Dataset Information

Evolution of Rifampin Resistance in Escherichia coli and Mycobacterium smegmatis Due to Substandard Drugs.

ABSTRACT: Poor-quality medicines undermine the treatment of infectious diseases, such as tuberculosis, which require months of treatment with rifampin and other drugs. Rifampin resistance is a critical concern for tuberculosis treatment. While subtherapeutic doses of medicine are known to select for antibiotic resistance, the effect of drug degradation products on the evolution of resistance is unknown. Here, we demonstrate that substandard drugs that contain degraded active pharmaceutical ingredients select for gene alterations that confer resistance to standard drugs. We generated drug-resistant Escherichia coli and Mycobacterium smegmatis strains by serially culturing bacteria in the presence of the rifampin degradation product rifampin quinone. We conducted Sanger sequencing to identify mutations in rifampin-resistant populations. Strains resistant to rifampin quinone developed cross-resistance to the standard drug rifampin, with some populations showing no growth inhibition at maximum concentrations of rifampin. Sequencing of the rifampin quinone-treated strains indicated that they acquired mutations in the DNA-dependent RNA polymerase B subunit. These mutations were localized in the rifampin resistance-determining region (RRDR), consistent with other reports of rifampin-resistant E. coli and mycobacteria. Rifampin quinone-treated mycobacteria also had cross-resistance to other rifamycin class drugs, including rifabutin and rifapentine. Our results strongly suggest that substandard drugs not only hinder individual patient outcomes but also restrict future treatment options by actively contributing to the development of resistance to standard medicines.

SUBMITTER: Weinstein ZB

PROVIDER: S-EPMC6325213 | biostudies-literature | 2019 Jan

REPOSITORIES: biostudies-literature

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Evolution of Rifampin Resistance in Escherichia coli and Mycobacterium smegmatis Due to Substandard Drugs.

Weinstein Zohar B ZB Zaman Muhammad H MH

Antimicrobial agents and chemotherapy 20181221 1

Poor-quality medicines undermine the treatment of infectious diseases, such as tuberculosis, which require months of treatment with rifampin and other drugs. Rifampin resistance is a critical concern for tuberculosis treatment. While subtherapeutic doses of medicine are known to select for antibiotic resistance, the effect of drug degradation products on the evolution of resistance is unknown. Here, we demonstrate that substandard drugs that contain degraded active pharmaceutical ingredients sel ...[more]

PMID: 30397062

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Project description:A series of structome analyses, that is, quantitative and three-dimensional structural analysis of a whole cell at the electron microscopic level, have already been achieved individually in Exophiala dermatitidis, Saccharomyces cerevisiae, Mycobacterium tuberculosis, Myojin spiral bacteria, and Escherichia coli. In these analyses, sample cells were processed through cryo-fixation and rapid freeze-substitution, resulting in the exquisite preservation of ultrastructures on the serial ultrathin sections examined by transmission electron microscopy. In this paper, structome analysis of non pathogenic Mycolicibacterium smegmatis, basonym Mycobacterium smegmatis, was performed. As M. smegmatis has often been used in molecular biological experiments and experimental tuberculosis as a substitute of highly pathogenic M. tuberculosis, it has been a task to compare two species in the same genus, Mycobacterium, by structome analysis. Seven M. smegmatis cells cut into serial ultrathin sections, and, totally, 220 serial ultrathin sections were examined by transmission electron microscopy. Cell profiles were measured, including cell length, diameter of cell and cytoplasm, surface area of outer membrane and plasma membrane, volume of whole cell, periplasm, and cytoplasm, and total ribosome number and density per 0.1 fl cytoplasm. These data are based on direct measurement and enumeration of exquisitely preserved single cell structures in the transmission electron microscopy images, and are not based on the calculation or assumptions from biochemical or molecular biological indirect data. All measurements in M. smegmatis, except cell length, are significantly higher than those of M. tuberculosis. In addition, these data may explain the more rapid growth of M. smegmatis than M. tuberculosis and contribute to the understanding of their structural properties, which are substantially different from M. tuberculosis, relating to the expression of antigenicity, acid-fastness, and the mechanism of drug resistance in relation to the ratio of the targets to the corresponding drugs. In addition, data obtained from cryo-transmission electron microscopy examination were used to support the validity of structome analysis. Finally, our data strongly support the most recent establishment of the novel genus Mycolicibacterium, into which basonym Mycobacterium smegmatis has been classified.

Project description:Background and Objectives:Metallic antimicrobial materials are of growing interest due to their potential to control pathogenic and multidrug-resistant bacteria. Yet we do not know if utilizing these materials can lead to genetic adaptations that produce even more dangerous bacterial varieties. Methodology:Here we utilize experimental evolution to produce strains of Escherichia coli K-12 MG1655 resistant to, the iron analog, gallium nitrate (Ga(NO3)3). Whole genome sequencing was utilized to determine genomic changes associated with gallium resistance. Computational modeling was utilized to propose potential molecular mechanisms of resistance. Results:By day 10 of evolution, increased gallium resistance was evident in populations cultured in medium containing a sublethal concentration of gallium. Furthermore, these populations showed increased resistance to ionic silver and iron (III), but not iron (II) and no increase in traditional antibiotic resistance compared with controls and the ancestral strain. In contrast, the control populations showed increased resistance to rifampicin relative to the gallium-resistant and ancestral population. Genomic analysis identified hard selective sweeps of mutations in several genes in the gallium (III)-resistant lines including: fecA (iron citrate outer membrane transporter), insl1 (IS30 tranposase) one intergenic mutations arsC ?/? yhiS; (arsenate reductase/pseudogene) and in one pseudogene yedN ?; (iapH/yopM family). Two additional significant intergenic polymorphisms were found at frequencies > 0.500 in fepD ?/? entS (iron-enterobactin transporter subunit/enterobactin exporter, iron-regulated) and yfgF ?/? yfgG (cyclic-di-GMP phosphodiesterase, anaerobic/uncharacterized protein). The control populations displayed mutations in the rpoB gene, a gene associated with rifampicin resistance. Conclusions:This study corroborates recent results observed in experiments utilizing pathogenic Pseudomonas strains that also showed that Gram-negative bacteria can rapidly evolve resistance to an atom that mimics an essential micronutrient and shows the pleiotropic consequences associated with this adaptation. Lay summary:We utilize experimental evolution to produce strains of Escherichia coli K-12 MG1655 resistant to, the iron analog, gallium nitrate (Ga(NO3)3). Whole genome sequencing was utilized to determine genomic changes associated with gallium resistance. Computational modeling was utilized to propose potential molecular mechanisms of resistance.

Dataset Information

Evolution of Rifampin Resistance in Escherichia coli and Mycobacterium smegmatis Due to Substandard Drugs.

Publications

Evolution of Rifampin Resistance in Escherichia coli and Mycobacterium smegmatis Due to Substandard Drugs.

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