Date Published: March 31, 2017
Author(s): Tarsizio Chikaonda, Irene Ketseoglou, Nelson Nguluwe, Robert Krysiak, Isaac Thengolose, Felix Nyakwawa, Nora E. Rosenberg, Christopher Stanley, James Mpunga, Irving F. Hoffman, Maria A. Papathanasopoulos, Mina Hosseinipour, Lesley Scott, Wendy Stevens.
Availability and access to the detection of resistance to anti-tuberculosis drugs remains a significant challenge in Malawi due to limited diagnostic services. The Xpert® MTB/RIF can detect Mycobacterium tuberculosis and resistance to rifampicin in a single, rapid assay. Rifampicin-resistant M. tuberculosis has not been well studied in Malawi.
We aimed to determine mutations in the rifampicin resistance determining region (RRDR) of the rpoB gene of M. tuberculosis strains which were defined as resistant to rifampicin by the Xpert MTB/RIF assay.
Rifampicin-resistant isolates from 43 adult patients (≥ 18 years) from various districts of Malawi were characterised for mutations in the RRDR (codons 507–533) of the rpoB gene by DNA sequencing.
Mutations were found in 37/43 (86%) of the resistant isolates in codons 511, 512, 513, 516, 522, 526 and 531. The most common mutations were in codons 526 (38%), 531 (29.7%) and 516 (16.2%). Mutations were not found in 6/43 (14%) of the resistant isolates. No novel rpoB mutations other than those previously described were found among the rifampicin-resistant M. tuberculosis complex strains.
This study is the first to characterise rifampicin resistance in Malawi. The chain-termination DNA sequencing employed in this study is a standard method for the determination of nucleotide sequences and can be used to confirm rifampicin resistance obtained using other assays, including the Xpert MTB/RIF. Further molecular cluster analysis, such as spoligotyping and DNA finger printing, is still required to determine transmission dynamics and the epidemiological link of the mutated strains.
Tuberculosis remains an important public health problem especially in the developing world. The global impact of tuberculosis is significant, with an annual estimate of 9.6 million tuberculosis cases and over 1.5 million deaths due to tuberculosis in 2014.1 The tuberculosis burden is worsened by the emergence and spread of multi-drug resistant (MDR) tuberculosis cases, defined as simultaneous resistance to at least rifampicin and isoniazid, with or without resistance to any other drug.1
The 644 specimens from NTRL were categorised clinically as previously treated cases (574/644; 89.1%), new (27/644; 4.2%), or not known (43/644; 6.7%). From Bwaila, 24/351 (6.8%) suspected tuberculosis cases were categorised as retreatment, and 327/351 (93.2%) were from patients presenting with tuberculosis for the first time.
This study is the first to characterise rifampicin resistance in Malawi. The majority of rpoB gene mutations in this study analysed by direct sequencing correlated well with rifampicin resistance observed on Xpert MTB/RIF. Our findings reveal that 86% of all rifampicin-resistant isolates harboured mutations in the RRDR of the rpoB gene with codon 526 (CAC → TAC) as the most frequent, followed by codon 531 (TCG → TTG/TAG). By contrast, mutations were not detected in 6/43 (14%) of the strains following nucleotide sequencing. False-positive rifampicin resistance was most likely considering the observed wild-type sequences in these strains. Data on drug-resistance mutations involving the RRDR of the rpoB gene, which relates highly to rifampicin resistance, is new for Malawi as no previous studies have analysed the rpoB gene among tuberculosis strains circulating in the country.