Alternative-to-antibiotics strategies to enhance disease resistance against necrotic enteritis

Hyun. S. Lillehoj 한국가금학회 2012 한국가금학회 심포지움 Vol.2012 No.5

Development of antibiotics free alternative strategies to enhance gut immunity and to reduce harmful inflammatory responses due to necrotic enteritis. Hyun Lillehoj. Animal and Natural Resources Institute. United States Department of Agriculture. Agricultural Research Service. Beltsville, MD. USA. In the United States, necrotic enteritis(NE) is among the most important infectious diseases in poultry. Recently, NE has reemerged as a significant problem as a result of restricted use of in-feed antibiotics, high-density housing conditions, and the reuse of litter. Thus, there is an urgent need to develop rational and alternative management strategies not only to control but also to prevent NE. Better understanding of host-pathogen interaction in NE will be required to realize these goals. Our laboratory recently developed a NE disease model using dual challenge of E.maxima and C.perfringens. Using this NE model, we are currently developing various drug-free alternative strategies to enhance gut immunity and to reduce gut damage caused by NE-induced inflammatory response. This presentation will highlight some of the recent findings from my laboratory.

In ovo vaccination using Eimeria profilin and Clostridium perfringens NetB proteins in Montanide IMS adjuvant increases protective immunity against experimentally-induced necrotic enteritis

Lillehoj, Hyun Soon,Jang, Seung Ik,Panebra, Alfredo,Lillehoj, Erik Peter,Dupuis, Laurent,Arous, Juliette Ben,Lee, Seung Kyoo,Oh, Sung Taek Asian Australasian Association of Animal Productio 2017 Animal Bioscience Vol.30 No.10

Objective: The effects of vaccinating 18-day-old chicken embryos with the combination of recombinant Eimeria profilin plus Clostridium perfringens (C. perfringens) NetB proteins mixed in the Montanide IMS adjuvant on the chicken immune response to necrotic enteritis (NE) were investigated using an Eimeria maxima (E. maxima)/C. perfringens co-infection NE disease model that we previously developed. Methods: Eighteen-day-old broiler embryos were injected with $100{\mu}L$ of phosphate-buffered saline, profilin, profilin plus necrotic enteritis B-like (NetB), profilin plus NetB/Montanide adjuvant (IMS 106), and profilin plus Net-B/Montanide adjuvant (IMS 101). After post-hatch birds were challenged with our NE experimental disease model, body weights, intestinal lesions, serum antibody levels to NetB, and proinflammatory cytokine and chemokine mRNA levels in intestinal intraepithelial lymphocytes were measured. Results: Chickens in ovo vaccinated with recombinant profilin plus NetB proteins/IMS106 and recombinant profilin plus NetB proteins/IMS101 showed significantly increased body weight gains and reduced gut damages compared with the profilin-only group, respectively. Greater antibody response to NetB toxin were observed in the profilin plus NetB/IMS 106, and profilin plus NetB/IMS 101 groups compared with the other three vaccine/adjuvant groups. Finally, diminished levels of transcripts encoding for proinflammatory cytokines such as lipopolysaccharide-induced tumor necrosis $factor-{\alpha}$ factor, tumor necrosis factor superfamily 15, and interleukin-8 were observed in the intestinal lymphocytes of chickens in ovo injected with profilin plus NetB toxin in combination with IMS 106, and profilin plus NetB toxin in combination with IMS 101 compared with profilin protein alone bird. Conclusion: These results suggest that the Montanide IMS adjuvants potentiate host immunity to experimentally-induced avian NE when administered in ovo in conjunction with the profilin and NetB proteins, and may reduce disease pathology by attenuating the expression of proinflammatory cytokines and chemokines implicated in disease pathogenesis.

Postnatal development of T-lymphocyte subpopulations in the intestinal intraepithelium and lamina propria in chickens

Lillehoj, Hyun, S.,Chung, Kyeong S. 충남대학교 약학대학 의약품개발연구소 1992 藥學論文集 Vol.8 No.-

Lillehoj, H.S. and Chung, K.S., l992. Postnatal developmemt of T-lymphocyte subpopulations in the intesinal intraepithelium and lamina propria ln chickens. Vet. Immunol. Immunopathol., 31:347-360. Postnatal development of various T-lymphocyte subpopulations expressing CD3, CD8, CD4, and antigen-specific TCR heterodimers αβ(TCR2) or γδ(TCRl) was investigated in two different inbred chicken strains, SC and TK. The ratios of jejunum T-cells expressing TCRl to TCR2 in the intraepithelium of SC and TK strains gradually increased after hatching and were 3.40 and 4.28 by l2 weeks in TK and SC chickens respectively. The ratios of TCRl^+ to TCR2^+-cells in intraepithelium and the lamina propria in SC chickens were 0.96 and l.23 at 8 weeks and 4.29 and 2.15 at 12 weeks, respectively. Jejunum intraepithelial lymphocytes expressing the CD8 antigen increased gradually until 4-6 weeks of age and subsequently declined as chickens aged. CD4^+-cells represented a minor subpopulalion among the intestinal lymphocyte subpopulations. Therefore, the composition of various T-cell subpopulations in the intestine depended upon host age, the regions of the gut examined and host genetic background. These results suggest that changes in T-cell subpopulations in the intestine may reflect age-related maturation of the gut-associated lymphoid tissues.

Application of DNA Marker Technology and Functional Genomics to the Development of coccidiosis Control Strategy

Lillehoj, H.S.,Zhu, J.,Min, W.G. The Korean Society of Poultry Science 2001 韓國家禽學會誌 Vol.28 No.2

조류의 장내 면역체계와 콕시듐(Eimeria)기생충들에 대한 국소면역 반응

Lillehoj, H.S. 한국가금학회 1999 韓國家禽學會誌 Vol.26 No.2

Coccidiosis, an intestinal infection caused by intracellular protozoan parasites belonging to several different species of Eimeria seriously impairs the growth and feed utilization of livestock and poultry. Due to complex life cycle of organism and intricate host immune responses to Elmeria, coccidia vaccine development has been difficult. Understanding of basic imunobiology of pertinent host-parasite interactions is necessary for the development of novel control strategy. Although chickens infected with Eimeria spp. produce parasite-specific antibodies in both the circulation and mucosal secretions, antibody mediated responses play a minor role in protection gainst coccidiosis. Rather, increasing evidence show that cell-mediated immunity plays a major role in resistance to coccidiosis. T-lymphocytes appear to respond to coccidiosis both through cytokine production and a direct cytotoxic attack on infected cells. The exact mechanisms by which T-cells eliminate the parasites, however, remain to be investigated. Since it is crucial to understand the intestinal immune system in order to develop an immunological control strategy against any intestinal immune system in order to develop an immunological control strategy against any intestinal diseases, this presentation will summarize our current understanding of the avian intestinal immune system and mucosal immune responses to Eimeria, to provide a conceptual overview of the complex molecular and cellular events involved in intestinal immune responses to enteric pathogens.

High-throughput Gene Expression Analysis to Investigate Host-pathogen Interaction in Avian Coccidiosis

Lillehoj Hyun, S. The Korean Society of Poultry Science 2007 韓國家禽學會誌 Vol.34 No.1

Poultry products including meat and eggs constitute a major protein source in the American diet and disease-causing pathogens represent major challenges to the poultry industry. More than 95% of pathogens enter the host through the mucosal surfaces of the respiratory, digestive and reproductive tracts and over the past few decades, the two main mechanisms used to control diseases have been the use of vaccines and antibiotics. However, in the poultry industry, there are mounting concerns over the ability of current vaccines to adequately protect against emerging hyper-virulent strains of pathogens and a lack of suitable, cost effective adjuvants. Thorough investigation of the immunogenetic responses involved in host-pathogen interactions will lead to the development of new and effective strategies for improving poultry health, food safety and the economic viability of the US poultry industry. In this paper, I describe the development of immunogenomic and proteomic tools to fundamentally determine and characterize the immunological mechanisms of the avian host to economically significant mucosal pathogens such as Eimeria. Recent completion of poultry genome sequencing and the development of several tissue-specific cDNA libraries in chickens are facilitating the rapid application of functional immunogenomics in the poultry disease research. Furthermore, research involving functional genomics, immunology and bioinformatics is providing novel insights into the processes of disease and immunity to microbial pathogens at mucosal surfaces. In this presentation, a new strategy of global gene expression using avian macrophage (AMM) to characterize the multiple pathways related to the variable immune responses of the host to Eimeria is described. This functional immunogenomics approach will increase current understanding of how mucosal immunity to infectious agents operates, and how it may be enhanced to enable the rational development of new and effective strategies against coccidiosis and other mucosal pathogens.

Prospects for Immunological Intervention for Coccidiosis

Lillehoj, H.S. The Korean Society of Poultry Science 1992 韓國家禽學會誌 Vol.19 No.3

콕시듐병은 감수성이 있는 숙주의 장에 주로 감염된 Eimeria에 의하여 일어나는 것으로 가축과 가금의 성장과 사료이용에 막대한 해를 미친다. Eimeria속에 속하는 원충은 주로 세포내에 기생하며 무성생식과 유성생식을 거쳐야 하는 매우 복잡한 생활사를 가진다. Eimeria에 대한 백신개발은 주로 방어면역의 기전과 백신에 사용될 수 있는 항원을 찾는 방면으로 활발한 연구가 이루어지고 있다. 원충은 질병 본래의 모습이나 전파방법이 독특하므로 원충성 질병의 방제에 있어서는 이러한 점이 충분히 숙고되어야 할 것이다. 난해하고 복잡한 세포나 세포 분비물질간의 작용은 콕시듐병의 병성기전 뿐 아니라 방어 면역의 형성에도 매우 깊이 관여하고 있다. 따라서, Eimeria의 감염에 따른 방어기전을 충분히 이해하는 것이 가장 좋은 백신을 개발하는 첩경이 될 것이다. Coccidiosis is caused by Eimeria infecting primarily the intestine of the susceptible host, thereby seriously impairing the growth and feed utilization of livestock and poultry. The genus Eimeria contains a number of obligate intracellular protozoan parasites with a complicated life-cycle involving both asexual and sexual stages of development. The desire to develop a vaccine against Eimeria has Promoted active research to elucidate the mechanisms of protective immunity and identification of candidate vaccine antigens. Protozoa are unique in their modes of transmission and nature of disease manifestations, the significance of which should be considered in the development of a control strategy. An intricate and complex interplay of different cell populations and cytokines is involved not only in the pathogenesis of coccidiosis but also in the development of protective immunity Thus, comprehensive understanding of the events leading to protection following Eimeria infection will be crucial for the development of an effective vaccine.

Airway Mucus: Its Components and Function

Lillehoj, Erik-P.,Kim, K.-Chul The Pharmaceutical Society of Korea 2002 Archives of Pharmacal Research Vol.25 No.6

The airway surface liquid (ASL), often referred to as mucus, is a thin layer of fluid covering the luminal surface of the airway. The major function of mucus is to protect the lung through mucociliary clearance against foreign particles and chemicals entering the lung. The mucus is comprised of water, ions, and various kinds of macromolecules some of which possess the protective functions such as anti-microbial, anti-protease, and anti-oxidant activity. Mucus glycoproteins or mucins are mainly responsible for the viscoelastic property of mucus, which is crucial for the effective mucociliary clearance. There are at least eight mucin genes identified in the human airways, which will potentially generate various kinds of mucin molecules. At present, neither the exact structures of mucin proteins nor their regulation are understood although it seems likely that different types of mucins are involved in different functions and might also be associated with certain airway diseases. The fact that mucins are tightly associated with various macromolecules present in ASL seems to suggest that the defensive role of ASL is determined not only by these individual components but rather by a combination of these components. Collectively, mucins in ASL may be compared to aircraft carriers carrying various types of weapons in defense of airborne enemies.

Recent Progress in Development of Vaccines against Avian Coccidiosis

Lillehoj, Hyun S. The Korean Society of Poultry Science 1999 韓國家禽學會誌 Vol.26 No.3

Protozoa of the genus Eimeria are the etiologic agents of avian coccidiosis, the most economically important Parasitic disease for the poultry industry. Coccidia multiply in intestinal epithelial cells of a wide range of hosts, including livestock in addition to poultry. Chemotherapy is extensively used to control coccidiosis. However, development of drug resistance by Eimeria parasites, the intensive cost and labor involved in the identification of new anticoccidial compounds and public awareness of drug residues in foods warrant alternative methods to prevent coccidiocic in the fast growing poultry industry. For these reasons, there is a great interest in developing vaccines against avian coccidiosis. Live Eimeria vaccines confer protective immunity, however a significant disadvantage of using these types of vaccines is their pathogenicity. Live parasites with attenuated pathogenicity also usually produce immunity but may revert back to a pathogenic form and may be contaminated with other pathogenic organisms. Killed Eimeria vaccines are safer but, unlike live attenuated vaccines, are not able to generate cytotoxic T lymphocyte responses. Recombinant vaccines are biochemically purified proteins produced by genetic engineering that consist of particular epitopes or metabolites of Eimeria. Unlike live attenuated organisms, recombinant vaccines do not possess as much risk and generally are able to induce both humoral and cell mediated immunity. DNA vaccines consist of genes encoding immunogenic proteins of pathogens that are directly administered into the host in a manner that the gene is expressed and the resulting protein generates a protective immune response. Although all of these different types of vaccines have been applied to coccidiosis, this disease continues to cause substantial morbidity and mortality in the poultry industry. Future development of an effective vaccine against coccidiosis will depend on further investigation of protective immunity to Eimeria infection and identification of important immundgenic parasite molecules.

Recent Progress in Understanding Host Mucosal Response to Avian Coccidiosis and Development of Alternative Strategies to Mitigate the Use of Antibiotics in Poultry Production

Lillehoj, Hyun-Soon,Lee, Sung-Hyen,Jang, Seung-Ik,Kim, Duk-Kyung,Lee, Kyung-Woo The Korean Society of Poultry Science 2011 韓國家禽學會誌 Vol.38 No.4

As the world population grows and developing countries become more affluent, the global consumption of meat will increase by more than 50% within the next 10 years. Confronting the increased demand for poultry food products are emerging field diseases, increasing regulatory bans of antimicrobial growth promoters, high-density growth conditions, and waste management. Although biotechnology offers solutions to some of these challenges, basic studies are needed to better understand the complex interaction between the intestinal microbiome, host immunity and the environment. This presentation will focus on emerging strategies to enhance gut immunity and to decrease economic losses due to poultry diseases. This presentation will highlight recent developments in coccidiosis research and provide information on host immunity, immunomodulation, and the latest advances in dietary and nutritional approaches against coccidiosis. Such information will magnify our understanding of host-parasite biology, mucosal immunology, and design of future nutritional interventions and vaccination strategies for coccidiosis.