Molecular analysis of clinical isolates of Mycobacterium tuberculosis collected from patients with persistent disease in the Khartoum region of Sudan.

SETTING: Patients with positive smears for acid-fast bacilli were enrolled at tuberculosis (TB) clinics in the Khartoum region of Sudan.

OBJECTIVE: To identify the presence of drug resistant genotypes in M. tuberculosis isolates which are difficult to treat.

METHODS: Genus specific PCR-SSCP was performed to confirm the presence of M. tuberculosis in clinical isolates. Genotypic drug resistance testing was performed by mutation analysis and spoligotyping was used to monitor transmission and to identify epidemic strains.

RESULTS: Fifty (48%) of the original 105 samples were classified as M. tuberculosis. Four (4%) of the samples were typed as mycobacteria other than TB, while the remaining (n =50) samples were refractory to further molecular analysis. The fifty amplifiable M. tuberculosis samples were used for subsequent mutation analysis and typing. Mutations were identified in the genes conferring resistance to INH (kat G, 12%), RIF (rpoB, 8%), SM (r psL and rrs, 30%) and EMB (embB, 4%). Two of the samples (4%) had mutations in genes associated to both INH and RIF and can be classified as MDR-TB. Thirty-three percent (13/39) of the persistant tuberculosis cases (5/18 treatment failure; 5/14 relapse; 3/7 defaulter) had mutations accounting for drug resistance. A total of 27 different spoligotypes were identified from 49/50 samples. Twenty-nine (59%) of the isolates were grouped into one of seven clusters, while 20 (41%) showed unique patterns. One patient was infected with M. bovis.

CONCLUSION: This is the first molecular approach to characterize clinical isolates of M. tuberculosis from Sudan. The results show that drug resistance is indeed a serious problem and it may compliment the efforts of the National Tuberculosis Programme to improve strategies to control this disease.