Synergistic effect of Carboxipterin and Methylene blue applied to antimicrobial photodynamic therapy against mature biofilm of Klebsiella pneumoniae

GASPAR TOSATO, MAIRA; SCHILARDI, P.L; FERNANDEZ LORENZO DE MELE, M; THOMAS, A; LORENTE, C; MIÑAN, A

Encuentro; 14th Encuentro Latinoamericano de Fotoquímica y Fotobiología, XIV ELAFOT; 2019

Multidrug resistance (MDR) bacteria is a serious and frequent health problem that results in the failure of infections treatment. Currently, new strategies for the control of the infections caused by these microorganisms are urgently needed. Notoriously, some MDR microorganisms generate complex structures or biofilms, which adhere to surfaces and confer extraordinary resistance properties that are fundamental challenges to control infections. One of the promising strategies for the control of MDR bacteria is antimicrobial photodynamic therapy that is a minimally invasive technique based on a photochemical process in which a non-toxic photosensitizer (PS) can be activated by low doses of radiation of a specific wavelength [1][2]. The absorbed energy generates radicals on biomolecules and highly reactive oxygen species (ROS), including superoxide anion (O2?-) and hydroxyl radicals (OH?) (Type I reactions) or singlet oxygen (1O2, type II reactions) that cause cytotoxic damage and cell death [3]. The selection of PSs is the key point to get successful results in the aPDT treatment. Habitual PDI treatments use only methylene blue (MB, 3,7-bis (dimethylamino)-phenothiazin-5-ium chloride) that under radiation it shows bactericidal effects against both Gram-positive and Gram-negative bacteria, but MDR microorganism eradication is not completely achieved. In particular, Klebsiella pneumoniae is an opportunistic Gram negative pathogen, responsible of several diseases, such as urinary tract infections, pneumonia, septicemias, and soft tissue infections [4]. Some K. pneumoniae strains are extremely resistant to antibiotic treatments due to their capacity to generate biofilms. The key result of this study revealed that a combination of two known PSs, 6-carboxypterin (Cap, 100 µM) and methylene blue (MB, 2.5-10 µM) exposed to ultraviolet and visible radiation, presents a synergistic effect on the eradication of a MDR Klebsiella pneumoniae strain. Similar effect was observed when the treatment was performed either with planktonic or biofilm growing cells. Moreover, it was found that after treatment the killing action continues in the absence of irradiation leading to the eradication of the microorganisms growing in biofilm.