Location: Infectious Bacterial Diseases Research
2017 Annual Report
Objectives
Objective 1: Define the immunopathogenesis of bovine tuberculosis at the lesion and cellular level by evaluating local cytokine and biomarker expression.
Subobjective 1.1. Characterize and compare known or implied relevant cytokine and biomarker expression in granulomas of different histopathologic stages (i.e. early vs. late) in lungs and lymph nodes from cattle experimentally inoculated with M. bovis.
Subobjective 1.2. Characterize cytokine and biomarker expression at the lesion level over time.
Subobjective 1.3. Characterize and compare lesion level cytokine and cellular responses between non-vaccinated cattle and cattle with vaccine-induced protective immune responses.
Objective 2: Using antigen mining and transcriptome analysis, develop novel diagnostic tests with improved sensitivity and specificity as compared to current methods.
Subjective 2.1. Improve specificity of diagnostic tests by developing diagnostic reagents from proteins found in M. bovis but not in non-tuberculous mycobacteria.
Subobjective 2.2: Identify proteins/genes expressed by M. bovis in vivo that may be considered as potential diagnostic test targets.
Subobjective 2.3: Use genomics/transcriptomics to characterize genes/gene profiles of M. bovis-infected vs non-infected cattle.
Objective 3: Develop novel vaccines, technologies and platforms (e.g. attenuated live vaccines and vectored vaccines) that can be used to reduce TB in cattle and white-tailed deer and interrupt disease transmission.
Subobjective 3.1. Examine duration of immunity to experimental infection provided by the vaccine M. bovis BCG in white-tailed deer.
Subobjective 3.2. Examine the effects of oral BCG vaccination of white-tailed deer on deer-to-deer transmission of virulent M. bovis.
Subjective 3.3. Determine the efficacy of simultaneous administration of parenteral BCG and a mucosally delivered bacterial-vectored subunit vaccine against aerosol M. bovis infection in neonatal calves.
Approach
Characterize and compare cytokine and biomarker expression (immune responses) at the cellular level in lungs and lymph nodes of Mycobacterium bovis-infected cattle. Comparing responses between tissues, as well as over time, will aid in understanding the host response to M. bovis within the environment where host and pathogen interact (granuloma). We aim to improve the specificity of diagnostic tests by developing diagnostic reagents from proteins found in M. bovis but not in non-tuberculous mycobacteria, thus avoiding cross-reactivity elicited by environmental mycobacteria that contributes to false positive results on cattle tuberculosis diagnostic tests. Similarly, we aim to identify proteins/genes expressed by M. bovis in vivo that may be considered as potential diagnostic test targets and to use genomics/transcriptomics to characterize genes/gene profiles of M. bovis-infected vs non-infected cattle. These data will aid diagnosis and provide insight into the immunopathogenesis of bovine tuberculosis. In terms of vaccine evaluation, we aim to examine duration of immunity to experimental infection provided by the vaccine M. bovis BCG in white-tailed deer and examine the effects of oral BCG vaccination on deer-to-deer transmission of virulent M. bovis. In cattle, we aim to determine the efficacy of simultaneous administration of parenteral M. bovis BCG and a mucosally delivered bacterial-vectored subunit vaccine against aerosol M. bovis infection in neonatal calves.
Progress Report
Nearly 100 years ago, in 1917, USDA initiated a bovine tuberculosis eradication program that is still in place today. Since that time, significant progress has been made, but eradication has proven elusive. Two significant obstacles to eradication include, 1) the lack of rapid and accurate diagnostic tests to detect animals infected with Mycobacterium bovis (the cause of tuberculosis in animals), and 2) wildlife acting as a source of infection for cattle. Consequently, research activities within the ARS Tuberculosis (TB) Project provide direct support for the TB eradication program directed by the Animal and Plant Health Inspection Service (APHIS), specifically discovery and development of diagnostics, vaccines, and animal disease models to protect and ensure food safety through enhanced capabilities to detect, prevent and control animal diseases.
During FY17 the project developed new tools to evaluate immune responses of cattle to M. bovis. These tools allow understanding of immune responses at the site of infection, where the outcome of infection is determined (elimination versus dissemination). The genomes of nonpathogenic environmental mycobacterial species that cause false positive tests were sequenced. Knowledge of the genetic sequences of these organisms allows development of novel diagnostic tests that will reduce false positive test results. During the past year scientists in the project also worked with APHIS program staff to resolve problems with a diagnostic test for tuberculosis in cattle.
Studies conducted by the ARS TB Project benefit the USDA regulatory program, state regulatory and natural resource agencies, and stakeholder groups (beef, dairy and deer producers). Research findings are used to create or modify state and federal regulations regarding TB testing of cattle and deer including approval of novel tests for the eradication of bovine tuberculosis.
Accomplishments
1. Improved tuberculosis diagnostic tests. Non-disease-causing mycobacteria often interfere with diagnosis of tuberculosis in cattle, resulting in unacceptably high numbers of false positive test results. False positive findings result in unnecessary removal and slaughter of uninfected cattle, often resulting in financial losses for producers, and impacting the bovine tuberculosis control program. ARS scientists at Ames, Iowa sequenced the genomes of the most common mycobacterial species that result in false positive results in the United States and compared their genome to that of virulent Mycobacterium bovis. Several proteins unique to M. bovis were identified. Using these unique proteins, novel diagnostic tests or modifications of existing tests can be developed that result in fewer false positive results while maintaining diagnostic sensitivity. More accurate diagnostic tests will expedite the eradication of bovine tuberculosis from the US, decrease financial losses to producers, and improve relations between producers and regulatory officials.
Review Publications
Palmer, M.V., Cox, R.J., Waters, W.R., Thacker, T.C., Whipple, D.L. 2017. Using white-tailed deer (Odocoileus virginianus) in infectious disease research. Journal of the American Association for Laboratory Animal Science. doi: no_doi/1494535970305.
Lyashchenko, K.P., Greenwald, R., Sikar-Gang, A., Sridhara, A.A., Johnathan, A., Lambotte, P., Esfandiari, J., Maggioli, M.F., Thacker, T.C., Palmer, M.V., Waters, W.R. 2017. Early detection of circulating antigen and IgM-associated immune complexes during experimental Mycobacterium bovis infection in cattle. Clinical and Vaccine Immunology. doi: 10.1128/CVI.00069-17.
Palmer, M.V., Thacker, T.C., Waters, W.R. 2015. Differential cytokine gene expression in granulomas from lungs and lymph nodes of cattle experimentally infected with aerosolized Mycobacterium bovis. PLoS One. 10(12):e0142287.
Maggioli, M.F., Palmer, M.V., Thacker, T.C., Vordermeier, H.M., McGill, J.L., Whelan, A.O., Larsen, M.H., Waters, W.R. 2016. Increased TNF-alpha/IFN-gamma/IL-2 and decreased TNF-alpha/IFN-gamma production by central memory T cells are associated with protective responses against bovine tuberculosis following BCG vaccination. Frontiers in Immunology. 7:421.
Palmer, M.V., Thacker, T.C., Waters, W.R. 2016. Multinucleated giant cell cytokine expression in pulmonary granulomas of cattle experimentally infected with Mycobacterium bovis. Veterinary Immunology and Immunopathology. 180:34-39.
Wanzala, S.I., Palmer, M.V., Waters, W.R., Thacker, T.C., Carstensen, M., Travis, D., Sreevatson, S. 2017. Evaluation of pathogen-specific biomarkers for the diagnosis of tuberculosis in white-tailed deer (Odocoileus virginianus). American Journal of Veterinary Research. 78(6):729-734. doi: 10.2460/ajvr.78.6.729.
