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The Canadian Journal of Infectious Diseases logoLink to The Canadian Journal of Infectious Diseases
. 2000 Jan-Feb;11(1):38–41. doi: 10.1155/2000/356317

Low prevalence of VRE gastrointestinal colonization of hospitalized patients in Manitoba tertiary care and community hospitals

George G Zhanel 1,2,3,5, Godfrey KM Harding 1,2,3,4,5,6,7, Stuart Rosser 1,5, Daryl J Hoban 1,3, James A Karlowsky 1,2,3, Michelle Alfa 1,7, Amin Kabani 1,3, John Embil 1,4,5, Alfred Gin 2,6, Trevor Williams 1,5, Lindsay E Nicolle 1,4,5
PMCID: PMC2094745  PMID: 18159264

Abstract

OBJECTIVE:

To determine the prevalence of vancomycin-resistant enterococci (VRE) bowel colonization in hospitalized patients in Manitoba who had stool specimens collected for Clostridium difficile toxin and/or culture testing.

DESIGN:

Two tertiary care and five community hospitals in Winnipeg and three rural Manitoba community hospitals participated in this study. From January 1 to December 31, 1997 stool specimens, one per patient, submitted to hospital microbiology laboratories for C difficile toxin and/or culture testing were screened for VRE on colistin-nalidixic acid-vancomycin (6 μg/mL) (CNAV) agar plates. The study was divided into six, eight-week intervals. Stool specimens received in the first two weeks of each eight week interval were screened for VRE.

MAIN RESULTS:

A total of 1408 stool specimens were submitted over the 48-week study period. Sixty-seven (4.8%) patients with VRE colonization of their lower gastrointestinal tract were identified. Three of the 67 (4.5%) VRE isolates were Enterococcus faecium, with the remaining 64 (95.5%) were Enterococcus gallinarum. The three vancomycin-resistant E faecium -VREF- (from two different Winnipeg hospitals) demonstrated the vanA genotype, and were resistant to vancomycin, teicoplanin and ampicillin. All three VREF isolates also demonstrated high level resistance to both gentamicin and streptomycin but were susceptible to quinuprisitin/dalfopristin and LY333328.

CONCLUSION:

VRE colonization in hospitalized patients in Manitoba is infrequent and most commonly due to E gallinarum. The prevalence of VREF colonization in the patients studied was 0.2% (three of 1408).

Key Words: Manitoba, Prevalence, Vancomycin-resistant enterococci


Vancomycin-resistant Enterococcus faecium (VREF) accounts for up to 65% of E faecium isolates in hospitalized patients across the United States and is endemic in many North American tertiary care institutions (1,2). The management of these infections presents a significant clinical challenge because species of the genus Enterococcus, and in particular E faecium, are frequently resistant to several antimicrobial agents (3). High level penicillin resistance, high level aminoglycoside resistance and most recently vancomycin resistance are emerging as significant concerns in the treatment of enterococcal infections. This has prompted the development and evaluation of new antimicrobial agents such as quinupristin/dalfopristin and LY333328, a glycopeptide, which may offer activity against enterococci resistant to conventional therapy (2).

VREF is not endemic in Manitoba hospitals, and infection with VREF is extremely rare (4). However, the prevalence of VREF lower gastrointestinal tract (GIT) carriage, which frequently precedes infection (5,6), is presently unknown for patients hospitalized in Manitoba. To determine whether the lack of VREF endemnicity correlated with an absence of lower GIT colonization, we assessed lower GIT carriage of VREF for patients hospitalized in 10 Manitoba hospitals from January 1 to December 31, 1997. Our study was consistent with Centers for Disease Control and Prevention guidelines (Atlanta, Georgia) that suggest surveillance programs for vancomycin-resistant enterococci (VRE) be undertaken on an intermittent basis in areas where VRE is not known to be endemic (6). Isolates of VREF identified were phenotypically and genotypically characterized, and tested for their susceptibilities against a panel of antimicrobial agents.

MATERIALS AND METHODS

Enterococcus species isolate collection:

Seven Winnipeg hospitals (Health Sciences Centre, St Boniface General Hospital, Misericordia General Hospital, Concordia General Hospital, Victoria General Hospital, Grace General Hospital, and Seven Oaks General Hospital) as well as hospitals in the rural Manitoba communities of Brandon, Thompson and Portage la Prairie participated. The Health Sciences Centre and St Boniface General Hospital are tertiary care teaching hospitals affiliated with the University of Manitoba. The remaining eight hospitals are community hospitals.

The study was divided into six, eight-week intervals, and extended from January 1 to December 31, 1997. Each hospital collected stool specimens submitted for Clostridium difficile toxin and/or culture testing in the first two weeks of each eight week interval from patients in their hospitals. One stool specimen per patient was considered. Stool specimens were frozen following collection and shipped to one of two central reference laboratories (Health Sciences Centre and the St Boniface General Hospital) where they were thawed and planted onto blood agar (BA), colistin-nalidixic acid agar (CNA), and colistinnalidixic acid agar supplemented with 6 μg/mL vancomycin (CNAV). All plates were incubated at 35°C for 48 h.

All morphotypes of suspected Enterococcus species were selected from BA, CNA or CNAV plates, stocked in skim milk at -80°C, and subsequently identified to species level using the following tests: Gram stain; motility assessment; catalase production; growth in 6.5% sodium chloride; L-pyrrolidonylb-naphthalamide hydrolysis; xylose, mannitol, arabinose and sorbitol utilization; bile/esculin growth/hydrolysis; pigment production; leucine aminopeptidase activity; and acidification of methyl-α-D-glucopyranoside (7,9). The identification of all discrepant organisms was determined using DNA sequencing (8,10).

Vancomycin resistance genotyping and epidemiological typing:

DNA lysates of all isolated VREF were subjected to multiplex polymerase chain reaction (PCR) to detect the presence of vanA, vanB, vanC-1 and vanC-2/vanC-3 genotypes (11), and pulse field gel electrophoresis (PFGE), following SmaI digestion, for epidemiological tracing purposes (12). Similar PFGE patterns were defined as those that differed from one another by no more than one or two clearly visible bands (12).

Antimicrobial agents and susceptibility testing:

All isolated VREF were tested for their susceptibility to vancomycin (Eli Lilly Canada Inc, Scarborough, Ontario), teicoplanin (Hoechst Marion Roussel Canada Inc, Montreal, Quebec), ampicillin (Wyeth Ayerst Canada Inc, North York), streptomycin (Pfizer Canada Inc, Kirkland, Quebec), gentamicin (Schering Canada Inc, Pointe-Claire, Quebec), ciprofloxacin (Bayer Inc, Etobicoke, Ontario), quinuprisitin/dalfopristin (Rhone Poulenc Rorer, Montreal, Quebec) and LY333328 (Lilly Research Laboratories, Indianapolis, Indiana) using the microdilution broth method and Mueller-Hinton broth as described by the National Committee of Clinical Laboratory Standards (Wayne, Pennsylvania (13). Before susceptibility testing all Enterococcus species were subcultured twice onto blood agar.

RESULTS

The number of stool specimens evaluated in the first two weeks of each eight week period between January 1 and December 31, 1997 ranged from 187 to 265. A total of 1408 stool specimens were tested. Sixty-seven of the 1408 (4.8%) hospitalized patients evaluated had VRE colonization of their lower GIT. Three of the 67 (4.5%) VRE identified were E faecium, while the remaining 64 isolates (95.5%) were Enterococcus gallinarum. Three of 1408 (0.2%) patients hospitalized in Manitoba, who had a stool specimen submitted for C difficile testing during the study period, were colonized with VREF. One isolate of VREF was identified at the Misericordia General Hospital, a Winnipeg community hospital, between April 23 and May 7, 1997. The two remaining isolates of VREF were identified between June 18 and July 2, 1997 at the Health Sciences Centre, a Winnipeg tertiary care hospital.

All three VREF identified possessed the vanA genotype (Table 1). The two isolates of VREF, from the Health Sciences Centre, possessed the same PFGE pattern while the third isolate, collected from a patient at the Misericordia General Hospital, had a different PFGE pattern (Table 1). Both PFGE patterns have been previously identified in VREF isolated in Manitoba (unpublished data), suggesting they may have been acquired locally or nosocomially. Using NCCLS serum breakpoints (14), the three isolates of VREF demonstrated uniform resistance to teicoplanin, ampicillin, streptomycin, gentamicin, and ciprofloxacin, but were sensitive to quinuprisitin/dalfopristin and LY333328 (Table 1).

TABLE 1.

Phenotypic and genotypic characteristics of the three vancomycin-resistant Enterococcus faecium (VREF) isolated identified as colonizers of the lower gastrointestinal tract of patients in Manitoba hospitals between January and December 1997.

MICS (μg/mL)
VREF isolate PFGE profile V T A S G C Q/D LY
1* vanA 1 128 32 64 >4000 >2000 >32 0.125 1
2 vanA 2 256 32 64 >4000 >2000 >32 0.125 0.5
3 vanA 2 256 32 64 >4000 >2000 >32 0.125 0.5
*

Isolate from a patient in Misericordia General Hospital, Winnipeg, Manitoba, collected between April 23 and May 7, 1997;

Isolate from a patient in Health Sciences Centre, Winnipeg, Manitoba, collected between June 18 and July 2, 1997.

A Ampicillin; C Ciprofloxacin; G Gentamicin; LY LY333328; Q/D Quinupristin/dalfopristin; S Streptomycin; T Teicoplanin; V Vancomycin

DISCUSSION

Enterococci are normal constituents of the human GIT, and may also colonize the oral cavity, vagina, biliary tract and upper respiratory tract of healthy people. Most clinical isolates of enterococci represent colonization, not infection (15). Enterococci may however cause infection, most commonly urinary tract infections (15). Cholecystitis, cholangitis, peritonitis, septicemia, endocardititis, meningitis and wound infections have also been documented (15). The GIT is the major reservoir for enterococci, and serves as the source of dissemination. However, the ability of enterococci to survive on environmental surfaces for extended periods (up to a week) can also facilitate their spread (15).

Traditionally, enterococcal infections have been treated with the penicillins, usually in combination with an aminoglycoside for serious infections. However, with penicillin and aminoglycoside resistance rates increasing vanconiycin is now often used to treat enterococcal infections. VRE were extremely rare before 1988 with just two isolates (MIC greatern than 100 μg/mL) documented in the previous two decades (16,17). In 1988, VRE began to emerge as a clinical concern, with the description of 55 clinical isolates of VRE (seven Enterococcus faecalis and 48 E faecium) (MICs greater than 64 μg/mL) from 22 British patients with renal impairment (18). In 1993, the American National Nosocomial Infections Surveillance (NNIS) system reported an increase in the incidence of nosocomial VRE from 0.3% in 1989 to 7.9% in 1993 (19). Many scientific publications from the United States and Europe have followed with reports of as many as 65% of E faecium isolates being vancomycin resistant in some American hospitals (1). The epidemiology of VRE in Canada before 1997 has been succinctly summarized (20), with the first isolate of VRE reported in Edmonton in 1993 and the first VRE outbreak, involving 38 patients, occurring in a Toronto hospital in 1995. A 1996 Canadian VRE surveillance study demonstrated a VRE (E faecalis or E faecium) point prevalence of 0.7% overall (26 of 3773), 0.1% in nonendemic hospitals, 3.7% in endemic hospitals and 5.3% in endemic patient groups within endemic hospitals (21). Twenty-three of the 26 patients identified in this study were from a single tertiary care institution with all isolates (three E faecalis and 23 E faecium) being colonizers (21).

VRE were not identified in a previous study over a four-month period in 1995 at the Health Sciences Centre in Winnipeg, one of the 10 Manitoba hospitals included in this survey (4). As a continuation of that study, we added six additional Winnipeg and three rural hospitals. Only three isolates, from two hospitals, representing VREF colonization of patients hospitalized in Manitoba were identified (0.2%). Isolates of vancomycin-resistant E faecalis were not identified.

CONCLUSIONS

We conclude that VRE colonization of patients hospitalized in Manitoba remains infrequent; however ongoing surveillance will be necessary to ensure adequate knowledge of changing trends in VRE epidemiology. A simple and effective technique to screen for the emergence of VRE is to evaluate stool specimens submitted for C difficile analysis.

Acknowledgments

This study was supported by a grant from the Manitoba Medical Service Foundation. Dr Zhanel is supported by a Merck Frosst Chair in Pharmaceutical Microbiology. We gratefully acknowledge each participating Manitoba hospital and their laboratory staff as well as the Cadham Provincial Laboratory in Winnipeg.

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