Muscovy duck reovirus (MDRV) infection induces serious immunosuppression and intestinal injury in Muscovy ducklings with a high morbidity and mortality, and Astragalus polysaccharide (APS) pretreatment could efficiently protect ducklings from MDRV infection, although the underlying immunoregulatory mechanisms remain unclear. Thus, the objective of this study was to investigate effects of APS on the intestinal mucosal immunity in MDRV-infected Muscovy ducklings. A total of 190 1-day-old healthy Muscovy ducklings were randomly assigned to 3 groups (n = 50): normal control group, APS pretreatment for MDRV-infected group, and cohabitation infection group, then pretreated with 0.6 g/L APS or only drinking water followed by MDRV cohabitation infection with the remaining 40 artificially infected ducklings, respectively. At the 2, 3, 4, 6, 9 and 15 d after cohabitation infection, the intestinal samples were prepared to measure intestinal parameters including villus length, villus length/crypt depth (V/C) ratio, and wall thickness, together with counts of intraepithelial lymphocyte (IEL) and goblet cell (GC) by hematoxylin–eosin staining. Meanwhile, ileal secretory IgA (sIgA) and duodenal cytokine levels of IL-4, IL-6, IL-15, tumor necrosis factor-alpha, and interferon gamma were detected by the ELISA and radioimmunoassay, respectively. The results showed that APS significantly improved intestinal injuries of villi length, V/C ratio, and wall thickness of the small intestine infected with MDRV, effectively inhibited the reduction of IEL and GC caused by MDRV infection, subsequently increased sIgA and all the cytokine secretions at most time points, suggesting that APS pretreatment can effectively stimulate mucosal immune function by improving intestinal morphology and repair MDRV caused injures of small intestinal mucosal immune barrier in infected ducklings. Our findings lay the foundation for further application of APS in prevention and treatment of MDRV infection.

Mycoplasma gallisepticum (MG) can induce persistent inflammatory damage to the tracheal mucosa of poultry and cause chronic respiratory diseases in chickens. To further investigate the mechanism of MG-induced injury to the tracheal mucosa, we used chick embryo tracheal organ culture (TOC) as a model to study the invasion and reproduction of MG, the effect of MG on tracheal morphology, and the potential factors that promote MG tissue invasion. The results showed that MG infection significantly damaged the tracheal epithelial structure and weakened tracheal epithelial barrier function; MG also increased the occurrence of bacterial displacement, with a significant (p < 0.05) increase in the bacterial load of the infected TOCs at 5 and 7 days post-infection. In addition, MG significantly (p < 0.05) increased the expression levels of inflammatory cytokines, such as TNF-α, interleukin-1β (IL-1β), and IL-6, and activated the NF-κB signalling pathway, leading to increased nuclear translocation of NF-κB p65. Simultaneously, the map kinase pathway (MAPK) was activated. This activation might be associated with increased myosin light chain (MLC) phosphorylation, which could lead to actin-myosin contraction and disruption of tight junction (TJ) protein function, potentially compromising epithelial barrier integrity and further catalysing MG migration into tissues. Overall, our results contribute to a better understanding of the interaction between MG and the host, provide insight into the mechanisms of damage to the tracheal mucosa induced by MG infection, and provide new insights into the possible pathways involved in Mycoplasma gallisepticum infection in vivo.

Mycoplasma gallisepticum (MG) is a pathogenic microorganism that causes serious harm to the poultry industry. It is mainly adsorbed on the cilia and mucosa of respiratory epithelial cells, causing tracheal mucosal damage or cilia loss, resulting in chronic respiratory disease. In order to study the effect of MG infection on chicken tracheal mucosa injury and explore its possible mechanism, specific pathogen-free chickens were challenged with MG wild-type strain MG-HY. Then, transcriptome sequencing analysis was performed to study the mechanism of MG tracheal mucosal damage. During infection, MG localizes and proliferates in the chicken trachea, and induces mucosal damage. A total of 3112 significantly (P < 0.01) differentially expressed genes were selected by RNA-seq, including 1646 up-regulated genes and 1466 down-regulated genes. Functional analysis showed increased expression levels of genes involved in the immune defense response and the mechanical barrier of the tracheal mucosa in infected chicks. The expression level of pro-inflammatory cytokines (TNF-α) increased, activating the upstream protein Ras of the extracellular signal regulated protein kinase (ERK)-myosin light chain kinase (MLCK) signalling pathway, causing ERK phosphorylation levels to rise and MLCK activation. This causes phosphorylation of myosin light chain, and further regulating the expression and mucous distribution of tight junction protein, leading to tracheal mucosal injury in chicks. The results of qRT-PCR assay and immunohistochemical analysis were consistent with the results of transcriptome analysis. Overall, our findings provide a basis for further research on the underlying mechanism of chick tracheal mucosal damage caused by MG infection, and help to understand how MG induces respiratory immune damage in birds.