NC_OLD1180: Control of Emerging and Re-emerging Poultry Respiratory Diseases in the United States

(Multistate Research Project)

Status: Inactive/Terminating

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SAES-422 Reports

Annual/Termination Reports:

[03/10/2010] [03/18/2011] [03/05/2012] [01/03/2013] [02/24/2014]

Date of Annual Report: 03/10/2010

Report Information

Annual Meeting Dates: 01/26/2010 - 01/27/2010
Period the Report Covers: 10/01/2008 - 09/01/2009

Participants

Advisor: Saif, Yehia (saif.1@osu.edu)

USDA representative: Johnson, Peter (PJOHNSON@CSREES.USDA.GOV)

State Station Representatives: Giambrone, Joe (giambjj@auburn.edu) Auburn University; Khan, Mazhar (mazhar.khan@uconn.edu) - University of Connecticut; Glisson, John (jglisson@vet.uga.edu) University of Georgia; Wu, Ching Ching (wuc@purdue.edu) Purdue University; Lee, Chang Won (lee.2854@osu.edu) Ohio State University; Zsak, Laszlo (Laszlo.Zsak@seprl.usda.gov) USDA, Southeast Poultry Research Lab.

Other participants: Keeler, Calvin (ckeeler@udel.edu) University of Delaware; Johnson, Timothy (joh04207@umn.edu) University of Minnesota; Toro, Haroldo (torohar@vetmed.auburn.edu) Auburn University; Garcia, Maricarmen (mcgarcia@uga.edu), Mundt, Egbert (emundt@uga.edu), Jackwood, Mark (mjackwoo@uga.edu), Sellers, Holly (hsellers@uga.edu), Ferguson-Noel, Naola (naolaf@uga.edu) - University of Georgia; Lin, Tsang Long (tllin@purdue.edu) Purdue University; Pantin-Jackwood, Mary (Mary.Pantin-Jackwood@ars.usda.gov), Yu, Qingzhong (Qingzhong.Yu@ars.usda.gov), Swayne, David (David.Swayne@ARS.USDA.GOV), Miller, Patty (Patti.Miller@ars.usda.gov) - USDA, Southeast Poultry Research Lab.

Brief Summary of Minutes

Minutes Attachment:

Attachment

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, which include hunter-killed or nesting waterfowl and shorebirds, starlings, and raptors, were accomplished. The enormous data obtained from different states (AL, DE, GA, MN, OH) are being shared. 2. Surveillance activities on the Delmarva Peninsula have yielded infectious laryngotracheitis (LT) virus and infectious bronchitis virus isolates from commercial broiler chickens. 3. In DE, LT incidence down in 2009 due to widespread vaccination. Severity of LT clinical signs and lesions are mild to moderate, very similar to that seen in adverse CEO vaccine reactions. 4. GA isolated and characterized current pathogenic respiratory viruses, bacteria, and mycoplasmas circulating within the poultry industry in Georgia. Identified at least 41 MG genotypes that are distinguishable from live vaccines and unique to individual countries or regions. 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 rapid, accurate, and economical method for ILTV detection using a molecular technique Loop-mediated isothermal amplification (LAMP). The LAMP could detect viral DNA directly from the tracheae of vaccine virus infected birds as well as ILTV plaques from embryonic eggs. 2. CT designed nine pairs of neuramidinase (NA) subtype-specific primers using Primer Hunter design tool and successfully used in real time RT-PCR with four primer-pool reactions to differentiate nine NA subtypes of AIV. 3. GA developed an indirect N1 and N2 ELISAs which were proven to be effective and rapid assay to identify exposure to challenge virus during a DIVA vaccination strategy. In addition, a speciesindependent competitive ELISA (cELISA) for the detection of H6, H7, H9 antibodies in several species was developed. 4. MN developed degenerate primer set for full-length amplification of four genes of influenza A viruses in a single reaction. 5. OH established the chloroform-Mag MAXTM method of viral RNA extraction followed by RRT-PCR which can be used as rapid and sensitive test to determine the titer of the viral RNA. Using this method, it was found that different commercial vaccines contain varied antigen contents. 6. SEPRL (USDA) developed two real time RT-PCR assays that allow the differentiation of North American H1N1 from pandemic H1N1. In addition, the current H7 RRT-PCR was improved to detect a broader range of H7 viruses that are found in Western hemisphere. 7. SEPRL demonstrated that NDV Matrix assay failed to detect a virulent NDV. If genotype VII virus was found in North America this assay could be used in the NALHN laboratories. 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. DE isolated 5038 IBDV isolate from commercial broiler chickens. Although unique based on VP2 sequencing and monoclonal antibody testing, may not be capable of breaking through maternal immunity in a laboratory designed trial may not be able to break through in real world progeny challenges. 2. GA identifed that temperature plays a pivotal role in the survivability of LPAI virus in feces and in contact with litter. GA also identified that a percentage of chickens receiving recombinant or TCO vaccines carry a significant amount of virulent ILTV in the trachea in the absence of clinical signs after being challenged with virulent ILTV. 3. Comparative genomic analysis of IBV 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. 4. OH identified amino acids contributing to antigenic drift in the Del-E infectious bursal disease virus. The short term implication this has for the poultry industry is that diagnostic assays designed to identify the 254 and 222 amino acids will discover viruses that have antigenically important mutations. 5. OH showed that two virulent infectious bursal disease viruses (vvIBDV) from California are identical and meet all the characteristics of a vvIBDV. Because they have the potential to spread rapidly and cause high mortality in chickens, the impact of these viruses on the U.S. broiler and layer industries could be considerable. 6. OH detected low pathogenic influenza viruses in albumin of eggs using real time RT-PCR and virus isolation in embryonated chicken eggs. Swabs from egg shells were also found positive by RRT-PCR. Objective IV. Develop new prevention and control strategies for poultry respiratory diseases. 1. AL developed two recombinant vaccines against H1N1 AIV (one DNA and the other in yeast) and found to induce a measurable immune response in young chickens. The DNA vaccine was given by injection and the yeast vaccine in the drinking water. 2. CT tested in ovo vaccination of recombinant DNA plasmid containing IBV spike gene with interferon-a which showed over 98% of protection rate against M41 field isolate challenge. 3. DE developed a second generation escape resistant RNAi constructs against avian influenza virus and found that avian-specific RNAi constructs against avian influenza virus did not increase the efficiency of RNAi inhibition. 4. IN demonstrated that IBDV large segment gene-based DNA can elicit specific immune response and provide protection of broiler chickens with maternally derived antibody against infection challenge. 5. OH developed NA- and NS-based DIVA vaccine strains using traditional reassortment as well as reverse genetics methods against H3N2 influenza in turkeys. The reassortant DIVA vaccines significantly reduced challenge virus shedding in the oviduct of breeder turkeys as well as trachea and cloaca of both young and old breeder turkeys, suggesting that proper vaccination could effectively prevent egg production drop and potential viral contamination of eggs in infected turkeys. 6. SERPL demonstrated that H7 AI vaccine may not protect against intercontinental H7 field viruses and vaccine may need to be from the same H7 lineage as field viruses to provide protection. In addition, turkeys vaccinated with commercial H1N1 vaccine have a low chance of being protected against swine-origin H1N1 infection. 7. SEPRL developed a model system for NDV vaccination which mimic egg production losses seen in Asia and Mexico in vaccinated poultry were developed and this system will allow the comparison of vaccines.

Publications

Publications Attachment:

Attachment

Impact Statements

Wild birds are a reservoir of AIVs and some species may serve as potential intermediate host. Viral detection should be done by passage of fecal swab material in embryos first then by RRT-PCR and should exclude AC-ELISA.
Composting of AIV infected eggs for as early as 24 hours and late as 52 hours can inactivate AIV. The internal temperature of the pile must reach 560 F for the inactivation to occur. The temperature is a function of the amount of pile turning and moisture. Presently, 7 days are used in the industry to perform this function.
Two real time RT-PCR assays that allow the differentiation of North American H1N1 from pandemic H1N1 were developed. The National Animal Health Laboratory Network adopted these tests.
Low pathogenic influenza viruses were detected from internal egg contents following experimental infection in turkeys. The possibility of hatchery contamination by egg borne influenza viruses and spread of virus during movement of contaminated cracked eggs and egg flats pose concerns regarding influenza viral dissemination.
ILTV is present in commercial poultry houses causing mild outbreaks. The viruses were found in the dust, litter, beetles, water, and rats. Heating of the house to 1000 F for 100 hours, composting of the litter for 3 days, improved beetle control, treatment of the drinking water system with commercial biofilm removers, and rodent control will reduce the amount of virus in the house.
Factors hindering control of ILT may be suboptimal immunization against ILT resulting from multivalent vaccinations. Reducing the number and diversity of live virus vaccines given concomitantly with ILT vaccines may optimize protection against ILTV and possibly against other viral respiratory diseases.
A high titer of ILTV vaccine is required for a prompt neutralizing immune response. Thus, vaccine fractionation would seem counterproductive.
Monitoring the ability of infectious bursal disease virus (IBDV) to break through maternal immunity in young broiler chickens is important to assess the immunosupproessive potential of the viruses.
IBDV large segment gene-based DNA vaccine has the potential for practical application to confer protection of chickens with maternal antibodies against IBD in the poultry industry.
Monitoring infectious bronchitis viruses from commercial broiler chickens is important for monitoring the effectiveness of vaccination programs and to isolate and characterize field viruses that break through vaccine induced immunity.
In-ovo DNA immunization may become one of the most important innovation in the DNA vaccination of poultry against IBV, allowing it to be used in commercial in-ovo vaccination as a much safer vaccine than the attenuating live IBV vaccines used currently.
Genomic characterization of fowlpox virus and other avianpox viruses for specific virulence markers e.g. full length REV can be done by PCR amplification of the genetic fragments with specific primers. In this regard, DNA isolated from formalin fixed paraffin-embedded tissue sections can be used effectively.

Date of Annual Report: 03/18/2011

Report Information

Annual Meeting Dates: 01/23/2011 - 01/24/2011
Period the Report Covers: 10/01/1999 - 09/01/2010

Participants

Participants:

Advisor: Saif, Yehia (saif.1@osu.edu)

USDA representative: Johnson, Peter (PJOHNSON@CSREES.USDA.GOV)

State Station Representatives: Kong, Byung-Whi (bkong@uark.edu) University of Arkansas, Giambrone, Joe (giambjj@auburn.edu) Auburn University; Khan, Mazhar (mazhar.khan@uconn.edu) - University of Connecticut; Gelb, Jack (jgelb@udel.edu) University of Delaware, Glisson, John (jglisson@vet.uga.edu) University of Georgia; Tripathy, Deoki (tripathy@uiuc.edu) - University of Illinois, Wu, Ching Ching (wuc@purdue.edu) Purdue University; Lee, Chang Won (lee.2854@osu.edu) Ohio State University; Zsak, Laszlo (Laszlo.Zsak@seprl.usda.gov) USDA, Southeast Poultry Research Lab.

Other participants: Keeler, Calvin (ckeeler@udel.edu) University of Delaware; Kakambi Nagaraja (nagar001@tc.umn.edu) University of Minnesota; Toro, Haroldo (torohar@vetmed.auburn.edu) Auburn University; van Santen, Vicky (vanvick@auburn.edu), Mundt - Auburn University, Egbert (emundt@uga.edu), Jackwood, Mark (mjackwoo@uga.edu), Sellers, Lin, Tsang Long (tllin@purdue.edu) Purdue University; Pantin-Jackwood, Mary (Mary.Pantin-Jackwood@ars.usda.gov), Yu, Qingzhong (Qingzhong.Yu@ars.usda.gov), Suarez, David (David.Suarez@ARS.USDA.GOV)

Brief Summary of Minutes

Minutes Attachment:

Attachment

Accomplishments

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.

Publications

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. Isolation, characterization, inactivation of H1N1 viruses from wild water fowl. Poult Science Association. Annual Meeting. Denver, CO. July 19-21. 13. Dormitorio, T.V. and J. J. Giambrone. 2010. Limiting dilution studies to detect avian influenza viruses from questionable. American Association of Avian Pathologist Annual Meeting, Atlanta, GA. Aug 1-4. 14. Giambrone, J. J., Sc. Ou, N. K. Singh, K. S. Gunn, H. Wu, and R, Singh. 2010. AIV H1N1 DNA vaccine induced humoral and cell-mediated immunity in SPF chickens. American Association of Avian Pathologist Annual Meeting, Atlanta, GA. Aug 1-4. 15. Ndegwa, E.N. and V.L. van Santen. Specific immune responses associated with viral subpopulations selected in chickens after vaccination with Ark-type infectious bronchitis vaccines. American Association of Avian Pathologists Annual Meeting, Atlanta, GA, August 1-4, 2010. 16. Gallardo, R.A., V.L. van Santen, and H. Toro. Effects of CAV and/or IBDV on IBV Replication and Phenotypic Drift. American Association of Avian Pathologists Annual Meeting, Atlanta, GA, August 1-4, 2010. 17. Gallardo, R.A., V.L. van Santen, F.J. Hoerr, and H. Toro. Effects of Infectious Bronchitis Virus on Chicken Testicles. (Poster) American Association of Avian Pathologists Annual Meeting, Atlanta, GA, August 1-4, 2010. 18. van Santen, V.L. and E. N. Ndegwa. Highly Localized Infections with Ark-type IBV Vaccines. (Poster) American Association of Avian Pathologists Annual Meeting, Atlanta, GA, August 1-4, 2010. 19. Bartlett, S. and V. van Santen. Mass IBV serotype vaccine predominates in chickens simultaneously vaccinated with Mass and Ark serotype vaccines. (Poster) Annual Merial NIH National Veterinary Scholars Symposium, Athens, GA, August 5-8, 2010. 20. Ndegwa, E.N. and V.L. van Santen. Transmission of IBV Ark serotype type vaccine viral subpopulations to non-vaccinated contact birds. (Poster) Annual Southeastern Branch ASM Conference, Montgomery, AL, Nov. 5-6, 2010. 21. Toro, H. D. C. Tang, D. L. Suarez, F. W. van Ginkel. Avian Influenza Vaccination with Non-Replicating Adenovirus Vector: Assessment of Protection after either Mucosal Delivery or Decreasing Dose Applied by the In Ovo Route. American Association of Avian Pathologists Annual Meeting, Atlanta, GA, August 1-4, 2010. 22. Toro, H., Minc, K., C. Bowman, S. Gulley, D.C. Tang, J. Hathcock. Avian Influenza Vaccination with Non-Replicating Adenovirus Vector: Target Tissues in Chicken Embryo. (Poster) American Association of Avian Pathologists Annual Meeting, Atlanta, GA, August 1-4, 2010. 23. Breedlove, C., F. W. van Ginkel, D. C. Tang, and H. Toro. Combined In Ovo-Vaccination with Non-Replicating Adenovirus-Vectored Avian Influenza and Mareks Disease Vaccines. (Poster) American Association of Avian Pathologists Annual Meeting, Atlanta, GA, August 1-4, 2010. 24. Mesonero Alexander, De-chu C. Tang, and Haroldo Toro. In Ovo-Vaccination with Non-Replicating Adenovirus-Vectored Avian Influenza: Maternal Immunity and Effects on Vaccination. (Poster) American Association of Avian Pathologists Annual Meeting, Atlanta, GA, August 1-4, 2010. 25. Chandra YG, Lee JY, and Kong B-W. 2011. Characterization of sequence variation in the viral genomes of infectious larygotracheitis virus (ILTV) using next generation sequencing. International Poultry Science Forum (IPSF), Atlanta, GA. January 24-25. 26. Lee JY, and Kong B-W. 2010. The analysis of gene expression of infectious laryngotracheitis virus during lytic replication phase in cultured cells. 29th Annual meeting of American Society for Viology. Montana State University, Bozeman, Montana. July 17-21. 27. Tripathy, D.N. and Bahaa, A.F.A. 2010. Differentiation of Avianpox viruses by PCR amplification of specific genes. (Abstract) AAAP, AVMA Conference, Atlanta, GA. 28. Tripathy, D.N. 2010. Fowlpox Virus Immunity, Interest of using such virus as vector. Ceva Vector Vaccines Symposium, San Diego, California, pp. 47. 29. Hariastuti, N.I., Babapoor, S., Girshick, T., and Khan, M.I. In-vitro inactivation of avian influenza viruses using caprylic acid and its derivatives. Precede 14th International Congress on Infectious Diseases, Miami, Florida, March 9-12, 2010, CDROM. 30. Huang, Y., Khan, M.I., and Mandoiu, I.I. Development of real time RT-PCR assays for neuraminidase subtyping of avian influenza virus. Precede 6th International Symposium on bioinformatics research and applications. Storrs, Connecticut. May 23-26, 2010. P19. 31. Wu, C.C., Hsieh, M.K., and Lin, T.L. Protection of broiler chickens against infectious bursal disease by DNA vaccination in the face of maternally derived antibodies. The Proceedings of the 147th Annual Meeting of the American Veterinary Medical Association and the 53th Annual Meeting of the American Association of Avian Pathologists. Atlanta, Georgia, July-August, 2010

Impact Statements

Avian paramyxovirus-1 isolates from wild waterfowl are capable of replicating in broiler chickens, turkeys and ducks and thus represent a risk to non-vaccinated poultry.
Monitoring infectious bronchitis viruses from commercial broiler chickens is important for monitoring the effectiveness of vaccination programs and to isolate and characterize field viruses that break through vaccine induced immunity.
The reassorting of California vvIBDV with an endemic serotype 2 virus suggests vvIBDV may have entered California earlier than originally thought. The control of vvIBDV and now these newly emerging reassortant viruses will be economically important to the future of the U.S. broiler and layer industries.
The persistence and distribution of IBDV in SPF chickens is significantly different in comparison with commercial chickens. Although the virus can persist in bursa for a month, it is very unlikely that the infectious virus will be present in the processed meat.
All strains of avian pox viruses showed the presence of photolyase gene in their genomes. Based upon the nucleotide sequence differences of a PCR amplified fragment of photolyase gene these viruses could be differentiated into four different groups.
Pandemic H1N1 virus is poorly adapted to poultry, therefore poultry would likely not serve as a reservoir and the virus has a minimal disease potential for young poultry. Turkey breeders could have been infected by infected insemination crews and that biosecurity practices for artificial insemination, which is universally practiced by the turkey industry, need to be modified.
Immunodeficiency caused by ubiquitous immunosuppressive viruses can have an effect on evolution and persistence of IBV in flocks.
In vitro analyses or virus subpopulation provide a benchmark for the screening of candidate live influenza vaccines and their potential as effective vaccines. Vaccine design may be improved by enhancement of attributes that are dominant in the effective vaccines.
IFN-alpha can protect chickens from disease associated with low pathogenic AIV and reduce the risk of transmission through decreased shedding.
IBDV large segment gene-based DNA vaccine has the potential for practical application to confer protection of chickens with maternal antibodies against IBD in the poultry industry.
NDV (B1) and IBV (ARK) vaccines and a multivalent vaccine constituted by NDV (B1) and IBV (ARK and MASS) do not interfere with the protection induced by the CEO ILTV vaccine. However, the NDV (B1) and the multivalent (B1/MASS/ARK) vaccines interfere with the protection induced by the TCO ILTV vaccine.
ILTV vaccines are wide spread in Northern Alabama and Georgia, causing a mild form of ILTV. New management techniques should be done to eliminate the reservoir viruses from rodents, beetles, and drinking waters.
Recombinant ILTV vaccines appeared to require at least 35 days post-vaccination before providing the best attainable protection. Protection provided by recombinant vaccines (HVT-vectored and fowlpox-vectored) was generally poor when vaccinated chickens were challenged at 28 days post-vaccination or less.
Infectious laryngotracheitis is an economic disease that also has important trade implications for the poultry industry. Vaccination using CEO and recombinant vaccines is helping control the disease but more research is warranted to develop improved vaccines and control strategies.

Date of Annual Report: 03/05/2012

Report Information

Annual Meeting Dates: 01/22/2012 - 01/23/2012
Period the Report Covers: 10/01/2010 - 09/01/2011

Participants

Advisor: Saif, Yehia (saif.1@osu.edu)
State Station Representatives: Haroldo, Toro (torohar@auburn.edu),Auburn University; Kong, Byung-Whi (bkong@uark.edu, University of Arkansas; Khan, Mazhar (mazhar.khan@uconn.edu) - University of Connecticut; Gelb, Jack (jgelb@udel.edu), University of Delaware; Jackwood, Mark (mjackwoo@uga.edu, University of Georgia; Tripathy, Deoki (tripath@uiuc.edu), University of Illinoise; Wu, Ching Ching (wuc@purdue.edu), Purdue University; Lee, Chang Won (lee.2854@osu.edu), Ohio State University; Johnson, Tim (joh04207@umn.edu)- University of Minnesota; Zsak, Laszlo (Laszlo.Zsak@ars.usda.gov), USDA, Southeast Poultry Research Lab.

Other participants: Keeler, Calvin (ckeeler@udel.edu), University of Delaware; Erin Brannick (brannick@udel.edu), University of Delaware; Joseph Giambrone (giambjj@auburn.edu), Auburn University; Maricarmen, Garcia (gmaricar@uga.edu), University of Georgia; Naola Ferguson-Noel (nferguson@uga.edu), University of Georgia; Lin, Tsang Long (tllin@purdue.edu), Purdue University; Mo Saif (saif.1@osu.edu ), Ohio State University; Pantin-Jackwood, Mary (Mary.Pantin-Jackwood@ars.usda.gov), Yu, Qingzhong (Qingzhong.Yu@ars.usda.gov)-USDA, Southeast Poultry Research Lab.

Brief Summary of Minutes

Minutes Attachment:

Attachment

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 surveillance data obtained from different states (AL, CT, DE) were shared. No AIV activity using USDA NAHLN-approved agent detection (real time RT-PCR and antigen capture on oropharyngeal swabs) were seen in commercial flocks (CT, DE). 2. Surveillance activities on the Delmarva Peninsula have yielded infectious laryngotracheitis (ILT) virus and infectious bronchitis virus isolates from commercial broiler chickens and Newcastle disease virus isolates from wild birds. 3. Delmarva has continued to observed ILT activity in 2010 and 2011. The severity of ILT clinical signs and lesions are mild to moderate, very similar to that seen in adverse CEO vaccine reactions. 4. Gene targeted sequencing was used to determine the source and spread of MG and MS isolates in the field. Approximately 123 MG and MS were analyzed in 2011 and the circulation of field strains within complexes and companies was identified. 5. SEPRL (USDA) obtained virulent Newcastle disease viruses from Mexico, Africa, China, Pakistan, and from U.S. wild birds which have been sequenced and characterized phylogenetically. The sequence data has allowed the improvement of the current diagnostic tests for NDV to ensure that the circulating viruses can be diagnosed. 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 TaqMan® real time polymerase chain reaction (PCR) and loop-mediated isothermal amplification (LAMP) assays. Both assays were specific, sensitive, and reproducible for ILTV detection. Although the sensitivity of LAMP was lower than real time PCR, it was faster, had a lower cost, and did not require a temperature cycler. This was the first report comparing these methods for ILTV DNA detection. 2. CT in collaboration with Guangxi Veterinary Institute, China developed loop-mediated isothermal amplification (LAMP) assays to detect the H3 subtype AIVs visually and rapid detection of group I avian adenoviruses. The newly developed H3-RT and group I avian adenoviruses LAMP assays are simple, sensitive, rapid and can identify H3 subtype AIVs and group I avian adenoviruses visually. Consequently, they will be very useful screening assays. 3. DE, SEPRL evaluated the effect of pooling 11 or 5 oropharyngeal swabbings on detecting avian influenza virus by real time reverse transcription PCR . 4. DE, GA and SEPRL using next generation sequencing technologies for rapid determination of the primary sequence of the infectious laryngotracheitis virus ( ILTV) genome. 5. GA developed a rapid multiplex microsphere assay for the simultaneous detection of all avian influenza viruses (AIV) as well as differentiation of H5, H7, N1 and N2 subtypes. 6. GA optimized duplex real-time PCR method for relative quantification of ILTV. 7. GA developed and validated N1 and N2 ELISAs as the assays that will be required for the implementation of a DIVA control strategy for H5N1, H5N2, H9N2, H3N2 and H1N1 poultry infections that will be required for the implementation of a DIVA control strategy for H5N1, H5N2, H9N2, H3N2 and H1N1 poultry infections. 8. GA developed and validated N1 and N2 ELISAs as the assays that will be required for the implementation of a DIVA control strategy for H5N1, H5N2, H9N2, H3N2 and H1N1 poultry infections. 9. IL developed a photolase gene specific PCR. Based on sequence information, avian pox viruses could be differentiated into four different groups. 10. OH developed 19-plex assay which can differentiate different HA subtypes of avian influenza viruses. 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 venereal transmission of IBV by artificially inseminating old hens either with semen from IBV infected roosters or with IBV suspended in naïve semen. IBV RNA was detected in the trachea of all hens inseminated with IBV-spiked semen and in 50% of hens inseminated with semen from IBV-infected males. These results provide experimental evidence for IBV venereal transmission. 2. AL investigated that the dominant genotype of the vaccine strain of IBV was rapidly negatively selected in all chicken groups [CAV, IBDV, CAV+IBDV, and immunocompetent]. These results suggest that the generation of genetic diversity in IBV is constrained. This finding constitutes further evidence for phenotypic drift occurring mainly as a result of selection. 3. GA examined and compared the genomes of pathogenic and attenuated strains of IBV and measure evolution of IBV by examining virus diversity and mutation rates. 4. GA investigated embryo lethal dose50 for a reliable indication of the virulence of MG isolates. The ELD50 of ts-11 like isolates tested correlated well with the history and previous pathogenicity testing. 5. IL genetically characterized a vaccine strain of fowlpox virus showed complete homology with the corresponding gene sequences and indicated absence of full-length REV in the genome of this virus. 6. MN identified molecular mechanisms enabling APEC to survive and grow in this critical host environment. 7. OH investigated the replication of swine and human influenza A viruses in juvenile and layer turkeys. 8. OH studied maternal immunity in limiting the spread or reducing the severity of the clinical disease caused by very virulent infectious bursal disease virus (vvIBDV). 9. OH studied amino acid sequence data acquired from sequencing of IBDV collected from bursa samples during a ten year period from 2002 2011. 10. OH investigated Fas/FasL and perforin systems as important mechanisms of T cell-mediated cytotoxicity in infectious bursal disease virus infected chickens. 11. SEPRL evaluated the pathogenicity of H5N1 HPAI viruses isolated in Egypt in domestic ducks. 12. SEPRL characterized pathogenicity of new NDV viral isolates from South America, Africa and US waterfowl in chickens. Objective IV. Develop new prevention and control strategies for poultry respiratory diseases. 1. AL evaluated protection conferred by mucosal vaccination with replication competent adenovirus (RCA)-free recombinant adenovirus expressing a codon-optimized avian influenza (AI) H5 gene fromA/turkey/WI/68 (AdTW68.H5ck). 2. AL developed a DNA vaccine consisted of the entire HA gene of an AIV H1N1 subtype (A/bluewinged teal/ AL/167/2007) cloned into the eukaryotic expression vector. The immunological responses induced by DNA vaccine against AIV were also investigated. 3. AR made comparison of ILTV genome sequences of two US CEO vaccines. 4. CT evaluated the level of protection of M2e-nanopartle based vaccine using quantitative real time PCR at 4, 6, and 8 days post-challenge with H5N2 LPAI by measuring virus shedding from trachea and cloaca. 5. IN conducted studies to determine if the combination of chicken calreticulin (CRT) gene and infectious bursal disease virus (IBDV) large segment (VP243) gene in a fusion gene or a chimeric DNA could enhance protection against IBD by DNA vaccination. 6. MN Correlated between virulence and MDR in avian E. coli and characterized the biology of the emergent IncA/C plasmid group. 7. OH in collaboration with the University of Cincinnati utilized flexible norovirus P particle as a novel influenza vaccine platform in vitro analysis of virus particle subpopulations in candidate live-attenuated influenza vaccines which could distinguish effective from ineffective vaccines. 8. SEPRL showed that a single vaccination can protect ducks and geese from avian influenza virus if the virus and vaccine are related. Reduction of pandemic H1N1 avian influenza growth with use of chicken interferon was investigated. 9. SEPRL generated and evaluated a bivalent vaccine against avian metapneumovirus and Newcastle disease viral diseases.

Publications

Publications Attachment:

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Impact Statements

Avian influenza subtype H5 and H7 were negative from the LBM and domestic poultry birds in New England states and in Delaware commercial farms. However wild birds do carry H5 subtypes in their population.
Infectious laryngotracheitis virus and infectious bronchitis virtues circulating in commercial broiler chickens flocks in Delaware.
Molecular Epidemiology reinforces the importance of surveillance for MG and MS isolates in poultry for the control of avian mycoplasmas.
The sequence data has allowed the improvement of the current diagnostic tests for NDV to ensure that the circulating viruses can be diagnosed.
A new diagnostic tests developed for ILTV, AIV and avian adenoviruses using loop-mediated isothermal amplification (LAMP) techniques will be faster, specific, sensitive and cost effective will not require sophisticated equipment.
Utilization of next generation sequencing technologies now permits the relatively rapid determination of the primary sequence of the ILTV genome.
Multiplex microsphere assay for detection of avian influenza viruses provides a rapid tool to identify multiple avian influenza types in the same sample.
Development of faster high-throughput serological assays for avian influenza (AI) that can complement a vaccination strategy to allow the rapid identification of infected flocks within large populations of vaccinated poultry. Identification of infected flocks is critical for control of AI outbreaks especially when vaccines are used.
Successfully developed 19-plex assay which can differentiate different HA subtypes of avian influenza viruses. With the multiplex capacity and feasibility of the assay, the multiplex branched DNA assay has a great potential in influenza research in addition to rapid diagnosis.
The egg internal and external quality was negatively affected in hens inseminated with semen containing IBV. These results provide experimental evidence for IBV venereal transmission.
Chickens infected with IBV and co-infected with CAV+IBDV will generate genetic diversity in IBV. This finding constitutes further evidence for phenotypic drift occurring mainly as a result of selection.
Poor vaccination against IBV infection contributes to the emergence of new IBV strains via mutation and/or selection. Under these conditions only IBV virus populations identical to the challenge virus was identified. From a broad perspective it indicates that selection is an important force driving IBV evolution.
Examine and compare gammacorona virus genomes for recombination, comparison data indicate that reticulate evolutionary change due to recombination in IBV, likely plays a major role in the origin and adaptation of the virus leading to new genetic types and strains of the virus. These data constitute a significant step forward in identifying pathogenicity genes in avian coronavirus infectious bronchitis.
In vitro expression of avian pathogenic Escherichia coli (APEC ) genes . This genome-wide analysis provides novel insight into processes that are important to the pathogenesis of APEC O1. Overall, these results indicate that a number of novel APEC virulence factors exist in APEC O1 that mediate systemic infection in the chicken host.
It was confirmed the susceptibility of both juvenile and layer turkeys to swine influenza viruses (SIVs) while the viruses replicated more efficiently in the reproductive tract of turkey hens compared to respiratory or digestive tracts.
Studies indicate the ability of vvIBDV to infect chickens is not affected by maternal immunity to IBDV strains typically found in commercial U.S. chickens. However maternal immunity did reduce the severity of the clinical signs and macroscopic lesions. These data suggest vvIBDV might be infecting chickens in California and other regions of the U.S. but they are going unnoticed because maternal immunity affects the clinical picture which does not include mortality and macroscopic lesions typical of a vvIBDV infection.
Data indicated that activated T cells may be involved in antiviral immunity and mediation of virus clearance from the bursa and spleen of IBDV-infected chickens. The findings of this study will help understanding the role of T cells in the pathogenesis of IBDV and designing effective control strategies against this immunosuppressive viral disease of chickens.
An increase in pathogenicity of AI in ducks observed with H5N1 HPAI viruses has implications for the control of the disease since vaccinated ducks infected with highly virulent strains shed more viruses and for longer periods of time, perpetuating the virus in the environment and increasing the possibility of transmission to susceptible birds.
Further comparison of US CEO vaccines to several ILTV genome sequences revealed that US CEO vaccines are genetically distinct from the two Australian-origin CEO vaccines, SA2 and A20, which showed close similarity. This information can be used to discriminate between vaccine ILTV strains and further, to identify newly emerging mutant strains of field isolates.
Preliminary studies suggest that the self-assembling polypeptide nanoparticle shows promise as a potential platform for a development of a universal vaccine against avian influenza type A.
Experimental studies indicated that live vaccines and bacterian can protect against ovarian regression as well as air sac and tracheal lesions.
It was shown that recombinant vaccines against ILTV provide some protection but do not prevent shedding, which can lead to continued spread of the virus, whereas the chicken embryo origin vaccine protected against both disease and virus shedding. This study is extremely important in the control of ILTV especially in the face of an outbreak.
Determining the unique sequences for chicken embryo origin (CEO) vaccines will enhance our ability to control the re-emerging epidemics ILTV in commercial chickens caused by CEO-related vaccines.
Study validated that the use of glycoprotein specific ELISAs as a tool to discriminate ILTV sero-conversion due to vaccination from infection. This work involves the serological differentiation of vaccinated and field virus exposed chickens which is critical for controlling ILTV epidemics.
IBDV large segment gene-based DNA can elicit specific immune response and provide protection of specific-pathogen-free and broiler chickens against infection challenge. The impact is that IBDV large segment gene-based DNA vaccine has the potential for practical application in providing protection of chickens against IBD in the poultry industry.
Evidence is mounting that IncA/C plasmids are widespread among enteric bacteria of production animals and these emergent plasmids have flexibility in their acquisition of MDR-encoding modules, necessitating further study to understand the evolutionary mechanisms involved in their dissemination and stability in bacterial populations.
Studies demonstrate that chicken interferon is biologically active against the pandemic H1N1 virus, is active in other avian species, and may be useful as therapy against avian influenza infection.
Potential bivalent recombinant vaccine candidate for NDV and aMPV was safe, stable and provided a complete protection against virulent NDV challenge and decreased the aMPV disease severity following experimental aMPV-C infection in turkeys.

Date of Annual Report: 01/03/2013

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Annual Meeting Dates: 12/02/2012 - 12/02/2012
Period the Report Covers: 10/01/2011 - 09/01/2012

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Accomplishments

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 surveillance data obtained from different states (CT, DE) were shared. No AIV activity using USDA NAHLN-approved agent detection (real time RT-PCR and antigen capture on oropharyngeal swabs) were seen in commercial flocks (CT, DE). 2. Surveillance activities on the Delmarva Peninsula have yielded infectious bronchitis viruses of Arkansas (57), Massachusetts (10), and Delaware 072 (10). Seven isolations of NDV were made during the period, of which three were concomitants of IBV isolations. 3. Delmarva has continued to observed ILT activity. The severity of LT clinical signs and lesions are mild to moderate, very similar to that seen in adverse CEO vaccine reactions. All suspect LT cases are evaluated by real time PCR and histopathology of eyelid and trachea for confirmation. 4. GA conducted Mycoplasma diagnostics/ surveillance in commercial poultry. Diagnostic tests conducted over the past year include 1076 cultures, 6884 HI tests, 3613 PCR tests and 141 sequencing reactions. 5. SEPRL on their international surveillance and characterization of avian influenza H5N1 subtypes indicated that the Egypt remains one of a handful of countries where the H5N1 bird flu continues to infect poultry 6. SEPR determined the recent H5N1 highly pathogenic avian influenza (HPAI) viruses circulating in Vietnam was evaluated in domestic ducks. One of the viruses, A/duck/Vietnam/NCVD-672/2011 (clade 2.3.2B), was highly virulent for ducks but the other virus, A/chicken/Vietnam/NCVD-675/2011 (clade 2.3.2A) was moderately pathogenic 7. SEPRL Strains of NDV obtained recently from Mexico, Indonesia, Malaysia, Venezuela, Pakistan, Vietnam, Belize, Dominican Republic, South Africa, and Peru and from wild birds from the U.S. have been sequenced and characterized genetically. Objective II: Develop improved diagnostic capabilities including real-time PCR as well as other rapid on-farm tests for economically important respiratory diseases. 1. CT in collaboration with Guangxi Veterinary Institute, China developed loop-mediated isothermal amplification (LAMP) assays to detect the Mycoplasma gallisepticum isolates. The newly developed LAMP assay is simple, sensitive, rapid and can identify Mycoplasma gallisepticum isolates visually. Consequently, this assay will be very useful screening assays. 2. DE, developed an IBV Arkansas genotype-specific real-time RT-PCR primer and probe set that could be seamlessly integrated with the published IBV real-time protocol within a multiplex real-time RT-PCR reaction 3. DE sequenced a recent 2011 ILTV field isolate from the Delmarva Peninsula that has demonstrated the capacity to break through a vaccinated flock. 4. GA Develop a multiplex assay to detect avian infectious bronchitis virus types. Four most common IBV serotypes diagnosed in the USA; Arkansas (Ark), Connecticut (Conn), Delaware (DE). 5. GA developed and validated a quantitative method for detection of infectious laryngotracheitis virus (ILTV) in clinical and laboratory samples. This methodology allows for the quantitation of virus copy numbers in a given clinical sample and it also allows for elimination of cumbersome classical virology and serology methods. 6. IL genetically characterized a vaccine strain of fowlpox virus involved in outbreaks in vaccinated flocks and determined that the reticuloendotheliosis virus genome was present in the fowlpox virus genes. 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. MN genetically analyzed the matrix (M) gene of avian influenza viruses isolated from wild birds and from the live bird markets indicated that independent evolution of M gene in the absence of antiviral drugs will lead to mutation causing resistance. 2. MN demonstrated during surveillance program that water borne transmission of influenza A virus likely occurs. 3. OH, investigated the replication of swine and human influenza A viruses in juvenile and layer turkeys. OH, noticed an enhanced replication of swine influenza viruses in immune compromised (dexamethasone-treated) juvenile and layer turkeys. 4. OH, assessed and quantified apoptosis and T cell mediated cytotoxicity in IBDV infected chickens. 5. OH, investigated an attachment of avian and mammalian influenza viruses in the respiratory and reproductive tracts of layer turkey hens 6. OH, demonstrated persistence and tissue distribution of infectious bursal disease virus in experimentally infected SPF and commercial broiler chickens 7. OH, showed the molecular evidence for a geographically restricted population of infectious bursal disease viruses. 8. OH, demonstrated the diversity of genome segment B from infectious bursal disease viruses in the United States SEPRL, Identified genetic and biological determinants of tissue tropism and transmission of avian influenza virus in chickens. 9. SPRL has determined gross and microscopic lesions from Newcastle disease virus (NDV) infections with novel circulating NDV of different genotypes in various species of birds. Objective IV: Develop new prevention and control strategies for poultry respiratory diseases. 1. AL developed a transgenic plat vaccine against avian influenza. 2. CT generated throughput gene sequence data of IBV field isolates from the commercial poultry flocks vaccine with various IBV vaccines. 3. DE generated a new generation ILT vaccine containing deletions in essential genes. 4. GA examined the dynamics of IBV vaccination and protecting poultry against Arkansas field strains of IBV.GA developed mutant vaccine against infectious laryngotrachitis infection. 5. IN investigated the infectious bursal disease kinetics using DNA vaccine in chickens. 6. SEPRL performed vaccine efficacy studies using circulating AI viruses from Vietnam. 7. SEPRL developed new vaccine platforms to control and prevent avian influenza outbreaks. 8. SEPRL determined the AI vaccine efficacy following vaccination with recombinant herpesvirus of turkey-vectored avian influenza vaccine against highly pathogenic H5N1 challenge.

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Impact Statements

1. Continuous surveillance and characterization of ILTVs from poultry house environments would help in the understanding of the origin, evolution, transmission and control of present and future ILTV outbreaks. Composting litter, a through cleanout out and disinfection of a house, and possibly the use of commercial recombinant vaccines given in ovo, will reduce the incidence and severity of subsequent ILTV outbreaks.
2. Avian influenza subtype H5 and H7 were negative from the LBM and domestic poultry birds in New England states in Delaware and Ohio commercial farms. However wild birds do carry H5 and H7 subtypes in their population and continued surveillance is warranted.
3. Infectious laryngotracheitis virus and infectious bronchitis virtues circulating in commercial broiler chickens flocks in Alabama, Delaware, and Georgia states. Surveillance activities on the Delmarva Peninsula have yielded infectious laryngotracheitis virus (ILTV) and infectious bronchitis virus (IBV) isolates from commercial broiler chickens and an avian paramyxovirus (APMV)-4 isolate from wild birds.
4. Quasi-species phenomenon in the IBV strains occurring in the field. This identification and characterization will be helpful for designing the better vaccines against IBV infections for poultry.
5. Continuous surveillance and characterization of MG from poultry would help in the understanding of the origin, evolution, transmission and control of present and future. Development of rapid tool loop mediated isothermal polymerase to identify MG infection will be very cost effective without the use of sophisticated and expensive thermal cyclers.
6. Multiplex assay to detect avian infectious bronchitis virus serotypes. For an additional $0.21 per reaction, multiplexing a Arkansas genotype specific with the universal infectious bronchitis virus (IBV) rRT-PCR assay permitted detection of the most common genotype in Delmarva broilers without impacting test sensitivity. Monitoring infectious bronchitis viruses from commercial broiler chickens is important for evaluating the effectiveness of vaccination programs and to isolate and characterize field viruses that break through vaccine induced immunity.
7. Method of delivery of Ark vaccines fully protects broilers. This is important for control of IBV Ark type viruses in the field.
8. Both traditional and recombinant-based approaches for the construction of the next generation of infectious laryngotracheitis virus (ILTV) live vaccines. Infectious laryngotracheitis is an economic disease that also has important trade implications for the U.S. poultry industry. Vaccination using CEO and recombinant vaccines is helping control the disease but more research is warranted to develop improved vaccines and control strategies.
9. Quantitative tool to detect ILTV in birds can be used to establish the viral load in chickens, which provides valuable data for estimating transmission and control.
10. IBDV large segment gene-based DNA vaccination in inhibiting and/or eliminating infectious bursal disease virus infection as illustrated by DNA vaccination kinetics and bursal transcriptome has the great potential for practical use in the field for protection of chickens against infectious bursal disease in the poultry industry.
11. PCR amplification of selected genomic fragments from DNA isolated from formalin fixed tissue sections of histologically positive cases of avianpox virus infection is convenient for genetic characterization of these viruses.
12. Rapid aptamer-based approach that will enable faster and cost-effective identification of influenza virus in animal samples.
13. Swine influenza viruses (SIVs) continue to be a threat for turkey industry and immunosuppression of the bird may enhance the transmission and adaptation of swine influenza viruses in turkeys through enhancement of virus replication, prolonged virus shedding, and possible decrease of infectious dose required to initiate infection.
14. Virus histochemistry can be applied as a useful in vitro screening tool to predict the in vivo replication of influenza virus which may help to reduce the use of live animals and research cost.
15. Studies provide new insights into the pathogenesis of IBDV and provide mechanistic evidence that the cytotoxic T cells may act through both Fas-FasL and perforin-granzyme pathways in mediating the clearance of virus-infected cells. The findings can be used to develop novel target for IBDV control.
16. IBDV RNA can be detected in thigh and breast muscles for short period of time. However, the presence of vRNA is not indicative of the presence of the infectious virus and does not necessarily correlate with virus isolation data. The first detailed report on the persistence and distribution of classic and variant strains of IBDV in different tissues of SPF and commercial chickens will be useful for risk assessment and develop prevention strategy.
17. The phylogeographic data suggest specific population of IBDV has been restricted for over 14 years to Northeast Ohio. Since commercially available classic and variant vaccines do not effectively control this population of IBDV, other alternatives are needed.
18. Molecular epidemiology study of IBDV shows the evidence of recombination events, in addition to reassortment, in creating genetic diversity both in variant and classic strains. Furthermore, the study shows importance and usefulness of analyzing genome segment B during routine molecular diagnosis of all IBDV strains.
19. Gene mutations detected in AIV in Egypt is more difficult to control outbreaks, because the vaccine is less effective against these mutant groups of AIV.
20. Serious concern for the control of H5N1 in Vietnam must consider the important role of domestic ducks in the epidemiology of H5N1 HPAI .
21. Information has implications for infection through artificial insemination and shows that the AI virus can replicate in the reproductive tract, which may mean the virus can be found in or on eggs.
22. An edible transgenic plant vaccine against the H5 and H7 AIV subtypes, which could be mixed in poultry feed, could be farther developed for use in controlling AIV in chickens, in 3rd world countries. This is important since these poorer countries are a constant source of AIV infections in poultry and swine populations. In addition, vaccines against animals are needed to prevent future pandemics in humans, which contain triple reassortments of AIVs from birds, humans, and swine. Recombinant vaccine can be used as an aid during AI eradication efforts in turkey species.
23. Proper identification of the disease signs, which are crucial to quickly preventing the spread NDV. The virulent NDV that are found in the U.S. in pigeons (genotype VIb) and cormorants (genotype V) and the virulent NDV (genotype V) from the last 2002 U.S. outbreak also produces few gross lesions upon infection of poultry, unlike what is seen world-wide from other virulent NDV (genotypes VII-X111).
24. New vaccine candidates are being evaluated by a vaccine company for distribution worldwide to improve NDV control. The benefit of these vaccines is their ability to decrease the amount of virus put into the environment by vaccinated birds infected with virulent NDV.

Date of Annual Report: 02/24/2014

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Annual Meeting Dates: 12/08/2013 - 12/08/2013
Period the Report Covers: 10/01/2008 - 12/01/2013

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NC1180's Termination Report is attached as the below "Copy of Minutes" File