Sensitivity profiling of genes responsible for multi-/extensively drug resistant mutations in Mycobacterium tuberculosis isolated from the Tshepong National Health Laboratory Service Referral Tuberculosis Laboratory in North-West

Abstract

Background: Tuberculosis (TB), an infection that is caused by Mycobacterium tuberculosis organisms, remains one of the leading infections causing morbidity and mortality globally, with the emergence of multidrug-resistant (MDR) and extensively drug-resistant (XDR) strains, causing a major public-health challenge. Tuberculosis is transmitted through the inhalation of airborne droplets expelled by a person with active TB when they cough or sneeze. Several diagnostic methods are used in the diagnosis of tuberculosis, including microscopy, culture, and molecular assays. The first-line drugs used in the treatment of TB are rifampicin, isoniazid, pyrazinamide, and ethambutol. The second-line drugs used in the treatment of multidrug-resistant tuberculosis (MDR-TB) include fluoroquinolones (such as levofloxacin and moxifloxacin), second-line injectable agents (such as amikacin, kanamycin, and capreomycin), and oral agents, including ethionamide, prothionamide, cycloserine, and para-aminosalicylic acid (PAS). Aim: To determine sensitivity profiling and detect the genes responsible for MDR/XDR mutations in tuberculosis isolated from the Tshepong National Health Laboratory Service (NHLS) TB laboratory from May 2023 to March 2025. Methods: Retrospective data were retrieved from the National Health Laboratory service through the Academic Affairs and Research Management Systems. The data were for the patients infected by Mycobacterium tuberculosis at Tshepong TB Referral Laboratory in North-West. The data were analysed using Microsoft 365 Excel, Descriptive statistics, Chi-square, Fisher’s exact test, and ANOVA. The results are presented in tables and figures. The results: A total of 340 Mycobacterium tuberculosis-positive isolates from the Tshepong NHLS TB Referral Laboratory in the North-West Province were analysed. The largest proportion of cases was observed in the 31–40-year age group (30%), reflecting the high burden of disease in the economically active population. MDR-TB represented the most frequent classification, accounting for 62.1% of all cases, indicating a substantial burden of multidrug resistance within the study population. Among the 340 specimens evaluated, 64.4% (n = 219) were resistant to rifampicin, while 21% (n = 70) were sensitive and 15% (n = 52) produced unsuccessful results. The highest levels of susceptibility were observed for amikacin (98.8%), capreomycin (98.8%), kanamycin (98.8%), and fluoroquinolones (93.2%), confirming the continued effectiveness of these second-line drugs. Ethionamide exhibited the highest resistance rate (12.7%). Molecular analysis revealed that the rpoB gene, associated with rifampicin resistance, was the most frequently affected locus (n = 219; 64.4%). The highest mean melting temperature was recorded for the inhA promoter gene (75 °C), followed by gyrA (72 °C) and rrs (71 °C). The lowest mean melting temperature (68 °C) was observed for the eis promoter gene associated with kanamycin resistance. Facility-level analysis demonstrated that Tshepong Hospital reported the highest number of TB cases (n = 155; 45.6%), followed by Moses Kotane Hospital (n = 45; 13.2%) and other small facilities (n = 56; 16.5%). The highest number of MDR-TB cases was also recorded at Tshepong (n = 95; 61.3%), followed by Moses Kotane (n = 32; 71.3%). Together, Tshepong and Moses Kotane Hospitals accounted for most MDR-TB cases in the cohort. XDR-TB cases were reported at Tshepong (3.2%), Moses Kotane (6.7%), other small facilities (1.8%), and the NIC Bodenstein Hospital (16.67%), indicating the presence of extensively drug-resistant TB in multiple facilities, although at low prevalence. Conclusion: These findings highlight the need for enhanced molecular diagnostic capacity and consistent monitoring of resistance trends in routine clinical practice. Targeted interventions at high-risk facilities should be prioritised to prevent further transmission. Future research should investigate the genetic diversity of circulating strains and evaluate the effectiveness of tailored treatment approaches for drug-resistant TB.