Journal of Antimicrobial Chemotherapy (2003) 51, 289–296
DOI: 10.1093/jac/dkg076
Advance Access publication 6 January 2003
Evaluation of selective and enrichment media for isolation of
glycopeptide-resistant enterococci from faecal specimens
Derek F. J. Brown* and Enid Walpole
Clinical Microbiology and Public Health Laboratory, Addenbrooke’s Hospital, Cambridge CB2 2QW, UK
Received 11 April 2002; returned 27 August 2002; revised 22 October 2002; accepted 4 November 2002
Objectives: Vancomycin-resistant enterococcus enrichment broth (VEB) and vancomycinresistant enterococcus selective agar with vancomycin 6 mg/L (VSA) are novel azide–aesculin
agar-based media that contain meropenem as an additional selective agent. The media were
compared with enterococcosel broth (EB) and enterococcosel agar with vancomycin 6 mg/L
(EA) for the isolation of glycopeptide-resistant enterococci (GRE) from routine faecal screening
specimens.
Methods: Two hundred and eighteen routine faecal screening specimens from patients at
Addenbrooke’s Hospital were examined. The majority were from patients on haematology wards
(155) or the intensive therapy unit (ITU) (21). Specimens were inoculated on to VSA and EA
directly, and after enrichment in VEB and EB, respectively.
Results: One hundred and twenty-eight GRE isolates were recovered from 93 (43%) specimens
with enterococci carrying vanA or vanB genes. There were no statistically significant differences between media (specimens positive; numbers of GRE isolates) on direct plating on VSA
(87; 104) or EA (86; 97) or following 24 h enrichment in VEB (89; 103) or EB (86; 98). There was no
significant advantage to enrichment compared with direct plating. Incubation of enrichment
broth cultures for only 6 h appeared detrimental. Enterococci with vanC were isolated
significantly less frequently from VEB and VSA than from EB and EA. Growth of organisms other
than GRE was more common on VSA than on EA.
Conclusions: VEB and VSA were at least as effective as EB and EA for the recovery of GRE
from faecal screening specimens, but substantially more non-GRE grew on VSA than on EA.
Enrichment culture offered no significant advantages over direct plating.
Introduction
Over the past 10 years there has been a rapid increase in
the incidence of infection and colonization of patients with
glycopeptide-resistant enterococci (GRE).1 Enterococci are
intrinsically resistant to many antimicrobial agents and the
emergence of strains resistant to commonly used agents,
including ampicillin, aminoglycosides and vancomycin, has
serious implications for immunosuppressed and debilitated
patients, who are at particular risk of infection. Enterococci
are found in the gastrointestinal tract, and many infections are
attributed to endogenous sources, although cross-infection
with GRE has been shown to occur.2 Colonization rates may
be high on intensive therapy unit (ITU),3,4 renal,5 oncology6–8
and transplant9 wards. Some patients carry multiple strains of
GRE10 and indistinguishable strains are isolated from different patients, suggesting cross-infection between patients
on the same wards.11,12
Faecal colonization has been found to precede infection13–16
and colonized patients may provide a reservoir for GRE, which
may cross-colonize and cause infection in other patients.
Once GRE have been isolated in a hospital, routine surveillance cultures of high-risk patients at weekly intervals have
been recommended to allow early identification of colonized
patients,13,16 and screening specimens should be cultured to
identify colonized patients during outbreaks of infection with
GRE.2,13
..................................................................................................................................................................................................................................................................
*Corresponding author. Tel: +44-1223-257020; Fax: +44-1223-242775; E-mail: [email protected]
...................................................................................................................................................................................................................................................................
289
© 2003 The British Society for Antimicrobial Chemotherapy
D. F. J. Brown and E. Walpole
Selective media containing vancomycin have been recommended for surveillance cultures for GRE.13 However, a range
of different media has been used, including 10% horse blood
agar with aztreonam and amphotericin,17 colistin nalidixic
acid agar,18 Mueller–Hinton agar with polymyxin and streptomycin,4 kanamycin aesculin azide agar,19 5% horse blood agar
with neomycin,20,21 bile aesculin azide agar,22,23 campylobacter blood agar with clindamycin,19 cefalexin aztreonam
arabinose agar24 and colistin nalidixic acid aesculin azide
agar.25 There is no consensus on which medium should be
used or the concentration of vancomycin to be included in the
medium. Concentrations of vancomycin from 4 to 64 mg/L
have been used, but 4 or 6 mg/L has been recommended to
facilitate the detection of low-level vancomycin resistance.26,27
Enrichment in broth followed by plating on vancomycinselective agar has been found to be more sensitive than
direct screening on selective agar media containing vancomycin;28 however, enrichment in broth has the disadvantage
that the procedure requires an extra day for incubation of the
broth.
PCR methods of screening for GRE in faeces have the
advantage that results can be available within a few hours of
receipt of the sample.29,30 However, extraction of DNA is
necessary to remove inhibitory substances in the sample, and
the method may not be cost-effective when dealing with large
numbers of specimens or when the prevalence is low.29 In
these situations, the use of PCR methods on enrichment broths
or colonies on selective medium may be appropriate.29,30 If
typing of isolates for epidemiological purposes is required,
culture will also be necessary.
There are no widely accepted standards for evaluation of
selective media for use in screening for GRE. However,
enterococcosel agar (EA) and broth (EB), which are bileaesculin-azide formulations, have been used in several studies.28,31,32 Oxoid vancomycin-resistant enterococcus enrichment broth (VEB) and selective agar (VSA) are novel in that
they are aesculin-azide media with meropenem as an additional selective agent. The objectives of this study were to
assess the performance of VEB and VSA in comparison with
EB and EA for the isolation of vancomycin-resistant enterococci (VRE) from faecal screening specimens. In addition, 6 h
enrichment was investigated to determine whether the extra
day required for broth enrichment could be avoided without
loss of sensitivity.
Materials and methods
Specimens
Two hundred and eighteen routine faecal screening specimens
from patients at Addenbrooke’s Hospital were included. The
majority were from patients on haematology wards (155) or
the ITU (21).
Media
The following media were used in screening for GRE. VEB
was VRE broth (Oxoid, Basingstoke, UK) containing meropenem 2 mg/L (meropenem supplement; Oxoid). VSA was
VSA agar (Oxoid) containing meropenem 1 mg/L and vancomycin 6 mg/L (meropenem and vancomycin supplements;
Oxoid). The antibiotic supplements in VEB and VSA were as
defined by the manufacturer. EB (Becton Dickinson, Oxford,
UK) was unsupplemented and EA (Becton Dickinson) was
supplemented with vancomycin 6 mg/L (Sigma, Poole, UK).
Screening methods
Specimens were inoculated directly on to VSA and EA media
by transferring faeces on a sterile swab and spreading with
a loop. Pea-sized portions of faeces were also transferred to
10 mL volumes of VEB and EB, vortexed and incubated at
37°C. After 6 and 24 h incubation the VEB was subcultured to
VRE selective agar. After 24 h incubation the EB was subcultured to EA. All plates were incubated in air at 37°C and
examined for colonies after 24 and 48 h. On both agar media
enterococci produce brown colonies with black/brown zones
around them. Different colony types, including colonies not
typical of enterococci, were subcultured to blood agar plates
from each selective plate. At least four separate colonies were
cultured to increase the chances of detecting mixtures of
organisms. Subcultures were incubated in air at 37°C for 24 h.
Isolates were tentatively identified as enterococci on the basis
of colony appearance, Gram’s stain and a positive test for
pyrrolidonylarylamidase activity. Identity and van gene
status were confirmed by a PCR method33,34 and antimicrobial
susceptibility was tested by disc diffusion.35 Differences in
the performance of media were tested statistically using the
χ2 method.
Results
Isolation of GRE with vanA or vanB
From the 218 specimens, GRE with vanA or vanB genotypes
were isolated from 93 (43%) (Table 1). Individual isolation
protocols detected GRE in between 86 (direct plating on EA,
enrichment for 6 h in VEB followed by plating on VSA, and
enrichment in EB followed by plating on EA) and 89 specimens (enrichment for 24 h in VEB followed by plating on
VSA) but there were no statistically significant differences
among the treatments.
More than one GRE with vanA or vanB genotypes was
isolated from several specimens and in total 128 isolates with
different identification or van gene status were recovered
(Table 1). Variation in susceptibility to agents other than vancomycin and teicoplanin was common with isolates from the
same specimen (data not shown). However, for the purposes
290
Isolation of glycopeptide-resistant enterococci
Table 1. Isolation of glycopeptide-resistant enterococci with vanA and vanB resistance from 218 faecal specimens
Culture method
Direct plating on VRE selective agar
Direct plating on enterococcosel agar
with vancomycin
6 h enrichment in VRE enrichment broth
and plating on VRE selective agar
24 h enrichment in VRE enrichment broth
and plating on VRE selective agar
24 h enrichment in enterococcosel broth
and plating on enterococcosel agar with
vancomycin
Total
aOne
Number of isolates
Number
of specimens
positive for
GRE
E. faecium
vanA
E. faecium
vanB
E. faecalis
vanA
E. gallinarum
vanA
total
87
86
37
37
54
46
12a
13
1
1
104
97
86
35
51
11a
1
98
89
40
52
10a
1
103
86
34
47
15
2
98
93
43
59
25
2
128
isolate gave colonies not typical of enterococci.
of this analysis, isolates were considered distinct if they were
different species and/or had different van gene types. Enterococcus faecium (102 isolates) was isolated more frequently
than Enterococcus faecalis (25 isolates). The vanB genotype
(59 isolates) was more common than vanA (43 isolates) in
E. faecium, but no isolates of E. faecalis with vanB resistance
were detected. There were two isolates of Enterococcus gallinarum with vanA, one being isolated on all media and one
only on EA following enrichment. Individual isolation protocols detected between 97 (direct plating on EA) and 104
(direct plating on VSE) GRE, but there were no statistically
significant differences among the treatments.
Enrichment in broth did not significantly increase the
isolation rates with either VEB or EB. It was notable that the
density of growth obtained on plates after 6 h incubation in
VEB was frequently less than that obtained on direct plating
or after 24 h enrichment. Where more than one GRE was
isolated, the predominant strain was not uncommonly different on direct plating and after enrichment. The majority of
isolates of GRE grew in large numbers on all media.
On all media, almost all GRE were detected after incubation of plates for 24 h. However, the colonies of GRE were
generally larger on VSA than on EA, in that, after incubation
for 24 h, colonies on VSA agar were generally at least 0.5 mm
in diameter, whereas colonies on EA were usually <0.5 mm in
diameter. On both the media, the brown/black colour greatly
assisted in indicating GRE colonies, but, particularly after
incubation of plates for 24 h, colonies on the same plate could
differ in appearance from very pale brown to black. One isolate of E. faecalis on VSA inoculated directly and following
enrichment for 6 or 24 h did not give colonies typical of
enterococci, in that they did not show any brown or black
pigment and would not have been subcultured as VRE in the
routine situation.
Isolation of vanC enterococci
E. gallinarum and Enterococcus casseliflavus with vanC
resistance were detected in eight and nine specimens,
respectively (Table 2). They were detected significantly less
frequently (P < 0.05) on VSA following enrichment than on
EA following enrichment, and significantly less frequently
(P < 0.05) on direct plating on VSA than on EA following
enrichment.
Isolation of vancomycin-susceptible enterococci
Vancomycin-susceptible enterococci, mostly E. faecalis,
were isolated from 36 specimens (Table 2). Differences
between media were not statistically significant (P > 0.05).
One isolate of E. faecalis on Oxoid VRE agar inoculated
following enrichment for 6 h and on enterococcosel agar
inoculated directly did not give colonies typical of enterococci, in that they did not show any brown or black pigment.
Isolation of organisms other than enterococci
Isolates that were not enterococci were not fully identified,
but were grouped on the basis of colony morphology and
Gram’s stain (Table 3). They were also grouped on the basis
of whether the colonies on the selective media appeared to be
similar to those of enterococci or clearly different.
Organisms other than GRE mostly grew with smaller numbers of colonies than VRE and were frequently not detected
until plates had been incubated for 48 h. Gram-positive bacilli
291
D. F. J. Brown and E. Walpole
Table 2. Positive isolates of enterococci other than vanA and vanB from 218 faecal specimens
Number of isolates
Culture method
Direct plating on VRE selective agar
Direct plating on enterococcosel agar
with vancomycin
6 h enrichment in VRE enrichment broth
and plating on VRE selective agar
24 h enrichment in VRE enrichment broth
and plating on VRE selective agar
24 h enrichment in enterococcosel broth
and plating on enterococcosel agar with
vancomycin
Total isolates
aOne
E. faecium
vancomycin
susceptible
E. faecalis
vancomycin
susceptible
E. gallinarum
E. casseliflavus
1
7
1
2
0
6a
3
6
3
4a
0
0
2
10
0
0
0
8
8
7
6
30
8
9
isolate gave colonies not typical of enterococci.
grew significantly more frequently following direct plating
on VSA than direct plating on EA (P < 0.01), enrichment in
VEB for 6 or 24 h followed by plating on VSA (P < 0.05),
direct plating on EA (P < 0.01) or enrichment in EB followed
by plating on EA (P < 0.001). Gram-positive bacilli grew
significantly more frequently on VSA following enrichment
in VEB than on EA following enrichment in EB (P < 0.05).
Colonies of Gram-positive bacilli were frequently, but not
always, smaller than GRE colonies, but they very frequently
had the characteristic appearance of enterococci. A few
staphylococci grew on the VSA in particular (Table 3). In
appearance of the colonies, about half were indistinguishable
from GRE. A few other Gram-positive cocci, which were not
enterococci, grew on both media. In appearance, they were
mostly indistinguishable from GRE. With Gram-positive
cocci other than enterococci, differences between media were
not statistically significant (P > 0.05). Yeasts were isolated
significantly more frequently on VSA, without enrichment
(P < 0.01), following enrichment for 6 h (P < 0.01) or enrichment for 24 h (P < 0.001), than on EA with or without
enrichment. However, colonies were usually atypical of
enterococci.
Discussion
Most of the specimens examined in this study were from
patients on haematology wards and the ITU. Patients on these
wards are known to have high GRE colonization rates, and the
high overall positivity rate (43%) is in line with that of previous publications.3–9 Multiple isolates of GRE from the same
specimen were not uncommon, in that 128 isolates of entero-
cocci with vanA or vanB were found in the 93 positive
specimens. Carriage of multiple strains of GRE has been
reported previously,10,36 but different colony morphologies
were generally not obvious on selective plates with either
medium in this study. Multiple strains were commonly
detected because different strains grew on different media, or
different strains grew on direct plating and enrichment. Some
were detected through routine selection of four colonies for
identification and susceptibility testing. In routine screening
for GRE in faecal specimens, multiple screening procedures
are unlikely to be used, and although this might be ameliorated to some extent by picking multiple colonies, some falsenegative reports appear to be inevitable.
There were more isolates of GRE (vanA and vanB) on
VSA, both with and without enrichment, than on EA with and
without enrichment, but differences were not statistically
significant. Enterococci with vanC include E. gallinarum and
E. casseliflavus, which were isolated less frequently from
VSA than from EA, particularly following enrichment in
VSB, which eliminated all enterococci with vanC except one
strain also carrying the vanA gene. As the concentrations of
vancomycin in VSA and EA were the same, the difference in
isolation rates of E. gallinarum and E. casseliflavus, which
are intrinsically low-level resistant to vancomycin, must be
related to other differences between the media. Enterococci
with vanC can cause significant infections,37 but this is rare
and they have not been implicated in nosocomial outbreaks.38
In most situations when screening for GRE it might therefore
be considered an advantage if enterococci with only the vanC
glycopeptide resistance genes did not grow on GRE screening
media. This would avoid unnecessary work involved in
292
Table 3. Organisms other than enterococci isolated from 218 faecal specimens
Number of isolates
Culture method
293
Direct plating on VRE selective
agar
Direct plating on enterococcosel
agar with vancomycin
6 h enrichment in VRE enrichment
broth and plating on VRE
selective agar
24 h enrichment in VRE enrichment
broth and plating on VRE
selective agar
24 h enrichment in enterococcosel
broth and plating on enterococcosel
agar with vancomycin
Total isolates
Gram-positive cocci
except staphylococci
Staphylococci
Yeasts
colonies like colonies not
colonies like colonies not
colonies like colonies not
colonies like colonies not
enterococci like enterococci enterococci like enterococci enterococci like enterococci enterococci like enterococci
55
7
11
1
2
4
3
29
25
3
7
4
0
2
0
1
32
2
5
2
1
3
2
24
31
6
3
3
6
8
8
32
14
0
3
4
0
1
0
0
65
15
18
9
7
8
11
49
Isolation of glycopeptide-resistant enterococci
Gram-positive bacilli
D. F. J. Brown and E. Walpole
identification and susceptibility testing of isolates that are
unlikely to cause infections or require infection control precautions.
Enrichment in VSB for 24 h did not significantly increase
the rate of isolation of GRE on VSA, although there was a
marginal increase in the number of specimens positive for
GRE; and enrichment in EB did not increase the rate of isolation of GRE on EA. These results conflict with a study by
Ieven et al.,28 which indicated that broth enrichment significantly increases isolation of GRE when screening faeces. The
difference may relate to several differences in the studies.
First, vancomycin was added to EB in the study by Ieven
et al.28 but not in ours. The benefits of adding vancomycin to
enrichment media have not been established and may depend
on the particular combination of enrichment broth and
selective agar. Klare et al.39,40 have reported isolation of some
GRE only after enrichment in a medium without vancomycin.
Secondly, the study by Ieven et al.28 included enterococci
with vanC, which accounted for 56% of their isolates.
Thirdly, there were differences in the patient populations
studied, in that Ieven et al.’s study28 included patients from all
hospital wards, whereas ours concentrated on patients known
to be at high risk of colonization with GRE. Patients in these
high-risk groups may be colonized with larger numbers of
GRE than other patients, in which case the benefit of a more
sensitive enrichment method would be reduced. Fourthly,
the study by Ieven et al.28 included rectal swabs if faecal
specimens were not available, and the numbers of GRE in
rectal swabs may be lower than in faecal samples, which again
might favour a more sensitive enrichment method. Detection
of GRE carriers colonized with low numbers of GRE may be
important in control of epidemic strains of GRE, so it would
be useful to extend the present work to other patient groups
and to rectal or peri-anal swab samples, as these are included
in some GRE screening protocols because swab samples are
easier to obtain.
The use of enrichment broth adds 24–48 h to the time taken
for screening, which is undesirable if screening is undertaken
for infection control purposes. In order to investigate whether
enrichment for a short period offered any benefit over direct
plating, VEB cultures were subcultured after incubation for
6 h. However, there were slightly fewer isolates of GRE when
VEB was subcultured after 6 h, and the density of growth was
frequently less than that obtained with direct plating or 24 h
enrichment. It is likely that 6 h is insufficient time for recovery
and growth of VRE following the initial dilution of organisms
in broth.
Growth of organisms other than GRE on GRE selective
media is undesirable because it leads to additional work in
checking isolates, and even if colonies are not typical of GRE
in appearance they can obscure GRE colonies or make it
difficult to subculture colonies for confirmation. A few
vancomycin-susceptible enterococci grew on both media.
Increasing the concentration of vancomycin in the media
would increase specificity, but isolation of GRE with lowlevel resistance is likely to be compromised and concentrations of vancomycin >6 mg/L are not recommended.26,27
Non-GRE colonies were isolated more frequently on VSA,
particularly on direct plating. While many of these organisms
could be distinguished because they did not give typical
colonies on the aesculin-based media, significant numbers
were not distinguishable from GRE solely on colony appearance. Non-GRE colonies commonly grew more slowly than
GRE, with colonies not clearly visible until 48 h incubation,
but this was also the case with some GRE, so the colonies
could not be discounted.
In conclusion, VSA and VEB were as effective as EA and
EB for the isolation of GRE from faecal specimens. The
media differed in that VSA and VEB were more effective in
preventing growth of enterococci with vanC, whereas EA and
EB were generally more effective at preventing growth of
other organisms. Enrichment was not shown significantly to
increase isolation of GRE in the largely high-risk population
studied, and enrichment for only 6 h may be deleterious.
Acknowledgements
This study was supported by a grant from Oxoid Ltd.
References
1. Martone, W. J. (1998). Spread of vancomycin-resistant enterococci: why did it happen in the United States? Infection Control and
Hospital Epidemiology 19, 539–45.
2. Hospital Infection Control Practices Advisory Committee.
(1995). Recommendations for preventing the spread of vancomycin
resistance. Infection Control and Hospital Epidemiology 16, 46–65.
3. Handwerger, S., Raucher, B., Altarac, D., Monka, J.,
Marchione, S., Singh, K. V. et al. (1993). Nosocomial outbreak due
to Enterococcus faecium highly resistant to vancomycin, penicillin
and gentamicin. Clinical Infectious Diseases 16, 750–5.
4. Livornese, L. L., Dias, S., Samel, C., Romanowski, B., Taylor, S.,
May, P. et al. (1992). Hospital acquired infection with vancomycinresistant Enterococcus faecium transmitted by electronic thermometers. Annals of Internal Medicine 117, 112–6.
5. Uttley, A. H. C., George, R. C., Naidoo, J., Woodford, N.,
Johnson, A. P., Collins, C. H. et al. (1989). High level vancomycinresistant enterococci causing hospital infections. Epidemiology and
Infection 31, 173–81.
6. Chadwick, P. R., Oppenheim, B. A., Fox, A., Woodford, N.,
Morgenstern, G. R. & Scarffe, J. H. (1996). Epidemiology of an outbreak due to vancomycin-resistant Enterococcus faecium on a
leukaemia unit. Journal of Hospital Infection 34, 171–82.
7. Edmond, M. B., Ober, J. F., Weinbaum, D. L., Pfaller, M. A.,
Hwang, T., Sanford, M. D. et al. (1995). Vancomycin-resistant
Enterococcus faecium bacteraemia: risk factors for infection.
Clinical Infectious Diseases 20, 1126–33.
294
Isolation of glycopeptide-resistant enterococci
8. Henning, K. J., deLencastre, H., Eagen, J., Boone, N., Brown,
A., Chung, M. et al. (1996). Vancomycin-resistant Enterococcus
faecium on a paediatric oncology ward. Paediatric Infectious
Diseases Journal 15, 848–54.
9. Papanicolaou, G. A., Meyers, B. R., Meyers, J., Mendelson, M.
H., Lou, W., Emre, S. et al. (1996). Nosocomial infections with
vancomycin-resistant Enterococcus faecium in liver transplant
recipients: risk factors for acquisition and mortality. Clinical Infectious Diseases 23, 760–6.
10. Mato, R., deLencestre, H., Carraher, M., Roberts, R. B. &
Tomasz, A. (1996). Multiplicity of genetic backgrounds among
vancomycin-resistant Enterococcus faecium isolates recovered
from an outbreak in a New York City Hospital. Microbial Drug
Resistance 2, 309–17.
11. Lam, S., Singer, C., Tucci, V., Morthland, V. H., Pfaller, M. A. &
Isenberg, H. D. (1995). The challenge of vancomycin-resistant
enterococci: a clinical and epidemiologic study. American Journal of
Infection Control 23, 170–80.
12. Boyce, J. M., Mermel, L. A., Zervos, M. J., Rice, L. B., PotterBynoe, G., Giorgio, C. et al. (1995). Controlling vancomycinresistant enterococci. Infection Control and Hospital Epidemiology
16, 634–7.
22. Barton, A. L. & Doern, G. V. (1995). Selective media for detecting gastrointestinal carriage of vancomycin-resistant enterococci.
Diagnostic Microbiology and Infectious Diseases 23, 119–22.
23. Sahm, D. F., Free, L., Smith, C., Eveland, M. & Mundy, L. M.
(1997). Rapid characterization schemes for surveillance isolates of
vancomycin-resistant enterococci. Journal of Clinical Microbiology
35, 2026–30.
24. Ford, M., Perry, J. D. & Gould, F. K. (1994). Use of cephalexin–
aztreonam–arabinose agar for selective identification of Enterococcus faecium. Journal of Clinical Microbiology 32, 2999–3001.
25. Nelson, R. R. S., McGregor, K. F., Brown, A. R., Amyes, S. G. B.
& Young, H.-K. (2000). Isolation and characterization of glycopeptideresistant enterococci from hospitalized patients over a 30-month
period. Journal of Clinical Microbiology 38, 2112–6.
26. Swenson, J. M., Clark, N. C., Ferraro, M. J., Sahm, D. F.,
Doern, G., Pfaller M. A. et al. (1994). Development of a standardized screening method for detection of vancomycin-resistant
enterococci. Journal of Clinical Microbiology 32, 1700–4.
27. Chadwick, P. R., Brown, D. F. J., Wilcox, M. H., Collyns, T. A.,
Walpole, E., Dillon, J. et al. (1997). Comparison of agar-based
media for the primary isolation of glycopeptide-resistant enterococci. Clinical Microbiology and Infection 3, 559–63.
13. Centres for Disease Control and Prevention. (1995). Recommendations for preventing the spread of vancomycin resistance:
recommendations of the Hospital Infection Control Practices
Advisory Committee (HICPAC). Morbidity and Mortality Weekly
Reports 44, 1–13.
28. Ieven, M., Vercauteren, E., Descheemaeker, P., van Laer, F.
& Goossens, H. (1999). Comparison of direct plating and broth
enrichment culture for the detection of intestinal colonization by
glycopeptide-resistant enterococci among hospitalized patients.
Journal of Clinical Microbiology 37, 1436–40.
14. Handwerger, S., Perlman, D. C. & Altarac, D. (1992). Concomitant high-level vancomycin and penicillin resistance in clinical
isolates of enterococci. Clinical Infectious Diseases 14, 655–61.
29. Satake, S., Clark, N., Rimland, D., Nolte, F. S. & Tenover, F. C.
(1997). Detection of vancomycin-resistant enterococci in fecal
samples by PCR. Journal of Clinical Microbiology 35, 2325–30.
15. Montecalvo, M. A., Horowitz, H. & Gedris, C. (1994). Outbreak
of vancomycin-, ampicillin- and aminoglycoside-resistant Enterococcus faecium bacteremia in an adult oncology unit. Antimicrobial
Agents and Chemotherapy 38, 1363–7.
30. Jayaratne, P. & Rutherford, C. (1999). Detection of clinically
relevant genotypes of vancomycin-resistant enterococci in nosocomial surveillance specimens by PCR. Journal of Clinical Microbiology 37, 2090–2.
16. Edmond, M. B., Ober, J. F. & Weinbaum, D. L. (1995). Vancomycin-resistant Enterococcus faecium bacteraemia: risk factors for
infection. Clinical Infectious Diseases, 20, 1126–33.
31. Van Horn, K. G., Gedris, C. A. & Rodney, K. M. (1996).
Selective isolation of vancomycin-resistant enterococci. Journal of
Clinical Microbiology 34, 924–7.
17. Gopal Rao, G., Ghanekar, K. & Ojo, F. (1996). Selective
medium for screening for vancomycin-resistant enterococci in
faeces. European Journal of Clinical Microbiology and Infectious
Diseases 15, 175–7.
32. Landman, D., Quale, J. M., Oydna, E., Willey, B., Ditore, V.,
Zaman, M. et al. (1996). Comparison of five selective media for
identifying fecal carriage of vancomycin resistant enterococci.
Journal of Clinical Microbiology 34, 751–2.
18. Karanfil, L. V., Murphy, M. & Josephson, A. (1992). A cluster of
vancomycin-resistant Enterococcus faecium in an intensive care
unit. Infection Control and Hospital Epidemiology 13, 195–200.
33. Dutka-Malen, S., Evers, S. & Courvalin, P. (1965). Detection of
glycopeptide resistance genotypes and identification to the species
level of clinically relevant enterococci by PCR. Journal of Clinical
Microbiology 33, 24–7.
19. Edberg, S. C., Hardalo, C. J., Kontnick, C. & Campbell S.
(1994). Rapid detection of vancomycin-resistant enterococci.
Journal of Clinical Microbiology 32, 2182–4.
20. Boyce, J. M., Opal, S. M., Chow, J. W., Zervos, M. J., PotterBynoe, G., Sherman, C. B. et al. (1994). Outbreak of multidrugresistant Enterococcus faecium with transferable vanB class
vancomycin resistance. Journal of Clinical Microbiology 32,
1148–53.
21. Chadwick, P. R. & Oppenheim, B. A. (1995). Neomycin agar as
a selective medium for Enterococcus faecium. Journal of Clinical
Pathology 48, 1068–70.
34. Jayaratne, P. & Rutherford, C. (1999). Detection of clinically
relevant genotypes of vancomycin-resistant enterococci in nosocomial surveillance specimens by PCR. Journal of Clinical Microbiology 37, 2090–2.
35. Andrews, J. M. for the BSAC Working Party on Susceptibility
Testing. (2001). BSAC standardized disc susceptibility testing
method. Journal of Antimicrobial Chemotherapy 48, Suppl. S1,
S43–58.
36. Tremlett, C. H., Brown, D. F. J. & Woodford, N. (1999). Variation in structure and location of VanA glycopeptide resistance
295
D. F. J. Brown and E. Walpole
elements among enterococci from a single patient. Journal of
Clinical Microbiology 37, 818–20.
37. Kaplan, A. H., Gilligan, P. H. & Facklam, R. R. (1988). Recovery
of resistant enterococci during vancomycin prophylaxis. Journal of
Clinical Microbiology 26, 1216–8.
39. Klare, I., Heier, H., Klaus, H., Bohme, R. G., Marin, S.,
Seltmann, G. et al. (1995). Enterococcus faecium strains with vanA
mediated high level resistance isolated from animal foodstuffs and
fecal samples of humans in the community. Microbial Drug Resistance 1, 265–72.
38. Toye, B., Shymanski, J., Bobrowska, M., Woods, W. &
Ramotar, K. (1997). Clinical and epidemiologic significance of
enterococci intrinsically resistant to vancomycin (possessing the
vanC genotype). Journal of Clinical Microbiology 35, 3166–70.
40. Klare, I., Heier, H., Klaus, H., Reissbrodt, R. & Witte, W. (1995).
vanA mediated high level glycopeptide resistance in Enterococcus
faecium from animal husbandry. FEMS Microbiology Letters 125,
165–71.
296