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Biofilms often engender persistent infections, heightened antibiotic resistance, and the recurrence of infections. Therefor, infections related to bacterial biofilms are often chronic and pose challenges in terms of treatment. The main transcription regulatory factor, CsgD, activates csgABC-encoded curli to participate in the composition of extracellular matrix, which is an important skeleton for biofilm development in enterobacteriaceae. In our previous study, a wide range of natural bioactive compounds that exhibit strong affinity to CsgD were screened and identified via molecular docking. Tannic acid (TA) was subsequently chosen, based on its potent biofilm inhibition effect as observed in crystal violet staining. Therefore, the aim of this study was to investigate the specific effects of TA on the biofilm formation of clinically isolated Escherichia coli (E. coli). Results demonstrated a significant inhibition of E. coli Ec032 biofilm formation by TA, while not substantially affecting the biofilm of the ΔcsgD strain. Moreover, deletion of the csgD gene led to a reduction in Ec032 biofilm formation, alongside diminished bacterial motility and curli synthesis inhibition. Transcriptomic analysis and RT-qPCR revealed that TA repressed genes associated with the csg operon and other biofilm-related genes. In conclusion, our results suggest that CsgD is one of the key targets for TA to inhibit E. coli biofilm formation. This work preliminarily elucidates the molecular mechanisms of TA inhibiting E. coli biofilm formation, which could provide a lead structure for the development of future antibiofilm drugs.

The aim of this study was to investigate the anti-Eimeria tenella mechanism of Qingchang Compound (QCC) and provide a basis for its clinical application. The active ingredients, active ingredient-disease intersection targets, and possible pathways of QCC for the treatment of chicken coccidiosis were analyzed, the binding ability of pharmacodynamic components and target proteins was determined by network pharmacology and the molecular docking, and a model of infection with coccidiosis was constructed to verify and analyze the mechanism of action of QCC against coccidiosis. Among the 57 components that met the screening conditions, the main bioactive components were quercetin, dichroine, and artemisinin, with IL-1β, IL-6, IL-10, IFN-γ, and IL-8 as the core targets. Simultaneously, the KEGG signaling pathway of QCC anti-coccidiosis in chickens was enriched, including cytokine-cytokine receptor interactions. The results showed that the main pharmacodynamic components of QCC and the core targets could bind well; artemisinin and alpine possessed the largest negative binding energies and presented the most stable binding states. In addition, in vivo studies showed that QCC reduced blood stool in chickens with coccidiosis, restored cecal injury, and significantly reduced the mRNA and protein expression levels of IL-1β, IL-10, and IFN-γ in ceca (p

Biofilm is a structured community of bacteria encased within a self-produced extracellular matrix. When bacteria form biofilms, they undergo a phenotypic shift that enhances their resistance to antimicrobial agents. Consequently, inducing the transition of biofilm bacteria to the planktonic state may offer a viable approach for addressing infections associated with biofilms. Our previous study has shown that the mouse antimicrobial peptide CRAMP-34 can disperse Pseudomonas aeruginosa (P. aeruginosa) biofilm, and the potential mechanism of CRAMP-34 eradicate P. aeruginosa biofilms was also investigated by combined omics. However, changes in bacterial extracellular metabolism have not been identified. To further explore the mechanism by which CRAMP-34 disperses biofilm, this study analyzed its effects on the extracellular metabolites of biofilm cells via metabolomics. The results demonstrated that a total of 258 significantly different metabolites were detected in the untargeted metabolomics, of which 73 were downregulated and 185 were upregulated. Pathway enrichment analysis of differential metabolites revealed that metabolic pathways are mainly related to the biosynthesis and metabolism of amino acids, and it also suggested that CRAMP-34 may alter the sensitivity of biofilm bacteria to antibiotics. Subsequently, it was confirmed that the combination of CRAMP-34 with vancomycin and colistin had a synergistic effect on dispersed cells. These results, along with our previous findings, suggest that CRAMP-34 may promote the transition of PAO1 bacteria from the biofilm state to the planktonic state by upregulating the extracellular glutamate and succinate metabolism and eventually leading to the dispersal of biofilm. In addition, increased extracellular metabolites of myoinositol, palmitic acid and oleic acid may enhance the susceptibility of the dispersed bacteria to the antibiotics colistin and vancomycin. CRAMP-34 also delayed the development of bacterial resistance to colistin and ciprofloxacin. These results suggest the promising development of CRAMP-34 in combination with antibiotics as a potential candidate to provide a novel therapeutic approach for the prevention and treatment of biofilm-associated infections.

Probiotic biofilms have been beneficial in the fight against infections, restoring the equilibrium of the host’s gut microbiota, and enhancing host health. They are considered a novel strategy for probiotic gut colonization. In this case, we evaluated the effects of various active substances from traditional Chinese medicine on Escherichia coli Nissle 1917 (EcN) to determine if they promote biofilm formation. It was shown that 8–64 μg/mL of oleanolic acid increased the development of EcN biofilm. Additionally, we observed that oleanolic acid can effectively suppress biofilm formation in pathogenic bacteria such as Salmonella and Staphylococcus aureus. Next, we assessed the amount of EcN extracellular polysaccharides, the number of live bacteria, their metabolic activity, the hydrophobicity of their surface, and the shape of their biofilms using laser confocal microscopy. Through transcriptome analysis, a total of 349 differentially expressed genes were identified, comprising 134 upregulated and 215 downregulated genes. GO functional enrichment analysis and KEGG pathway enrichment analysis revealed that oleanolic acid functions are through the regulation of bacterial motility, the iron absorption system, the two-component system, and adhesion pathways. These findings suggest that the main effects of oleanolic acid are to prevent bacterial motility, increase initial adhesion, and encourage the development of EcN biofilms. In addition, oleanolic acid interacts with iron absorption to cooperatively control the production of EcN biofilms within an optimal concentration range. Taking these results together, this study suggests that oleanolic acid may enhance probiotic biofilm formation in the intestines, presenting new avenues for probiotic product development.

Abstract Background Paratuberculosis is a widespread chronic infection of Mycobacterium avium subspecies paratuberculosis (MAP) that causes significant economic losses to the sheep industry. The current study investigated this disease, which causes diarrhea in sheep, particularly, in Bayannaoer, Inner Mongolia, China. Diagnosis was based on clinical symptoms, pathological autopsy, histopathological inspection, and serological and molecular methods. Results MAP was confirmed using polymerase chain reaction using DNA extracted from tissue and fecal samples. Serum samples from 472 individual sheep were obtained to detect antibodies against MAP using an enzyme-linked immunosorbent assay. MAP antibodies were separately detected in 17.86% (35/196) and 18.48% (51/276) of sheep herds at approximately 6 months and ≥ 1 year of age, respectively. The tissue lesion and pathological section results were consistent with paratuberculosis infection. Conclusions To our knowledge, this is the first report of Mycobacterium avium subspecies paratuberculosis seroprevalence in Bayannaoer sheep in Inner Mongolia. Our findings show that MAP is not only prevalent, but also a potential threat to this region. Further investigations, including long-term epidemiological surveillance and isolation are needed for the awareness and effective treatment of paratuberculosis in sheep of Inner Mongolia.

The influenza A virus replicates in a broad range of avian and mammalian species by hijacking cellular factors and processes. Avian influenza A viruses (AIVs) generally propagated poorly in mammalian cells, but some mutants of virus-encoded RNA polymerase components, especially PB2 subunit, can overcome host restriction. Host factors associated with PB2 may be essential for efficient AIV replication in mammalian cells. Here, we infected human cells with the PB2 Flag-tagged replication-competent recombinant AIV and identified cellular proteins that coprecipitate with PB2 protein by mass spectrometry. We confirmed one of the coprecipitating host factors, DEAD-box protein eIF4A3, that interacts with viral PB2, PB1, and NP proteins. Depletion of endogenous eIF4A3 significantly reduced virus replication. Later studies showed that eIF4A3 is essential for viral RNA polymerase activity and viral RNAs synthesis. Upon systematic dissection of the influenza virus progeny mRNA generation, from pre-mRNA processing to nuclear export, we found that the depletion of eIF4A3 resulted in significant defects in the ratio of M2 to M1 and NS2 to NS1, and the proportion of viral spliced mRNA in the nucleus increased, indicating that eIF4A3 plays a significant function in viral nascent intron mRNA splicing and spliced mRNA (M2 and NS2) nuclear export. Additionally, we confirmed that in specific deletion of eIF4A3, the synthesis of reduced NS2 can significantly impair neo-synthetized viral ribonucleoprotein (vRNP) nuclear export. Taken together, our findings revealed that eIF4A3 is a key mediator of AIV polymerase activity, mRNA splicing, and spliced mRNA nuclear export.

The H5 subtype virus of Highly Pathogenic Avian Influenza Virus has caused huge economic losses to the poultry industry and is a threat to human health. Until 2010, H5N1 subtype virus was the major genotype in China. Since 2011, reassortant H5N2, H5N6, and H5N8 viruses were identified in domestic poultry in China. The clade 2.3.4.4 H5N6 and H5N8 AIV has now spread to most of China. Clade 2.3.4.4 H5N6 virus has caused 17 human deaths. However, the prevalence, pathogenicity, and transmissibility of the distinct NA reassortment with H5 subtypes viruses (H5Nx) is unknown. We constructed five clade 2.3.4.4 reassortant H5Nx viruses that shared the same HA and six internal gene segments. The NA gene segment was replaced with N1, N2, N6, ΔN6 (with an 11 amino acid deletion at the 58th to 68th of NA stalk region), and N8 strains, respectively. The reassortant viruses with distinct NAs of clade 2.3.4.4 H5 subtype had different degrees of fitness. All reassortant H5Nx viruses formed plaques on MDCK cell monolayers, but the ΔH5N6 grew more efficiently in mammalian and avian cells. The reassortant H5Nx viruses were more virulent in mice as compared to the H5N2 virus. The H5N6 and H5N8 reassortant viruses exhibited enhanced pathogenicity and transmissibility in chickens as compared to the H5N1 reassortant virus. We suggest that comprehensive surveillance work should be undertaken to monitor the H5Nx viruses.