Dataset Information

In vitro and in vivo fitness costs associated with Mycobacterium tuberculosis RpoB mutation H526D.

ABSTRACT:

Aim

There is controversy regarding the potential fitness costs of rifampicin (RIF) resistance-conferring mutations in the Mycobacterium tuberculosis (Mtb) rpoB gene. We characterized the pathogenicity of an Mtb RpoB H526D mutant.

Materials & methods

A mutant containing the RpoB H526D mutation was isolated from wild-type Mtb grown on RIF-containing plates and complemented for determination of in vitro and in vivo fitness costs.

Results

The RpoB H526D mutant showed reduced survival relative to control strains during progressive hypoxia and delayed growth following resuscitation from nutrient starvation (p < 0.05), which was associated with reduced expression of the resuscitation-promoting factor genes rpfB, rpfC and rpfE. Relative to the isogenic wild-type strain, the mutant showed significantly attenuated growth and long-term survival as well as reduced inflammation in mouse lungs. Conclusion & future perspective: Our data suggest that RpoB H526D mutation confers a fitness cost during growth-limiting conditions in vitro and in mouse lungs.

SUBMITTER: Rifat D

PROVIDER: S-EPMC5506874 | biostudies-literature | 2017 Jul

REPOSITORIES: biostudies-literature

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Publications

In vitro and in vivo fitness costs associated with Mycobacterium tuberculosis RpoB mutation H526D.

Rifat Dalin D Campodónico Victoria L VL Tao Jing J Miller James A JA Alp Alpaslan A Yao Yufeng Y Karakousis Petros C PC

Future microbiology 20170327

<h4>Aim</h4>There is controversy regarding the potential fitness costs of rifampicin (RIF) resistance-conferring mutations in the Mycobacterium tuberculosis (Mtb) rpoB gene. We characterized the pathogenicity of an Mtb RpoB H526D mutant.<h4>Materials & methods</h4>A mutant containing the RpoB H526D mutation was isolated from wild-type Mtb grown on RIF-containing plates and complemented for determination of in vitro and in vivo fitness costs.<h4>Results</h4>The RpoB H526D mutant showed reduced su ...[more]

PMID: 28343421

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Project description:Discordant results between genotypic drug susceptibility testing (gDST) and phenotypic DST (pDST) for Mycobacterium tuberculosis isolates with disputed (discordance between gDST and pDST results) mutations affect rifampin (RIF)-resistant (RR) and multidrug-resistant (MDR) tuberculosis (TB) treatments due to a lack of practical clinical guidelines. To investigate the role of disputed rpoB mutations in M. tuberculosis and TB treatment outcomes, initial isolates of 837 clinical RR- or MDR-TB cases confirmed during 2014 to 2018 were retested using agar-based RIF pDST and rpoB gene sequencing. MICs were determined for isolates with disputed rpoB mutations. Disputed rpoB mutations were identified in 77 (9.2%) M. tuberculosis isolates, including 50 (64.9%) and 14 (18.2%) phenotypically RIF- and rifabutin (RFB)-resistant isolates, respectively. The predominant single mutations were those encoding L533P (a change of L to P at position 533) (44.2%) and L511P (20.8%). Most of the isolates harboring mutations encoding L511P (87.5%), H526N (100%), D516Y (70.0%), and L533P (63.6%) had MICs of ≤1 mg/liter, whereas isolates harboring the mutation encoding H526L (75%) had a MIC of >1 mg/liter. Of the 63 cases with treatment outcomes available, 11 (17.5%) cases died, 1 (1.6%) case transferred out, and 51 (81%) cases had favorable outcomes, including 8 and 20 cases treated with standard-dose RIF- and RFB-containing regimens, respectively. Excluding cases that transferred out or received no or 1-day treatment, we observed statistically significant differences between the outcomes using active and inactive fluoroquinolones (FQs) (P = 0.008, odds ratio = 0.05 [95% confidence interval, 0.01 to 0.38]) in 57 cases (where active means a case susceptible to the drug and inactive means a case resistant to the drug or drug not used). We concluded that disputed rpoB mutations are not rare. Depending on the resources available, sequencing and/or MIC testing is recommended for better management of RR- and MDR-TB cases.

Project description:Tuberculosis (TB) is one of the leading causes of death due to an infectious disease. The rise of multi-drug resistance (MDR) in Mycobacterium tuberculosis (Mtb), the causative agent of TB, presents a significant obstacle to TB control. While human studies report dysregulated immune responses during MDR TB, a clear understanding of the host-pathogen interactions of MDR Mtb is lacking. Here, we studied the immune responses induced by Mtb strains carrying two of the most common rifampicin drug-resistance (RDR)-conferring single-nucleotide polymorphisms (SNPs) in the RNA polymerase gene of Mtb, which accounts for nearly 90% of drug-resistance mutations found clinically in Mtb. During Mtb infection of primary human macrophages, we found that pro-inflammatory cytokine production was reduced upon infection with Mtb carrying the H445Y SNP but not the S450L SNP. Using a mouse model, we also characterized the host immune response in vivo following infection. Despite similar establishment of Mtb infection in the lung and dissemination to the peripheral organs, we show that infection with the RDR Mtb rpoB-H445Y strain, but not with the rpoB-S450L strain, resulted in a suppressed lung myeloid and lymphoid immune responses through type I IFN-dependent pathways, relative to wt Mtb. This suppressed host immunity had functional consequences in limiting control of RDR Mtb strains harboring rpoB-H445Y and led to worse pathology during chronic TB disease. Collectively, our results suggest that disease pathogenesis may be associated with specific RDR mutations in Mtb, which may differentially regulate immune responses. IMPORTANCE This study highlights the impact of specific rifampicin-resistance-conferring mutations on the host immune response to Mycobacterium tuberculosis (Mtb), the causative agent of tuberculosis (TB). Clinical reports have previously suggested that multi-drug-resistant) TB patients exhibit altered peripheral immune responses as compared with their drug-sensitive TB counterparts. The murine model of infection with Mtb strains carrying drug-resistance-conferring mutations recapitulated these findings and allowed us to mechanistically interrogate the pathways responsible for driving the divergent immune responses. Our findings underscore the need for greater investigation into bacterial heterogeneity to better appreciate the diversity in host-pathogen interactions during TB disease.

Dataset Information

In vitro and in vivo fitness costs associated with Mycobacterium tuberculosis RpoB mutation H526D.

Aim

Materials & methods

Results

Publications

In vitro and in vivo fitness costs associated with Mycobacterium tuberculosis RpoB mutation H526D.

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