Location: Infectious Bacterial Diseases Research
2023 Annual Report
Objectives
Objective 1: Develop rationally-designed vaccines, including recombinants, that prevent disease or mitigate disease spread with an emphasis on platforms that allow for extended antigen release and remote delivery for cattle and elk.
Subobjective 1A: Assess the potential of polyanhydride polymers as extended release vaccine platform using killed bacterial preparations to generate cell-mediated immune responses against Brucella in cattle and elk.
Subobjective 1B: Evaluate immunogenicity/efficacy of a new recombinant vaccine in elk.
Subobjective 1C: Develop new DIVA vaccine strains with targeted mutations using CRISPR.
Objective 2: Improve current diagnostic tests for brucellosis that differentiate between Brucella species and can be used in multiple livestock and wildlife species.
Subobjective 2A: Characterize sensitivity and specificity of diagnostic tests using synthetic Brucella antigens.
Subobjective 2B: Identify new Brucella epitopes recognized by humoral responses of B. suis infected animals using phage library expression.
Objective 3: Using transcriptomic approaches, define the immunopathogenesis of brucellosis at the tissue and cellular level by analyzing gene expression of peripheral immune responses and local immune responses to inform vaccine and therapeutic development.
Subobjective 3A: Using a transposon generated mutant library of Brucella abortus, characterize how bacterial populations are limited during in vivo infection and identify bacterial genes critical for establishing infection in cattle.
Subobjective 3B: Using a transposon-generated mutant library of Brucella abortus, characterize how pregnancy influences diversity of bacterial populations during in vivo infection in cattle.
Subobjective 3C: Characterize the gene expression profiles of the peripheral immune response to Brucella in cattle, bison, and elk to determine species-specific differences in response to vaccination.
Approach
The long-term goals of this project are to facilitate the completion of brucellosis eradication programs in domestic livestock, and prevent reintroduction of brucellosis into livestock from wildlife reservoirs. Specifically, fundamental knowledge on Brucella pathogenesis will be gained, efficacious vaccination systems will be identified, and sensitive and specific diagnostic tools will be developed to aid eradication programs. Immunogenicity of vaccination strategies in targeted hosts (cattle, bison, elk, and swine), including novel vaccine platforms, will be evaluated in targeted species and efficacy characterized by experimental challenge. In addition, the project will try to improve the standard experimental challenge model for elk to better replicate the clinical effects of brucellosis under field conditions. By simultaneously characterizing the in vivo transcriptome of B. abortus and natural host during infection, we will develop knowledge of molecular mechanisms involved in regulation of host responses to infection, and genes expressed by the pathogen under in vivo conditions. This basic knowledge will identify future targets for development of new vaccines, diagnostics, immunomodulation, and possibly therapeutics. New diagnostics will be developed and analyzed for their ability to detect brucellosis in swine and cattle, and may allow differentiation of which Brucella spp. is associated with infection. The research will help resolve the risk of re-infection of domestic livestock from wildlife reservoirs of brucellosis, protect the financial investment that has been made in the U.S. brucellosis eradication program, and provide public health benefits by reducing the risk of zoonotic infection.
Progress Report
During the FY23 year, progress was made in meeting the objectives of the brucellosis research project despite significant financial challenges. As vaccination is a critical intervention tool for the control of brucellosis, immune responses to killed RB51 antigens in a sustained release platform were evaluated for Objective 1. Progress is being made on new vaccine candidates but has not progressed to where recombinant strains can be evaluated under in vivo conditions. In studies under Objective 2, a new test using synthetic O-polysaccharide as antigen was evaluated. Antigen discovery using phage libraries or with collaborators is ongoing to identify new antigens that may improve sensitivity or specificity in brucellosis diagnostic tests. For Objective 3, open and pregnant cattle were infected with a defined transposon mutant Brucella abortus library and differences in tissue localization and strain colonization were characterized. In addition, studies were conducted on immune gene expression in Brucella-infected elk to help understand the molecular basis for differences in immune function between species. Comparative transcriptomics and genomics continue to be utilized to identify molecular differences between natural hosts of Brucella that may contribute to observed differences in disease pathogenesis.
Accomplishments
1. Mucosal and systemic responses of Elk to Brucella abortus infection. Brucellosis causes reproductive losses in domestic livestock and can also cause human infection. Elk are important reservoirs for brucellosis in Yellowstone National Park, and surrounding areas, and can transmit infection to domestic livestock. Although vaccination is frequently used to prevent brucellosis, current vaccines are not efficacious in elk. To understand immune responses in elk, ARS scientists in Ames, Iowa, compared mucosal and system antibody responses after experimental brucellosis infection. Data demonstrated that elk have increased IgG and IgA titers in conjunctival fluids after experimental infection that was not influenced by vaccination status. Systemic (serum) IgG antibody responses increased after infection but differed from mucosal responses. These data suggest differences between systemic and mucosal immune regulation in elk that may influence disease pathogenesis. These data will be of interest to regulator officials, wildlife managers, and scientists with interests in brucellosis in wildlife reservoirs.
2. Tissue localization of recombinant strains of Brucella in cattle. Brucellosis is a zoonotic disease that causes reproductive losses in domestic livestock. To understand the pathogenesis of brucellosis in a natural reservoir, ARS scientists in Ames, Iowa, infected non-pregnant and pregnant cattle by mucosal or intravenous routes using a defined Brucella abortus transposon library. Cattle infected by the intravenous route primarily had localization in internal tissues (bronchial and hepatic lymph nodes) whereas animals infected by the conjunctival route primarily had localization in lymphatic tissues of the head and neck. Pregnant animals had more diffuse tissue localization than non-pregnant animals. Data suggests differences in disease pathogenesis by route of challenge and between open and pregnant animals. Results also demonstrated that some recombinant strains could infect tissues if they did not have to cross a mucosal surface. This work provides information on the mechanisms used by Brucella to infect cattle and will be of interest to regulatory officials and scientists with an interest in the pathogenesis of brucellosis.
3. Sodium hypocholoride identified as a disinfectant for the inactivation of the potential bioweapon Brucella. Brucella abortus is a potential bioweapon that is listed as a Select Agent within the United States. Select Agent regulations require validation of procedures or methods for inactivating the agent. To develop safe and cost-effective methods to disinfect the effluent waste system of a Biolevel 3 facility prior to maintenance, ARS scientists in Ames, Iowa, evaluated the inactivation properties of diluted sodium hydroxide and sodium hypochlorite solutions on Brucella abortus viability. Data demonstrated that highly diluted solutions of sodium hypochlorite are highly effective in killing high concentrations of Brucella abortus. Although sodium hydroxide would be effective in removing organic matter from the system, it was not as effective as sodium hypochlorite as an inactivation agent for Brucella. This data demonstrates the disinfectant capabilities of sodium hypochlorite that can be utilized for Biolevel 3 facility procedures. This data will be of interest to managers of high containment facilities, regulatory officials, producers, and scientists working with brucellosis for use in disinfecting facilities or equipment in a manner that is safe for personnel and the environment.
Review Publications
Falkenberg, S.M., Dassanayake, R.P., Crawford, L., Sarlo Davila, K.M., Boggiatto, P.M. 2023. Response to bovine viral diarrhea virus in heifers vaccinated with a combination of multivalent modified live and inactivated viral vaccines. Viruses. 15(3). Article 703.. https://doi.org/10.3390/v15030703.
Kanipe, C.R., Boggiatto, P.M., Putz, E.J., Palmer, M.V. 2022. Histopathologic differences in granulomas of BCG vaccinated and non-vaccinated cattle with bovine tuberculosis. Frontiers in Microbiology. 13, Article 1048648. https://doi.org/10.3389/fmicb.2022.1048648.
Boggiatto, P.M., Olsen, S.C., Palmer, M.V. 2022. Pulmonary hamartoma in an elk calf (Cervus elaphus canadensis). Journal of Veterinary Diagnostic Investigation. 35(2). https://doi.org/10.1177/10406387221141091.
Olsen, S.C., Boggiatto, P.M. 2022. Characterization of the duration of immunity of Brucella abortus strain RB51 vaccination in cattle after experimental challenge. Preventive Veterinary Medicine. 206. Article 105705. https://doi.org/10.1016/j.prevetmed.2022.105705.
Crawford, L., Falkenberg, S., Putz, E.J., Olsen, S.C., Boggiatto, P.M. 2023. Effects of concurrent administration of modified live viral vaccines with RB51 on immune responses to RB51. Frontiers in Veterinary Science. 10. eArticle 1105485. https://doi.org/10.3389/fvets.2023.1105485.
Palmer, M.V., Kanipe, C.R., Boggiatto, P.M. 2022. The bovine tuberculoid granuloma. Pathogens. 11(1). Article 61. https://doi.org/10.3390/pathogens11010061.
