NE1701: Mycobacterial Diseases of Animals

(Multistate Research Project)

Status: Active

NE1701: Mycobacterial Diseases of Animals

Duration: 10/01/2017 to 09/30/2022

Administrative Advisor(s):


NIFA Reps:


Statement of Issues and Justification

The proposed multi-state initiative will focus on two of the most important mycobacterial diseases of animals; paratuberculosis (Johne’s disease; JD) and the bovine tuberculosis complex (TB). These two mycobacterial diseases represent some of the most prevalent and economically significant infections of livestock, and each has a long and rich history. A brief background, including significance and need for work, on each of these diseases, is provided below. Johne’s Disease (JD) is a chronic granulomatous inflammatory intestinal disease that results from infection with Mycobacterium avium subspecies paratuberculosis. JD is recognized as a serious economic and animal health problem in domesticated ruminants including dairy and beef cattle, sheep, and goats throughout the world. It results in more than $200 million in annual losses to the United States (US) dairy industry each year with additional losses incurred by the other species. The growing recognition of M. paratuberculosis infection in wildlife species is also of considerable concern. Similarly, recent evidence of the presence of M. paratuberculosis in retail milk sources is of concern from a milk quality and potential food safety standpoint. The growing recognition of MAP infection in wildlife species is also of considerable concern, as well as contaminations in environments such as grassland, soil, and even water-supply systems. Despite considerable efforts, JD remains a major concern for producers with very high prevalence rates (68% of all US dairy herds and 95% of those with over 500 cows have at least one JD positive animal) based on culturing of fecal samples. MAP and JD are now considered endemic in the US and in most dairy producing nations, and without major breakthroughs, efforts at controlling pathogen are likely to remain salutary and the disease will continue to spread unabated.


There have been considerable ongoing efforts made to identify knowledge gaps, define research priorities, and develop recommendations for implementing JD control measures in the field. For instance, a 2003 report from the National Research Council of the US National Academies of Sciences on JD comprehensively reviewed the literature, identified major gaps in knowledge, and provided clear recommendations for future research priorities and strategies for the prevention and control of JD. In brief, the report concluded that JD is a significant animal health problem whose study and control deserves high priority from the USDA. It was recognized that the problems associated with JD stem from: (i) difficulties in diagnosis because of an unusually long incubation period and a lack of specific and sensitive diagnostic tests for detecting early infections; (ii) a lack of vaccines or other effective measures for infection control; and, (iii) general lack of awareness of the disease and its true economic and animal health consequences by producers and veterinarians. The report made 25 specific recommendations regarding implementation of strategies for the control of JD, educating and training of producers and veterinarians, and filling of key gaps in knowledge relating to JD. In 2005 and 2006, specialty working groups were formulated by the USDA-APHIS-VS and the Johne’s Disease Integrated Program (JDIP; http://mycobacterialdiseases.org/) to review knowledge gaps and opportunities for research, extension, and training in JD.


Some of the community needs that were identified as gaps included: (i) the development of new and improved diagnostics and candidate vaccines; (ii) improving research efficiencies by developing shared resources and guidelines for basic and translational research in JD; and, (iii) developing strong education and extension programs. While considerable progress has been made in all areas, the proposed multi-state initiative will facilitate meeting remaining major unmet needs.


The TB complex of diseases of livestock results from infection of animals with mycobacterial pathogens, primarily M. bovis and Mycobacterium avium subspecies avium (MAA). These organisms can cause disease in multiple livestock and wild animal species and can be readily transmitted to humans. M. bovis, whose disease and infections will be the primary focus of the activities proposed in this multi-state initiative, is closely related to the organism that causes human tuberculosis, Mycobacterium tuberculosis (MTB).


TB is a disease of antiquity that has resulted in a considerable economic loss to animal agriculture and, as a zoonotic disease, contributed greatly to human suffering prior to the widespread requirement for milk pasteurization. In fact, at the turn of the 20th century, M. bovis was considered to be the cause of greater economic losses to livestock production than all other infectious diseases combined. The implementation of rigorous control and disease eradication programs, including test and slaughter or test and segregate programs, have reduced or eliminated tuberculosis in cattle in the US and most developed countries. However, reservoirs in wildlife have precluded complete eradication. TB continues to be a significant recurring concern in many countries, including Ireland, the United Kingdom (UK) and New Zealand. In addition, both bovine tuberculosis and M. bovis infections in humans remain common in less developed countries, resulting in considerable economic losses due to disease and trade restrictions.


While TB incidence in the US remains low, there is considerable concern that we may be experiencing a resurgence of this disease in livestock species, primarily cattle. In 1994, a white-tailed deer (WT deer) from northeastern Michigan was found to be infected with M. bovis. This led to the wide-scale testing of cattle and deer with subsequent identification of M. bovis in both populations within this area. The spread of M. bovis in Michigan was slowed by a strict policy of total herd depopulation upon identification of positive cattle, as well as large-scale hunter education programs and a massive testing initiative in WT deer. Still, in Michigan, over 650 cases of M. bovis infection in WT deer and 49 positive cattle herds have been identified to date. Alarmingly, M. bovis has now spread to other states. M. bovis was recently detected in 27 WT deer and 12 cattle herds in Minnesota and has been confirmed in cattle from Colorado, Nebraska, Indiana, Kentucky, North Dakota, South Dakota, New Mexico, and California. Detection of M. bovis infection has lead to quarantine and depopulation of nearly all affected herds. Clearly, this disease is continuing its resurgence throughout the US, particularly where cattle and WT deer commingle.


A second major source of M. bovis infected cattle in the US is imported animals from other countries where the disease is endemic, particularly Mexico. Indeed, molecular epidemiology studies have demonstrated that M. bovis cases in all states other than Michigan are likely of Mexican origin. Although USDA regulations stipulate that imported cattle must be tested within 60 days of import, the low sensitivity of most approved M. bovis diagnostic tests suggests that some infected animals will be missed.   Because cattle are only held at the border for 48 to 72 hours, there is little time to conduct additional testing at the point of entry. In addition, the lack of mandatory animal identification in the US limits the ability to track cattle after introduction into the country. Clearly, it is crucial to have rapid diagnostics with improved sensitivity that could be deployed at points of entry. It is equally important to improve information on cattle movements to control importation of M. bovis infected cattle.



  1. bovis is of significant concern to government agencies and cattle industries due to associated economic, social and potential public health problems. The inclusion of M. bovis research, teaching and extension in this multi-state project will address serious concerns from cattle industry representatives, government agencies, and public health officials that the US is experiencing a resurgence of M. bovis that will have devastating economic effects, cause a disruption or severe restrictions in movements of cattle including exports, and have profound effects on producers, who own positive herds and must suffer depopulation or quarantine.


Finally, the generation of new knowledge relative to the diagnosis, management, and control of mycobacterial diseases of animals is critical if we are to prevent the spread, lower the prevalence and minimize the impact of the diseases in our livestock populations. USDA NAHMS studies and other work, including the National Dairy Producer Johne’s survey, have shown that while producers are increasingly aware of the diseases, they often lack knowledge relative to their management and control. Therefore, there is a critical need for developing coordinated approaches for education and outreach programs related to mycobacterial diseases of animals.


Taken together, the proposed multi-state initiative described below will facilitate the development of shared research as well as the leveraging of intellectual and physical resources to address some of the most important mycobacterial diseases of animals.


 

Related, Current and Previous Work

The proposed multi-state initiative will focus on two of the most important mycobacterial diseases of animals; paratuberculosis (Johne’s disease; JD) and the bovine tuberculosis complex (TB). These two mycobacterial diseases represent some of the most prevalent and economically significant infections of livestock, and each has a long and rich history. A brief background, including significance and need for work, on each of these diseases is provided below. Johne’s Disease (JD) is a chronic granulomatous inflammatory intestinal disease that results from infection with Mycobacterium avium subspecies paratuberculosis. JD is recognized as a serious economic and animal health problem in domesticated ruminants including dairy and beef cattle, sheep, and goats throughout the world. It results in more than $200 million in annual losses to the United States (US) dairy industry each year with additional losses incurred by the other species. The growing recognition of M. paratuberculosis infection in wildlife species is also of considerable concern. Similarly, recent evidence of the presence of M. paratuberculosis in retail milk sources is of concern from a milk quality and potential food safety standpoint. The growing recognition of MAP infection in wildlife species is also of considerable concern, as well as contaminations in environments such as grassland, soil, and even water-supply systems. Despite considerable efforts, JD remains a major concern for producers with very high prevalence rates (68% of all US dairy herds and 95% of those with over 500 cows have at least one JD positive animal) based on culturing of fecal samples. MAP and JD are now considered endemic in the US and in most dairy producing nations, and without major breakthroughs, efforts at controlling pathogen are likely to remain salutary and the disease will continue to spread unabated.


There have been considerable ongoing efforts made to identify knowledge gaps, define research priorities, and develop recommendations for implementing JD control measures in the field. For instance, a 2003 report from the National Research Council of the US National Academies of Sciences on JD comprehensively reviewed the literature, identified major gaps in knowledge, and provided clear recommendations for future research priorities and strategies for the prevention and control of JD. In brief, the report concluded that JD is a significant animal-health problem whose study and control deserves high priority from the USDA. It was recognized that the problems associated with JD stem from: (i) difficulties in diagnosis because of an unusually long incubation period and a lack of specific and sensitive diagnostic tests for detecting early infections; (ii) a lack of vaccines or other effective measures for infection control; and, (iii) general lack of awareness of the disease and its true economic and animal-health consequences by producers and veterinarians. The report made 25 specific recommendations regarding implementation of strategies for the control of JD, educating and training of producers and veterinarians, and filling of key gaps in knowledge relating to JD. In 2005 and 2006, specialty-working groups were formulated by the USDA-APHIS-VS and the Johne’s Disease Integrated Program (JDIP; http://mycobacterialdiseases.org/) to review knowledge-gaps and opportunities for research, extension and training in JD.


Some of the community needs that were identified as gaps included: (i) the development of new and improved diagnostics and candidate vaccines; (ii) improving research efficiencies by developing shared resources and guidelines for basic and translational research in JD; and, (iii) developing strong education and extension programs. While considerable progress has been made in all areas, the proposed multi-state initiative will facilitate meeting remaining major unmet needs.


The TB complex of diseases of livestock results from infection of animals with mycobacterial pathogens, primarily M. bovis and Mycobacterium avium subspecies avium (MAA). These organisms can cause disease in multiple livestock and wild animal species and can be readily transmitted to humans. M. bovis, whose disease and infections will be the primary focus of the activities proposed in this multi-state initiative, is closely related to the organism that causes human tuberculosis, Mycobacterium tuberculosis (MTB).


TB is a disease of antiquity that has resulted in considerable economic loss to animal agriculture and, as a zoonotic disease, contributed greatly to human suffering prior to the widespread requirement for milk pasteurization. In fact, at the turn of the 20th century, M. bovis was considered to be the cause of greater economic losses to livestock production than all other infectious diseases combined. The implementation of rigorous control and disease eradication programs, including test and slaughter or test and segregate programs, have reduced or eliminated tuberculosis in cattle in the US and most developed countries. However, reservoirs in wildlife have precluded complete eradication. TB continues to be a significant recurring concern in many countries, including Ireland, the United Kingdom (UK) and New Zealand. In addition, both bovine tuberculosis and M. bovis infections in humans remain common in less developed countries, resulting in considerable economic losses due to disease and trade restrictions.


While TB incidence in the US remains low, there is considerable concern that we may be experiencing a resurgence of this disease in livestock species, primarily cattle. In 1994, a white-tailed deer (WT deer) from northeastern Michigan was found to be infected with M. bovis. This led to wide-scale testing of cattle and deer with subsequent identification of M. bovis in both populations within this area. The spread of M. bovis in Michigan was slowed by a strict policy of total herd depopulation upon identification of positive cattle, as well as large-scale hunter education programs and a massive testing initiative in WT deer. Still, in Michigan, over 650 cases of M. bovis infection in WT deer and 49 positive cattle herds have been identified to date. Alarmingly, M. bovis has now spread to other states. M. bovis was recently detected in 27 WT deer and 12 cattle herds in Minnesota and has been confirmed in cattle from Colorado, Nebraska, Indiana, Kentucky, North Dakota, South Dakota, New Mexico, and California. Detection of M. bovis infection has lead to quarantine and depopulation of nearly all affected herds. Clearly, this disease is continuing its resurgence throughout the US, particularly where cattle and WT deer commingle.


A second major source of M. bovis infected cattle in the US is imported animals from other countries where the disease is endemic, particularly Mexico. Indeed, molecular epidemiology studies have demonstrated that M. bovis cases in all states other than Michigan are likely of Mexican origin. Although USDA regulations stipulate that imported cattle must be tested within 60 days of import, the low sensitivity of most approved M. bovis diagnostic tests suggests that some infected animals will be missed.   Because cattle are only held at the border for 48 to 72 hours, there is little time to conduct additional testing at the point of entry. In addition, the lack of mandatory animal identification in the US limits the ability to track cattle after introduction into the country. Clearly, it is crucial to have rapid diagnostics with improved sensitivity that could be deployed at points of entry. It is equally important to improve information on cattle movements to control importation of M. bovis infected cattle.



  1. bovis is of significant concern to government agencies and cattle industries due to associated economic, social and potential public health problems. The inclusion of M. bovis research, teaching and extension in this multi-state project will address serious concerns from cattle industry representatives, government agencies, and public health officials that the US is experiencing a resurgence of M. bovis that will have devastating economic effects, cause a disruption or severe restrictions in movements of cattle including exports, and have profound effects on producers, who own positive herds and must suffer depopulation or quarantine.


Finally, the generation of new knowledge relative to the diagnosis, management and control of mycobacterial diseases of animals is critical if we are to prevent the spread, lower the prevalence and minimize the impact of the diseases in our livestock populations. USDA NAHMS studies and other work, including the National Dairy Producer Johne’s survey, have shown that while producers are increasingly aware of the diseases, they often lack knowledge relative to their management and control. Therefore, there is a critical need for developing coordinated approaches for education and outreach programs related to mycobacterial diseases of animals.


Taken together, the proposed multi-state initiative described below will facilitate the development of shared research as well as the leveraging of intellectual and physical resources to address some of the most important mycobacterial diseases of animals.


 


Related Current and Previous Work


In terms of prior and current related work, during the fall of 2004, the USDA-CSREES-NRI’s Coordinated Agricultural Projects (CAP) helped bring together leading scientists in the field of JD to form a comprehensive, multi-institutional, interdisciplinary Johne’s Disease Integrated Program for research, education, and extension, or JDIP. We started with a team of approximately 70 scientists from two-dozen leading academic and government institutions in the US, who represented the diverse disciplines of microbiology, immunology, pathology, molecular and cellular biology, genomics, proteomics, epidemiology, clinical veterinary medicine, public health, extension, and public policy. Since inception, membership in MDA has grown to more than 220, and the program has become international in scope.


Based on the success of the program, JDIP was renewed in 2008, and the Multistate Initiative program in the Mycobacterial Diseases of Animals-Multistate initiatives enabled JD research, education, and extension to rapidly move forward in a manner that would not be possible through traditional funding mechanisms from the USDA. In particular, the founding and continued support of MDA has enabled the community, for the first time since JD was described more than a century ago, to develop an integrated and coordinated program with a focus on developing a strong translational pipeline of new diagnostic tests, vaccine candidates, strategies to manage, prevent and control the disease, and the formulation of an outstanding education and training program. As detailed in the sections below, in the brief period since the founding of the program, JDIP investigators have conducted path-breaking research and development that has resulted in:


 



  • A better understanding of paratuberculosis on-farm transmission dynamics that is helping identify critical control points in the transmission chain.

  • The development of alternative sampling and testing strategies for detection of infected animals and herds that are being adopted by the national voluntary control program for JD.

  • The optimization and standardization of laboratory protocols for paratuberculosis culture and PCR for reducing timelines for rapid and sensitive detection of infected animals.

  • Characterization of genetic differences between isolates of paratuberculosis for molecular epidemiologic analyses and tracking of strains in infected animals and the environment.

  • Development of standards for animal challenge models with paratuberculosis for the evaluation of vaccine efficacy.

  • Identification of key genes, proteins and lipids unique to paratuberculosis for development of the next generation of diagnostic tests and vaccines.

  • Development and widespread use of an on-line JD veterinary certification program.

  • Development of educational modules for producers as well as field and laboratory technicians providing milk ELISA tests for producers.

  • Development of community resources including paratuberculosis isolates, serum samples and other clinical material for the development and validation of diagnostic tests, genomic microarrays, recombinant proteins, and mutant strain banks of M. paratuberculosis for identification of potential vaccine candidates.

  • Development an individual-based dairy herd model by incorporating basic herd dynamics in a closed herd environment where no new animals have been bought from outside.

  • Development of a useful platform for gene discovery and analysis by isolating three novel mutants for each transposon.

  • Establishment of high quality longitudinal data collection which turned out to be an essential tool in our understanding of pathobiology and epidemiology of MAP infections in dairy herds

  • Development of a peptide-based vaccine for cattle using the PLGA NP delivery systems

  • Evaluation of the Bovine Leukemia Virus and Mycobacterium avium subsp. paratuberculosis relationship with Shiga Toxin-Producing Escherichia coli Shedding in Cattle

  • Evaluation of the humoral immunity and atypical cell-mediated immunity in response to vaccination in cows naturally infected with bovine leukemia virus

  • Screen the bovine serum samples with MTB and MAP protein microarray for antigen discovery

  • Research on the evaluation of prevention of infection by stimulating innate response using Mycobacterium bovis as the model of infection.

  • Establishment of model systems that can be used to obtain crucial information that would unveil key aspects of MAP pathogenesis, and would enable the researchers to compare the different phases of the disease between in vitro and in vivo systems.

  • Determining the role of luxR homolog gene in invasion of MAP into epithelial cells using Mycobacterium smegmatis as a model of infection.

  • Investigation of the phenotypic diversity in the immune response against Mycobacterium avium paratuberculosis in MAP-infected dairy cows.

  • Identification of several candidate MAP proteins of potential utility for the early detection of MAP infection.

  • Detection of pathogens and control pathogen transmission, both within-herd transmission and between-herd transmission.

  • Development of a quantitative methodology for incorporating whole genome sequence (WGS) data into bacterial transmission models for infectious diseases incorporating ecology, economics, molecular biology, and epidemiology.

  • Better understanding of the principles and dynamics governing transmission of mycobacterial infection.

  • Development, assessment, and implementation of vaccines for JD and bTB.

  • Providing veterinarians, producers of potentially impacted species, state and federal policy makers, and other stakeholders with accurate, high quality, up to date, and easy to access information and education to assist efforts that will effectively address mycobacterial diseases.


 


In addition to our research accomplishments, we have developed a strong communications and extension plan that includes workshops, newsletters, regular conference calls, and an annual conference of JD researchers. Hence, MDA has brought together scientists and stakeholders with a shared vision and well-defined plan to support and facilitate research, extension and education activities and enhance animal health through biosecurity by addressing well-documented and emerging needs in JD.


Workshop on Accelerating bovine Tuberculosis (bTB) Control in Developing Countries


With funding from Bill & Melinda Gates Foundation, The University of Georgia, Cornell University, and The


Pennsylvania State University, a Workshop on Accelerating bovine Tuberculosis (bTB) Control in Developing Countries was conducted on December 8-10, 2015 in Rabat, Morocco. The workshop was a representation of the collective efforts of a committed and diverse global group of bTB experts who convened to develop a shared vision and forward-looking research agenda for developing and implementing effective bTB control strategies in developing countries.


The workshop was co-chaired by Vivek Kapur (Penn State, US), Martin Vodermeier (Animal and Plant Health Agency, UK), Yrjo Grohn (Cornell, US), and Fred Quinn (UGA, US). The workshop brought together a diverse group of 40 leading bTB investigators from 16 countries, which worked with policy makers and funding agency representatives to develop a shared vision and strategic framework for the implementation of bTB control programs in developing countries in which the disease is endemic in livestock, humans, and wildlife.


Participant presentations and discussions provided key insights on seven topical areas including: (1) vaccines and diagnostics, (2) the zoonotic impact of bTB, (3) bTB control efforts that have worked to date, (4) the World Organization for Animal Health (OIE) perspective, (5) the African perspective, (6) implications of bTB in wildlife, and (7) the India and China perspectives. Participants generated an initial 175 insights and 154 questions through discussions, and an “idea sorting” round-robin exercise worked to enhance the robustness of the knowledge base and identify the top five most critical insights and questions for each topic area. The group developed an integrated strategy map and detailed five-year action plan to help meet these three key inter-dependent and inter-related needs: (i) Establishment of the business case through rigorous bTB risk and economic impact assessments and the development of advocacy tools for bTB control programs, (ii) Establishment of technical capabilities to ensure the widespread availability of and access to fit-for purpose diagnostic tests and vaccines, (iii) Establishment of key market and public investment operational drivers and the creation of value-chain for bTB control by small-holder farmers. 


Hence, the renewed multi-state proposal seeks to continue to build the considerable progress we have made during the past two phases of JDIP so that we can continue to leverage the financial and scientific resources even after the completion of the second Phase of the program. We are convinced that the accomplishments of this CAP project thus far have created a momentum that will continue to grow through the proposed multi-state initiative that expands the focus from JD to include TB and mycobacterial diseases in animals.

Objectives

  1. Objective 1 will focus on understanding the epidemiology and transmission of JD and TB in animals through the application of predictive modeling and assessment of recommended control practices.
    Comments: To accomplish our overall objective of developing a better understanding of the epidemiology and transmission of JD and TB.
  2. Objective 2 will seek to develop and implement new generations of diagnostic tests for JD and TB.
    Comments: Improved methods for the rapid, specific, sensitive, and cost-efficient diagnosis of JD or TB-infected remain a major priority.
  3. Objective 3 will focus on improving our understanding of biology and pathogenesis of Mycobacterial diseases, as well as the host response to infection
    Comments: It is well recognized that the ability to identify the route of invasion and the host-pathogen interactions at a molecular level is important for the future development of strategies to prevent infections or to limit the spread of the infection. Similarly, the elucidation of gene products specific to in vivo growth holds great promise in identifying new antigens for diagnostics or vaccine development, as well as products essential to pathogenesis. Hence, as part of the proposed multi-state initiative, we envision studies of the basic biology of the causative organisms of JD and TB and their interaction with the host. Specifically, we anticipate studies that will employ state-of-the-art microbiological, molecular biology, genomic, proteomic, metabolomic, immunology, and or bioinformatic approaches.
  4. Objective 4 will focus on development of programs to create and evaluate and develop new generations of vaccines for JD and TB.
    Comments: Under the auspices of this multi-state initiative, we propose specific research projects to help achieve each of the 4 objectives and include a strong education and extension plan. We envision many of the projects to be crosscutting in nature (i.e. cut across objectives and/or address both diseases) that will together help address the major animal, human, and societal issues surrounding detection and control of mycobacterial diseases in animals. It is important to note that our research objectives are closely linked and coordinated with our education, extension and outreach plan.

Methods

Objective 1 will focus on understanding the epidemiology and transmission of Mycobacterial diseases in animals. To accomplish our overall objective of developing a better understanding of the epidemiology and transmission of JD and TB, we propose studies that include:

  • Continued development of mathematical models of JD and TB transmission dynamics, including within-host, between individuals, within and between domesticated dairy and beef herds and wildlife, as well as on an ecological scale. For example, several investigators have initiated the process of development of mathematical models for JD and TB (2-4) and we will continue the process with studies such as estimating the performance of JD vaccines, defining the impact of wildlife infection on JD and TB dynamics, analyzing the spread of JD and TB through cattle trading networks, and finding economically optimal JD and TB control strategies.  Examples of the types of investigations that will be carried out are presented in(5-7).
  • Characterization of herd and environmental distribution of specific genotypes of paratuberculosis and M. bovis using state-of-the-art methods for strain differentiation using simple sequence repeats and or single nucleotide-based typing approaches and applying this knowledge to characterize the genetic diversity and molecular epidemiology of M. paratuberculosis and M. bovis infections;
  • Delineation of mycobacterial disease transmission dynamics, including paratuberculosis transmission within calf-rearing systems, risk of M. paratuberculosis transmission from infected dams to daughters, and risk of M. paratuberculosis infection associated with ‘super-shedders’ and calf-to-calf transmission;
  • Clarification and delineation of critical management practices for control, prevention, and eradication of mycobacterial diseases; and,
  • Identification and optimization of surveillance methods and strategies.

Taken together, these studies will significantly advance our understanding of the epidemiology and transmission dynamics of mycobacterial diseases of animals.

Objective 2 will seek to develop and implement new generations of diagnostic tests for JD and TB. Improved methods for the rapid, specific, sensitive, and cost-efficient diagnosis of JD or TB infected remain a major priority. Hence, as part of this multi-state initiative, we anticipate carrying out investigations that include:

  • Development of methods for the early detection of paratuberculosis and M. bovis infected animals, including newer generations of molecular, serological and microbiological assays with greater sensitivity, specificity, speed, and or ease-of-use, by using state-of-the art molecular biological, immunological, and materials science and engineering methods and approaches; and,
  • Development of resources for validation and standardization of diagnostic assays, including well-accessioned biological sample collections (strains, tissue, clinical samples, etc.), and processes to make these accessible to the scientific community.

Together, these studies and efforts will facilitate the development, validation, and implementation of the next-generation of improved diagnostic tests for mycobacterial diseases of animals.

Objective 3 will focus on improving our understanding of biology and pathogenesis of Mycobacterial diseases of animals, as well as the host response to infection. Our understanding of the basic biology and mechanisms of pathogenesis of M. paratuberculosis and M. bovis is far from complete. It is well recognized that the ability to identify the route of invasion and the host-pathogen interactions at a molecular level is important for the future development of strategies to prevent infections or to limit the spread of the infection. Similarly, the elucidation of gene products specific to in vivo growth holds great promise in identifying new antigens for diagnostics or vaccine development, as well as products essential to pathogenesis.

Hence, as part of the proposed multi-state initiative, we envision studies of the basic biology of the causative organisms of JD and TB and their interaction with the host.  Specifically, we anticipate studies that will employ state-of-the art microbiological, molecular biology, genomic, proteomic, metabolomic, immunology, and or bioinformatic approaches to carry out studies that include:

  • Investigations into the basic mechanisms of pathogen invasion of host cells and tissue using state-of the art methods in mycobacteriology, cell biology, and genomics;
  • Identification of mycobacterial genes and proteins whose inactivation or alternated expression results in reduced virulence. This will be accomplished by screening large libraries of mutants, as well as by characterizing these mutant strains using state-of-the art genomics and proteomics based methods and will also lead to the identification of genes associated with the ability of the pathogen to survive in the host as markers for virulence and pathogenicity; and,
  • Characterization of the microbial factors that contribute to the innate and adaptive immune response using sophisticated in vitro cellular immunologic assays and animal models of infection.
  • Exploitation of knowledge from immune response studies to create new methods of diagnosis.

Taken together, we anticipate that these investigations will reveal important insights on the basic biology of the causative organisms of JD and TB and their interaction with their hosts.

Objective 4 will focus on the evaluation and development of new generations of vaccines for JD and TB. It is well recognized that defining the host genetic, cellular and molecular events associated with susceptibility to JD and TB is essential for the development of candidate vaccines and host genetic selection for resistance. For TB in particular, the experience in the UK and elsewhere have shown that traditional test/slaughter and abattoir inspection campaigns fail to control the spread of bovine TB (bTB), most likely due to the presence of a wildlife reservoir. Vaccine research must become a priority. Similarly, in the US where a wildlife reservoir exists, control efforts have not eradicated bTB and are unlikely to do so. Hence, the development of a vaccine against bTB is required to control disease. Under the auspices of this multi-state initiative, we envision projects that will seek to develop candidate vaccines, identify genes and markers associated with susceptibility of animals to mycobacterial infection, and define the cellular and molecular events associated with development of immune responses to M. paratuberculosis and M. bovis in cattle. Specifically, we anticipate the development of projects that will:

  • Analyze the early immune response to infection as well as the host response to animals at different stages of disease using well-characterized in vitro models and animal experimentation;
  • Develop and validate animal models for vaccine development;
  • Identify genetic markers for susceptibility to infection in cattle using genome wide association studies with well-defined resource populations. A combination of candidate gene identification with whole genome SNP typing promises to rapidly identify a set of markers that could be used to select for resistance to disease caused by mycobacteria;
  • Compare the efficacy of candidate vaccines in animal models of infection. We hypothesize that live attenuated vaccines are likely to elicit a protective response superior to the response elicited by currently available killed vaccines. However, it will be essential to develop vaccine candidates that are able to differentiate vaccinated from naturally infected animals. To test this hypothesis, we anticipate studies that include: (a) use of flow cytometry, long-oligo microarrays, and real time RT-PCR to compare immune responses elicited by candidate mutant vaccines; (b) Determine if mutant vaccines elicit development of effector memory CD4 and/or CD8 T cells that kill infected autologous macrophages or arrest replication of intracellular bacteria; and, (c) Determine if animal immunized with mutant vaccines are protected against challenge;
  • Evaluate the ability of recombinant or vector expressed proteins and mycobacterial lipids to elicit effector T cells with the capacity to kill infected macrophages or arrest replication of intracellular bacteria. The working hypothesis is that modification of mycobacterial antigens by attachment of Trojan peptides will selectively enhance development of long-lived memory CD4 and/or CD8 effector T cells and may be suitable candidate antigens for use as subunit vaccines; and,
  • Determine the role of regulatory T cells in the immunopathogenesis of mycobacterial infections in animals. The working hypothesis is that dysregulation of the immune response to paratuberculosis and M. bovis is, at least in part, attributable to development of regulatory T cells (Tregs). Evidence suggests that Tregs may be responsible for down-regulating effector memory CD4 cells in an antigen-specific manner. This hypothesis will be tested by characterizing cell surface markers of Tregs using flow-cytometeric and expression analysis techniques.

 Taken together, we anticipate that these investigations will reveal important insights into the immune response of animals to mycobacterial infections, as well as lead to the identification and evaluation of candidate vaccines.

Measurement of Progress and Results

Outputs

  • The outputs, including research data, methods Comments: a. A better understanding of the epidemiology and transmission of JD and TB in animals, and the development of predictive models of infection; b. New generations of diagnostic tests for JD and TB that are sensitive, specific, rapid, and cost-efficient; c. Improved understanding of the biology and pathogenesis of mycobacterial diseases of animals, as well as the host response to infection; d. Development and evaluation of new generations of vaccines for JD and TB; e. Development of shared resources and protocols; and, f. Development of education materials and delivery plan to provide veterinarians, producers of potentially impacted species, state and federal policy makers and other stakeholders with accurate, high quality, up to date, and easy to access information related to mycobacterial diseases of animals.

Outcomes or Projected Impacts

Milestones

(0):We anticipate the following programmatic milestones. a. Each of the four objectives and the outreach and education plan will start during year 1 and continue through the duration of the project. b. An annual meeting of investigators. c. During year 3, working in concert with our stakeholders, we anticipate carrying out a needs assessment for both the research and outreach components of the program. d. Year 4 will involve a comprehensive evaluation of progress of the multi-state initiative, and focus on developing renewal application.

Projected Participation

View Appendix E: Participation

Outreach Plan

We recognize and appreciate that outreach and education efforts are vital components in achieving the objectives of this multi-state initiative, as described above. The underlying mission of our outreach plan is to provide veterinarians, producers of potentially impacted species, state and federal policy makers, and other stakeholders with accurate, high quality, up to date, and easy to access information and education to assist efforts that will effectively address mycobacterial diseases. To accomplish this, we need to better understand the factors that encourage or deter veterinarians and their producer clients from adopting JD and TB control or eradication practices, as well as the educational needs of these populations, to develop educational materials based on current, evidence-based information and deliver these materials in a flexible, convenient, cost-effective, and readily available manner.


As part of our education and outreach plan for this multi-state initiative, we envision using currently available and relevant information as well as generating new information.   In addition, we will seek to use and enhance existing information distribution systems but also develop new tools for this purpose. The following are objectives for the education and outreach component of this multi-state initiative:



  • Create an internet portal to provide access to information related to mycobacterial diseases, specifically JD and bTB. Internet access provides the most rapid, cost effective means to sharing information with a widely distributed audience. The site will provide convenient access to information generated through the initiative and seek to be as comprehensive as possible by sharing previously developed information through links to existing sites such as jdip.org, www.johnes.org, and www.johnesdisease.org. Links to international sites will allow US scientists, producers, and policy makers access to information on the success of domestic herd and wildlife control programs, such as the badger vaccination program in Ireland. These sites already exist and are supported from various extramural and intramural sources, and we anticipate that we will continue to seek funding for the development, management, and curation of these web-sites.

  • Encourage, monitor and increase awareness of the publication of work of initiative collaborators in peer-reviewed journals and through other scientific outlets. Publication of research results in peer reviewed journals is important to the initiative and to those who collaborate in the effort, since it validates the credibility of the work and makes it more widely available. The Education/Outreach team will strongly encourage publication of initiative research in appropriate journals. We will seek to make others in the industry aware of work as it is published and also monitor the publications for work that may be shared with producers and others through the initiative. Current Johne’s efforts have developed a strong international network of scientists and interested professionals, through the International Association for Paratuberculosis (IAP), who are effectively sharing information as they work to address this world-wide disease. Efforts in other nations are also looking to address a wider range of mycobacterial diseases, so this initiative will fit well into expanding international efforts. We will seek to maintain and enhance current working relationships and explore new ones that will allow the most effective use of existing resources.

  • Enhance and strengthen working relationships and communication links with producer and professional organizations. While many good working relationships currently exist, expanding these networks will increase awareness of the initiative, build confidence in the results and help to make them more readily available to our target audiences. It is anticipated that activities in this area will include:

    1. Partnering with the Animal Health committee for the Joint Annual Meeting (JAM) of the American Dairy Science Association and the American Society of Animal Science to include specific oral and poster presentation sections for mycobacterial diseases at the JAM. Include, as appropriate, mycobacterial sessions/symposia in the scientific sessions of the American Association of Bovine Practitioners (AABP), the Association of Veterinary Consultants (AVC) and the American Veterinary Medical Association (AVMA). This will provide an opportunity to reach large and very important target audiences in a cost effective manner. It will also assure inclusion of abstracts of the work presented in highly respected journals that are readily available nationally and internationally.

    2. Holding “Interest Group” meetings at the JAM, the annual meeting of the American Association of Bovine Practitioners (AABP), the Association of Veterinary Consultants (AVC) and similar meetings to reach extension and industry professionals with interests in this area by providing them with information from the initiative, seeking input on current and planned activities, and inviting their participation in the initiative.

    3. Coordinate preconference seminars, or clinical forums, on a periodic basis at the annual conference of the AABP to reach professionals who are on the farm with timely information and solicit their input on additional needs that the initiative is equipped to address.

    4. Facilitate discussion with government and industry to consider expansion of the National Johne’s Work Group (NJWG), currently a subcommittee of the US Animal Health Association (USAHA)’s Johne’s Disease Committee, to become a Mycobacterial Disease Work Group, working with the Tuberculosis and other appropriate USAHA committees. It is anticipated that this group would meet annually at the USAHA’s annual meeting and “as needed” at the annual meeting of the National Institute for Animal Agriculture (NIAA) to share information and identify additional research and education needs.

    5. Partner with relevant organizations in organizing scientific and educational information sessions for producers focused on relevant topics. Potential collaborators include:

      1. NCBA Cattlemen’s College

      2. National Dairy Herd Information Association (NDHIA)







  • World Dairy Expo



  1. The Joint Annual Meeting of the National Milk Producers Federation (NMPF), the National Dairy Board (NDB) and the United Dairy Industry Association (UDIA)

  2. Dairy and beef breed associations

  3. The American Farm Bureau Federation (AFBF)



  1. Partner with USDA to assist in training programs on related diseases

  2. Organize, with industry, extension, and government agency collaboration, a national symposium on mycobacterial diseases of animals every five years

  3. Develop and conduct webinar’s on “high interest” topics in conjunction with extension and or other industry partners



  • Provide convenient access to comprehensive, high quality, and consistent education materials for veterinarians, producers and others. We will seek out and use existing tools, such as those currently available at http://ce.vetmed.wisc.edu/Johnes_Disease,that are developed and reviewed by experts in the field. Additional information that is needed will be identified and resources/collaborators needed to produce and deliver the material will be identified. Materials will be delivered electronically, but will include supporting material that can be printed locally.

  • Leverage existing information/education delivery mechanisms to more comprehensively reach target audiences with information about mycobacterial diseases. We will work actively with trade media and partner with groups like the Johne’s Education Initiative (JEI), DAIReXNET, the eXtension Wildlife Damage Management Community of Practice, and the Internet Center for Wildlife Damage Management (ICWDM) in this effort.

  • Reach non-traditional audiences, including policy makers and interested members of the public, with accurate and timely information relative to mycobacterial diseases in livestock and serve as a point of contact for further information needs. Social media tools such as “Linked In” and “Facebook” will be used to reach these audiences. We will seek to partner with and draw on expertise from industry groups to make the most effective use of these tools in a timely manner as this effort moves forward.


  • ICP – Coauthored presentation on JD programs in the U.S.


    2016 JAM




  • Annual Meeting – MDA interest session, material available in press room and registration




  • World Dairy Expo – met with 10 dairy trade publications, material available




  • USAHA – Display and presentations to JD Committee, State, extension and Federal vets


    Sample


 Taken together, the above approach will help us achieve our objectives of providing veterinarians, producers, and other stakeholders with high quality, up-to-date information and education to foster a cost-effective approach of managing JD and TB risk and preventing and controlling mycobacterial diseases in animals.

Organization/Governance

We build on our experience with the JDIP and TB-CAP initiatives and have formulated a robust plan for the administration of the multi-state initiative.


In brief, we have proposed the formation of an Executive Committee that will be responsible for all strategic, scientific, and management policy decisions for this multi-state initiative, and serve to advise the Administrative Advisor of the program. The Chair of the Executive Committee is responsible for the implementation and facilitation of programmatic goals and will serve as the primary liaison with the USDA, Experiment Station Directors, and external stakeholders. We also propose the formulation of an External Advisory Board, which will consist of public and private stakeholders (regulatory agencies, members of industry, and prominent scientists from related disciplines and Experiment Station Directors), to provide advice on programmatic matters, and ensure that the initiative stays true to its mission. The Chair of the External Advisory Board will be a member of the Executive Committee. The composition, membership and voting structure of the Executive Committee is described below:


Executive Committee. The initial Executive Committee will be comprised of a total of nine members, representing individuals with leadership in Mycobacterial disease research, extension, and education, a documented commitment to helping the community realize a shared vision, and a history of working together as a team. The proposed members of the Executive Committee are:



  • John Bannantine (National Animal Disease Center, USDA-ARS).

  • Luiz Bermudez (Oregon State University, OR).

  • Paul Coussens (Michigan State University, MI).

  • Ian Gardner (University of Prince Edward Island, Canada).

  • Yrjö Gröhn (Cornell University, NY).

  • Vivek Kapur (Penn State, PA). Initial Chair of the multi-state initiative.

  • Don Lein. (Cornell University, NY). Initial Chair of the External Advisory Board.

  • Kenneth Olson (KEO Consulting, IL).

  • Scott Wells (University of Minnesota, MN).


 


Governance of the Executive Committee:


 



  1. Chair: The chair of the committee is responsible for organizing the meeting agenda, conducting the meeting, and assuring that task assignments are completed. The chair will be elected for at least a two-year term to provide continuity and be eligible for reelection.


 



  1. Chair-elect: The chair-elect will succeed the chair, and is expected to support the chair by carrying out duties assigned by the chair. The chair-elect serves as the chair in the absence of the elected chair. Normally the chair-elect is elected for at least two years and be eligible for reelection.


 



  1. Secretary: The secretary is responsible for the distribution of documents prior to the meeting and is responsible for keeping the minutes, preparing the accomplishments report (i.e., the SAES-422). The secretary will succeed the chair-elect and be eligible for reelection.

  2. Responsibilities of the Executive Committee. The Executive Committee is responsible for the overall management and administration of the program and will make all responsible efforts to achieve unanimous consent or make decisions through simple majority vote. The executive committee may appoint sub-committees (that may comprise of any member of the multi-state initiative) whenever needed in order to make flexible and informed decisions and provide guidance to the program chair and executive committee and will nominate and vote on the composition of the external advisory board.


 


Program Members: In addition to carrying out the agreed research collaboration, research coordination, information exchange, or advisory activities, project members are responsible for reporting progress, contributing to the ongoing progress of the activity, and communicating their accomplishments to the committee’s members and their respective employing institutions.

Literature Cited

Below are our 2016 publications:


1. Al-Mamun, M. A., Smith, R. L., Schukken, Y. H. and Grohn, Y. T. (2016) Modeling of Mycobacterium avium subsp. paratuberculosis dynamics in a dairy herd: An individual based approach. J Theor Biol 408: 105-117.


2. Bannantine, J. P., Lingle, C. K., Adam, P. R., Ramyar, K. X., McWhorter, W. J., Stabel, J. R., Picking, W. D. and Geisbrecht, B. V. (2016) NlpC/P60 domain-containing proteins of Mycobacterium avium subspecies paratuberculosis that differentially bind and hydrolyze peptidoglycan. Protein Sci 25: 840-851.


3. Beaver, A., Cazer, C. L., Ruegg, P. L., Grohn, Y. T. and Schukken, Y. H. (2016) Implications of PCR and ELISA results on the routes of bulk-tank contamination with Mycobacterium avium ssp. paratuberculosis. J Dairy Sci 99: 1391-1405.


4. Beaver, A., Ruegg, P. L., Grohn, Y. T. and Schukken, Y. H. (2016) Comparative risk assessment for new cow-level Mycobacterium avium ssp. paratuberculosis infections between 3 dairy production types: Organic, conventional, and conventional-grazing systems. J Dairy Sci.


5. Capsel, R. T., Thoen, C. O., Reinhardt, T. A., Lippolis, J. D., Olsen, R., Stabel, J. R. and Bannantine, J. P. (2016) Composition and Potency Characterization of Mycobacterium avium subsp. paratuberculosis Purified Protein Derivatives. PLoS One 11: e0154685.


6. Espejo, L. A., Zagmutt, F. J., Groenendaal, H., Munoz-Zanzi, C. and Wells, S. J. (2015) Evaluation of performance of bacterial culture of feces and serum ELISA across stages of Johne's disease in cattle using a Bayesian latent class model. J Dairy Sci 98: 8227-8239.


7. Franklin, R. K., Marcus, S. A., Talaat, A. M., KuKanich, B. K., Sullivan, R., Krugner-Higby, L. A. and Heath, T. D. (2015) Correction to: ''A Novel Loading Method for Doxycycline Liposomes for Intracellular Drug Delivery: Characterization of In Vitro and In Vivo Release Kinetics and Efficacy in a J774A.1 Cell Line Model of Mycobacterium smegmatis Infection". Drug Metab Dispos 43: 1805.


8. Ghosh, P., Shippy, D. C. and Talaat, A. M. (2015) Superior protection elicited by live-attenuated vaccines in the murine model of paratuberculosis. Vaccine 33: 7262-7270.


9. Hempel, R. J., Bannantine, J. P. and Stabel, J. R. (2016) Transcriptional Profiling of Ileocecal Valve of Holstein Dairy Cows Infected with Mycobacterium avium subsp. Paratuberculosis. PLoS One 11: e0153932.


10. Kincaid, V. A., London, N., Wangkanont, K., Wesener, D. A., Marcus, S. A., Heroux, A., Nedyalkova, L., Talaat, A. M., Forest, K. T., Shoichet, B. K. and Kiessling, L. L. (2015) Virtual Screening for UDP-Galactopyranose Mutase Ligands Identifies a New Class of Antimycobacterial Agents. ACS Chem Biol 10: 2209-2218.


11. Krueger, L. A., Beitz, D. C., Humphrey, S. B. and Stabel, J. R. (2016) Gamma delta T cells are early responders to Mycobacterium avium ssp. paratuberculosis in colostrum-replete Holstein calves. J Dairy Sci 99: 9040-9050.


12. Krueger, L. A., Reinhardt, T. A., Beitz, D. C., Stuart, R. L. and Stabel, J. R. (2016) Effects of fractionated colostrum replacer and vitamins A, D, and E on haptoglobin and clinical health in neonatal Holstein calves challenged with Mycobacterium avium ssp. paratuberculosis. J Dairy Sci 99: 2884-2895.


13. Kugadas, A., Lamont, E. A., Bannantine, J. P., Shoyama, F. M., Brenner, E., Janagama, H. K. and Sreevatsan, S. (2016) A Mycobacterium avium subsp. paratuberculosis Predicted Serine Protease Is Associated with Acid Stress and Intraphagosomal Survival. Front Cell Infect Microbiol 6: 85.


14. Li, L., Katani, R., Schilling, M. and Kapur, V. (2016) Molecular Epidemiology of Mycobacterium avium subsp. paratuberculosis on Dairy Farms. Annu Rev Anim Biosci 4: 155-176.


15. Magombedze, G., Eda, S. and Koets, A. (2016) Can Immune Response Mechanisms Explain the Fecal Shedding Patterns of Cattle Infected with Mycobacterium avium Subspecies paratuberculosis? PLoS One 11: e0146844.


16. Magombedze, G., Eda, S. and Stabel, J. (2015) Predicting the Role of IL-10 in the Regulation of the Adaptive Immune Responses in Mycobacterium avium Subsp. paratuberculosis Infections Using Mathematical Models. PLoS One 10: e0141539.


17. Marcus, S. A., Sidiropoulos, S. W., Steinberg, H. and Talaat, A. M. (2016) CsoR Is Essential for Maintaining Copper Homeostasis in Mycobacterium tuberculosis. PLoS One 11: e0151816.


18. Marcus, S. A., Steinberg, H. and Talaat, A. M. (2015) Protection by novel vaccine candidates, Mycobacterium tuberculosis DeltamosR and DeltaechA7, against challenge with a Mycobacterium tuberculosis Beijing strain. Vaccine 33: 5633-5639.


19. McDaniel, M. M., Krishna, N., Handagama, W. G., Eda, S. and Ganusov, V. V. (2016) Quantifying Limits on Replication, Death, and Quiescence of Mycobacterium tuberculosis in Mice. Front Microbiol 7: 862.


20. McDonald, J. L., Bailey, T., Delahay, R. J., McDonald, R. A., Smith, G. C. and Hodgson, D. J. (2016) Demographic buffering and compensatory recruitment promotes the persistence of disease in a wildlife population. Ecol Lett 19: 443-449.


21. Mitachi, K., Sharma Gautam, L. N., Rice, J. H., Eda, K., Wadhwa, A., Momotani, E., Hlopak, J. P., Eda, S. and Kurosu, M. (2016) Structure determination of lipopeptides from Mycobacterium avium subspecies paratuberculosis and identification of antigenic lipopeptide probes. Anal Biochem 505: 29-35.


22. Otsubo, S., Cossu, D., Eda, S., Otsubo, Y., Sechi, L. A., Suzuki, T., Iwao, Y., Yamamoto, S., Kuribayashi, T. and Momotani, E. (2015) Seroprevalence of IgG1 and IgG4 class antibodies against Mycobacterium avium subsp. paratuberculosis in Japanese population. Foodborne Pathog Dis 12: 851-856.


23. Peterz, M., Butot, S., Jagadeesan, B., Bakker, D. and Donaghy, J. (2016) Thermal Inactivation of Mycobacterium avium subsp. paratuberculosis in Artificially Contaminated Milk by Direct Steam Injection. Appl Environ Microbiol 82: 2800-2808.


24. Pomeroy B, Sipka A, Klaessig S, Schukken Y (2016) Longitudinal characterization of bovine monocyte-derived dendritic cells from mid-gestation into subsequent lactation reveals nadir in phenotypic maturation and macrophage-like cytokine profile in late gestation. J Reprod Immunol 118: 1-8.


25. Rathnaiah, G., Bannantine, J. P., Bayles, D. O., Zinniel, D. K., Stabel, J. R., Grohn, Y. T. and Barletta, R. G. (2016) Analysis of Mycobacterium avium subsp. paratuberculosis Mutant Libraries Reveals Loci-dependent Transposition Biases and Strategies to Novel Mutant Discovery. Microbiology.


26. Ribeiro-Lima, J., Carstensen, M., Cornicelli, L., Forester, J. D. and Wells, S. J. (2016) Patterns of Cattle Farm Visitation by White-Tailed Deer in Relation to Risk of Disease Transmission in a Previously Infected Area with Bovine Tuberculosis in Minnesota, USA. Transbound Emerg Dis.


27. Ribeiro-Lima, J., Schwabenlander, S., Oakes, M., Thompson, B. and Wells, S. J. (2016) Risk profiling of cattle farms as a potential tool in risk-based surveillance for Mycobacterium bovis infection among cattle in tuberculosis-free areas. J Am Vet Med Assoc 248: 1404-1413.


28. Roussey, J. A., Oliveira, L. J., Langohr, I. M., Sledge, D. G. and Coussens, P. M. (2016) Regulatory T cells and immune profiling in johne's disease lesions. Vet Immunol Immunopathol.


29. Shaughnessy, R. G., Farrell, D., Riepema, K., Bakker, D. and Gordon, S. V. (2015) Analysis of Biobanked Serum from a Mycobacterium avium subsp paratuberculosis Bovine Infection Model Confirms the Remarkable Stability of Circulating miRNA Profiles and Defines a Bovine Serum miRNA Repertoire. PLoS One 10: e0145089.


30. Slater, N., Mitchell, R. M., Whitlock, R. H., Fyock, T., Pradhan, A. K., Knupfer, E., Schukken, Y. H. and Louzoun, Y. (2016) Impact of the shedding level on transmission of persistent infections in Mycobacterium avium subspecies paratuberculosis (MAP). Vet Res 47: 38.


31. Smith, R. L., Grohn, Y. T., Pradhan, A. K., Whitlock, R. H., Van Kessel, J. S., Smith, J. M., Wolfgang, D. R. and Schukken, Y. H. (2016) The effects of progressing and nonprogressing Mycobacterium avium ssp. paratuberculosis infection on milk production in dairy cows. J Dairy Sci 99: 1383-1390.


32. Smith, R. L., Schukken, Y. H. and Grohn, Y. T. (2015) A new compartmental model of Mycobacterium avium subsp. paratuberculosis infection dynamics in cattle. Prev Vet Med 122: 298-305.


33. Wanzala, S. I., Nakavuma, J., Travis, D. A., Kia, P., Ogwang, S. and Sreevatsan, S. (2015) Draft Genome Sequences of Mycobacterium bovis BZ 31150 and Mycobacterium bovis B2 7505, Pathogenic Bacteria Isolated from Archived Captive Animal Bronchial Washes and Human Sputum Samples in Uganda. Genome Announc 3.


34. Winthrop, K., Rivera, A., Engelmann, F., Rose, S., Lewis, A., Ku, J., Bermudez, L. and Messaoudi, I. (2016) A Rhesus Macaque Model of Pulmonary Nontuberculous Mycobacterial Disease. Am J Respir Cell Mol Biol 54: 170-176.


35. Wolf, T. M., Sreevatsan, S., Singer, R. S., Lipende, I., Collins, A., Gillespie, T. R., Lonsdorf, E. V. and Travis, D. A. (2016) Noninvasive Tuberculosis Screening in Free-Living Primate Populations in Gombe National Park, Tanzania. Ecohealth 13: 139-144.

Attachments

Land Grant Participating States/Institutions

IL, MI, NE, PA

Non Land Grant Participating States/Institutions

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