Progress 10/01/11 to 09/30/12
Outputs
OUTPUTS: The goal of this seed research was to provide proof-of-concept demonstrating that a novel bioinformatics approach developed by Epivax Inc. for human pathogens can be used to identify immunoprotective epitopes that could be used in the development of effective, inexpensive, and safe vaccines for prevention of bacterial diseases in aquaculture. The goals of this research were to: 1) Survey multiple stakeholders to determine which pathogen and finfish species should be targeted for our proof-of-concept; 2) Target eight genes that are highly conserved among the selected pathogens; and 3) Determine immune response in fish against the predicted epitopes. We performed a survey targeted to the Northeast US region that included 3 shellfish pathologists, 5 extension agents (ME, NY, NH, RI, MA, CT, 4 responded), and 2 farmers (Cooke Aquaculture from Maine and Great Bay Aquaculture from New Hampshire). Input from regional stakeholders regarding the suitability of the approach and their needs regarding vaccines for major diseases affecting their facilities was also gathered through a presentation of our concept in vaccine design at the Northeast Aquaculture Conference and Expo held in Plymouth, MA, in December 2010. Farmers were intrigued by the approach and expressed a large need for tools to prevent diseases like Infectious Salmon Anemia in salmon, and vibriosis in salmon, summer flounder, and cod. PARTICIPANTS: This research brought together a diverse team of experts from academia and industry. The PI, Dr. Denice Spero is the Co-Director of the iCubed and a Research Professor at URI. She has 18 years of experience working in the pharmaceutical industry and has expertise in moving drug candidates from the discovery stage into development. Denice was responsible for the oversight of the program and coordinated the research. Dr. Annie De Groot M.D. is CEO and Chief Scientific Officer of EpiVax. She is a specialist in vaccine development and licensed the EpiMatrix vaccine design technology and established EpiVax in 1998. She participates in the study design. Dr. William Martin is the COO and Chief Information Officer at EpiVax. He was responsible for the development of algorithms for the identification of immunogenic epitopes in the sequences of Vibrio anguillarum. Dr. Lenny Moise is Assistant Research Professor at the Institute for Immunology and Informatics (iCubed) at the University of Rhode Island. He has 7 years experience in translating immunoinformatic epitope predictions to immunogenic and efficacious vaccines using a genomes-to-vaccine approach. He developed the DNA vaccines used in this research. Dr. David Nelson, a microbiologist at URI, provided the sequences and strains of V. anguillarum used in this research. Dr. Gomez-Chiarri, an aquatic pathologist, was responsible for the design of the immunization and challenge experiments and supervised the students performing the experiments. Rachel Bone, a masters student at URI,performed these experiments, with the help of undergraduate student Danielle Aguirre. TARGET AUDIENCES: Target audiences for this research include researchers, veterinarians, and immunologists interested in aquatic animal health, and vaccine companies. Outreach effort of this research specially targeted fish farmers in the New England region, as well as fish pathologists and extension specialists. PROJECT MODIFICATIONS: Major changes include: 1) the use of a challenge experiment to test the candidate vaccines, since isogenic trout available were too small to collect samples for immunological analysis; and 2) performance of an additional experiment with outbred trout so samples for immunological analysis could be collected.
Impacts
Outcomes of this seed research on the development of an immunoinformatics approach to the development of vaccines for fish pathogens included the development of a tool for the identification of vaccine candidates (epitope vaccines) for the bacterial pathogen of fish Vibrio anguillarum based on the identification of epitopes that are predicted to best bind pockets in the trout MHC critical to T cell epitope binding. EpiVax's analysis included alignment of rainbow trout Major Histocompatibility Complex (MHC) to a human reference allele (HLA) and extraction of pocket profiles. Weak homologues within a database of human MHC structures were successfully identified. First generation predictive models were developed and immunogenic epitopes in selected sequences of V. anguillarum genes were predicted. In order to test these predictions, 3 epitope-based DNA vaccines were developed each containing a set of 10 peptides predicted to have high (VibVaxI), medium (VibVaxII), and low (VibVax III) immunogenicity. Two vaccination trials were performed to test the efficacy of these DNA vaccines (delivered by intramuscular injection), compared to a control heat-killed Vibrio anguillarum vaccine (delivered by immersion). One of these trials was done using isogenic fish (Arlee and OSU strains) provided by G. Thorgaard (Washington State University), while the other was done using outbred rainbow trout from the URI Aquaculture Center. Triplicate groups of 6 - 20 trout each were immunized using vaccine vehicle (negative control), the candidate DNA vaccines (trial 2 only), a mix of the 3 DNA vaccines, and a heat-killed V. anguillarum vaccine (positive control). In trial 2, 10 fish per treatment were euthanized 5 weeks after immunization and samples were collected for analysis of immunological responses to the vaccines (in storage at -80C pending further analysis). In addition, four (trial 1) and 5 (trial 2) weeks after immunization, duplicate tanks for each treatment containing 6-10 fish were challenged by intraperitoneal injection of Vibrio anguillarum and mortality was evaluated daily for 2 weeks. An increase in survival compared to control non-vaccinated fish was detected in fish vaccinated with a mix of the three DNA vaccines (11% cumulative percent mortality compared to 44% in controls) in trial 1. These results should be interpreted with caution since they were only evident in one of the isogenic strains (OSU) and due to the impact of a concurrent infection by a water mold. The DNA vaccines did not provide protection against challenge with V. anguillarum in trial 2. These apparently conflicting results indicate that further research needs to be done to determine the efficacy of the immunoinformatics approach. The research team has developed several recommendations on how to proceed to further refine the prediction algorithms and the use of a better model to validate the approach. This research has led to the training of a masters graduate student and an undergraduate student on fish vaccination and disease challenge methods.
Publications
- No publications reported this period
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