Evaluation of the analytical performance of the PURE-TB-LAMP assay for tuberculosis detection

Abstract

Given the prevalence and lethality of tuberculosis (TB) in developing countries, there is an ongoing need for rapid, simple, and low-cost detection method that nonetheless sensitive and highly specific. The present study evaluated the basic performance of a novel TB detection method combining procedure for ultra-rapid extraction (PURE) and loop-mediated isothermal amplification for TB (TB-LAMP). The PUER-TB-LAMP assay detected four Mycobacterium tuberculosis complexes and did not show any cross-reactivity with 18 species of non-TB mycobacteria (NTM) or with 10 species of other bacteria that cause respiratory tract infections such as Streptococcus pneumonia, underscoring its high specificity for TB detection. The analytical sensitivity of the assay was 100 CFU/ml for M. tuberculosis strain H37Rv cell and was unaffected by the presence of excess amounts of M. avium cells (a typical NTM species) or human genomic DNA. Moreover, when used with a range of artificial specimens prepared by spiking known amounts of cultured M. tuberculosis cells, the PURE method efficiently removed various inhibitory materials from a variety of samples such as sputum, urine, simulated gastric fluid, and whole blood, demonstrating the applicability of this assay to these samples. These results suggest that the PURE-TB-LAMP assay is a highly effective and accessible TB detection method that can be useful in resource-limited communities.

Evaluation of Loopamp Assay for the Diagnosis of Pulmonary Tuberculosis in Cambodia

“…A 30 μL aliquot of purified DNA was dispensed in the LAMP reaction tube containing the lyophilised LAMP reagents (Bst DNA polymerase, dNTPs, calcein, and MTBC-specific primers targeting DNA gyrase subunit B and insertion sequence 6110). High specificity was reported by Yuki regarding the combination of primers included in the kit [7]. The dried LAMP reagents (stored in the cap of the reaction tube) were reconstituted by mixing by inversion with the DNA solution.…”