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United States Department of Agriculture

Agricultural Research Service

Related Topics

Research Project: Functional Genomics Approaches for Controlling Diseases of Poultry

Location: Animal Biosciences and Biotechnology Laboratory

2013 Annual Report


1a.Objectives (from AD-416):
Objective 1: Develop immune reagents to detect host effector molecules controlling immune responses and determine the role of host effector molecules in disease resistance to discover biological determinants associated with disease resistance to infectious diseases of poultry. Sub-objective 1a: Develop immune reagents to detect host effector molecules controlling immune responses to NE. Sub-objective 1b: Determine the role of host effector molecular in NE disease resistance. Objective 2: Discover effective immune intervention strategies to prevent and control infectious diseases of poultry through use of nutrients as immune modulators to enhance gut health and develop strategies for their use in increasing production efficiency. Identify effector molecules of innate immunity and develop strategies for their use in reducing economic losses associated with enteric diseases of poultry. Sub-objective 2a: Use nutrients as immune modulators to enhance gut health and develop strategies for their use in increasing production efficiency. Sub-objective 2b: Identify effector molecules of innate immunity and develop strategies for their use in reducing economic losses associated with enteric diseases of poultry.


1b.Approach (from AD-416):
Develop immune reagents to detect host effector molecules controlling immune responses. Determine the role of the host effector molecules in disease resistance. Discover biological determinants associated with disease resistance to infectious diseases of poultry.


3.Progress Report:
Under Objective 1, progress was made to develop immune reagents to identify molecules in poultry that mediate immune responses in two major infectious diseases of the intestine in poultry. Several mouse-derived monoclonal antibodies that identify biological markers of chicken dendritic cells (DCs) were developed and used to characterize DCs of poultry. Aside from this report, much less is known about the expression and function of biological markers for DCs in chicken compared with its mammalian counterpart. Using the chicken monoclonal antibodies, a putative marker that was primarily expressed on mature DCs, CD83, was identified. The chicken CD83 monoclonal antibodies will be useful for future investigations of chicken immune cell maturation and mechanisms of action. Major progress was also made in the characterization of a novel chicken immune regulating factor (cytokine) interleukin-17F, which promotes inflammation and helps maintain a stable physiological balance in the gut. In general, IL-17F cytokine is expressed when cell-mediated immunity is activated by pathogens and its expression indicates the importance of IL-17F cytokine in local immune regulation. Our study showed that upon intestinal infection with the parasites Eimeria maxima and Eimeria tenella that cause the parasitic disease Coccidiosis, the level of interleukin-17F was differentially regulated.

Under Objective 2, novel antibiotic-free alternative disease control strategies against Coccidiosis and Necrotic enteritis, a parasitic and bacteria caused disease, respectively, were developed using various nutritional immune modulation strategies in collaboration with several food and nutrition companies. Our recent studies documented that the dietary immunomodulation of gut immunity in broiler chickens using natural dietary supplements and plant-derived phytochemicals that interact with innate sensing molecules to stimulate innate immunity, is a promising alternative strategy that can be applied to many infectious diseases where traditional prevention methods show limitations. In our study, we identified three plant-derived extracts that act in synergy to enhance gut immunity and protect against Coccidiosis in birds and Necrotic Enteritis in commercial broiler chickens. This finding led to the development of a commercial product for Coccidiosis. Major progress was also made in identifying specialized immune enhancing additives that increase the immune response to the Coccidiosis recombinant vaccine and provide cross protection against multiple species of the parasite Eimeria causing Coccidiosis. These additives (Montanide adjuvants: ISA 70, ISA 71, ISA 201, and ISA 206) have shown superior efficacy with a variety of human and animal vaccines. In collaboration with a private company, the effectiveness of the four additives in combination with a distinct vaccination against Coccidiosis in birds was investigated. Immunization with the profilin-based vaccine and ISA 70 or ISA 71 increased body weight gain, compared to the vaccine alone following experimental infection with the parasite Eimeria. The vaccine in combination with ISA 71 also decreased parasite survival.


4.Accomplishments
1. Developed, characterized, and commercialized critical immune reagents that detect major components of the poultry immune system. In poultry, the lack of adequate immune reagents to identify critical immune cells and the factors they release that mediate the immune response of poultry to infectious agents hinders progress in vaccine development. The ability to measure major molecules providing a protective immune response against specific infectious agents in the intestine, that include parasites of the Eimeria family and a bacterium called Clostridium, will enhance our ability to develop novel disease control strategies against them. As the poultry species Director of the NIFA-funded U.S. Veterinary Immune Reagent Network (VIRN), 35 chicken-specific immune reagents were developed and transferred to a commercial partner for commercialization and worldwide distribution. These reagents will allow scientists to assess poultry immunity to infection or vaccination and will be used to study basic or applied immunology in poultry.

2. Two major toxin proteins from Clostridium perfringens identified as potential vaccine candidates against Necrotic enteritis in chicken. In the United States, Necrotic enteritis caused by the bacterium Clostridium Perfringens is among the most important infectious diseases of the intestine in commercial broiler chickens. Globally, the economic loss due to Necrotic enteritis is estimated to cost the United States $3 billion annually largely due to medical treatments and impaired growth performance. Recently, Necrotic enteritis has re-emerged as a significant problem as a result of the restricted use of in-feed antibiotics, high-density housing conditions, and the re-use of litter. Thus, there is an urgent need to develop rational and antibiotic-free alternative disease control strategies to not only control, but to also prevent Necrotic enteritis. In order to develop an antibiotic-free strategy, an ARS scientist partnered with private industry to evaluate the vaccine potential of two major toxin proteins made by Clostridium perfringens that are involved in development of the disease in chickens. Trials using various vaccine formulations and novel delivery methods showed vaccinated chickens had significant protection against live infection challenges with Clostridium perfringens. This study provided important information on the nature of the vaccine formulation which elicited strong protective immunity against Necrotic enteritis and a patent is being filed based on the outcome of this study.

3. Discovery of a parasite-derived protein that induces protection against multiple species of the chicken parasite Eimeria. Coccidiosis in birds is a complex intestinal disease of major economic importance in chickens that is caused by multiple species of the parasite, Eimeria (Coccidia). Infection by Coccidia parasites has an enormous impact on worldwide poultry production due to the morbidity, mortality, and reduced body weight gain as a result of infection. Conventional disease control methods have relied on the use of drugs or on vaccination with live disease causing or non-disease causing forms of parasites. However, alternative methods of disease mitigation are needed due to increasing government restrictions on the use of antibiotics which kill Coccidia, the emergence of drug resistant parasites, and the high costs of new drug development. Particularly, there is an urgent need to develop a vaccine strategy against Coccidiosis in birds that is safe and effective against all relevant parasites. In collaboration with a vaccine company, a novel vaccine formulation was developed using four different combinations of immune-enhancing agents, and when given to day-old chickens, a protective immune response was elicited. Multiple in vivo trials consistently demonstrated that immunization with this special vaccine formulation induced significant protection against multiple species of Coccidia and reduced body weight loss caused by Coccidiosis. This work led to the filing of a new patent on the engineered Coccidiosis vaccine to reduce economic losses caused by Coccidiosis.

4. Characterization of a novel factor that regulates the immune response to intestinal parasites in chicken. A parasitic disease in poultry called Coccidiosis inflicts significant economic loss to the commercial poultry industry due to decreased nutrient absorption, a retarded growth rate, reduced egg production, and increased mortality. Although drugs have been traditionally used for disease control, the emergence of drug-resistant parasites and legislative bans on the use of in-feed antibiotic growth promoters and non-therapeutic antimicrobial feed additives encourage the development of alternative Coccidiosis control strategies. A scientist at ARS applied technologies and identified a new immune factor and its biological mode of action associated with the immune response in the gut of the chicken to better identify potential target sites of intervention. This new information will expand our understanding of host-parasite interactions in Coccidiosis of birds and contribute to the development of novel vaccine strategies to reduce economic losses due to gut parasites.

5. Commercialization of dietary plant extracts to enhance gut immunity and to reduce antibiotic usage in poultry production. Although the widespread use of antibiotic-based growth promoters has improved the efficiency of poultry production worldwide, there is an increasing interest in developing alternative strategies to antibiotics to control infectious diseases in livestock and poultry due to the emergence of drug-resistant disease causing organisms. Phytogenics are a group of natural growth promoters derived from herbs, spices or other plants and many medicinal foods that are highly effective in enhancing an organism’s defense against microbial infections. In collaboration with a private company, phytogenics were identified and shown to stimulate immunity and reduce damage in the intestine of poultry caused by bacteria called Clostridium perfringens and intestinal parasites. The ARS scientist went on to demonstrate that chickens fed a diet supplemented with phytogenics showed reduced intestinal lesions and improved growth or well-being following an infection with parasites. This increased protection coincided with a change in the expression of genes that regulate the immune response, supporting the well-known medicinal effects of phytogenics. These results showed that dietary supplementation of young broiler chickens with plant extracts can lead to the enhanced host resistance to parasitic and bacterial infections.


Review Publications
Lee, S.H., Lillehoj, H.S., Jang, S.I., Lee, K., Baldwin, C., Tomkpins, D., Wagener, B., Lillehoj, E., Hong, Y. 2012. Development and characterization of mouse monoclonal antibodies reactive with chicken CD83. Veterinary Immunology and Immunopathology. 145(1-2):527-533.

Lee, K.W., Hong, Y.H., Lee, S.H., Jang, S.I., Park, M.S., Bautista, D.A., Ritter, D.G., Jeong, W., Jeoung, H.Y., An, D.J., Lillehoj, E.P., Lillehoj, H.S. 2013. Effects of anticoccidial and antibiotic growth promoter programs on broiler performance and immune status. Research in Veterinary Science. 93:721-728.

Lee, K.W., Lillehoj, H.S., Park, M.S., Jang, S., Ritter, D.G., Hong, Y.H., Jeong, W., Jeoung, H.Y., An, D.J., Lillehoj, E.P. 2013. Clostridium Perfringens a-Toxin and NetB Toxin Antibodies and their possible role in protection against Necrotic Enteritis and Gangrenous Dermatitis in broiler chickens. Avian Diseases. 56:230-233.

Jeong, J., Kim, W.H., Yoo, J., Lee, C., Kim, S., Cho, J., Jang, H., Kim, D.W., Lillehoj, H.S., Min, W. 2013. Identification and comparative expression analysis of interleukin 2/15 receptor B chain in chickens infected with E. tenella. PLoS One. 7:e37704.

Kim, W.H., Jeong, J., Park, A.R., Yim, D., Kim, Y., Kim, K.D., Chang, H.H., Lillehoj, H.S., Min, W. 2013. Chicken IL-17F: Identification and comparative expression analysis in Eimeria-Infected chickens. Developmental and Comparative Immunology. 38:401-409.

Kim, D., Lillehoj, H.S., Lee, K., Neumann, A., Siragusa, G., Lillehoj, E. 2013. Genome-Wide differential gene expression profiles in broiler chickens with gangrenous dermatitis. BioMed Central (BMC) Genetics. 56:670-679.

Yim, D., Kang, S.S., Lillehoj, H.S., Min, W. 2013. A simple and efficient method for isolation of a single Eimeria oocyst from poultry litter using a micromanipulator. Research in Veterinary Science. 90:260–261.

Jang, S.I., Kim, D., Lillehoj, H.S., Lee, S.H., Lee, K.W., Bertrand, F., Dupuis, L., Deville, S., Arous, J.B., Lillehoj, E.P. 2013. Evaluation of Montanide ISA 71 VG adjuvant during profilin vaccination against experimental coccidiosis. PLoS One. 8(4):e59786.

Jang, S.I., Lillehoj, H.S., Lee, S.H., Lee, K.W., Lillehoj, E.P., Bertrand, F., Dupuis, L., Deville, S. 2013. Montanide ISA 71 VG adjuvant enhances antibody and cell-mediated immune responses to profilin subunit antigen vaccination and promotes protection against Eimeria acervulina and Eimeria tenella. Experimental Parasitology. 127:178-183.

Seal, B.S., Gay, C.G., Lillehoj, H.S., Donovan, D.M. 2013. Alternatives to antibiotics: a symposium on the challenges and solutions for animal production. Animal Health Research Reviews. 14(1):78-87.

Last Modified: 8/22/2014
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