Location: Infectious Bacterial Diseases Research
2020 Annual Report
Objectives
These studies are focused on developing an understanding of how Leptospira and Treponema species interact with their hosts to establish colonization, infection, and clinical disease. A combination of genomic, proteomic and immunological methodologies will be used to analyze how the host responds to infection and how bacteria respond to the host, with the long-term goal of identifying pathways that can be targeted to alter disease outcomes or exploited to induce protective immunity.
Objective 1. Identify and characterize the Leptospira and Treponema sp. circulating in livestock.
Subobjective l.1 - Determine prevalence of leptospires circulating in local bovine herds.
Subobjective l.2 - Characterize clonal isolates of Treponema from bovine digital dermatitis at the phenotypic, genomic and proteomic level.
Objective 2. Develop animal models that will mimic infection, facilitate characterization of interactions between host and pathogen, and allow development of assays that will accurately identify infected individuals.
Subobjective 2.1 - Characterize urinary immunoglobulin from reservoir hosts of leptospirosis.
Subobjective 2.2 - Characterize the cellular immune response of reservoir hosts of leptospirosis.
Subobjective 2.3 - Characterize and refine an ovine model of bovine digital dermatitis.
Objective 3. Characterize spirochete antigens including those that are differentially expressed during infection.
Subobjective 3.1 - Identification and characterization of leptospiral proteins that are expressed in response to mammalian host signals.
Subobjective 3.2 - Characterize host humoral responses to outer membrane protein antigens derived from bacteria associated with digital dermatitis.
Approach
Objective 1: This objective seeks to identify and characterize species of Leptospira and Treponema sp. circulating in livestock. Studies will be conducted to determine the prevalence of leptospires circulating in local bovine herds (Sub-objective 1.1); and to characterize clonal isolates of Treponema from bovine digital dermatitis at the phenotypic, genomic and proteomic levels (Sub-objective 1.2). We expect these studies to determine if serovars of leptospires currently circulating in bovine populations of the Mid-West have changed over the last 20 years and to demonstrate that different phylotypes of Treponema derived from bovine digital dermatitis have unique genomic, proteomic and virulence factors.
Objective 2: Development of animal models that mimic infection will facilitate characterization of interactions between host and pathogen, and allow development of assays that will accurately identify infected individuals. Urinary immunoglobulins from reservoir hosts of leptospirosis will be collected and characterized (Sub-objective 2.1); the cellular immune response of reservoir hosts of leptospirosis will also be characterized (Sub-objective 2.2); and an ovine model of bovine digital dermatitis will be further characterized and refined (Sub-objective 2.3). We will also evaluate immune activation pathways in a reservoir host model of leptospirosis using the inbred Fisher 344 rat. Studies conducted will advance the use of sheep as a ruminant model to understand the pathogenic mechanisms and involvement of treponemes in digital dermatitis.
Objective 3: Characterize spirochete antigens including those that are differentially expressed during infection. Studies will be conducted to identify and characterize leptospiral proteins that are expressed in response to mammalian host signals (Sub-objective 3.1) and to characterize host humoral responses to outer membrane protein antigens derived from bacteria associated with digital dermatitis (Sub-objective 3.2).
Progress Report
Work related to Objective 1 led to the development of novel microbiology growth media that enhances the isolation and propagation of leptospires. In collaboration with scientists at the National Veterinary Services Laboratory and the Center for Disease Control and Prevention, the media is being used to isolate Leptospira bacteria and determine disease prevalence in domestic and wild animal populations. Also related to Objective 1, completion of the genomes of Treponema brennaborense and T. phagedenis led to novel insights into the genetics of this bacterial species, including identification of a tetracycline antibiotic resistant gene (tetM) in T. brennaborense that is not present in other species of Treponema. In addition, a new media for isolation and propagation of Treponema was developed.
In work related to Objective 2, ARS scientists used gel electrophoresis and immunoblotting to demonstrate the presence of antibodies against pathogenic leptospires in the urine of cattle naturally infected with L. borgpetersenii serovar Hardjo. Work in a chronic rat model of leptospirosis identified changes in gene expression pathways associated with innate immunity. The bacterial community (microbiome) of digital dermatitis was characterized in an experimental ovine (sheep) model and naturally infected ungulates (hoofed mammals). In collaborative research, the pathology and immunological responses of free-roaming wild elk with a digital dermatitis-like disease in the Pacific Northwest were characterized.
In studies related to Objective 3, data demonstrated that leptospires shed in urine have modified proteins (post-translational modifications) as compared to bacteria grown under in vitro conditions.
Accomplishments
1. Development of leptospira vaccines. Leptospirosis is a zoonotic bacterial infection that causes reproductive losses in cattle and clinical disease in humans. ARS scientists in Ames, Iowa, completed studies to: 1) characterize immune response and efficacy of standard bacterin vaccines (suspension of killed or live attenuated bacteria), and 2) determine if using different vaccine adjuvants would enhance efficacy. Results demonstrated that effective vaccines elicited production of pro-inflammatory cytokines (cellular-signaling molecules) in memory T cells. Increased gamma-delta T cells, an effector type of T-cell, were detected following experimental infection. It is believed that both the cytokine signals from the memory cells and circulating effector gamma-delta T cells are necessary for clearance of Leptospira from resident tissues. The novel adjuvants evaluated in the study induced greater immune responses than those induced by current vaccines. This research will be used by other scientists, vaccine companies, and producers for creating better leptospirosis vaccines for cattle and bacterial diseases of livestock.
Review Publications
Nally, J.E., Ahmed, A., Putz, E.J., Palmquist, D.E., Goris, M. 2020. Comparison of real-time PCR, bacteriologic culture and fluorescent antibody test for the detection of Leptospira borgpetersenii in urine of naturally infected cattle. Veterinary Sciences. 7(2):66. https://doi.org/10.3390/vetsci7020066.
O'Neill, L., Keane, O.M., Ross, P.J., Nally, J.E., Seshu, J., Markey, B.K. 2019. Evaluation of protective and immune responses following vaccination with recombinant MIP and CPAF from Chlamydia abortus as novel vaccines for enzootic abortion of ewes. Vaccine. 37(36):5428-5438. https://doi.org/10.1016/j.vaccine.2019.06.088.
Hornsby, R.L., Alt, D.P., Nally, J.E. 2020. Isolation and propagation of leptospires at 37 degree C directly from the mammalian host. Scientific Reports. 10(9620). https://doi.org/10.1038/s41598-020-66526-4.
Wilson-Welder, J.H., Boggiatto, P.M., Nally, J.E., Wafa, E.I., Alt, D.P., Hornsby, R.L., Frank, A.T., Jones, D.E., Olsen, S.C., Bowden, N.B., Salem, A.K. 2020. Bovine immune response to leptospiral antigen in different novel adjuvants and vaccine delivery platforms. Vaccine. 38(18):3464-3473. https://doi.org/10.1016/j.vaccine.2020.02.086.
