Research letters
9 Poirel L, Lagrutta E, Taylor P et al. Emergence of metallo-b-lactamase NDM-1-producing multidrug-resistant Escherichia coli in Australia. Antimicrob Agents Chemother 2010; 54: 4914–6. 10 Peirano G, Ahmed-Bentley J, Woodford N et al. The characteristics of a metallo-b-lactamase-producing Escherichia coli isolated in Canada from a patient with recent travel to India. In: Abstracts of the Fiftieth Interscience Conference on Antimicrobial Agents and Chemotherapy, Boston, USA, 2010. Abstract C1-675a. American Society of Microbiology, Washington, DC, USA. 11 Wirth T, Falush D, Lan R et al. Sex and virulence in Escherichia coli: an evolutionary perspective. Mol Microbiol 2006; 60: 1136 –51. 12 Diancourt L, Passet V, Verhoef J et al. Multilocus sequence typing of Klebsiella pneumoniae nosocomial isolates. J Clin Microbiol 2005; 43: 4178– 82. 13 Nordmann P, Cuzon G, Naas T. The real threat of Klebsiella pneumoniae carbapenemase-producing bacteria. Lancet Infect Dis 2009; 9: 228–36.
15 Vading M, Samuelsen Ø, Haldorsen B et al. Comparison of disk diffusion, Etest and VITEK2 for detection of carbapenemase-producing Klebsiella pneumoniae with the EUCAST and CLSI breakpoint systems. Clin Microbiol Infect 2010; doi:10.1111/j.1469-0691.2010.03299.x.
J Antimicrob Chemother 2011 doi:10.1093/jac/dkq467 Advance Access publication 21 December 2010
Plasmid-encoded OXA-48 carbapenemase in Escherichia coli from Israel Moran G. Goren, Inna Chmelnitsky, Yehuda Carmeli and Shiri Navon-Venezia* Molecular Epidemiology and Antibiotic Resistance Laboratory, Division of Epidemiology, Tel Aviv Medical Center, affiliated to the Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv, Israel *Corresponding author. Tel: +972-3-6925644; Fax: +972-3-6974332; E-mail:
[email protected]
Keywords: imipenem resistance, oxacillinases, Tn1999, plasmids,
class D b-lactamases Sir, Carbapenem resistance among Enterobacteriaceae in Israel emerged in 2004 and was observed mainly in Klebsiella pneumoniae, but also in Enterobacter species and Escherichia coli. Since its emergence, carbapenem resistance in these species, both in clinical and colonizing isolates, has been rendered by the production of plasmid-mediated K. pneumoniae carbapenemase (KPC). In late 2007, a woman in her early thirties previously diagnosed with acute lymphoblastic leukaemia, was admitted to the haematology ward in Tel Aviv Sourasky Medical Center. The patient had arrived in Israel from Jordan in order to undergo chemotherapy
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14 Karthikeyan K, Thirunarayan MA, Krishnan P. Coexistence of blaOXA-23 with blaNDM-1 and armA in clinical isolates of Acinetobacter baumannii from India. J Antimicrob Chemother 2010; 65: 2253– 4.
and, later, bone marrow transplantation. During the first month after transplantation the patient was intermittently febrile and was treated with various antimicrobials, including piperacillin/tazobactam, amikacin, ciprofloxacin, vancomycin, imipenem and voriconazole. Two months after admission, while being treated with imipenem and voriconazole, the patient suffered from fever, dyspnoea and renal failure, and was transferred to the intensive care unit. During that period a carbapenem-resistant E. coli strain (E. coli 1736) was isolated from a Hickman catheter, leading to removal of the catheter and further treatment with ceftazidime and colistin. Treatment with these antibacterial agents cleared the OXA-48-producing E. coli, yet unfortunately the patient died 3 months later from a systemic Pseudomonas infection. E. coli 1736 was multidrug resistant, showing resistance to penicillins, piperacillin/tazobactam, aminoglycosides, quinolones and carbapenems, but susceptibility to all cephalosporins, aztreonam, tigecycline and colistin (Table 1). Analytical isoelectric focusing (IEF) performed on crude enzyme preparations revealed the presence of two b-lactamases with pIs of 5.4 and 7.2 (data not shown). PCR screening for the presence of b-lactamases in E. coli 1736 indicated the presence of blaTEM-1 and blaOXA-48 genes corresponding to the pIs of the b-lactamases visualized by IEF. Plasmid analysis1 of E. coli 1736 revealed four plasmids, three of around ≤50 kb in size and a larger plasmid of around 100 kb. Plasmid DNA was purified and transformed into E. coli DH10B. Transformant colonies that were screened positive for blaOXA-48 by PCR harboured the 50 kb plasmid. Southern analysis of plasmid DNA derived from these transformants using a blaOXA-48-labelled probe demonstrated the presence of blaOXA-48 on the acquired 50 kb plasmid. Acquisition of this plasmid increased the MICs of imipenem, meropenem and ertapenem without conferring full resistance (Table 1). PCR mapping of the genetic environment surrounding blaOXA-48 was performed in collaboration with the laboratory of Professor P. Nordmann (Hospital de Bicetre, Paris, France). blaOXA-48 was found to be located inside Tn1999.2, similar to the structure described for other enteric strains, such as the E. coli strain from Turkey and the K. pneumoniae strain from Lebanon.2 E. coli 1736 was not resistant to all b-lactam antibiotics as reported for other OXA-48-producing strains,2 yet it presented a high level of resistance to the commonly used carbapenems (MICs ≥16 mg/L), higher than usually seen in KPC-producing E. coli strains isolated in our hospital.3 These high carbapenem MICs suggested the presence of additional resistance mechanisms together with OXA-48 carbapenemase. Outer membrane protein (OMP) produced by E. coli 1736 was determined by PCR and sequencing of ompA, ompC and ompF genes. Further OMP analysis was performed by protein extraction and separation on Tris –Tricine gels using SDS-PAGE followed by mass spectrometry (performed in the Biological Mass Spectrometry Facility at the Weizmann Institute of Science). Both methods indicated the absence of at least one major porin, OmpC. Until the isolation of E. coli 1736, carbapenem resistance in E. coli in our country was exclusively attributed to the Ambler class A carbapenemase KPC.3 This is the first identified Enterobacteriaceae isolate in our country possessing a carbapenem-hydrolysing oxacillinase. To seek the possible origin of this strain we performed multilocus sequence typing (MLST; http://www.pasteur.fr/recherche/genopole/PF8/mlst/EColi. html), which genotyped the strain as sequence type (ST) 2, an E. coli ST that has never been recorded previously in our
JAC
Research letters
Table 1. Antibiotic susceptibility of E. coli 1736, E. coli transformant and E. coli recipient strain DH10B MIC for strain (mg/L)a Antibiotic
E. coli transformant
E. coli DH10B
.32 0.25 0.38 ,1 ,1 .128 64
.32 0.19 0.19 ,1 ,1 .128 64
8 0.095 0.095 ,1 ,1 ,4 ,1
8 4 .4 .8 16 16 .32 4 0.25 0.38
1.5 ,1 ,0.25 ,0.25 1 0.5 1 ,0.5 0.125 0.047
,1 ,1 ,0.25 ,0.25 0.25 0.023 0.008 ,0.5 0.125 0.047
a
Susceptibility testing of all antibiotics was performed using Vitek-2; carbapenem MICs were additionally tested by agar dilution; MICs of imipenem, meropenem and ertapenem below 0.5 mg/L and MICs of ceftriaxone, ceftazidime, tigecycline and colistin were determined by Etest.
nosocomial E. coli collection. The origin of the blaOXA-48-encoding plasmid is also uncertain; restriction fragment length polymorphism analysis showed that it differed from previously studied blaKPC-encoding plasmids carried by other carbapenem-resistant E. coli clones in our hospital.3 Since the emergence of carbapenem-resistant K. pneumoniae in Israel during 2006, an active surveillance programme was implemented in high-risk patients newly admitted to our institution, and at the national level in long-term care facilities. blaOXA-48 was not detected in Enterobacteriaceae prior to and after this single report. Thus, although the patient was hospitalized in our hospital 2 months prior to the isolation of the OXA-48-producing E. coli strain, it is most likely that the strain or the plasmid was carried prior to hospitalization. Class D OXA-48 carbapenemase among Enterobacteriaceae was first reported in 2004 in K. pneumoniae from Turkey4 and is continuously spreading in the Mediterranean area,5,6 as well as in other countries in Europe. Recent reports in the Middle East region stress the urgent need for regional collaboration to confront the spread of resistance.
This work was supported in part by the European Commission Research grant FP7: SATURN—Impact of Specific Antibiotic Therapies on the Prevalence of Human Host Resistant Bacteria (grant no. 241796). This work was performed in partial fulfilment of the requirements for an MSc degree of M. G. G., Sackler Faculty of Medicine, Tel Aviv University, Israel.
Transparency declarations None to declare.
References 1 Barton BM, Harding GP, Zuccarelli AJ. A general method for detecting and sizing large plasmids. Anal Biochem 1995; 226: 235– 40. 2 Carrer A, Poirel L, Yilmaz M et al. Spread of OXA-48-encoding plasmid in Turkey and beyond. Antimicrob Agents Chemother 2010; 54: 1369 –73. 3 Goren MG, Navon-Venezia S, Chmelnitsky I et al. Carbapenem-resistant KPC-2-producing Escherichia coli in a Tel Aviv Medical Center, 2005 to 2008. Antimicrob Agents Chemother 2010; 54: 2687– 91. 4 Poirel L, Heritier C, Tolun V et al. Emergence of oxacillinase-mediated resistance to imipenem in Klebsiella pneumoniae. Antimicrob Agents Chemother 2004; 48: 15 –22. 5 Benouda A, Touzani O, Khairallah MT et al. First detection of oxacillinase-mediated resistance to carbapenems in Klebsiella pneumoniae from Morocco. Ann Trop Med Parasitol 2010; 104: 327–30. 6 Cuzon G, Naas T, Lesenne A et al. Plasmid-mediated carbapenemhydrolysing OXA-48 b-lactamase in Klebsiella pneumoniae from Tunisia. Int J Antimicrob Agents 2010; 36: 91– 3.
J Antimicrob Chemother 2011 doi:10.1093/jac/dkq473 Advance Access publication 1 December 2010
Molecular characterization of high-level fluoroquinolone resistance in a clinical isolate of Haemophilus parainfluenzae Jose´-Manuel Rodrı´guez-Martı´nez1*, Inmaculada Lo´pezHerna´ndez2 and A´lvaro Pascual1,3 1
Department of Microbiology, University of Seville, Seville, Spain; LABSUR, Cruz Roja Victoria Eugenia Hospital, Seville, Spain; 3 University Hospital Virgen Macarena, Seville, Spain 2
*Corresponding author. Tel: +34-954-552862; Fax: +34-954-377413; E-mail:
[email protected]
Acknowledgements The study was presented in part at the Forty-ninth Interscience Conference on Antimicrobial Agents and Chemotherapy, San Francisco, CA, 2009 (Abstract C1-1765). We would like to thank the laboratory of Professor P. Nordmann, Hospital de Bicetre, Paris, France, for confirming the results on Tn1999.2.
Keywords: prostatitis, quinolones, QRDRs, quinolone resistance-
determining regions Sir, Haemophilus parainfluenzae is commonly implicated as a causative organism in respiratory tract infections.1 H. parainfluenzae has also been frequently associated with the genitourinary tract,
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Ampicillin Ceftriaxone Ceftazidime Cefepime Aztreonam Piperacillin Piperacillin/ tazobactam Amikacin Gentamicin Ciprofloxacin Levofloxacin Imipenem Meropenem Ertapenem Doripenem Tigecycline Colistin
E. coli 1736
Funding
