Glycopeptide-resistant Enterococci Research Articles

Significant Increase in the Isolation of Glycopeptide-Resistant Enterococci From Patients Hospitalized in the Transplant Surgery Ward in 2004–2005

Enterococci despite their low pathogenicity are the third cause of hospital infections. Enterococci resistant to glycopeptides present special risks. The aim of this work was to determine the frequency of isolates of all enterococci versus enterococci resistant to glycopeptides from patients in the Transplant Surgery Ward. Moreover, vancomycin-resistant enterococci (VRE) were characterized with respect to the type of van and ddl genes as well as vancomycin and teicoplanin MIC values. Among 160 enterococcal strains isolated in 2004, only 2 were resistant to glycopeptides (1.3%). In 2005, among 244 enterococci, 44 strains were resistant (18%). All resistant strains were Enterococcus faecium, as confirmed by detection of the ddl gene specific for E. faecium. Moreover, among all enterococci isolated from these patients, E. faecium dominated (over 50% in 5 subsequent years). All examined VRE possessed VanA type of resistance with high vancomycin and teicoplanin MIC values. All of them possessed the vanA ligase gene. The investigated VRE were characterized by high resistance to most antibiotics: penicillin and amoxicillin, rifampicin, ciprofloxacin, and high concentrations of streptomycin, but susceptible to linezolid and quinupristine/dalfopristine. Strains differed in their susceptibility to tetracycline, nitrofurantoin, and high concentrations of gentamicin.

Glycopeptide-resistant enterococci: deciphering virulence, resistance and epidemicity

Since their first discovery, glycopeptide-resistant enterococci have emerged as important nosocomial pathogens first in the US, followed by the rest of the world. In this review the most recent findings that relate to enterococcal epidemiology, virulence and glycopeptide-resistance maintenance will be discussed. Frequent horizontal gene transfer and recombination, resulting in high-level genome plasticity, facilitating rapid responsiveness of enterococci to changing environmental conditions may have contributed to the worldwide emergence. For Enterococcus faecium this has resulted in the development of a distinct genetic subspecies, clonal complex 17, responsible for the majority of glycopeptide-resistant enterococci-related hospital burden. Preliminary data also suggest that such high-risk enterococcal clonal complexes may exist within Enterococcus faecalis. The last 2 years have not only disclosed novel determinants implicated in enterococcal pathogenesis, but also showed that enterococci are able to sense their environment and regulate virulence gene expression accordingly. Linkage of glycopeptide resistance in enterococci to plasmid maintenance systems holds a doomed perspective for controlling antibiotic resistance emergence. Recent developments have improved our understanding of enterococcal population structure, pathogenesis and glycopeptide-resistance maintenance. This may contribute to the development of novel intervention strategies to prevent enterococcal infections and contain the spread of glycopeptide-resistant enterococci.

Multidrug-Resistant Bacteria in Southeastern Austria

To the Editor: In many parts of the world, the proportions of methicillin-resistant Staphylococcus aureus (MRSA), vancomycin-resistant enterococci, and extended-spectrum β-lactamase (ESBL)–producing organisms in the family Enterobacteriaceae have increased remarkably during recent years (1). However, proportions of antimicrobial drug resistance vary substantially at national and regional levels. We describe antimicrobial drug resistance data for hospitalized patients and outpatients in southeast Austria. A total of 690,967 clinical samples were collected from hospitalized patients and outpatients and analyzed at the microbiology laboratory of the Medical University of Graz during 1997–2006. Selected for resistance surveillance were nonduplicate isolates of S. aureus, Enterococcus faecium, E. faecalis, Escherichia coli, and Klebsiella spp. Antibiotic susceptibilities were determined by using disk diffusion and the VITEK2 system (bioMerieux, Marcy l’Etoile, France) with specific susceptibility test cards. Etest (AB Biodisk, Solna, Sweden) was used to confirm results. Test results were interpreted according to the recommendations of the Clinical and Laboratory Standards Institute (2). During the study period, the proportion of patients with MRSA remained stable (2.5%–4.9%) (Figure, panel A). The prevalence of MRSA among invasive S. aureus isolates ranges between 0.5% and 44.4% in European countries and has increased in recent years (3). We found MRSA predominantly in samples from hospitalized patients (median 72.1%); however, the incidence of community-acquired MRSA increased slightly during recent years, similar to that of other central European countries (4). Vancomycin resistance was not noted during the study period; however, 4 vancomycin-intermediate MRSA isolates were noted in 2004, 2005, and 2006, in concordance with the sporadic occurrence of MRSA with intermediate susceptibility to glycopeptides recently reported for other European countries (3). The percentage of patients with vancomycin-resistant E. faecium and E. faecalis was low (median 0.4%) (Figure, panel A). In total, 10 E. faecium and 4 E. faecalis isolates with resistance to vancomycin were reported; most were from hospitalized patients. As in most European countries, human infections due to glycopeptide-resistant enterococci remain rare in Austria, although a high proportion of glycopeptide-resistant E. faecium was reported recently from animals used in food production (5). Among E. coli isolates, no ESBL producers were noted in 1997. From 1998 through 2002, proportions of ESBL-producing E. coli were 0.06%–0.13%, which corresponds to 3–6 isolates per year. A subsequent increase of ESBL-producing E. coli isolates was noted, from 19 (0.3%) in 2003 to 148 (2.4%) in 2006 (Figure, panel B). Most (67%) ESBL-producing isolates found during 2003–2006 originated from community-acquired urinary tract infections. Resistance of E. coli to carbapenems was not reported during the study period. Among Klebsiella spp. isolates, 2 (0.2%) ESBL producers were observed during 1997. From 1998 through 2004, the prevalence of ESBLs among Klebsiella spp. ranged between 0.6% and 1.6%. In 2005 and 2006, the rate of ESBL-producing Klebsiella spp. increased to 3.8% (44 isolates) and 4.5% (55 isolates), respectively, and originated mainly from intensive care units (Figure, panel B). In 2005, a single Klebsiella pneumoniae isolate showed reduced susceptibility to imipenem (MIC 2 μg/mL) and to meropenem (MIC 4 μg/mL) and resistance to ertapenem (MIC >16 μg/mL). Nevertheless, production of ESBL by Enterobacteriaceae organisms is still rare in southeast Austria compared with other European countries (6). However, a dramatic increase of ESBL-producing E. coli and Klebsiella spp. has been observed during recent years. The increase of ESBL-producing E. coli isolates in outpatients with urinary tract infections leads to serious treatment problems. Results from a recent study indicate that the increase of ESBL-producers in southeast Austria is caused mainly by the emergence of CTX-M–type ESBLs, which are increasingly being isolated from outpatients (7). The K. pneumoniae isolate found in 2005 represents the first ESBL-producing isolate not susceptible to carbapenems reported from Austria. Development of resistance to carbapenems in Enterobacteriaceae organisms has been reported increasingly, which substantially limits treatment options for persons with multidrug-resistant gram-negative infections (8). Our data show insignificant changes in prevalence of MRSA and vancomycin-resistant enterococci in southeast Austria during the past decade but an alarming increase of multidrug-resistant ESBL-producing E. coli and Klebsiella spp. isolates in recent years. Detection of an ESBL-producing K. pneumoniae isolate with reduced susceptibility to carbapenems shows that pathogens with new mechanisms of resistance are emerging in this region. Figure A) Proportion of methicillin resistance in Staphylococcus aureus and vancomycin resistance in Enterococcus faecium and E. faecalis in southeastern Austria, 1997–2006. B) Proportion of extended-spectrum β-lactamase–producing (ESBL) ...

Novel Semisynthetic Derivative of Antibiotic Eremomycin Active against Drug-Resistant Gram-Positive Pathogens Including Bacillus anthracis

Five adamantyl-containing carboxamides of eremomycin or vancomycin were synthesized and their antibacterial activities against some Gram-positive clinical isolates were investigated in vitro and in vivo. The adamantyl-2 amide of glycopeptide antibiotic eremomycin (1a in Chart 1, AN0900) was the most active compound and showed high activity against several Gram-positive pathogens: vancomycin-susceptible staphylococci and enterococci, glycopeptide-intermediate-resistant Staphylococcus aureus, and glycopeptide-resistant enterococci. Compound 1a was equally active in vitro against both Ciprofloxacin-susceptible and -resistant Bacillus anthracis strains (MICs 0.25-0.5 microg/mL). It was distinguished by having a 2.8 h half-life (t1/2) in mice and a volume of distribution of 2.18 L/kg. Compound 1a was active against Staphylococcus aureus in mice (iv) and provided complete protection against a lethal intravenous challenge with vegetative B. anthracis bacilli and also in a murine pulmonary anthrax model in which mice were challenged with Bacillus anthracis spores.

Three novel highly charged copper-based biocides: safety and efficacy against healthcare-associated organisms

We investigated three novel highly charged copper-based inorganic biocidal formulations for their activity against organisms highly relevant to healthcare-associated infection. The three copper-based formulations were tested: (i) against clinical isolates of methicillin-resistant Staphylococcus aureus (MRSA), Legionella pneumophila, Acinetobacter calcoaceticus/baumannii (ACCB), glycopeptide-resistant Enterococcus and spores of Clostridium difficile in time-kill assays; (ii) for their ability to decontaminate ultramicrofibre (UMF) cloths; and (iii) for their cytotoxicity to human skin and intestinal epithelial cells. All three copper-based formulations were potently biocidal down to concentrations of 1 ppm for both stationary- and log-phase organisms, and they were all active against C. difficile spores. At 150 ppm, they achieved a complete (>6 log10) kill of MRSA and ACCB mostly within 1 h. This biocidal activity was not achieved by copper sulphate or the inorganic binders used in the formulations. All three copper-based formulations completely decontaminated UMF cloths containing MRSA, ACCB or C. difficile spores, suggesting that any of these copper-based formulations would be highly beneficial in the healthcare environment. All three copper-based formulations and copper sulphate were not cytotoxic to human epithelial cells up to concentrations of 100-200 ppm. All three of the novel copper-based biocidal formulations, but not their components (copper sulphate and inorganic binders), have potent activity against organisms highly relevant to healthcare-associated infections.

The challenge of healthcare-associated infection outside hospital

Infection control is an area of healthcare practice subject to rapid development in line with changes in the behaviour of pathogens, advances in technology, the altered profile of people undergoing healthcare, the settings in which that care is delivered and the impact of new ways of working throughout the National Health Service. This article describes how micro-organisms responsible for healthcare-associated infection can be transferred between hospital and community settings and explains the risks of disseminating infection in relation to two types of pathogen carrying resistance to antibiotics: methicillin resistant Staphylococcus aureus and glycopeptide resistant enterococci. Ability to read current literature critically, to be aware of changing policy and its impact on the risk of infection forms an important part of the community nursing role.

N′-(α-Aminoacyl)- and N′-α-(N-Alkylamino)acyl Derivatives of Vancomycin and Eremomycin

The antibacterial activities of the series of novel N'-(alpha-aminoacyl)- and N'-alpha-(N-akylamino)acyl derivatives of eremomycin and vancomycin containing hydrophobic moieties have been investigated. The N'-(N-alkylglycyl) derivatives of vancomycin are more active against vancomycin-susceptible staphylococci and enterococci and glycopeptide intermediate-resistant Staphylococcus aureus (GISA) than the corresponding eremomycin derivatives, but except for N'-[N-(p-octyloxybenzyl)glycyl-vancomycin] (28) and N'-[N-(p-octyloxybenzyl)-L-alanyl-vancomycin (33)--they are less active against glycopeptide-resistant enterococci (GRE). Derivatives 28 and 33 are the most active compounds (MIC's for glycopeptide-sensitive staphylococci and enterococci are 0.25 approximately 1 microg/ml, for GISA 1 approximately 2 microg/ml, for GRE 2 approximately 6 microg/ml). In in vivo studies, derivative 28 was active against S. aureus infections in mice with ED(50) 1 mg/kg versus 2 mg/kg for vancomycin (iv). In general N'-(N-alkylglycyl)-derivatives of vancomycin and eremomycin were more active than the corresponding N'-aminoacylated derivatives of these antibiotics containing other than glycin amino acids (L-Lys, L-Met, L-Orn, L- and D-Ala) and also L- and D-Phe or benzyl-O-L-Tyr.

VanA-Type Enterococci from Humans, Animals, and Food: Species Distribution, Population Structure, Tn 1546 Typing and Location, and Virulence Determinants

VanA-type human (n=69), animal (n=49), and food (n=36) glycopeptide-resistant enterococci (GRE) from different geographic areas were investigated to study their possible reservoirs and transmission routes. Pulsed-field gel electrophoresis (PFGE) revealed two small genetically related clusters, M39 (n=4) and M49 (n=13), representing Enterococcus faecium isolates from animal and human feces and from clinical and fecal human samples. Multilocus sequence typing showed that both belonged to the epidemic lineage of CC17. purK allele analysis of 28 selected isolates revealed that type 1 was prevalent in human strains (8/11) and types 6 and 3 (14/15) were prevalent in poultry (animals and meat). One hundred and five of the 154 VanA GRE isolates, encompassing different species, origins, and PFGE types, were examined for Tn1546 type and location (plasmid or chromosome) and the incidence of virulence determinants. Hybridization of S1- and I-CeuI-digested total DNA revealed a plasmid location in 98% of the isolates. Human intestinal and animal E. faecium isolates bore large (>150 kb) vanA plasmids. Results of PCR-restriction fragment length polymorphism and sequencing showed the presence of prototype Tn1546 in 80% of strains and the G-to-T mutation at position 8234 in three human intestinal and two pork E. faecium isolates. There were no significant associations (P>0.5) between Tn1546 type and GRE source or enterococcal species. Virulence determinants were detected in all reservoirs but were significantly more frequent (P<0.02) among clinical strains. Multiple determinants were found in clinical and meat Enterococcus faecalis isolates. The presence of indistinguishable vanA elements (mostly plasmid borne) and virulence determinants in different species and PFGE-diverse populations in the presence of host-specific purK housekeeping genes suggested that all GRE might be potential reservoirs of resistance determinants and virulence traits transferable to human-adapted clusters.

Linezolid treatment of glycopeptide-resistant Enterococcus faecium in very low birth weight premature neonates

Glycopeptide-resistant Enterococcus faecium has emerged as an important nosocomial pathogen with limited therapeutic options. Here we report the successful treatment of glycopeptide-resistant E. faecium infection in two very low birth weight premature infants with the new oxazolidinone linezolid. Treatment with linezolid at a dosage of 10 mg/kg every 8 h intravenously for a duration of 16 days and 14 days, respectively, was well tolerated and led to complete clinical recovery and clearance of the organism from all body sites. The two cases support the clinical efficacy and safety of linezolid in very low birth weight premature neonates with glycopeptide-resistant E. faecium infections.

Prevalence, persistence, and molecular characterization of glycopeptide-resistant enterococci in Norwegian poultry and poultry farmers 3 to 8 years after the ban on avoparcin.

Environmental reservoirs of glycopeptide-resistant enterococci (GRE) in Norway have been linked to former growth promoting use of the glycopeptide avoparcin in poultry production. We have examined the prevalence of fecal GRE in poultry and poultry farmers 3 to 8 years after the Norwegian avoparcin ban in 1995 and performed molecular analyses of the GRE population. Fecal samples from poultry farmers and their flocks on 29 previously avoparcin-exposed farms were collected on five occasions during the study period (1998 to 2003). All flocks (100%) were GRE positive in 1998. Throughout the study period, 78.5% of the poultry samples were GRE positive. Glycopeptide-resistant Enterococcus faecium (GREF) was isolated from 27.6% of the farmer samples in 1998 and from 27.8% of the samples collected between 1998 and 2003. The prevalence of fecal GRE in poultry declined significantly during the study period, but prevalence in samples from the farmers did not decline. PCR analysis revealed a specific Tn1546-plasmid junction fragment in 93.9% of E. faecium isolates. A putative postsegregation killing (PSK) system linked to Tn1546 was detected in 97.1% of the isolates examined. Multilocus sequence typing of glycopeptide-susceptible (n = 10) and -resistant (n = 10) E. faecium isolates from humans (n = 10) and poultry (n = 10) on two farms displayed 17 different sequence types. The study confirms the continuing persistence of a widespread common plasmid-mediated vanA-pRE25-PSK element within a heterogeneous GRE population on Norwegian poultry farms 8 years after the avoparcin ban. Moreover, it suggests an important role of PSK systems in the maintenance of antimicrobial resistance determinants in reservoirs without apparent antimicrobial selection.

Clinical Glycopeptide-Resistant Enterococci Isolated From Patients After Solid Organ Transplantation

The aim of this study was to confirm the identification and resistance to vancomycin and teicoplanin of nosocomial enterococcal strains using molecular biology methods. Glycopeptide-resistant enterococci (GRE) strains were isolated from clinical specimens of hospitalized patients. Bacterial identification was performed in an automatic ATB Expression system (bioMérieux SA). Susceptibility to glycopeptides was determined by the disc diffusion method and Etest (AB BIODISK, Sweden). We performed polymerase chain reactions (PCR) for Enterococcus faecium and E. faecalis identification and van genes detection. Fifteen GRE strains were cultured over 2 years (2003–2004). Fourteen isolates were highly resistant to vancomycin (MIC range, 128–>256 mg/L) and teicoplanin (MIC range, 32–>256 mg/L). Twelve strains harbored van A gene (Van A phenotype). Seven isolates were identified as E. faecium and seven as E. faecalis by the multiplex-PCR method. One strain— E. casseliflavus—showed low resistance to vancomycin (MIC 8 mg/L) with retained susceptibility to teicoplanin (MIC 4 mg/L). It harbored the van C2/C3 gene and was identified as the Van C2/C3 phenotype. GRE strains were more often isolated from hospitalized patients in Poland. Constant monitoring by reliable microbiological methods has become necessary to prevent the spread of these strains in the hospital environment.

National Glycopeptide-Resistant Enterococcal Bacteraemia Surveillance Working Group Report to the Department of Health — August 2004

<h2>Summary</h2> <ul><li>1.From September 2003, acute National Health Service trusts in England have been required by the Department of Health to undertake mandatory surveillance of bacteraemias caused by glycopeptide-resistant enterococci (GRE).</li><li>2.In total, 4855 enterococcal bacteraemias (including 256 reports of ‘Group D streptococci') were reported through the voluntary reporting system to the Communicable Disease Surveillance Centre (CDSC) in England and Wales in 2002, and 8.9% of the reports that contained information on vancomycin susceptibility indicated that the isolate was resistant to vancomycin. Most hospitals reported fewer than five GRE bacteraemias per year to the CDSC, with only 13 hospitals reporting more than five episodes per year, although there is undoubtedly an element of under-reporting.</li><li>3.This Working Group was established to review the methods used to identify and test the susceptibility of enterococci to glycopeptides, and to make recommendations on the reporting of GRE bacteraemias, as there were indications that current methods would not provide a suitable basis for this surveillance.</li><li>4.A review of methods was undertaken, including a national questionnaire on methods, approaches to testing and risk factors for GRE bacteraemia. This, together with data from routine laboratory reports and special surveys, revealed the following.<li>–A significant proportion of enterococci are not identified to species level.</li><li>–There are problems with identification of enterococci. Commercially available kits do not identify some enterococcal species reliably.</li><li>–Not all laboratories test vancomycin susceptibility of enterococci from bacteraemia. This does not allow a robust assessment of GRE bacteraemia rates because the prevalence of glycopeptide resistance will be underestimated if laboratories test teicoplanin alone or no glycopeptides against enterococci.</li><li>–There are technical difficulties in detecting non-VanA resistance in enterococci and this will also contribute to under-reporting of glycopeptide resistance.</li></li><li>5.Clinically significant GRE infection is associated with a worse prognosis than infection with susceptible strains, but this factor has not been clearly separated from the possible influence of co-morbidities and antibiotic usage.</li><li>6.GRE bacteraemia occurs mainly on specialist units, particularly transplantation, renal, haematological malignancy and intensive care units. Hospitals with such units will inevitably have a higher risk of featuring prominently in surveillance studies. Hence, meaningful denominators for surveillance of GRE bacteraemia are required.</li></ul> <h3>Recommendations for GRE bacteraemia surveillance</h3> <ul><li>1.As the mandatory surveillance of glycopeptide-resistant enterococci (GRE) bacteraemias has already started, this report recommends a phased approach to some of the developments, such as using meaningful denominators for the bacteraemia rates.</li><li>2.Rates of GRE bacteraemias in trusts should not be used as a performance indicator because the numbers are too small for valid analysis in this way.</li><li>3.GRE bacteraemias as a proportion of all clinically significant bacteraemias in the trust should be measured as an indicator of changing trends.</li><li>4.Since most GRE are likely to be associated with specialist units, reports of GRE bacteraemia should indicate the specialty in which the patient acquired the infection.</li><li>5.Data on clinical activity in the special units where patients acquired GRE bacteraemias should be obtained from trusts with more than five GRE bacteraemias in a year.</li><li>6.The significance of blood cultures containing GRE should be assessed clinically. If the bacteraemia that includes GRE is deemed to be clinically significant (according to the case definitions for bacteraemia), whether mono- or polymicrobial, it should be reported as a GRE bacteraemia.</li><li>7.Enterococci from blood cultures should be identified to species level to provide data on resistance rates in different species.</li><li>8.Biochemical identification should be augmented by antimicrobial susceptibility patterns. The identification is suspect if Enterococcus faecalis appears to be ampicillin resistant and/or quinupristin/dalfopristin susceptible, or if Enterococcus faecium appears to be ampicillin susceptible and/or quinupristin/dalfopristin resistant. The identification and susceptibility of such isolates should be checked and, if confirmed, the isolate should be sent to a reference laboratory for further investigation.</li><li>9.Enterococci from blood cultures should be tested for susceptibility to vancomycin. Teicoplanin is not an acceptable alternative to vancomycin for these purposes.</li><li>10.Standardized methods should be used for glycopeptide susceptibility testing of enterococci, and the technical requirements of the methods must be adhered to.</li><li>11.The situation should be reviewed after a year and the methodology should be amended as appropriate.</li></ul> <h3>Recommendations for further research</h3> <ul><li>1.A group should be established to explore methods of data collection for global and individual risk factors for GRE bacteraemia.</li><li>2.Further studies on the performance of commercial identification kits with a range of enterococcal species are necessary.</li><li>3.Further studies to improve the reliability of susceptibility testing methods for detection of GRE are necessary.</li></ul>

Antibiotics and gastrointestinal colonization by vancomycin-resistant enterococci

Although several classes of antimicrobial agents have been associated with colonization or infection with glycopeptide-resistant enterococci (GRE) in individual clinical studies, the agents most commonly implicated are extended-spectrum cephalosporins and compounds with potent activity against anaerobic bacteria, including ticarcillin-clavulanic acid. In some clinical studies, formulary alterations designed to minimize the use of extended-spectrum cephalosporins or ticarcillin-clavulanic acid have resulted in significant decreases in colonization and infection by GRE. Experimental data using a mouse model of GRE gastrointestinal colonization indicate that persistence of high-level GRE colonization of the mouse gastrointestinal tract is promoted by exposure to agents with potent activity against anaerobic bacteria, suggesting that reduction of competing flora is the major factor leading to persistence of high-level colonization. One study performed in humans is consistent with this model and suggests that high levels of colonization may promote spread of resistant organisms in the nosocomial setting. Establishing colonization with GRE in uncolonized mice correlates with exposure to agents that are (a) secreted into the bile in significant concentrations and (b) have negligible activity against the colonizing enterococcal strain. Differences between piperacillin-tazobactam and ceftriaxone in the establishment model can be attributed directly to differences in their anti-enterococcal activity. Modification of antimicrobial prescribing practices may play an important role in facilitating successful infection control efforts to limit GRE in the nosocomial setting.

Vancomycin-resistant enterococci (VRE) in meat and environmental samples

We investigated the spread of vancomycin-resistant enterococci (VRE) in strains from meat and environmental samples and the location of glycopeptide-resistance determinants in VanA isolates. VRE and VSE (vancomycin-sensitive enterococci) resistance patterns to six antimicrobials were also evaluated. A total of 59 meat isolates (35%) and 119 environmental isolates (26.5%) were glycopeptide resistant enterococci. In particular, 10.7% meat isolates belonged to the VanA, 8.3% to VanB and 16% to VanC phenotypes. Environmental samples presented 0.7% VanA, 14.5% VanB, and 11.4% VanC strains. Evident differences were not observed among the resistance patterns of VRE and VSE isolates. Neither an important difference was observed comparing the resistance patterns in enterococci from meat and environment. In particular a low incidence of β-lactamic resistant strains was found, whereas high rates of resistance were observed for streptomycin (85.7% and 92.8%), kanamycin (79.7% and 96%) and gentamycin (85.1% and 91.7%). An intermediate rate of resistant bacteria emerged for erythromycin (35.1% and 10.5%). All VanA isolates independent of origin had more plasmids with different molecular weights. PCR amplification of the 732 bp fragment in plasmids from the VanA strains confirmed affiliation to the vanA gene cluster and the extrachromosomal location of the glycopeptide-resistance determinants. Our study suggests that food and environment play a potential role as reservoirs of resistance determinants, prompting the need to undertake epidemiological and molecular studies to evaluate the mobility of these genes.

Dutch Guideline for Preventing Nosocomial Transmission of Highly Resistant Microorganisms (HRMO)

Hospitals are faced with the increasingly rapid emergence and dissemination of antimicrobial-resistant microorganisms. US and European guidelines on the prevention of antimicrobial resistance in hospitals were, until recently, mainly directed at methicillin-resistant Staphylococcus aureus (MRSA). In 2004, the Dutch Working Party on Infection Prevention issued a guideline on the prevention of nosocomial transmission of highly resistant microorganisms (HRMO), in order to fulfill the growing need for additional guidance on the control of other pathogens with acquired resistance and the potential to spread within hospitals (such as glycopeptide-resistant Enterococcus faecium, penicillin-resistant Streptococcus pneumoniae, extendedspectrum beta-lactamase producing Enterobacteriaceae, and other (multi)drug-resistant gram-negatives). In addition to providing criteria for defining HRMO, the Dutch guideline provides recommendations on isolation of patients, active surveillance, and contact tracing. The guideline will enable the comparison of HRMO rates between hospitals, and may be used to evaluate the efficacy of programs to control antibiotic use and/or nosocomial transmission of resistant pathogens. The eventual success of nationwide implementation of this guideline remains to be established in the coming years.

Identification of Glycopeptide-resistant enterococci by VITEK 2 system and conventional and real-time polymerase chain reaction

Glycopeptide-resistant enterococci (GRE) are important causes of nosocomial infections in the United States and in Europe. Rapid detection of GRE is essential for the implementation of appropriate control measures to prevent the spread of GRE. In this study, we compared the reliability of 3 different methods, VITEK 2 automated system (bioMérieux), a conventional multiplex polymerase chain reaction (m-PCR) protocol, and a real-time PCR protocol performed on the LightCycler system (Roche) for identification of GRE in the routine microbiology laboratory. Species identification and glycopeptide resistance determination was tested with 80 enterococcal isolates with different glycopeptide resistance phenotypes. With the VITEK 2 system, 39% of the strains were correctly identified to species level. Resistance to vancomycin was detected in all isolates; however, discrepancies occurred in the correct detection of teicoplanin resistance. The PCR protocols proved to be suitable for detecting clinically relevant GRE; 90% of the isolates studied were correctly identified with the conventional m-PCR and 100% of vanA and vanB isolates with the real-time PCR protocol, respectively. High specificity and rapidity make the real-time PCR assays superior tools for identification of GRE in clinical samples; however, they do not have the ability to detect vanC.

VanB-VanC1Enterococcus gallinarum, Italy

To the Editor: We report detecting a vanB determinant in Enterococcus gallinarum in poultry in Italy. High-level vanA-mediated glycopeptide resistance has been described for E. gallinarum and E. casseliflavus (1–4), and vanB-mediated vancomycin resistance has been frequently described for E. faecalis and E. faecium. However, vanB-mediated resistance in isolates of E. gallinarum has been described only in sporadic nosocomial cases of infection or colonization (5,6). In January 2005, a study of contamination by foodborne organisms in slaughtered broiler carcasses was conducted in Sicily. To detect glycopeptide-resistant enterococci (GRE), each carcass was placed in a bag with 100 mL sterile buffered peptone water and shaken vigorously for 60 sec. After overnight incubation at 37°C, 0.5 mL rinsate was added in duplicate to 5 mL ethyl violet azide broth (Oxoid, Basingstoke, United Kingdom) with 4 mg/L vancomycin. Broth cultures were further incubated at 37°C for 48 h, and 0.1 mL aliquots were spread onto duplicate plates of VRE (commercial denomination product, Oxoid) agar. A vancomycin-resistant isolate of E. gallinarum was identified in a carcass from a broiler farm in eastern Sicily. The biochemical tests of API 20 Strep (bioMerieux, Marcy l'Etoile, France) and motility test at 30°C were used to characterize the isolate at the species level. The MICs of vancomycin and teicoplanin were 64 μg/mL and 1 μg/mL, respectively. The isolate was subjected to a multiplex polymerase chain reaction followed by an endonuclease cleavage of amplicons by MspI (Invitrogen, Carlsbad, CA, USA) as previously described (7) to detect van gene determinants; this process demonstrated a simultaneous presence of vanC1 and vanB determinants. E. gallinarum and the other motile enterococci are thought to infrequently cause infection. However, the recent involvement of vanC1-vanA E. gallinarum in person-to-person spread in a long-term-care facility (8) and in an intensive care unit (2), along with identification of vanC1-vanB isolates in some patients treated with prolonged courses of glycopeptides (5,6), suggests reassessment of their possible pathogenic role. For the first time, 1 isolate of E. gallinarum has been found harboring the vanB gene in poultry. Our findings confirm that E. gallinarum can capture the genetic determinants of high-level glycopeptide resistance, probably under selective pressure conditions that do not permit survival of a host organism with constitutive low-level resistance (3). Previous studies have demonstrated that E. gallinarum can transfer these determinants to E. faecium by conjugation (2). The role of food animals as reservoirs of GRE and the causes of their persistently high prevalence in poultry carcasses in some European countries are being investigated (9). Moreover, the public health risk associated with consumer exposure to GRE when handling raw animal foods is poorly understood. In Europe, the food chain is thought to be the major source of GRE since avoparcin was used as a food additive for animals until the European Union ban in 1997. Previous studies in Italy showed that avoparcin withdrawal successfully reduced GRE contamination of poultry meat products (10). However, our finding, 7 years after the European Union ban, highlights that resistance genotypes in motile enterococci should be closely monitored (11).

A Survey of Antimicrobial Resistance in a Tertiary Referral Hospital in the United Arab Emirates

Published data on antimicrobial resistance are lacking for the United Arab Emirates (UAE). Knowledge of local patterns of resistance is essential to optimize guidelines for empirical antimicrobial treatment. Awareness of susceptibility patterns in other parts of the world may be important for determining empirical antimicrobial therapy for travelers or patients transferred from abroad. Antibiograms from unique isolates of common hospital and community bacteria are described in a tertiary referral hospital in the UAE, which had no antimicrobial restrictions and limited resources. Antimicrobial resistance rates were unpredictable and were high for Gram-negative organisms, such as Escherichia coli, Shigella sonnei, Campylobacter spp, and also Streptococcus pneumoniae. In contrast, methicillin-resistant Staphylococcus aureus (MRSA) comprised less than 5% of S. aureus isolates and glycopeptide-resistant enterococci (GRE) were not detected during the surveillance period. Our results provide data on antimicrobial resistance for the UAE and highlight the need for resources for surveillance of antimicrobial resistance.

Caracterización molecular de un brote por Enterococcus faecalis resistente a los glucopéptidos en una unidad de cuidados intensivos

Between January and August 2003 glycopeptide-resistant Enterococcus faecalis was isolated from eight patients admitted to the intensive care unit (ICU).Antibiotic susceptibility testing was performed by disk diffusion and the Etest, clonal relatedness of the isolates was studied by pulsed-field gel electrophoresis (PFGE), and the presence of vanA was investigated by PCR.All the isolates were vanA-positive and had an identical PFGE pattern, showing that an outbreak had occurred in our ICU.

Clonality AmongEnterococcus faeciumClinical Isolates

In the course of a survey to determine the epidemiology of enterococcal infections in Italy, a sudden increment, in a 1-year time, was noted in the number of glycopeptide resistant Enterococcus faecium isolated from different wards of the University Hospital in Rome, Italy. The isolates were characterized for clonal relatedness by comparing SmaI gel electropherotypes, presence of vancomycin-resistance genes, and expression of virulence factors. PFGE identified in a single pulsed type all the glycopeptide-resistant isolates but one. Resistance to high levels of aminoglycosides was expressed by these same isolates, which also included a majority of non biofilm-forming strains. Two esp gene-carrying strains were also identified in different PFGE types. Data indicates that a specific clone acquired, in the clinical setting, the genetic determinant for glycopeptide resistance, thus improving environmental adaptation and favoring its persistence and spread.