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Background:
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.
Objectives:
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.
Description:
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.
Importance
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)
Achievements
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.
Website:
http://www.mtc.ki.se/Mtchome/groups/Mollby/fair.htm
Coordinator
Roland
MÖLLBY
Karolinska
Institute
Theorells
väg 3
S-171
77 Stockholm
Tel.:
+46 87 287 155/ 8 345 704
Fax:
+46 8 331 547
E-mail:
roland.mollby@mtc.ki.se
Partners
- Huw TAYLOR
University of Brighton
Lewes Road
UK-BN2 4GJ Brighton
Tel.: +44 1273 64 22 88
Fax: +44 1273 64 22 85
E-mail: h.d.taylor@bton.ac.uk
- 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
E-mail: anicet@bio.ub.es
- Frank Møller AARESTRUP
Danish Veterinary Laboratory
Bülowsvej 27
DK-1790 Copenhagen V
Tel.: +45 35 30 01 00
Fax: +45 35 30 01 20
E-mail: faa@svs.dk
- Anders FRANKLIN
National Veterinary Institute
Ulls Väg 2b
S-750 07 Uppsala
Tel.: +46 18 67 40 00
Fax: +46 18 30 91 62
E-mail: anders.franklin@sva.se
- 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
E-mail: mamoreno@eucmax.sim.ucm.es
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