In vitro responses of chicken macrophage-like monocytes following exposure to pathogenic and non-pathogenic E. coli ghosts loaded with a rational design of conserved genetic materials of influenza and Newcastle disease viruses

• A construct contains the most conserved part of avian influenza and Newcastle disease viruses was rationally designed and developed.
• Bacterial Ghosts platform was used as delivery vehicle and immunoadjuvant for the construct.
• Delivery efficiency reaches over 90%.
• The loaded ghosts could effectively stimulate the chicken macrophage-like monocyte cells.

Avian influenza virus (AIV) and Newcastle disease virus (NDV) are two important viral diseases in the poultry industry. Therefore, new disease-fighting strategies, especially effective genetic vaccination, are in high demand. Bacterial Ghost (BG) is a promising platform for delivering genetic materials to macrophages, cells that are among the first to encounter these viruses. However, there is no investigation on the immune response of these macrophage-targeted treatments.Here, we investigated the effect of genetic materials of AIV and NDV on the gene expression profile of important pro-inflammatory cytokines, a chemokine, a transcription factor, major histocompatibility complexes, and the viability of the chicken macrophage-like monocyte cells (CMM). Our genetic construct contained the external domain of matrix protein 2 and nucleoprotein gene of AIV, and immunodominant epitopes of fusion and hemagglutinin-neuraminidase proteins of NDV (hereinafter referred to as pAIV-Vax), delivered via the pathogenic and non-pathogenic BGs (Escherichia coli O78K80 and E. coli TOP10 respectively).The results demonstrated that both types of BGs were able to efficiently deliver the construct to the CMM, although the pathogenic strain derived BG was a significantly better stimulant and delivery vehicle. Both BGs were safe regarding LPS toxicity and did not induce any cell death. Furthermore, the loaded BGs were more powerful in modulating the pro-inflammatory cytokines’ responses and antigen presentation systems in comparison to the unloaded BGs. Nitric oxide production of the BG-stimulated cells was also comparable to those challenged by the live bacteria.According to the results, the combination of pAIV-Vax construct and E. coli O78K80 BG is promising in inducing a considerable innate and adaptive immune response against AIV-NDV and perhaps the pathogenic E. coli, provided that the current combination be a potential candidate for in vivo testing regarding the development of an effective trivalent DNA vaccine against avian influenza and Newcastle disease, as well as a bacterial ghost vaccine against avian pathogenic E. coli (APEC).

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