Tea Polyphenols Protects Tracheal Epithelial Tight Junctions in Lung during Actinobacillus pleuropneumoniae Infection via Suppressing TLR-4/MAPK/PKC-MLCK Signaling.

Actinobacillus pleuropneumoniae (APP) is the causative pathogen of porcine pleuropneumonia, a highly contagious respiratory disease in the pig industry. The increasingly severe antimicrobial resistance in APP urgently requires novel antibacterial alternatives for the treatment of APP infection. In this study, we investigated the effect of tea polyphenols (TP) against APP. MIC and MBC of TP showed significant inhibitory effects on bacteria growth and caused cellular damage to APP. Furthermore, TP decreased adherent activity of APP to the newborn pig tracheal epithelial cells (NPTr) and the destruction of the tight adherence junction proteins ß-catenin and occludin. Moreover, TP improved the survival rate of APP infected mice but also attenuated the release of the inflammation-related cytokines IL-6, IL-8, and TNF-α. TP inhibited activation of the TLR/MAPK/PKC-MLCK signaling for down-regulated TLR-2, TLR4, p-JNK, p-p38, p-PKC-α, and MLCK in cells triggered by APP. Collectively, our data suggest that TP represents a promising therapeutic agent in the treatment of APP infection.

Short-Chain Fatty Acids Alleviate Vancomycin-Caused Humoral Immunity Attenuation in Rabies-Vaccinated Mice by Promoting the Generation of Plasma Cells via Akt-mTOR Pathway.

Mounting evidence suggests that gut microbial composition and its metabolites, including short-chain fatty acids (SCFAs), have beneficial effects in regulating host immunogenicity to vaccines. However, it remains unknown whether and how SCFAs improve the immunogenicity of the rabies vaccine. In this study, we investigated the effect of SCFAs on the immune response to rabies vaccine in vancomycin (Vanco)-treated mice and found that oral gavage with butyrate-producing bacteria (C. butyricum) and butyrate supplementation elevated RABV-specific IgM, IgG, and virus-neutralizing antibodies (VNAs) in Vanco-treated mice. Supplementation with butyrate expanded antigen-specific CD4+ T cells and IFN-γ-secreting cells, augmented germinal center (GC) B cell recruitment, promoted plasma cells (PCs) and RABV-specific antibody-secreting cells (ASCs) generation in Vanco-treated mice. Mechanistically, butyrate enhanced mitochondrial function and activated the Akt-mTOR pathway in primary B cells isolated from Vanco-treated mice, ultimately promoting B lymphocyte-induced maturation protein-1 (Blimp-1) expression and CD138+ PCs generation. These results highlight the important role of butyrate in alleviating Vanco-caused humoral immunity attenuation in rabies-vaccinated mice and maintaining host immune homeostasis. IMPORTANCE The gut microbiome plays many crucial roles in the maintenance of immune homeostasis. Alteration of the gut microbiome and metabolites has been shown to impact vaccine efficacy. SCFAs can act as an energy source for B-cells, thereby promoting both mucosal and systemic immunity in the host by inhibiting HDACs and activation of GPR receptors. This study investigates the impact of orally administered butyrate, an SCFA, on the immunogenicity of rabies vaccines in Vanco-treated mice. The results showed that butyrate ameliorated humoral immunity by facilitating the generation of plasma cells via the Akt-mTOR in Vanco-treated mice. These findings unveil the impact of SCFAs on the immune response of the rabies vaccine and confirm the crucial role of butyrate in regulating immunogenicity to rabies vaccines in antibiotic-treated mice. This study provides a fresh insight into the relationship of microbial metabolites and rabies vaccination.

The adcA and lmb Genes Play an Important Role in Drug Resistance and Full Virulence of Streptococcus suis.

Streptococcus suis is an recognized zoonotic pathogen of swine and severely threatens human health. Zinc is the second most abundant transition metal in biological systems. Here, we investigated the contribution of zinc to the drug resistance and pathogenesis of S. suis. We knocked out the genes of AdcACB and Lmb, two Zn-binding lipoproteins. Compared to the wild-type strain, we found that the survival rate of this double-mutant strain (ΔadcAΔlmb) was reduced in Zinc-limited medium, but not in Zinc-supplemented medium. Additionally, phenotypic experiments showed that the ΔadcAΔlmb strain displayed impaired adhesion to and invasion of cells, biofilm formation, and tolerance of cell envelope-targeting antibiotics. In a murine infection model, deletion of the adcA and lmb genes in S. suis resulted in a significant decrease in strain virulence, including survival rate, tissue bacterial load, inflammatory cytokine levels, and histopathological damage. These findings show that AdcA and Lmb are important for biofilm formation, drug resistance, and virulence in S. suis. IMPORTANCE Transition metals are important micronutrients for bacterial growth. Zn is necessary for the catalytic activity and structural integrity of various metalloproteins involved in bacterial pathogenic processes. However, how these invaders adapt to host-imposed metal starvation and overcome nutritional immunity remains unknown. Thus, pathogenic bacteria must acquire Zn during infection in order to successfully survive and multiply. The host uses nutritional immunity to limit the uptake of Zn by the invading bacteria. The bacterium uses a set of high-affinity Zn uptake systems to overcome this host metal restriction. Here, we identified two Zn uptake transporters in S. suis, AdcA and Lmb, by bioinformatics analysis and found that an adcA and lmb double-mutant strain could not grow in Zn-deficient medium and was more sensitive to cell envelope-targeting antibiotics. It is worth noting that the Zn uptake system is essential for biofilm formation, drug resistance, and virulence in S. suis. The Zn uptake system is expected to be a target for the development of novel antimicrobial therapies.

Mutations in troABCD against Copper Overload in a copA Mutant of Streptococcus suis.

Streptococcus suis is a major swine pathogen that is increasingly recognized as a porcine zoonotic pathogen that threatens the health of both pigs and humans. Metal homeostasis plays a critical role during the process of bacterial infection. In this study, RNA sequencing was used to identify potential candidate genes involved in the maintenance of intracellular copper homeostasis. CopA was identified as the primary copper exporter in S. suis. The copA deletion mutant strain was found to be more sensitive to copper and accumulated more intracellular copper than the wild-type (WT) parent strain. In addition, adding manganese increased the ability of S. suis to resist copper, and the manganese transporter, TroABCD, was involved in tolerance to copper. The copA deletion mutant strain accumulated less copper when supplemented with manganese. Furthermore, when cultured with copper, the double deletion mutant (ΔcopAΔtroA) exhibited improved growth compared to the copA deletion mutant strain. In addition, the double deletion mutant (ΔcopAΔtroA) accumulated less copper than the copA deletion mutant strain. These data were consistent with a model wherein defective TroABCD resulted in decreased cellular copper accumulation and protected the strain against copper poisoning. IMPORTANCE Metal homeostasis plays a critical role during the process of bacterial infection. We identified three important potential candidate genes involved in maintenance of intracellular copper homeostasis. CopA was demonstrated to be the main copper exporter in Streptococcus suis, and manganese increased the tolerance of S. suis to copper. The double deletion mutant (ΔcopAΔtroA) improved growth ability over the copA deletion mutant strain in the presence of high concentrations of copper and accumulated less copper. These findings are consistent with a model wherein defective TroABCD resulted in decreased cellular accumulation of copper and protected the strain against copper poisoning.

Metabolomics and proteomics analyses revealed mechanistic insights on the antimicrobial activity of epigallocatechin gallate against Streptococcus suis.

Streptococcus suis (S. suis) is a highly virulent zoonotic pathogen and causes severe economic losses to the swine industry worldwide. Public health security is also threatened by the rapidly growing antimicrobial resistance in S. suis. Therefore, there is an urgent need to develop new and safe antibacterial alternatives against S. suis. The green tea polyphenol epigallocatechin gallate (EGCG) with a number of potential health benefits is known for its antibacterial effect; however, the mechanism of its bactericidal action remains unclear. In the present, EGCG at minimal inhibitory concentration (MIC) showed significant inhibitory effects on S. suis growth, hemolytic activity, and biofilm formation, and caused damage to S. suis cells in vitro. EGCG also reduced S. suis pathogenicity in Galleria mellonella larvae in vivo. Metabolomics and proteomics analyses were performed to investigate the underlying mechanism of antibacterial activity of EGCG at MIC. Many differentially expressed proteins involved in DNA replication, synthesis of cell wall, and cell membrane, and virulence were down-regulated after the treatment of S. suis with EGCG. EGCG not only significantly reduced the hemolytic activity of S. suis but also down-regulated the expression of suilysin (Sly). The top three shared KEGG pathways between metabolomics and proteomics analysis were ABC transporters, glycolysis/gluconeogenesis, and aminoacyl-tRNA biosynthesis. Taken together, these data suggest that EGCG could be a potential phytochemical compound for treating S. suis infection.

Green tea polyphenols inhibit growth, pathogenicity and metabolomics profiles of Streptococcus suis.

Streptococcus suis (SS) is an important pathogen in pigs and can also cause severe infection in humans. Currently, more and more drug resistance is reported, resulting in the search for new drugs being needed urgently. Green tea polyphenols (GTP) was reported to inhibit many bacteria. However, SS response to GTP has not been studied before. In this report, the effect of GTP on growth, cell integrity, pathogenicity and metabolic pathway of SS was examined. The GTP inhibited growth, led to cellular damage, and attenuated pathogenicity of SS. Finally, GTP affected many important metabolic pathways of SS, such as ABC transporters, pyrimidine metabolism, protein digestion and absorption. The results provide new insight into the prevention and control of SS infection.

Anti-inflammatory activity of arctigenin against PCV2 infection in a mouse model.

Arctigenin (ACT) is a novel anti-inflammatory lignan extracted from Arctium lappa L, a herb commonly used in traditional Chinese herbal medicine. In this study, we investigated the molecular mechanism whereby ACT inhibits PCV2 infection-induced proinflammatory cytokine production in vitro and in vivo. We observed that in PCV2 infection+ACT treated PK-15 cells, proinflammatory cytokine production was significantly reduced, compared to the PCV2-infected cells. The transfection and luciferase reporter assay confirmed that ACT suppressed NF-κB signalling pathway activation following PCV2 infection in PK-15 cells. Furthermore, western blotting demonstrated that ACT suppressed the NF-κB signal pathway in PCV2 infection-stimulated PK-15 cells by inhibiting the translocation of p65 from the cytoplasm to the nucleus and IκBα phosphorylation. BALB/c mice were used as a model to evaluate the anti-inflammatory effect of ACT in vivo. We found that the BALB/c mice inoculated with PCV2 infection + ACT treated showed a significant reduction of proinflammatory cytokine production in serum, lung and spleen tissue, compared to the PCV2-infected mice. Western blotting confirmed that ACT suppressed the NF-κB signal pathway in PCV2-infected mice by inhibiting the translocation of p65 from the cytoplasm to the nucleus and IκBα phosphorylation in lung tissue. Our studies first demonstrate that ACT inhibits PCV2 infection-induced proinflammatory cytokine production by suppressing the phosphorylation and nuclear translocation of NF-κB in vitro and in vivo. These results will help further develop ACT as a Traditional Chinese herbal medicine remedy in the treatment of porcine circovirus-associated diseases.

Tea polyphenols inhibit the growth and virulence of ETEC K88.

Diarrhea caused by Enterotoxigenic Escherichia coli (ETEC) causes high levels of morbidity and mortality in neonatal piglets. Owing to the abuse of antibiotics and emergence of drug resistance, antibiotics are no longer considered only beneficial, but also potentially harmful drugs. Supplements that can inhibit the growth of bacteria are expected to replace antibiotics. Tea polyphenols have numerous important biological functions, including antibacterial, antiviral, antioxidative, anti-inflammatory, and antihypertensive effects. We investigated the role of tea polyphenols in ETEC K88 infection using a mouse model. Pretreating with tea polyphenols attenuated the symptoms induced by ETEC K88. Furthermore, in a cell adherence assay, tea polyphenols inhibited ETEC K88 adherence to IPEC-J2 cells. When cells were infected with ETEC K88, mRNA and protein levels of claudin-1 were significantly decreased compared with those of control cells. However, when cells were pretreated with tea polyphenols, claudin-1 mRNA and protein levels were higher than those in cells without pretreatment upon cell infection with ETEC K88. TLR2 mRNA levels were also higher following cell infection with ETEC K88 when cells were pretreated with tea polyphenols. These data revealed that tea polyphenols could increase the barrier integrity of IPEC-J2 cells by upregulating expression of claudin-1 through activation of TLR2. Tea polyphenols had beneficial effects on epithelial barrier function. Therefore, tea polyphenols could be used as a novel strategy to control and treat pig infections caused by ETEC K88.