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Ingo Klare

Publications and source records attributed to Ingo Klare.

17 recordsLinked to original sources
PubMed2006

Molecular epidemiology of hospital-acquired vancomycin-resistant enterococci.

Vancomycin-resistant Enterococcus faecium strains are a significant cause of nosocomial infections in predisposed patients. Multiple-locus variable-number tandem repeat analysis (MLVA) has been validated recently by use of a global strain collection. In this report, we applied MLVA together with multilocus sequence typing (MLST) and pulsed-field gel electrophoresis (PFGE) to type 14 isolates from three clusters of patients colonized or infected with vancomycin-resistant Enterococcus faecium and another 10 epidemiologically unrelated isolates from the same hospital. The clusters could be distinguished by all three typing methods, which proved to be concordant. PFGE patterns provided the highest resolution. We observed seven sequence types (ST), six MLVA types (MT), and nine distinct ST/MT combinations. The combination of MLST and MLVA may be an alternative to PFGE in hospital epidemiology, providing the benefits of high accuracy, reproducibility, and portability.

Bacterial Typing Techniques↗
PubMed2005

Outbreak of vancomycin-resistant enterococci (VRE) in a hematological oncology ward and hygienic preventive measures. A long-term study.

BACKGROUND: We report on two endemic outbreaks of VRE (vancomycin-resistant enterococci, vanA type Enterococcus faecium) on a hematological oncology ward in a university hospital and a new strategic concept to fight against spread of VRE. PATIENTS AND METHODS: During management of the outbreak, a total of 1,124 patients and 1,700 specimens were investigated from June 1999 to December 2001. Protective measures were instituted. Intensive prophylactic infection control was practiced until December 2003. RESULTS: Of the 1,124 patients, 44 were VRE-positive. From 1,700 environment specimens, 110 VRE isolates were obtained from the inanimate environment, 5 from the living environment. Molecular biological typing identified 5 different clones which had spread on the ward in different locations. CONCLUSION: The study shows the need for prophylactic microbiological screening investigations in the risk group of hematological oncology patients and consistent implementation of protective hygiene measures to prevent the spread of multiresistant infectious pathogens on risk wards. It is also shown that intervention can to control such an endemic outbreak.

Anti-Bacterial Agents↗
PubMed2005

How many infections are caused by patient-to-patient transmission in intensive care units?

OBJECTIVE: The proportion of intensive care unit (ICU)-acquired infections that are a consequence of nosocomial cross-transmission between patients in tertiary ICUs is unknown. Such information would be useful for the implementation of appropriate infection control measures. DESIGN: A prospective cohort study during 18 months. SETTING: Five ICUs from two university hospitals. PATIENTS: All patients admitted for >/=48 hrs. MEASUREMENT: ICU-acquired infections were ascertained during daily bedside patient and chart reviews. Episodes of potential cross-transmission were identified by highly discriminating genetic typing of all clinical and surveillance isolates of the ten bacterial species most frequently associated with nosocomial infections in ICUs. Isolation of indistinguishable isolates in two or more patients defined potential transmission episodes. MAIN RESULTS: During 28,498 patient days, 431 ICU-acquired infections and 141 episodes of nosocomial transmissions were identified. A total of 278 infections were caused by the ten species that were genotyped, and 41 of these (14.5%) could be associated with transmissions between patients. CONCLUSION: Infections acquired during treatment in modern tertiary ICUs are common, but a causative role of direct patient-to-patient transmission can only be ascertained for a minority of these infections on the basis of routine microbiological investigations.

Bacterial Infections↗
PubMed2005

Evaluation of new broth media for microdilution antibiotic susceptibility testing of Lactobacilli, Pediococci, Lactococci, and Bifidobacteria.

Nine pure or mixed broth media were evaluated for their suitabilities to determine MICs in a microdilution test of 19 antibacterial agents for lactic acid bacteria (LAB) of the genera Lactobacillus, Pediococcus, Lactococcus, and Bifidobacterium. A mixed formulation of Iso-Sensitest broth (90%) and deMan-Rogosa-Sharpe broth (10%) with or without supplementation with L-cysteine, referred to as the LAB susceptibility test medium, provided the most optimal medium basis in terms of growth support of nonenterococcal LAB and correct indication of MICs of international control strains.

Anti-Bacterial Agents↗
PubMed2004

Conjugative transfer of the virulence gene, esp, among isolates of Enterococcus faecium and Enterococcus faecalis.

OBJECTIVES: The enterococcal surface protein gene, esp, is a major putative pathogenicity marker in clinical isolates of Enterococcus faecium and Enterococcus faecalis. This study demonstrates in vitro conjugative transfer of the esp gene among E. faecium and E. faecalis. MATERIALS AND METHODS: Enterococcal isolates from clinical samples, positive for esp, were mated on filters with enterococcal recipients. Transconjugants were checked for transfer of antibiotic resistance determinants and co-mobilization of the esp gene. They were also characterized by PCR and plasmid profiling/PFGE typing including Southern hybridizations with labelled esp probes. Transfer as triggered by excision was tested using Taqman PCR. RESULTS: Two of five E. faecalis and five of nine E. faecium transferred antibiotic resistance determinants into a recipient. Of the transconjugants analysed by PCR for acquisition of esp, only isolates from two E. faecalis and a single E. faecium mating were positive. In the donor strains, the esp gene was located on the chromosome. Molecular analysis revealed a plasmid localization of esp in the E. faecium transconjugant and chromosome-to-chromosome transfer in E. faecalis. CONCLUSION: The esp gene is transferable by conjugation among enterococcal isolates.

Conjugation, Genetic↗
PubMed2004

Occurrence of antibiotic-resistant enterobacteria in agricultural foodstuffs.

Antibiotic-resistant bacteria or their corresponding resistance determinants are known to spread from animals to humans via the food chain. We screened 20 vegetable foods for antibiotic-resistant coliform bacteria and enterococci. Isolates were directly selected on antibiotic-containing selective agar (color detection). Thirteen "common vegetables" (tomato, mushrooms, salad) possessed 10(4)-10(7) cfu/g vegetable of coliform bacteria including only few antibiotic-resistant variants (0-10(5) cfu/g). All seven sprout samples showed a some orders of magnitude higher contamination with coliform bacteria (10(7)-10(9) cfu/g) including a remarkable amount of resistant isolates (up to 10(7) cfu/g). Multiple resistances (up to 9) in single isolates were more common in sprout isolates. Resistant bacteria did not originate from sprout seeds. The most common genera among 92 isolates were: 25 Enterobacter spp. (19 E. cloacae), 22 Citrobacter spp. (8 C. freundii), and 21 Klebsiella spp. (9 K. pneumoniae). Most common resistance phenotypes were: tetracycline (43%), streptomycin (37%), kanamycin (26%), chloramphenicol (29%), co-trimoxazol (9%), and gentamicin (4%). The four gentamicin-resistant isolates were investigated in molecular details. Only three (chloramphenicol) resistant, typical plant-associated enterococci were isolated from overnight enrichment cultures. In conclusion, a contribution of sprouts contaminated with multiresistant, Gram-negative enterobacteria to a common gene pool among human commensal and pathogenic bacteria cannot be excluded.

Chloramphenicol Resistance↗
PubMed2004

Two episodes of vancomycin-resistant Enterococcus faecium outbreaks caused by two genetically different clones in a newborn intensive care unit.

In 2001 two outbreak episodes (January-March and June-July) caused by vancomycin-resistant E. faecium (VRE) of the VanA-type were observed at a neonatal intensive care unit (NICU) of a university hospital in south-west Germany. To identify the initial source and the route of transmission environmental samples were examined as well as stool samples from patients and the staff. VRE was not found in environmental samples. However, stool samples from 24 hospitalised children tested positive and bacterial clonality was assessed by Sma1-based macro restriction analysis. Furthermore, esp gene and vancomycin resistance gene carriage were examined as well as bacteriocin production. PCR analysis showed that all 24 isolates carried vanA gene cluster, encoding resistance to vancomycin and teicoplanin. However, five of the vanA-positive isolates were resistant to vancomycin but not to teicoplanin. Only these five isolates produced bacteriocin, but in none of the isolates esp gene was detected. PFGE revealed that both outbreaks were caused by two different clones. The patient initiating the first episode, was identified whereas the origin of the second episode remained unknown. From one of the 40 staff stool samples VRE was isolated. This strain was related to the clone of the summer outbreak. In conclusion there were two independent episodes of self limiting VRE outbreaks and transmission on the ward is highly probable.

Bacterial Proteins↗
PubMed2004

Molecular detection of linezolid resistance in Enterococcus faecium and Enterococcus faecalis by use of 5' nuclease real-time PCR compared to a modified classical approach.

A nucleotide transversion from guanine to uracil in the 23S rRNA confers linezolid resistance. We describe a real-time PCR using two Taqman probes that detects a single mutated allele among the genomes of Enterococcus faecium and Enterococcus faecalis. Results were confirmed by a classical approach involving LabChip technology assayed with an Agilent Bioanalyzer 2100.

Acetamides↗
PubMed2003

Occurrence and spread of antibiotic resistances in Enterococcus faecium.

Enterococci are the second to third most important bacterial genus in hospital infections. Especially Enterococcus (E.) faecium possesses a broad spectrum of natural and acquired antibiotic resistances which are presented in detail in this paper. From medical point of view, the transferable resistances to glycopeptides (e.g., vancomycin, VAN, or teicoplanin, TPL) and streptogramins (e.g., quinupristin/dalfopristin, Q/D) in enterococci are of special interest. The VanA type of enterococcal glycopeptide resistance is the most important one (VAN-r, TPL-r); its main reservoir is E. faecium. Glycopeptide-resistant E. faecium (GREF) can be found in hospitals and outside of them, namely in European commercial animal husbandry in which the glycopeptide avoparcin (AVO) was used as growth promoter in the past. There are identical types of the vanA gene clusters in enterococci from different ecological origins (faecal samples of animals, animal feed, patients in hospitals, persons in the community, waste water samples). Obviously, across the food chain (by GREF-contaminated meat products), these multiple-resistant bacteria or their vanA gene clusters can reach humans. In hospital infections, widespread epidemic-virulent E. faecium isolates of the same clone with or without glycopeptide resistance can occur; these strains often harbour different plasmids and the esp gene. This indicates that hospital-adapted epidemic-virulent E. faecium strains have picked up the vanA gene cluster after they were already widely spread. The streptogramin virginiamycin was also used as feed additive in commercial animal husbandry in Europe for more than 20 years, and it created reservoirs for streptogramin-resistant E. faecium (SREF). In 1998/1999, SREF could be isolated in Germany from waste water of sewage treatment plants, from faecal samples and meat products of animals that were fed virginiamycin (cross resistance to Q/D), from stools of humans in the community, and from clinical samples. These isolations of SREF occurred in a time before the streptogramin combination Q/D was introduced for therapeutic purposes in German hospitals in May 2000, while other streptogramins were not used in German clinics. This seems to indicate that the origin of these SREF or their streptogramin resistance gene(s) originated from other sources outside the hospitals, probably from commercial animal husbandry. In order to prevent the dissemination of multiple antibiotic-resistant enterococci or their transferable resistance genes, a prudent use of antibiotics is necessary in human and veterinary medicine, and in animal husbandry.

Animal Husbandry↗
PubMed2003

Intra-hospital dissemination of quinupristin/dalfopristin- and vancomycin-resistant Enterococcus faecium in a paediatric ward of a German hospital.

OBJECTIVES: To demonstrate nosocomial transmission of Enterococcus faecium resistant to quinupristin/dalfopristin and vancomycin/teicoplanin among paediatric patients in a German hospital ward. MATERIALS AND METHODS: Multiply-resistant E. faecium were isolated from three female patients aged 9 months, 2 and 15 years during a 10 day time span. Antibiotic susceptibilities were determined by microbroth dilution. Clonal relatedness among the isolates was investigated via SmaI-macrorestriction analysis by PFGE, multilocus sequence typing (MLST), and plasmid profiling. Presence of virulence and resistance determinants was tested by polymerase chain reaction (PCR). Selected resistance genes were localized by Southern hybridizations. RESULTS: A single E. faecium isolate per patient was investigated. All exhibited resistances to quinupristin/dalfopristin, vancomycin/teicoplanin, streptomycin (high-level), penicillin/ampicillin, erythromycin, oxytetracycline, chloramphenicol, rifampicin and fusidic acid. The isolates were susceptible to linezolid only and intermediately resistant to fluoroquinolones including moxifloxacin. PFGE revealed identical patterns for all three isolates. PCRs for virulence determinants hyaluronidase and enterococcal surface protein, esp, were negative, whereas PCR for the enterocin A gene was positive. MLST identified clonal type [8-5-1-1-1-1-1] belonging to a clonal subgroup C1 of hospital- and outbreak-related E. faecium. Southern hybridizations located several resistance genes (erm(B), vat(D), vanA) on a large plasmid, which was transferable in mating experiments with an E. faecium recipient. CONCLUSIONS: These data show routes of dissemination of resistance to multiple antibiotics including streptogramins and glycopeptides in E. faecium via vertical and/or horizontal gene transfer. The isolates spread in the absence of a direct selective pressure, as none of the patients had received earlier streptogramin or glycopeptide therapy.

Anti-Bacterial Agents↗
PubMed2003

A potential virulence gene, hylEfm, predominates in Enterococcus faecium of clinical origin.

An open reading frame (hyl(Efm)) with homologies to previously described hyaluronidase genes has been identified in nonstool isolates of Enterococcus faecium. E. faecium isolates (n=577) from diverse sources were screened for the presence of hyl(Efm) and esp(Efm), a putative virulence gene associated with epidemic E. faecium strains. The presence of esp(Efm) was roughly twice that of hyl(Efm), but both were found primarily in vancomycin-resistant E. faecium isolates in nonstool cultures obtained from patients hospitalized in the United States. These data suggest that specific E. faecium strains may be enriched in determinants that make them more likely to cause clinical infections. Differences in the prevalence of these strains may help explain variations in the clinical importance of multiresistant E. faecium across different continents.

Anti-Bacterial Agents↗
PubMed2003

Influence of transferable genetic determinants on the outcome of typing methods commonly used for Enterococcus faecium.

A variety of methods is used for a molecular typing of Enterococcus spp. and related gram-positive bacteria including macrorestriction analysis using pulsed-field gel electrophoresis (PFGE), ribotyping, rapid amplification of polymorphic DNA (RAPD), and amplified fragment length polymorphism (AFLP). To test the influence of transferable determinants on the outcome of different typing methods commonly used for enterococci, we established a homogenous strain collection of 24 transconjugants resulting from filter matings with antibiotic-resistant Enterococcus faecium. As expected, AFLP, RAPD, and PFGE all identified our model bacteria as strongly related. However, distinct differences in the resolving and discriminatory power of the tested methods could be clearly addressed. In PFGE, 22 of 24 transconjugants possessed less than a three-band difference to the recipient pattern and would be regarded as strongly related. Three different RAPD PCRs were tested; in two reactions, identical patterns for all transconjugants and the recipient were produced. One RAPD PCR produced an identical pattern for 18 transconjugants and the recipient and a clearly different pattern for the remaining 6 transconjugants due to a newly appearing fragment resulting from acquisition of the tetL gene. AFLP clusters all transconjugants into a group of major relatedness. Percent similarities were highly dependent on the method used for calculating the similarity coefficient (curve-based versus band-based similarity coefficient). Fragment patterns of digested plasmids showed the possession of nonidentical plasmids in most transconjugants. PFGE still could be recommended as the method of choice. Nevertheless, the more-modern AFLP approach produces patterns of comparable discriminatory power while possessing some advantages over PFGE (less-time-consuming internal standards). Plasmid fingerprints can be included to subdifferentiate enterococcal isolates possessing identical macrorestriction and PCR typing patterns.

Anti-Bacterial Agents↗
PubMed2002

Molecular analysis of streptogramin resistance in enterococci.

The new semi-synthetic streptogramin antibiotic combination quinupristin/dalfopristin (Synercid) is a promising alternative for a treatment of infections with multiple resistant gram-positive pathogens, e.g. glycopeptide- and multi-resistant Enterococcus faecium. Streptogramins consist of two unrelated compounds, a streptogramin A and B, which act synergistically when given in combination. Mechanisms conferring resistance against both components are essential for resistance against the combination in E. faecium. In this species resistance to streptogramin A compounds is mediated via related acetyltransferases VatD and VatE. Resistance against streptogramins B is either encoded by the widespread ermB gene cluster conferring resistance to macrolide-lincosamide-streptogramin B antibiotics or via expression of the vgbA gene, which encodes a staphylococcal-type lactonase. E. faecalis is intrinsically resistant to streptogramins. Due to a wide use of streptogramins (virginiamycins S/M) in commercial animal farming a reservoir of streptogramin-resistant E. faecium isolates had already been selected. Determinants for streptogramin resistance are localized on plasmids that can be transferred into an E. faecium recipient both in vitro in filter-matings and in vivo in the digestive tracts of rats. Hybridization and sequencing experiments revealed a linkage of resistance determinants for streptogramins A and B on definite plasmid fragments.

Anti-Bacterial Agents↗
PubMed1999

Identification of enterococci and determination of their glycopeptide resistance in German and Austrian clinical microbiology laboratories.

OBJECTIVE: To determine exactly how German and Austrian routine laboratories perform tests for the identification of enterococci and determination of their glycopoptide resistance. METHODS: Six enterococcal test strains with different types of glycopeptide resistance (Enterococcus faecium, VanA; E. faecalis, VanA; E. faecium, VanB; E. faecalis, VanB; E. gallinarum, VanC1; E. casseliflavus, VanC2) were sent as anonymous isolates to 73 clinical microbiology laboratories (65 in Germany; eight in Austria). The participating laboratories had to identify the strains up to the species level and to determine their antibiotic susceptibilities to the glycopeptides vancomycin and teicoplanin by the test method(s) that are used daily in the corresponding laboratories. RESULTS: The analysis of the results received from 62 laboratories (56 from Germany, six from Austria) demonstrated that the most used routine method in susceptibility testing was the agar diffusion test, followed by the Etest, and the microbroth dilution procedure. The majority of participants had no difficulties in susceptibility testing with the VanA-type strains. However, the agar diffusion test was often not able to recognize clearly the VanB and VanC strains; some problems also arose with VanB isolates in the Etest. With the microbroth dilution method, the corresponding type of glycopeptide resistance was correctly determined in the majority of enterococcal test strains. Difficulties also arose in identification, especially with the VanC strains (E. gallinarum and E. casseliflavus), which were often falsely identified as E. faecium. The reasons for these errors are obviously based on the lack of important tests (such as motility and presence of a yellow pigment) in some commercially available identification test kits.

Journal Article↗