Induction of innate immune response following infectious bronchitis corona virus infection in the respiratory tract of chickens

Abstract

Infectious bronchitis virus (IBV) replicates in the epithelial cells of trachea and lungs of chicken, however the mechanism of generation of innate immune response against IBV infection in these tissues has not been fully characterized. Our objective was to study innate responses induced early following IBV infection in chickens. Initiation of the transcription of selected innate immune genes such as TLR3, TLR7, MyD88, IL-1β and IFN-β, as well as recruitment of macrophages, were evident following an initial down regulation of some of the observed genes (TLR3, IL-1β, and IFN-γ) in trachea and lung. This initial down-regulation followed by the induction of innate immune response to IBV infection appears to be inadequate for the control of IBV genome accumulation and consequent histopathological changes in these tissues. Potential induction of innate immunity before infection occurs may be necessary to reduce the consequences since vaccine induced immunity is slow to develop.

Keywords: Avian; Cytokine; Infectious bronchitis virus; Macrophage; Toll-like receptor.

Fig. 1

Relative TLR3 and TLR7 mRNA expression in trachea and lungs of chickens infected with the Conn A5968 strain of IBV. (A) and (B) represent relative TLR3 and TLR7 mRNA expression in trachea, respectively. (C) and (D) represent TLR3 and TLR7 mRNA expression in the lung, respectively. Chickens were infected intra-tracheally with the Conn A5968 strain of IBV at 6 days of age and trachea and lung tissues were collected at 12, 24, 48, and 72 h post-infection (hpi). There were five IBV-infected chickens at each time point (six animals were sampled at 12 hpi) and five PBS-treated chickens used as controls for each time point (only two animals were sampled at 72 hpi for the control group). Target mRNA expression was normalized via the geometrical mean of Eff.(uninfected control)^Cp for all control genes (β-actin and ubiquitin). Error bars represent standard error of the mean (SEM). ⁎=relative mRNA expression is significantly up-regulated when compared to the uninfected controls and ⁎⁎=relative mRNA expression is significantly down-regulated when compared to controls.

Fig. 2

Relative MyD88 mRNA expression in trachea and lungs of chickens infected with the Conn A5968 strain of IBV. (A) and (B) represent relative MyD88 mRNA expression in trachea and lung, respectively. Experimental design was as indicated in the legend of Fig. 1. Target mRNA expression was normalized via the geometrical mean of Eff.(uninfected control)^Cp for all control genes (β-actin and ubiquitin). Error bars represent SEM. ⁎=relative mRNA expression is significantly up-regulated when compared to the uninfected controls.

Fig. 3

Relative IL-1β, IFN-β and -γ mRNA expression in trachea and lungs of chickens infected with the Conn A5968 strain of IBV. (A–C) represents relative IL-1β, IFN-β, and -γ mRNA expression in trachea, respectively. (D–F) represents relative IL-1β, IFN-β, and -γ mRNA expression in the lung, respectively. Experimental design was as indicated in the legend of Fig. 1. Target mRNA expression was normalized via the geometrical mean of Eff.(uninfected control)^Cp for all control genes (β-actin and ubiquitin). Error bars represent SEM. ⁎=relative mRNA expression is significantly up-regulated when compared to the uninfected controls, and ⁎⁎=relative mRNA expression is significantly down-regulated when compared to uninfected controls.

Fig. 4

Quantification of macrophages present in trachea and lung. Chickens were infected intra-tracheally with Conn A5968 at 6 days of age and trachea and lung tissues were collected at 12, 24, and 48 hpi. There were three IBV-infected and three PBS-treated chickens used at each time point for the lung macrophage quantification, and in a separate experiment with the same experimental design, four IBV-infected and five PBS-treated chickens used at each time point for quantification of macrophages in trachea. (A) and (B) represent the percent of macrophages in control and IBV infected trachea respectively at 24 hpi. (D) and (E) represent the percent of macrophages in IBV infected and control lung respectively for 24 hpi. (C) and (F) are graphical representations of the percentage of macrophage numbers at each time point for trachea and lungs, respectively. Error bars represent SEM. ⁎=significant increase when compared to uninfected controls at 12, 24, and 48 hpi in the lungs and ⁎⁎=significant increase when compared to all other time points in the trachea.