Glaesserella parasuis (G. parasuis) is a commensal bacterium in the upper respiratory tract of pigs that can also cause the swine Glässer disease, which induces an intensive inflammatory response and results in significant economic losses to the swine industry worldwide. G. parasuis can cause disease through infection of the respiratory tract, resulting in systemic infection, but the mechanism is largely unknown. Recently we showed that Glaesserella parasuis serotype 4 (GPS4) increased swine tracheal epithelial barrier permeability, resulting in easier bacterial translocation. Tight junction proteins (TJ) play a crucial role in maintaining the integrity and impermeability of the epithelial barrier. GPS4 decreased the expression of the TJ ZO-1 and occludin in swine tracheal epithelial cells (STEC). Furthermore, the proinflammatory cytokines IL-6, IL-8 and TNF-α were significantly upregulated in GPS4-infected STEC, and both the MAPK and NF-κB signaling pathways were activated and contributed to the expression of TNF-α. We demonstrate that the production of proinflammatory cytokines, especially TNF-α, during GPS4 infection was involved in barrier dysfunction. Additionally, animal challenge experiments confirmed that GPS4 infection downregulated TJ in the lungs of piglets and induced a severe inflammatory response. In general, G. parasuis infection downregulated the expression of TJ and induced massive secretion of proinflammatory cytokines, resulting in epithelial barrier disruption and favoring bacterial infection. This study allowed us to better understand the mechanism by which G. parasuis crosses the respiratory tract of pigs.

Since 2010, the Tembusu virus (TMUV) has been highly prevalent in China, causing significant economic losses to the poultry industry. In 2022, a suspected outbreak of TMUV occurred at a goose farm located in Anhui Province. A strain of TMUV, TMUV HQ-22, was isolated from the infected geese. Phylogenetic analysis using the E gene of the HQ-22 strain demonstrated its affiliation with cluster 3, a less commonly reported cluster in comparison to the main circulating cluster, cluster 2. Through a comparison of the envelope (E) protein of HQ-22 with other typical TMUV strains, a mutation at the 157th amino acid position was identified, wherein valine (V) in cluster 3 changed to alanine (A), a characteristic that is unique to cluster 2. These findings highlight the diversity and complexity of the TMUV strains circulating in China. In our experimental analysis, an injection of TMUV HQ-22 into the muscles of 3-day-old goslings resulted in severe neurological symptoms and a mortality rate of 60%. Similarly, the intracranial or intranasal infection of 3-week-old ICR mice with TMUV HQ-22 led to severe neurological symptoms and respective mortality rates of 100% or 10%. In summary, our study isolated a TMUV strain, TMUV HQ-22, from geese that belongs to cluster 3 and exhibits significant pathogenicity in both goslings and ICR mice. These results emphasize the genetic diversity of the TMUV circulating in China and expand the host range beyond mosquitoes to include ducks, chickens, geese, and even mice. It is crucial to not underestimate the risk of TMUV infection in mammals, warranting our utmost attention.

Duck Tembusu virus (DTMUV) is an avian pathogenic flavivirus that causes substantial economic losses to the poultry industry in China and Southeast Asia. The current antiviral drugs for the prevention and treatment of DTMUV infection are limited. Quercetin is a natural flavonoid widely found in fruits and vegetables and has been reported to exhibit antiviral activity against many viruses. However, whether quercetin possesses antiviral activity against DTMUV remains unknown. Therefore, the present study aimed to investigate the antiviral efficacy of quercetin against DTMUV. A Cell Counting Kit-8 (CCK-8) assay was performed to evaluate the cytotoxicity of quercetin in BHK-21 cells. The antiviral activity of quercetin was assessed using 50% tissue culture infectious dose (TCID₅₀) assay, real-time quantitative PCR (RT-qPCR), Western blotting and indirect immunofluorescence assay (IFA). Molecular docking analysis was conducted to predict potential viral targets. Moreover, the antiviral efficacy of quercetin in vivo was further evaluated in DTMUV-infected goslings. The results of the CCK-8 assay revealed that quercetin at concentrations up to 40 μM had no significant effect on BHK-21 cells. Quercetin significantly inhibited DTMUV replication in BHK-21 cells in a dose- and time-dependent manner, as demonstrated by reduced viral titres, RNA copy numbers, and E protein expression. Direct virucidal assays revealed that quercetin could directly inactivate DTMUV particles. Adsorption and post-adsorption assays revealed that quercetin inhibited DTMUV infection during the post-adsorption stage. Furthermore, molecular docking predictions indicate that the viral NS5 protein is a potential target for quercetin. In vivo, quercetin treatment significantly improved the survival rate of infected goslings; reduced viral loads in the liver, spleen, and brain; and alleviated gross pathological lesions and histopathological damage caused by DTMUV infection. In conclusion, quercetin exhibits potent anti-DTMUV activity both in vitro and in vivo. These findings provide a theoretical basis for the development of quercetin as a natural antiviral agent and highlight its potential application in the prevention and control of DTMUV infection.

Duck Tembusu virus (DTMUV) is a neurotropic virus belonging to the genus Flavivirus, and causes massive economic losses to the poultry industry in China and neighbouring countries. Autophagy plays a pivotal role in cellular responses to pathogens and viral pathogenesis. However, little is known about the roles of autophagy in DTMUV replication and virus pathogenesis, especially in neuropathogenesis. In this study, mouse neuroblastoma cells (Neuro-2a) were used to establish a DTMUV infection cell model. Our experiments indicated that DTMUV infection induced incomplete autophagy in Neuro-2a cells. Then, autophagy induced by DTMUV was altered with different autophagy regulators, and we found that incomplete autophagy promoted DTMUV replication. Furthermore, we showed that DTMUV infection activated both the ERK and AMPK pathways, resulting in decreased phosphorylation levels of the autophagy repressor mTOR, subsequently leading to autophagic induction. In addition, we utilized ICR mice as the DTMUV-infected animal model to evaluate the autophagic responses in brain tissues and investigate the effects of autophagy on viral replication and tissue lesions. As a result, we confirmed that DTMUV induced incomplete autophagy in mouse brain tissues and that autophagy inducer treatment promoted DTMUV replication and aggravated DTMUV-induced lesions, whereas autophagy inhibitor treatment led to the opposite effects. Collectively, DTMUV infection induced incomplete autophagy through the ERK/mTOR and AMPK/mTOR signalling pathways to promote viral replication in mouse neuronal cells, and DTMUV-induced incomplete autophagy contributed to the neuropathogenesis of DTMUV.