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Evaluation of the effectiveness of Dnase associated with photodynamic therapy in inactivating biofilms from clinical isolates of Candida albicans resistant to fluconazole

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César Augusto Abreu Pereira
Total Authors: 1
Document type: Master's Dissertation
Press: Araraquara. 2021-08-02.
Institution: Universidade Estadual Paulista (Unesp). Faculdade de Odontologia. Araraquara
Defense date:
Advisor: Ana Claudia Pavarina; Marlise Inêz Klein Furlan

Antimicrobial photodynamic therapy (PDT) promotes microorganisms’ inactivation in planktonic cultures; however, the same effect does not occur in biofilms. The resistance of Candida spp. biofilms is multifactorial and it has been associated to the protective effect of biofilm’s matrix extracellular (MEC) that prevents the penetration of photosensitizing agent (PA) and drugs. The objective of this study was to evaluate the effectiveness of DNase I enzyme associated with PDT, mediated to Photodithazine® (200 mg/L) and LED light (660 nm; 50 J/cm2 ), on biofilms of fluconazole-resistant C. albicans’ clinical isolates (CaR). 48h-old biofilms were formed from the strains of CaR [ATCC96901; and two clinical isolates (R14 and R70)]. DNase was used in the aim of degrading the MEC of the biofiilms and potentializing the effect of PDT. Biofilms were exposed to pre-incubation with DNase for 5 minutes, followed by photosensitizer aplication (P) and the ligh (L), either in isolated or associated form, giving rise for 8 groups of treatment (P+L+, P-L+, P+L-, P-L-, P-L-DNase, P+L+DNase, P+L-DNase and P-L+DNase; n=12). The effectiviness of treatments was evaluated trough the following methods: colony forming unit (CFU) counting, quantification of: total and isoluble dry-weight, soluble and insoluble proteins, water-soluble polysaccharides from (WSP) and alkali-soluble polysaccharides (ASP), and extracellular DNA (eDNA). The data were analyzed by three-way ANOVA test with Bonferroni's post-test. The DNase treatment associated with PDT (P+L+DNase) showed a reduction of 1.92; 1.65 e 1.29 in log10 of cell viability to ATCC 96901, R14 and R70 strains, respectively. It was also noticed a reduction of biofilms MEC’s components, like WSP and eDNA, as well as its insoluble biomass when it’s been compared with control group (P-L-). The total dry-weight and the proteins quantification showed no statistical diference between treatments. All of the strains showed the same standard behavior when treating with DNase, and MEC’s components were affected, improving PDT’s effectiveness. (AU)

FAPESP's process: 19/11602-8 - Evaluation of the efficacy of DNase associated with photodynamic therapy against fluconazole resistant Candida albicans biofilm
Grantee:César Augusto Abreu Pereira
Support Opportunities: Scholarships in Brazil - Master