Study on the reduction of MRSA biofilm formation by Brazilin through the inhibition of SarA in a PIA-dependent manner
Objective The goal was to find out how Brazilin(BN)inhibited methicillin-resistant Staphylococcus aureus(MRSA)from making biofilms and to give new ideas for using traditional Chinese medicine to prevent and treat MRSA infections.Methods A biofilm model of the MRSA ATCC43300 strain was established.The experiment was divided into BN 8,16 and 32 μg/mL groups as well as a control group.The inhibitory effect of BN on MRSA biofilm was determined through the crystal violet method.The Congo red agar plate method and the sulfuric acid-phenol method were employed to detect the effect of BN on the synthesis of polysaccharide intercellular adhesion(PIA).Quantitative real-time PCR(qRT-PCR)was used to verify the transcription levels of the icaA and icaD gene clusters involved in intercellular adhesion.High-throughput transcriptomic sequencing was performed to identify potential biofilm-inhibitory genes.qRT-PCR and Western blot validated the relevant genes and proteins.The mouse model of MRSA biofilm infection through subcutaneous placement of an intrvenous catheter was established to further validate the efficacy of BN.Results Compared to the control group,BN at different concentrations significantly reduced MRSA biofilm and PIA formation.Specifically,at the lowest dose of 8 μg/mL BN,MRSA biofilm formation was decreased by 40.17％(P＜0.01)and PIA by 55.56％(P＜0.01).qRT-PCR verification indicated that BN at different concentrations significantly inhibited the expression of the icaA and icaD genes.Compared to the control group,the expression levels of icaA and icaD genes were reduced by 59.03％(P＜0.01)and 48.33％at an 8 μg/mL concentration(P＜0.05),respectively.High-throughput sequencing revealed a sarA downregulation of-1.458 log2.qRT-PCR and Western blot results showed that BN significantly inhibited the transcription and protein expression of sarA.In vivo experiments indicated that BN effectively inhibited MRSA biofilm formation in mice.Conclusion BN reduces the synthesis of PIA by inhibiting the expression of SarA and thereby intervening in biofilm formation.
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