| Summary: | With the culture of marine fish species increasing, interest in ways to combat infection in these species has grown. One such avenue is the search for novel antimicrobials. Antimicrobial peptides (AMPs) are host derived molecules with a significant role in innate immune responses and have been studied extensively in mammals. Well known AMP families include β-defensins, cathelicidins, liver expressed antimicrobial peptides and, specific to fish, the piscidins. While these molecules have been found in some fish species mining of expressed sequence tag (EST) databases was carried out to identify them in other fishes. In this study we have found new members of the β-defensin, cathelicidin and piscidin families of AMPs, while also identifying new hepcidins and LEAP-2 molecules in a diverse range of fish species. The rest of this thesis focused on one major AMP family, the cathelicidins, which in fish have been mainly identified in salmonids. They are stored in secondary granules of neutrophils as inactive pro-peptides and when infection occurs, a cleavage event releases the active mature form that effects microbicidal activity. Characterization of the cathelicidin genes present in Atlantic cod (Gadus morhua) and haddock (Melanogrammus aeglefinus) was undertaken. A number of ESTs with significant homology to cathelicidins were found in Atlantic cod. Primers were designed to amplify these sequences by PCR in both gadoid species, and the gene products were cloned and sequenced. One and two cathelicidin genes were identified in cod (gmCath1) and haddock (maCath1; maCath2), respectively, which differ in the length of their active mature peptides. Gadoid cathelicidins were constitutively expressed in many tissues and were upregulated by a range of immunostimulants. Analysis of the gmCath1 promoter revealed numerous putative transcription factor binding sites, some of which were used in an expression vector construct to show upregulation of gene expression when stimulated with the viral RNA mimic Poly I:C. Synthetic and recombinant gadoid cathelicidins were produced but did not appear to display antimicrobial activity, so this requires further investigation. The discovery of new members of the major AMP families in such a diverse range of fish species bodes well for future research into the use of these antimicrobials as potential disease control molecules in the aquaculture industry. Particularly for Atlantic cod this is crucial as the major cod farms in Norway are experiencing difficulties with bacterial infections for which there is no vaccine at present.
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