1 Department of Microbiology, School of Medicine, University of Thessalia, Papakyriazi 22, Larissa; 2 Department of Microbiology, School of Medicine, University of Patras; 3 Department of Microbiology, General Hospital of Athens G. Genimatas, Athens; 4 Department of Microbiology, General Hospital Asclepeion, Voula, Athens, Greece
Received 23 July 2003; returned 1 September 2003; revised 23 September 2003; accepted 10 October 2003
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Abstract |
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Materials: A total of 1200 Staphylococcus aureus and 2760 coagulase-negative staphylococci (CoNS), consecutively collected from four Greek hospitals located in different geographical areas, were tested for susceptibility to mupirocin using the Etest and a reference agar dilution method.
Results: Twenty-four S. aureus (2%) and 532 CoNS (19.2%) were found to be mupirocin-resistant during the study period. High-level mupirocin resistance was detected in 20 S. aureus (1.6%) and in 440 CoNS (15.9%), respectively. No variations in the rates of mupirocin-resistant S. aureus in relation to the year of collection were observed. In contrast, the rate of mupirocin-resistant CoNS increased dramatically from 9% in 1999, to 14% in 2000, 20% in 2001 and reached 33% in 2002. PFGE analysis revealed the presence of one main clone (A) among mupirocin-resistant S. aureus and two main clones (i and a) among Staphylococcus epidermidis isolates.
Conclusions: In Greece, the rate of mupirocin-resistant S. aureus has remained low and steady since 1999. The high rate of mupirocin-resistant CoNS (33%) in 2002 was due mainly to clonal dissemination of epidemic hospital clones.
Keywords: Staphylococcus aureus, Staphylococcus epidermidis, mupirocin resistance, epidemic clones, Greece
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Introduction |
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In Greece, mupirocin is only used to eradicate nasal carriage of MRSA in patients and staff. The antibiotic is not used for the treatment of staphylococcal skin infections or for the prevention of bacterial colonization due to coagulase-negative staphylococci. In the present study, we investigated the rate of development of mupirocin-resistant staphylococci (S. aureus and CoNS) in Greek hospitals during 19992002.
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Materials and methods |
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A total of 3960 staphylococcicomprising 1200 S. aureus and 2760 CoNS, consecutively isolated during January 1999December 2002, associated with blood, skin and soft tissue infections, and recovered from clinically significant specimenswere included in the study. The samples were collected from four tertiary care Greek hospitals, located in three geographical areas (Athens, Central Greece and Southwestern Greece). Isolates recovered from different cultures (blood, catheter etc.) from the same patient with the same SmaI pulsotype and the same antibiotic resistance profile were included once. Identification at the species level was carried out by Gram stain, catalase and coagulase tests, and by the API Staph System (bioMerieux, SA Lyon, France).
Susceptibility tests
All isolates were tested using the mupirocin Etest (AB BIODISK, Solna, Sweden), and interpretation of susceptibility test results was conducted following the recommendations of the mupirocin manufacturer. Susceptibility results obtained by Etest were compared with those obtained after MIC determination using the reference agar dilution method.4 Potential co-resistance to 14 antimicrobial agents (ampicillin, oxacillin, trimethoprim/sulfamethoxazole, ofloxacin, clindamycin, erythromycin, gentamicin, tobramycin, rifampicin, tetracycline, fusidic acid, vancomycin, linezolid and quinupristin/dalfopristin) was also determined by the agar diffusion method.5
Detection of ileS-2 and mecA genes
All isolates were tested for the presence of ileS-2 and mecA genes by PCR, as described previously.6 The predicted size of the PCR products were 456 bp and 310 bp for the ileS-2 and mecA fragments, respectively.
PFGE analysis
Molecular typing of the mupirocin-resistant isolates was performed by PFGE analysis.7,8 The banding patterns of the strains were compared visually following the criteria of Tenover et al.7
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Results |
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Among the 1200 S. aureus isolates, 24 (2%) expressed mupirocin resistance during the study period. These 24 isolates were collected from patients; none of them had taken mupirocin treatment for nasal carriage. The distribution of low- and high-level mupirocin resistance in relation to time of isolation is described in Table 1. The rate of mupirocin resistance among S. aureus isolates was low and has remained steady since 1999. MuL was detected only in four mecA-positive S. aureus isolates (MIC 32 mg/L), belonging to clones A (three) and B (one), which have spread in several Greek hospitals.8 MuH (MIC 512 mg/L) was detected in 20 S. aureus isolates (14 mecA-positive), sporadically isolated in two of the four participating hospitals. PFGE analysis revealed that all of the MuH strains belonged to clone A, which expressed a relatively susceptible phenotype (Table 2).
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As expected, all MuH isolates carried the ileS-2 gene, which was not detected in any MuL isolate. Analysis by PFGE showed that, although MuH S. epidermidis strains fell into six distinct clones (i, d, b, g, l, c), the great majority of isolates, 286 out 436 (65.6%), belonged to clone i (Figure 1).9 Before 1999, strains belonging to this clone did not carry the ileS-2 gene, so the resistant mutants have emerged in the last 4 years (data not shown). The MuL S. epidermidis strains were grouped into six different clones (a, e, i, d, c, b), the most dominant being clone a, comprising 64 out of 92 strains (69.56%). PFGE types a, b, i and l have been characterized previously as epidemic clones.9 PFGE types d, e, c and g emerged later, after 2000.
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Discussion |
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Mupirocin resistance is relatively unusual in S. aureus, but it is common and increasing in CoNS. It varies greatly from institution to institution regardless of geographic region monitored. According to the SENTRY antimicrobial surveillance programme 2000, mupirocin resistance rates from bloodstream infections varied by geographic area (USA, Canada, Latin America and Europe) for S. aureus from 1.9% to 5.6% and for CoNS from 12.8% to 39.9%.2 A previous study in 19 European hospitals in 12 countries reported high-level resistance in 1.6% of S. aureus and 5.6% of CoNS isolates, and low-level resistance in 2.3% of S. aureus and 7.2% of CoNS isolates.3
The prevalence of mupirocin-resistant S. aureus in Greek hospitals in this study is lower than that reported in a previous study (1.8% in 2002 versus 4.5% in 1997).10 However, the rate of mupirocin-resistant CoNS has increased dramatically, ranging from 9% in 1999 to 33% in 2002. The predominance of the clones A (among MuH S. aureus, which has spread in several Greek hospitals),8 and i and a (among MuH and MuL S. epidermidis strains), already characterized as epidemic clones,9 suggests that a limited number of mupirocin-resistant clones has been disseminated in the Greek hospital environment. This is not surprising for chromosomally mediated MuL, but is less expected for plasmid-mediated MuH, where horizontal spread of the plasmid among genetically diverse strains is likely. The high prevalence of mupirocin-resistant staphylococci was due mainly to clonal dissemination and to a lesser extent to gene spread.
The resistance profiles of the isolates have shown that the overwhelming majority of these were resistant to methicillin. Linezolid, quinupristin/dalfopristin and vancomycin maintained high activity against essentially all mupirocin-resistant strains.
In the period 19992002 in Greece, a rising incidence of mupirocin-resistant CoNS has been observed. In contrast, mupirocin resistance in S. aureus has remained more constant. In our hospitals, the use of mupirocin is limited and it is only used for controlling the spread of MRSA. The low-rate of mupirocin-resistant S. aureus is due to the limited MRSA exposure to mupirocin and any subsequent development of resistance. On the other hand, the finding that mupirocin resistance is more common among S. epidermidis than S. aureus could be explained by the capacity of certain clones (i, a) to spread widely. Thus, the increased rate of mupirocin-resistant CoNS in Greece is related to the spread of methicillin-resistant epidemic hospital clones rather than the consumption of mupirocin. Measures to combat this spread, such as effective control of hospital clones, would appear to be prudent.
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Footnotes |
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These authors contributed equally to this work.
¶ Corresponding author. Fax: +30-41-0682508; E-mail: anm{at}otenet.gr
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References |
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