Horse immune response of recombinant Lawsonia intracellularis vaccine: Assessing the immunomodulatory impact of probiotic supplementation.

Overview

• This study investigates the immune response in horses to a recombinant vaccine targeting Lawsonia intracellularis, the cause of equine proliferative enteropathy (EPE).
• It also evaluates how supplementing horses with the probiotic Bacillus toyonensis affects the vaccine’s ability to stimulate immune responses.

Background and Significance

• EPE is a disease in horses caused by Lawsonia intracellularis, leading to thickening of the intestinal lining (hyperplasia).
• This results in symptoms like poor nutrient absorption (malabsorption), weight loss, abdominal pain (colic), and diarrhea.
• There is currently no vaccine specifically approved for horses against this disease, despite its clinical and economic impact.
• Recombinant subunit vaccines, which use specific protein fragments rather than whole organisms, are considered safer and easier to produce but often have weaker immune responses.
• Using probiotics as immunomodulators is a potential strategy to enhance vaccine efficacy.
• Bacillus toyonensis is one such probiotic that may boost immune responses when given alongside vaccines.

Study Objectives

• Develop a recombinant vaccine antigen (rLiTT) derived from a specific gene of L. intracellularis (li0902) fused with a portion of tetanus toxin to improve immune recognition.
• Test whether supplementing horses with B. toyonensis affects humoral (antibody-mediated) and cellular immune responses to this vaccine candidate.

Methods

• Production of rLiTT protein was conducted using the bacterium Escherichia coli BL21 (DE3), ensuring proper expression and confirmation through SDS-PAGE and Western blot techniques.
• Horses were vaccinated with rLiTT and split into groups with or without B. toyonensis supplementation.
• Serum samples were collected to measure total Immunoglobulin G (IgG) and its subtypes (IgGa, IgGb, IgGT) using ELISA assays.
• To assess cellular immune response, peripheral blood mononuclear cells (PBMCs) were analyzed through quantitative PCR (qPCR) to measure gene expression of several cytokines involved in immune signaling pathways.

Key Findings

• Vaccination with rLiTT induced a specific antibody response in all vaccinated horses, indicating the vaccine candidate is immunogenic.
• Probiotic supplementation with B. toyonensis significantly amplified total IgG antibody levels, especially IgGb, and influenced the profiles of IgGa and IgGT antibodies, suggesting altered antibody subclass distribution.
• Cytokine gene transcription analysis revealed:
• A strong increase (~9-fold) in il6 mRNA; IL-6 is a pro-inflammatory cytokine important for acute immune activation.
• Upregulation of il4, tnf-α (~5-fold), and il10 transcripts with probiotic treatment; these cytokines are involved in immune regulation, inflammation modulation, and anti-inflammatory effects.
• A decrease in il12 and il1 transcript levels, indicating a reduction in early pro-inflammatory signaling which may help balance immune responses to prevent excessive inflammation.

Implications and Conclusions

• The recombinant rLiTT protein shows promise as a vaccine antigen against L. intracellularis in horses, addressing a current gap in equine healthcare.
• Administering the probiotic B. toyonensis alongside vaccination enhances both antibody and cellular immune responses, highlighting its role as a beneficial adjuvant.
• This combined approach could improve vaccine effectiveness against EPE by boosting the necessary immune mechanisms without increasing harmful inflammation.
• Future equine vaccination strategies could incorporate probiotics like B. toyonensis to achieve stronger, balanced immunity with potentially better protection outcomes.