Reznick Yana, Banin Ehud, Lipovsky Anat, Lubart Rachel, Polak Pazit, Zalevsky Zeev
Department of Physics, Bar-Ilan University.
J Vis Exp. 2013 Jul 2(77):e4370. doi: 10.3791/4370.
Recently there were several publications on the bactericidal effect of visible light, most of them claiming that blue part of the spectrum (400 nm-500 nm) is responsible for killing various pathogens(1-5). The phototoxic effect of blue light was suggested to be a result of light-induced reactive oxygen species (ROS) formation by endogenous bacterial photosensitizers which mostly absorb light in the blue region(4,6,7). There are also reports of biocidal effect of red and near infra red(8) as well as green light(9). In the present study, we developed a method that allowed us to characterize the effect of high power green (wavelength of 532 nm) continuous (CW) and pulsed Q-switched (Q-S) light on Pseudomonas aeruginosa. Using this method we also studied the effect of green light combined with antibiotic treatment (gentamycin) on the bacteria viability. P. aeruginosa is a common noscomial opportunistic pathogen causing various diseases. The strain is fairly resistant to various antibiotics and contains many predicted AcrB/Mex-type RND multidrug efflux systems(10). The method utilized free-living stationary phase Gram-negative bacteria (P. aeruginosa strain PAO1), grown in Luria Broth (LB) medium exposed to Q-switched and/or CW lasers with and without the addition of the antibiotic gentamycin. Cell viability was determined at different time points. The obtained results showed that laser treatment alone did not reduce cell viability compared to untreated control and that gentamycin treatment alone only resulted in a 0.5 log reduction in the viable count for P. aeruginosa. The combined laser and gentamycin treatment, however, resulted in a synergistic effect and the viability of P. aeruginosa was reduced by 8 log's. The proposed method can further be implemented via the development of catheter like device capable of injecting an antibiotic solution into the infected organ while simultaneously illuminating the area with light.
最近有几篇关于可见光杀菌作用的出版物，其中大部分声称光谱的蓝色部分（400纳米 - 500纳米）负责杀死各种病原体（1 - 5）。蓝光的光毒性作用被认为是内源性细菌光敏剂光诱导活性氧（ROS）形成的结果，这些光敏剂大多在蓝色区域吸收光（4,6,7）。也有关于红光和近红外光（8）以及绿光（9）杀菌作用的报道。在本研究中，我们开发了一种方法，使我们能够表征高功率绿色（波长532纳米）连续（CW）和脉冲调Q（Q - S）光对铜绿假单胞菌的影响。使用这种方法，我们还研究了绿光与抗生素治疗（庆大霉素）联合对细菌活力的影响。铜绿假单胞菌是一种常见的医院获得性机会致病菌，可引起各种疾病。该菌株对各种抗生素相当耐药，并含有许多预测的AcrB/Mex型RND多药外排系统（10）。该方法利用在含有和不含有抗生素庆大霉素的情况下，暴露于调Q和/或连续波激光器的Luria肉汤（LB）培养基中生长的自由生活的稳定期革兰氏阴性细菌（铜绿假单胞菌菌株PAO1）。在不同时间点测定细胞活力。所得结果表明，与未处理的对照相比，单独激光处理不会降低细胞活力，单独庆大霉素处理仅使铜绿假单胞菌的活菌数减少0.5个对数。然而，激光和庆大霉素联合处理产生了协同效应，铜绿假单胞菌的活力降低了8个对数。所提出的方法可以通过开发一种类似导管的装置进一步实施，该装置能够在向感染器官注射抗生素溶液的同时用光照射该区域。
