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Research article
Fangyou Yu
Tongji University Affiliated Shanghai Pulmonary Hospital
Corresponding Author
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Background: Resistance to rifampin (RIF) in Mycobacterium tuberculosis infection is associated with mutations in the rpoB gene coding for the beta-subunit of RNA polymerase. The contribution of many individual rpoB mutations to the development and level of RIF resistance remains elusive. Our objective for this study was to investigate the relationship between specific rpoB mutations and the minimum inhibitory concentrations (MICs) of RIF and rifabutin (RFB) against M.tuberculosis. Methods: We collected 195 clinical isolates of M. tuberculosis including 105 RIF-resistant and 90 RIF-susceptible isolates from Shanghai Pulmonary Hospital in China. The MICs of antituberculosis drugs in 7H10 Middlebrook medium for clinical isolates of M. tuberculosis were determined. Strains were screened for rpoB mutations by DNA extraction, rpoB gene amplification, and DNA sequencing analysis. Results: Twenty different types of mutations were identified in the rpoB gene. One hundred isolates (95.24%) were found to have mutations in the RIF resistance-determining region (RRDR) of the rpoB gene. Three rpoB mutations were identified in 90 RIF-susceptible isolates. Out of 105 isolates, 86 (81.90%) were cross-resistant to both RIF and RFB. The most frequent mutation occurred at codon 531 (65.71%), followed by 526 (8.57%). We also found a novel nine-nucleotide (ATCATGCAT) deletion (between positions 1543 and 1551) in the rpoB gene among two strains (1.90%) with resistance to RIF, but susceptibility to RFB. In addition, the mutation frequency at codon 531 was significantly higher in RIF-resistant/RFB-resistant (RIFR/RFBR) strains than in RIFR/RFBS strains (75.58% versus 21.05%), whereas the mutation frequency at codon 516 was significantly lower in RIFR/RFBR strains than in RIFR/RFBS strains (1.16% versus 26.32%). The MICs of RIF against 87.62% (92/105) of the M.tuberculosis isolates were ≥ 16 µg/mL. Conclusions: Our data supported previous findings that various rpoB mutations are associated with differential levels of resistance to RIF. The specific mutations of the rpoB gene in RIFR/RFBR isolates differed from those in RIFR/RFBS isolates. A novel deletion mutation in the RRDR might be associated with resistance to RIF, but not to RFB. Further clinical studies are required to investigate the efficacy of RFB in the treatment of M. tuberculosis infections, which harbor the mutations.
Keywords
M. tuberculosis, Rifampin, Rifabutin, rpoB mutations, MICs
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A new preprint design is coming!
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This is a preprint, a preliminary version of a manuscript that has not completed peer review at a journal. Research Square does not conduct peer review prior to posting preprints. The posting of a preprint on this server should not be interpreted as an endorsement of its validity or suitability for dissemination as established information or for guiding clinical practice.
Research article
Fangyou Yu
Tongji University Affiliated Shanghai Pulmonary Hospital
Corresponding Author
Badges
Prescreen
This manuscript passed our Prescreen assessment
Complete author information
Appropriate declaration statements
Potential risks to human health
Prescreen
History
CURRENT STATUS: Posted
Version 1
Posted 23 Jul, 2019
No community comments so far
Metrics
Comments: 0
PDF Downloads: ...
HTML Views: ...
Subject Areas
Infectious Diseases
License:
This work is licensed under a CC BY 4.0 License. Read Full License
Background: Resistance to rifampin (RIF) in Mycobacterium tuberculosis infection is associated with mutations in the rpoB gene coding for the beta-subunit of RNA polymerase. The contribution of many individual rpoB mutations to the development and level of RIF resistance remains elusive. Our objective for this study was to investigate the relationship between specific rpoB mutations and the minimum inhibitory concentrations (MICs) of RIF and rifabutin (RFB) against M.tuberculosis. Methods: We collected 195 clinical isolates of M. tuberculosis including 105 RIF-resistant and 90 RIF-susceptible isolates from Shanghai Pulmonary Hospital in China. The MICs of antituberculosis drugs in 7H10 Middlebrook medium for clinical isolates of M. tuberculosis were determined. Strains were screened for rpoB mutations by DNA extraction, rpoB gene amplification, and DNA sequencing analysis. Results: Twenty different types of mutations were identified in the rpoB gene. One hundred isolates (95.24%) were found to have mutations in the RIF resistance-determining region (RRDR) of the rpoB gene. Three rpoB mutations were identified in 90 RIF-susceptible isolates. Out of 105 isolates, 86 (81.90%) were cross-resistant to both RIF and RFB. The most frequent mutation occurred at codon 531 (65.71%), followed by 526 (8.57%). We also found a novel nine-nucleotide (ATCATGCAT) deletion (between positions 1543 and 1551) in the rpoB gene among two strains (1.90%) with resistance to RIF, but susceptibility to RFB. In addition, the mutation frequency at codon 531 was significantly higher in RIF-resistant/RFB-resistant (RIFR/RFBR) strains than in RIFR/RFBS strains (75.58% versus 21.05%), whereas the mutation frequency at codon 516 was significantly lower in RIFR/RFBR strains than in RIFR/RFBS strains (1.16% versus 26.32%). The MICs of RIF against 87.62% (92/105) of the M.tuberculosis isolates were ≥ 16 µg/mL. Conclusions: Our data supported previous findings that various rpoB mutations are associated with differential levels of resistance to RIF. The specific mutations of the rpoB gene in RIFR/RFBR isolates differed from those in RIFR/RFBS isolates. A novel deletion mutation in the RRDR might be associated with resistance to RIF, but not to RFB. Further clinical studies are required to investigate the efficacy of RFB in the treatment of M. tuberculosis infections, which harbor the mutations.
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