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Potential of tetracycline resistance proteins to evolve tigecycline resistance
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. (Dan I. Andersson)
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. (Dan I. Andersson)
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. (Dan I. Andersson)
2016 (English)In: Antimicrobial Agents and Chemotherapy, ISSN 0066-4804, E-ISSN 1098-6596, Vol. 60, no 2, 789-796 p.Article in journal (Refereed) Published
Abstract [en]

Tigecycline is a glycylcycline antibiotic active against multidrug-resistant bacterial pathogens. The objectives of our study were to examine the potential of the Tet(A), Tet(K), Tet(M), and Tet(X) tetracycline resistance proteins to acquire mutations causing tigecycline resistance and to determine how this affects resistance to earlier classes of tetracyclines. Mutations in all four tet genes caused a significant increase in the tigecycline MIC in Escherichia coli, and strains expressing mutant Tet(A) and Tet(X) variants reached clinically relevant MICs (2 mg/liter and 3 mg/liter, respectively). Mutations predominantly accumulated in transmembrane domains of the efflux pumps, most likely increasing the accommodation of tigecycline as a substrate. All selected Tet(M) mutants contained at least one mutation in the functionally most important loop III of domain IV. Deletion of leucine 505 of this loop led to the highest increase of the tigecycline MIC (0.5 mg/liter) among Tet(M) mutants. It also caused collateral sensitivity to earlier classes of tetracyclines. A majority of the Tet(X) mutants showed increased activity against all three classes of tetracylines. All tested Tet proteins have the potential to acquire mutations leading to increased MICs of tigecycline. As tet genes are widely found in pathogenic bacteria and spread easily by horizontal gene transfer, resistance development by alteration of existing Tet proteins might compromise the future medical use of tigecycline. We predict that Tet(X) might become the most problematic future Tet determinant, since its weak intrinsic tigecycline activity can be mutationally improved to reach clinically relevant levels without collateral loss in activity to other tetracyclines.

Place, publisher, year, edition, pages
2016. Vol. 60, no 2, 789-796 p.
National Category
Microbiology in the medical area
URN: urn:nbn:se:uu:diva-257805DOI: 10.1128/AAC.02465-15ISI: 000369159800009OAI: oai:DiVA.org:uu-257805DiVA: diva2:843645
EU, European Research Council, 282004Swedish Research Council, K2013-56X-13025-15- 5Swedish Research Council, 2013-5476-25194-9Swedish Research Council, K2013-99X-22208-01-5
Available from: 2015-07-30 Created: 2015-07-08 Last updated: 2016-03-02Bibliographically approved
In thesis
1. Evolution and Mechanisms of Tigecycline Resistance in Escherichia coli
Open this publication in new window or tab >>Evolution and Mechanisms of Tigecycline Resistance in Escherichia coli
2015 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Antibiotic resistance is an ongoing global medical crisis and we are in great need of new antibacterial agents to combat rapidly emerging resistant pathogens. Tigecycline is one of few drugs that have been introduced into medicine during the last two decades. It is a broad-spectrum third generation tetracycline that is active against multidrug-resistant bacteria that cause complicated infections.

In this thesis I examined the development of tigecycline resistance in Escherichia coli and associated in vitro and in vivo fitness effects. Selections of spontaneous E. coli mutants revealed relatively high accumulation rates of changes in the multidrug efflux system AcrAB-TolC regulation network and in heptose biosynthesis and transport pathways important for lipopolysaccharide (LPS) synthesis. Both groups of mutations led to reduced susceptibility to tigecycline and slower growth compared to the wild-type bacteria. Additional in vitro fitness assays and in vivo competitions showed that LPS mutants were less fit than efflux mutants, providing a possible explanation for why up-regulation of multidrug efflux pumps is the main tigecycline resistance mechanism reported in clinical isolates.

Tigecycline was designed to evade the two most common tetracycline resistance mechanisms conferred by Tet proteins, efflux and ribosomal protection. However, tigecycline is a substrate for the tetracycline modifying enzyme Tet(X). Screening of Tet protein mutant libraries showed that it is possible to select Tet mutants with minimal inhibitory concentrations of tigecycline that reach clinically relevant levels. Mutations in Tet proteins that permitted a better protection from tigecycline frequently exhibited reduced activity against earlier generations of tetracyclines, except for the Tet(X) enzyme mutants, which were better at inactivating all tested tetracyclines. This is particularly worrisome because different variants of Tet(X) have recently spread to multidrug-resistant pathogens through horizontal gene transfer. Therefore, Tet(X) mutants with improved activity threaten the medical future of tetracyclines.

Multidrug resistance is easily disseminated through horizontally spreading conjugative plasmids. pUUH239.2 is an example of a successful conjugative plasmid that caused the first clonal outbreak of extended spectrum β-lactamase-producing Klebsiella pneumoniae in Scandinavia. This plasmid was formed after rearrangements between two different plasmid backbones and it carries resistance genes to multiple antibiotic classes, heavy metals, and detergents.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2015. 58 p.
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine, ISSN 1651-6206 ; 1121
Tigecycline, Bacterial resistance, Fitness, Escherichia coli, AcrAB, LPS, Tet proteins, Tet(A), Tet(K), Tet(M), Tet(X), tet genes, pUUH239.2
National Category
Medical and Health Sciences Natural Sciences
Research subject
urn:nbn:se:uu:diva-259226 (URN)978-91-554-9285-4 (ISBN)
Public defence
2015-09-25, B42, BMC, Husargatan 3, Uppsala, 09:00 (English)
Available from: 2015-08-26 Created: 2015-07-30 Last updated: 2015-10-01

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