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<OL> <LI> Complete a comprehensive literature review of pre, pro and synbiotics used in animals
<LI> Determination of the inhibitory spectrum of probiotics against multi-resistant S. typhimurium in
co-culture studies
<LI> In vitro batch experiments using mixed culture inocula containing probiotics and prebiotics
<LI> Continuous culture model systems with synbiotics – salmonellae challenge experiments
<LI> Development of analytical tools: Development of antimicrobial resistance gene probes for in
vitro and in vivo real time studies
<LI> Verification of the reduction of penta-resistant S. typhimurium in the pig model by treatment
with pre and probiotic
<LI> Dissemination of information
Progress: The removal of antibiotics from feed stuffs is regarded as a major control measure that should lead to the reduction of the carriage by farmed animals of organisms that harbour resistance to antimicrobials. Whilst there is supportive evidence that removal equates with reduction in resistance, there is an increasing body of evidence that indicates resistance to antimicrobials is retained without antimicrobial selective pressure. Indeed, resistance is present in organically produced animals where there is no use of antimicrobials other than those used sparingly for chemotherapeutic purposes (data from OD2006, Pleydell, VLA). <P>
There are hypotheses that may explain the retention of resistance such as linkage of gene encoding fitness factors to resistance genes, co-selection through dietary factors, application of heavy metals and linkage of heavy metal resistance genes and so forth. Whilst the underlying cause of retention of resistance requires investigation, there is a more pressing need to adopt simple, economically sound and farm friendly measures to reduce the burden of resistance. In order to achieve this goal, this study will investigate potential approaches and one approach in depth experimentally in a model that focuses upon the pig that currently is associated with high levels of infecion with multi-resistant Salmonella enterica serotype Typhimurium.
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A major public health objective is the reduction, preferably elimination, of antimicrobial resistance entering the food chain from farmed animal species. This study will focus on simple dietary modifications including the use of pre, pro and syn-biotics that will be aimed at modulating the bacterial flora of the host animal. So far pre, pro and syn-biotics have been used to achieve two general goals in animal health and welfare which may be summarised as protection from gastro-intestinal pathogens and enhanced 'thrift'. Thus far, there has been no serious evalutaion of the effect of pre, pro and syn-biotics in the reduction of resistance carried by the many bacterial species in animal hosts. <P>The expected benefits of this study will be the generation of scientifically sound evidence describing the potential of pre, pro and syn-biotics to contribute to the reduction of resistance in farmed animals. Additional outputs will be the presentation of such information to a wider audience (incl. farmers) and simple recommendations on the use of these agents in conjunction with other control measures (vaccine and biosecurity for pathogen control) for sustainable production. <P>To achieve this goal, tools will be developed for the rapid analysis of resistance genes in gut microbial populations. These will be of wider use for subsequent monitoring and assessment of resistance genes in animals. <P>The scientific literature gives many examples of the use of pre, pro and syn-biotics for use in reducing colonisation of enteric pathogens and for host innate immunity modulation but there is very little information on the use of these agents for possible control of resistance carrying pathogens or commensals. <P>The first part of the study will be a desk exercise to define the range of products used in animals, and for competitive exclusion agents define the targets against which they have been used. This review will focus on the potential application of these agents for control of resistance and will identify those agents that are confirmed free of exogenous resistance genes.The desk exercise will not be limited to just pre, pro and syn-biotics but consider other potentially synergistic interventions such as use of lysins and lytic bacteriophages. <P> A key objective of the study is to gain definitive scientific evidence that demonstrates the effect of pre, pro and syn-biotics upon carriage of resistance in animals. To do this appropriate tools are needed which can rapidly assess the type of resistance genes carried in an entire bacterial gut population and also specifically locate resistance genes to bacterial hosts in situ. A nano-array platform is in the final stages of development in project VMO2136 which will be used for screening isolates for resistance genes (VLA). It is intended to further develop this tool to detect resistance genes in bacterial populations and complex matrices such as luminal contents and faeces. Fluorescent In Situ Hybridisation (FISH) methods have been developed (Reading University) for the detection and quantification of specific species in complex matrices. This technology will be exploited also and further developed to detect specific antibiotic resistance genes [a limited number can be screened simultaneously]. <P>Once the tools for resistance gene detection have been constructed and validated, these will be used to detect and quantify resistance in model systems in which pre, pro and syn-biotics are used. Two model systems will be studied. The first is the artificial gut fermentation system developed by Glen Gibson at Reading University. The system will be primed with pig derived gut flora and stabilised. The inoculum will be tested for resistance profile and spiked with pig derived multi-resistant Salmonella enteric serotype Typhimurium as required. Resistance genes will be monitored in the bacterial flora with and without use of pre, pro and syn-biotic combinations. <P>The second model will be mini pigs (snatched at birth/colustrum deprived) inoculated with the same flora and treated with or without pre, pro and syn-biotics. Faeces will be monitored (as will gut content by cannulation; subject to ethical approval and HO licence). The number of combinations of treatment trialed in these models will be limited by budgetary constraints. The availability of the models will also permit more specific studies on the effect of pre, pro and syn-biotics on the reduction of targeted pathogens irrespective of carriage of resistance, such as Salmonella and C. coli/jejuni and this will be considered within the brief of this study subject to budgetary constraints. <P> The findings of these studies will be collated and peer reviewed publications will be prepared and reports and/or presentations for Defra, the industry and probiotic/pharmaceutical companies.
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