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Investigations into the poor recovery of sulphadimidine from medicated animal feeds have shown that irreversible drug-feed binding, and not drug degradation, is responsible for the poor recoveries. The experimental work involved the novel use of C-sulphadimidine in analytical studies and in autoradiography of C-sulphadimidine-bound feed. The latter showed that the drug was not bound preferentially to specific feed constituents but was widely distributed on nearly all the feed particles. Further work on sulphadimidine recovery from feeds demonstrated an inverse relationship between drug recovery and feed moisture content. The role of moisture in the binding mechanism was then considered, and experiments conducted on the adsorption of moisture by feeds showed that the rate controlling mechanism was diffusion. A hypothesis is presented in which sulphadimidine is partially dissolved by the moisture in the feed and the resulting solution then diffuses into the internal regions of the feed particles via pores and cracks in the constituent particles. The deep penetration of the drug into the feed prevents the drug from being recovered by the extraction solution. Experimental evidence was found to support this hypothesis. Experimental work also investigated the causes of poor recoveries of sulphadiazine, trimethoprim and dinitolmide.

Fasciola hepatica is responsible for substantial economic losses and animal welfare issues within the agricultural sector worldwide. The increasing incidence of fasciolosis, coupled with the emergence of flukicide resistance, makes vaccination an attractive alternative control strategy. Hidden antigens extracted from the gut of blood feeding parasites have proven to be excellent vaccine candidates against haematophagous parasites, most notably Haemonchus contortus and Rhipicephalus (Boophilus) microplus. Here, as a first step towards a prototype liverfluke vaccine an attempt to identify hidden gut antigens in F. hepatica was made. Proteomic analysis on extracts of adult F. hepatica was used to identify molecules exclusively found within the membrane-bound fraction including four proteases; cathepsin B2, legumain-2, a putative lysosomal pro-x-carboxypeptidase precursor and a saposin-like protein. Histological sections of adult F. hepatica were screened with a panel of 21 lectins to identify those with an affinity for glycoproteins on the parasite’s gut and to inform subsequent lectin affinity chromatography. Seven lectins showed affinity for the gut region, with peanut (PNA) and jacalin (JAC) lectins binding to glycoproteins on either the gastrodermal cells or gut lamellae, respectively. PNA and JAC were then used to purify glycoproteins from the crude S3 extract by affinity chromatography. The resultant fractions were separated by SDS-PAGE and the protein profiles analysed by mass spectrometry. The enriched lectin-binding fractions shared a number of proteins but one of note that was exclusively identified in the PNA-binding fraction was a cathepsin D-like aspartyl protease, which had not previously been studied in F. hepatica. The proteolytic activities of three somatic extracts of adult F. hepatica were therefore investigated. The ability of the respective fractions to digest haemoglobin, a potential food source, was measured in the presence/absence of class-specific enzyme inhibitors. These analyses confirmed the presence of cathepsin D-like aspartyl protease activity capable of digesting haemoglobin optimally at pH 2 - 2.5. Further characterisation of the cathepsin D-like aspartyl (FhCatD) protease revealed it to be highly conserved within trematodes, to be localized to the gastrodermis of immature (10 day) and adult fluke, and to be more highly expressed, at the RNA level, in the Newly Excysted Juveniles (NEJ) than adult stages. Western blot analysis of native somatic extracts, enriched lectin-binding fractions and recombinant FhCatD using antisera from naturally infected sheep, showed some recognition of the recombinant FhCatD but did not provide clear evidence that the cathepsin D is strongly antigenic during natural infection.

Avian influenza viruses are considered to be key contributors to the emergence of human influenza pandemics. While aquatic birds and ducks are the major reservoir for influenza viruses, they are typically resistant to the effects of viral infection, in contrast to the frequently severe disease observed in chickens. In order to understand whether differences in receptors might contribute to this observation, anatomical distribution of influenza virus receptors (sialic acid SAα2,3-Gal and SA α2,6-Gal) in key organs of both species was studied using lectin histochemistry with linkage specific lectins, and receptor binding assays with swine H1N1 (classical A/sw/Iowa/15/30) and avian H2N3 (A/mallard duck/England/7277/06) influenza viruses. Widespread presence of both SAα 2,6-Gal and SAα2,3-Gal receptors were found in all major organs examined in both chickens and ducks. Interestingly, the predominant receptor type in chicken tracheal epithelium (TE) was SAα2,6-Gal whereas SAα2,3-Gal receptors were most abundant in duck TE. Paradoxically, infection of primary cell cultures (duck and chicken lung cells and embryo fibroblasts) with the swine H1N1, the low pathogenicity avian H2N3, and a highly pathogenic H5N1 (A/turkey/England/50-92/91) virus resulted in more extensive and rapid cell death in duck cells than in chicken cells. Infected duck cells displayed morphological features of apoptosis, increased DNA fragmentation and activation of caspase-3/7. Infected duck cells produced comparable levels of viral RNA but less infectious virus than infected chicken cells. Notably, such rapid cell death was not observed in duck cells infected with a contemporary Eurasian lineage H5N1 virus (A/turkey/Turkey/1/05) which has been shown to be fatal to ducks. Gene expression profiling of infected chicken and duck cells, 24hrs post-infection, with a chicken Affymetrix microarray platform revealed differential transcription of many genes between the two avian species. In particular, the array results suggested a possible role of BCoR, HSPA-9, STAT-3, AVEN, BCLAF1, IL-18, IFN-α, and TNF-α genes in mediating the contrasting species phenotypic response to influenza infections. In summary, rapid cell death in duck cells, mediated at least in part by apoptosis, results in reduced infective virus production and may well be an important protective host response of resistant ducks. By contrast, longer surviving infected chicken cells produce much higher infective virus load along with high levels of pro-inflammatory cytokines which could account for the susceptibility of chickens to influenza infections.