Characterization of Mycobacterium tuberculosis heteroresistance by genotyping

Background: Heteroresistance is the coexistence of susceptible and resistant strains in the same individual, considered the preliminary step for total resistance, and can stem from mixed infection or clonal heterogeneity. The aim of this study was to evaluate the heteroresistance of Mycobacterium tuberculosis to rifampicin and isoniazid and its characterization. Method: GenoType MTBDRplus®; Sanger sequencing of the rpoB, katG, and inhA genes; and Mycobacterial Interspersed Repetitive Unit – Variable Number Tandem Repeat (MIRU-VNTR) were performed. Results: In a total of 654 isolates, 530 were resistant, 124 were susceptible, and 29 were heteroresistant to a first-line drug. GenoType MTBDRplus® detected heteroresistance in the rpoB gene in 26/29 (89.6%), as compared to 5/29 (17.2%) in the katG gene and 2/29 (6.8%) in the inhA gene. Four isolates showed heteroresistance in these genes. The Sanger sequencing detected heteroresistance in the rpoB gene in 7/29 (24.1%), as compared to 3/29 (10.3%) in the katG gene. In one isolate, heteroresistance was concomitant in both the rpoB and katG genes. MIRU-VNTR detected mixed infection in three heteroresistant isolates, while four isolates showed clonal heterogeneity. Conclusions: GenoType MTBDRplus® detected more cases of heteroresistance when compared to sequencing. It was also possible to characterize mixed infection and clonal heterogeneity by MIRU-VNTR.