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Transfer of an Escherichia coli ST131 multiresistance cassette has created a Klebsiella pneumoniae-specific plasmid associated with a major nosocomial outbreak
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 Sciences, Clinical Bacteriology. (Åsa Melhus)
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Clinical Bacteriology.
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2012 (English)In: Journal of Antimicrobial Chemotherapy, ISSN 0305-7453, E-ISSN 1460-2091, Vol. 67, no 1, 74-83 p.Article in journal (Refereed) Published
Abstract [en]


To characterize the complete sequence, horizontal spread and stability of the CTX-M-15-encoding multiresistance plasmid of a Klebsiella pneumoniae strain involved in a large nosocomial outbreak.


The 220 kbp plasmid pUUH239.2 was completely sequenced using 454 technology. The conjugational host range, conjugation frequencies, plasmid stability and fitness cost of plasmid carriage were studied in vitro. Conjugational spread during the outbreak was assessed retrospectively by multiplex PCR screening, restriction fragment length polymorphism and PFGE.


Plasmid pUUH239.2 encodes resistance to β-lactams (blaCTX-M-15, blaTEM-1 and blaOXA-1), aminoglycosides [aac-(6′)-1b-cr and aadA2], tetracyclines [tet(A) and tetR], trimethoprim (dhfrXII), sulphonamides (sul1) quaternary ammonium compounds (qacEΔ1), macrolides [mph(A)-mxr-mphR(A)] and heavy metal ions (silver, copper and arsenic). The plasmid consists of a backbone, highly similar to the K. pneumoniae plasmid pKPN3, and a 41 kbp resistance region, highly similar to the resistance regions of plasmids pEK499 and pC15-1a previously isolated from Escherichia coli strains belonging to the outbreak lineage ST131 (where ST stands for sequence type). The pUUH239.2 plasmid is stable in K. pneumoniae but unstable in E. coli and confers a fitness cost when introduced into a naive host cell. Transfer of pUUH239.2 from the outbreak K. pneumoniae clone to the E. coli of the patients’ intestinal floras has occurred on multiple occasions during the outbreak.


The plasmid pUUH239.2 is a composite of the pKPN3 K. pneumoniae plasmid backbone and the blaCTX-M-15-encoding multiresistance cassette associated with the internationally recognized outbreak strain E. coli ST131. The resulting plasmid differs in stability between K. pneumoniae and E. coli, and this has probably limited the spread of this plasmid during the outbreak.

Place, publisher, year, edition, pages
2012. Vol. 67, no 1, 74-83 p.
Keyword [en]
O25:H4-ST131, ESBL, outbreak, horizontal transfer, fitness-cost, evolution, CTX-M-15
National Category
Microbiology in the medical area
Research subject
URN: urn:nbn:se:uu:diva-169437DOI: 10.1093/jac/dkr405ISI: 000300833700012OAI: oai:DiVA.org:uu-169437DiVA: diva2:506693
Available from: 2012-02-29 Created: 2012-02-29 Last updated: 2015-08-28
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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Sandegren, LinusLinkevicius, MariusLytsy, BirgittaÅsa, MelhusAndersson, Dan I.
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