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<p>The application of sortase-mediated surface display technology on <i>Bacillus subtilis</i> represents a novel approach in protein engineering. We developed and evaluated a recombinant chimeric protein (R-anz) integrating truncated forms of three <i>Clostridium perfringens</i> toxins (CPA, NetB, ZMP) displayed on <i>B. subtilis</i>. Bioinformatics analyses using docking demonstrated that the 3D structure of R-anz chimeric protein has the potential to interact with chicken Toll-like receptor 21 (TLR21). Successful expression of the chimeric antigen was confirmed through SDS-PAGE and Western blotting, revealing a 78&#xa0;kDa band in lysozyme-treated, xylose-induced samples. Oral immunization of chickens with this live bacterial vaccine significantly elevated IgY antibody levels, as assessed by ELISA. Cytokine profiling demonstrated a robust immune response, with marked upregulation of IFN-γ, IL-4, IL-17, and IL-22. Post-challenge with virulent <i>C. perfringens</i> strains, immunized chickens exhibited significantly reduced intestinal lesion scores, indicating partial protection. This study highlights the dual activation of humoral and cellular immune pathways, evidenced by elevated IgY levels and enhanced cytokine responses, particularly IFN-γ, IL-4, and IL-22. While the results demonstrate the potential of the R-anz chimeric protein to mitigate necrotic enteritis (NE), further research is essential to refine its efficacy, explore commercial feasibility, and address industry-specific challenges. This work paves the way for advancing NE vaccines using innovative sortase-mediated surface display technology, offering a promising strategy for sustainable poultry health management.</p>

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Sortase-mediated surface display of a chimeric immunogen on Bacillus subtilis: a chicken necrotic enteritis vaccine candidate

  • 期刊论文

  • 出版模式：

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  • 发表日期：

    2025年7月1日

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  • Bassima Al-Aneed,
  • Hoang Duc Nguyen,
  • Ali Akbar Masoudi,
  • Saghi Nooraei,
  • Abbas Hajizade,
  • Hossein Tarrahimofrad,
  • Gholamreza Ahmadian,
  • Jamshid Razmyar,
  • Moeen Khatami

摘要

The application of sortase-mediated surface display technology on Bacillus subtilis represents a novel approach in protein engineering. We developed and evaluated a recombinant chimeric protein (R-anz) integrating truncated forms of three Clostridium perfringens toxins (CPA, NetB, ZMP) displayed on B. subtilis. Bioinformatics analyses using docking demonstrated that the 3D structure of R-anz chimeric protein has the potential to interact with chicken Toll-like receptor 21 (TLR21). Successful expression of the chimeric antigen was confirmed through SDS-PAGE and Western blotting, revealing a 78 kDa band in lysozyme-treated, xylose-induced samples. Oral immunization of chickens with this live bacterial vaccine significantly elevated IgY antibody levels, as assessed by ELISA. Cytokine profiling demonstrated a robust immune response, with marked upregulation of IFN-γ, IL-4, IL-17, and IL-22. Post-challenge with virulent C. perfringens strains, immunized chickens exhibited significantly reduced intestinal lesion scores, indicating partial protection. This study highlights the dual activation of humoral and cellular immune pathways, evidenced by elevated IgY levels and enhanced cytokine responses, particularly IFN-γ, IL-4, and IL-22. While the results demonstrate the potential of the R-anz chimeric protein to mitigate necrotic enteritis (NE), further research is essential to refine its efficacy, explore commercial feasibility, and address industry-specific challenges. This work paves the way for advancing NE vaccines using innovative sortase-mediated surface display technology, offering a promising strategy for sustainable poultry health management.