Abstract：
OBJECTIVE: Candida albicans is the most prevalent pathogenic fungus, exhibiting escalating multidrug resistance (MDR). Antimicrobial peptides (AMPs) represent promising candidates for addressing this issue. In this research, five antimicrobial peptides, ACP1 to ACP5 which named ACPs were studied as alternative fungicidal molecules.
METHODS: CD assay was used to analyze the 2D structures, Absorbance method was used to test the antimicrobial activity, haemolytic activity, time-kill kinetics, biofilm inhibition and reduction activity, resistance induction activity and assessment against fluconazole-resistant C. albicans. SEM, TEM, CLSM, flow cytometer and FM were carried out to provide insight into the mechanisms of anti-Candida action.
RESULTS: ACPs possessed an α-helical structure and strong anti-Candida activities, with minimum inhibitory concentrations (MICs) from 3.9 to 15.6 μg/mL. In addition, ACPs did not produce hemolysis at concentrations lower than 10 or 62 × MIC, indicating their low cytotoxicity. Fungicidal kinetics showed that they completely killed C. albicans within 8 h at 2 to 4 × MIC. Notably, ACPs were highly fungicidal against fluconazole-resistant C. albicans and showed low resistance. In addition, they were effective in inhibiting mycelium and biofilm formation. Fluorescence microscopy revealed that while fluconazole had minimal to no inhibitory effect on biofilm-forming cells, ACPs induced apoptosis in all of them. The research on mechanism of action revealed that ACPs disrupted the cell membranes, with ROS increasing and cellular mitochondrial membrane potential decreasing.
CONCLUSIONS: ACPs could be promising candidates for combating fluconazole-resistant C. albicans infections.
METHODS: CD assay was used to analyze the 2D structures, Absorbance method was used to test the antimicrobial activity, haemolytic activity, time-kill kinetics, biofilm inhibition and reduction activity, resistance induction activity and assessment against fluconazole-resistant C. albicans. SEM, TEM, CLSM, flow cytometer and FM were carried out to provide insight into the mechanisms of anti-Candida action.
RESULTS: ACPs possessed an α-helical structure and strong anti-Candida activities, with minimum inhibitory concentrations (MICs) from 3.9 to 15.6 μg/mL. In addition, ACPs did not produce hemolysis at concentrations lower than 10 or 62 × MIC, indicating their low cytotoxicity. Fungicidal kinetics showed that they completely killed C. albicans within 8 h at 2 to 4 × MIC. Notably, ACPs were highly fungicidal against fluconazole-resistant C. albicans and showed low resistance. In addition, they were effective in inhibiting mycelium and biofilm formation. Fluorescence microscopy revealed that while fluconazole had minimal to no inhibitory effect on biofilm-forming cells, ACPs induced apoptosis in all of them. The research on mechanism of action revealed that ACPs disrupted the cell membranes, with ROS increasing and cellular mitochondrial membrane potential decreasing.
CONCLUSIONS: ACPs could be promising candidates for combating fluconazole-resistant C. albicans infections.
摘要：
目的：白色念珠菌是最常见的病原真菌，表现出不断升级的多药耐药性（MDR）。抗微生物肽(AMP)代表了用于解决该问题的有希望的候选物。在这项研究中,五种抗菌肽，研究了命名为ACP的ACP1至ACP5作为替代杀真菌分子。
方法：CD测定用于分析2D结构，采用吸光度法检测抗菌活性,溶血活性，时间-杀伤动力学，生物膜抑制和还原活性，对氟康唑耐药白色念珠菌的耐药诱导活性及评价。SEM,TEM,CLSM,进行流式细胞仪和FM以深入了解抗念珠菌作用的机制。
结果：ACPs具有α-螺旋结构和较强的抗念珠菌活性，最低抑制浓度（MIC）为3.9至15.6μg/mL。此外,ACP在浓度低于10或62×MIC时不产生溶血，表明它们的细胞毒性低。杀真菌动力学表明，它们在8小时内以2至4×MIC完全杀死白色念珠菌。值得注意的是,ACP对氟康唑耐药的白色念珠菌具有高度杀真菌性，并且显示出低抗性。此外,它们能有效抑制菌丝体和生物膜的形成。荧光显微镜显示，虽然氟康唑对生物膜形成细胞有最小的抑制作用，ACPs均诱导细胞凋亡。作用机制的研究表明，ACP破坏了细胞膜，随着ROS的增加和细胞线粒体膜电位的降低。
结论：ACP可能是对抗氟康唑耐药白色念珠菌感染的有希望的候选药物。
方法：CD测定用于分析2D结构，采用吸光度法检测抗菌活性,溶血活性，时间-杀伤动力学，生物膜抑制和还原活性，对氟康唑耐药白色念珠菌的耐药诱导活性及评价。SEM,TEM,CLSM,进行流式细胞仪和FM以深入了解抗念珠菌作用的机制。
结果：ACPs具有α-螺旋结构和较强的抗念珠菌活性，最低抑制浓度（MIC）为3.9至15.6μg/mL。此外,ACP在浓度低于10或62×MIC时不产生溶血，表明它们的细胞毒性低。杀真菌动力学表明，它们在8小时内以2至4×MIC完全杀死白色念珠菌。值得注意的是,ACP对氟康唑耐药的白色念珠菌具有高度杀真菌性，并且显示出低抗性。此外,它们能有效抑制菌丝体和生物膜的形成。荧光显微镜显示，虽然氟康唑对生物膜形成细胞有最小的抑制作用，ACPs均诱导细胞凋亡。作用机制的研究表明，ACP破坏了细胞膜，随着ROS的增加和细胞线粒体膜电位的降低。
结论：ACP可能是对抗氟康唑耐药白色念珠菌感染的有希望的候选药物。
