Photodynamic inactivation of Candida tropicalis biofilms by parietin, a natural dihydroxyanthraquinone

MARIONI J; MUGAS ML; BORSARELLI CD; NÚÑEZ MONTOYA SC

Congreso; XIV ENCUENTRO LATINOAMERICANO DE FOTOQUÍMICA Y FOTOBIOLOGÍA (ELAFOT); 2019

ENCUENTRO LATINOAMERICANO DE FOTOQUÍMICA Y FOTOBIOLOGÍA

The 1,8-dihydroxy-3-methoxy-6-methylanthraquinone (Parietin, PTN), a blue-light absorbing pigment present in several lichens of the genus Teloschistes spp., shows photosensitizing properties through both Type I (superoxide anion, O2?-) and Type II (singlet molecular oxygen, 1O2) mechanism, with antibacterial effects in vitro when it was photo-stimulated [1]. The aim of this work was to evaluate the in vitro antifungal activity of PTN, analyzing whether this effect could be increased by actinic irradiation, and establishing if the photoinactivation mechanism involves the production of Reactive Oxygen Species (ROS).PTN was isolated from the lichen Teoloschistes nodulifer (Nyl.) Hillman (Teloschistaceae) and it was purified by recrystallization from the acetone extract, and its purity was determined by HPLC. Minimum Inhibitory Concentration (MIC) of PTN was determined by following the protocols of the Clinical and Laboratory Standards Institute [2], testing twelve concentrations (0.24 to 500 µg/mL) in planktonic cultures of Candida tropicalis NCPF 3111 (103 cells/mL). PTN aqueous solution (ethanol as co-solvent <1%) was added to the fungal suspension following the additive serial double dilutions method, and immediately the assay was performed simultaneously under darkness and illumination during 15 min with a Philips TL/03 actinic lamp emitting in the range 380-480 nm. Under darkness, PTN did not decrease meaningfully the growth of planktonic cells of C. tropicalis, but under irradiation the MIC was 0.98 µg/mL.The antifungal activity of PTN was also tested on C. tropicalis biofilms at three concentrations in triplicate: MIC, MIC/2 and MICx2, under darkness and same irradiation conditions than in the case of yeast suspensions. After 48 h of incubation, aliquots of biofilm were removed and serially diluted. The number of viable yeasts was determined also in triplicate, and the counting of colony forming units (CFU/mL) was log-transformed. In addition, the scavenging effect of 200 mM sodium azide (1O2 quencher) or Tiron (O2?- quencher) on biofilm photoinactivation by PTN was evaluated. Results show that PTN did not produced antibiofilm activity in darkness, but under blue light a 3 log reduction of the CFU at the MIC2 was observed. Moreover, the antibiofilm photodynamic activity of PTN was completely suppressed in the presence of sodium azide or Tiron.In conclusion, PTN was able to inactivate C. tropicalis biofilms under the action of blue-light, by means of Type I and II photodynamic mechanisms. Therefore, PTN can be a promising photosensitizer candidate for application in photodynamic therapy of Candida infections.