【摘要】 目的利用透射电子显微镜和流式细胞仪等技术手段,对重组柞蚕抗菌肽AD抗菌作用机制进行初步研究。方法用微量肉汤法鉴定重组柞蚕抗菌肽AD对E.coli敏感质控标准菌株ATCC25922和产β-内酰胺酶菌株ATCC35218的抑菌作用后,利用透射电子显微镜观察经重组柞蚕抗菌肽AD处理前后E.coli超微结构变化,再用流式细胞Ca2+荧光探针技术检测经重组柞蚕抗菌肽AD处理前后E.coli细胞膜内游离钙离子浓度的变化。结果微量肉汤法证明重组柞蚕抗菌肽AD对E.coli敏感株ATCC25922和耐药株ATCC35218的MIC值均为8μg/m L;透射电镜观察到重组柞蚕抗菌肽AD对E.coli的作用主要表现为破坏细菌胞膜的完整性;流式细胞Ca2+荧光探针检测发现,经重组柞蚕抗菌肽AD作用后,细菌胞内表面上的二价阳离子Ca2+浓度增高。结论重组柞蚕抗菌肽AD对E.col i的抗菌机制可表现为膜完整性的破坏,并伴随胞内Ca2+浓度的增高。更多还原

【Abstract】 Objective To study the antibacterial mechanism of recombinant antimicrobial peptides Cecropin AD fromAntherea pernyi against E.coli by the technology of transmission electron microscopy and flow cytometry. Methods Afteridentifying the antibacterial action of antimicrobial peptides against both E.coli ATCC25922 and ESBLs- producing strainATCC35218, the change of ultrastructure of E.coli after Cecropin AD treatment were observed by transmission electronmicroscopy. Then, FCM calcium fluorescence probe technology was used to detect the change of intracellular free calcium.Result The MIC of Cecropin AD against E.coli ATCC25922 and ESBLs-producing strain ATCC35218 were 8μg/m L. Themechanism of cecropin AD against E.coli observed by transmission electron microscopy was mainly performanced bydestruction of the integrity of bacterial cell membrane. The concentration of divalent cation calcium was increased afterCecropin AD treatemt as determined by flow cytometry fluoscent probing. Conclusion With the increase of intracellular freecalcium concentration, membrane-destrcution might be one of the antibacterial mechanisms of Cecropin AD.更多还原