A Novel Vaccine For Aquatic Mycobacteriosis     David J. Pasnik, Stephen A. Smith, Gerhard Schurig and Ramesh Vemulapalli   Aquatic Medicine Laboratory, Department of Biomedical Sciences and Pathobiology, Virginia-Maryland Regional College of Veterinary Medicine, Blacksburg, VA 24061 USA     Mycobacteriosis is an economically important bacterial disease of virtually all species of freshwater and marine fish.  It is usually caused by Mycobacterium marinum, M. fortuitum, or M. chelonae and is a chronic progressive disease, leading to systemic infections and ultimately death.  While the immune response against aquatic Mycobacterium spp. has been characterized, an effective vaccine against mycobacteriosis has not been developed.  A vaccine for aquatic mycobacteriosis was created using a Mycobacterium spp. 85A antigen and Brucella abortus as the delivery vector.  Previous studies in this laboratory have established that striped bass, Morone saxatilis, are particularly susceptible to Mycobacterium spp.   Therefore, juvenile striped bass were used as a model to test the vaccine.  Intraperitoneal vaccinations were administered on days 0, 14, and 35 with bacterial concentrations ranging from 106 through 1010, along with a sham-inoculated saline control group.  On days 0, 14, 35, and 49, blood was drawn from the caudal vessels of the fish for ELISA evaluation.  Data from the ELISA exhibited a significant humoral antibody response to the 85A antigen and B. abortus antigen among the 108, 109, and 1010 groups.  On day 49, fish were challenged with an intramuscular injection of M. marinum at a concentration of  1.6 x 106 colony forming units per fish.  Moribund fish were euthanized and routine histological examination performed to assess the protection conferred by the vaccine.  In addition, on day 56, the anterior kidney harvested from a sub-sample of each group of fish for a lymphoproliferative assay.  This was used to assess the cell-mediate immune response elicited by the vaccine.  The long-term goal of this project is to create a safe, effective, marketable vaccine which will: 1) decrease financial losses due to mycobacteriosis, 2) improve the overall health and immunocompetence of economically-important cultured food fish, 3) decrease antibiotic use in food fish, and 4) provide a protocol for developing fish vaccines against other significant intracellular pathogens.