# Administration of a Recombinant Fusion Protein of IFN-γ and CD154 Inhibited the Infection of Chicks with Salmonella enterica

## TL;DR

A fusion protein combining chicken IFN-γ and CD154 was found to protect chickens from Salmonella infection by boosting immunity and reducing tissue damage.

## Contribution

The study introduces a novel fusion protein (chIFN-γ-chCD154) that synergistically enhances immunity and reduces Salmonella infection in chickens.

## Key findings

- The fusion protein significantly improved survival rates and reduced bacterial loads in infected chickens.
- It enhanced gut barrier function and immune responses while reducing inflammation and tissue damage.
- The fusion protein modulated key immune pathways, including TLR4/MyD88/NF-κB and IFN-γ/STAT/IRF1/GBP1.

## Abstract

Salmonella infections pose a significant risk to poultry health, affecting both animal welfare and food safety. While the immune functions of two proteins, IFN-γ and CD154, are well known in mice, their roles in chickens have not been thoroughly explored. This study focuses on a fusion protein combining chicken IFN-γ (chIFN-γ) and CD154 (chCD154) and its protective effects in chickens infected with Salmonella. Our results demonstrate that the fusion protein significantly improved survival rates, reduced bacterial loads, and lessened tissue damage compared with the use of either protein alone. Additionally, it enhanced immune responses and strengthened the gut barrier, which is crucial for protecting against infection. This research highlights the potential of chIFN-γ-chCD154 as a promising approach to boost immunity and control Salmonella infection in poultry.

The cytokines IFN-γ and CD154 have been well established, and they play pivotal roles in immune protection against Salmonella in mice, but their effects and specific mechanisms in Salmonella-infected chickens are less understood. In this study, we conducted animal experiments to screen the highly immunoprotective chIFN-γ-chCD154 fusion protein compared with single protein chIFN-γ or chCD154 in white Leghorn chickens. The results showed that compared with separate pretreatments with chIFN-γ and chCD154, the fusion protein, chIFN-γ-chCD154, synergistically increased survival of infected chickens, reduced bacterial load in feces and organs, and attenuated pathological damage to the liver and cecum. Pretreatment with chIFN-γ-chCD154 also increased humoral immune responses, expression of the tight junction proteins zo-1, occludin, and claudin-1, and the relative abundance of Enterococcus_cecorum, Lactobacillus_helveticus, and Lactobacillus_agilis, which protect against intestinal inflammation. Compared with single protein pretreatment, chIFN-γ-chCD154 significantly upregulated STAT1, IRF1, and GBP1 in infected chickens while decreasing mRNA expression of TLR4, MyD88, NF-κB, TNF-α, IL-6, and IL-1β. In summary, damage to the cecal epithelial barrier and the inflammation induced by S. typhimurium infection was alleviated by chIFN-γ-chCD154 pretreatment through a mechanism involving the TLR4/MyD88/NF-κB and IFN-γ/STAT/IRF1/GBP1 pathways.