Mastitis is a major disease in the modern dairy cow with respect to both cow welfare and the quality and quantity of milk produced. The dry period is increasingly being recognised as a critical time point within the lactation cycle for the control of mastitis-causing pathogens.

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The principal aims of the work presented in this thesis were to further investigate the pathophysiology of the acute phase proteins in bovine mastitis and to document the changes in the protein composition of milk during mastitis using modem proteomic methods. Mastitis is one of the most important diseases affecting cattle worldwide. It has adverse effects on the economics of milk production through reducing the quality and quantity of milk. Acute phase proteins (APPs) are valuable markers of inflammatory lesions and are widely used in the veterinary field for disease diagnosis. This study further investigated the presence of APPs, haptoglobin (Hp) 1 I and mammary- associated serum amyloid A3 (M-SAA3) in the milk of cows with mastitis and evaluated the possibility of using these proteins to detect mastitis. An Enzyme linked immunosorbent assay (ELISA) was developed and validated for the measurement of milk Hp. A commercially available ELISA was used for the detection ofM-SAA3 in milk. Both Hp and M-SAA3 showed similar kinetics to somatic cell counts and can be considered as sensitive and reliable markers of mastitis. Advanced proteomic methods were developed and used to further characterise the changes taking place in the proteins expressed in normal and mastitic milk. The methodology for analysis of the bovine milk proteome was successfully established. This part of the work concluded that the patterns of protein expression of clinically mastitic milk showed clear differences from that of normal milk. Further analysis of milk samples from a mild subclinical model of Staphylococcus aureus mastitis showed that alterations to the milk p:oteome were only minimal. However this study was useful in identifying areas worthy of future research. Recent publications on APP research speculate upon the local production ofM-SAA3 in the bovine mammary gland. The final objective of this study was therefore to investigate the expression of MSAA3 in bovine mammary tissue. Our studies confirm the local production of M-SAA3 in the bovine mammary gland and its up-regulation during bovine mastitis. This finding may provide further information on a possible role for this protein. Overall the findings detailed in this thesis indicate that APPs in milk are valuable markers of mastitis and additionally that the ease of collection of milk by non-invasive methods suggest that milk has potential for the discovery of disease biomarkers.

Mastitis is an important disease causing health, productivity and welfare problems in dairy cattle. Over the past 40 years, control programs have successfully revolved around prophylactic and therapeutic antibiotic usage. This has led to a substantial decrease in the prevalence of subclinical mastitis and the suggestion that antibiotic prophylaxis should be reduced. This thesis describes a study that compared the efficacy of a non-antibiotic internal teat sealer containing bismuth subnitrate to antibiotic dry cow therapy (DCT). In cows uninfected at drying off, the teat sealer was significantly better than antibiotic DCT at preventing new dry period intra-mammary infections caused by Escherichia coli, all Enterobacteriaceae species and all major pathogens. Animals that received antibiotic DCT suffered numerically more cases of clinical mastitis during the dry period and next lactation. Significantly fewer Corynebacterium bovis intra-mammary infections (III) were cured in the teat sealer group during the dry period. A novel method of speciation based on endonucleaser estriction analysis of the 16S rRNA gene sequence was successfully developed and utilized to differentiate C. bovis for other Corynebacterium species. A novel lipophilic Corynebacterium species, named "C. langfordif', was identified, typed by 16S rRNA gene sequencing and described. Re-analysis of the study database demonstrated for the first time that quarters infected with C. bovis during the dry period were significantly less likely to acquire a major pathogen BE. It appeared that IMI with C. bovis "protected" quarters from infection with mastitis pathogens. In vitro studies on solid and in liquid media demonstrated that metabolic products of C. bovis could inhibit the growth of some mastitis pathogens. The inhibitory factor was partially and almost completely inactivated by heating to 100°C and treatment with Proteinase K respectively. Production of an inhibitory factor (possibly a bacteriocin) is proposed as one explanation for the protective effect demonstrated in vivo