Prevalence and patterns of rifampicin and isoniazid resistance conferring mutations in <i>Mycobacterium tuberculosis</i> isolates from Uganda

<div><p>Background</p><p>Accurate diagnosis of tuberculosis, especially by using rapid molecular assays, can reduce transmission of drug resistant tuberculosis in communities. However, the frequency of resistance conferring mutations varies with geographic location of <i>Mycobacterium tuberculosis</i>, and this affects the efficiency of rapid molecular assays in detecting resistance. This has created need for characterizing drug resistant isolates from different settings to investigate frequencies of resistance conferring mutations. Here, we describe the prevalence and patterns of rifampicin- and isoniazid- resistance conferring mutations in isolates from Uganda, which could be useful in the management of MDR-TB patients in Uganda and other countries in sub-Saharan Africa.</p><p>Results</p><p>Ninety seven <i>M</i>. <i>tuberculosis</i> isolates were characterized, of which 38 were MDR, seven rifampicin-resistant, 12 isoniazid-mono-resistant, and 40 susceptible to rifampicin and isoniazid. Sequence analysis of the <i>rpoB</i> rifampicin-resistance determining region (<i>rpoB</i>/RRDR) revealed mutations in six codons: 588, 531, 526, 516, 513, and 511, of which Ser531Leu was the most frequent (40%, 18/45). Overall, the three mutations (Ser531Leu, His526Tyr, Asp516Tyr) frequently associated with rifampicin-resistance occurred in 76% of the rifampicin resistant isolates while 18% (8/45) of the rifampicin-resistant isolates lacked mutations in <i>rpoB</i>/RRDR. Furthermore, sequence analysis of <i>katG</i> and <i>inhA</i> gene promoter revealed mainly the Ser315Thr (76%, 38/50) and C(-15)T (8%, 4/50) mutations, respectively. These two mutations combined, which are frequently associated with isoniazid-resistance, occurred in 88% of the isoniazid resistant isolates. However, 20% (10/50) of the isoniazid-resistant isolates lacked mutations both in <i>katG</i> and <i>inhA</i> gene promoter. The sensitivity of sequence analysis of <i>rpoB</i>/RRDR for rifampicin-resistance via detection of high confidence mutations (Ser531Leu, His526Tyr, Asp516Tyr) was 81%, while it was 77% for analysis of <i>katG</i> and <i>inhA</i> gene promoter to detect isoniazid-resistance via detection of high confidence mutations (Ser315Thr, C(-15)T, T(-8)C). Furthermore, considering the circulating TB genotypes in Uganda, the isoniazid-resistance conferring mutations were more frequent in <i>M</i>. <i>tuberculosis</i> lineage 4/sub-lineage Uganda, perhaps explaining why this genotype is weakly associated with MDR-TB.</p><p>Conclusion</p><p>Sequence analysis of <i>rpoB</i>/RRDR, <i>katG</i> and <i>inhA</i> gene promoter is useful in detecting rifampicin/isoniazid resistant <i>M</i>. <i>tuberculosis</i> isolates in Uganda however, about ≤20% of the resistant isolates lack known resistance-conferring mutations hence rapid molecular assays may not detect them as resistant.</p></div

Frequency of rifampicin/isoniazid-resistance conferring mutations in the circulating <i>M</i>. <i>tuberculosis</i> genotypes/sub-lineages in Uganda (2009–2011).

<p>Frequency of rifampicin/isoniazid-resistance conferring mutations in the circulating <i>M</i>. <i>tuberculosis</i> genotypes/sub-lineages in Uganda (2009–2011).</p

Flow Chart of patient enrollment in the National Anti-TB drug resistance survey in Uganda; December 2009–February 2011.

<p>Figure legend: **NTM = Non Tuberculous Mycobacteria.</p

Frequency and patterns of resistance conferring mutations in <i>M</i>. <i>tuberculosis</i> isolates that were resistant to rifampicin (n = 45) and isoniazid (n = 50).

<p>Frequency and patterns of resistance conferring mutations in <i>M</i>. <i>tuberculosis</i> isolates that were resistant to rifampicin (n = 45) and isoniazid (n = 50).</p

Analysis of factors associated with Multi drug resistance in Uganda; December 2009–February 2011.

<p>Variable included in the multivariate model were, age, sex, residence and previous history of TB treatment.</p>**<p>Analysis limited to univariate level as inclusion at multivariate level masked the apparent association between MDR and potential risk factors.</p