Duration: 10/01/2001 to 09/30/2006

Administrative Advisor(s):

NIFA Reps:

Non-Technical Summary

Statement of Issues and Justification

Bovine respiratory disease (BRD) is one of the most important disease problems facing the cattle industry. This respiratory disease complex caused by a variety of bacteria and viruses results in losses of over $3 billion to the US cattle economy annually (Kapil and Basaraba, 1997). NC-107 Bovine Respiratory Disease (BRD) Committee has a long and productive history in BRD research including the development of diagnostic methods and vaccines. The importance of BRD research was recently reaffirmed by stakeholders at the USDA-CREES Stakeholder Priorities Workshop for Animal Agriculture, Nutrition and Food Safety held in Washington DC in December 1999. In a list of high priority research areas presented by the Animal Agriculture Coalition, bovine respiratory disease and two bovine respiratory viruses, bovine respiratory syncytial virus (BRSV) and bovine viral diarrhea virus (BVD) were listed. In the Executive Summary of the Protection of Animal Health Session of the Workshop, these diseases along with developing new diagnostics, increasing surveillance and elucidating microbe genetics were key points of Stakeholder input for research areas. Understanding BRD pathogenesis, patterns and developing new diagnostic and control approaches are the heart of NC-107 committee efforts.

BRD continues to be major problem in spite of 40 years of control measures using antibiotics and vaccination. Even with up to 50% of all animals being vaccinated, BRD lesions are seen in 30-90% of cattle at slaughter. Failing to continue a BRD research program will result in more severe losses to BRD as no new information on pathogens or pathogenesis would be available. This coupled with the need to minimize antibiotic usage for prevention and control of BRD would make beef production more expensive and the consumption of beef prohibitive to many because of the increased cost of beef.

BRD is a complicated and incompletely defined syndrome and is best described as a disease complex. Numerous factors are important in BRD including environmental components (stressors), infectious agents (viral, bacterial, mycoplasmal and chlamydial agents) and unique anatomical and physiological and immunological aspects of the bovine respiratory tract. These factors interact in an extremely complex and intricate manner resulting in BRD. Developing approaches to control BRD requires a coordinated research effort investigating multiple aspect of this disease complex. Areas requiring further study include identifying and characterizing emerging and reemerging pathogens through the improvement of surveillance and detection methods, defining mechanisms and intervention targets in pathogenesis of BRD at the molecular, cellular and host level and developing intervention strategies for critical control points to reduce impact of BRD.

This must be done while ensuring that control strategies contribute to the production of wholesome food.

This NC-107 project addresses the seven cross cut priorities and thirteen objectives developed by the North Central Directors in the Crosscutting Research Priorities. This research project will enhance beef and dairy production; provide genetic information on disease pathogens; develop detection and control strategies to decrease disease incidence; decrease the spread of disease to wildlife; decrease antibiotic usage and antibiotic resistance; develop new disease detection and control technologies; provide important examples of applications of biotechnology for rural America; and enhance food safety by decreasing antibiotic usage and finding antimicrobial activity from phytochemicals. The Specific Crosscutting priorities and objectives addressed are: 1) Agricultural Production, Processing and Distribution, Objective 2 -Develop improved animal, plant and microbial production, processing and marketing systems that are competitive, profitable and environmentally sound over the long term; and Objective 5 -Assemble and maintain regional, national and international data bases on production systems and use them for modeling and decision support; 2) Genetic Resources Development and Manipulation (Genomics and Germplasm), Objective 2- Broaden and enrich the knowledge base about genomics; Objective 3- Collect, preserve, share, enhance and evaluate germplasm at the molecular, cellular and/or organismal levels; and Objective 5- Develop increased knowledge of the interactions and interrelationships of the various life forms. 3) Integrated Pest Management, Objective 1- Develop alternative controls based on biological control and cultural practices; Objective 2 -Investigate the genetics of pests and hosts to identify new and different vulnerabilities that can be exploited in pest control strategies; and Objective 4 -Refine and develop rapid and positive pest detection and identification techniques to enhance the capability to predict the occurrence and magnitude of pest populations/infestations/infection. 4) Natural Resources and the Environment, Objective 6 - Understand and identify factors that influence the ecological relationships among production agriculture, wildlife management and human health. 5) Economic Development and Policy, Objective 1 - Develop profitable technologies and systems. 6) Social Change and Development, Objective 3 - Determine barriers to use of appropriate technologies and increase the adoption of environmentally, socially and economically sustainable agricultural and community practices; evaluate social impacts of technological changes on rural residents. 7) Food and Nutrition, Objective 3 - Enhance food safety by expanding research efforts to identify and control food borne pathogens at all stages of the food system from producer to consumer and to develop and evaluate effective food safety programs for both producers and consumers; and Objective 5 -Elucidate health benefits associated with functional or phytochemical properties of food constituents.

The NC-107 Committee consists of veterinary clinicians, virologists, bacteriologists, epidemiologists, immunologists and pathologists. The expertise of the committee ranges from clinical trials to cellular immunology to molecular pathogenesis. This group has tremendous breadth in basic and applied bovine research. All the methodology described in the proposal is within the scope of this group of researchers. There are many long-standing collaborations between many of the investigators. There are three large feedlot multistate studies with each study involving multiple stations and researchers. The complicated nature of BRD research and the cost of cattle studies make it impossible for a single Agricultural Experiment Station to have the expertise or resources to investigate BRD. This project allows individual expertise of each station to be coordinated in a cooperative effort that will maximize the BRD research effort. Many benefits will be realized from the new project. The new project will generate data on pathogen patterns and better diagnostic methods that can be used for better treatment and control measures. Understanding the mechanisms used by the various agents of BRD to persist on the host, and produce inflammatory and immune responses are needed for the characterization of novel intervention targets that can minimize the use of antibiotics. Specific targets to be studied will be the ability to block latency of bovine herpesvirus 1 (BHV-1), replication of bovine viral diarrhea virus (BVDV), attachment of Mannheimia haemolytica, as examples of blocking persistence of a pathogen in the host. Blocking of inflammatory response cytokine action on bovine respiratory tissue affected by leukotoxin, as well as blocking of apoptotic effect of Haemophilus somnus on endothelial cells are examples of targeting to the inflammatory/cytotoxic responses of affected cattle. Finally, development of protective immune responses devoid of sensitizing responses is needed to improve current vaccines against bovine respiratory syncytial virus (BRSV) and H. somnus. Protective immune responses that utilize primarily cytotoxic T lymphocytes may be needed to control persistent or latent viral infections. The studies not only have implication for controlling BRD but also may be useful as animal models of comparable human diseases

NC-107 Crosscutting Areas for Regional Research

The NCA-02 assigned the following percentages for the proposed NC-107 project

1. Agricultural Production, Processing and Distribution


2. Genetic Resources Development and Manipulation


3. Integrated Pest Management


4. Natural Resources and the Environment


5. Economic Development and Policy


6. Social Change and Development


7. Food and Nutrition

Related, Current and Previous Work

A search of the CRIS project database using bovine and respiratory and disease as keywords found 24 projects. Eighteen of these projects were with members of the NC-107 committee and related directly to this project. Only one of the remaining 6 projects dealt with basic research with BRD and it was at an NC-107 experiment station. There is no duplication between, this project and existing USDA funded research.

The focus of this proposal on bovine respiratory disease includes a broad perspective of the causes of this disease. This is required in part by the often complex etiology involved, and by the stated goals, of identifying multiple emerging and re-emerging agents involved in BRD, characterizing mechanisms, and developing intervention strategies for BRD.

As a top priority disease for US beef cattle producers, the impact of BRD on production has been the subject of assessment by NAHMS surveys of beef cattle producers.¹ Recent estimates of the losses caused by BRD place the production value lost at over 3 billion dollars annually.² Other studies have focused on lung lesions at slaughter as an index of BRD loss, and a recent study indicated that even though 29% of beef cattle were treated for BRD in feedlots, and 35% of beef cattle were treated for BRD during all stages of growth, these treatments appeared inadequate for controlling production losses associated with BRD.³ The widespread use of vaccines for the prevention of viral and bacterial agents is also well documented for beef cattle,⁴ and appears to also have only partially beneficial effects.³ Beyond the measurement of production losses, the quantification of functional losses associated with BRD is also undergoing change,⁴ with findings that implicate subclinical disease in production losses. These findings imply that much work has still to be done to fully control production losses caused by BRD.

Other related work is providing genomic information about important bacterial pathogens in BRD. A recently completed sequencing effort has produced the full genome sequence of avian- pathogenic Pasteurella multicida, and this, together with sequencing efforts (both supported by USDA-NRICGP) on the M. haemolytica genome will provide comprehensive databases from which to extend many aspects of our proposal. The generation of isogenic mutants of M haemolytica,⁵ and the demonstration that leukotoxin mutants of M haemolytica are apathogenic⁶ were also important milestones in the development of a knowledge base about the pathogenesis of BRD. Research on respiratory pathogens conducted in other continents also provides context. For example, the finding that BRSV can produce severe respiratory disease in adult cattle in England⁷ underscores the need for strain characterization as well as understanding of virulence mechanisms of important viruses involved in BRD.

Notable among novel intervention strategies has been the use of metaphylaxis targeted towards M. haemolytica.⁹ Other strategies use novel manipulations to redirect immune responses,⁹ a concept that is also employed for several pathogens in our current proposal.

Occurrence studies performed by stations in the NC-107 project have provided invaluable baseline information on which to assess emergence or re-emergence of pathogens. For example, identification of the serotypes and biotypes ofM haemolytica and P. multocida ¹⁰ leads to studies on the emerging role ofM. haemolytica of serotypes other than Al, and the increasing frequency of P. multocida presentations. Recognition of bovine respiratory coronavirus as an agent of BRD,¹¹ is expanded to further surveys as well as to a genome characterization project. The emerging role of Mycobacterium bovis in regional outbreaks of respiratory disease has prompted studies on the nature of the bovine immune response to this pathogen.¹² Other emerging diseases are associated with host responses to a pathogen, and a model of vaccine- enhanced BRSV disease¹³ will be used to study acute interstitial pneumonia under experimental and field conditions. Since H. somnus bacterin was shown to play a role in enhanced H. somnus disease,¹⁴ this will also be further investigated.

Studies on the pathogenesis of viral infections in BRD are evolving from a focus on receptors and cell tropism, through effects on host cells, to effects on host organs and systems and immune response components. The tyrosine kinase cell signaling pathway used by BHV-1¹⁵ is a potential intervention site using the phytochemical, genistein, will be further investigated. Findings on the latency mechanisms of BHV-1¹⁶ as well as novel sites of latency¹⁷ will be fully exploited in future work to develop intervention targets. Studies on the thrombocytopenic effect of BVDV virus indicated that both numbers as well as functionality of platelets were impaired,¹⁸ and these effects require further characterization. Models to study local lung and systemic immune responses to BRSV have been developed,¹⁹ and studies to extend these findings to severe disease forms as well as to concurrent infection models are planned.

The leukocyte receptor for the M. haemolytica leukotoxin was identified and characterized through a cooperative effort of several stations in the project.²⁰ In addition, cytokine induction by leukotoxin effect on bovine macrophages was characterized.²¹ These efforts are now the basis for a systematic effort at developing these receptors as intervention targets in the new proposal. The detection of antimicrobial anionic peptides in the lung of sheep was reported,²² and this work has led to plans to use a sheep model to study the role of antimicrobial peptides in M. haemolytica pneumonia.

Cooperative studies on treatment and control of BRD have been completed in several stations. Recombinant BHV-1 vaccines have been tested,²³ and will undergo more complete evaluation. Several cooperative studies have been completed on the immunogenicity of BVDV vaccines.²⁴ These have been complemented with virus biotyping studies,²⁵ These studies will project into various field studies designed to measure protective efficacy of BVDV vaccines using new test methodologies. The effect of novel antimicrobials on reduction of horizontal spread ofM haemolytica infection was studied.²⁶ Integration of metaphylactic and vaccination protocols has then been proposed and is currently under study. In particular, it is considered important to establish if metaphylaxis is a sustainable management practice, and if it has effects on normal flora that may be of concern. Comparison of aerosol and injectable vaccines for M haemolytica has also been proposed, as a follow-up of extensive studies on nasal immunity to M. haemolytica infection.

Objectives

1. Identify emerging and re-emerging agents and develop diagnostic methods for BRD.
   
2. Characterize mechanisms and intervention targets in pathogenesis of BRD at the molecular, cellular and host level
   
3. Develop intervention strategies for critical control points to reduce impact of BRD
   
4. 
   

Methods

Objective 1

This objective has two components-identifying emerging and re-emerging agents and developing diagnostic methods for BRD. MN and SD will collect and summarize BRD pathogen data from the veterinary diagnostic laboratories (AL, CA, CO, GA, IL, IA, KS, MN, MS, OK, SD, TX, WI). These states account for more than 85% of all cattle fed in the US. OK will collect isolates from BRD cases from large scale field trials (with TX, LA and NADC) and will characterize strain/serotype differences. These agents will include M. haemolytica, P. multocida, H. somnus, BVDV, BHV and bovine adenoviruses. OK will determine the prevalence of bacteria and viruses involved in transported cattle with BRD morbidity/mortality (in collaboration with TX and NADC). SD and LA will optimize antemortem testing methods for bovine respiratory viruses and M haemolytica respectively. SD and AL will screen and type BVDV field isolates (in collaboration with NE). NADC and MN will type H. somnus and M. haemolytica field isolates (in collaboration with MI, IA and SD). MS will perform diagnostic workups on chronic BRD to identify factors responsible for poor therapeutic response. GA will undertake a study of bacterial and viral pathogens associated with acute interstitial pneumonia (AIP) in 3 western US feedlots.

The development of new BRD diagnostic methodologies include MI with a cowside "dipstick" BVDV test (in collaboration with AL), LA and KS with bovine respiratory coronavirus (in collaboration with OK and TX), GA and CA with BRSV and IA with M. bovis (in collaboration with MN and SD). IA will develop ribotyping ofM bovis isolates for epidemiological tracing and for diagnostic identification. IA will develop serological methods to assess M. bovis infection status among acutely or endemically infected herds. IA will facilitate with MN, NADC, OK, LA, TX the use of this ribotyping technology for M haemolytica, P. multocida, H. somnus. SD (in collaboration with MI and AL) will develop "molecular beacon" BVDV diagnostic testing. SD will develop BVDV signature sequences using multiplex PCR (in collaboration with AL) to differentiate vaccine and field BVDV strains. NE will validate the BVDV skin immunohistochemistry detection test and determine if the test can be used to differentiate between vaccination and field infections. The NC-107 group will present these BRD diagnostic approaches in a symposium to be held in conjunction with Academy of Veterinary Consultants in December 20001.

Objective 2

NE will continue investigations on the mechanism by which BHV-1 gives rise to latent infections. KS and NE will determine the molecular basis of differential pathogenesis in bovine herpesvirus types 1 and 5 infections. The high degree of homology between BHV-1 and BHV-5 offers a powerful system to analyze functional domains of the constituent glycoproteins. SD will continue work on cell signaling with BHV-1 and BVDV and will also work on the interactions of macrophages with BHV-1 and BVDV. WI will continue work on the role of BHV-1 glycoproteins in pathogenesis and immunity. NE will characterize down-regulation of MHC class I molecules by BHV-1 infection, which result in reduced cytotoxic lymphocyte responses. This work will be complemented with continuing efforts at development of epitope vaccines based on directing cytotoxic lymphocyte responses towards complexes of BHV-1 glycoprotein peptides and host heat shock protein. KS will collaborate with LA on bovine immunodeficiency virus (BIV) and Jembrana disease virus (JDV) investigations. Both bovine lentiviruses are closely related genetically, and a unique capsid protein epitope of BIV will be studied.

GA will carry out a study of risk factors for feedlot acute interstitial pneumonia (AIP) in 3 western US feedlots. CA will cooperate in this project studying multi-etiology factors such as mycotic challenge combined with BRSV infection, BVDV and BRSV infection, and H. somnus plus BRSV infection. GA will continue to collaborate with CA on studies of the cellular and molecular events associated with protection against severe disease due to BRSV. GA will evaluate cytokine expression in lung and regional lymphoid tissue. CA will continue studies on immune regulation and antigen presentation in the lung of BRSV-infected vs normal calves. CA will also study factors involved in protective vs sensitizing responses, as well as immunological memory in BRSV.

MI will determine the effect of BVDV-persistently infected calves on performance of calves in a feedlot (in collaboration with AL), and continue studies together with KS to determine the mechanism by which acute BVDV infection induces thrombocytopenia. KS will study apoptotic effects of BVDV, and SD (in collaboration with AL) will identify virulence-associated BVDV genome regions. MI will determine the mechanism of virulence in acute BVDV infection (in collaboration with NADC), and will study correlates of viremia and immune responses with KS. NE will utilize engineered BVDV genomes expressing reporter tags in conjunction with gene expression profiling methods (bovine microarrays) to elucidate the molecular basis of BVDV virulence. MI will also characterize mucosal immunity to BVDV in the respiratory tract and investigate the effects of BVDV infection on endocrine function. OK (in collaboration with NADC) will examine the role of a specific BVDV envelope protein on the host immune defense, and NE will identify candidate gene correlates of protective immunity to BVDV in cattle. KS will continue exploring the use of transgenic plants to immunize against BVDV. CA and GA plan to use methodologies to evaluate protective vs sensitizing responses to BVDV infection.

OK will further investigate mechanisms of bacterial pathogen adherence to the respiratory tract, including effects of viruses and different strains ofM haemolytica from sheep and cattle. Genes for virulence factors ofM haemolytica and P. multocida will be identified by OK and KS. These two stations will continue to investigate the role of oxygen concentration on the quantity of M haemolytica leukotoxin production. Co-regulation of P. multocida capsule expression and a membrane lipoprotein will continue to be studied by MS. This station will also evaluate dam gene regulation of P. multocida virulence. WI will evaluate a DNA vaccine using the leukotoxin gene.

MN and WI will continue to investigate molecular and cellular events by M. haemolytica leukotoxin induces the host inflammatory response leading to lung injury. In particular, the role ofLFA-1 and Gi-coupled signaling receptors of bovine leukocytes will be defined. Studies on M. haemolytica leukotoxin and LPS, and their interactions on bovine leukocytes will continue in several stations. The role of bovine CD 18 as a receptor M. haemolytica leukotoxin will be investigated further by MN. Host responses to gene modified bacteria (isogenic mutants jointly developed by NADC and Texas Health Center) will be compared to wild-type strains by MN and WI. IA will develop an ovine model to characterize production of inducible and non-inducible antimicrobial peptides by respiratory epithelial cells. IA will also determine mast cell-substance P fiber axis activity and its relationship to antimicrobial peptide production during the development ofM haemolytica pneumonia in vivo. The effect of sub-MIC concentrations of antibiotics M. haemolytica and H. somnus will continue to be studied by MS. MS will also evaluate immunomodulatory effects of various hormones, combination of hormones, or other immunomodulatory products to see if they have potential for vaccine enhancement or boosting of innate resistance to BRD agents.

WI will investigate the interaction of H. somnus lipooligosaccharide with bovine endothelial cells, in particular, the role of purinergic receptor activation on endothelial apoptosis. IA will characterize molecular events involved in systemic M. bovis infection, and correlate these with virulence genes of the mycoplasma, and will participate in a genome sequencing project to characterize virulence genes of M bovis.

From the Objective 2 studies, specific intervention targets are highlighted for further characterization. Specific types of immune responses will be targeted for BHV-1 (cytolytic by NE), and BRSV (protective rather than sensitizing by GA and CA). Host resistance to viral infection will also be studied (for BVDV by NE). Competitive binding with anti-leukotoxin receptors will be approached by MN, and similar competition with H. somnus lipooligosaccharide receptors on endothelial cells will be pursued by WI. Signal transduction pathways of neutrophils and macrophages reacted with leukotoxin will be evaluated as targets by MN. In addition, inflammatory cytokine expression in lung will be targeted for down-regulation in studies by MN. TX will also study anti-inflammatory agents to understand their effect on BRD-associated inflammation.

Objective 3

Methods: With the identification of co-mingling as a primary management control point, the influence of different methods and treatments will be examined to identify specific management practices that will significantly reduce bovine respiratory tract disease. This will be done by promoting high health to reduce disease risks, vaccination and follow-up surveillance serology to reduce risk, improved disease recognition, evaluation of vaccination programs prior to co-mingling, administration of immune modulators, biosecurity measures, and metaphylaxis. The evaluation and incorporation of novel vaccination strategies will be a primary focus of this production period. Emphasis will be on preventive medicine. OK, TX, USDA/NADC will evaluate the ability of modified live viral and bacterial (M haemolytica) vaccines to reduce bovine respiratory disease in transported stocker/feeder cattle as well as in cattle entering the feedlot by determining morbidity and morality and treatment and prevention costs. The utility of an intranasal BRSV vaccination will be evaluated by GA. OK, AL, and MI in collaboration with USDA/NADC, and TX will investigate management factors from individual herds contributing to bovine respiratory disease as cattle will be followed from the brood cow operations, through the feedyard, and eventually by slaughter check for lung/liver lesions including transfer of maternal immunity; nutrition, postweaning management and vaccines. The concept of pathogen reduction will be testing by MI and AL in determining the outcome of co-mingling with calves persistently infected with bovine viral diarrhea virus. MI will evaluate the optimal time of vaccine administration in preconditioning programs and determine the effect of BVDV PI calves on performance of calves in the feedlot. In addition, specific and non specific host factors will be examined. TX will evaluate the genetic basis of interferon induction and action in cattle and host response to modifiers such as cytokines and adjuvants. TN and TX will characterize the micro- environment response to various novel agents. MS and TX will evaluate hormonal and immunomodulatory compounds and oral vaccination with live M. haemolytica organisms and M. haemolytica leukotoxin.

In line with concerns of increasing antimicrobial sensitivity, the practice of metaphylaxis will be examined. These practices will be evaluated by AL and MI in relationship to potential antimicrobial resistance by characterizing the effect of antimicrobial use on the genetic diversity of nontarget (non-respiratory pathogens) organisms. MI and AL will investigate metaphylaxis and its effect on antimicrobial susceptibility of nontarget bacteria. TX will improve the effectiveness and safety of metaphylactic treatment by examination of timing of administration and the effect on antimicrobial resistance in the feedyard.

Measurement of Progress and Results

Outputs

• See attached "Measurements of Progress and Results"
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Outcomes or Projected Impacts

Milestones

(0):0

Projected Participation

View Appendix E: Participation

Outreach Plan

The results of this project will be published in peer reviewed journals. A BRD symposium will be held in Year 1 and results will also be presented in newsletters, veterinarian and producer meetings and lay publications.

Organization/Governance

The Research Technical Committee shall consist of one technical committee representative from each cooperating agency as appointed or otherwise designated by the respective organization, an administrative advisor appointed by the Association of North Central Experiment State Directors, and a representative of the Cooperative State Research, Education and Extension Service (CSREES).


The executive committee shall consist of a chair, chair-elect and secretary, two additional technical committee members plus administrative advisor and representative of CSREES. Members of the executive committee are elected annually and may succeed themselves. Chair, chair-elect and secretary will serve a three-year succession term beginning with secretary in year 1, chair-elect in year 2 and chair in year 3.



Meetings will be held annually at the time and place mutually agreed upon by the technical committee, or as designated by the executive committee with the approval of the administrative advisor.



The secretary will normally record the minutes of the annual meeting. Two copies of the minutes with original signature of recommendation by the secretary will be sent to the Administrative Advisor with an approval block for his/her signature. The Administrative Advisor will distribute copies to appropriate individual.



The chairperson will normally prepare the annual report summarized from material supplied to him/her by the technical committee member from each participating station or agency. The chairperson will send the final draft of the annual report to the Administrative Advisor for his/her approval. The Administrative Advisor will assume responsibility for its distribution.



Coordination states are identified for the cooperative procedures within each objective. Overall coordinators for each objective are identified. The executive committee is responsible for an overall coordination of the project. Such coordination will occur continually throughout the project period. The annual meeting of the Technical Committee will serve as a monitoring session for coordination.

Literature Cited

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2. Kapil S, Basaraba RJ. Infectious bovine rhinotracheitis, parainfluenza-3, and respiratory coronavirus. Bovine respiratory disease update. In Clinics of North America: Large Animal 1997; 13:455-469.


3. Wittum TE, NE Woollen, LJ Perino ET Littledike. 1996. Relationship among treatment for respiratory tract disease, pulmonary lesions evident at slaughter, and rate of weight gain in feedlot cattle. JAVMA 209:814-818.


4. Cough J, C Uystepruyst, F Butreau, ML van de Weerdt, P Lekeux. New techniques to evaluate functional impact of bovine respiratory disease. In; 1998 Proceedings of the Association of Bovine Practitioners, pp 247-249, Spokane, WA.


5. Fedorova ND, SK Highlander. 1997. Generation of targeted nonpolar gene insertions and operon fusions in Pasteurella haemolytica and creation of a strain that produces and secretes inactive leukotoxin. Infect Immun. 65:2593-2598.


6. Highlander SK, ND Fedorova, DM Dusek, R Panciera, LE Alvarez, C Rinehart. 2000. Inactivation of Pasteurella (Mannheimia) haemolytica leukotoxin causes partial attenuation of virulence in a calf challenge model. Infect Immun. 68:3916-3922.


7. Elvander M. 1996. Severe respiratory disease in dairy cows caused by infection with bovine respiratory syncytial virus. Vet. Rec. 138:101-105.


8. Young C. 1995. Antimicrobial metaphylaxis for undifferentiated bovine respiratory disease. The Compendium on Continuing Education for the Practicing Veterinarian 17:133-142.


9. Freidag BL, GB Melton, F Collins, DM Klinman, A Cheever, L Stobie, W Suen, RA Seder. 2000. CpG oligonucelotides and interleukin-12 improve the efficacy of Mycobacterium bovis BCG vaccination in mice challenged with M. tuberculosis. Infect. Immun. 68:2948-2953.


10. Purdy CW, RH Raleigh, JK Collins, JL Watts, DC Straus. 1997. Serotyping and enzyme characterization of Pasteur ella haemolytica and Pasteurella multocida isolates recovered from pneumonic lungs of stressed feeder calves. Curr. Microbiol. 34:244-249.


11. Stortz J, CW Purdy, XQ Lin, M Burrell, RE Truax, RE Briggs, GH Frank, RW Loan. 2000. Isolation of bovine respiratory coronavirus, other cytocidal viruses, and Pasteurella spp from cattle involved in two natural outbreaks of shipping fever. JAVMA 216:1599-1604.


12. Smith RA, JM Kreeger, AJ Alvarez, JC Goin, WC Davis, DL Whipple, DM Estes. 1999. Role ofCD8+ and WC-1'" gamma/delta T cells in resistance to Mycobacterium bovis infection in the SCID-bo mouse. J. Leuk. Biol. 65:28-34.


13. Gershwin LJ, ES Schlegle, RA Gunther, ML Anderson, AR Woolums, DR Larochelle, GA Boyle, KE Friebertshausen, RS Singer. 1998. A bovine model of vaccine enhanced respiratory syncytial virus pathophysiology. Vaccine 16:1225-1236.


14. Ruby KW, RW Griffith, LJ Gershwin, ML Kaeberle. 2000. Haemophilus somnus-mduced IgE in calves vaccinated with commercial monovalent H. somnus bacterins. Vet. Microbiol. 76:373-383.


15. Shaw, A.M., L. Braun, T. Frew, DJ Hurley, RRR Rowland, CCL Chase. 2000. A role for bovine herpesvirus 1 (BHV-1) glycoprotein E (gE) tyrosine phosphorylation in replication of BHV-1 wild-type virus but not BHV-1 gE deletion mutant virus. Virology 268:159-166.


16. Ciacci-Zanella J, M Stone, C Jones. 1999. The latency related gene of bovine herpesvirus 1inhibits programmed cell death. J. Virol. 73:9734-9740.


17. Winkler MTC, A Doster, C Jones. 2000. Persistence and reactivation of bovine herpesvirus 1 in the tonsils of latently infected calves. J. Virol. 74:5337-5346.


18. Waltz PH, BA Steficek, JC Baker, L Kaiser, TG Bell. 1999. Effect of experimentally induced type II bovine viral diarrhea virus infection on platelet function in experimentally infected calves. AJVR 60:1396-1401.


19. Gershwin LJ, RA Gunther, ML Anderson, AR Woolums, K McArthur-Vaughan, R Kandel, GA Boyle, KE Friebertshauser, PS Mclntuff. 2000. Bovine respiratory syncytial virus specific IgE is associated with IL-2, IL-4, and interferon gamma expression in pulmonary lymph of experimentally infected calves. AJVR 61:291-298.


20. Jeyaseelan S, SL Hsuan, MS Kannan, B Walcheck, JF Wang, ME Kehrii, ET Lally, GC Siek, SK Maheswaran. 2000. Lymphocyte function associated antigen 1 is a receptor for 'Pasteurella haemolytica leukotoxin. Infect. Immun. 68:72-79.


21. Hsuan SL, MS Kannan, YS Prakash, S Jeyaseelan, C Malazdrewich. MS Abrahamsen, GC Stiek, SK Maheswaran. 1999. Pasteurella hemolytica leukotoxin and endotoxin-induced cytokine gene expression in bovine alveolar macrophages requires calcium elevation and NF-kB translocation. Microb. Pathog. 26:263-273.


22. Brogden KA, MR Ackermann, KM Huttner. 1998. Detection of anionic antimicrobial peptides in ovine bronchoalveolar lavage fluid and respiratory epithelium. Infect. Immun. 66:5948-5954.


23. Belknap EB, LM Walters, CL Kelling, VK Ayers, J Norris, J McMillen, C, Hayhow, M. Cochran, DN Reddy, J Wright, JK Collins. 1999. Immunogenicity and protective efficacy of a gE, gG, and US2 gene-deleted bovine herpesvirus 1 (BHV-1) vaccine. Vaccine 17:2297-2305.


24. Fulton RW, LJ Burge. 2000. Bovine viral diarrhea virus type 1 and 2 antibody response in calves receiving modified live virus or inactivated vaccines. Vaccine 19:264-274.


25. Fulton RW, JM d'Offay, JT Saliki, LJ Burge, RG Helman, AW Confer, SR Bolin, JF Ridpath. 1999. Nested reverse transcription-polymerase chain reaction (RT-PCR) for typing ruminant pestiviruses: bovine viral diarrhea viruses and Border disease virus. Can. J Vet Res 63:276-281.


26. Frank GH, RE Briggs, RW Loan, CW Purdy, ES Zehr. 2000. Effects of tilmicosin treatment on Pasteurella haemolytica organisms in nasal secretion specimens of calves with respiratory tract disease. AJVR 61:525-529.

Attachments

Land Grant Participating States/Institutions

AL, CA, IA, KS, LA, MI, MN, MO, MS, OK, SD, WI

Non Land Grant Participating States/Institutions

USDA/NADC