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Mutation supply and relative fitness shape the genotypes of ciprofloxacin-resistant Escherichia coli
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
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2017 (English)In: Molecular biology and evolution, ISSN 0737-4038, E-ISSN 1537-1719, Vol. 34, no 5, 1029-1039 p.Article in journal (Refereed) Published
Abstract [en]

Ciprofloxacin is an important antibacterial drug targeting Type II topoisomerases, highly active against Gram-negatives including Escherichia coli. The evolution of resistance to ciprofloxacin in E. coli always requires multiple genetic changes, usually including mutations affecting two different drug target genes, gyrA and parC. Resistant mutants selected in vitro or in vivo can have many different mutations in target genes and efflux regulator genes that contribute to resistance. Among resistant clinical isolates the genotype, gyrA S83L D87N, parC S80I is significantly overrepresented suggesting that it has a selective advantage. However, the evolutionary or functional significance of this high frequency resistance genotype is not fully understood. By combining experimental data and mathematical modeling, we addressed the reasons for the predominance of this specific genotype. The experimental data were used to model trajectories of mutational resistance evolution under different conditions of drug exposure and population bottlenecks. We identified the order in which specific mutations are selected in the clinical genotype, showed that the high frequency genotype could be selected over a range of drug selective pressures, and was strongly influenced by the relative fitness of alternative mutations and factors affecting mutation supply. Our data map for the first time the fitness landscape that constrains the evolutionary trajectories taken during the development of clinical resistance to ciprofloxacin and explain the predominance of the most frequently selected genotype. This study provides strong support for the use of in vitro competition assays as a tool to trace evolutionary trajectories, not only in the antibiotic resistance field.

Place, publisher, year, edition, pages
2017. Vol. 34, no 5, 1029-1039 p.
Keyword [en]
ciprofloxacin, multistep evolution, population bottleneck, modeling evolution, clinical isolates
National Category
Biochemistry and Molecular Biology
Identifiers
URN: urn:nbn:se:uu:diva-264874DOI: 10.1093/molbev/msx052PubMedID: 28087782OAI: oai:DiVA.org:uu-264874DiVA: diva2:862228
Funder
Swedish Research CouncilSwedish Foundation for Strategic Research Knut and Alice Wallenberg Foundation
Available from: 2015-10-20 Created: 2015-10-19 Last updated: 2017-05-11Bibliographically approved
In thesis
1. Evolution of Antibiotic Resistance
Open this publication in new window or tab >>Evolution of Antibiotic Resistance
2015 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

The emergence of antimicrobial resistance is a major global threat to modern medicine. The rapid dissemination of resistant pathogens and the associated loss of efficacy of many important drugs needs to be met with the development of new antibiotics and alternative treatment options. A better understanding of the evolution of resistance could help in developing strategies to slow down the spread of antimicrobial drug resistance.

In this thesis we investigated the evolution of resistance to two important antibiotics, rifampicin and ciprofloxacin, paying special attention to the resistance patterns occurring with high frequency in clinical isolates.

Rifampicin is a first-line drug in tuberculosis treatment and resistance to this valuable drug limits treatment options. Our work on rifampicin resistance helps to explain the extreme bias seen in the frequency of specific resistance mutations in resistant clinical isolates of M. tuberculosis. We identified an important interplay between the level of resistance, relative fitness and selection of fitness-compensatory mutations among the most common resistant isolates.

Fluoroquinlones are widely used to treat infections with Gram-negatives and the frequency of resistance to these important drugs is increasing. Resistance to fluoroquinolones is the result of a multi-step evolutionary process. Our studies on the development of resistance to the fluoroquinolone drug ciprofloxacin provide insights into the evolutionary trajectories and reveal the order in which susceptible wild-type E. coli acquire multiple mutations leading to high level of resistance. We found that the evolution of ciprofloxacin resistance is strongly influenced by the mutation supply rate and by the relative fitness of competing strains at each successive step in the evolution. Our data show that different classes of resistance mutations arise in a particular, predictable order during drug selection. We also uncovered strong evidence for the existence of a novel class of mutations affecting transcription and translation, which contribute to the evolution of resistance to ciprofloxacin.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2015. 67 p.
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine, ISSN 1651-6206 ; 1150
Keyword
ciprofloxacin, rifampicin, bacterial fitness, compensation, mutation rate
National Category
Natural Sciences
Research subject
Biology with specialization in Evolutionary Genetics
Identifiers
urn:nbn:se:uu:diva-265018 (URN)978-91-554-9380-6 (ISBN)
Public defence
2015-12-11, A1:111a, BMC, Husargatan 3, Uppsala, 09:15 (English)
Opponent
Supervisors
Available from: 2015-11-18 Created: 2015-10-20 Last updated: 2016-01-13
2. Biased Evolution: Causes and Consequences
Open this publication in new window or tab >>Biased Evolution: Causes and Consequences
2016 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

In evolution alternative genetic trajectories can potentially lead to similar phenotypic outcomes. However, certain trajectories are preferred over others. These preferences bias the genomes of living organisms and the underlying processes can be observed in ongoing evolution.

We have studied a variety of biases that can be found in bacterial chromosomes and determined the selective causes and functional consequences for the cell. We have quantified codon usage bias in highly expressed genes and shown that it is selected to optimise translational speed. We further demonstrated that the resulting differences in decoding speed can be used to regulate gene expression, and that the use of ‘non-optimal’ codons can be detrimental to reading frame maintenance. Biased gene location on the chromosome favours recombination between genes within gene families and leads to co-evolution. We have shown that such recombinational events can protect these gene families from inactivation by mobile genetic elements, and that chromosome organization can be selectively maintained because inversions can lead to the formation of unstable hybrid operons.

We have used the development of antibiotic resistance to study how different bacterial lifestyles influence evolutionary trajectories. For this we used two distinct pairs of antibiotics and disease-causing bacteria, namely (i) Mycobacterium tuberculosis that is treated with rifampicin and (ii) Escherichia coli that is treated with ciprofloxacin. We have shown that in the slow-growing Mycobacterium tuberculosis, resistance mutations are selected for high-level resistance. Fitness is initially less important, and over time fitness costs can be ameliorated by compensatory mutations. The need for rapid growth causes the selection of ciprofloxacin resistance in Escherichia coli not only to be selected on the basis of high-level resistance but also on high fitness. Compensatory evolution is therefore not required and is not observed.

Taken together, our results show that the evolution of a phenotype is the product of multiple steps and that many factors influence which trajectory is the most likely to occur and be most beneficial. Over time, selection will favour this particular trajectory and lead to biased evolution, affecting genome sequence and organization.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2016. 48 p.
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine, ISSN 1651-6206 ; 1198
Keyword
Evolution, Codon usage bias, Post-transcriptional regulation, Recombination, Inversion, EF-Tu, Frameshift suppression, Antibiotic resistance, Rifampicin, Ciprofloxacin, Compensatory evolution, Drug efflux, RNA polymerase, DNA gyrase
National Category
Microbiology
Research subject
Microbiology
Identifiers
urn:nbn:se:uu:diva-276456 (URN)978-91-554-9518-3 (ISBN)
Public defence
2016-05-09, A1:107a, BMC, Husargatan 3, Uppsala, 09:00 (English)
Opponent
Supervisors
Available from: 2016-04-13 Created: 2016-02-13 Last updated: 2016-04-21

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Huseby, Douglas L.Pietsch, FranziskaBrandis, GerritGaroff, LinnéaTegehall, AngelicaHughes, Diarmaid

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