Expression analysis of efflux pump genes among drug-susceptible and multidrug-resistant Mycobacterium tuberculosis clinical isolates and reference strains

Calgin, Mustafa Kerem; Şahin, Fikret; Turegun, Buse; Gerçeker, Devran; Atasever, Melike; Köksal, Deniz; Karasartova, Djursun; Kıyan, Mehmet

doi:10.1016/j.diagmicrobio.2013.02.033

Cited by 41 publications

(30 citation statements)

References 21 publications

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“…Several recent studies have demonstrated the importance of the overexpression of efflux pump genes in MDR and XDR M . tuberculosis clinical strains (Calgin et al, 2013; Coelho et al, 2015; Li et al, 2015a; Yamchi et al, 2015; Kanji et al, 2016; Machado et al, 2016; Oh et al, 2017), in rifampicin monoresistant strains (Li et al, 2015b), or in the H37Rv susceptible strain after exposure to drugs (Garima et al, 2015; Caleffi-Ferracioli et al, 2016). Nevertheless, most of these studies are based on the simple assessment and evaluation of the levels of expression of M. tuberculosis efflux pump genes, and few have determined the effect of efflux inhibitors on the MICs of the antituberculosis drugs and have quantified the activity of the overexpressed efflux systems.…”

Section: Discussionmentioning

confidence: 99%

Interplay between Mutations and Efflux in Drug Resistant Clinical Isolates of Mycobacterium tuberculosis

et al. 2017

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Numerous studies show efflux as a universal bacterial mechanism contributing to antibiotic resistance and also that the activity of the antibiotics subject to efflux can be enhanced by the combined use of efflux inhibitors. Nevertheless, the contribution of efflux to the overall drug resistance levels of clinical isolates of Mycobacterium tuberculosis is poorly understood and still is ignored by many. Here, we evaluated the contribution of drug efflux plus target-gene mutations to the drug resistance levels in clinical isolates of M. tuberculosis. A panel of 17 M. tuberculosis clinical strains were characterized for drug resistance associated mutations and antibiotic profiles in the presence and absence of efflux inhibitors. The correlation between the effect of the efflux inhibitors and the resistance levels was assessed by quantitative drug susceptibility testing. The bacterial growth/survival vs. growth inhibition was analyzed through the comparison between the time of growth in the presence and absence of an inhibitor. For the same mutation conferring antibiotic resistance, different MICs were observed and the different resistance levels found could be reduced by efflux inhibitors. Although susceptibility was not restored, the results demonstrate the existence of a broad-spectrum synergistic interaction between antibiotics and efflux inhibitors. The existence of efflux activity was confirmed by real-time fluorometry. Moreover, the efflux pump genes mmr, mmpL7, Rv1258c, p55, and efpA were shown to be overexpressed in the presence of antibiotics, demonstrating the contribution of these efflux pumps to the overall resistance phenotype of the M. tuberculosis clinical isolates studied, independently of the genotype of the strains. These results showed that the drug resistance levels of multi- and extensively-drug resistant M. tuberculosis clinical strains are a combination between drug efflux and the presence of target-gene mutations, a reality that is often disregarded by the tuberculosis specialists in favor of the almost undisputed importance of antibiotic target-gene mutations for the resistance in M. tuberculosis.

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Section: Discussionmentioning

confidence: 99%

Interplay between Mutations and Efflux in Drug Resistant Clinical Isolates of Mycobacterium tuberculosis

et al. 2017

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“…It was reported previously that mutations or overexpression of Rv0194 and Rv2686c are associated with increased resistance to multiple drugs in M. tuberculosis [26, 27]. But according to another recent study which aimed to compare the differences of the expression of 15 putative multidrug efflux pump genes in clinically isolated drug sensitive and MDR M. tuberculosis isolates, all the tested putative multidrug efflux pump genes in the drug-sensitive and MDR M. tuberculosis isolates have similar rates of expression [28]. Thus, the existence of mutations and over expression of the efflux pump genes might not be necessarily associated with increased drug resistance.…”

Section: Discussionmentioning

confidence: 99%

Comparative genomic analysis of Mycobacterium tuberculosis clinical isolates

Liu

Wang

et al. 2014

BMC Genomics

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BackgroundDue to excessive antibiotic use, drug-resistant Mycobacterium tuberculosis has become a serious public health threat and a major obstacle to disease control in many countries. To better understand the evolution of drug-resistant M. tuberculosis strains, we performed whole genome sequencing for 7 M. tuberculosis clinical isolates with different antibiotic resistance profiles and conducted comparative genomic analysis of gene variations among them.ResultsWe observed that all 7 M. tuberculosis clinical isolates with different levels of drug resistance harbored similar numbers of SNPs, ranging from 1409–1464. The numbers of insertion/deletions (Indels) identified in the 7 isolates were also similar, ranging from 56 to 101. A total of 39 types of mutations were identified in drug resistance-associated loci, including 14 previously reported ones and 25 newly identified ones. Sixteen of the identified large Indels spanned PE-PPE-PGRS genes, which represents a major source of antigenic variability. Aside from SNPs and Indels, a CRISPR locus with varied spacers was observed in all 7 clinical isolates, suggesting that they might play an important role in plasticity of the M. tuberculosis genome. The nucleotide diversity (Л value) and selection intensity (dN/dS value) of the whole genome sequences of the 7 isolates were similar. The dN/dS values were less than 1 for all 7 isolates (range from 0.608885 to 0.637365), supporting the notion that M. tuberculosis genomes undergo purifying selection. The Л values and dN/dS values were comparable between drug-susceptible and drug-resistant strains.ConclusionsIn this study, we show that clinical M. tuberculosis isolates exhibit distinct variations in terms of the distribution of SNP, Indels, CRISPR-cas locus, as well as the nucleotide diversity and selection intensity, but there are no generalizable differences between drug-susceptible and drug-resistant isolates on the genomic scale. Our study provides evidence strengthening the notion that the evolution of drug resistance among clinical M. tuberculosis isolates is clearly a complex and diversified process.Electronic supplementary materialThe online version of this article (doi: 10.1186/1471-2164-15-469) contains supplementary material, which is available to authorized users.

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“…tuberculosis drug-resistant strains have been described to increase the expression of efflux pumps to enable their survival and persistence in the presence of antibiotics (41)(42)(43)(44). Although we did not explore drug-resistant strains in this study, additional research to try and reverse drug resistance using TIM and clinical isolates of M. tuberculosis and evaluation of these in broth and infected macrophage cultures, as well as in vivo studies, are warranted.…”

Section: Discussionmentioning

confidence: 99%

The Efflux Pump Inhibitor Timcodar Improves the Potency of Antimycobacterial Agents

Grossman

Shoen

Jones

et al. 2015

Antimicrob Agents Chemother

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bPrevious studies indicated that inhibition of efflux pumps augments tuberculosis therapy. In this study, we used timcodar (formerly VX-853) to determine if this efflux pump inhibitor could increase the potency of antituberculosis (anti-TB) drugs against Mycobacterium tuberculosis in in vitro and in vivo combination studies. When used alone, timcodar weakly inhibited M. tuberculosis growth in broth culture (MIC, 19 g/ml); however, it demonstrated synergism in drug combination studies with rifampin, bedaquiline, and clofazimine but not with other anti-TB agents. When M. tuberculosis was cultured in host macrophage cells, timcodar had about a 10-fold increase (50% inhibitory concentration, 1.9 g/ml) in the growth inhibition of M. tuberculosis and demonstrated synergy with rifampin, moxifloxacin, and bedaquiline. In a mouse model of tuberculosis lung infection, timcodar potentiated the efficacies of rifampin and isoniazid, conferring 1.0 and 0.4 log 10 reductions in bacterial burden in lung, respectively, compared to the efficacy of each drug alone. Furthermore, timcodar reduced the likelihood of a relapse infection when evaluated in a mouse model of long-term, chronic infection with treatment with a combination of rifampin, isoniazid, and timcodar. Although timcodar had no effect on the pharmacokinetics of rifampin in plasma and lung, it did increase the plasma exposure of bedaquiline. These data suggest that the antimycobacterial drug-potentiating activity of timcodar is complex and drug dependent and involves both bacterial and host-targeted mechanisms. Further study of the improvement of the potency of antimycobacterial drugs and drug candidates when used in combination with timcodar is warranted.

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Expression analysis of efflux pump genes among drug-susceptible and multidrug-resistant Mycobacterium tuberculosis clinical isolates and reference strains

Cited by 41 publications

References 21 publications

Interplay between Mutations and Efflux in Drug Resistant Clinical Isolates of Mycobacterium tuberculosis

Interplay between Mutations and Efflux in Drug Resistant Clinical Isolates of Mycobacterium tuberculosis

Comparative genomic analysis of Mycobacterium tuberculosis clinical isolates

The Efflux Pump Inhibitor Timcodar Improves the Potency of Antimycobacterial Agents

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