Towards the development of a vaccine for proliferative kidney disease

This proposal will study an important disease that affects the rainbow trout aquaculture industry, proliferative kidney disease (PKD), with the objective of making significant progress towards the development of a vaccine. The disease is caused by a microscopic parasite known as a myxozoan, which causes a severe immune response in fish characterized by a chronic kidney pathology. Our recent work led to the discovery that colonial freshwater invertebrates known as bryozoans act as a reservoir of parasites infective to fish. Recently we have found that increasing temperatures cause the parasite to proliferate in bryozoans with greater numbers of released spores, suggesting that that the disease is likely to become more problematic in farmed and wild salmonid stocks as water temperatures increase due to climate change. There is already evidence for this effect in wild brown trout populations in Switzerland and wild Atlantic salmon populations in Norway and we are aware that PKD has increased in severity over the past few years in trout farms in Southern England. Currently, there are no current treatments to prevent or control the disease. However, an important characteristic of PKD is that fish that do survive an initial infection with the parasite are resistance to a subsequent exposure, and thus priming of the immune system with a vaccine is expected to be one way to control this disease. The key is to find an appropriate molecule that can trigger a protective immune response. In recent studies we have identified a number of PKD molecules, several of which have characteristics in common with molecules that elicit protection in other host-pathogen models. So in this proposal we will study the effectiveness of these molecules. We will use a recent advance in vaccine technology, and use for the most part DNA vaccines rather than protein based vaccines, since the former are a cost-effective way to screen our vaccine candidates and have been shown previously to work well in fish. Fish will be vaccinated at a commercial rainbow trout farm, that suffers from this disease every year, and the results will be assessed in terms of survival and the immune response elicited. However, we will not assume that the best vaccine candidates will be amongst the genes we have already sequenced, and so will take an alternative approach as well. The second approach will use pathogen material isolated from the fish host, as well as an intermediate host, and using molecular biology techniques we will create 'gene libraries' from this material that will contain many of the genes that can be expressed by this pathogen. In the same way as above, we will use a DNA vaccine approach to screen these gene libraries for vaccine candidates, but in this case will use 'pools' of many genes. Fish vaccinated with these different 'pools' will be assessed for disease resistance as above, and batches that show beneficial affects will be subdivided and retested, to eventually give small numbers of potential candidate molecules to look at in more detail. Since pathogen strain variation can affect the molecular composition of molecules, we will undertake a survey of some of the candidate genes from the initial approach and see if they are invariant and suitable for all strains, or whether they vary and that an eventual vaccine would require to be derived from multiple sources. In this manner, we will take a systematic approach to vaccine development for this disease, based on our past success in discovering parasite genes, and our existing expertise in molecular biology, PKD biology and fish vaccination.

Proliferative kidney disease (PKD) is one of the most economically devastating diseases currently affecting the aquaculture industry in the UK. Increasing water temperatures due to climate change processes are likely to exacerbate PKD outbreaks, owing to their impact on parasite development in bryozoan populations. Currently, there are no treatments for this disease, although the possibility of vaccination exists as recovering fish are known to have resistance to re-infection. Our recent work uncovered a large resource of novel genes from the PKD parasite, several of which are homologous to or have similar molecular characteristics with known protective antigens in other parasite models. Here we will determine the protective efficacy of genes of therapeutic interest as single-gene DNA vaccines. DNA vaccine technologies have become firmly established in fish vaccinology, offering high levels of protection efficacy, particularly against viruses. The most immune-protective antigen genes in this study will also be further tested by fusing each gene to a suitable molecular adjuvant, namely a parasite-derived Hsp70 fragment and by assessing the protective efficacy of the corresponding recombinant proteins. We will also undertake an unbiased approach to the discovery of protective antigens by employing expression library immunization. Full length cDNA libraries will be generated from both fish and bryozoan-derived parasite material, and sub-libraries used for immunisation. Protective sub-libraries will be sequentially fractionated and genes present in small protective gene pools finally sequenced and characterised. Looking towards the future, we will determine the sequence diversity of protective antigens in the most divergent parasite strains, as determined by ITS-1 sequence analysis. Favourable antigens for strain-transcending vaccines will be those with very low sequence diversity and very low sequence homology to known fish genes.