Molecular characterization of mutations associated with resistance to second-line tuberculosis drug among multidrug-resistant tuberculosis patients from high prevalence tuberculosis city in Morocco

Abstract

BackgroundThe emergence of extensively drug-resistant tuberculosis (XDR-TB) has raised public health concern for global TB control. Although multi drug-resistant tuberculosis (MDR- TB) prevalence and associated genetic mutations in Morocco are well documented, scarce information on XDR TB is available. Hence, the evaluation of pre-XDR and XDR prevalence, as well as the mutation status of gyrA, gyrB, rrs, tlyA genes and eis promoter region, associated with resistance to second line drugs, is of great value for better management of M/XDR TB in Morocco.ObjectivesTo evaluate pre-XDR and XDR prevalence, as well as the mutation status of gyrA, gyrB, rrs, tlyA genes and eis promoter region, associated with resistance to second line drug resistance, in 703 clinical isolates from TB patients recruited in Casablanca, and to assess the usefulness of molecular tools in clinical laboratories for better management of M/XDR TB in Morocco.MethodsDrug susceptibility testing (DST) was performed by the proportional method for first line drugs, and then the selected MDR isolates were tested for second line drugs (Ofloxacin, Kanamycin, Amikacin and Capreomycin). Along with DST, all samples were subjected to rpoB, katG and p-inhA mutation analysis by PCR and DNA sequencing. MDR isolates as well as 30 pan-susceptible strains were subjected to PCR and DNA sequencing of gyrA, gyrB, rrs, tlyA genes and eis promoter, associated with resistance to fluoroquinolones and injectable drugs.ResultsAmong the 703 analysed strains, 12.8% were MDR; Ser531Leu and Ser315Thr being the most common recorded mutations within rpoB and katG genes associated with RIF and INH resistance respectively. Drug susceptibility testing for second line drugs showed that among the 90 MDR strains, 22.2% (20/90) were resistant to OFX, 2.22% (2/90) to KAN, 3.33% (3/90) to AMK and 1.11% (1/90) to CAP. Genotypic analysis revealed that 19 MDR strains harbored mutations in the gyrA gene; the most recorded mutation being Asp91Ala accounting for 47.6% (10/21), and 2 isolates harbored mutations in the promoter region of eis gene. No mutation was found in gyrB, rrs and tlyA genes. Moreover, none of the pan-susceptible isolates displayed mutations in targeted genes.ConclusionMost of mutations associated with SLD resistance occurred in gyrA gene (codons 90-94) and eis promoter region. These findings highlight the impact of mutations in gyrA on the development of fluroquinolones resistance and provide the first estimates of the proportion of pre-XDR-TB among MDR-TB cases in Morocco.

Whole-genome sequencing analysis of multidrug-resistant Mycobacterium tuberculosis from Java, Indonesia

“…Multidrug-resistant TB (MDR-TB, resistant to at least isoniazid and rifampicin), pre-extensively drug-resistant TB [pre-XDR-TB, resistant to at least isoniazid, rifampicin, and a fluoroquinolone (FQ) or a second-line injectable agent] and extensively drug-resistant TB (XDR-TB, resistant to at least isoniazid, rifampicin, an FQ and a second-line injectable agent) remains a public health crisis [3]. Infections with MDR-TB, pre-XDR-TB and XDR-TB incur higher costs of treatment, lower treatment success rates and higher mortality rates among TB patients [3, 4].…”

“…Drug susceptibility testing (DST) is required to guide effective treatment [5]. The gold-standard phenotypic testing method is technically laborious, has a long turnaround time of around 4–6 weeks and has a high rate of contamination causing delays in effective treatment [3, 6].…”

Detection of Second Line Drug Resistance among Drug Resistant Mycobacterium Tuberculosis Isolates in Botswana

“…There were no mutations detected in the gyrB gene in any of the isolates in this study. The mutations in the gyrB gene are usually associated with low level resistance to fluoroquinolones and are not as common as those in the gyrA gene [2]. The rrs MUT1 A1401G mutation which leads to a high level second-line injectable drug resistance was detected among 5% of the MDR-TB isolates.…”

“…In the same year, an increase in cases of rifampicin monoresistant and multidrug resistant TB (MDR-TB defined as TB that is resistant to rifampicin and isoniazid) from 490,000 in 2016 to 558,000 in 2017 was observed [1]. The increasing numbers of rifampicin and MDR-TB cases poses a risk to TB control programs throughout the world [2]. Rifampicin monoresistance is considered to be a precursor to MDR-TB and there are often concerns about rifampicin monoresistant TB patients acquiring MDR-TB [3,4].…”

“…Rifampicin monoresistance is considered to be a precursor to MDR-TB and there are often concerns about rifampicin monoresistant TB patients acquiring MDR-TB [3,4]. The standardized World Health Organization (WHO) MDR-TB treatment regimen recommends the use of second-line injectable drugs (SLIDs) in combination with flouroquinolones as part of the standardized MDR-TB treatment regimen [2]. The resistance to a fluoroquinolone and a SLID negatively impacts treatment outcome and has been defined as extensively drug resistant TB (XDR-TB) [5][6][7].…”