JAC Advance Access originally published online on March 16, 2005
Journal of Antimicrobial Chemotherapy 2005 55(5):810-811; doi:10.1093/jac/dki080
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Correspondence |
Panresistance in VIM-1-producing Klebsiella pneumoniae
1 Laboratory of Bacteriology, Hellenic Pasteur Institute, Vas. Sofias 127, Athens 11521, 2 First Department of Propaedeutic Medicine and 3 Department of Microbiology, Medical School, University of Athens, Athens, Greece
* Corresponding author. Tel: +30-210-6478810; Fax: +30-210-6423498; Email: miriagou{at}mail.pasteur.gr
Keywords: K. pneumoniae , carbapenems , resistance , metallo-ß-lactamases , VIM-1
Sir,
The term ‘panresistance’ usually refers to non-fermenting bacilli, such as Pseudomonas and Acinetobacter spp., resistant to all clinically available antimicrobials, except colistin. We report here on four Klebsiella pneumoniae clinical isolates exhibiting this pattern of resistance. The isolates, derived from patients treated in intensive care units in three teaching hospitals in Athens during 2002–2004, were positive for metallo-ß-lactamase (MBL) production. Determination of antibiotic susceptibility by the microdilution method showed high levels of resistance to carboxy- and ureido-penicillins, penicillin–inhibitor combinations, cefoxitin, aztreonam, and late-generation cephalosporins, including cefepime and cefpirome (all MICs were 
128 mg/L). MICs of imipenem and meropenem were 32 mg/L. MIC values above the respective resistance breakpoints were also observed for aminoglycosides (AMGs) (gentamicin, tobramycin, netilmicin and amikacin), ciprofloxacin, trimethoprim, sulphonamides and chloramphenicol.
Isolates were compared by PFGE in 1% agarose, after digestion of total DNA with XbaI. The resulting fingerprints differed by three or fewer DNA fragments, indicating a possible common ancestry, according to established criteria.1
The observed PFGE patterns are also common among VIM-producing K. pneumoniae exhibiting resistance phenotypes other than the one described here. Additionally, conjugative transfer experiments, using a highly rifampicin-resistant Escherichia coli laboratory strain as recipient, followed by plasmid DNA analysis, showed that all isolates harboured the same two plasmids, with approximate sizes of 110 and 50 kb. The plasmids were readily segregated, were both self-transmissible, and conferred distinct multiresistant phenotypes. The 110 kb plasmid was similar to the SHV-5-encoding plasmids that belong to incompatibility group L/M (IncL/M) and are widely disseminated among clinical enterobacteria in Europe, and especially in the Mediterranean region. PCR assays based on published sequences of the above mentioned IncL/M plasmids (AJ245670
[GenBank]
and AF550679
[GenBank]
in GenBank) and sequencing of the amplicons, showed that the 110 kb plasmid carried a blaSHV-5 gene adjacent to an integron similar to In-T3, that contained aacC1, dhfrI, aadA and sulI.2
This plasmid also mediated resistance to chloramphenicol, but the respective determinant was not examined. The antibiotic resistance region of the 50 kb plasmid included an intact class 1 integron containing blaVIM-1, aacA4, dhfrI, aadA and sulI, as described previously (AY339625
[GenBank]
in GenBank).3
This plasmid was similar to a group of IncN, VIM-1-encoding plasmids, found recently in clinical enterobacteria in Greece.3,4
In addition, a mutation in the gyrA gene associated with resistance to quinolones (Ser-83
Phe) was detected in all isolates. gyrB and parC were not examined.
Panresistant K. pneumoniae have been noticed in the intensive care units of various Athens hospitals since 2001, coinciding with the emergence of MBL-positive enterobacteria. We have shown that this phenotype can be caused by the acquisition of two multiresistant plasmids, able to co-exist within a single cell, since they belong to different incompatibility groups. Until now, VIM-1-producing K. pneumoniae were susceptible to aztreonam—a poor substrate for the VIM-1 MBL5— and gentamicin, which remains unaffected by the AMG-modifying enzymes encoded by the VIM-1 plasmids. However, in the isolates presented here, these two drugs are also rendered inactive, by expression of blaSHV-5 and aacC1, carried by the 110 kb plasmid. Another notable characteristic of these isolates was their high-level resistance to carbapenems, contrary to the majority of VIM-producing enterobacteria, where carbapenem MICs remain within the susceptible range. Carbapenemase activity of cell extracts, assessed by spectrophotometry, did not reveal significant differences in the VIM-1 amounts produced by these highly resistant isolates, compared with the less resistant ones. Therefore, it is possible that mutations affecting intracellular carbapenem accumulation contribute to the elevated MICs, but this was not examined further.
PFGE typing suggested that the isolates may be genetically related. This clonal hypothesis is not contradicted by the identical gyrA mutations. Despite the propitious and ubiquitous antibiotic selection pressure in the nosocomial environment, these panresistant strains still only account for a small fraction of all VIM-positive K. pneumoniae isolated. Nevertheless, emergence of other enterobacterial clones with a similar phenotype can be expected, given that both plasmids are endemic in this country.
The limited number of cases and the serious nature of patients’ conditions that led to frequent changes in antibiotic treatment, do not allow us at present to make any firm suggestions for an optimal therapeutic scheme. So far, colistin, various antibiotic combinations, and non-conventional dosing schemes have been proposed for the treatment of life-threatening infections caused by panresistant non-fermenting bacilli. Our isolates appeared susceptible to colistin by disc diffusion. However, we also determined the activity of aztreonam in the presence of clavulanic acid (2 mg/L) and tazobactam (4 mg/L), and found that both inhibitors partly restored activity of the antibiotic, lowering the MICs to 1–2 mg/L. This approach could therefore also be of clinical relevance.
References
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Villa, L., Pezzela, C., Tosini, F. et al. (2000). Multiple-antibiotic resistance mediated by structurally related IncL/M plasmids carrying an extended-spectrum ß-lactamase gene and a class 1 integron. Antimicrobial Agents and Chemotherapy 44, 2911–4.
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Miriagou, V., Tzelepi, E., Gianneli, D. et al. (2003). Escherichia coli with a self-transferable, multiresistant plasmid coding for the metallo-ß-lactamase VIM-1. Antimicrobial Agents and Chemotherapy 47, 395–7.
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Giakkoupi, P., Xanthaki, A., Kanelopoulou, M. et al. (2003). VIM-1 metallo-ß-lactamase-producing Klebsiella pneumoniae strains in Greek hospitals. Journal of Clinical Microbiology 41, 3893–6.
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Franceschini, N., Caravelli, B., Docquier, J.-D. et al. (2000). Purification and biochemical characterization of the VIM-1 metallo-ß-lactamase. Antimicrobial Agents and Chemotherapy 44, 3003–7.
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