Study On The Inhibitory Effect Of Total Flavonoids From Ilex Rotunda Thunb. On Multidrug Resistant Bacteria

In this study,the object was Ilex Rotunda Thunb..The extraction of total flavonoids from Ilex Rotunda Thunb.and bacteriostasis of multi drug resistance bacteria of total flavonoids,especially against ESBLs-producing E.coli were the main resear-ch.The minimal inhibitory concentrations of total flavonoids from Ilex Rotunda Thunb.and antimicrobial agents against ESBLs-producing E.coli were determined by two-fold microdilution methods.Both the normal growth curve and that of E.coli treated with different concentrations of total flavonoids of Ilex Rotunda Thunb.were detected by using Colony Forming Units(CFU).The antibacterial effect of the total flavonoids combined with antimicrobial agents was determined by the combined passage method and microdilution checkerboard techniques.Agarose gel electrophoresis was used to analyze the effect of total flavonoids of Ilex Rotunda Thunb.to bacterial genome DNA.Flow cytometry analysis was used for the character of apoptosis on E.coli treated with different concentrations of total flavonoids of Ilex Rotunda Thunb.through Annexin-V and PI staining.The effect of total flavonoids of Ilex Rotunda Thunb.on the protein bands of E.coli was analyzed by protein spectrum analysis.The main results were as follows:1?Escherichia coli,Streptococcus,Pseudomonas aeruginosa,Enterococcus faecalis,Proteus vulgaris,Staphylococcus aureus,Acinetobacter baumannii were isolated.The E.coli was confirmed containing drug-resistant genes,such as fosA3,CTX-M and TEM and identified as ESBLs-producing E.coli.2.The bacter:ia isolated were all inhibited by total flavonoids from Ilex Rotunda Thunb.And the rate of bacterial growth became delayed.The total flavonoids could significantly inhibit the growth of ESBLs-producing E.coli,and the degree of inhibition of growth would be positively correlated with the concentration of total flavonoids.The total flavonoids and antimicrobial agents were combined to inhibit different resistant bacteria.For different resistant bacteria,the total flavonoids would exhibit synergistic or additive with different antibiotics.3.The amount of the genome DNA of E.coli treated with the total flavonoids of Ilex Rotunda Thunb.was less than the control groups.The total flavonoids of Ilex Rotunda Thunb.could reduce the amount of genome DNA of the ESBLs-producing E.coli,which was consistent with the change of growth curve of ESBLs-producing E.coli treated with total flavonoids of Ilex Rotunda Thunb.4.The percentage of apoptotic cells increased with addition of total flavonoids of Ilex Rotunda Thunb.and the percentage of apoptosis bacteria was increased.This increasing trendency was statistical significance.5.The results of SDS-PAGE indicated the total flavonoids could inhibit or eliminate the synthesis of total protein of E.coli,and it mainly influenced the fragment of about 35-40 kDa.Protein spectrum analysis by matrix assisted laser desorption mass spectrometry analysis showed that total flavonoids of Ilex Rotund Thunb.could mainly inhibit the content of ADP-glycerol-mannose-6-epimerase and fructose diphosphate aldolase.