Duration: 10/01/2024 to 09/30/2029

Administrative Advisor(s):

NIFA Reps:

Non-Technical Summary

The United States (U.S.) is the world's largest poultry producer, the second-largest exporter of poultry meat, and a major producer of eggs and egg products. This project is focused on the detection, monitoring, prevention, and control of infectious diseases of commercial and small flock poultry production.  Infectious diseases are a major concern for poultry producers. Losses induced by infectious diseases of poultry have a major economic impact affecting local, national, and global economies. Many endemic infectious diseases in the U.S. continue to erode profitability and production efficiency. Catastrophic diseases, such as avian influenza, result in more immediate and major financial losses to poultry producers. The USDA reported that the cost of infectious diseases for broilers, eggs, turkeys, and sales of chickens for 2021 exceeded $3.6 billion of the production value of $46.1 billion. Taxpayers and consumers ultimately pay the cost of poultry infectious diseases. For example, consumers paid higher prices for poultry products during the avian influenza outbreak of 2021-22 as evidenced by the increased price of eggs and turkey products during that period. In recent times the use of traditional prevention and treatment regiments have fallen into disfavor due to concerns such as antimicrobial resistance. This has led to an increased impetus on using other strategies such as vaccines, natural products and other prevention and control methods.  This project focuses on those infectious disease that are most important and critical to poultry producers with focus on disease detection, monitoring, prevention and control methods and strategies.

Statement of Issues and Justification

The ongoing need for this work, as indicated by stakeholder input. The United States (U.S.) is the world's largest poultry producer, the second-largest exporter of poultry meat, and a major producer of eggs and egg products. The significant integration and growth over time places the poultry industry as one of the major contributors to U.S. animal agriculture and to the national economy overall. The combined value of production from broilers, eggs, turkeys, and the value of sales from chickens in 2022 was $77.0 billion, up 67% from $46.1 billion in 2022 (USPEA, 2023). Chicken eggs and meat are an important source of dietary protein and micronutrients for the U.S. population. Ten billion broiler chickens were hatched in 2022 up 1 percent from 2021 (USDA, NASS hatchery production summary, April 2023) and are now raised for meat consumption each year on over 25,000 U.S. farms (National Chicken Council data, 2022). American per capita consumption of all poultry continues to increase annually, estimated to be 113.9 pounds in 2022, with chicken being the top animal protein consumed nationally. Additionally, U.S. egg operations produced over 96 billion eggs per year, 69.2% are domestically consumed (retail and food service shell eggs, United Egg Producers data, 2020). Per capita egg consumption has increased significantly (~15%) over the last decade from 250 to 286. The U.S. has the largest broiler chicken industry and turkey industry globally, producing over 9.78 billion chickens and 210 million turkeys annually in 2023 (USPEA, 2023). Over 59 billion pounds live weight of broiler chickens is grown annually, with 16.6% of production being exported to other countries.            

Infectious diseases continue to be a major deterrent for poultry producers. Losses induced by infectious diseases of poultry have a major economic impact affecting local, national, and global economies. Many endemic infectious diseases in the U.S. continue to decrease the profitability of commercial poultry production. Losses from mortality and morbidity, at processing (via condemnations) and overall poor flock performance (increased feed conversion, medication costs, etc.) are the compounded effects of infectious diseases. Environmental factors and novel production systems may also contribute to the increased incidence, and severity of infectious diseases. Thus, preventative strategies are critical to control infectious diseases. Since infectious diseases are multifactorial in nature, improved understanding of the underlying factors that influence disease ecology in poultry production and the interrelatedness of disease agents is necessary knowledge for effective poultry disease prevention and control.  Studying emerging and reemerging infectious disease pathogens and pathogenesis improves detection, control, and prevention of infectious poultry diseases. The renewal of the NC-1180 project will further the understanding of the epidemiology and ecology of re-emerging and emerging diseases by exploring the complexity of polymicrobial relationships that occur between pathogens, the normal microbiota, and immunity during the disease and convalescent processes.  Of primary concern are infectious disease resurgences that periodically threaten the U.S. poultry industry such as avian influenza. However, control of other significant endemic poultry infectious diseases will be strongly addressed by the participants of this project. For example, the consumer driven demand for antibiotic-free animal protein sources, has led to the emergence and reemergence of persistent enteric and immunosuppressive poultry diseases that were previously of minor importance. Determining the economic impact of poultry infectious diseases is challenging. The “cost of infectious diseases” to the poultry industry varies widely from year to year depending on the endemic, emergent, and reemerging diseases present within a given year. The USDA reported that the cost of infectious diseases for broilers, eggs, turkeys, and sales of chickens for 2021 exceeded $3.6 billions of the production value of $46.1 billions. Taxpayers and consumers ultimately pay the cost of poultry infectious diseases. Recent cases of highly pathogenic avian influenza (HPAI) provide an excellent example. Consumers paid higher prices for poultry products during the HPAI outbreak of 2021-22 as evidenced by the increased price of eggs and turkey products during that period. As part of the disease control response by the USDA, the government compensates producers for those flocks that are depopulated due to avian influenza infections which further increases the cost to the taxpayer.

            Unfortunately, HPAI emerged again in late winter of 2021 and early spring of 2022 and is still sporadically appearing in U.S. flocks and other commercial operations around the world.  By comparison, as of this writing, in the 2021-23 HPAI outbreak an estimated 58.8 million birds were lost encompassing 325 commercial flocks and 511 backyard flocks and the full economic impacts are yet to be determined but are estimated to reach 3.5 billion dollars.  On May 17^th, 2023, National Turkey Federation Vice Chairman John Zimmerman testified about the industry’s priorities before the US House Agriculture Subcommittee on Livestock, Dairy and Poultry.  He related that “avian influenza vaccination is the number one priority”. He testified “An effective vaccine would be an important tool for eradication … But we must work together to identify potential vaccines and to modernize our trade agreements. No one should be afraid of sound scientific research and open negotiation with our trading partners.”

            The export market comprises a large portion of the U.S. poultry meat and egg production. Export markets are subject to immediate restrictions when infectious diseases such as HPAI or virulent Newcastle disease (VND) are diagnosed and reported in commercial flocks. HPAI and VND have continued to emerge/reemerge. The magnitude of losses caused by HPAI were well illustrated during the 2014-2015 outbreak. According to the USDA’s final report “Approximately 7.4 million turkeys and 43 million egg-layers/pullet chickens, as well as a limited number of mixed poultry flocks, were affected by HPAI and died from the disease or were depopulated as part of the USDA’s control response.

            Although HPAI remains one of the biggest threats to the U.S. poultry industry, other endemic, emerging, and reemerging poultry diseases compromise the profitability of poultry production in the U.S. The U.S. Animal Health Association (USAHA) Committee on Poultry and Other Avian Species (CPAS) 2022 report (Sato & Neil) separately ranked the most significant infectious disease issues of poultry commodities (broilers, breeders, pullets, layers, and turkeys). We consolidated the rankings of common infectious diseases among poultry commodities into one graph.   The highest-ranked infectious diseases are located at the bottom of the graph. Overall, this project will address one or various aspects (epidemiology, diagnosis, host interactions, and disease control) of eleven (indicated as red bars) of the twenty most significant current poultry diseases.

Bars in red indicate the diseases that will be addressed in the renewal project. Please see the Attachments section for this figure.

             Based on the USAHA report variants of infectious bronchitis virus (IBV), infectious laryngotracheitis virus (ILTV), lentogenic and velogenic Newcastle disease virus (NDV), Avibacterium paragallinarum (AP) the causative agent of Infectious coryza, avian mycoplasmas (Mycoplasma gallisepticum), Ornithobacterium rhinotracheale (ORT), Escherichia coli, and Pasteurella multocida are among the more impactful respiratory agents that have the potential to damage protective airway mucosa and either themselves, or secondary opportunistic pathogens, invade body tissues. Potentially these infections may lead to systemic spread (sepsis/septicemia) and consequently to significant economic losses due to poor performance (increased feed conversion and medication costs, decreased growth), bird mortality (death), or condemnation of carcasses at processing. Environmental factors and novel production systems may also promote the incidence of these pathogens or exacerbate clinical signs and lesions. Thus, preventative strategies including poultry management are critical to control respiratory diseases of poultry. As these diseases are multifactorial in nature, improved understanding of the underlying factors driving the ‘respiratory disease complex’ are necessary, including understanding the role of immunosuppression in disease development and understanding the polymicrobial relationships that occur between pathogens and the normal microbiota during the disease process.

             Among the immunosuppressive diseases of poultry, a well-recognized immunosuppressive agent is infectious bursal disease virus (IBDV) which can severely deplete immune cells responsible for naturally detecting and controlling infections. Birds with immunosuppressive diseases are unable to respond properly when other infectious agents are encountered and are more severely affected when secondary infections occur.  Another continuous battle of the poultry industry is to maintain flocks adequately protected through vaccination against the alpha-herpesvirus Marek’s disease virus (MDV) the cause of a common lymphoproliferative disease of chickens. Although MDV vaccines protect against tumor formation, virulent strains frequently induce immunosuppression altering the susceptibility of chickens to other diseases. Another debilitating viral disease that significantly affects poultry production is caused by avian reoviruses (ARV). Increased incidence of tenosynovitis/viral arthritis in broiler chickens due to the emergence of new genotypes and serotypes of ARVs are problematic. Thus, studying immune responses under immunosuppressive conditions in association with disease pathogenesis furthers our understanding on control of the pathogen and disease. In turn, we can tease apart complicated, multifactorial infectious disease expressions. It is imperative to engage in active surveillance and continued research of infectious poultry diseases using a holistic approach while developing strategies for enhancing biosecurity, vaccination, and control for these economically important diseases. This renewal of the NC-1180 project will take a more synergist and holistic approach exploring the “disease triad” of the interplay between host, environment, and disease agents.       

 b) The importance of the work. In our NC1180 2023 meeting in August we invited poultry experts from the layer, broiler, and turkey sectors to present and discuss those poultry health / disease issues that are of highest priority. After the stakeholders’ presentations the NC1180 group summarized the stakeholders input and determined how we could have the greatest impact. Six main aims were identified: Need for broadly protective vaccines; b. better understanding of the pathology and epidemiology of reovirus infections; c. the role of infectious diseases in early chick mortality; d. detection and reduction of pathogen in the chicken house environment; e. understanding the role of current emergent immunosuppressive diseases in multifactorial infections of poultry; f. improving the mass administration of poultry vaccines.  We ascertain that the renewal project will address these aims with the expectation of having significant impact. The NC1180 participants will continue to address diagnosis, control, and prevention of emerging and endemic infectious poultry diseases. Vaccine development and use of novel vaccine technologies will also be addressed. Complacency and compliance are often the downfall of programs that are essential for the control of infectious diseases. Participants of this project will actively continue to educate poultry producers and veterinarians about best practices for disease surveillance, biosecurity, and control of infectious diseases of poultry. Additionally, the NC-1180 group is committed to hold a “stakeholder listening sessions” during their annual meetings in order for the group to maintain its relevance and continue to advance knowledge and efforts to protect our nation's food supply and the economic well-being of poultry producers considering the newer challenges posed by emerging and reemerging poultry infectious diseases and producing poultry without use of antimicrobials. 

 c) The Technical Feasibility of This Project. We realize there are many technical challenges to achieve the goals and objectives of this project. Although the “disease triad”, often used to illustrate the relationships between host, environment, and disease agent, seems straight forward and clear-cut, often it is a complex entity that requires alternative and holistic approaches. The objectives of this project are feasible and realistic because the project participants have the experience, training, and expertise in a range of disciplines, and they will be interacting collaboratively to achieve mutual success. The current NC1180 group includes a diverse group of scientists with expertise in poultry medicine, virology, microbiology, vaccinology, immunology, genomics and bioinformatics, biosecurity, and extension. The scientist within this project lead research in at least 28 independent laboratories representing nine institutions across 10 U.S. states [AL (5), CA (3), DE (6), GA (5), IL (1), IA (5), MD (4), MN (1), NE (2)]. Seven USDA scientists (SEPRL) collaborate on the project. Conducting this work as a multistate effort allows for the greatest efficiency of resource use among these scientists.

 d) Advantages for doing the work as a multistate effort (The Essential Collaborative Nature of this Project). This project is focused on infectious diseases of poultry which encompasses numerous poultry pathogens in a diversity of host systems. The complexity of disease pathogenesis and control of infectious diseases requires multiple disciplines and approaches and thus lends itself to collaborative multistate research. The multistate effort achieves synergism by participants sharing and exchanging ideas, biological materials, facilities, research equipment, expertise, and coordinated support. No single station possesses all the expertise and facilities needed to address all the poultry infectious disease and management issues included in this project. The use of a team approach enables us to perform cross-disciplinary and multiple poultry industry (i.e., broiler, layer, turkey, etc.) research with translatable value. The current NC-1180 project has served as a venue for gathering, discussing, and providing critical information on respiratory disease status of poultry at the state and national level which has incentivized research projects and collaborations among members of the group, with industry, and state and federal personnel. Major impacts from the current project include new approaches to detect respiratory pathogens using next generation sequencing for the rapid identification and prediction of pathogenicity of bacterial and viral pathogens. Host resistance aspects to IBV and ILTV have been uncovered, knowledge on the adaptation process of waterfowl avian influenza to poultry, and effects of viral immunosuppression on AIV shedding. Novel vaccine candidates for IB, ILT, and NDV are among some of the contributions of the current NC-1180 project on respiratory diseases of poultry (2019 – 2023). In the new proposed effort, we will further expand the impact towards the understanding of enteric and immunosuppressive diseases and how these intensify the persistence of respiratory diseases. Note that the NC-1180 project objectives synergize with the NE-1834, Genetic Bases for Resistance and Immunity to Avian Diseases. The participation of some of the members in both projects allows for excellent coordination between the two multi-state projects, ensuring good communication and collaboration while avoiding duplicative efforts. We recognize that there are other multistate research projects, like the NC-1202, which emphasizes food safety and the control of enteric diseases of all livestock and poultry species. The NC-1180 focus will be to explore the interactions of enteric, respiratory, immunosuppressive, and other pathogens to promote flock health and well-being.e) Outcomes and Impacts of this Project. The overall impact of a successful outcome will be improved understanding of the ecology, pathobiology, epidemiology, diagnosis, and control of major poultry infectious diseases. These outcomes will not only benefit the poultry industry but also the food security of our country. Impact of this research will be derived from identification and characterization of disease agents, their reservoirs, factors involved in agent transmission to poultry, development and delivery of novel, fast and accurate molecular and protein-based diagnostics, determination of infection status, rapid pathogen and strain identification or characterization, evaluation and development of vaccines, and the design and implementation of eradication protocols for select agents. Findings from this project will assist poultry producers to remain competitive and profitable while promoting poultry health and well-being.

Scientists of the NC-1180 Project, Endemic and Emerging Infectious Diseases of Poultry in the United States

Station	Abbreviation	Scientists Technical Committee	Num Scientists Per Unit	Collaboration Between NC1180 units
Auburn University	AL	1	5	yes
Univ. of California-Davis	CA	1	3	yes
Univ. of Delaware	DE	1	6	yes
Univ. of Georgia	GA	1	5	yes
Univ. of Illinois Urbana Champaign	IL	1	2	yes
Iowa State Univ.	IA	1	5	yes
Univ. of Maryland	MD	2	4
Univ. of Minnesota	MN	1	2	yes
Univ. of Nebraska	NE	1	2	yes
USDA SEPRL Exotic	USDA Exotic	1	4	yes
USDA SEPRL Endemic	USDA Endemic	1	3	yes
TOTAL		12	39

 

Related, Current and Previous Work

Objective 1. To investigate the ecology of infectious diseases of poultry including detection methods, pathogen characterization (e.g., genotyping, serotyping, etc.) and epidemiology. Previous projects on the ecology of respiratory diseases are strongly represented in this objective with contributions from CA monitoring the prevalence of IBV genotypes and from those in the USDA monitoring and identifying emerging strains of AIV and NDV in the US. With regards to bacterial respiratory diseases IA and CA are collaborating to understand the emergence of Avibacterium paragallinarum (AP).   In addition to these respiratory diseases, participants from MN are studying the ecology of E. coli (EC) in the chicken gut and how it relates to the barn environment. NE is studying the prevalence of Enterococcus faecalis (EF) in embryos and unhatched eggs to understand the role this bacterium may have in early chicken mortality.  Surveillance of IBDV and reovirus has been performed by MD and CA respectively, to monitor the emergence of new viral variants. SEPRL continues to study the ecology and epidemiology of emerging AIV and NDV strains and to determine the genetic changes important for transmission and pathogenicity in poultry and wild birds.

Objective 2. To develop new and improved diagnostic tools for infectious diseases of poultry. GA has developed detection and genotyping methods for IBV which have significantly improved detection and rapid genotyping of samples with multiple IBV strains. They will continue the development of, and improving, IBV genotyping assays. IA, MD, and CA have contributed to new diagnostic tools for the rapid detection of AP and the genotyping of these bacterial strains directly from clinical samples. Additionally, GA has, and will continue to, develop new and more accurate genotyping methods for ILTV. DE and SEPRL continue to update their molecular diagnostic tools for detection and identification of IBV, AIV, and NDV.

Objective 3. To elucidate host-pathogen interactions of infectious diseases of poultry.

AL is studying the susceptibility of wild aquatic birds to different strains to low pathogenic and high pathogenic AIV strains to better understand the adaptation of these viruses to domestic poultry species. AL is also studying the early immune responses elicited by novel NDV genotypes with the long-term goal of developing more effective vaccines against newly emerging NDV genotypes. CA is studying the role of passive immunity (maternal antibodies) in preventing the development of chronic microscopic lesions in kidneys and oviducts of mature layers caused by the early exposure to IBV. Their findings indicate that without adequate maternal antibodies, early IBV vaccination, or challenge with field strains, will cause microscopic lesions that could increase the incidence of disease throughout the life of the bird. GA has discovered that the ILTV chicken embryo origin (CEO) vaccines, as well as virulent ILTV strains, inhibit Type I interferon (IFN) immune responses in the upper respiratory tract. This work may lead to the discovery of the viral genes that are responsible for blocking the local Type I INF responses. IL has identified the glycoprotein C gene (gC) of MDV that is essential for viral transmission and that the gC of a virulent MDV strain can compensate for gC function of MD vaccines. It may be possible to “swap” the MDV vaccine gC for the MD gC to enhance the MDV-specific immunity without affecting vaccine replication. MD is studying the pathogenesis and immunosuppressive potential of currently circulating IBDV isolates and analyzing composition of cytoplasmic virus factories (VFs). MD is also assessing the replication and pathogenicity patterns of avian reoviruses (ARVs) and avian influenza viruses (AIVs) in primary chicken intestinal cells. NE hypothesizes that virulent strains of E. faecalis account for some of the decreased hatchability that is attributed to infertility and unhatched chicks / embryos and will determine if E. faecalis virulence is associated with the production of a bacterial exotoxin. With the aid of viral transcriptomics SEPRL in collaboration with GA has identified novel ILTV transcripts encoding viral immuno-modulators. The strategy is to generate knock-out mutants which lack homologous viral proteins and to study their pathogenesis in chickens. SEPRL will continue studies intended to find novel AIV and NDV molecular markers of infectivity, pathogenicity, and transmissibility that allow these viruses to evolve and adapt to the environment and new hosts.

Objective 4. To develop control and prevention strategies against infectious diseases of poultry.

Development of novel vaccines and vaccination strategies: AL has provided strong evidence to show that the use of IBV attenuated vaccines contribute to perpetuating the emergence of new virus variants by the selection of vaccine subpopulations and/or through naturally occurring recombination events. Therefore, new vaccination strategies with more widely protecting IBV vaccines are being developed and evaluated. DE has assessed the efficacy of existing commercial poultry disease control methods and has developed new vaccination and therapeutic strategies for viral, bacterial, and protozoal agents. GA is investigating novel, safe and affordable methods to do mucosal immunizations in the hatchery against ILTV with ILTV DNA vaccines. GA has demonstrated the feasibility of developing modified live vaccines (MLV) against H9 subtype AIVs. Such MLVs are unable to reassort with wild-type strains in vitro. GA is currently testing multiple spray vaccination machines to understand the deficiency of the process during IBV vaccination in the field.  MD has developed non-replicative adenoviral vectors (AdV) expressing avian influenza virus H5 or H7 proteins. The vectored vaccines immunogenicity and efficacy will be investigated.  SEPRL is evaluating three vaccine platforms for in ovo delivery. One platform utilizes NDV La Sota strain as a vector. This construct can be safely administered in ovo and it was shown to induce significant protection against MDV when expressing MDV gB. The second vaccine platform utilizes MDV serotype II strain as a vector to share expression of multiple antigens when co-administered with HVT.  The third vaccine platform are mRNA-based vaccines expressing immunogens of ILTV and IBV.

Outreach Programs: MD will conduct biosecurity compliance audits to prevent outbreaks of HPAI and risk-based planning to improve outbreak response. NE will continue the use and update of the “Big Red Biosecurity Program” web page to provided information on how to improve biosecurity to avoid the introduction of Avian Influenza.  CA continues its outreach programs for disease prevention of Game-fowl in Southern California.

 

Objectives

1. Objective 1. To investigate the ecology of infectious diseases of poultry including detection methods, pathogen characterization (e.g., genotyping, serotyping, etc.) and epidemiology.
   
2. Objective 2. To develop new and improved diagnostic tools for infectious diseases of poultry.
   
3. Objective 3. To elucidate host-pathogen interactions of infectious diseases of poultry.
   
4. Objective 4. To develop control and prevention strategies against infectious diseases of poultry.
   

Methods

Objective 1. To investigate the ecology of infectious diseases of poultry including detection methods, pathogen characterization (e.g., genotyping, serotyping, etc.) and epidemiology. CA, DE, GA, MD, and IA will collect tracheal / choanal swabs, cecal tonsils, bursas, tendons, tracheas, conjunctivas, hearts, and synovial fluid and perform detection and isolation methods for the following poultry pathogens: IBV, Reovirus, IBDV, AIV, NDV, ILTV, and Avibacterium paragallinarum. To investigate the ecology and epidemiology of these diseases, conventional and real time PCR will be utilized for detection from clinical samples followed by either Sanger or next generation sequencing of the pathogen’s amplified genome. To study the ecology of E. coli MN will use E. coli isolates collected from multiple sites within the bird and between birds within the same flock and between flocks. NE will determine the prevalence of E. faecalis in infertile/unhatched commercial hatchery eggs and embryos and determine whether transovarial egg transmission and / or eggshell transmission occurs. Researchers at AL, CA, DE, GA, MD, NE, MN, and IA have the depositories of samples and pathogens that will be collected during the project. Not all the laboratories have the expertise to genetically characterize these pathogens. As part of Objective 1 we will establish a network for laboratory exchange of samples to guarantee that most of the relevant pathogens isolated during the project get properly genotyped either by Sanger or third generation sequencing methods.

Objective 2. To develop new and improved diagnostic tools for infectious diseases of poultry. CA will focus on streamlining the genotyping assay of Avibacterium paragallinarum (AP) the causative agent of infectious coryza and IA will optimize a qPCR strategy for the rapid detection of AP. GA will utilize multiplex PCR and MinION sequencing to increase the discriminatory power of ILTV and IBVs genotyping assays. Bioinformatic pipelines will be used to sort, collate, and analyze next generation sequencing (NGS) data into a direct diagnostic report. GA also will do target sequences of avian reovirus Sigma C gene by MinION sequencing to elucidate the level of mixed viral populations present in clinical cases. Illumina sequencing will be utilized to generate whole genome sequences for variant reoviruses strains isolated from clinical cases.  MN will use the Mid-Central Research and Outreach Center at the University of Minnesota to collect clinical isolates of E. coli from various poultry producers across the US. The isolates will be subjected to a refined multiplex PCR schemes that identifies the proportions of high-risk bacterial clones in the sample. Whole genome sequencing will be applied to identify new and emergent clones to continue the refinement of the avian pathogenic E. coli (APEC) typing tool. NE will determine virulent from avirulent strains of E. faecalis by using an embryo lethality assay.  They will then use these strains to develop a matrix-assisted laser desorption/ionization–time of flight (MALDI-TOF) mass spectroscopy protocol to differentiate virulent from avirulent isolates of E. faecalis. SEPRL will evaluate the current real-time RT-PCR tests for the detection and differentiation of virulent and non-virulent AIV or NDV strains and update primers, probes, and assay conditions as needed.  They will also improve the non-targeted viral sequencing approach using NGS and MinION for rapid and precise characterization of pathogens from respiratory clinical samples. AL will collaborate with SEPRL and exchange reagents to establish histochemistry assays to determine the tropism and virulence of newly emerging NDV strains.  Also, SEPRL will provide NDV and AIV RNAs to AL scientists to develop and validate next generation sequencing protocols and to all participating NC1180 participating institutions to develop and validate novel diagnostic assays. GA will transfer protocols and validation data of the newly develop ILTV genotyping MinION sequencing assay to IA so they can do similar surveillance studies in layer populations. The collaboration between IA, MD, and CA to improve diagnosis and facilitate the differentiation among Avibacterium paragallinarun. GA will utilize the newly develop whole blood IL6 protein assay developed by NE to test innate responses elicited by ILTV vaccines.

Objective 3. To elucidate host-pathogen interactions of infectious diseases of poultry. To evaluate markers of resistance and susceptibility to low pathogenic AIV infections, AL will infect immortalized cell lines followed by subsequent classical virology techniques and RNA-seq. They will also develop an in vitro cell-based system to select and identify chicken immune cells by flow cytometry and evaluate immune regulation in infected cells. AL will characterize early immune gene responses elicited after vaccination with the LaSota strain of NDV in chickens with or without maternal antibodies.  Harderian glands and tracheas will be collected to determine relative gene expression by RNAseq and viral load by RT-qPCR. In parallel, the influence of maternal antibodies on the success of active vaccination will be evaluated. CA will test the efficacy of IBV maternal antibodies when administered to newly hatched chicks through feasible mass delivery routes to determine their efficacy after IBV challenge. CA will also test tissues of genetically distinct chicken lines before and after AIV and NDV infections and epigenomic assays will be conducted to understand the role of regulatory elements in host genetics to virus infections. GA will utilize long and short-read next-generation RNA sequencing to obtain transcriptomic maps of the ILTV chicken embryo origin (CEO) vaccine and virulent strains from in vitro and in vivo grown viruses. Using Illumina RNA sequencing, viral genes with relatively high expression will be compared to homologous alpha-herpesvirus genes known to interfere with type I IFN responses. Long-reads (PacBio) will facilitate the identification of transcript variations through alternative splicing and identify novel transcripts or transcript isoforms. IL will identify cellular targets for viral proteins that are required for host-to-host transmission by determining the viral transcriptome and proteome of infected tissues that are ports of viral entry and shedding. Transmission-competent and -incompetent viruses will be compared. MD will develop IBDV strains using an in-house reverse genetics system to determine the relative contribution of mutations in the hypervariable regions (HVR) of the viral genome. These recombinant viruses will allow estimates to what levels these immune escape mutants have advantage and allow infections to persist. IBDV-infected cells will be subject to live cell imaging, and fluorescence recovery after photobleaching (FRAP), and the composition of cytoplasmic virus factories (VFs) will be determined by mass-spectrometry. In addition, primary chicken intestinal organoids will be cultured, infected with ARV or AIV, and subject to RT-PCR, and confocal microscopy. NE will assess the embryo lethality of E. faecalis. The bacteria will be propagated in culture and/or embryonated eggs.  Bacterial filtrates will then be inoculated into embryos to assess embryo lethality. SEPRL will investigate host-specific factors associated with the infectivity, pathogenicity, and transmissibility in different avian species of emerging AIVs and NDVs including species, breed, age, and physiological state of the bird, and concomitant infections. SEPRL and GA will investigate ILTV-specific factors associated with immune evasion by generating knock-out mutants and studying the virus pathogenesis and host immune responses. SEPRL will investigate the efficacy of an NDV vector vaccine against MDV and ILTV as well as the feasibility of immunization against ILTV using a glycoprotein B mRNA vaccine. GA will perform host and ILTV transcriptome studies which will inform and guide projects by SEPRL and IL to identify potential ILTV and MDV genes that immunomodulate host responses and viral genes that are associated with facilitating viral transmission. NE will characterize the virulence of E. faecalis isolates and in collaboration with MN will genetically characterize isolates of E. fecalis with different degrees of virulence. CA will provide Hepatitis E virus to MD to join efforts to in vitro propagate and characterize this virus. SEPRL, after confirming low virulence, will provide wild birds origin avian paramyxovirus (APMV-1) isolates to AL scientist that will study the adaptation process of these isolates to poultry.

Objective 4. To develop control and prevention strategies against infectious diseases of poultry. AL will evaluate natural IBV recombinant viruses of the Ark-type and the ArkDPI vaccine-specific subpopulations for their pathogenicity and ability to break through immunity elicited by vaccination with a commercial ArkDPI vaccine. Pathogenicity and protection will be evaluated by clinical signs, viral load, and histopathology in naïve and vaccinated chickens. Sequencing of the spike gene will confirm the stability of the relevant changes in the viruses in vivo. CA will elucidate the antigenic maps of ARV variants’ antigenicity to determine the cross protective ability of different antigenic variants through molecular bioinformatic tools. GA will utilize adjuvanted DNA plasmids expressing ILTV antigens in calcium phosphate nanoparticles for eye drop and in ovo vaccination as a strategy to enhance mucosal protection in primary sites of ILTV infection. IBV vaccines will be designed using infectious clone technology. Chickens will be vaccinated with these clones and then challenged with various heterologous IBV types to evaluate protection levels. Also, GA will vaccinate pullets against IBV and ILTV with several different standard field spray vaccination machines and individual parameters (droplet size, spray volume, vaccine type) to determine where the deficiencies lie in field spray applications. GA will develop modified live virus (MLV) vaccines against avian influenza viruses of worldwide significance through genome rearrangement using the Flu-RAM strategy. SPF chickens will be vaccinated against AI via the drinking water with different doses of the vaccines. Viral load, antibody levels, and protection from challenge will be assessed. IL will test modified live attenuated Marek disease (MD) vaccines to antagonize viral shedding and transmission of the challenge virus. Reduction or block of herpesvirus transmission to chickens will be measured. MD will test the immunogenicity of a non-replicative adenoviral (AdV) vector expressing avian influenza virus H5 or H7 by in-house ELISA using recombinant AIV HA proteins. NE will use the whole-blood innate immunity assay as a tool to evaluate vaccines, vaccine adjuvants, vaccine programs, and other compounds that may enhance host protection. SEPRL will evaluate currently licensed and experimental vaccines against emerging AIVs and NDVs in different hosts and experimental conditions and study different aspects of immunity (innate, mucosal, antibody, and cell-mediated immunity) to predict protection conferred by vaccination and how vaccine-induced immunity affects the evolution of viruses. SEPRL will evaluate novel vaccine platforms for the in ovo delivery of NDV vectored- and mRNA-based vaccines expressing immunogens of MDV and ILTV. Also, SEPRL will construct and evaluate MDV vector vaccine candidates against poultry respiratory viral diseases. e.g., infectious laryngotracheitis, avian influenza using newly constructed MDV-BACs, HVT-BAC, and MDV 301B BAC. Recombinant MDV will be prepared using MDV-BACs to express protective antigens of respiratory disease pathogens using the BAC recombineering process. In addition, they will construct MDV-BAC with chicken cytokines to stimulate immune responses against vector vaccines. CA, GA, and SEPRL will exchange expertise in the development of avian Reovirus (ARV) vaccines and data on antigenic mapping of ARV. GA and SEPRL are collaborating in the development of live genetically attenuated ILTV vaccines as well as ILTV DNA and RNA vaccination against ILTV. SEPRL and IL are collaborating in the development of an MDV vaccine that blocks viral shedding and the next generation of MDV vectors that allow multivalent vaccination. SEPRL will develop assays for new immunological parameters to measure protection against virulent NDV that will share with all members of the NC1180 group. MD, NE, CA will join efforts to continue expanding their outreach platforms on biosecurity.

Studying the ecology of infectious diseases, developing tools, understanding the host-pathogen interactions, and developing control tools based on what we have learned about infectious disease interactions of poultry will improve our ability to control and prevent these diseases. Using a plethora of methods in this project across numerous institutions and stations will provide a robust collaborative effort to develop intervention and preventative strategies targeting important infectious diseases of poultry.

Measurement of Progress and Results

Outputs

• Improve and develop assays for the identification and characterization of poultry pathogens to determine prevalence of these agents in commercial poultry.
• Detect and monitor emerging and endemic diseases across economically important poultry dense geographic regions of the U.S. and beyond.
• Enhance the understanding of the ecology of emerging poultry pathogens (with focus on avian influenza viruses and Newcastle disease viruses) in pathogen transmission from wild bird reservoirs to poultry.
• Determine pathogen virulence factors and host specific factors (e.gs., species, breed, age, physiological state, presence of concomitants infections, immune status) which determine susceptibility or resistance to disease.
• Develop and evaluate different poultry vaccine platforms including DNA and mRNA vaccines, live attenuated vaccines, and viral vectored vaccines.

Outcomes or Projected Impacts

• New and improved assays for the identification and characterization of poultry pathogens will allow diagnostic laboratories, poultry health experts and epidemiologists earlier and more accurate detection methods to establish more effective control and prevention strategies for poultry diseases.
• The detection and monitoring of emerging and endemic poultry diseases will be paramount in protecting our poultry flocks against such reoccurring diseases as avian influenza and Newcastle disease.
• Wild birds (e.g., waterfowl) are important reservoirs for poultry diseases such as avian influenza and mycoplasmosis. Enhancing our understanding of how these pathogens persist in wild bird populations and how they are spread and transmitted will be key in prevention and control strategies to protect poultry flocks.
• Determining virulence factors of pathogens and host specific disease/resistance factors are important in developing effective disease prevention and control approaches.
• The increased impetus on decreasing the use of antibiotics and other chemotherapeutics in animal production agriculture has placed an emphasis on vaccine development. New and improved vaccines will be important for prevention and control of poultry and zoonotic diseases.

Milestones

(2024):All activities proposed in the four Objectives will be carried out simultaneously and completed in the next five years (2024-2029). Note to NCRA reviewers from Christina Hamilton, NCRA Asst Director: Please be aware that our NC Guidelines state, “For NC projects, this section (Milestones) should be eliminated and replaced with a timeline of each objective common to most competitive grants," we believe this section is acceptable as written.

Projected Participation

View Appendix E: Participation

Outreach Plan

The NC1180 renewal project proposes to expand its’ focus from respiratory diseases to infectious diseases of poultry. This expansion of the project’s focus reflects stakeholder input that was gathered during the project renewal process and the recognition that poultry diseases are multifactorial in nature.  Over the next 5-year period, the project will continue to provide information to veterinarians, researchers, animal scientists, diagnostic laboratory personnel, medical professionals, scientific extension personnel, poultry producers, industry organizations, students, and the public using oral, written and web-based formats. We anticipate that continuing education through national meetings will be an important component of our outreach activities. The NC1180 project is committed to inviting industry stakeholders to attend the annual project meetings and encourage these stakeholders to report on the most prevalent emerging and reemerging diseases affecting the various sectors of the poultry industry. The attendance of these stakeholders will facilitate information exchange which will lead to successful collaborations and will improve the impact of the NC-1180 project. The NC1180 participants have demonstrated repeated successes in these outreach activities. Additionally, the NC1180 members collaborate on a regular basis to enhance these efforts. The following summarizes our major outreach plan:

• Scientific peer-reviewed journal articles. A primary form of dissemination of research results and description of important outcomes and impacts will be through scientific peer-reviewed journal articles.


• Scientific meetings. Scientific meetings will continue to be the major venue for sharing our new findings with presentations (talks, posters) at international, national, regional, and local scientific conferences.


• Seminars/seminar series. Members of the NC1180 group actively give seminars each year to stakeholder groups reporting the results and impacts of their research.


• Graduate student training. Graduate education is a continuing commitment to train the next generation of scientists in poultry infectious diseases.


• Hybrid and virtual meetings will be used to enhance member and stakeholder participation and collaboration.

Organization/Governance

In accordance with the Guidelines for Multistate Research Activities the recommended governance for the NC1180 project includes the election of a Chair and a Secretary (also the Chair-Elect).  All officers are to be elected for at least two-year terms to provide continuity. Administrative guidance will be provided by an assigned Administrative Advisor and a CSREES Representative.  Chairs are required to organize and run the annual meeting during their term. The Secretary is responsible for the distribution of documents in advance of the annual meeting, documenting the progress and content of the meeting, communicating the results to the membership, and preparing NC1180 annual reports for submission to NIMSS. The NC1180 participants are responsible for working together to ensure the overall success of the NC-1180 project by completing their proposed research, fostering collaborations, providing timely progress reports and updates, participating in information exchanges, and attending the annual meetings.    

Literature Cited

National Chicken Council. 2022. Per-capita-consumption-of-poultry-and-livestock. https://www.nationalchickencouncil.org/about-the-industry/statistics/per-capita-consumption-of-poultry-and-livestock-1965-to-estimated-2012-in-pounds/

Animal and Plant Health Inspection Service U.S. DEPARTMENT OF AGRICULTURE. https://www.aphis.usda.gov/aphis/ourfocus/animalhealth/animal-disease-information/avian/avian-influenza/hpai-2022/2022-hpai-commercial-backyard-flocks

Livestock, Dairy, and Poultry Outlook: June 2023, LDP-M-348, U.S. Department of Agriculture, Economic Research Service, June 15, 2023

Committee of transmissible diseases of poultry & Avian Species, U.S. Animal and Health Association. Report 2022. https://www.usaha.org/transmissible-diseases-of-poultry-avian-species

US Poultry & Egg Association economic data website. https://www.uspoultry.org/economic-data/

USDA, NASS Hatchery Production Summary. April 2023. https://downloads.usda.library.cornell.edu/usda-esmis/files/9306sz28s/5999ph75k/t722jp16k/htpdan23.pdf

Attachments

Land Grant Participating States/Institutions

AL, CA, DE, IL, IN, MD, NE, OH

Non Land Grant Participating States/Institutions

Southeast Poultry Research Laboratory, USDA-ARS/Georgia