Brief Summary of Minutes of Annual Meeting

Accomplishments: Objective I: Identify reservoirs of infectious respiratory disease agents in wild birds and poultry. 1. Isolation and characterization of avian influenza viruses (AIV) from wild birds and commercial poultry flocks which include live bird markets and backyard flocks were accomplished. The enormous data obtained from different states (AL, CT, DE, MN) were shared. 2. Surveillance activities on the Delmarva Peninsula have yielded infectious laryngotracheitis (LT) virus and infectious bronchitis virus isolates from commercial broiler chickens and Newcastle disease virus isolates from wild birds. 3. In DE, the incidence of LT vaccine reaction and vaccinal LT clinical cases increased in 2010 compared to 2009. No new IBV variants were found during the period. 4. Using gene targeted sequencing and random amplified polymorphic DNA analysis, GA identified the circulation of field strains within complex and companies and analyzed 170 MG and MS strains in 2010. 5. SEPRL (USDA) characterized new avian paramyxovirus isolated from penguins. It was determined that the viruses corresponded to a new serotype (serotype 10). Objective II. Develop improved diagnostic capabilities including real-time PCR as well as other rapid on-farm tests for economically important respiratory diseases. 1. AL developed a method to detect CEO ILTV vaccines in drinking water lines which detects ILTV DNA in the biofilm collected from the water system by real-time PCR. 2. AK and DE used next generation sequencing technologies (Illumina) which permit the relatively rapid determination of the primary sequence of the ILTV genome. AK determined genomes of one wild type and two vaccine ILTV strains. 3. GA developed a multiplex detection of avian influenza HA (H5 & H7) and NA (N1 & N2) subtypes using a microsphere-based assay. 4. GA developed a species-independent competitive ELISA (cELISA) for the detection of influenza A antibodies directed to H6, H7, and H9. 5. GA made H9 specific monoclonal antibodies and further developed H9 subtype specific ELISA systems. 6. IL developed a photolase gene specific PCR. Based on sequence information, avian pox viruses could be differentiated into four different groups. 7. OH developed 19-plex assay which can differentiate different HA subtypes of avian influenza viruses. 8. SEPRL developed an enzyme-linked immunospot assay which can detect avian influenza specific antibody-secreting B cells in chickens. 9. SEPRL identified that the optimal detection methods for avian influenza virus from wild birds depend on the prevalence of virus. Objective III. Investigate the pathogenesis and polymicrobial interactions of specific infectious agents associated with poultry respiratory diseases (this includes interactions with underlying immunosuppressive agents). 1. AL investigated effects of immunosuppressive viruses, chicken anemia virus (CAV) and/or infectious bursal disease virus (IBDV), on evolution of infectious bronchitis virus (IBV). 2. GA conducted comparative genomic analysis of IBV which indicates that the replicase protein in addition to the already recognized spike gene of coronaviuses plays a key role in pathogenicity. GA have identified regions in the replicase that likely effects cleavage and assembly of the enzyme. 3. MN studied the host-pathogen interactions during E. coli infection in the broiler chicken. The genes differentially expressed in air sac tissue did not involve any of the typical APEC virulence factors, and instead involved a large number of chromosome-encoded transport system genes and genes of unknown function. 4. OH studied two isolates of vvIBDV from California which were identified to contain a vvIBDV genome segment A but instead of a serotype 1 vvIBDV genome segment B, their genome segment B was most closely related to a serotype 2 IBDV. 5. OH studied the persistence of classical (STC) and variant (IN) IBDVs. The two strains were detected much longer in bursal tissues (upto 8 weeks) followed by spleen, thymus and bone marrow. In non-lymphoid tissues both strains persisted longer in cecum followed by liver, kidney, pancreas, lungs, thigh and breast muscles. 6. SEPRL demonstrate that the pandemic H1N1 influenza virus does not easily infect young poultry. However, laying turkey hens were susceptible to pandemic H1N1 virus by reproductive tract exposure. 7. SEPRL demonstrated that aMPV-C wild bird isolates induced typical aMPV/C disease in the domestic turkeys. This result suggests that the wild birds may play a role in the spread of the aMPV-C virus. SEPRL also showed that the M2-2 gene is not essential for virus replication in cell culture, but required for efficient virus replication in turkeys to counteract the hosts natural defense and immunity. Objective IV. Develop new prevention and control strategies for poultry respiratory diseases. 1. AL showed for the first time that a DNA vaccine containing an HA gene of an AIV produced cellular immune responses in chickens with a T-helper 1 (Th1) preference. AL also developed an H1 vaccine in transgenic Arabidopsis thallenia. Arabidopsis is a commonly used small weed, whose genome has been sequenced. 2. CT developed nanoparticle-based vaccines carrying M2e of influenza virus and demonstrated the immunogenicity and protection induced by M2e-based vaccine by challenge studies. 3. DE utilizes both traditional and recombinant-based approaches for the construction of the next generation of ILTV live vaccines. 4. GA determined the baseline coverage of four different commercial IBV vaccines (Ark, Mass, GA98 and Mass/Conn) tested at a full dose in 1-day old broilers. 5. GA studied aerosol delivery of a virus-like-particle (VLP) vaccine against H5N1 avian influenza in Poultry which showed for the first time that non-replicating influenza VLPs might be used for mass aerosol vaccination in chickens. 6. IN showed that a prime-boost approach for protection of broiler chickens with maternally derived antibody against IBDV infection by DNA vaccination can be achieved by priming with a high dose of DNA carrying IBDV large segment gene and boosting with a single dose of killed IBD vaccine. 7. IN showed that DNA vaccination confers protection against IBDV challenge by delayed appearance and rapid clearance of the invading viruses. 8. OH used in vitro analysis of virus particle subpopulations in candidate live-attenuated influenza vaccines which could distinguish effective from ineffective vaccines. 9. SEPRL showed that intranasal administration of alpha interferon reduced morbidity associated with low pathogenic avian influenza virus infection. 10. SEPRL demonstrated that commercial influenza vaccines have variable efficacy for protecting chickens and ducks against H5N1 highly pathogenic avian influenza (HPAI) viruses. Work Planned for Next Year 1. Continue surveillance, screening, and characterization of respiratory pathogen from wild and domestic bird populations; 2. Continue development and refinement of diagnostic assays to detect and differentiate poultry pathogens 3. Continue to study polymicrobial infection in poultry using a co-infection model; and continue to study E. coli as a primary or secondary pathogen of poultry; 4. Continue development, refinement and testing of vaccine against influenza, ILT, IBDV, ORT, E. coli, and other respiratory pathogens of poultry. 5. The molecular basis for antigenicity, pathogenicity, and transmission of respiratory pathogens will be studied using naturally occurring viruses and reverse genetically created viruses. 6. Collaborative work will continue with a number of national and international partners.

Publication in Journals: 1. Jindal N, Patnayak DP, Chander Y, Ziegler AF, Goyal SM. 2010. Detection and molecular characterization of enteric viruses from poult enteritis syndrome in turkeys. Poult Sci. 89:217-26. 2. Goyal S, Jindal N, Chander Y, Ramakrishnan M, Redig P, Sreevatsan S. 2010. Isolation of mixed subtypes of influenza A virus from a bald eagle (Haliaeetus leucocephalus). Virology Journal 7:174. 3. Ramakrishnan M, Wang P, Abin M, Yang M, Goyal S, Gramar M, Redig P, Fuhrman M, Sreevatsan S. 2010. Triple reassortment swine influenza A (H3N2) virus in waterfowl. Emerg Infect Dis. 16:728-729. 4. Ladman, B. S., C. P. Driscoll, C. R. Pope, R. D. Slemons, and J. Gelb Jr. Potential of low pathogenicity avian influenza viruses of wild bird origin to establish experimental infections in turkeys and chickens. Avian Diseases 54:10911094. 2010. 5. Spackman, Erica, Jack Gelb, Lauren Preskenis, Brian Ladman, Conrad Pope, Mary Pantin-Jackwood and Enid McKinley. The pathogenesis of low pathogenicity H7 avian influenza viruses in chickens, ducks and turkeys. Virology Journal 7:331 2010. 6. Marcus PI, Ngunjiri JM, Sekellick MJ, Wang L, Lee CW. In Vitro Analysis of Virus Particle Subpopulations in Candidate Live-Attenuated Influenza Vaccines Distinguishes Effective from Ineffective Vaccines. Journal of Virology. 84(21):10974-81. 2010. 7. Yassine HM, Khatri M, Lee CW, Saif YM. Potential role of viral surface glycoproteins in the replication of H3N2 triple reassortant influenza A viruses in swine and turkeys. Vet Microbiol. In Press. 8. Yassine HM, Khatri M, Lee CW, Saif YM. Characterization of an H3N2 triple reassortant influenza virus with a mutation at the receptor binding domain (D190A) that occurred upon virus transmission from turkeys to pigs. Virology Journal. 7(1):258. 2010. 9. Qin Z, Clements T, Wang L, Khatri M, Pillai SPS, Zhang Y, LeJeune JT, Lee CW. Detection of Influenza Viral Gene in European Starlings and Experimental Infection. Influenza and Other Respiratory Viruses. In Press. 10. Pillai SPS, Pantin-Jackwood M, Suarez DL, Saif YM , Lee CW. Pathobiological characterization of low pathogenicity H5 avian influenza viruses of diverse origins in chickens, ducks and turkeys. Arch Virol. 155(9): 1439-51. 2010. 11. W Cha, Y Ma, YM Saif, Lee CW. Development of microsphere-based multiplex branched DNA assay for the detection and differentiation of avian influenza virus. J Clin Microbiol. Vol. 7, no. 48: 2575-2577. 2010. 12. Pillai SPS, Saif YM, Lee CW. Detection of influenza A viruses in eggs laid by infected turkeys. Avian Dis. 54(2):830-3. 2010. 13. Pillai SPS & Lee CW. Species and age related differences in the type and distribution of influenza virus receptors in different tissues of chickens, ducks and turkeys. Virol J. 7:5. 2010. 14. Wang L, Yassine HM, Saif YM, Lee CW. Developing Live Attenuated Avian Influenza Virus In Ovo Vaccines for Poultry. Avian Dis. 54:297301, 2010. 15. Pillai SPS, Pantin-Jackwood M, Yassine HM, Saif YM, Lee CW. The high susceptibility of turkeys to Influenza viruses of different origins implies their importance as potential intermediate hosts. Avian Dis. 54:522526, 2010. 16. Avellaneda G, Mundt E, Lee CW, Jadhao S, Suarez DL. Differentiation of Infected and Vaccinated Animals (DIVA) Using the NS1 Protein of Avian Influenza Virus. Avian Dis. 54:278286. 2010. 17. Avellaneda G, Lee CW, Suarez DL. A Heterologous Neuraminidase Subtype Strategy for the Differentiation of Infected and Vaccinated Animals (DIVA) for Avian Influenza Virus Using an Alternative Neuraminidase Inhibition Test. Avian Dis. 54:272277. 2010. 18. Yassine HM, Lee CW, Gourapura R, Saif YM.. Review of Interspecies and Intraspecies Transmission of InfluenzaViruses: Viral, Host, and Environmental Factors. Animal Health Research Reviews. Vol. 1, no. 11: 53-72. 2010. 19. Abdul Rauf, Mahesh Khatri , Maria V. Murgia, Yehia M. Saif. Expression of perforin-granzyme pathway genes in the bursa of infectious bursal disease virus-infected chickens. Developmental and Comparative Immunology. In Press. 20. Thomas, C., Swayne, D.E. 2009. Thermal inactivation of H5N2 high pathogenicity avian influenza virus in dried egg white with 7.5% moisture. Journal of Food Protection. 72(9):1997-2000. 21. Swayne, D.E., Pantin Jackwood, M.J., Kapczynski, D.R., Spackman, E., Suarez, D.L. 2009. Limited susceptibility of Japanese quail (Coturnix japonica) and resistance of other poultry species to the 2009 novel H1N1 influenza A virus. Emerging Infectious Diseases. 15(12):2061-2063. 22. Spackman, E., Swayne, D.E., Joly, D., Gilbert, M., Karesh, W., Suarez, D.L., Sodnomdarjaa, R., Cardona, C. 2009. Characterization of low pathogenicity avian influenza viruses isolated from wild birds in Mongolia 2005 through 2007. Virology Journal. 6:190-198. 23. Kapczynski, D.R., Swayne, D.E. 2009. Influenza vaccines for avian species. In: Compans, R.W., Orenstein, W.A., editors. Vaccines for Pandemic Influenza, Current Topics in Microbiology and Immunology. Berlin: Springer-Verlag. p.133-152. 24. Petkov, D.I., Linneman, E.G., Kapczynski, D.R., Sellers, H.S. 2009. Identification and characterization of two distinct bursal B-cell subpopulations following infectious bursal disease virus infection of White Leghorn chickens. Avian Diseases. 53(3):347-355. 25. Das, A., Spackman, E., Pantin Jackwood, M.J., Suarez, D.L. 2009. Removal of real-time reverse transcription polymerase chain (RT-PCR) inhibitors associated with cloacal swab samples and tissues for improved diagnosis of avian influenza virus by RT-PCR. Journal of Veterinary Diagnostic Investigation. 21:771-778. 26. Sylte, M.J., Suarez, D.L. 2009. Influenza neuraminidase as a vaccine antigen. In: Compans, R.W., Orenstein, W.A., editors. Vaccines for Pandemic Influenza. New York, NY: Springer. p. 227-242. 27. Jadhao, S.J., Lee, C., Sylte, M.J., Suarez, D.L. 2009. Comparative efficacy of North American and antigenically matched reverse genetics derived H5N9 DIVA marker vaccines against highly pathogenic Asian H5N1 avian influenza in chickens. Vaccine. 27:6247-6260. 28. Pfeiffer, J., Suarez, D.L., Sarmento, L., To, T., Nguyen, T., Pantin Jackwood, M.J. 2010. Efficacy of commercial vaccines in protecting chickens and ducks against H5N1 highly pathogenic avian influenza viruses from Vietnam. Avian Diseases. 54:262-271. 29. Moresco, K.A., Stallknecht, D., Swayne, D.E. 2010. Evaluation and attempted optimization of avian embryos and cell culture methods for efficient isolation and propagation of low pathogenicity avian influenza viruses. Avian Diseases. 54:622-626. 30. Lira, J., Moresco, K.A., Stallknecht, D., Swayne, D.E., Fisher, D.S. 2010. Single and combination diagnostic test efficiency and cost analysis for detection and isolation of avian influenza virus from wild bird cloacal swabs. Avian Diseases. 54:606-612. 31. Kwon, Y., Thomas, C., Swayne, D.E. 2010. Variability in pathobiology of South Korean H5N1 high-pathogenicity avian influenza virus infection for 5 species of migratory waterfowl. Veterinary Pathology. 47(3):495-506. 32. Jadhao, S., Suarez, D.L. 2010. New approach to delist highly pathogenic avian influenza viruses from BSL3+ select agents to BSL2 non-select status for diagnostics and vaccines. Avian Diseases. 54:302-306. 33. Arafa, A., Suarez, D.L., Aly, M.M., Hassan, M.K. 2010. Phylogenetic analysis of hemagglutinin and neuraminidase genes of highly pathogenic avian influenza H5N1 Egyptian strains isolated from 2006 to 2008 indicates heterogeneity with multiple distinct sublineages. Avian Diseases. 54:345-349. 34. Wasilenko, J.L., Sarmento, L., Spatz, S.J., Pantin Jackwood, M.J. 2010. Cell surface display of highly pathogenic avian influenza hemagglutinin on the surface of Pichia pastoris cells using alpha-agglutinin for production of oral vaccines. Biotechnology Progress. 26(2):542-547. 35. Pantin Jackwood, M.J., Wasilenko, J.L., Spackman, E., Suarez, D.L., Swayne, D.E. 2010. Susceptibility of turkeys to pandemic H1N1 virus by reproductive tract insemination. Virology Journal. 7:27. 36. Eggert, D.L., Thomas, C., Spackman, E., Pritchard, N., R0jo, F., Bublot, M.,Swayne, D.E. 2010. Characterization and efficacy determination of commercially available Central American H5N2 avian influenza vaccines for poultry. Vaccine.28:4609-4615. 37. Abbas, M.A., Spackman, E., Swayne, D.E., Ahmed, Z., Sarmento, L., Siddique, N., Naeem, K., Hameed, A., Rehmani, S. 2010. Sequence and phylogenetic analysis of H7N3 avian influenza viruses isolated from poultry in Pakistan 1995-2004. Virology Journal. 7:137. 38. Avellaneda, G.E., Sylte, M.J., Lee, C., Suarez, D.L. 2010. A heterologous neuraminidase subtype strategy for the differentiation of infected and vaccinated animals (DIVA) for avian influenza virus using an alternative neuraminidase inhibition test. Avian Diseases. 54:272-277. 39. Liljebjelke, K.A., Petkov, D., Kapczynski, D.R. 2010. Mucosal vaccination with a codon-optimized hemagglutinin gene expressed by attenuated Salmonella elicits a protective immune response in chickens against highly pathogenic avian influenza. Vaccine. 28(27):4430-4437. 40. Avellaneda, G.E., Mundt, E., Lee, C., Jadhao, S., Suarez, D.L. 2010. Differentiation of infected and vaccinated animals (DIVA) using the NS1 protein of avian influenza virus. Avian Diseases. 54:278-286. 41. Sarmento, L., Wasilenko, J.L., Pantin Jackwood, M.J. 2010. The effects of NS gene exchange on the pathogenicity of H5N1 HPAI viruses in ducks. Avian Diseases. 54:532-537. 42. Suarez, D.L. 2010. Avian Influenza: Our current understanding. Animal Health Research Reviews. 11(1):19-33. 43. Miller, P.J., Decanini, E.L., Afonso, C.L. 2010. Newcastle disease: Evolution of genotypes and the related diagnostic challenges. Infection, Genetics and Evolution. 10(1):26-35. 44. Khan, T.A., Rue, C.A., Rehmani, S.F., Ahmed, A., Wasilenko, J.L., Miller, P.J., Afonso, C.L. 2010. Phylogenetic and pathological characterization of Newcastle disease virus isolates from Pakistan. Journal of Clinical Microbiology. 48(5):1892-1894. 45. Rue, C.A., Susta, L., Brown, C.C., Pasick, J.M., Swafford, S.R., Wolf, P.C., Killian, M.L., Pedersen, J.C., Miller, P.J., Afonso, C.L. 2010. Evolutionary changes effecting rapid diagnostics of 2009 Newcastle disease viruses isolated from Double-crested Cormorants. Journal of Clinical Microbiology. 48(7):2440- 2448. 46. Susta, L., Miller, P. J., Estevez, C., Yu, Q., Zhang, J., Brown, C.C. 2010. Pathogenicity evaluation of different Newcastle disease virus chimeras in 4-week-old chickens. Trop Animal Health Prod. 42(8):1785-95. 47. Miller, P. J., Afonso, C. L., Spackman, E., Scott, M. A., Pedersen, J. C., Senne, D. A., Brown, J. D., Fuller, C. M., Uhart, M. M., Karesh, W. B., Brown, I. H., Alexander, D. J., Swayne, and D. E. 2010 Evidence for a new avian paramyxovirus serotype-10 detected in Rockhopper Penguins from the Falkland Islands. Journal of Virology. 84(21): 11496-11504. 48. Susta, L., Miller, P.J., Afonso, C.L., and Brown, C.C. 2010 Clinicopathological characterization in poultry of three strains of Newcastle disease viruses isolated from recent outbreaks in four-week old SPF Leghorns. Veterinary Pathology. E pub, August 2010. 49. Mesonero, A.*, D. L. Suarez, E. van Santen, D. C. Tang, H. Toro. Avian Influenza In Ovo Vaccination with Replication-Defective Recombinant Adenovirus in Chickens: Vaccine Potency, Antibody Persistence, and Maternal Antibody Transfer Avian Diseases. Submitted 11/22/2010 50. Toro, H,, D. L. Suarez, D. C. Tang, F.W. van Ginkel, C. Breedlove. Avian Influenza Mucosal Vaccination in Chickens with Replication-Defective Recombinant Adenovirus Vaccine Avian Diseases (in press). 51. Toro, H., F.W. van Ginkel D.C. Tang, B. Schemera, S. Rodning, J. Newton (2010). Avian Influenza Vaccination with in chickens and pigs with replication competent adenovirus free human recombinant adenovirus 5. Avian Diseases, Supplement 54: 224-231 52. Toro, H. (2010) Infectious Bronchitis Virus: Dominance of ArkDPI-type Strains in the United States Broiler Industry during the Last Decade Brazilian Journal of Poultry Science 12: 79-86 53. Singh S*, Toro H., Tang DC, Briles WE, Yates LM, Kopulos RT, Collisson EW. (2010). Non-replicating adenovirus vectors expressing avian influenza virus hemagglutinin and nucleocapsid proteins induce chicken specific effector, memory and effector memory CD8(+) T lymphocytes.405: 62-9 54. Gallardo, R.A.*, F.J. Hoerr, W.D. Berry, V.L. van Santen, H. Toro. Infectious Bronchitis Virus in Testicles and Venereal Transmission. Avian Diseases, (submitted 10/29/2010). 55. Gallardo, R. A.*, V. L. van Santen, H. Toro (2010). Host Intraspatial Selection of Infectious Bronchitis Virus Populations. Avian Diseases 54: 807-813 56. Arathy, D.S., Tripathy, D.N., Sabrinath, G.P., Bhaiyat, M.I., Chikweto, A. Mathew, V. and Sharma, R.N. 2010. Preliminary Molecular Characterization of a Fowl Poxvirus Isolate in Grenada. Avian Diseases, 54: 1081-1085 57. Tripathy, D.N. 2010. Fowlpox. Chapter in the Merck Veterinary Manual, Tenth Edition, 2426-2429. 58. Xie, Z., C. Qina, L. Xie, J. Liua, Y. Pang, X. Deng, Z. Xie, M. I Khan. Recombinant protein-based ELISA for detection and differentiation of antibodies against avian reovirus in vaccinated and non-vaccinated chickens. J. Virol Meth.165: 108-111. 2010. 59. Wei Zhu " Jianbao Dong " Zhixun Xie Qi Liu " Mazhar I. Khan. Phylogenetic and pathogenic analysis of Newcastle disease virus isolated from house sparrow (Passer domesticus) living around poultry farm in southern China. Virus Genes. 231235. 2010. 60. Susta, L., Miller, P. J., Afonso, C. L., Estevez, C., Yu, Q., Brown, C.C. 2010. Pathogenicity evaluation of different Newcastle disease virus chimeras in 4-week-old chickens. Trop.Anim.Health Prod. 42:1785-1795. 61. Estevez, C., King, D.J., Luo, M., and Yu, Q. 2011. A Single Amino Acid Substitution in the Hemagglutinin-Neuraminidase Protein of Newcastle Disease Virus Results in Increased Fusion promotion and Decreased Neuraminidase Activities without Changes in Virus Pathotype. Journal of General Virology, In press. 62. Weng, Y., Lu, W., Harmon, A., Xiang, X., Deng, Q., Song, M., Wang, D., Yu, Q., and Li, F. 2011. The cellular ESCRT pathway is not involved in avian metapneumovirus budding in a virus-like-particle expression system. Journal of General Virology, In press. 63. Yu, Q., Estevez, C.N., Roth, J.P., Hu, H., and Zsak, L. 2011. Deletion of the M2-2 gene from avian metapneumovirus subgroup C (aMPV-C) impairs virus replication and immunogenicity in turkeys. Virus Genes, In press. 64. Hsieh, M.K., Wu, C.C., and Lin, T.L. 2010. DNA-mediated vaccination conferring protection against infectious bursal disease in broiler chickens in the presence of maternal antibody. Vaccine, 28: 3936-3943. 65. Jackwood, M. W., D. L. Suarez, D. Hilt, M. J. Pantin-Jackwood, E. Spackman, P. Woolcock, and C. Cardona. Biological Characterization of Chicken-Derived H6N2 Low Pathogenic Avian Influenza Viruses in Chickens and Ducks. Avian Diseases 54:120-125, 2010. 66. Jackwood, M. W., T. O. Boynton, D. A. Hilt, E. T. McKinley, J. C. Kissinger, A. H. Paterson, J. Robertson, c. Lemke, A. W. McCall, S. M. Williams, J. W. Jackwood, and L. A. Byrd. Emergence of a Group 3 Coronavirus Through Recombination. Virology, 398:98-108. 2010. 67. Jackwood, M. W., R. Rosenbloom, M. Petteruti, D. A. Hilt, A. W. McCall, and S. M. Williams. Avian Coronavirus Infectious Bronchitis Virus Susceptibility to Botanical Oleoresins and Essential Oils In Vitro and In Vivo. Virus Research, 149:86-94. 2010. 68. Jackwood, M. W., D. A. Hilt, H. S. Sellers, S. M. Williams, and H. N. Lasher. Rapid Heat-Treatment Attenuation of Infectious Bronchitis Virus. Avian Pathology 39:227-233, 2010. 69. Panshin, A., N. Golender, I. Davidson, S. Nagar, M. Garcia, M. W. Jackwood, E. Mundt, A. Alturi, S. Perk. Variavility of NS1 proteins among H9N2 avian influenza viruses isolated in Israel during 2000-2009. Virus Genes 41:396-405, 2010. 70. Gay, L., and E. Mundt. (2010). Testing of a New Disinfectant Process for Poultry Viruses. Avian Diseases 54: 763767. 71. Dlugolenski, D., Hauck, R., Hogan, R. J., Michel, F., and E. Mundt. (2010). Production of H5 specific monoclonal antibodies and the development of a competitive ELISA for detection of H5 antibodies in multiple species. Avian Diseases 54: 644649. 72. Liu, Y., Mundt E., Mundt A., Sylte M., Swayne D., and M. García (2010). Development and evaluation of an avian influenza (AI) neuraminidase subtype 1 (N1) based serological ELISA for poultry using the differentiation of infected and vaccinated animals (DIVA) control strategy. Avian Diseases 54: 613621. 73. Avellaneda, G., Mundt, E., Lee, C-W, and Suarez, D. L. (2010). Differentiation of infected and vaccinated animals (DIVA) using the NS1 protein of avian influenza virus. Avian Diseases 54:278286. 74. Vagnozzi A., M. García, S. M. Riblet, and G. Zavala*. Protection Induced by Infectious Laryngotracheitis Virus Vaccines Alone and Combined with Newcastle Disease Virus and/or Infectious Bronchitis Virus Vaccines. Avian Diseases December 2010, Vol. 54, No. 4: 1210-1219. 75. Johnson D. I., A. Vagnozzi, F. Dorea, S. M. Riblet, A. Mundt, G. Zavala, and M. García*. Protection Against Infectious Laryngotracheitis by In Ovo Vaccination with Commercially Available Viral Vector Recombinant Vaccines. Avian Diseases December 2010, Vol. 54, No. 4: 1251-1259. 76. Liu, Y., M. Sylte, D. Swayne, E. Mundt, and M. García. Development and evaluation of an avian influenza (AI) neuraminidase subtype 1 (N1) based ELISA for poultry using the differentiation of infected and vaccinated animals (DIVA) approach. Submitted to Avian Diseases. 77. McKinley, E. T., D. A. Hilt, and M. W. Jackwood. Avian coronavirus infectious bronchitis attenuated live vaccines undergo selection of subpopulations and mutations following vaccination. Vaccine. 26:1274-1284, 2008. 78. Warke, A., L. Appleby, and E. Mundt. Prevalence of Antibodies to Different Avian Paramyxoviruses in Commercial Poultry in the United States. Avian Diseases 52:549, 2008. 79. Oldoni, I., A. Rodríguez-Avila, S. M. Riblet, G. Zavala, and M. García. Pathogenicity and Growth Characteristics of selected infectious laryngotracheitis virus (ILTV) strains from the United States. Avian Pathology, in press. Abstracts, Presentations, etc: 1. Rauf Abdul, Maria V. Murgia, M. Khatri, A. Rodriguez-Palacios, C-W. Lee and Y.M. Saif. Distribution and Persistence of Infectious Bursal Disease Virus in Chickens. 61st North Central Avian Disease Conference. St. Paul, Minnesota. March 15 & 16. 2010. 2. Jackwood, D. J., S. E. Sommer-Wagner, S. T. Stoute, P. R. Woolcock, B. M. Crossley, S. K. Hietala and B. R. Charlton. The very virulent infectious bursal disease virus (vvIBDV) strain of birnavirus in California: Identification and pathogenicity of a reassortant virus. Abstr. #41, 29th Annual Am. Society for Virol. Meet. 2010. 3. Jackwood, D. J., S. E. Sommer-Wagner and S. T. Stoute. Morbidity, mortality and pathology caused by different challenge doses of vvIBDV. Abstr. 9370, Poster #51, 147th AVMA meeting. 2010. 4. Rauf A, Murgia MV, Rodriguez-Palacios A, Khatri M, Lee CW, Saif YM. Persistence and distribution of infectious bursal disease virus in SPF and commercial broiler chickens. OARDC Conference. Wooster, Ohio. April 22. 2010. 5. Ngunjiri JM, Marcus PI, Sekellick MJ, Wang L, Lee CW. In vitro analysis of virus particle subpopulations in candidate live-attenuated influenza vaccines distinguishes effective from ineffective vaccines. American Society of Virology Annual Meeting. Bozeman, Montana. 2010. 6. Rauf Abdul, Maria V. Murgia, Lee CW, M. Khatri, and Y.M. Saif. Persistence and distribution of infectious bursal disease virus in SPF and commercial broiler chickens. 147th AVMA Annual Convention. Atlanta, GA. July 30August 4, 2010. 7. Lee CW, Qin Z, Clements T, Wang L, Khatri M, Zhang Y, LeJeune JT. Influenza infection in starlings. 147th AVMA Annual Convention. Atlanta, GA. July 30August 4, 2010. 8. Rauf Abdul, Maria V. Murgia, C-W. Lee, M. Khatri and Y.M Saif. Persistence and distribution of infectious bursal disease virus in SPF and commercial broiler chickens. 147th AVMA Annual Convention. Atlanta, GA. July 30August 4, 2010. 9. Rauf Abdul, M. Khatri, Maria V. Murgia and Y.M. Saif. Viral induced inflammatory cytokine, toll like receptors and cytotoxic T cells components in infectious bursal disease infected chickens. Conference for Research workers in animal science at Chicago, December 5  7, 2010. 10. Giambrone, K. Guo, and T. V. Dormitoriiso.2010. Detection and Differentiation of avian reoviruses using SYBER-Green I based two step real time RT-PCR with melting curve analysis. Southern Conference on Avian Diseases. Atlanta Ga. Jan 26. 11. Ou, Shan-Chia, T. V. Dormitorio, and J. J. Giambrone. 2010. Detection of infectious laryngotracheitis virus by loop mediated isothermal amplification (LAMP). Southern Conference on Avian Diseases. Atlanta Ga. Jan 26. 12. Dormitorio, T.V., Giambrone, J. J., and K. Guo. 2010. 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