JAC Advance Access originally published online on November 14, 2005
Journal of Antimicrobial Chemotherapy 2006 57(1):146-149; doi:10.1093/jac/dki413
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
A 1 year surveillance study of glycopeptide-intermediate Staphylococcus aureus strains in a French hospital
1 Laboratoire de Bactériologie-Virologie-Hygiène, CHU Dupuytren, EA 3175, Limoges, France; 2 Department of Microbiology, University of Bristol, Bristol, UK; 3 AB BIODISK, Solna, Sweden
* Corresponding author. Tel: +33-555-05-63-48; Fax: +33-555-05-67-22; E-mail: fabien.garnier{at}unilim.fr
Received 20 April 2005; returned 15 June 2005; revised 7 October 2005; accepted 18 October 2005
 |
Abstract |
|---|
 Top Abstract IntroductionMaterials and methodsResultsDiscussionTransparency declarationsReferences |
|---|
Objectives: Glycopeptides are the drugs of choice to treat infections due to methicillin-resistant Staphylococcus aureus, but since 1995, glycopeptide-intermediate S. aureus (GISA) and heterogeneous GISA (hGISA) have been reported worldwide. Detection of reduced susceptibility to glycopeptides in S. aureus is very difficult in a routine clinical laboratory. The aim of this study was to investigate the prevalence of hGISA/GISA strains using a three-step approach during a 1 year period.
Methods: The following algorithm was adopted: (i) brain heart infusion agar with 4 mg/L teicoplanin was used to screen S. aureus strains for reduced susceptibility to glycopeptides; (ii) for each agar screen-positive strain, an Etest macromethod using modified cut-off values (vancomycin and teicoplanin 
4 mg/L) was used to detect potential hGISA/GISA; and (iii) the population analysis profile (PAP) method was finally used to confirm the hGISA/GISA phenotype.
Results: In total, 2300 strains of S. aureus were screened and 255 (11%) were categorized as hGISA with the PAP method, whereas no GISA strains were detected. Standard MIC values and current MIC breakpoints could not discriminate the hGISA/GISA phenotype from glycopeptide-susceptible S. aureus. Thus laboratories using currently standardized MIC methods cannot be expected to detect S. aureus strains that may exhibit reduced susceptibility to glycopeptides. Molecular typing by PFGE revealed that 238 strains belonged to the same clone.
Conclusions: A clonal hGISA strain has disseminated within our hospital. The method described in this study has to be further investigated to see if it is applicable to other S. aureus strains.
Keywords: glycopeptide resistance , S. aureus , Etest macromethod , population analysis profile method
|
Introduction |
|---|
|
TopAbstractIntroductionMaterials and methodsResultsDiscussionTransparency declarationsReferences |
|---|
Glycopeptides remain the drugs of choice to treat infections due to methicillin-resistant Staphylococcus aureus (MRSA). The first glycopeptide-intermediate S. aureus (GISA) strain was isolated in France in 1995.1 Since then, there have been further reports of GISA in Japan, USA and Europe.2,3 Two phenotypes of strains with reduced susceptibility are described: (i) GISA sensu stricto for which vancomycin and teicoplanin MICs are >4 mg/L such as S. aureus Mu50 and LIM-2 strains;1,2 and (ii) heterogeneous GISA (hGISA) for which vancomycin and teicoplanin MICs are
4 mg/L, but which contain a subpopulation that can grow in the presence of >4 mg/L vancomycin, such as S. aureus Mu3 strain.2 Reduced susceptibility to glycopeptides in S. aureus appears to be complex and difficult to detect in the clinical laboratories. The aim of this study was to investigate the prevalence of hGISA/GISA strains during a 1 year period in the teaching hospital where the GISA strain LIM-2 was characterized and to determine which detection method could be used for routine clinical testing. |
Materials and methods |
|---|
|
TopAbstractIntroductionMaterials and methodsResultsDiscussionTransparency declarationsReferences |
|---|
Bacterial isolates
From July 2001 to June 2002, all S. aureus strains isolated from clinical samples, at the Limoges Teaching Hospital, a 2000 bed hospital, were collected. Duplicate strains for each patient were excluded.
S. aureus ATCC 25923, Mu3 and S. aureus LIM-2 and Mu501,2 were used as control strains to represent various phenotypes, i.e. vancomycin-susceptible, hGISA and GISA.
Screening for glycopeptide resistance
All isolates were screened for reduced susceptibility to glycopeptides by inoculating 10 µL of a 2 McFarland suspension onto brain heart infusion agar (BHIA) (AES Laboratoire) supplemented with 4 mg/L teicoplanin (prepared in-house) and incubated at 35°C for 24 h.
Antimicrobial susceptibility testing
For each screen-positive strain, vancomycin and teicoplanin susceptibilities were determined using the Etest (AB Biodisk) macromethod with a 2 McFarland suspension. This macromethod was performed twice in two independent laboratories, using BHIA from AES Laboratoire at the Limoges hospital laboratory in France and from Becton Dickinson at the AB Biodisk research laboratory in Sweden. In this study, modified interpretive criteria different from the manufacturer's recommendations were used. All isolates with Etest macromethod vancomycin and teicoplanin values of 4 mg/L were considered as potential hGISA/GISA strains as previously described.4 MICs of vancomycin and teicoplanin were determined by the standardized agar dilution method using MHA (Bio-Rad Life Science group) and BHIA (AES Laboratoire) according to the Antibiogram Committee of the French Society of Microbiology recommendations.
Antimicrobial susceptibility testing was performed by the Vitek 2 system (bioMérieux).
Population analysis profile
For each strain with Etest vancomycin and teicoplanin values of 4 mg/L, the population analysis profile (PAP) method was performed at the Limoges hospital laboratory as described by Wootton et al.5 and the PAP–area under the curve (PAP–AUC) ratio method was performed at the Department of Microbiology at Southmead Hospital, Bristol, UK as previously described.3
Genotyping
All hGISA strains were typed by PFGE after DNA digestion by SmaI as previously described.4 Comparison of the fingerprinting was performed by the unweighted pair group method using programs of the Taxotron package (Taxolab, Institut Pasteur, Paris, France).
|
Results |
|---|
|
TopAbstractIntroductionMaterials and methodsResultsDiscussionTransparency declarationsReferences |
|---|
Screening test
From July 2001 to June 2002, 2300 strains were screened and 555 strains (24.1%) grew on BHIA with 4 mg/L teicoplanin and were suspected to exhibit reduced susceptibility to glycopeptides.
Antimicrobial susceptibility
For the 555 screen-positive strains, the Etest macromethod was first performed in our laboratory. In total, 294 strains of the 555 screen positives (53%) were considered as potential hGISA/GISA with the modified cut-off values (vancomycin and teicoplanin 4 mg/L) (Table 1). Retrospectively, these 294 strains were tested at the AB Biodisk research laboratory. The Etest macromethod values between the French and Swedish laboratories had a concordance of 97.6% within ±1 dilution (data not shown).
|
Among these 294 potential hGISA/GISA strains, 91 were isolated from pus, 77 from tracheal aspirates, 33 from superficial wounds, 31 from urines, 22 from sputum, 16 from blood cultures, 13 from catheters and 11 from deep wounds.
Agar dilution MIC values on MHA of vancomycin ranged from 1 to 8 mg/L and those of teicoplanin ranged from 1 to 24 mg/L, whereas on BHIA corresponding values were 2–12 mg/L and 2–24 mg/L, respectively (Table 1).
Population analysis profile
The 294 potential hGISA/GISA strains were further studied by the PAP method. Among these 294 strains no GISA phenotype was detected, whereas 255 (86.7%) were identified as hGISA and 39 as susceptible to glycopeptides (Table 1). Thus, the incidence of hGISA strains in our hospital was found to be 11% (255/2300).
To confirm the PAP data, the PAP–AUC method was performed at the Bristol laboratory and showed that 238 out of the 294 strains were defined as hGISA. Therefore, the detection of hGISA by the French and the English laboratories showed a good concordance of 93.3%.
Genotyping
Among the 255 hGISA, 225 strains (88.2%) harboured the pattern A (Table 1). Surprisingly, this profile was identical with the pattern of the first GISA isolate, LIM-2, described in our hospital in 1995 (Figure 1).1 Thirteen glycopeptide-susceptible strains also presented the same pulsotype (Table 1).
|
|
Discussion |
|---|
|
TopAbstractIntroductionMaterials and methodsResultsDiscussionTransparency declarationsReferences |
|---|
In this study we developed and used a three-step procedure to detect hGISA/GISA strains: (i) initial screening on BHIA with 4 mg/L teicoplanin; (ii) all agar screen positives were then subjected to the Etest macromethod with modified vancomycin and teicoplanin positive cut-offs of
4 mg/L; and (iii) Etest macromethod based hGISA/GISA suspects were finally analysed with the PAP method.
From July 2001 to June 2002, 2300 S. aureus strains were screened and 555 strains were suspected of potentially exhibiting reduced susceptibility to glycopeptides. After assessment with the Etest macromethod, 294 strains could be considered as potential hGISA/GISA. The PAP method confirmed that 255 out of these 294 strains were hGISA and no GISA phenotype was found. The rate of false-positives by the modified Etest macromethod was therefore 1.7% compared with the PAP analysis method, which is comparable to the results previously described.3 We knowingly chose lower Etest macromethod cut-off values to increase detection sensitivity in order to improve the capture of hGISA/GISA that may prevail in our clinical environment. If the manufacturer-recommended interpretive criteria for the Etest macromethod were used (vancomycin and teicoplanin 8 mg/L, or teicoplanin alone >12 mg/L), only 247 strains would have qualified as potential hGISA/GISA and with a false-positive rate of 0.3% (7/2300), however, 22 hGISA strains (according to PAP) would not have been detected, thus incurring a potential false-negative rate of 0.95%. With yet another Etest cut-off of vancomycin 4 and teicoplanin 8 mg/L used to analyse our data, 264 strains were defined as potential hGISA/GISA giving 0.8% false-positives and 0.4% false-negatives compared with the PAP method.
Among the 255 hGISA, seven strains (2.75%) were susceptible to methicillin as already described,6 which demonstrated that all S. aureus strains, and not only MRSA as is often recommended, should be screened.
In our study, the incidence of hGISA strains was 11% (255/2300). This is much higher than the prevalence previously reported in two studies (0.5 and 0.6%) where consecutive isolates of S. aureus were screened with the same testing algorithm used by us.6,7 However, higher prevalence rates have been previously described in a few studies involving a selected population of S. aureus or MRSA strains.7,8 The difference in the study populations of the S. aureus strains as well as clinical exposure to glycopeptide treatment and detection methods may account for the differences in the reported prevalence levels.
PFGE analysis showed that 225 out of the 255 hGISA strains belonged to the same pattern which is identical with the one of the first GISA isolate (LIM-2) in our hospital in 1995. Thus, the high incidence of hGISA strains in our hospital was more likely due to the spread of a predominant clone, as described in previous French studies.4,9,10 The committee for nosocomial infection surveillance and the hygiene unit were aware of the dissemination of this hGISA clone. Accordingly, different interventions were adopted: (i) a reinforcement of the infection control policies and practices for the prevention and the control of multidrug-resistant bacteria; (ii) an improvement of dosage regimens for glycopeptide use; and (iii) the use of alternative antibiotics, such as linezolid, to treat infections caused by hGISA strains.
While the three-step hGISA/GISA detection algorithm described in this study may appear to better detect and discriminate the aforementioned clonal types, it may not be appropriate to other strains until further investigated.
Moreover, this screening method is time consuming and the PAP method requires significant experience and is not easy to set up in a clinical laboratory. Therefore, this screening protocol may not be feasible for all strains in routine laboratories and could be reserved for patients with severe infections, and for strains isolated from sterile sites.
|
Transparency declarations |
|---|
|
TopAbstractIntroductionMaterials and methodsResultsDiscussionTransparency declarationsReferences |
|---|
No declarations were made by the authors of this paper.
|
Acknowledgements |
|---|
We thank I. Podglajen for her helpful advice. This work was supported by a grant from the Direction de la Recherche Clinique, CHU Dupuytren, Limoges, France.
|
References |
|---|
|
TopAbstractIntroductionMaterials and methodsResultsDiscussionTransparency declarationsReferences |
|---|
1. Ploy MC, Grelaud C, Martin C et al. First clinical isolate of vancomycin-intermediate Staphylococcus aureus in a French hospital. Lancet 1998; 340: 493–501.[CrossRef]
2. Hiramatsu K, Aritaka N, Hanaki H et al. Dissemination in Japanese hospitals of strains of Staphylococcus aureus heterogeneously resistant to vancomycin. Lancet 1997; 350: 1670–3.[CrossRef][Web of Science][Medline]
3.
Walsh TR, Bolmström A, Qwärnström A et al. Evaluation of current methods for detection of staphylococci with reduced susceptibility to glycopeptides. J Clin Microbiol 2001; 39: 2439–44.
4.
Guerin F, Buu-Hoï A, Mainardi JL et al. Outbreak of methicillin-resistant Staphylococcus aureus with reduced susceptibility to glycopeptides in a parisian hospital. J Clin Microbiol 2000; 38: 2985–8.
5.
Wootton M, Howe RA, Hillman R et al. A modified population analysis profile (PAP) method to detect hetero-resistance to vancomycin in Staphylococcus aureus in a UK hospital. J Antimicrob Chemother 2001; 47: 399–403.
6. Reverdy ME, Jarraud S, Bobin-Dubreux S et al. Incidence of Staphylococcus aureus with reduced susceptibility to glycopeptides in two French hospitals. Clin Microbiol Infect 2001; 7: 267–72.[CrossRef][Web of Science][Medline]
7.
Kim MN, Hwang SH, Pyo YJ et al. Clonal spread of Staphylococcus aureus heterogeneously resistant to vancomycin in a university hospital of Korea. J Clin Microbiol 2002; 40: 1376–80.
8.
Bert F, Clarissou J, Durand F et al. Prevalence, molecular epidemiology, and clinical significance of heterogeneous glycopeptide-intermediate Staphylococcus aureus in liver transplant. J Clin Microbiol 2003; 41: 5147–52.
9. Cartalano GL, Cheron M, Benabid D et al. Methicillin-resistant Staphylococcus aureus (MRSA) with reduced susceptibility to glycopeptides (GISA) in 63 French general hospitals. Clin Microbiol Infect 2004; 10: 448–51.[CrossRef][Web of Science][Medline]
10. Mallaval FO, Carricajo A, Delavenna F et al. Detection of an outbreak of methicillin-resistant Staphylococcus aureus with reduced susceptibility to glycopeptides in a French hospital. Clin Microbiol Infect 2004; 10: 459–70.[CrossRef][Web of Science][Medline]
CiteULike 
Connotea 
Del.icio.us What's this?
This article has been cited by other articles:
|
|
 |
|
  S. N. Leonard, C. Vidaillac, and M. J. Rybak Activity of Telavancin against Staphylococcus aureus Strains with Various Vancomycin Susceptibilities in an In Vitro Pharmacokinetic/Pharmacodynamic Model with Simulated Endocardial Vegetations Antimicrob. Agents Chemother., July 1, 2009; 53(7): 2928 - 2933. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 S. N. Leonard, K. L. Rossi, K. L. Newton, and M. J. Rybak Evaluation of the Etest GRD for the detection of Staphylococcus aureus with reduced susceptibility to glycopeptides J. Antimicrob. Chemother., March 1, 2009; 63(3): 489 - 492. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. N. Leonard and M. J. Rybak Evaluation of vancomycin and daptomycin against methicillin-resistant Staphylococcus aureus and heterogeneously vancomycin-intermediate S. aureus in an in vitro pharmacokinetic/pharmacodynamic model with simulated endocardial vegetations J. Antimicrob. Chemother., January 1, 2009; 63(1): 155 - 160. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
K. Kosowska-Shick, L. M. Ednie, P. McGhee, K. Smith, C. D. Todd, A. Wehler, and P. C. Appelbaum Incidence and Characteristics of Vancomycin Nonsusceptible Strains of Methicillin-Resistant Staphylococcus aureus at Hershey Medical Center Antimicrob. Agents Chemother., December 1, 2008; 52(12): 4510 - 4513. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 M. J. Rybak, S. N. Leonard, K. L. Rossi, C. M. Cheung, H. S. Sadar, and R. N. Jones Characterization of Vancomycin-Heteroresistant Staphylococcus aureus from the Metropolitan Area of Detroit, Michigan, over a 22-Year Period (1986 to 2007) J. Clin. Microbiol., September 1, 2008; 46(9): 2950 - 2954. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
Y. Maor, G. Rahav, N. Belausov, D. Ben-David, G. Smollan, and N. Keller Prevalence and Characteristics of Heteroresistant Vancomycin-Intermediate Staphylococcus aureus Bacteremia in a Tertiary Care Center J. Clin. Microbiol., May 1, 2007; 45(5): 1511 - 1514. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
K. Credito, G. Lin, and P. C. Appelbaum Activity of Daptomycin Alone and in Combination with Rifampin and Gentamicin against Staphylococcus aureus Assessed by Time-Kill Methodology Antimicrob. Agents Chemother., April 1, 2007; 51(4): 1504 - 1507. [Abstract] [Full Text] [PDF]
|
|
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||