| Avian infectious bronchitis (IB) is an acute highly contagious respiratory disease caused by avian infectious bronchitis virus (IBV) and results in serious economic losses in poultry industry. The serotypes of the virus is complicated and the cross protection between them are poor. The strategy of IBV prevention by traditional vaccine faces challenging. Therefore, further studies on pathogenesis and immunologic mechanism of IBV to control IB are significant.Innate immunity is the first defensive line to protect from the infection of virus. It is an effective antiviral mechanism and plays an important role in antiviral immunity. Host receives signal of virus invasion and produces series of defensive reaction to virus infection through pattern recognition receptor (PRR) in innate immunity. Secretory immunoglobulin A (SIgA) is an important molecule in mucosal immune. Presently, there are limited studies on the influence of IBV infection on SIgA, TLR3 and transcription molecules nuclear factor-kappa B (NF-kB and interferon regulatory factor 7 (IRF-7) in the downstream of TLR3 signal pathway.In order to explore the distribution rule of IBV M41 and the influence of IBV infection on TLR signal pathway and SIgA, reveal the pathogenesis and defense mechanism of host antiviral innate immune response for IBV, the TLR3, NF-kB, IRF-7, SIgA and virus in trachea, kidney and gland of harder at 1,3,5,8,11,14,21 and 28 days post infection (DPI) by IBV M41 were detected by immunohistochemistry technique (IHC) in the present study. The histopathological changes were observed in the three tissues in the same time. The results are listed as follows.1. Trachea, kidney and gland of harder of SPF chicken showed varying degrees pathological change after infection by IBV M41. From 1DPI to 8DPI, trachea showed degeneration and necrosis in mucosal epithelial cells and lymphocyte infiltration in submucosa. The virus loads were kept at high level during the course, and reached the peak at 5DPI. From 11 DPI to 14DPI, the mucosal epithelial cells were repaired and regenerated, and lymphocyte infiltration was alleviated, at the same time the virus loads decreased sharply. From 21DPI to 28 DPI, viruses were cleared, and the organization structure of trachea was recovered. Kidney showed obvious pathological change from 5DPI to 11DPI. Kidney tubules occurred degeneration and necrosis and slight lymphocyte infiltration in interstitial substance. The virus loads were high during this period but lower than those in trachea, and reached the peak at 8DPI. The virus loads decreased after 14DPI. The pathological changes of kidney were relieved and the structure was recovered. From 1DPI to 3DPI, there were no virus and no obvious pathological lesion in gland of harder. From 5DPI to 14DPI, the histocytes showed degeneration and necrosis, and a few viruses were detected. The viruses were disappeared and structure of tissue was become normal after 21 DPI. This study showed the histopathological changes lesion of trachea was more serious than those of kidney and gland of harder, and the virus loads of trachea were also higher than those in other two tissues. Therefore, IBV M41 strain had different tropism to different tissues and organs and causing varying degrees of histopathological changes, the degrees of histopathological changes were related to virus loads.2. The IHC results showed that TLR3, NF-κB and IRF-7 are mainly located in cytoplasm in trachea mucosal and tubules epithelial cells and a small number of nucleuses also have NF-κB and IRF-7 expression. The expression levels of TLR3, NF-κB and IRF-7 in trachea were increased markedly from 3DPI to 8DPI, and reached the peak at 5DPI,5DPI and 8DPI (p<0.01). The number of IRF-7 in nucleus was increased. From 11DPI to 14DPI, the expression levels of TLR3 were still high (P＜0.05), but expression levels of NF-κB and IRF-7 were became normal. After 21DPI, expression level of TLR3 became normal too. Expression levels of TLR3 and NF-κB in kidney of treatment group were increased markedly from 5DPI to 11DPI, reached the peak at 8DPI and 5DPI (P<0.01). Expression levels of IRF-7 were increased slightly from 3DPI to 5DPI, and the expression in nucleus was increased. After 14DPI, expression levels of TLR3 and NF-kB in kidney were became normal, and expression level of IRF-7 became normal at 8DPI, but the expression in nucleus kept at high level until 21DPI.3. In the early stage of IBV M41 infection (1DPI to 11 DPI), there was no SIgA positive cell in gland of harder from both treatment group and control group. From 14DPI to 28DPI, there were some SIgA positive cells in gland of harder from both two groups. The SIgA positive cells in treatment group were bigger, full of brow granules and the number increased locally, but the SIgA positive cells were small, triangle and oval in control group. From 21DPI to 28DPI, the cell shape and number of SIgA positive cells from the treatment group were similar with those from control group. No SIgA positive cell could be detected in the trachea from both groups throughout the experiment.Based on the above results, in the early stage of IBV M41 infection, the trachea of SPF chicken showed obvious pathological change. The virus loads of trachea began to rise from 1DPI and kept at high level between 3DPI to 8DPI, and reached the peak at 5DPI. During this period, the expression levels of TLR3, NF-kB and IRF-7 of trachea in treatment group were rising at various degrees, and TLR3 was up-regulated more obviously and the lasted time was the longest. Compared to control group, the expression levels of TLR3 were significant difierence from 3DPI to 14DPI. The expression levels of NF-kB and IRF-7 were up-regulated from 3DPI to 8DPI and 5DPI to 8DPI, respectively. The expression levels of NF-kB and IRF-7 in nucleus were increased and the expression levels IRF-7 were especially high. In the end of infection, with the recovering of histological structure and the decrease and disappear of virus loads, the expression levels of TLR3, NF-kB and IRF-7 in treatment group recovered to normal level. The pathological changes of kidney were mild and occurred later, which was from 5DPI to 14DPI. Meanwhile, the changeable trends of virus load, TLR3, NF-kB and IRF-7 expression levels in kidney were consistent with those in trachea, but the degrees and lasting times were slighter and shorter than those in trachea. The peaks of virus load and expression levels of TLR3, NF-kB and IRF-7 occurred during 5DPI to 8DPI. The SIgA positive cells in gland of harder were bigger and the number increased locally in the middle stage of IBV M41 infection. Therefore, this study indicated that the infection of IBV M41 could cause the change of expression levels of TLR3, NF-kB and IRF-7, and their changeable trends were consistent with that of virus load. IBV infection caused the change of shape and quantity of SIgA positive cells,In summary, the present study demonstrated that in the early stage of IBV M41 infection, the expression levels of innate immunity related molecules in SPF chickens could be activated and had a significant effect on the innate immune, indicating an important role in the defense mechanism of host antiviral innate immune response for IBV. |
