Abstract: OBJECTIVE: To observe the effects of New Dayuan powder(NDYP) on Methicillin-resistant Staphylococcus aureus(MRSA) biofilms and the embedded bacteria in vitro.METHODS: 2,3-Bis-(2-methoxy-4-nitro-5-sulfophenyl)-2 H-tetrazolium-5-carboxanilide(XTT) assays were used to study the effects of NDYP on developing MRSA biofilms: 100 μL of bacterial culture and100 μL drug solution were added to wells of96-well plates. After 24 h of incubation, the plates were washed and XTT-phenazine methyl sulfate(PMS) was added to enable counting of the number of live bacteria in biofilms using a microplate reader. XTT assays were also used to explore the effects of NDYP on mature MRSA biofilms: 100 μL of bacterial culture were added to wells of 96-well plates. Bacteria were cultured in the plates for 24 h,and then drug solution was added. The plates were cultured for another 24 h, and then XTT-PMS was added to detect the number of live bacteria in the biofilms. Scanning electron microscopy(SEM) was used to observe the effects of NDYP on mature MRSA biofilms: washed and sterilized glass coverslips were added to 24-well plates. Bacterial culture was added. After 24 h of incubation, drug solution was added. After another 24 h of incubation, the samples were observed by SEM.RESULTS: XTT assays showed that the number of live bacteria in both developing and mature MRSA biofilms decreased significantly(P < 0.01) after the administration of NDYP. SEM images showed that NDYP could destroy the structure of the bacteria and resulted in uneven thickness of MRSA biofilms.CONCLUSION: In vitro, NDYP has obvious inhibitory effects on the formation of MRSA biofilms and on mature biofilms.

Key words: Methicillin-resistant Staphylococcus aureus, Biofilms, In vitro techniques, New Dayuan powder