for DST are cost-effective and should be considered for use by national tuberculosis programs in middle-income countries.

Operational feasibility of using loop-mediated isothermal amplification for diagnosis of pulmonary tuberculosis in microscopy centers of developing countries.

The characteristics of loop-mediated isothermal amplification (LAMP) make it a promising platform for the molecular detection of tuberculosis (TB) in developing countries. Here, we report on the first clinical evaluation of LAMP for the detection of pulmonary TB in microscopy centers in Peru, Bangladesh, and Tanzania to determine its operational applicability in such settings. A prototype LAMP assay with simplified manual DNA extraction was evaluated for accuracy and ease of use. The sensitivity of LAMP in smear- and culture-positive sputum specimens was 97.7% (173/177 specimens; 95% confidence interval [CI], 95.5 to 99.9%), and the sensitivity in smear-negative, culture-positive specimens was 48.8% (21/43 specimens; CI, 33.9 to 63.7%). The specificity in culture-negative samples was 99% (500/505 specimens; CI, 98.1 to 99.9%). The average hands-on time for testing six samples and two controls was 54 min, similar to that of sputum smear microscopy. The optimal amplification time was 40 min. No indeterminate results were reported, and the interreader variability was 0.4%. Despite the use of a single room without biosafety cabinets for all procedures, no DNA contamination was observed. The assay was robust, with high end-point stability and low rates of test failure. Technicians with no prior molecular experience easily performed the assay after 1 week of training, and opportunities for further simplification of the assay were identified.