Varying linezolid susceptibility of vancomycin-resistant Enterococcus faecium isolates during therapy: a case report
Stefanie Swoboda1,*,
Stefan Fritz2,
Marcus E. Martignoni2,
Rita A. Feldhues3,
Torsten Hoppe-Tichy1,
Markus W. Buchler2 and
Heinrich K. Geiss3
1 Pharmacy Department, University of Heidelberg, Im Neuenheimer Feld 670, D-69120 Heidelberg, Germany; 2 Department of Surgery, University of Heidelberg, Im Neuenheimer Feld 120, D-69120 Heidelberg, Germany; 3 Institute of Hygiene, University of Heidelberg, Im Neuenheimer Feld 670, D-69120 Heidelberg, Germany
* Corresponding author. Tel: +49-6221-56-32320; Fax: +49-6221-56-5343; E-mail: stefanie.swoboda{at}med.uni-heidelberg.de
Received 6 May 2005; returned 29 June 2005; revised 3 August 2005; accepted 12 August 2005
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Abstract
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Objectives: Linezolid is an oxazolidinone antibiotic used in the treatment of infections caused by vancomycin-resistant enterococci. Resistance to linezolid has been associated with a G2576U mutation in domain V of the 23S rRNA.
Patient and methods: We present clinical details and susceptibility data from multiple Enterococcus faecium strains isolated from a liver transplant patient over 13 months. MICs of linezolid, vancomycin and quinupristin/dalfopristin were determined using Etest. Molecular typing was performed by pulsed-field gel electrophoresis. Domain V of the 23S rRNA gene in the vancomycin-resistant Enterococcus faecium was amplified. Linezolid concentrations were analysed by HPLC.
Results: We report the emergence of resistance to linezolid in a vancomycin-resistant Enterococcus faecium during linezolid treatment. After discontinuation of the linezolid therapy, the isolate reverted to susceptibility. However, after re-administration of linezolid the vancomycin-resistant Enterococcus faecium became resistant to linezolid again. The isolates that were resistant to linezolid had a G2576T mutation in their 23S rDNA.
Conclusion: We describe a clinical case that shows the shift of a vancomycin-resistant Enterococcus faecium from linezolid resistance to susceptibility and then back to resistance again related to linezolid therapy.
Keywords:
resistance
,
pharmacokinetics
,
pharmacodynamics
,
liver transplantations
,
mutations
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Introduction
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Linezolid is an important therapeutic option for the treatment of infections caused by multiresistant Gram-positive bacteria. Linezolid is active against vancomycin-resistant enterococci, methicillin-resistant Staphylococcus aureus (MRSA) and glycopeptide-intermediate Staphylococcus aureus (GISA).1 However, the clinical benefit of linezolid is threatened by the emergence of resistant strains of MRSA and vancomycin-resistant enterococci reported in North America and the UK.26 To date, a clinical isolate of a linezolid-resistant vancomycin-resistant Enterococcus faecium (VRE) has not yet been reported in Germany. We describe the case of a liver transplant patient suffering from vanishing bile duct syndrome, who developed chronic subhepatic abscesses. From those abscesses, we isolated E. faecium which became vancomycin-resistant. During prolonged therapy with linezolid, this enterococcal strain became linezolid-resistant. After discontinuation of linezolid, this strain became susceptible to linezolid, but the linezolid MIC rose again (>16 mg/L) when treatment resumed.
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Patient and methods
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Identification and antimicrobial susceptibility testing was performed by the Phoenix system (BD Diagnostic Systems, Heidelberg, Germany). Additionally, MIC values for vancomycin, linezolid and quinupristin/dalfopristin were confirmed using Etest (AB Biodisk, Solna, Sweden). MIC of linezolid was 2 mg/L for linezolid-susceptible and >16 mg/L for linezolid-resistant isolates. Molecular characterization for clonal identity of VRE isolates was performed by means of pulsed-field gel electrophoresis (PFGE) of SmaI-macrorestricted genomic DNA, and restriction-fragment-length polymorphism analysis of PCR-amplified intragenic fragments of the VRE gene.
Linezolid concentrations were determined in serum and abscess fluid samples by a modified HPLC analytical method with UV absorbance detection (
max 251 nm) as described by Peng et al.7 and Tobin et al.8 Briefly, samples were prepared by protein precipitation with acetonitrile. After centrifugation (10°C, 7826 g, 10 min), 100 µL supernatant was injected into the analytical column (phenomenex Luna C8). The mobile phase consisted of water and acetonitrile (8:2 v/v). The analysis was performed at room temperature, at a flow rate of 1.4 mL/min.
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Results
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A 42-year-old man underwent liver transplantation in June 2003 because of primary sclerosing cholangitis with liver cirrhosis. Immunosuppressive therapy was performed with tacrolimus and methylprednisolone. Vanishing bile duct syndrome led to malfunctioning of the liver graft and the patient had to be re-transplanted in December 2003. In February 2004, he suffered from fever caused by a liver abscess followed by severe septic complications. After radiology-guided interventional abscess drainage and long-term antibiotic therapy, the patient recovered slowly. Since the cholestatic parameters were raised during May 2004, a percutaneous transhepatic cholangiodrainage was performed. In the following 4 months, recurrent liver abscesses caused by different bacterial species (ESBL-positive Escherichia coli and Klebsiella oxytoca, Proteus spp., Enterobacter spp., Stenotrophomonas maltophilia, E. faecium and different Candida spp.) necessitated multiple antibiotic treatment courses in varying combinations including meropenem, piperacillin/tazobactam, amoxicillin/clavulanic acid), ceftriaxone, ciprofloxacin, vancomycin (two treatment courses of 11 and 14 days), and linezolid (four therapy cycles of 1014 days each). The steady worsening of the liver function led to another transplantation in August 2004, when for the first time a VRE (VanA-positive) was isolated from an intra-operative swab (ascites) and the catheter. Linezolid treatment was given thereafter for 21 days (VRE was sensitive to linezolid). Five weeks after the first VRE was isolated, an indistinguishable VRE strain was isolated from an aspirate of the subhepatic abscess which showed an MIC of >16 mg/L for linezolid. Sequencing the PCR product of domain V in the 23S rRNA using the primers 5'-GACGGAAAGACCCCATGG-3' and 5'-ACACTTAGATGCTTT-3' corresponding to bp 2049 to 2767 showed the previously described G2576T mutation.3 Treatment was changed to quinupristin/dalfopristin for 6 weeks. The clinical situation improved during this therapy and the patient was discharged to home in November 2004. However, all subhepatic drainage samples were still positive for ESBL-positive E. coli and VRE. In January 2005, the patient was readmitted because he suffered another septic episode originating from sub- and intra-hepatic abscesses. The VRE isolate now exhibited an MIC value against linezolid of 2 mg/L. The patient's condition worsened in February 2005 leading to renewed linezolid therapy for 19 days. Molecular analysis was negative for the G2576T mutation. On the fourth day of linezolid treatment (600 mg twice a day intravenously), the linezolid concentration in blood 2 h after the start of infusion was 32 mg/L and 5 mg/L in abscess fluid 5 h after the start of infusion. At the end of this treatment cycle, the VRE strain was linezolid resistant. This strain was again positive for the G2576T mutation. The patient still suffered from multiple liver abscesses but due to the patient's unstable condition surgical re-intervention was impossible. At the end of February 2005, antibiotic therapy was finally stopped and the patient died within weeks.
Table 1 summarizes the relevant data on the different E. faecium isolates, their origins and the corresponding MICs of vancomycin, linezolid and quinupristin/dalfopristin. Four linezolid-susceptible and three linezolid-resistant isolates had identical susceptibility profiles and were indistinguishable by PFGE.
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Discussion
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Several factors contributing to antibiotic resistance have been described of which prolonged exposure to antibiotics is the most striking. Although new substances promising a low propensity to develop resistance are frequently introduced in medical practice, clinical reality differs. Several authors have published on the development of resistance to linezolid in MRSA and VRE.2,3,9 To our knowledge, however, we describe here for the first time the changing susceptibility in a clinical VRE isolate during treatment. Owing to a hospital-wide surveillance system of patients harbouring VRE, MRSA and ESBL-positive Enterobacteriaceae, we were able to retrieve almost all isolates from our patient and to check for clonal identity of the enterococcal strains. Additionally, this case is unique in that our patient had to be treated several times with linezolid because the chronic sub- and intrahepatic abscesses persisted as the result of vanishing bile duct syndrome, a rare condition after liver transplantation.
Treatment failure was related to the low ratio of concentration of linezolid in the abscess fluid to the MIC, a PK/PD parameter which is known to be relevant in oxazolidinones.10 The changing susceptibility pattern of the VRE may be due to the MIC values being correlated with the number of mutated 23S rRNA alleles in resistant strains.10,11 The discontinuation of linezolid therapy most probably led to an incomplete back mutation. Interestingly, isolates with an MIC value of 4 mg/L still showed a single mutated allele exhibiting the G2576U transversion, which may explain the rapid re-emergence of MIC values of 16 mg/L and higher after re-treatment with linezolid.
This case demonstrates that MIC determination for linezolid in strains which previously were categorized as resistant against this substance may lead to false-susceptible results. Possibly molecular characterization should be performed as a predictor of clinical failure in these cases.
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Transparency declarations
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There were no conflicts of interest.
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Acknowledgements
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We thank Magdalena Geiss, Institute for Hygiene, University of Heidelberg, for the molecular technical support with PFGE and sequence analysis.
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References
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