National Tuberculosis Reference Laboratories (NTRLs) are central to tuberculosis (TB) control programs. Between 2018 and 2024, the Republic of Congo, a country of 6 million inhabitants, achieved a transformative strengthening of its TB diagnostic system, coordinated by the NTRL. Strategic investments, supported mainly by international partners, enabled a substantial decentralization of services, expanding the diagnostic network from 38 to 113 diagnostic and testing centers and increasing GeneXpert sites from 3 to 31. The expansion of the diagnostic network and specimen referral system was associated with a reduced structural gap in diagnostic coverage by extending access to GeneXpert testing to a larger number of peripheral and previously underserved centers. Critically, the establishment of a BSL-3 laboratory and the deployment of advanced assays like Xpert MTB/XDR ended the reliance on overseas testing by introducing in-country capacity for multidrug-resistant and pre-extensively drug-resistant TB detection. These systemic improvements were associated with significant positive outcomes, including an annual molecular testing surging from 11,609 in 2022 to over 27,000 in 2024 and bacteriological confirmation rates rising from 34 to 73%. This comprehensive laboratory systems strengthening, which also facilitated cross-programmatic initiatives like HIV and Mpox testing integration, underscores how sustained investment in infrastructure, logistics, and quality management is fundamental to improving case detection, surveillance, and progress toward the WHO End TB Strategy milestones.

Tuberculosis (TB) remains a major public health challenge, particularly in sub-Saharan Africa, where the emergence of drug-resistant MTB undermines current control efforts. This study aimed to characterize genetic mutations associated with resistance to first- and second-line injectable drugs in MTB isolates from pulmonary TB patients in the Republic of Congo. A total of 45 culture-positive MTB isolates collected between 2018 and 2019 were analyzed using targeted next-generation sequencing on the MinION device. Resistance-associated genes, including rpoB, katG, embB, gyrA, and rrs, were examined. Overall, 22/45 isolates (48%) harbored at least one mutation associated with drug resistance. Among resistant isolates, rifampicin resistance was detected in 15 of 22 (68.2%), predominantly associated with mutations in the rpoB rifampicin resistance–determining region, including p.Ser531Leu, p.Asp516Val, and p.His526Tyr.Ethambutol resistance was observed in 11 of 22 isolates (50.0%) and was mainly associated with the embB p.Met306Val mutation. Isoniazid resistance was detected in 9 of 22 isolates (40.9%), primarily associated with the KatG p.Ser315Thr mutation. Fluoroquinolone resistance was identified in 2 of 22 isolates (9.1%), mainly associated with gyrA p.Ala90Val and p.Asp94Gly mutations. Resistance to second-line injectable drugs was detected in 1 of 22 isolates (4.5%) and was associated with the rrs p.Ala1401Gly mutation. These findings underscore the need to enhance molecular surveillance and strengthen treatment protocols for drug-resistant TB in the Republic of Congo to effectively combat this evolving health crisis.