【摘要】 目的 研究植物源光敏剂竹红菌素衍生物（HB-TP）介导的光动力对创伤弧菌的体外杀伤作用以及对细菌侵袭力的影响。方法 （1）体外杀伤效应评价：设空白对照组（C组）、光敏剂对照组（P组）、光照对照组（L组）和光动力组（PDT组）。其中，PDT组菌悬液分别采用1.25μmol/L、2.5μmol/L、5μmol/L和10μmol/L的HB-TP孵育30 min后行波长630 nm激光照射（根据功率密度40 mW/cm~2和100 mW/cm~2再分为两组，能量密度均为72 J/cm~2）。各组处理后采用稀释平板法计数各组菌落形成单位，得出亚致死PDT剂量后，对PDT组和对照组进行超微结构的观察，分析鞭毛等超微结构变化。（2）对细菌侵袭力影响的检测：将亚致死PDT剂量处理后的菌悬液加入贴壁生长于24孔板的Hela细胞，孵育1 h后，洗涤、裂解细胞以释放细胞内细菌，稀释平板法计数各组菌落形成单位，计算侵袭率。与未行PDT处理的对照组细菌的侵袭率比较，分析侵袭力的变化。结果 在相同HB-TP孵育浓度和相同能量密度下，功率密度100 mW/cm~2对创伤弧菌的杀伤作用强于功率密度40 mW/cm~2(P<0.05)，且PDT两个组的细菌存活对数值均随HB-TP孵育浓度增加而增加，达有效杀菌作用的孵育浓度分别为5μmol/L和10μmol/L。亚致死PDT剂量处理后，可观察到创伤弧菌鞭毛缺失、荚膜减少或消失、细胞壁连续性破坏及空泡形成；创伤弧菌在Hela的侵袭率由阳性对照组的80.2%降低为0.0%(P<0.05)。结论 HB-TP介导的光动力疗法可有效杀伤体外培养的创伤弧菌，破坏其鞭毛等超微结构，降低其侵袭力。更多还原

【Abstract】 Objective To study the in vitro killing effect of the phytogenic photosensitizer(HB-TP) mediated PDT on Vibrio vulnificus and assess the damaging effect on bacterial flagella and invasive abilities.Methods （1） In vitro killing effect evaluation: Four groups, namely, blank control group(group C), photosensitizer group(group P), light group(group L) and photodynamic group(group PDT), were set. The vibrio vulnificus suspension in the PDT groups were incubated with 1. 25 μmol/L, 2. 5 μmol/L, 5 μmol/L and 10 μmol/L of HB-TP for 30 min respectively and then irradiated with 630 nm laser. The PDT group was further divided into two groups by power density(40 mW/cm~2 and 100 mW/cm~2 at the same energy density 72 J/cm~2). After treatment, the colony forming unit of each group was counted via dilution plate method. Immediately after the treatment with the sublethal aPDT dose obtained in part(1), the bacterial suspension was centrifuged, collected and observed under transmission electron microscope after negative staining. Based on a comparison with the blank control group, the ultrastructural changes such as flagella were analyzed.(2) Detection of the influence on bacterial invasive abilities: The bacterial suspension treated with sublethal aPDT dose was added to Hela cells growing on a 24-well plate. After incubation for 1 hour, the cells were lysed to release intracellular bacteria. Colony forming units of each group were counted via dilution plate method, and the invasion rate was calculated. The invasion rate of bacteria was compared with that of the control group, and the changes of invasion ability were analyzed.Results Under the same HB-TP incubation concentration and the same energy density, the killing effect of 100 mW/cm~2 to Vibrio vulnificus was stronger than 40 mW/cm~2(P<0. 05), and the effective bactericidal photosensitizer incubation concentrations were 5 μM and 10 μM respectively. After sublethal aPDT dose treatment, Vibrio vulnificus flagella loss, capsule reduction or disappearance, cell wall continuity disruption and vacuole formation were observed. The invasion rate of Vibrio vulnificus in Hela decreased from 80. 2%(positive control group) to 0. 0%. Conclusions HB-TP mediated photodynamic therapy will effectively kill Vibrio vulnificus in vitro, destroy its flagellar structure and reduce its invasiveness.更多还原