The Antibacterial Activity Of Venenum Bufonis In Combination With Aminoglycosides Against Multi-drug Resistant Acinetobacter Baumannii

Objective To explore the antibacterial activity of Venenum Bufonis in combination with aminoglycosides against Acinetobacter Baumannii,and the correlation of drug resistant phenotype and genotype in multi-drug resistant Acinetobacter Baumannii.To discuss the impacts of different times of Venenum Bufonis on the expression of aminoglycosides resistance gene.Method Minimum inhibitory concentrations(MICs)of Venenum Bufonis(alcohol extract or aqueous extract),Amikacin,Gentamicin,and Netilmicin were examined by using MH agar plates,PDA agar plates and mini-dilution,meanwhile MIC of Venenum Bufonis in combination with Amikacin.The resistance gene of 40 strains of Acinetobacter Baumannii by PCR,the impacts of different times of Venenum Bufonis(two extracting solution)on the expression of aminoglycosides resistance gene by RT-PCR.Result MIC50 of Venenum Bufonis(alcohol extract)was 0.625mg/ml,MIC90 was 1.25mg/ml.MIC50 of Venenum Bufonis(aqueous extract)was 1.25mg/ml,MIC90 was 2.5mg/ml.MIC50 of Amikacin was 128μg/ml,MIC90 was 512μg/ml,combining with Venenum Bufonis(alcohol extract)MIC50 was 32μg/ml,MIC90 was 256g/ml,while combining with Venenum Bufonis(aqueous extract)MIC50 was 64μg/ml,MIC90 was 512g/ml;MIC50 of Gentamicin was 256μg/ml,MIC90 was 1024μg/ml,c.ombining with Venenum Bufonis(alcohol extract)MIC50 was 128μg/ml,MIC90 was 512g/ml,while combining with Venenum Bufonis(aqueous extract)MIC50 was 128μg/ml,MIC90 was 1024g/ml;MIC50 of Netilmicin was 512μg/ml,MIC90 was 1024μg/ml,combining with Venenum Bufonis(alcohol extract)MIC50 was 256μg/ml,MIC90 was 1024g/ml,while combining with Venenum Bufonis(aqueous extract)MIC50 was 256μg/ml,MIC90 was 1024g/ml.Aminoglycosides resistance gene of Acinetobacter Baumannii(40 strains)were detected.The positive of aac(6’)-Ⅱ,AMI-strA-B,AMI-aphA1,ant(3")-I,aac(6’)I,AMI-aadA1,AMI-AadDA1,ant(2")-Ⅰ,aadAlwere 35(87.5%),29(72.5%),27(67.5%),23(57.5%),22(55.0%),20(50.0%),19(47.5%),16(40.0%)and13(32.5%),respectively.The expression of aac(6’)-Ⅱ and aac(6’)-Ⅰ were decreased after 16 to 24 hours with Venenum Bufonis(alcohol extract and aqueous extract),especially alcohol.Venenum Bufonis(alcohol extract)has weak inhibition effects on the expression of ant(3")I after 16 hours.The inhibition effects of Venenum Bufonis(alcohol extract)on AMI-aphA1 and AMIaadA1 was coincident with Venenum Bufonis(aqueous extract)for two hours.Conclusion Venenum Bufonis has synergistic antibacterial effect with aminoglycosides.Aminoglycosides-resistant Acinetobacter Baumannii is related to the high expression of transacetylase aac(6’)-Ⅱ and aac(6’)-Ⅰ.Venenum bufonis(especially alcohol extract)and aminoglycosides against multi-drug resistant Acinetobacter Baumannii relative to the inhibition of transacetylase aac(6’)-Ⅱ and aac(6’)-Ⅰ.