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Epidemiology and ecology of enterococci, with special reference to antibiotic resistant strains, in animals, humans and the environment

Contract nr: FAIR-CT97-3709
Project nr: 3709
Project type: SC
Starting date: 01/01/1998
Duration: 42 months
Total cost: 1,226,584 EUR
EC Contribution: 924,000 EUR
Scientific Officer: Isabel Minguez-Tudela
Rtesearch topic: Animal health

Enterococci are bacteria that are normal inhabitants of the intestinal floras of humans and warm blooded animals. As such, they are released into the environment by animal waste and fertilizers of animal origin. In contrast to coliforms and other intestinal indicator bacteria, the enterococci are rather tough and can survive for long periods of time in soil and water, and thus re-enter the food chain. Enterococci that are resistant to all presently available antibiotics have started to appear, possibly due to the use of drugs such as avoparcine in agriculture. This is regarded as very alarming. Due to the high risk that resistant enterococci or resistance genes will spread via the food chain to humans from animal waste and natural fertilizers, we need to know more about the epidemiology and ecology of both normal and resistant enterococci.

The overall objectives of the present proposal are to generate knowledge of the ecological and epidemiological role of enterococci in the food chain, and of their possible threat to human health due to resistance development and transfer of resistance.
The project will involve studies on:

1) the population structure of enterococci from farm land, animal feed, animals, animal carcasses, vegetables, humans, and sewage in different areas within the EC;
2) the antibiotic resistance patterns of enterococci from the above sources;
3) the population structure of antibiotic resistant enterococci versus non-resistant strains; and
4) the diversity and the epidemiology of resistance genes and the resistance mechanisms among enterococci.


Working plan
Samples from different reservoirs in the food chain (natural and artificial fertilised soil, sewage, water), animal fodder, animal faecal material, food (meat and vegetables), as well as human faecal materials (healthy individuals and hospitalised patients) are screened for both normal and highly resistant enterococcal populations. The enterococci from all samples are first characterised by a simple phenotyping method (the PhP system) to measure the diversity of enterococcal populations in individual samples and the similarities between related samples. Isolates representing dominating and/or vancomycin resistant phenotypes are saved and characterised centrally by genotyping (PFGE, RAPD) to find out their clonal relations; species are thereby identified (biochemical methods and PCR techniques), and their resistance patterns are evaluated (MIC method). In vancomycin-resistant isolates the resistance gene is characterised.

Our investigations will result in knowledge of the diversity and the dynamics of both drug-resistant and normal populations of enterococci from different environments. This knowledge will also result in a better understanding of the fate of all bacteria released into the environment by animal waste and fertilizers, their further fate in the food chain, and the dangers of antibiotic use in agriculture. The knowledge may thus serve as a basis for future actions regarding the policy of the antibiotic use and the treatment of animal waste within the EU. The knowledge can also serve as a basis for the current discussions on using the enterococci as indicators of faecal pollutions.

Current situation/results:


State of progress
During the project second year, the main tasks have been sampling and preliminary phenotyping of enterococci, using the protocol that was created during the project first year. The sampling part of the project is now almost finished. The common culture collection of enterococcal strains is in the process of being built up. Some analysis on the saved isolates have also been initiated (MIC determination, species identification, PFGE typing)

The PhenePlate system has been set up for large-scale typing of enterococci. The software has been installed in the involved laboratories. The project has also resulted in the development of improved methods for identification and typing of enterococci, for resistance gene characterisation, and a protocol for MIC determination of enterococci.
Samples have been collected according to schedule, and during the project second year, a total of 751 samples has been collected. Thus, including the samples collected during the first project year, we now have data on 1,527 samples.

Presumed enterococci were found in 1,148 out of the 1,527 samples analysed (75%). Presumed enterococcal isolates resistant to 8 mg of vancomycin have been isolated in 178 out of the 1,527 samples (12 %). In most samples vancomycin resistant enterococci were only found after enrichment in liquid medium containing vancomycin. Most of these isolates have are still to be assayed for resistance towards 20 mg/l vancomycin.
About 16,000 isolates from the project (representing the total enterococcal floras as well as antibiotic resistant floras) have so far been subject to PhP typing with the PhP-RF plate. In addition some 4,500 enterococcal isolates from related projects have been subject to PhP-typing, and thus our database now consists of typing data on more than 20,000 isolates.


Karolinska Institute
Theorells väg 3
S-171 77 Stockholm
Tel.: +46 87 287 155/ 8 345 704
Fax: +46 8 331 547


  • Huw TAYLOR
    University of Brighton
    Lewes Road
    UK-BN2 4GJ Brighton
    Tel.: +44 1273 64 22 88
    Fax: +44 1273 64 22 85

  • Anicet BLANCH
    Universidad de Barcelona
    Gran Via de les Corts Catalanes 585
    E-08071 Barcelona
    Tel.: +34 93 402 90 12
    Fax: +34 93 411 05 92

  • Frank Møller AARESTRUP
    Danish Veterinary Laboratory
    Bülowsvej 27
    DK-1790 Copenhagen V
    Tel.: +45 35 30 01 00
    Fax: +45 35 30 01 20

  • Anders FRANKLIN
    National Veterinary Institute
    Ulls Väg 2b
    S-750 07 Uppsala
    Tel.: +46 18 67 40 00
    Fax: +46 18 30 91 62

  • Miguel Angel MORENO
    Complutense University of Madrid
    Department of Animal Pathology I (Animal Health),
    Avenida de Puerta de Hierro, s/n,
    E-28040 Madrid
    Tel.: +34 91 394 3721
    Fax: +34 91 394 3908

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