In silico design, optimization, and evaluation of a multi-epitope vaccine targeting the clostridium perfringens collagen adhesin protein.

Authors: DHIRAJ, CHUNDRA - University Of Maryland; SHAILES, BHATTRAI - US Department Of Agriculture (USDA); MADHUSUDAN, TIMILSINA - University Of Maryland; MOSTAFA, GHANEM - University Of Maryland; Lillehoj, Hyun; Sun, Zhifeng; Li, Charles

Submitted to: Microorganisms
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 5/10/2025
Publication Date: 5/16/2025
Citation: Dhiraj, C., Shailes, B., N N 3. Madhusudan, T., Mostafa, G., Lillehoj, H.S., Sun, Z., Li, C.Z. 2025. Immunoinformatic-guided design and refinement of multi-epitope vaccines targeting Clostridium perfringens collagen adhesin protein (CNA) through CD4+ Th cells, CD8+ CTL, and B cell epitopes. Microbial Pathogenesis. 13(5). Article e1147. https://doi.org/10.3390/ microorganisms13051147.
DOI: https://doi.org/10.3390/ microorganisms13051147

Interpretive Summary: Necrotic enteritis (NE) is a serious gut disease in poultry caused by an infection from a bacterium called Clostridium perfringens. This disease leads to major financial losses of estimated 6 billion US dollars in the global poultry industry. One reason for the increase in NE cases is the recent trend of reducing or removing antibiotics from animal feed, which had previously helped control such infections. As a result, developing effective vaccines against NE has become a top priority. The pathogenic bacterium Clostridium perfringens is a significant health threat to both humans and animals. One key factor in its ability to cause NE is a protein of 647 amino acids in length called collagen adhesion (CNA). Targeting CNA for vaccine development could be a promising way to protect poultry from this disease. In our approach, we developed a multi-pieces vaccine (MEV) designed to target CNA. This vaccine includes pieces of CNA that can stimulate the immune system, specifically for the cell-mediated system and antibody production. Using advanced computer tools, we identified and selected the best parts of CNA to include in the vaccine, ensuring it would be effective while minimizing allergic reactions. We created a vaccine containing 115 amino acids from CNA (about 1/5 of the native protein in length), representing the most important functional parts of the protein. This new vaccine shows strong potential in protecting poultry against Clostridium perfringens infections, offering similar immunity as vaccines based on the entire CNA protein.

Technical Abstract: Necrotic enteritis (NE), caused by pathogenic Clostridium perfringens, poses a significant threat to global poultry health, with estimated annual losses exceeding USD 6 billion. The rising incidence of NE has been associated with the reduced use of antibiotic growth promoters, underscoring the urgent need for alternative control measures such as vaccination. Collagen adhesin protein (CNA), a key virulence factor in NE pathogenesis, represents a promising vaccine target. The US Food and Drug Administration has begun phasing out animal testing requirements for biologics and monoclonal antibody drugs. In this study, a computational multi-epitope vaccine (MEV) targeting CNA was designed by integrating predicted Cluster of Differentiation (CD)4+ helper T lymphocyte (Th), CD8+ cytotoxic T lymphocyte (CTL), and B-cell epitopes. Bioinformatics tools were used to identify immunogenic, antigenic, and non-allergenic epitopes assembled into a 115-amino-acid peptide vaccine construct. The candidate demonstrated strong stability and solubility. In silico immune simulation predicted robust immune responses, including elevated IgG and IgM antibody levels, plasma cell proliferation, Th memory formation, and CTL activation, comparable to responses elicited by a full-length CNA. These findings support the potential of the designed peptide as one of the multiple effective NE vaccine components, offering a promising alternative to antibiotic-based approaches in poultry disease management.