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目的:探讨低频超声对血卟啉单甲醚(Hematoporphyrin monomerthyl ether,HMME)在金黄色葡萄球菌生物膜中渗透效果的影响。方法:将直径25 mm,孔径0.22μm的微孔滤膜平铺于LB琼脂培养基表面,再将纯种的金黄色葡萄球菌(ATCC6538)菌液,通过均匀涂布接种到滤膜表面,37℃恒温培养48 h获得实验所需的生物膜模型。将培养所得生物膜样本随机分为空白对照组,单独超声组,单独药物组和实验组,其中实验组按超声强度(0.5 w/cm~2或1 w/cm~2)和超声作用时间(1 min、2 min或3 min)不同又分为实验(A、B、C、D、E、F)组。所用渗透药物为20μg/m L的HMME溶液。对照组使用等量的生理盐水孵育,孵育时间为3 min,单独超声组也用生理盐水孵育并加0.5 W/cm~2的超声作用3 min。HMME在405 mm激发光下有特征性光谱,通过测量光谱曲线下积分面积,可间接反映生物膜中HMME渗入量。结果:实验A组(0.5 w/cm~2,1 min)的荧光光谱曲线下积分面积显著高于单独药物组(P<0.05)。声强相同时,随作用时间增加,所得光谱曲线下积分面积显著增加(P<0.01或P<0.05);作用时间相同时,声强越大,光谱曲线下积分面积亦越大(P<0.01)。结论:低频超声可以显著增加HMME渗透进入金黄色葡萄球菌生物膜的量,且渗透效果与超声作用时间和超声声强正相关。
Objective: To investigate the effect of low frequency ultrasound on the permeation of Hematoporphyrin monomerthyl ether (HMME) in S. aureus biofilm. Methods: A microporous membrane with a diameter of 25 mm and a pore size of 0.22 μm was plated on the surface of LB agar medium. The pure strain of Staphylococcus aureus (ATCC6538) was inoculated onto the surface of the filter membrane through uniform coating. 37 Incubated at 48 ℃ for 48 h to obtain the experimental biofilm model. The cultured biofilm samples were randomly divided into blank control group, ultrasound alone group, separate drug group and experimental group, the experimental group according to ultrasound intensity (0.5w / cm ~ 2 or 1w / cm ~ 2) and sonication time 1 min, 2 min or 3 min) was divided into experimental group (A, B, C, D, E, F). The osmotic drug used was a 20 μg / mL HMME solution. The control group was incubated with the same amount of saline, the incubation time was 3 min, the ultrasound group was also incubated with saline and 0.5 W / cm ~ 2 ultrasound for 3 min. HMME has a characteristic spectrum under 405 mm excitation light. By measuring the integral area under the spectral curve, the amount of HMME infiltration in the biofilm can be indirectly reflected. Results: The integral area under the fluorescence spectrum of experimental group A (0.5 w / cm ~ 2,1 min) was significantly higher than that of single drug group (P <0.05). With the same sound intensity, the integral area under the spectral curve increased significantly (P <0.01 or P <0.05) with the increase of the action time. The larger the sound intensity and the larger the integral area under the spectral curve (P <0.01 or P <0.01) ). Conclusion: Low-frequency ultrasound can significantly increase the amount of HMME infiltrated into S. aureus biofilm, and the infiltration effect is positively correlated with the duration of ultrasound and the intensity of ultrasound.