RpoB Gene-Based Characterization of Non-Tuberculous Mycobacteria in Zimbabwe
Wadzanai Manjeese *
Department of Applied Biology and Biochemistry, National University of Science and Technology (NUST), P.O.Box AC 939, Ascot, Bulawayo, Zimbabwe and Department of Technical, National Biotechnology Authority, 21 Princess Drive, Newlands, Harare, Zimbabwe
Boniface Muzividzi
National Microbiology Reference Laboratory, Harare Central Hospital, P.O.Box ST 14, Southerton, Harare, Zimbabwe
Joshua Mbanga
Department of Applied Biology and Biochemistry, National University of Science and Technology (NUST), P.O.Box AC 939, Ascot, Bulawayo, Zimbabwe
Jonathan Mufandaedza
Department of Technical, National Biotechnology Authority, 21 Princess Drive, Newlands, Harare, Zimbabwe
Nyasha Chin’ombe
Molecular Microbiology Laboratory, Department of Medical Microbiology, University of Zimbabwe, P.O.Box A178, Avondale, Harare, Zimbabwe
*Author to whom correspondence should be addressed.
Abstract
Aims: To characterize archived nontuberculous Mycobacteria (NTM) samples previously isolated from humans across Zimbabwe during the 2014 national tuberculosis survey. The rpoB gene of the isolates was targeted for characterization.
Study Design: The research was an observational study where NTM samples previously isolated from Zimbabwean population were analysed by rpoB gene sequencing and phylogeny to identify NTM species.
Place and Duration of Study: Department of Medical Microbiology, University of Zimbabwe between September 2015 and July 2016.
Methodology: We obtained 99 NTM DNA samples from 963 NTM at NMRL, which we characterised using rpoB gene analysis after PCR amplification. The amplicons were sequenced and bioinformatics tools were used for speciation. The rpoB gene of DNA extracts from the NTM was amplified and the sequences were analysed using bioinformatics tools to identify the NTM to species level.
Results: From the 99 NTM isolates, 40 were sequenced and analyzed. The NTM were identified as belonging to 13 species. The species were M. palustre (14.8%), M. aroisense (29.6%), M. parascrofulaceum (3.7%), M. arupense (7.4%), M. asiaticum (3.7%), M. malmoense (3.7%), M. lacus (3.7%), M. avium (7.4%), M. nonchromogenicum (3.7%), M. gordonae (3.7%), M. aromaticivorans (3.7%), M. novocastrense (3.7%), M. bourgelatii (3.7%). One sample (3.7%) belonged to Mtb complex species (3.7%) and another one (3.7%) was closely related to S. oryzae. The most common species belonged to M. aroisense and M. palustre. The species showed a high degree of rpoB sequence diversity. Sequence analysis of the rifampicin resistance determining region (RRDR) showed the existence of only silent mutations.
Conclusion: Species of NTM from Zimbabwe showed a high degree of rpoB gene sequence diversity. This characteristic feature can therefore be used in diagnosis and identification of NTM in clinical specimens.
Keywords: rpoB gene, nontuberculous mycobacteria, sequencing, alignments, RRDR, Mutations