Alves Fernanda, de Oliveira Mima Ewerton Garcia, Passador Renata Caroline Polato, Bagnato Vanderlei Salvador, Jorge Janaína Habib, Pavarina Ana Cláudia
Department of Dental Materials and Prosthodontics, Araraquara Dental School, UNESP - Univ. Estadual Paulista, Rua Humaitá, 1680, 14801-903, Araraquara, SP, Brazil.
Biophotonics Lab, Group of Optics, Physics Institute of São Carlos, University of São Paulo - USP, Av. Trabalhador São-carlense, 400, 13566-590, São Carlos, SP, Brazil.
Lasers Med Sci. 2017 May;32(4):815-826. doi: 10.1007/s10103-017-2177-y. Epub 2017 Mar 9.
This study evaluated the effects of antimicrobial photodynamic therapy (aPDT) mediated by Photodithazine® (PDZ) and LED light on the virulence factors of fluconazole-susceptible (CaS) and fluconazole-resistant (CaR) Candida albicans. Standardized suspensions of strains were prepared (10), and after 48 h of biofilm formation, these strains were incubated with PDZ (100 mg/L) for 20 min and exposed to LED light (660 nm, 37.5 J/cm). Additional samples were treated with PDZ or light only, and the control consisted of biofilms that received no treatment. After aPDT, the cells were recovered and the virulence factors were evaluated. To analyze the capacity of adhesion, cells were recovered after aPDT and submitted to the adhesion process in the bottom of a 96-well plate. After this, metabolic activity tests (XTT assay) and cell viability (colony forming units per milliliter, CFU/mL) were applied. To evaluate the biofilm-forming ability after aPDT, the cells recovered were submitted to biofilm formation procedures, and the biofilm formed was evaluated by XTT, CFU/mL, and total biomass (crystal violet) tests. Lastly, the capacity for synthesizing protease and phospholipase enzymes after aPDT was evaluated by fluorimetric tests. Data were analyzed by two- or three-way ANOVA tests (p ≤ 0.05). It was verified that aPDT reduced the viability of both strains, fluconazole-susceptible and fluconazole-resistant C. albicans. It was also observed that the CaR strain had lower susceptibility to the aPDT when compared with the CaS strain. However, regarding the virulence factors evaluated, it was demonstrated that aPDT did not alter the adherence and biofilm formation ability and enzymatic production.
本研究评估了由Photodithazine®(PDZ)和LED光介导的抗菌光动力疗法(aPDT)对氟康唑敏感(CaS)和氟康唑耐药(CaR)白色念珠菌毒力因子的影响。制备菌株的标准化悬液(10个),在生物膜形成48小时后,将这些菌株与PDZ(100mg/L)孵育20分钟,然后暴露于LED光(660nm,37.5J/cm)。另外的样本仅用PDZ或光处理,对照组为未处理的生物膜。aPDT处理后,回收细胞并评估毒力因子。为分析黏附能力,aPDT处理后回收细胞,并将其置于96孔板底部进行黏附过程。之后,进行代谢活性测试(XTT测定)和细胞活力(每毫升菌落形成单位,CFU/mL)检测。为评估aPDT后的生物膜形成能力,将回收的细胞进行生物膜形成程序,通过XTT、CFU/mL和总生物量(结晶紫)测试评估形成的生物膜。最后,通过荧光测试评估aPDT后合成蛋白酶和磷脂酶的能力。数据通过双向或三向方差分析检验(p≤0.05)。结果证实,aPDT降低了氟康唑敏感和氟康唑耐药白色念珠菌这两种菌株的活力。还观察到,与CaS菌株相比,CaR菌株对aPDT的敏感性较低。然而,关于所评估的毒力因子,结果表明aPDT并未改变黏附、生物膜形成能力和酶的产生。