Inhibition Of Staphylococcus Aureus α-toxin And Enterotoxin Expression Through Natural Compound Sereening

Staphylococcus aureus, a virulent gram-positive pathogen, leads to both human andanimal diseases, including soft tissue infections, keratitis, pneumonia, bacteremia,endocarditis, ect. As a foodborne pathogen, S. aureus could cause food poisoning andgastroenteritis through the digestive tract. S. aureus is the leading cause of mastitis indairy cows, seriously affecting the quality of dairy products when the infectionoccurring. Therefore, S. aureus is the hidden trouble to jeopardy the public health andfood safety. In recent decades, due to the tremendous pressure on treatment of theinfection of methicillin-resistant S. aureus, MRSA and emergence of othermulti-resistant strains, treatment of S. aureus infection has become increasinglydifficult.Antimicrobial agents has played an role in the treatment of bacterial infection,accelerating the development in bacterial resistance at the same time. There arecomplex and multi-resistant strain contribute to the current dilmma, of which in newantibacterial drugs research and development. The relationship between thedevelopment of bacterial resistance and the antimicrobial usage lead to a transform ofthe study point in anti-infection drug development. Recent years, more and moreinstitutions prefer to intervene the infection, rather than to investigate thebacteriostatic and bactericidal capability. It received the widespread attention oftargeting pathogen virulence factors in developing new antibacterial drugs. Targetingvirulence factors strategy could lessen the bacterial resistance, because they are notessential for bacteria grow with low selection pressure.S. aureus secrete various of virulence factors, including extracellular toxins,enzymes and surface proteins, and these virulence factors determinate the pathogen ofBacteria. S. aureus α-toxin is a significant virulence factor, secreting at thepost-exponential phase. As sensitive to red blood cells, α-toxin induce cell lysis anddead by form a mushroom to the heptamer on cell surface. α-toxin play an important role in causing diseases. For example, S. aureus strains lacking α-toxin show lessvirulence in the model of animals pneumonia by knocking out hla gene. However, thevirulence of S. aureus didn’t decline in which knocking out other genes.Staphylococcus food poisoning does not result from the ingestion of S. aureus itself,but rather from ingestion S. aureus enterotoxins in food, with vomiting and diarrhoea.S. aureus enterotoxins are stable when heat treatment and are resistant to treatmentwith strong acids and alkalis. It hard to damaged the S. aureus enterotoxins bytraditional food processing. Additionally, S. aureus enterotoxins stimulate T-cellactivation and the release of T-cell-derived cytokines.In our study, we screen five natural compounds for both not affect the growth ofS. aureus and has ability to anti-virulence factors activity. We use MIC, hemolysis,T-cell proliferation assay and western blot to preliminary screenin. We found thatsilibinin does not affect growth of S. aureus, can markedly inhibit the hemolysisactivity of α-toxin at low concentrations and reduce T-cell proliferation. And westernblot prove that silibinin inhibits the production of α-toxin,SEA and SEB. Then wefound that silibinin decrease the expression of hla,sea,seb and RNAIII of agr byRT-PCR. Therefore, we infer that the reduction of α-toxin, SEA and SEB in S. aureusin the presence of silibinin, in part, originate from the inhibition of the agrtwo-component system. In cytotoxicity assays, confocal laser scanning microscopeand LDH assays were employed to investigate for both qualitative analysis andquantitative analysis, and we found silibinin attenuates α-toxin-mediated injury ofhuman alveolar epithelial cells (A549). The result of Enzyme-linked immunosorbentassay also demonstrated that silibinin could reduce TNF-α release caused by S.aureussupernatants.