Susceptibility to quinupristin/dalfopristin and other antibiotics of vancomycin-resistant enterococci from the UK, 1997 to mid-1999

Alan P. Johnson*, Marina Warner, Gill Hallas and David M. Livermore

Antibiotic Resistance Monitoring and Reference Laboratory, Central Public Health Laboratory, Colindale, London NW9 5HT, UK


    Abstract
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 References
 
Susceptibility to quinupristin/dalfopristin and other antibiotics was studied for clinical isolates of vancomycin-resistant enterococci (VRE) referred by UK hospitals between January 1997 and June 1999. Single isolates of VRE from 858 patients in 136 hospitals were received, of which 76% were Enterococcus faecium and 21% were Enterococcus faecalis, the remainder comprising minor species. Most isolates were multi-resistant. After allowing for the effect of blood, which raised the MICs of quinupristin/dalfopristin four-fold, 98.3% of E. faecalis isolates and all the Enterococcus avium, Enterococcus casseliflavus and Enterococcus gallinarum appeared resistant to quinupristin/dalfopristin, whereas 98.8% of the E. faecium isolates and the single Enterococcus raffinosus isolate were susceptible.


    Introduction
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 References
 
Enterococci are important opportunistic nosocomial pathogens and have intrinsic and acquired resistance to many antibiotics. During the past decade, a major interest has been the emergence and spread of enterococci resistant to vancomycin. Following their initial recognition in a south London hospital in 1987, vancomycin-resistant enterococci (VRE) had been isolated in at least 71 UK hospitals by 1995.1 They are an even greater problem in the USA, and have been isolated in many other countries.2 Although 85–90% of clinical isolates of enterococci are Enterococcus faecalis, most VRE are Enterococcus faecium.2

The streptogramin quinupristin/dalfopristin (Synercid) was recently licensed in the UK and the USA, with treatment of infections caused by glycopeptide-resistant E. faecium as an indication. The PHLS Antibiotic Resistance Monitoring and Reference Laboratory (ARMRL) regularly receives isolates of VRE from many hospitals throughout the UK. We report on the activity of quinupristin/dalfopristin and other agents against clinical isolates of VRE referred between January 1997 and June 1999, immediately before the drug's launch.


    Materials and methods
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 References
 
Susceptibility testing

For the past decade, Gram-positive bacteria (comprising predominantly pneumococci, enterococci, viridans streptococci and staphylococci) submitted to the ARMRL have been tested for resistance to ampicillin, penicillin, erythromycin, gentamicin, vancomycin, teicoplanin, rifampicin, tetracycline, ciprofloxacin and chloramphenicol. Quinupristin/dalfopristin was added to this panel in January 1997. MICs were determined on Diagnostic Sensitivity Test (DST) agar (Oxoid, Basingstoke, UK) containing 5% lysed horse blood (TCS Microbiology, Buckingham, UK) to promote the growth of fastidious species. Inocula comprise 104–105 cfu/spot, and plates are incubated for 18 h at 37°C in air. Originally, the MICs were recorded by eye, but since May 1997 they have been read with a Domino image analyser (Perceptive Instruments, Haverhill, UK). Before analysis, records for isolates from non-human sources (e.g. those from food, animals or the environment) were deleted, as were those for repeat isolates from individual patients. Isolates were categorized as susceptible or resistant using breakpoints recommended by the British Society for Antimicrobial Chemotherapy (BSAC)3,4 as follows: ampicillin, 8 mg/L; chloramphenicol, 8 mg/L; ciprofloxacin, 1 mg/L; erythromycin, 0.5 mg/L; gentamicin (high-level), 500 mg/L; rifampicin, 1 mg/L; teicoplanin, 4 mg/L; tetracycline, 1 mg/L; vancomycin, 4 mg/L.


    Results
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 References
 
Referred isolates of VRE

Over the 2.5 year study period, isolates of VRE from 858 different patients in 136 hospitals were referred to the ARMRL. Speciation of the VRE by PCR,5 undertaken in the Epidemiological Typing Unit of the Laboratory of Hospital Infection at Colindale, showed that 650 (76%) were E. faecium, 179 (21%) E. faecalis, and the remainder comprised Enterococcus avium (n = 3), Enterococcus casseliflavus (n = 7), Enterococcus gallinarum (n = 18) and Enterococcus raffinosus (n = 1). The number of hospitals referring isolates of vancomycin-resistant E. faecium increased from 52 in 1997 to 76 in 1998, with 43 referring isolates in the first 6 months of 1999. Corresponding figures for E. faecalis were 28, 41 and 15. Many of the referred E. faecium and E. faecalis isolates were resistant to multiple antibiotics (Table IGo). Approximately 83% of vancomycin-resistant isolates of both these species were resistant to teicoplanin, over 96% to erythromycin, 52% to high levels of gentamicin and 99% to ciprofloxacin. Approximately 99% of E. faecalis isolates, but only 37.5% of E. faecium, were resistant to tetracycline. All E. faecalis isolates, but none of E. faecium, were susceptible to ampicillin. All 29 isolates of the four minor species were sensitive to ampicillin (MICs 0.5–8 mg/L), with 32% also sensitive to erythromycin; four (14%) showed high-level resistance to gentamicin. The MICs of vancomycin for E. casseliflavus and E. gallinarum isolates were 8–16 mg/L, but these species were sensitive to teicoplanin (MICs 0.5–2 mg/L), as is typical of the VanC phenotype.2 The three isolates of E. avium and the one E. raffinosus were resistant to both vancomycin and teicoplanin (MICs > 32 mg/L).


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Table I. Antimicrobial susceptibility of isolates of vancomycin-resistant E. faecium and E. faecalis referred to Antibiotic Resistance Monitoring and Reference Laboratory January 1997 to June 1999
 
Activity of quinupristin/dalfopristin

MIC distributions of quinupristin/dalfopristin on DST agar containing 5% lysed horse blood are shown in Table IIGo. Of the 650 E. faecium isolates, 77.8% were susceptible to quinupristin/dalfopristin at <=2 mg/L, and the MIC90 was 4 mg/L. In contrast, MICs for 98.3% of the isolates of E. faecalis were >=8 mg/L, as were those for 14 of 18 E. gallinarum isolates. The MICs for the E. faecium isolates appeared higher than those reported elsewhere6,7 and, as several groups have shown that quinupristin/dalfopristin MICs are influenced by blood,79 50 selected E. faecium isolates were re-tested with quinupristin/dalfopristin on DST with and without lysed blood. As shown in Table IIGo, MICs without blood were about four-fold lower than those obtained with 5% blood.


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Table II. Activity of quinupristin/dalfopristin against vancomycin-resistant enterococci
 

    Discussion
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 References
 
Quinupristin/dalfopristin was recently licensed in the UK, inter alia, for treatment of infections due to vancomycin-resistant E. faecium. We outline here its activity against a large collection of clinical VRE isolates (predominantly E. faecium, many multiply resistant), referred to the ARMRL from a sizeable number of UK hospitals, in the 2.5 years immediately before its launch.

As noted by others,6,7 quinupristin/dalfopristin was more active against E. faecium than against E. faecalis. However, determination of the proportion of isolates resistant to quinupristin/dalfopristin posed some difficulty. When this study was initiated, there was no defined breakpoint for quinupristin/dalfopristin. In 1998, the BSAC advocated a breakpoint of 2 mg/L for enterococci,4 and on this basis, only 77.8% of the referred isolates of E. faecium would be regarded as susceptible. However, the BSAC guidelines note that the in vitro activity of quinupristin/dalfopristin is reduced by blood, as reported also by others.79 This effect was confirmed here, the MICs of quinupristin/dalfopristin on blood-free DST agar being about four-fold lower than on DST agar with 5% lysed blood. The BSAC guidelines advocate that the disc zone diameter breakpoint of >=20 mm for quinupristin/dalfopristin should be reduced to >=15 mm if the medium contains blood. If the same broad principle is applied here, and the breakpoint raised four-fold to 8 mg/L to compensate for the effect of the blood, about 98.8% of the vancomycin-resistant E. faecium isolates would be regarded as sensitive. On this basis we confirm that quinupristin/dalfopristin is highly active against most vancomycin-resistant E. faecium from UK hospitals. Nevertheless, the occurrence of a few resistant E. faecium isolates gives some cause for concern. Resistance has already been noted in clinical E. faecium isolates and those from foodstuffs,10 with the latter perhaps reflecting the use of a related streptogramin (virginiamycin) as a growth promoter. Resistance may become more prevalent under the selective pressure of increasing quinupristin/dalfopristin usage; monitoring of this situation will require the inclusion of quinupristin/dalfopristin in resistance surveillance programmes.


    Acknowledgments
 
We thank Terri Parsons for technical assistance and our colleagues in the Epidemiological Typing Unit, Central Public Health Laboratory, for confirming the identification of the isolates tested.


    Notes
 
* Corresponding author. Tel: +44-181-200-4400, ext. 4237; Fax: +44-181-200-7449; E-mail: ajohnson{at}phls.nhs.uk Back


    References
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 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 References
 
1 . Public Health Laboratory Service. (1996). Vancomycin resistant enterococci in hospitals in the United Kingdom—1995. Communicable Disease Report Weekly 6, 267.

2 . Woodford, N., Johnson, A. P., Morrison, D. & Speller, D. C. E. (1995). Current perspectives on glycopeptide resistance. Clinical Microbiology Reviews 8, 585–615.[Abstract]

3 . Phillips, I., Andrews, J. M., Bridson, E., Cooke, E. M., Holt, H. A., Spencer, R. C. et al. (1991). A guide to sensitivity testing. Report of the working party on antibiotic sensitivity testing of the British Society for Antimicrobial Chemotherapy. Journal of Antimicrobial Chemotherapy 27, Suppl. D, 5–6.

4 . Working Party on Sensitivity Testing of the British Society for Antimicrobial Chemotherapy. (1998). BSAC standardized disc sensitivity testing method. Newsletter of the British Society for Antimicrobial Chemotherapy, Summer 1998.

5 . Dutka-Malen, S., Evers, S. & Courvalin, P. (1995). Detection of glycopeptide resistance genotypes and identification to the species level of clinically relevant enterococci by PCR. Journal of Clinical Microbiology 33, 24–7.[Abstract]

6 . Jones, R. N., Ballow, C. H., Biedenbach, D. J., Deinhart, J. A. & Schentag, J. J. (1998). Antimicrobial activity of quinupristin/dalfopristin (RP 59500, Synercid) tested against over 28,000 recent clinical isolates from 200 medical centers in the United States and Canada. Diagnostic Microbiology and Infectious Disease 31, 437–51.[ISI][Medline]

7 . King, A., May, J. & Phillips, I. (1998). Comparative activity of quinupristin/dalfopristin and RPR 106972 and the effect of medium on in-vitro test results. Journal of Antimicrobial Chemotherapy 42, 711–9.[Abstract]

8 . Jones, R. N. (1996). Medium and supplement effects on the antimicrobial activity of quinupristin/dalfopristin tested by agar dilution and Etest methods. Diagnostic Microbiology and Infectious Disease 26, 99–102.[ISI][Medline]

9 . Andrews, J. M., Ashby, J. P. & Wise, R. (1999). Study to assess the reliability of a disc diffusion method for determining the sensitivity of Gram-positive pathogens to dalfopristin/quinupristin. Journal of Antimicrobial Chemotherapy 43, 141–3.[Abstract/Free Full Text]

10 . Woodford, N., Palepou, M.-F., Johnson, A. P., Chadwick, P. R. & Bates, J. (1997). Methicillin-resistant Staphylococcus aureus and vancomycin-resistant enterococci. Lancet 350, 738.[ISI][Medline]

Received 16 November 1999; returned 23 January 2000; revised 4 February 2000; accepted 25 February 2000