Location: Poultry Research
2025 Annual Report
Objectives
1. Use proteomics, genomics, and systems biology approaches to identify molecular determinants of pathogenesis, strain variation, and tissue tropism of different E. coli strains.
2. Identify immunological targets that will confer cross-protection against prevalent E. coli strains in poultry production and develop vaccine platforms that are effective in very young birds, provide cross- protection, and can be easily administered.
2.a. Identify genetic determinants for antigenicity and pathogenicity of E. coli through comparative genomics and analyses.
2.b. Identification of immunological targets will provide a cross protection against different strains of Avian Pathogenic Escherichia coli (APEC).
2.c. Assess in ovo vaccination technology for delivery of live attenuated APEC vaccines.
3. Develop systems-level capabilities to evaluate the effects of commercial-scale, poultry management practices on animal health and production; microbial ecology, development of antimicrobial resistance and bacterial pathogen transmission to develop mitigation strategies.
3.a.1. Evaluate performance of three bio-aerosol samplers for collecting airborne E. coli attached to dust particles from poultry production environments.
3.a.2. Quantify concentration and size distribution of airborne E. coli in representative US broiler and layer houses.
3.a.3. Evaluate electrostatic particle ionization (EPI) and ultraviolet (UV) radiation to reduce airborne E. coli.
3.b.1. Evaluate effects of litter amendment application rate on E. coli populations in broiler litter.
3.b.2. Assess E. coli populations and antibiotic growth promoter (AGP) residue concentrations in biochar-treated and untreated litter (live study).
3.b.3. Evaluate effects of litter management [top-dressed (TD) vs non-top-dressed] and bedding type (pine vs switchgrass) on litter E. coli populations over successive flocks.
Approach
Proteomic, genomic, and systems biology approaches will be applied to identify molecular determinants of pathogenesis, strain variation, and tissue tropism of different E. coli strains. The E. coli strains analyzed will be isolated from varying diseased poultry flocks and strain genomic & proteomic characteristics and isolate epidemiological factors will be applied model development for greater understanding of pathogenic E. coli and associated disease. To further protect against pathogenic E. coli, immunological targets will be identified that will confer cross-protection against prevalent E. coli strains in poultry production. The genomic and plasmid sequences of various E. coli strains will be aligned, and antigenic factors will be determined. Immunological targets will be identified and assessed via challenge models that provide cross-protection against pathogenic E. coli. In addition, vaccination platforms that are effective in very young birds, provide cross-protection, and can be easily administered will be developed. In ovo technologies will be assessed for delivery of protective vaccines and associated protocols developed. To increase the understanding of environmental E. coli and evaluate risks to poultry and associated antimicrobial resistance, studies will evaluate airborne E. coli associated with dust particles and E. coli linked to poultry litter. Further, mitigation means will be assessed for their impact on environmental E. coli populations.
Progress Report
Per sub-objective 2A, comparative genomic analysis along with in vitro testing revealed pagP to play a strong role in APEC infections with potential to serve as a subunit vaccine. A pagP knockout APEC strain was created and used in three independent APEC infection trials to assess its role in embryonic, subcutaneous, and intratracheal infections. Results found pagP to play a role in systemic, but not local, infections with the potential to be effective as a subcutaneously or intramuscularly injected vaccine. In line with Sub-Objective 2B, a novel Oxford Nanopore Technologies (ONT) sequencing-based method for E. coli O-serogroup typing was developed and shown to have high accuracy. Additionally, work continued on optimizing intratracheal and subcutaneous infection models for broiler and layer chickens during multiple developmental stages. Per Sub-Objective 3B1, next generation sequencing was performed to identify the effect of litter treatments on E. coli populations as well as the overall bacterial and fungal populations. Concurrent sequencing of chicken microbiomes is being used to determine a link between litter treatments and poultry intestinal health.
Accomplishments
1. Effective disease models for Avian Pathogenic E. coli. Effective Avian Pathogenic E. coli (APEC) intratracheal disease models representative of industry were developed for layer and broiler chickens. In layer chickens, models were developed for both young (7 day-old) chicks and pre-lay (18 week-old) hens, while the broiler model focused on early life infections (7 day-old chicks). Accurate and reproducible disease models are the cornerstone of effective disease management as they provide reliable information necessary for the development of effective means of control. These models are critical to continued work focused on characterizing host immune response to APEC infection and testing of novel vaccine candidates.
2. Avian pathogenic E. coli electron beam killed vaccine. A novel Electron Beam irradiated Avian Pathogenic E. coli (APEC) vaccine was tested in two individual disease trials to determine its ability to protect against APEC infection in layer chickens. APEC is responsible for over $40 million in economic losses to the U.S. broiler industry each year due to carcass condemnation alone. Current APEC vaccines are incapable of providing broad spectrum control due to the wide genetic variation among field strains. Electron beam technology has the potential to be used to create a broad spectrum vaccine due to its ability to inactivate pathogens without disrupting the cell membrane and the ease at which multivalent vaccines can be created with the technology. These studies provide important initial information on the applicability of electron beam killed vaccines for APEC infections, potentially laying the groundwork for the development of an urgently needed solution for the poultry industry.
3. Comprehensive assessment of pagP as a subunit vaccine candidate for Avian Pathogenic E. coli. Subunit vaccines are a promising vaccine strategy for live animal production as they can be designed using the most immunogenic targets, are stable, and are less likely to produce negative side effects. Our team has begun work to develop a subunit vaccine for Avian Pathogenic E. coli (APEC) to provide an urgently needed therapeutic to the poultry industry. We identified the gene pagP as a promising vaccine in cell line experiments due to its strong upregulation during infection. To determine the safety of pagP for subunit vaccine development we sought to characterize its role in APEC infections. Studies using an APEC strain containing a pagP deletion found that it is most likely important during systemic infections but not local respiratory infections indicating a potential for a pagP subunit vaccine to be effective when applied via intramuscular or subcutaneous injection.
4. Novel sequencing-based method for E. coli O-Serogroup typing. The wide genetic variation among Avian Pathogenic E. coli (APEC) strains has made rapid strain identification an integral part of identifying and controlling emerging APEC infections. Current methods of strain identification are time and labor intensive often requiring samples to be collected and shipped to a state lab for testing. Here, a reliable sequencing-based method for APEC O-serogroup typing was developed using Oxford Nanopore Technologies (ONT). Testing revealed the method was able to successfully identify the correct O-serogroup in 24 of 25 E. coli isolates tested. This ONT-based approach offers a promising tool for early detection of pathogenic serotypes in poultry, with potential to support rapid disease intervention and minimize economic losses.
Review Publications
Leigh, S.A., Evans, J.D. 2024. Genome sequence of Mycoplasma gallisepticum strain A5969. Microbiology Resource Announcements. 13:e00813-24. https://doi.org/10.1128/mra.00813-24.
Collins Elliott, K.E., Lindsey, L.L., Evans, J.D., Leigh, S.A., Robinson, K., Fatemi, S.A., Mousstaaid, A., Gerard, P.D., Purswell, J.L., Peebles, E.D. 2024. Effects of the in ovo injection of an Escherichia coli vaccine on the hatchability and subsequent early post hatch characteristics of commercial layer chicks. Poultry Science. 104:104562. https://doi.org/10.1016/j.psj.2024.104562.
Linhoss, J., Mohammadi-Aragh, M., Evans, J.D. 2023. Influence of pine and miscanthus biochar on the water activity and moisture sorption isotherms of used broiler litter. Heliyon. 9:e22618. https://doi.org/10.1016/j.heliyon.2023.e22618.
Joseph, J., Zhang, L., Adhikari, P., Evans, J.D., Ramachandran, R. 2023. Avian pathogenic Escherichia coli (APEC) in broiler breeders: An overview. Heliyon. 12:1280. https://doi.org/10.3390/pathogens12111280.
Jia, L., Arick Ii, M.A., Hsu, C., Peterson, D.G., Evans, J.D., Robinson, K., Sukumaran, A., Ramachandran, R., Adhikari, P., Zhang, L. 2024. High-throughput Oxford Nanopore sequencing-based approach for the multilocus sequence typing analysis of large-scale avian Escherichia coli study in Mississippi. Poultry Science. 103:104067. https://doi.org/10.1016/j.psj.2024.104067.