<OL> <LI> To set up model systems to simulate storage, treatment and spreading of organic wastes
<LI> To use marked antimicrobial resistant organisms with known plasmid borne or chromosomal genetic resistance determinants as donors in the above waste processing models
<LI> To use marked antimicrobial sensitive organisms to act as recipients in artificially contaminated and natural organic wastes
<LI> To quantify the frequency of transfer of resistance and the resulting levels of resistant organisms in organic wastes
<LI> To develop quantitative risk assessment (QRA) models to describe the risk of transfer of antimicrobial resistance genes between different bacteria in stored and spread farm wastes
<LI> To assess the potential for survival of naked DNA after death of the organisms and transfer of antimicrobial resistance genes to other organisms in stored and treated wastes
Progress: Main findings of the research: <BR>
No conjugation occurred for all the organisms tested, under all the experimental conditions described within poultry, cattle and pig faecal waste laboratory storage models. No difference was found between sterile and untreated fresh waste models, which implies that chemical composition and nutrient deficient conditions may be the cause of inhibition rather than competition by resident flora. <BR> <BR>
Similarly, there was no conjugation for all the organisms tested, under all the experimental conditions described within cattle and pig farm-scale storage models. In the case of the poultry farm-scale models, conjugation was detected after 5 and 24 h, however the organisms were non-detectable after 48 h probably due to the rapid increase of the temperature in
the manure heaps. <BR> <BR>
Conjugation was detected after 24 h and up to 7 days in the laboratory models simulating spread of all 3 waste types.
No conjugation was detected from the field-scale model experiments in March, however the second experiments in June did show that conjugation was possible. This difference may imply that temperature as well as other climatic conditions may influence the outcome of conjugation in field conditions. We may conclude that warmer temperatures in the summer
season favoured the transfer of the resistance determinants by conjugation..
<BR> <BR> In summary, results from our experiments suggest that stored slurry is not a good environment for conjugation.
However, when applied to pasture, animal manure could provide a favourable environment for exchange of genetic
material between bacteria, facilitating the spread of antimicrobial resistance. <BR> <BR>
Studies on transfer of naked DNA:
The naked DNA experiments have not yielded positive results under the experimental conditions so far employed. We
therefore consider that on the basis of results so far achieved, the uptake of naked DNA by pathogenic bacteria in stored
organic wastes is unlikely. Annex 10 provides the full details on these experiments.
<BR> <BR> Treatment models:
Initially, models simulating stirring tanks, anaerobic or aerobic digestion, lime addition were thought to be adequate for
this section of the project. However, most farms would struggle to finance treatments of this sort. Most farms only have 1
slurry store and this store is added to on a continual basis. As a consequence a more realistic approach was followed.
Storage without extra additions is the cheapest and most pragmatic way to treat pathogen-containing manures. What we
have done is to assess the effectiveness of a very cheap treatment method for manures by following long-term storage up
to 96 days. In spite of the presence of “natural” transferable elements in the waste, and of the external source of donor
strains added in the model, no transfer of resistance determinants to our recipient strains could be detected. <BR> <BR>
Using the data from the experimental work carried out at the Veterinary Laboratories Agency and the Health Protection
Agency, risk assessment models were developed to estimate the risk of antimicrobial resistance gene transfer in stored
and spread farm waste. Each model describes the storage and spreading practices of farm waste and investigates the
impact of such practices on the transfer of antimicrobial resistance genes. The models predict that the risk of
antimicrobial resistance gene transfer in stored and spread waste is low, and that the risk to livestock grazing on land
spread with farm waste is equally low. However, the models indicated that this risk could be reduced further by, for
example, increasing the time between the last spreading of waste and the time of turnout.
<BR> <BR> Prime options for new work:
Having demonstrated some risk in association with spread waste a wider survey of the potential of recipient organisms
to acquire resistance from freshly spread waste and waste that has been left for the recommended period before grazing.
The risk of uptake of such organisms by wildlife and contamination of salad crops should also be determined. The
question remains what is the risk for transfer of resistance once the organisms have been acquired by grazing animals in
the field. A collaborative project with Bristol University has been proposed looking at transfer or resistance genes in
Salmonella in vivo pig models. The experience gathered using the strains in OD2008 will be invaluable for the new
proposed work. Similar controlled studies should also be carried out in grazing animals. Additional work should also be
carried out to determine whether antimicrobial pollution in the environment, particularly in conditions simulating
aquaculture in the Far East, may enhance transfer of antimicrobial resistance and linked virulence genes.