JAC Advance Access originally published online on April 3, 2006
Journal of Antimicrobial Chemotherapy 2006 57(6):1026-1029; doi:10.1093/jac/dkl110
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Leading articles |
Fluoroquinolone resistance and plasmid addiction systems: self-imposed selection pressure?
Antibiotic Resistance Monitoring and Reference Laboratory, Centre for Infections, Health Protection Agency 61 Colindale Avenue, London NW9 5EQ, UK
*Corresponding author. Tel: +44-208-8327-7236; Fax +44-208-8327-6264; E-mail: matthew.ellington{at}hpa.org.uk
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Multi-antibiotic-resistant Gram-negative pathogens are becoming more prevalent and an association exists between chromosomally conferred fluoroquinolone resistance and the presence of plasmid-borne resistances, such as extended spectrum ß-lactamases. This link is not wholly explained by strain spread or the presence of fluoroquinolone-modifying enzymes. Plasmid-encoded toxin–antitoxin addiction systems enforce plasmid maintenance in bacteria and, like fluoroquinolones, some toxins target DNA gyrase. Bacteria can develop resistance to these toxins, which would free the cell of the plasmid addiction and allow it to ditch the ‘excess baggage’. We hypothesize that these plasmid-encoded gyrase toxins might contribute to, or predispose towards, clinically significant fluoroquinolone resistance, and that the plasmid-encoded quinolone resistance determinant, Qnr, may facilitate this. Establishing the extent and mechanisms of cross-resistance to toxins and fluoroquinolones will aid the management of resistance and may contribute to the development of novel antimicrobials.
Keywords: post-segregational killing , toxin–antitoxin systems , toxin resistance , Qnr , DNA gyrase
Multi-antibiotic-resistant Gram-negative pathogens are becoming more prevalent and are threatening public health in the face of a dearth of new antibiotics against them.1 Resistance to the widely used fluoroquinolone class of antibacterials occurs in 15–20% of >16 000 bacteraemias caused by Escherichia coli in the UK per annum, and is similarly prevalent in other common Enterobacteriaceae such as Klebsiella spp. and Enterobacter spp.2 In many bacterial species there is a strong association between clinically significant fluoroquinolone resistance, contingent on chromosomal mutation, and the carriage of plasmids, including those that encode extended-spectrum ß-lactamases.3,4 This correlation between chromosomal and plasmid-mediated resistances occurs also in Gram-positive species, such as in successful strains of methicillin-resistant Staphylococcus aureus5 and highly gentamicin-resistant Enterococcus faecalis.6,7 Moreover, as multiple strains of many species show this association it is apparent that clonal strain success is not the only factor involved, raising the question of what else is driving this relationship.
Fluoroquinolones target topoisomerase IV and DNA gyrase and, until recently, resistance was considered to be entirely mediated by chromosomal mutations. Despite this, conflicting data exist amongst clinical isolates for a link between hypermutability and the emergence of fluoroquinolone resistance.8,9 Most clinically significant fluoroquinolone resistance is are associated with mutations in the quinolone resistance determining regions (QRDRs) of gyrA-gyrB or parC-parE (encoding DNA gyrase and topoisomerase IV subunits, respectively).10 Relatively little is known of the importance of gyr and par mutations outside of the QRDRs, but these have recently been associated with fluoroquinolone resistance.11,12 This raises the possibility that changes at other sites in DNA gyrase or topoisomerase IV could play either a direct role in fluoroquinolone resistance or in predisposition towards its development. In E. coli little consideration has been given to such possibilities thus far.
Low-level and incremental fluoroquinolone resistance can also be conferred by chromosomal mutations that cause overexpression of the Acr system, and can increase antibiotic efflux.13,14 Recently, other mechanisms of low-level fluoroquinolone resistance associated with plasmids have impacted upon our understanding. The gene encoding QnrA has been found on multiresistance plasmids in Enterobacteriaceae worldwide.15,16 QnrA interacts with DNA gyrase and topoisomerase IV,17,18 protecting against fluoroquinolone binding and conferring a modest increase in fluoroquinolone MICs. This widens the mutant selection window by increasing the mutant prevention concentration 
10-fold, facilitating an increase in the frequency of the emergence of clinically significant high-level resistance via QRDR mutations.19 The occurrence of QnrA on multiresistance plasmids might explain the association between high-level quinolone resistance and plasmid-mediated resistances. However, large variations in Qnr prevalence have been reported worldwide, and large European surveys have found qnr genes in only 0.5% of quinolone-resistant isolates in France, and of quinolone- or cephalosporin-resistant isolates in Germany,15 and 3% of cephalosporin-resistant Enterobacteriaceae in the UK.20 Thus QnrA appears to be a relatively minor factor contributing to the association between fluoroquinolone resistance and multiresistance. The global prevalence of other Qnr determinants such as QnrB and QnrS15 is currently unknown. A further plasmid-mediated determinant—a novel variant of a common aminoglycoside-acetyltransferase [aac(6')-Ib-cr]—was found in 51% of 71 ciprofloxacin-resistant clinical isolates from Shanghai. This variant modifies ciprofloxacin and confers a 4-fold decrease in ciprofloxacin susceptibility, in addition to kanamycin, tobramycin and amikacin resistance.21 Like QnrA, this mechanism also eases the development of higher-level fluoroquinolone resistance in E. coli. Despite its high prevalence in the sample set tested, its global prevalence is currently unknown although it has been reported in North America and China.19 Only 21% of the 466 ciprofloxacin-resistant E. coli referred to the Antibiotic Resistance Monitoring and Reference Laboratory (ARMRL) at the Health Protection Agency's Centre for Infections in London, between 2004 and 2006, were resistant to both amikacin and tobramycin compared with 7% of the 168 ciprofloxacin-susceptible isolates. Thus, even when the prevalences of QnrA and AAC(6')-Ib-cr are considered together, these plasmidic mechanisms are unlikely to account entirely for the association between ciprofloxacin resistance and multiresistance in E. coli or other bacteria. It is likely that other factors also contribute to this association.
Successful strains of pathogenic bacteria often carry multiple plasmids and theoretically are disposed to elevated rates of plasmid loss as a result of partitioning mistakes during cell division. In reality plasmid loss may be prevented by plasmid-encoded ‘addiction systems’ [also called post-segregation killing or toxin–antitoxin (TA) systems], which work as TA pairs. The greater longevity of the toxin compared with the antitoxin means that daughter cells that lose the plasmid are no longer able to produce the antitoxin, and become exposed to the residual toxin with fatal consequences.22 TA systems not only enforce plasmid stability, but also act to exclude other plasmids of the same compatibility group. While they have been known for some time,23,24 they have only been identified on resistance plasmids relatively recently.25 At least six plasmid-mediated TA systems have been identified in E. coli.22 The molecular target of three of these systems has been identified. The CcdB and ParE toxins (distinct from the ParE subunit of topoisomerase IV) from plasmids F and RK2, respectively, both target and interact with the GyrA subunit of DNA gyrase.26,27 In this functional respect, they resemble fluoroquinolones and QnrA. While the CcdB toxin and its antitoxin CcdA form a tight complex,28 exposure to even a small amount of free toxin would be analogous with exposure to subinhibitory concentrations of an antibiotic and might select for resistance. Resistance to such toxins would potentially allow the bacterium to circumvent plasmid addiction and ditch the ‘excess baggage’. It would even allow the bacterium to acquire a new plasmid of the same compatibility group, permitting gene flux amongst the total cellular DNA—providing an advantage in competitive and stressful environments. Indeed, resistance to these ‘addictive’ and ‘parasitic’22 toxins can occur via changes in the CcdB toxin itself, which can attenuate the toxic effect of the protein.29,30 In addition, target changes can occur; an amino acid substitution in the GyrA protein (Arg-462
Cys) confers resistance to CcdB, but this position is situated far from the QRDR of GyrA, and the Cys substitution does not affect fluoroquinolone susceptibility.31 Nevertheless, the small number of toxin resistance mutations so far described is unlikely to be comprehensive, and it is possible that amino acid substitutions at non-QRDR sites in DNA gyrase may provide a degree of cross-resistance to both toxins and fluoroquinolones, providing a further putative link between fluoroquinolone resistance and plasmidic resistances. At least one route of toxin-fluoroquinolone cross-resistance is already known; E. coli can resist or tolerate the CcdB toxin via up-regulation of the intrinsic chromosomally encoded protein GyrI, which also confers decreased susceptibility to fluoroquinolones.32,33 Like QnrA, GyrI interacts with DNA gyrase and inhibits formation of a drug–gyrase–DNA ternary complex. By analogy, QnrA, a DNA gyrase inhibitor known to confer quinolone resistance, might also influence the development of toxin resistance. Moreover, given that quinolones are synthetic compounds, it seems fortuitous that a natural protein, Qnr, confers resistance to them, and tempts speculation that toxin resistance could be one of the (currently unknown) true physiological roles of Qnr-like proteins. A precedent for protection against such self-encoded or ‘self-imposed’ pressure is set by another pentapeptide repeat protein related to Qnr, McbG, which also inhibits DNA gyrase, and contributes to immunity to the colicin ‘microcin B17’,34–36 effectively protecting bacteria producing this toxin from self-inhibition.
Clearly there are multiple natural and synthetic inhibitors of DNA gyrase, each with different effects. These can be encoded by the cell's own genome or by the genomes of other cells, or they may be present as synthetic antibacterials. To date, there is no simple explanation for the association between multiple plasmid-mediated resistances and high-level fluoroquinolone resistance. It is likely to be multifactorial, and we propose that this association may, in part, represent self-imposed resistance development, exacerbated by the presence of multiple plasmids which may encode TA systems that target DNA gyrase—which is a critical target for fluoroquinolone antibiotics. Importantly, the effect of mechanisms developed to resist DNA-gyrase-targeting toxins on the development of mutations affecting fluoroquinolone resistance remains unknown. However, the hypothesis could be tested if plasmids encoding gyrase-targeting toxins were identified and the frequency of the emergence of fluoroquinolone-resistant mutants determined in the absence and presence of the plasmids. But in a naive host this may be complicated if mutational toxin resistance is required in a multistep pathway before predisposition towards quinolone resistance is detectable. Further additional interesting questions include the matter of plasmid stability and the functional relationship between Qnr and GyrI for the evolution and maintenance of resistance. ‘Plasmid addiction’ toxins are a focus for the development of novel antimicrobials through structural studies,37,38 and a novel bacteriophage delivery technology to facilitate their use has been developed.39 It is of utmost importance to understand the development of resistance to these toxins in order to assess their efficacy or longevity as potential novel agents. In addition, the toxins have fundamental relevance, not only to multiresistant E. coli, which is a major pathogen causing increasing public health concern as it becomes more multiresistant, but also to other Gram-negative and Gram-positive pathogens where plasmidic resistance such as high-level gentamicin-resistance is associated with fluoroquinolone resistance.
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We confirm no conflicting financial interests.
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We would like to thank Dr Russell Hope (ARMRL) for supplying aminoglycoside susceptibility data.
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1 Livermore DM. (2003) The threat from the pink corner. Ann Med 35:226–34.[CrossRef][Web of Science][Medline]
2 CDR Weekly Anonymous. (2004) Escherichia coli bacteraemias, England, Wales, and Northern Ireland: 2003. 14:No 38 Available at: http://www.hpa.org.uk/cdr/PDFfiles/2004/Bact_3804.pdf (5 February 2006, date last accessed).
3 Livermore DM. (2000) Epidemiology of antibiotic resistance. Intensive Care Med 26:Suppl 1, S14–21.[Medline]
4 Paterson DL, Mulazimoglu L, Casellas JM, et al. (2000) Epidemiology of ciprofloxacin resistance and its relationship to extended-spectrum ß-lactamase production in Klebsiella pneumoniae isolates causing bacteremia. Clin Infect Dis 30:473–8.[CrossRef][Web of Science][Medline]
5 Cox RA, Mallaghan C, Conquest C, et al. (1995) Epidemic methicillin-resistant Staphylococcus aureus: controlling the spread outside hospital. J Hosp Infect 29:107–19.[CrossRef][Web of Science][Medline]
6
Woodford N, Reynolds R, Turton J, et al. (2003) Two widely disseminated strains of Enterococcus faecalis highly resistant to gentamicin and ciprofloxacin from bacteraemias in the UK and Ireland. J Antimicrob Chemother 52:711–4.
7
Hallgren A, Saeedi B, Nilsson M, et al. (2003) Genetic relatedness among Enterococcus faecalis with transposon-mediated high-level gentamicin resistance in Swedish intensive care units. J Antimicrob Chemother 52:162–7.
8
Komp Lindgren P, Karlsson A, Hughes D. (2003) Mutation rate and evolution of fluoroquinolone resistance in Escherichia coli isolates from patients with urinary tract infections. Antimicrob Agents Chemother 47:3222–32.
9 Baquero M, Nilsson AI, Turrientes MC, et al. The frequency of weak-hypermutable E. coli strains correlates with pathogenic and geographical locations but not with quinolone-resistance. Abstracts of the Forty-fourth Interscience Conference on Antimicrobial Agents and Chemotherapy, Washington, DC, 2004(American Society for Microbiology, Washington, DC, USA) Abstract C2-1888, p. 125.
10
Ruiz J. (2003) Mechanisms of resistance to quinolones: target alterations, decreased accumulation and DNA gyrase protection. J Antimicrob Chemother 51:1109–17.
11
Aubry A, Veziris N, Cambau E, et al. (2006) Novel gyrase mutations in quinolone-resistant and -hypersusceptible clinical isolates of Mycobacterium tuberculosis: functional analysis of mutant enzymes. Antimicrob Agents Chemother 50:104–12.
12
Friedman SM, Lu T, Drlica K. (2001) Mutation in the DNA gyrase A gene of Escherichia coli that expands the quinolone resistance-determining region. Antimicrob Agents Chemother 45:2378–80.
13
Wang H, Dzink-Fox JL, Chen M, et al. (2001) Genetic characterization of highly fluoroquinolone-resistant clinical Escherichia coli strains from China: role of acrR mutations. Antimicrob Agents Chemother 45:1515–21.
14
Webber MA and Piddock LJV. (2003) The importance of efflux pumps in bacterial antibiotic resistance. J Antimicrob Chemother 51:9–11.
15
Nordmann P and Poirel L. (2005) Emergence of plasmid-mediated resistance to quinolones in Enterobacteriaceae. J Antimicrob Chemother 56:463–9.
16
Corkill JE, Anson JJ, Hart CA. (2005) High prevalence of the plasmid-mediated quinolone resistance determinant qnrA in multidrug-resistant Enterobacteriaceae from blood cultures in Liverpool, UK. J Antimicrob Chemother 56:1115–7.
17
Tran JH and Jacoby GA. (2002) Mechanism of plasmid-mediated quinolone resistance. Proc Natl Acad Sci USA 99:5638–42.
18
Tran JH, Jacoby GA, Hooper DC. (2005) Interaction of the plasmid-encoded quinolone resistance protein Qnr with Escherichia coli DNA gyrase. Antimicrob Agents Chemother 49:118–25.
19 Jacoby GA. (2005) Mechanisms of resistance to quinolones. Clin Infect Dis 41:S120–6.[CrossRef]
20 Ellington MJ, Hope R, Woodford R, et al. Detection of QnrA amongst extended-spectrum and AmpC ß-lacatamase producers from South East England. Abstracts of the Forty-fifth Interscience Conference on Antimicrobial Agents and Chemotherapy, Washington, DC, 2005(American Society for Microbiology, Washington, DC, USA) Abstract C2-786, p. 119.
21 Robicsek A, Strahilevitz J, Jacoby GA, et al. (2006) Fluoroquinolone-modifying enzyme: a new adaptation of a common aminoglycoside acetyltransferase. Nat Med 12:83–8.[CrossRef][Web of Science][Medline]
22
Hayes F. (2003) Toxins-antitoxins: plasmid maintenance, programmed cell death, and cell cycle arrest. Science 301:1496–9.
23 Lehnherr H, Maguin E, Jafri S, et al. (1993) Plasmid addiction genes of bacteriophage P1: doc, which causes cell death on curing of prophage, and phd, which prevents host death when prophage is retained. J Mol Biol 233:414–28.[CrossRef][Web of Science][Medline]
24 Weaver KE and Tritle DJ. (1994) Identification and characterization of an Enterococcus faecalis plasmid pAD1-encoded stability determinant which produces two small RNA molecules necessary for its function. Plasmid 32:168–81.[CrossRef][Medline]
25 Grady R and Hayes F. (2003) Axe-Txe, a broad-spectrum proteic toxin-antitoxin system specified by a multidrug-resistant, clinical isolate of Enterococcus faecium. Mol Microbiol 47:1419–32.[CrossRef][Web of Science][Medline]
26 Kampranis SC, Howells AJ, Maxwell A. (1999) The interaction of DNA gyrase with the bacterial toxin CcdB: evidence for the existence of two gyrase-CcdB complexes. J Mol Biol 293:733–44.[CrossRef][Web of Science][Medline]
27 Jiang Y, Pogliano J, Helinski DR, et al. (2002) ParE toxin encoded by the broad-host-range plasmid RK2 is an inhibitor of Escherichia coli gyrase. Mol Microbiol 44:971–9.[CrossRef][Web of Science][Medline]
28 Bernard P and Couturier M. (1992) Cell killing by the F plasmid CcdB protein involves poisoning of DNA-topoisomerase II complexes. J Mol Biol 226:735–45.[CrossRef][Web of Science][Medline]
29 Bahassi EM, Salmon MA, Van Melderen L, et al. (1995) F plasmid CcdB killer protein: ccdB gene mutants coding for non-cytotoxic proteins which retain their regulatory functions. Mol Microbiol 15:1031–7.[Web of Science][Medline]
30
Bahassi EM, O'Dea MH, Allali N, et al. (1999) Interactions of CcdB with DNA gyrase. Inactivation of GyrA, poisoning of the gyrase-DNA complex, and the antidote action of CcdA. J Biol Chem 274:10936–44.
31
Scheirer KE and Higgins NP. (1997) The DNA cleavage reaction of DNA gyrase. Comparison of stable ternary complexes formed with enoxacin and CcdB protein. J Biol Chem 272:27202–9.
32 Chatterji M, Sengupta S, Nagaraja V. (2003) Chromosomally encoded gyrase inhibitor GyrI protects Escherichia coli against DNA-damaging agents. Arch Microbiol 180:339–46.[CrossRef][Web of Science][Medline]
33 Chatterji M and Nagaraja V. (2002) GyrI: a counter-defensive strategy against proteinaceous inhibitors of DNA gyrase. EMBO Rep 3:261–7.[CrossRef][Web of Science][Medline]
34 Heddle JG, Blance SJ, Zamble DB, et al. (2001) The antibiotic microcin B17 is a DNA gyrase poison: characterisation of the mode of inhibition. J Mol Biol 307:1223–34.[CrossRef][Web of Science][Medline]
35
Zamble DB, Miller DA, Heddle JG, et al. (2001) In vitro characterization of DNA gyrase inhibition by microcin B17 analogs with altered bisheterocyclic sites. Proc Natl Acad Sci USA 98:7712–7.
36 Garrido MC, Herrero M, Kolter R, et al. (1988) The export of the DNA replication inhibitor microcin B17 provides immunity for the host cell. EMBO J 7:1853–62.[Web of Science][Medline]
37 DeNap JC and Hergenrother PJ. (2005) Bacterial death comes full circle: targeting plasmid replication in drug-resistant bacteria. Org Biomol Chem 3:959–66.[CrossRef][Web of Science][Medline]
38 Dao-Thi MH, Van Melderen L, De Genst E, et al. (2005) Molecular basis of gyrase poisoning by the addiction toxin CcdB. J Mol Biol 348:1091–102.[CrossRef][Web of Science][Medline]
39
Westwater C, Kasman LM, Schofield DA, et al. (2003) Use of genetically engineered phage to deliver antimicrobial agents to bacteria: an alternative therapy for treatment of bacterial infections. Antimicrob Agents Chemother 47:1301–7.
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