Association of Mycobacterium tuberculosis genotypes and treatment outcome in pulmonary tuberculosis patients in Tshwane Metropolitan area

Abstract

Tuberculosis (TB) is a major cause of death worldwide, especially in Asia and Africa. Genotyping methods such as insertion sequence 6110-restriction fragment length polymorphism (IS6110-RFLP), spoligotyping and mycobacterial interspersed repetitive units variable number of tandem repeats (MIRU-VNTR) have expanded the ability to investigate and understand TB. Genotyping of M. tuberculosis has shown that certain M. tuberculosis strains are more prevalent in certain geographic regions, thus raising questions about bacterial factors in pathogenesis, presentation of disease and treatment response. Given the diversity in insertions, deletions and single nucleotide polymorphisms seen in M. tuberculosis, it is plausible that the genetics of the pathogen play a role in presentation of disease and response to treatment. However, there is very limited evidence on the influence of M. tuberculosis genotypes on treatment outcome of TB around the world. Whether response to TB treatment differs among the different genotypes is unknown. It is important to investigate the influence M. tuberculosis genotypes may have on treatment response; to develop effective control strategies for tuberculosis. This study aimed to determine the genetic diversity of M. tuberculosis strains in Tshwane metropolitan area and to determine the association of those genotypes with the laboratory based-treatment response using the eight weeks culture result as a marker for treatment response. In addition the study explored the association of genotypes with drug resistance and patient demographics (age and gender). In this study a total of 108 consecutive M. tuberculosis isolates resistant to either rifampicin and/or isoniazid were collected between September 2011 and December 2011from the National Health Laboratory Services (NHLS), Tshwane academic division laboratory . Two isolates were excluded from the study due to contamination. The study population included 50 females (47.17 %) and 47 males (44.34 %) with 9 patients (8.49 %) that did not have gender available. The age range for the study population was from 2 days to 79 years. Genotyping was performed using two different PCR-based methods; spoligotyping, and 24 loci MIRU-VNTR typing. Comparison of Spoligotyping results of the 106 M. tuberculosis isolates with the SpolDB4 database showed that 73% (83/106) isolates belonged to 21 previously described shared types (ST); while 27% (23/106) were not found in the SpolDB4 database and were considered as orphans. Spoligotyping identified seven major distinct families of TB including Beijing, East African Indian (EAI), Latin American and Mediterranean (LAM), T family, X family, CAS as well as Manu 2 families. Spoligotyping identified 14 clusters, giving a clustering rate of 50%. However no cluster was identified with 24-loci MIRU-VNTR typing. The study did not find any as association of. In this study only 46 patients out of the 106 of the study population had the 8 weeks follow up result and no genotypes were associated with treatment outcome in this population. The study also did not find any association of specific genotype with gender, age or drug resistance to INH or Rif. Resistance to RIF was associated with treatment failure (patients failed to convert after 8 weeks), with P value of 0.006. However, no significant association was found between INH resistance and treatment outcome. In conclusion the study showed high genetic diversity of strains in Tshwane Metropolitan area. No association was found between genotypes and treatment response. However more than 50% did not have eight weeks follow up result. The study did however find Rifampicin (Rif) mono resistance to be associated with treatment failure. Further studies are needed to confirm the present study finding.