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ARS Home » Plains Area » Clay Center, Nebraska » U.S. Meat Animal Research Center » Animal Health Genomics » Research » Research Project #432111

Research Project: Genomic Intervention Strategies to Prevent and/or Treat Respiratory Diseases of Ruminants

Location: Animal Health Genomics

2017 Annual Report

Objective 1. Elucidate host response associated with the bovine respiratory disease complex (BRDC) and protective immunity, including discovering genetic and biological determinants associated with bovine respiratory disease susceptibility, tolerance, or resistance, and discovering genetic and biologic determinants associated with good responders to bovine respiratory disease vaccines. Sub-objective 1.A: BVD viral infections play an integral and complicated role in BRDC. Current available technology for preventing BVD virus infection includes vaccination, biosecurity, and the elimination of persistently infected cattle. However, if available, genetic selection for animals less likely to become persistently infected would facilitate control and eradication of BVD. The proposed research will test for genetic risk factors associated with BVD virus infection. Sub-objective 1.B: Ovine progressive pneumonia is one of the most economically important diseases in sheep. A major gene TMEM154 was recently discovered that influences susceptibility to OPP in sheep. However, there are no ovine cell lines with defined TMEM154 diplotypes available to study OPP virus infection in vitro. The proposed research will develop cell lines to enable the study of TMEM154 variants in OPP virus infection.S Objective 2. Develop genomics-based strategies to control respiratory diseases of ruminants, including identifying antibiotic-resistance genes and other virulence determinants of bacteria that associate with increased BRDC severity, and developing intervention strategies to reduce antibiotic use and BRDC severity based on genetic typing of bacteria and cattle. Sub-objective 2.A: M. haemolytica of North America place into two major genotypes (1 and 2). Genotype 2 associates with BRDC and genotype 1 does not. The proposed research will identify genomic determinants specific to genotype 2 that may lead to intervention strategies that reduce the incidence of BRDC caused by genotype 2 M. haemolytica. Sub-objective 2.B: Current interventions for BRDC in beef calves include vaccination and metaphylactic use of antibiotics. However, if we had knowledge of the disease-causing potential of nasopharyngeal bacteria in calves, alternative interventions could be designed to reduce the impact of BRDC outbreaks. The proposed research is designed to identify genetic and biological determinants that may influence the disease-causing potential of nasopharyngeal bacteria. Sub-objective 2.C: BCV is involved in the etiology of three distinct clinical syndromes: calf diarrhea, winter dysentery with hemorrhagic diarrhea in adults, and respiratory infections in cattle of all ages. The biological mechanisms underlying disease presentation and variation in their severity are not well understood. The proposed research will determine the influence of serum antibodies, virus strain, and co-infection with other respiratory pathogens on BCV disease presentation and severity of disease.

Infectious respiratory diseases of ruminants are a serious health and economic problem for U.S. agriculture. In cattle alone, the costs of bovine respiratory disease complex (BRDC) exceed one billion dollars annually. Therefore, this research focuses primarily on BRDC with an additional component targeting ovine respiratory disease. Our project vision is to reduce the prevalence and severity of respiratory diseases, thereby promoting livestock welfare, enhancing producer efficiency, and reducing antibiotic use. BRDC is a multi-component disease caused by complex interactions among viral and bacterial pathogens, stress and environmental factors, and host genetics. Consequently, we have developed a multi-component approach focused on the host-pathogen interface to study respiratory disease. On the host side, a genome-wide association study will test for genetic risk factors for bovine viral diarrhea (BVD) virus susceptibility. On the bacterial pathogen side, genomics combined with phenomics will identify the spectrum of genetic determinants of M. haemolytica and other bacteria that associate with BRDC. On the viral pathogen side, genomics combined with serology, and microbial diagnostic testing will determine the contribution of bovine coronavirus (BCV) to BRDC. Lastly, novel ovine cell lines will be developed to test host and virus genetic risk factors for ovine progressive pneumonia (OPP). The knowledge gained from this research will be valuable for developing new intervention strategies for controlling BRDC and producing healthier livestock, and could ultimately benefit animals, producers, veterinarians, diagnostic laboratories, pharmaceutical companies, genetic testing laboratories, and regulatory agencies.

Progress Report
Objective 1.1a: We have completed the WGS for each of the 96 cases with a minimum of 11x genome coverage (minimum 40 Gb, average 51 Gb, maximum 104 Gb, SD 11, and 4.9 Tb total sequence). Objective 1.1b: The 2016 lamb crop from the U.S. Meat Animal Research Center (USMARC), Clay Center, Nebraska ovine-progressive pneumonia (OPP)-free flock were tested to determine OPP Virus status and TMEM154 genotype. Individual lambs that were OPPV negative with the TMEM-154 genotypes “1,1”, “2,2” or “3,3”, and “4,4” were selected for terminal tissue collection. Animal use protocols were reviewed by the Center Veterinarian and approved by the U.S. Meat Animal Research Center Animal Care and Use Committee. Objective 2.2a: The genomes of 35 genotype 1 Mannheimia haemolytica isolates have all been sequenced and assembled into complete, error-corrected, circularized chromosomes. All five known subtypes of genotype 1 are well-represented by the sequenced isolates. All 35 isolate chromosomes have been annotated through the National Center for Biotechnology Information (NCBI) prokaryotic genome annotation pipeline and will become publically available through the NCBI website immediately following the publication of a relevant manuscript. Objective 2.2b: The closed genomes of over 20 H. somni, P. multocida and other species have been sequenced and deposited in GenBank, some of which are publically available at Objective 2.2c: Serial blood samples and nasal swabs were collected from 890 beef calves from four herds at predetermined times from birth though the feed yard. Serum anti-bovine coronavirus (BCV) antibody abundance was measured by enzyme linked immuno sorbant assay (ELISA) and BCV and other viral and bacterial bovine respiratory disease (BRD) pathogens were detected by real-time polymerase chain reaction (PCR). Test results were compared among herds, over time, and between calves that did and did not develop bovine respiratory disease complex (BRDC). The associations of various herd and calf factors with test results were also evaluated. In addition, six full-length BCV genomes were sequenced and annotated.

1. Development and publication of a test that identifies strains of Mannheimia haemolytica that cause bovine respiratory disease. M. haemolytica is the principle bacterial species that causes respiratory disease in cattle. However, not all strains associate equally with disease or antibiotic resistance. In collaboration between ARS scientists in Clay Center, Nebraska and the University of Nebraska - Lincoln, a test was developed that distinguishes between strains predominantly associated with bovine respiratory disease and antibiotic resistance and those that associate more as commensals. Available for use without restriction, the test can assist in the detection and control of pathogenic M. haemolytica strains in cattle.

Review Publications
Workman, A.M., Dickey, A.M., Heaton, M.P., Clawson, M.L., Smith, T.P.L. 2017. Complete genome sequences of two genotype A2 small ruminant lentiviruses isolated from infected U.S. sheep. Genome Announcements. 5(13):e00109-17.
Capik, S.F., White, B.J., Lubbers, B.V., Apley, M.D., DeDonder, K.D., Larson, R.L., Harhay, G.P., Chitko-McKown, C.G., Harhay, D.M., Kalbfleisch, T.S., Schuller, G., Clawson, M.L. 2017. Comparison of the diagnostic performance of bacterial culture of nasopharyngeal swab and bronchoalveolar lavage fluid samples obtained from calves with bovine respiratory disease. American Journal of Veterinary Research. 78(3):350-358.
Clawson, M.L., Murray, R.W., Sweeney, M.T., Apley, M.D., DeDonder, K.D., Capik, S.F., Larson, R.L., Lubbers, B.V., White, B.J., Kalbfleisch, T.S., Schuller, G., Dickey, A.M., Harhay, G.P., Heaton, M.P., Chitko-McKown, C.G., Brichta-Harhay, D.M., Bono, J.L., Smith, T.P.L. 2016. Genomic signatures of Mannheimia haemolytica that associate with the lungs of cattle with respiratory disease, an integrative conjugative element, and antibiotic resistance genes. BMC Genomics. 17:982. doi: 10.1186/s12864-016-3316-8.
Harhay, D.M., Bono, J.L., Smith, T.P., Fields, P.I., Dinsmore, B.A., Santovenia, M., Kelley, C.M., Wang, R., Harhay, G.P. 2016. Complete closed genome sequences of Salmonella enterica subsp. enterica serotypes Anatum, Montevideo, Typhimurium and Newport, isolated from beef, cattle, and humans. Genome Announcements. 4(1):e01683-15. doi:10.1128/genomeA.01683-15.
Nguyen, S.V., Harhay, D.M., Bono, J.L., Smith, T.P.L., Fields, P.I., Dinsmore, B.A., Santovenia, M., Kelley, C.M., Wang, R., Bosilevac, J.M., Harhay, G.P. 2016. Complete, closed genome sequences of 10 Salmonella enterica subsp. enterica serovar Typhimurium strains isolated from human and bovine sources. Genome Announcements. 4(6):e01212-16. doi:10.1128/genomeA.01212-16.
Zhu, L., Workman, A.M., Jones, C. 2017. Potential role for a B-catenin coactivator (high mobility group AT-hook 1 protein) during the latency-reactivation cycle of bovine herpesverus 1. Journal of Virology. 91(5):e02132-16.
Brown, D., Lampe, A.T., Workman, A.M. 2016. The differentiation and protective function of cytolytic CD4 T cells in influenza infection. Frontiers in Immunology. 7:93. doi:10.3389/fimmu.2016.00093
McNeel, A.K., Ondrak, J.D., Amundson, O.L., Fountain, T.H., Wright, E.C., Whitman, K.J., Chitko-McKown, C.G., Jones, S., Chase, C.C., Cushman, R.A. 2017. Timing of transcriptomic and proteomic changes in the bovine placentome after parturition. Theriogenology. 100:1-7. doi: 10.1016/j.theriogenology.2017.05.020.
Loy, J.D., Clawson, M.L. 2017. Rapid typing of Mannheimia haemolytica major genotypes 1 and 2 using MALDI-TOF mass spectrometry. Journal of Microbiological Methods. 136:30-31.