Abstract: In order to study the effects of berberine on the elimination of drug resistance of E.coli with ultra-broad spectrum β-lactamase（ESBLs）, the MIC changes of 3 ESBLs E. coli strains before and after berberine treatment were measured by double dilution method. Then, the changes of R plasmid, ESBLs activity, biofilm formation and the transcription levels of AcrA mRNA and AcrB mRNA in efflux pump of E.coli after berberine treatment were detected by agalose gel electrophoresis, ELISA and real-time fluorescence quantitative PCR. The results showed that the MIC of 3 E.colis to berberine was 2.00 mg/mL. After treatment with berberine, the MIC of 2 ESBLs E.colis against cefotaxime decreased from 32.00 mg/mL to 8.00 mg/mL, and the MIC of 1 ESBLs E.colis against cefotaxime decreased from 32.00 mg/mL to 16.00 mg/mL. After treatment with berberine, the plasmid bands of 2 strains of ESBLs disappeared, and the plasmid bands of 1 strain of ESBLs disappeared. The ESBLs activity of 3 ESBLs strains was significantly decreased（P＜0.01）. Alkaline phosphatase activity was significantly increased（P＜0.01）. The transcription level of AcrA mRNA in efflux pump of ESBLs was significantly decreased in 3 strains（P＜0.01）, and the transcription level of AcrB mRNA in efflux pump of ESBLs was decreased in 2 strains（P＜0.05）. These results indicated that berberine had a good effect on the 4 drug resistance elimination mechanisms of ESBLs E. coli.