Molecular characterization of rpoB gene mutations in isolates from tuberculosis patients in Cubal, Republic of Angola

Angola; Rifampicina; Mutaciones rpoBAngola; Rifampicina; Mutacions rpoBAngola; Rifampicin; rpoB mutationsBackground The importance of Mycobacterium tuberculosis strains with disputed rpoB mutations remains to be defined. This study aimed to assess the frequency and types of rpoB mutations in M. tuberculosis isolates from Cubal, Angola, a country with a high incidence of tuberculosis. Methods All isolates included (n = 308) were analyzed using phenotypic drug susceptibility testing and GenoType MTBDRplus assay. DNA sequencing of the rpoB gene and determination of rifampicin MIC by macrodilution method were additionally performed on isolates yielding discordant results (n = 12) and those in which the mutation detected was not characterized (n = 8). Results In total, 85.1% (74/87) of rifampicin-resistant strains had undisputed rpoB mutations -S450L (49), D435V (15), H445D (3), H445Y (2), Q432ins (1), L449M plus S450F (1), S450F (1), S450W (1) and S450Y (1)-; 10.3% (9/87) had disputed rpoB mutations—L430P plus S493L (1), N437del (1), H445L (3), D435Y (2), L452P (2)-, 2.3% (2.3%) showed no rpoB mutations and 2.3% (2/87) showed heteroresistance—D435Y plus L452P and L430P plus S493L-. Conclusion Disputed rpoB mutations were common, occurring in 10.3% of rifampicin resistant isolates. Current phenotyping techniques may be unable to detect this resistance pattern. To increase their sensitivity, a lower concentration of RIF could be used in these tests or alternatively, rpoB mutations could be screened and characterized in all M. tuberculosis strains.This work was supported by Probitas Foundation. Thanks to the financial support received from Probitas Foundation it was possible not only purchase the equipment and reagents to launch the study but to strengthen the capacity of the laboratory and local staff

Molecular characterization of rpoB gene mutations in isolates from tuberculosis patients in Cubal, Republic of Angola

Background: The importance of Mycobacterium tuberculosis strains with disputed rpoB mutations remains to be defined. This study aimed to assess the frequency and types of rpoB mutations in M. tuberculosis isolates from Cubal, Angola, a country with a high incidence of tuberculosis. Methods: All isolates included (n = 308) were analyzed using phenotypic drug susceptibility testing and GenoType MTBDRplus assay. DNA sequencing of the rpoB gene and determination of rifampicin MIC by macrodilution method were additionally performed on isolates yielding discordant results (n = 12) and those in which the mutation detected was not characterized (n = 8). Results: In total, 85.1% (74/87) of rifampicin-resistant strains had undisputed rpoB mutations -S450L (49), D435V (15), H445D (3), H445Y (2), Q432ins (1), L449M plus S450F (1), S450F (1), S450W (1) and S450Y (1)-; 10.3% (9/87) had disputed rpoB mutations-L430P plus S493L (1), N437del (1), H445L (3), D435Y (2), L452P (2)-, 2.3% (2.3%) showed no rpoB mutations and 2.3% (2/87) showed heteroresistance-D435Y plus L452P and L430P plus S493L-. Conclusion: Disputed rpoB mutations were common, occurring in 10.3% of rifampicin resistant isolates. Current phenotyping techniques may be unable to detect this resistance pattern. To increase their sensitivity, a lower concentration of RIF could be used in these tests or alternatively, rpoB mutations could be screened and characterized in all M. tuberculosis strains