Source: UNIVERSITY OF DELAWARE submitted to
CONTROL OF EMERGING AND RE-EMERGING POULTRY RESPIRATORY DISEASES IN THE UNITED STATES
Sponsoring Institution
National Institute of Food and Agriculture
Project Status
TERMINATED
Funding Source
HATCH
Reporting Frequency
Annual
Accession No.
0220195
Grant No.
(N/A)
Project No.
DEL00664
Proposal No.
(N/A)
Multistate No.
NC-_OLD1180
Program Code
(N/A)
Project Start Date
Oct 1, 2009
Project End Date
Sep 30, 2014
Grant Year
(N/A)
Project Director
Gelb, JA.
Recipient Organization
UNIVERSITY OF DELAWARE
(N/A)
NEWARK,DE 19717
Performing Department
Animal And Food Sciences
Non Technical Summary
The control of emerging and re-emerging poultry respiratory diseases in the United States is a major challenge facing scientists and producers. Our project will focus on performing ongoing surveillance of respiratory disease and the major viral etiologies. Our efforts will then focus on understanding how viruses cause respiratory disease and how the diseases may be prevented or controlled to sustain poultry health. A major emphasis will be placed on three classic viral diseases that continue to cause widespread disease, infectious laryngotracheitis and infectious bronchitis or pose a threat, avian influenza virus. Laboratory in vitro studies will be combined with animal studies to develop the best data for basing conclusions that will lead to improved prevention and control measures.
Animal Health Component
100%
Research Effort Categories
Basic
(N/A)
Applied
(N/A)
Developmental
(N/A)
Classification

Knowledge Area (KA)Subject of Investigation (SOI)Field of Science (FOS)Percent
3113220104025%
3113220106025%
3113220109015%
3113220110135%
Knowledge Area
311 - Animal Diseases;

Subject Of Investigation
3220 - Meat-type chicken, live animal;

Field Of Science
1040 - Molecular biology; 1090 - Immunology; 1060 - Biology (whole systems); 1101 - Virology;
Goals / Objectives
Identify reservoirs of infectious respiratory disease agents in wild birds and poultry. Develop improved diagnostic capabilities including real time PCR as well as other rapid on-farm tests for economically important respiratory diseases. Investigate the pathogenesis and polymicrobial interactions of specific infectious agents associated with poultry respiratory diseases (this includes interactions with underlying immunosuppressive agents). Develop new prevention and control strategies for poultry respiratory diseases.
Project Methods
1. Identify reservoirs of infectious respiratory disease agents in wild birds and poultry. Delaware will perform surveillance of avian influenza virus in wild aquatic birds, backyard poultry and commercial poultry. (Gelb, Ladman, and Bautista). Delaware will conduct surveillance of infectious laryngotracheitis virus, infectious bronchitis virus, Newcastle disease virus, infectious bursal disease virus (immunosuppressive agent). (Bautista, Dohms, Gelb, Keeler, Ladman, Ruano) 2. Develop improved diagnostic capabilities including real time PCR as well as other rapid on-farm tests for economically important respiratory diseases. Delaware will identify key ILTV gene targets for diagnostic application (Schmidt, Keeler, Parcells, and Ruano). 3. Investigate the pathogenesis and polymicrobial interactions of specific infectious agents associated with poultry respiratory diseases (this includes interactions with underlying immunosuppressive agents). Delaware will evaluated the pathogenicity of avian influenza virus isolates for various poultry species. (Gelb, Ladman, Pope). Delaware will develop an ILTV transmission model for comparing isolates, reference strains and CEO vaccines (Ruano and Gelb). Delaware will characterize the pathogenic and antigenic properties of IBDV field strains (Gelb, Ladman and Bautista). 4. Develop new prevention and control strategies for poultry respiratory diseases. Delaware will evaluate the efficacy of IBV vaccines to protect vs. variant virus challenge (Gelb, Ladman, and Pope). Delaware will evaluate the efficacy of recombinant vaccines to protect vs. ND and ILT challenges (Bernberg, Morgan, Gelb, and Ladman).

Progress 10/01/09 to 09/30/14

Outputs
Target Audience:Poultry Production Companies Poultry Biologics/Vaccine Companies Other Poultry Scientists including Researchers of this Multistate Project Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?We regularly attended the American Assn. of Avian Pathologists (AAAP)/AVMA combined annual meetings during the period of performance. We also regularly attended the National Meeting on Poultry Health and Processing (NMPHP) meetings during the period. How have the results been disseminated to communities of interest?Yes. Results have been disseminated at AAAP/AVMA, NMPHP via abstracts, conference proceedings, and presentations. We have also met with broiler production companies, Allen Harim Foods, Amick Farms, Mountaire Farms, Perdue Farms, and Tyson Farms with operations on the Delmarva peninsula to share important findings. What do you plan to do during the next reporting period to accomplish the goals? Nothing Reported

Impacts
What was accomplished under these goals? What was accomplished under these goals? Obj. 1. General Ongoing diagnostic surveillance efforts on the Delmarva Peninsula though the University of Delaware Poultry Health System (UDPHS) have yielded high numbers of infectious laryngotracheitis virus (ILTV) and infectious bronchitis virus (IBV) isolates from commercial broiler chickens, and avian influenza virus isolates from wild birds. Lentogenic Newcastle disease virus (NDV) is also detected in clinical samples albeit at a much lower rate compared to IBV and ILTV. Detection of AIV, IBV, ILTV and NDV is performed using molecular (PCR based) testing. The awareness of these avian viruses provides justification for biosecurity and vaccination programs as well as continuing research. Avian Influenza Virus (AIV) Real time RT-PCR detection of low pathogenicity AIV in pools of 11 individual oropharyngeal (O/P) swabbings from broiler chickens was readily detected suggesting that this single flock pooling procedure may be used instead of testing two pools of five and six O/P swabbings per flock. The practice of using a single pool of 11 O/P swabbings has been adopted by USDA APHIS for performing AIV mandated real time RT-PCR surveillance nationwide. Clearly, the use of the single pool will substantially reduce the testing costs and save millions of dollars annually. National Animal Health Laboratory Network avian influenza surveillance testing at the UDPHS detected an H7 subtype in non-commercial backyard auction poultry in Delaware in 2014. Trace forward and backward efforts lead to successful resolution. No Delmarva commercial flocks were found to be AIV positive. Surveillance testing is vital to protecting the region's $2 billion/year broiler industry. From 2006 - 2010, 46 low path (LP) AIV isolates were isolated from wild waterfowl and the shorebird species Ruddy Turnstone. The most commonly isolated H subtype was H6 (six isolates) while the most common N subtype was N2 (nine isolates). Five of the six H6 isolates were N2 subtype viruses. Three isolates of the H7 and H5 subtypes (one H7N3 and two H5N2) were recovered. Based upon their HA sequence, none of the three viruses contained HPAIV cleavage site sequence motifs. Many other H and N subtype combinations were also isolated. The study shows that wild birds are able to harbor subtypes that are important to the poultry industry and reinforces the notion that limiting the interaction between wild birds and commercial poultry is an important aspect of influenza control. H5N2 and H7N3 subtypes, of particular concern because of their potential to become highly pathogenic. Avian influenza virus research demonstrated serial passage of a duck origin low path H5N1 North American virus in broiler chickens expanded the genomic diversity and favored accumulation of basic amino acids at the HA cleavage site in the broiler chicken-adapted virus. Avian Paramyxovirus (APMV) In 2010, an APMV-4 was isolated from a hunter-harvested American black duck in Delaware. In 2012, the genome of the isolate, APMV-4/duck/Delaware/549227/2010 was characterized and found to be 15,048 nucleotides (nt), which is shorter by 6 nt from all previously reported strains. To date complete genome sequences of APMV-4 are available for strains that were isolated from ducks in Hong Kong, Korea, Belgium, and South Africa, over a period of 37 years, but this is the first for a North America isolate. Phylogenetic analysis revealed the Delaware strain formed a distant unique cluster, demonstrating genetic diversity of APMV-4. The GenBank accession number of the Delaware strain is 73 JX987283. Infectious Bronchitis Virus (IBV) A nephropathogenic strain of IBV referred to as DMV/1639/11 appears to have been introduced into Delmarva peninsula region from Pennsylvania and has now become a widespread epornitic in the region causing renal as well as respiratory forms of IB. Vaccination with commercially available live vaccines offer only partial protection against challenge with DMV/1639/11. An experimental autogenous attenuated 1639 type vaccine has been developed and will be field-tested in winter of 2015-2016. Infectious Laryngotracheitis Virus (ILTV) Next generation sequencing technologies now permit the relatively rapid determination of the primary sequence of the ILTV genome. "Next Gen "data will greatly enhance epiemiological investigations in the future. Obj. 2. IBV Since 2005, Arkansas (Ark) genotype IBVs were isolated from diagnostic specimens at a greater rate than any other genotype. Given the high frequency of Ark virus isolations, we sought to develop an alternative method of Ark detection. We developed an Ark genotype-specific real-time RT-PCR primer and probe set that was integrated with the published (Callison et al., 2006) IBV real-time protocol within a multiplex real-time RT-PCR reaction. With excellent specificity, good sensitivity, and a much faster turn-around-time than traditional VI followed by sequencing, the UDPHS has been able to offer rapid genotyping results to the poultry industry. Primers for real-time RT-PCR for IBV nephropathogenic strain DMV/1639/11 were developed and validated. The test was added to our UDPHS molecular panel for rapid diagnosis of respiratory viral diseases. ILTV We are continuing efforts to sequence complete genomes of ILTV field isolates from the Delmarva Peninsula over a 25 year period (1985-2012). The sequences will be used to determine the molecular phylogeny of isolates from the region. In 2012, a 2011 ILTV field isolate from Delmarva that has demonstrated the capacity to break through a vaccinated flock was sequenced and is undergoing sequence analysis. Obj. 3 No major accomplishments to report. Obj. 4 IBV Vaccination with IBV vaccines Massachusetts serotype Ma5 + Delaware 072 offered no protection against challenge with Arkansas DPI. The lack of protection supports field observations and the high numbers of Arkansas virus isolations from Delmarva broiler chicken respiratory disease cases in Fall of 2013 and Winter of 2014. ILTV Both traditional and recombinant-based approaches are being used for the construction of the next generation of ILTV live vaccines. Drinking water vaccination is the primary method of administration of infectious laryngotracheitis virus (ILTV) chicken embryo origin vaccines. A controlled laboratory study indicated a commercial vaccine given at the recommended 1X dose did not achieve flock coverage on day 5 post vaccination compared to broilers given a 10X dose. Higher doses of vaccine may be required to achieve more complete coverage in drinking water applications to avoid potential back passage of vaccine and associated disease side effects. The application of this research has lead to improved ILTV vaccination methods and results in Delmarva commercial broiler chickens. Serial passage of infectious laryngotracheitis virus (ILTV) at a reduced incubation temperature of 30°C for 32 passages gave encouraging results as a method to attenuate the virus. A patent application has been filed. See "Product" section of this final report.

Publications

  • Type: Journal Articles Status: Published Year Published: 2013 Citation: Parthiban, M., K. Manimaran, S. Xiao, B. Nayak, A. Paldurai, S. Kim, B.S. Ladman, L.A. Preskenis, J. Gelb Jr., P.L. Collins and S.K. Samal. Complete genome sequence of an avian paramyxovirus type 4 from North America reveals a shorter genome and new genotype. Genome Announcements (genome A). January/February 2013 vol. 1 no. 1 e00075-12.
  • Type: Journal Articles Status: Published Year Published: 2013 Citation: Spackman, E., J.C. Pedersen, E.T. McKinley and J. Gelb Jr. Optimal specimen collection and transport methods for the detection of avian influenza virus and Newcastle disease virus. BMC Vet Res. 2013 22:9:35. doi: 10.1186/1746-6148-9-35.
  • Type: Journal Articles Status: Published Year Published: 2013 Citation: Wang, Y., B. Ladman, C. Wu, J. Gelb Jr. and S. Golovan. Comparison of vRNA and cRNA based reporters for detection of influenza replication. Antiviral Research. 98: 76084. 2013.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2011 Citation: Ladman, B. S., G. V. Oldfield, C. R. Pope, L. A. Preskenis, S. K. Samal, and J. Gelb, Jr. Characterization of avian paramyxoviruses isolated from migratory waterfowl in chickens, turkeys and ducks. Proc. 148th American Veterinary Medical Assn./American Assn. Avian Pathologists Ann. Mtg. St. Louis, Missouri, July 16-19, 2011.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2012 Citation: Gelb, Jack, Jr., Brian Ladman, Conrad Pope, Miguel Ruano, Erin Brannick, and Daniel Bautista, Characterization of nephropathogenic infectious bronchitis virus from Delmarva broiler chickens 2011. Proc. 149th American Veterinary Medical Assn./American Assn. Avian Pathologists Ann. Mtg. San Diego, California, August, 2012.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2012 Citation: Ladman, B. S., L. A. Preskenis, M. Murphy, M. Ruano, B. F. Sample, D. A. Bautista and J. Gelb, Jr. Development and field evaluation of a multiplex real-time RT-PCR assay for detecting Arkansas infectious bronchitis virus. Proc. 149th American Veterinary Medical Assn./American Assn. Avian Pathologists Ann. Mtg. San Diego, California August 4-7, 2012.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2013 Citation: Ladman, B., J. Gelb, Jr., C. Pope, E. Brannick, E. Spackman, and R. Slemons. Evaluating the Genetic Diversity of a Wild Bird Origin H5N1 Low Path Avian Influenza Viral Genome After Adaptation to Different Poultry. Proc. 150th American Veterinary Medical Assn./American Assn. Avian Pathologists Ann. Mtg. Chicago, Illinois, July 21-23, 2013.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2014 Citation: Gelb, Jack, Jr., Daral J. Jackwood, Brian S. Ladman, and Erin M. Brannick. Studies on the efficacy of recombinant HVT-IBD vaccines-Vaxxitek trial. Proc. 151st American Veterinary Medical Assn./American Assn. Avian Pathologists Ann. Mtg. Denver, Colorado, July 26-29, 2014.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2014 Citation: Jackwood, Daral J., Brian S. Ladman, and Erin Brannick. Studies on the efficacy of recombinant HVT-IBD vaccines-Vectormune trial. Proc. 151st American Veterinary Medical Assn./American Assn. Avian Pathologists Ann. Mtg. Denver, Colorado, July 26-29, 2014.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2014 Citation: Ladman, Brian, Lauren Preskenis, Marcy Murphy, Miguel Ruano, Daniel Bautista & Jack Gelb, Jr. Infectious bronchitis virus isolate DMV/2392/12 from Delmarva broiler chickens-Epidemiology and protection afforded by the Delaware 072 and Georgia 98 commercial vaccines. Proc. 151st American Veterinary Medical Assn./American Assn. Avian Pathologists Ann. Mtg. Denver, Colorado, July 26-29, 2014.
  • Type: Journal Articles Status: Published Year Published: 2012 Citation: Gelb J., Jr., D. J. Jackwood, E. Mundt, C. R. Pope, R. Hein, G. Slacum, J. M. Harris, B. S. Ladman, P. Lynch, D. A. Bautista, J. M. Ruano, and M. M. Troeber. Characterization of infectious bursal disease viruses isolated in 2007 from Delmarva commercial broiler chickens. Avian Dis. 56:82-89. 2012.
  • Type: Journal Articles Status: Published Year Published: 2012 Citation: Ladman, B. S., E. Spackman, J. Gelb, Jr. Comparison of pooling eleven or five oropharyngeal swabbings for detecting avian influenza virus by real-time RT-PCR in broiler chickens. Avian Dis. 56:227-229. 2012.
  • Type: Journal Articles Status: Published Year Published: 2012 Citation: Spatz, S. J., J. D Volkening, and C. L Keeler, G. F. Kutish, S. M. Riblet, C. M. Boettger, K. F. Clark, L. Zsak, C. L. Afonso, E. S. Mundt, D.L. Rock, and M. Garcia. Comparative full genome analysis of four infectious laryngotracheitis virus (Gallid herpesvirus-1) virulent isolates from the United States. Virus Genes 44:273-285. 2012.
  • Type: Journal Articles Status: Published Year Published: 2013 Citation: Gelb, J. Jr., B.S. Ladman, C.R. Pope, J.M. Ruano, E.M. Brannick, D.A. Bautista, C.M. Coughlin and L.A. Preskenis. Characterization of nephropathogenic infectious bronchitis virus DMV/1639/11 recovered from Delmarva broiler chickens in 2011. Avian Dis. 57:65-70. 2013
  • Type: Journal Articles Status: Published Year Published: 2013 Citation: Maughan, M.N., L.S. Dougherty, L.A. Preskenis, B.S. Ladman, J. Gelb, Jr., E. Spackman and C. Keeler, Jr., Transcriptional analysis of the innate immune response of ducks to different species-of-origin low pathogenic H7 avian influenza viruses. Virology Journal. 2013 Mar 23:10:94. doi: 10.1186/1743-422X-10-94.


Progress 10/01/13 to 09/30/14

Outputs
Target Audience:U.S. poultry industry, specifically chicken broiler and egg producers. Other researchers participating in same or similar multistate USDA project(s). Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided? Nothing Reported How have the results been disseminated to communities of interest?Yes. Gelb, Ladman, and Bautista met with broiler industry personnel to convey findings. What do you plan to do during the next reporting period to accomplish the goals?Will continue work on the above disease agents.

Impacts
What was accomplished under these goals? Obj. 1 In 2013-14, the combination of an extremely cold winter, very high numbers of IBV, ILTV or IBV/ILTV (co-infection) respiratory disease cases made the winter of 2013-14 one of the worst in recent memory for broiler chicken production on the Delmarva peninsula. Moreover, a major change in IBV broiler vaccination programs appeared to contribute to problem. National Animal Health Laboratory Network Avian influenza surveillance testing at the University of Delaware detected an H7 subtype in non-commercial backyard auction poultry in Delaware in 2014. Trace forward and backward efforts lead to successful resolution. No commercial flocks were infected in Delmarva. Surveillance testing is vital to protecting the region's $2 billion broiler industry. Avian influenza wild bird surveillance from 2007-2010 in Delaware yielded 39 isolates from waterfowl and shorebird species Ruddy Turnstone This study shows that wild birds are able to harbor subtypes that are important to the poultry industry and reinforces the notion that limiting the interaction between wild birds and commercial poultry is an important aspect of influenza control. H5N2 and H7N3 subtypes, of particular concern because of their potential to become highly pathogenic, were recovered from waterfowl. Many other H and N subtype combinations were also isolated. Obj. 2 Primers for real-time RT-PCR for IBV variant DMV/1639/11 were developed and validated and the test was added to our molecular respiratory panel for rapid diagnosis. The ability to sequence the ILTV genome from primary clinical samples will assist in determining the molecular ecology of this pathogen. Obj. 3 Vaccination with IBV vaccines Massachusetts serotype Ma5 + Delaware 072 offered no protection against challenge with Arkansas DPI. The lack of protection supports field observations and the high numbers of Arkansas virus isolations from Delmarva broiler chicken respiratory disease cases in Fall of 2013 and Winter of 2014. Drinking water vaccination is the primary method of administration of infectious laryngotracheitis virus (ILTV) chicken embryo origin vaccines. A controlled laboratory study indicated a commercial vaccine given at the recommended 1X dose did not achieve flock coverage on day 5 post vaccination compared to broilers given a 10X dose. Higher doses of vaccine may be required to achieve more complete coverage in drinking water Serial passage of infectious laryngotracheitis virus (ILTV) at a reduced incubation temperature of 30°C for 32 passages gave encouraging results as a method to attenuate the virus.

Publications

  • Type: Conference Papers and Presentations Status: Accepted Year Published: 2014 Citation: Gelb, Jack, Jr., Daral J. Jackwood, Brian S. Ladman, and Erin M. Brannick. Studies on the efficacy of recombinant HVT-IBD vaccines-Vaxxitek trial. Proc. 151st American Veterinary Medical Assn./American Assn. Avian Pathologists Ann. Mtg. Denver, Colorado, July 26-29, 2014.
  • Type: Conference Papers and Presentations Status: Accepted Year Published: 2014 Citation: Jackwood, Daral J., Brian S. Ladman, and Erin Brannick. Studies on the efficacy of recombinant HVT-IBD vaccines-Vectormune trial. Proc. 151st American Veterinary Medical Assn./American Assn. Avian Pathologists Ann. Mtg. Denver, Colorado, July 26-29, 2014.
  • Type: Conference Papers and Presentations Status: Accepted Year Published: 2014 Citation: Ladman, Brian, Lauren Preskenis, Marcy Murphy, Miguel Ruano, Daniel Bautista & Jack Gelb, Jr. Infectious bronchitis virus isolate DMV/2392/12 from Delmarva broiler chickens-Epidemiology and protection afforded by the Delaware 072 and Georgia 98 commercial vaccines. Proc. 151st American Veterinary Medical Assn./American Assn. Avian Pathologists Ann. Mtg. Denver, Colorado, July 26-29, 2014.


Progress 01/01/13 to 09/30/13

Outputs
Target Audience: Poultry Production Companies PoultryBiologics/Vaccine Companies Other Poultry Scientists Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided? Ladman, B., J. Gelb, Jr., C. Pope, E. Brannick, E. Spackman, and R. Slemons. Evaluating the Genetic Diversity of a Wild Bird Origin H5N1 Low Path Avian Influenza Viral Genome After Adaptation to Different Poultry. Proc. 150th American Veterinary Medical Assn./American Assn. Avian Pathologists Ann. Mtg. Chicago, Illinois, July 21-23, 2013. How have the results been disseminated to communities of interest? IV. APPLICATION OF FINDINGS: Avian influenza virus Avian influenza virus subpopulations change as the virus adapts to other poultry hosts. Surveillance of AIV in wild bird and poultry populations is warranted so that monitoring of the H and other genes can be used to possibly predict a change in pathogenicity. Infectious bronchitis virus Monitoring infectious bronchitis viruses from commercial broiler chickens is important for assessing the effectiveness of vaccination programs. In 2012, IBV DMV/2392/12 was isolated from a Delmarva broiler chicken flock. The virus was genetically characterized and found to be related (~90% by S1 gene sequence analysis) to reference strain DE/072/92. In previous studies, we have found that percent relatedness values less than 90% may not correlate with protection in a vaccinated chicken. A vaccination – challenge of immunity study was performed to evaluate potential protection provided by commercial vaccine strain Delaware 072 and Georgia 98 vs. challenge with DMV/2392/12. The findings suggested that vaccination with Shore-Bron-D (DE072) or Mildvac-GA-98 (GA98) afforded broiler chickens 86% and 79% protection, respectively, against a challenge of the field strain DMV/2392/12. These results suggest that current live IBV vaccines Shore-Bron-D (DE072) and Mildvac-GA-98 (GA98) may be used to protect commercial flocks against a challenge DMV/2392/12. Infectious laryngotracheitis virus Infectious laryngotracheitis is an economic disease that also has important trade implications for the poultry industry. Vaccination using Chicken-embryo-origin (CEO) and recombinant vaccines is helping control the disease but more research is warranted to develop improved vaccines and control strategies. Improvements in sequencing technologies have enhanced our ability to monitor ILTV field isolates. CEO strains that exhibit reduced pathogenicity are being characterized for their potential utility as live attenuated vaccines. What do you plan to do during the next reporting period to accomplish the goals? V. FUTURE WORK: Avian influenza, infectious bronchitis, infectious laryngotracheitis, E. coli, and infectious bursal disease. Will continue work on the above disease agents.

Impacts
What was accomplished under these goals? Surveillance activities on the Delmarva Peninsula have yielded infectious bronchitis virus isolates from commercial broiler chickens. In 2012-13, a virus related to the Delaware 072 genotype virus was frequently recovered. Avian influenza virus research demonstrated serial passage of a duck origin low path H5N1 North American virus in other poultry species expanded the genomic diversity and favored accumulation of basic amino acids at the HA cleavage site in broiler chicken-adapted virus. The respiratory PCR panel provides a way to rapidly identify IBV vaccine strains and provides a way to identify potential non-vaccine genotypes. Next generation sequencing technologies permit the relatively rapid determination of the primary sequence of the ILTV genome from egg-passaged material.

Publications

  • Type: Journal Articles Status: Published Year Published: 2013 Citation: Gelb, J. Jr., B.S. Ladman, C.R. Pope, J.M. Ruano, E.M. Brannick, D.A. Bautista, C.M. Coughlin and L.A. Preskenis. Characterization of nephropathogenic infectious bronchitis virus DMV/1639/11 recovered from Delmarva broiler chickens in 2011. Avian Dis. 57:65-70
  • Type: Conference Papers and Presentations Status: Published Year Published: 2013 Citation: Ladman, B., J. Gelb, Jr., C. Pope, E. Brannick, E. Spackman, and R. Slemons. Evaluating the Genetic Diversity of a Wild Bird Origin H5N1 Low Path Avian Influenza Viral Genome After Adaptation to Different Poultry. Proc. 150th American Veterinary Medical Assn./American Assn. Avian Pathologists Ann. Mtg. Chicago, Illinois, July 21-23, 2013
  • Type: Journal Articles Status: Published Year Published: 2013 Citation: Maughan, M.N., L.S. Dougherty, L.A. Preskenis, B.S. Ladman, J. Gelb, Jr., E. Spackman and C. Keeler, Jr., Transcriptional analysis of the innate immune response of ducks to different species-of-origin low pathogenic H7 avian influenza viruses. Virology Journal. 2013 Mar 23:10:94. doi: 10.1186/1743-422X-10-94.
  • Type: Journal Articles Status: Published Year Published: 2013 Citation: Parthiban, M., K. Manimaran, S. Xiao, B. Nayak, A. Paldurai, S. Kim, B.S. Ladman, L.A. Preskenis, J. Gelb Jr., P.L. Collins and S.K. Samal. Complete genome sequence of an avian paramyxovirus type 4 from North America reveals a shorter genome and new genotype. Genome Announcements (genome A). January/February 2013 vol. 1 no. 1 e00075-12.
  • Type: Journal Articles Status: Published Year Published: 2013 Citation: Spackman, E., J.C. Pedersen, E.T. McKinley and J. Gelb Jr. Optimal specimen collection and transport methods for the detection of avian influenza virus and Newcastle disease virus. BMC Vet Res. 2013 22:9:35. doi: 10.1186/1746-6148-9-35.
  • Type: Journal Articles Status: Published Year Published: 2013 Citation: Wang, Y., B. Ladman, C. Wu, J. Gelb Jr. and S. Golovan. Comparison of vRNA and cRNA based reporters for detection of influenza replication. Antiviral Research. 98: 76084. 2013.


Progress 01/01/12 to 12/31/12

Outputs
OUTPUTS: Obj. 1 In spring 2012, disease surveillance in Delmarva commercial broiler chicken flocks identified five infectious bronchitis virus isolates of the Delaware 072 genotype. Four of five isolates were highly related by S-1 subunit spike (S) gene sequence analysis while DMV/9422/12 appeared in a different clade. DMV/9422/12 displayed a 89% S1 amino acid identity with reference strains Delaware 072 and Georgia 98, however the findings also demonstrated a 99% identity to a 2012 isolate from Pennsylvania, PA/3365A/12, from a commercial broiler flock. In 2010, an avian paramyxovirus type 4 (APMV-4) was isolated from a hunter-harvested American black duck in Delaware. In 2012, the genome of the isolate, APMV-4/duck/Delaware/549227/2010 was characterized and found to be 15,048 nucleotides (nt), which is shorter by 6 nt from all previously reported strains. To date complete genome sequences of APMV-4 are available for strains that were isolated from ducks in Hong Kong, Korea, Belgium, and South Africa, over a period of 37 years, but this is the first for a North America isolate. Phylogenetic analysis revealed the Delaware strain formed a distant unique cluster, demonstrating genetic diversity of APMV-4. The GenBank accession number of the Delaware strain is 73 JX987283. From 2006 - 2010, 46 low path (LP) avian influenza virus isolates were isolated from wild waterfowl. Twenty-two isolates have been typed to date. The most commonly isolated HA subtype was H6 (six isolates) while the most common N subtype was N2 (nine isolates). Five of the six H6 isolates were N2 subtype viruses. Only three isolates of the H7 and H5 subtypes (one H7N3 and two H5N2) were recovered. Based upon their HA sequence, none of the three viruses contained HPAIV cleavage site sequence motifs. Obj. 2 Since 2005, Arkansas (Ark) genotype IBVs were isolated from diagnostic specimens at a greater rate than any other genotype. Given the high frequency of Ark virus isolations, we sought to develop an alternative method of Ark detection. We developed an Ark genotype-specific real-time RT-PCR primer and probe set that was integrated with the published (Callison et al., 2006) IBV real-time protocol within a multiplex real-time RT-PCR reaction. With excellent specificity, good sensitivity, and a much faster turn-around-time than traditional VI followed by sequencing, the UDPHS has been able to offer rapid genotyping results to the poultry industry. We are continuing in efforts to sequence complete genomes of ILTV field isolates from the Delmarva Peninsula over a 25 year period (1985-2012). The sequences will be used to determine the molecular phylogeny of isolates from the region. In 2012, a 2011 ILTV field isolate from Delmarva that has demonstrated the capacity to break through a vaccinated flock was sequenced and is undergoing sequence analysis. Obj.4 Both traditional and recombinant-based approaches are being used for the construction of the next generation of ILTV live vaccines. PARTICIPANTS: J Gelb is the PI for this project and coordinated the AIV, IBV, and APMV-4 research. C. Keeler directed the ILTV research. B. Ladman, M. Ruano, D. Bautista and E. Brannick contributed significantly to this research and report. Collaborators with C. Keeler: S. Spatz (USDA ARS SEPRL) and M. Garcia (University of Georgia). TARGET AUDIENCES: Scientists/health professionals in the poultry industry, academia and government. PROJECT MODIFICATIONS: Not relevant to this project.

Impacts
Avian influenza virus Monitoring avian influenza virus in wild birds will continue. The H and N genes will be sequenced and selected virus isolates will be evaluated for their potential pathogenicity in poultry. Infectious bronchitis virus 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. Infectious laryngotracheitis virus 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.

Publications

  • Ladman, B. S., L. A. Preskenis, M. Murphy, M. Ruano, B. F. Sample, D. A. Bautista and J. Gelb, Jr. Development and field evaluation of a multiplex real-time RT-PCR assay for detecting Arkansas infectious bronchitis virus. Proc. 149th American Veterinary Medical Assn./American Assn. Avian Pathologists Ann. Mtg. San Diego, California August 4-7, 2012.
  • Spatz, S. J., J. D Volkening, and C. L Keeler, G. F. Kutish, S. M. Riblet, C. M. Boettger, K. F. Clark, L. Zsak, C. L. Afonso, E. S. Mundt, D.L. Rock, and M. Garcia. Comparative full genome analysis of four infectious laryngotracheitis virus (Gallid herpesvirus-1) virulent isolates from the United States. Virus Genes 44:273-285. 2012.
  • Gelb, J. Jr., B. S. Ladman, C. R. Pope, J. M. Ruano, E. M. Brannick, D. A. Bautista, C. M. Coughlin, and L. A. Preskenis. 2013 Characterization of nephropathogenic infectious bronchitis virus DMV/1639/11 recovered from Delmarva broiler chickens in 2011. Avian Diseases (In press)
  • Gelb, Jack, Jr., Brian Ladman, Conrad Pope, Miguel Ruano, Erin Brannick, and Daniel Bautista, Characterization of nephropathogenic infectious bronchitis virus from Delmarva broiler chickens 2011. Proc. 149th American Veterinary Medical Assn./American Assn. Avian Pathologists Ann. Mtg. San Diego, California, August, 2012.
  • Gelb, Jack, Jr., Brian Ladman, Conrad Pope, Miguel Ruano, Erin Brannick, Daniel Bautista, Marcy Troeber, and Lauren Preskenis. Nephrogenic bronchitis. Laboratory characterization. Proc. 46th National Mtg. on Poultry Health and Processing. Ocean City, Maryland. October, 2011.
  • Parthiban, M., K. Manimaran, S. Xiao, B. Nayak, A. Paldurai, S. Kim, B. S. Ladman, L. A. Preskenis, J. Gelb Jr., P. L. Collins and S. K Samal. (2013) Complete genome sequence of an avian paramyxovirus type 4 from North America reveals a shorter genome and new genotype. Genome Announcements (genome A). (In press)


Progress 01/01/11 to 12/31/11

Outputs
OUTPUTS: Routine virus isolation attempts from respiratory disease accessions from Delmarva commercial broiler chickens yielded; infectious bronchitis (IB) vaccine related spike (S) genotypes; Arkansas (59), Massachusetts (13), and Connecticut (0). A limited outbreak of nephrogenic infectious bronchitis (NIB) occurred in three Delmarva commercial broiler chicken flocks in Spring 2011. Findings indicated that the isolates were identical to each other and highly related to a 2010 isolate (PA/9579A/10) from commercial poultry in Pennsylvania that had been sequenced by our laboratory last October. NIB is not new to Pennsylvania. An outbreak of NIB was reported in Lancaster and nearby counties in 1997-2000. The 2010 PA isolate and the 2011 Delmarva isolates appear to have evolved from the earlier PA viruses. No isolations of Newcastle disease virus were made during the period from commercial poultry. No avian influenza (AI) activity using USDA NAHLN-approved agent detection (real time RT-PCR and antigen capture on oropharyngeal swabs) or antibody detection assays were observed in active (pre-slaughter) or passive (clinical disease cases) surveillance samples in commercial broilers or backyard flocks during the period. Delmarva continued to observe significant infectious laryngotracheitis virus (ILTV) activity in 2010 and 2011. 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. Five AIV isolates were recovered from wild birds during the period. Both traditional and recombinant-based approaches are being used for the construction of the next generation of ILTV live vaccines. PARTICIPANTS: Both traditional and recombinant-based approaches are being used for the construction of the next generation of ILTV live vaccines. Collaborator: B.-W. Kong (U. Arkansas). Next generation sequencing technologies now permit the relatively rapid determination of the primary sequence of the ILTV genome. Collaborators: Dr. Maricarmen Garcia (University of Georgia) and Dr. Stephen Spatz (USDA ARS SEPRL). Wild Birds. Five AIV isolates were recovered from wild birds during the period. Collaborators: Delaware Department of Natural Resources & Environmental Control, USDA Wildlife Services Real time RT-PCR detection of low pathogenicity avian influenza virus in pools of 11 individual oropharyngeal (O/P) swabbings from broiler chickens was readily detected suggesting that this single flock pooling procedure may be used instead of testing two pools of five and six O/P swabbings per flock. Collaborator: E. Spackman, USDA-ARS, Athens, Georgia. TARGET AUDIENCES: Nothing significant to report during this reporting period. PROJECT MODIFICATIONS: Not relevant to this project.

Impacts
Surveillance activities on the Delmarva Peninsula have yielded infectious laryngotracheitis virus and infectious bronchitis virus isolates from commercial broiler chickens and avian influenza virus isolates from wild birds. The awareness of the presence of these viruses provides justification for biosecurity and vaccination programs in poultry. A nephropathogenic strain of infectious bronchitis virus appears to have been introduced into Delmarva peninsula region from Pennsylvania and cause three outbreaks on commercial broiler farms. Vaccination with commercially available live vaccines offer partial protection against challenge with DMV/1639/11. Real time RT-PCR detection of low pathogenicity avian influenza virus in pools of 11 individual oropharyngeal (O/P) swabbings from broiler chickens was readily detected suggesting that this single flock pooling procedure may be used instead of testing two pools of five and six O/P swabbings per flock. This practice could susbstantially reduce the costs of surveillance programs. Next generation sequencing technologies now permit the relatively rapid determination of the primary sequence of the ILTV genome. The data will enhance epiemiological investigations.

Publications

  • Gelb J., Jr., D. J. Jackwood, E. Mundt, C. R. Pope, R. Hein, G. Slacum, J. M. Harris, B. S. Ladman, P. Lynch, D. A. Bautista, J. M. Ruano, and M. M. Troeber. Characterization of infectious bursal disease viruses isolated in 2007 from Delmarva commercial broiler chickens. Avian Dis. In press. 2012.
  • Ladman, B. S., G. V. Oldfield, C. R. Pope, L. A. Preskenis, S. K. Samal, and J. Gelb, Jr. Characterization of avian paramyxoviruses isolated from migratory waterfowl in chickens, turkeys and ducks. Proc. 148th American Veterinary Medical Assn./American Assn. Avian Pathologists Ann. Mtg. St. Louis, Missouri, July 16-19, 2011.
  • Ladman, B. S., J. Gelb, Jr., R. Slemons, C. R. Pope, & E. Spackman. Effects of Interspecies adaptation to different poultry on a wild bird origin H5N1 low path avian influenza viral genome. Proc. 148th American Veterinary Medical Assn./American Assn. Avian Pathologists Ann. Mtg. St. Louis, Missouri, July 16-19, 2011.
  • Ladman, B. S., Spackman, E., Gelb,J., Jr. Comparison of pooling eleven or five oropharyngeal swabbings for detecting avian influenza virus by real-time RT-PCR in broiler chickens. Avian Dis. In press. 2012