Effects of Growth Phase and Extracellular Slime on Photodynamic Inactivation of Gram-Positive Pathogenic Bacteria

Abstract

The emergence of antibiotic resistance among pathogenic bacteria has led to efforts to find alternative antimicrobial therapeutics to which bacteria will not be easily able to develop resistance. One of these may be the combination of nontoxic dyes (photosensitizers [PS]) and visible light, known as photodynamic therapy, and we have reported its use to treat localized infections in animal models. While it is known that gram-positive species are generally susceptible to photodynamic inactivation (PDI), the factors that govern variation in degrees of killing are unknown. We used isogenic pairs of wild-type and transposon mutants deficient in capsular polysaccharide and slime production generated from Staphylococcus epidermidis and Staphylococcus aureus to examine the effects of extracellular slime on susceptibility to PDI mediated by two cationic PS (a polylysine-chlorin e6 conjugate, pL-c e6 , and methylene blue [MB]) and an anionic molecule, free c e6 , and subsequent exposure to 665-nm light at 0 to 40 J/cm 2 . Free c e6 gave more killing of mutant strains than wild type, despite the latter taking up more PS. Log-phase cultures were killed more than stationary-phase cultures, and this correlated with increased uptake. The cationic pL-c e6 and MB gave similar uptakes and killing despite a 50-fold difference in incubation concentration. Differences in susceptibility between strains and between growth phases observed with free c e6 largely disappeared with the cationic compounds despite significant differences in uptake. These data suggest that slime production and stationary phase can be obstacles against PDI for gram-positive bacteria but that these obstacles can be overcome by using cationic PS.

Inativação fotodinâmica de microrganismos

“…tem sido proposta como uma terapia antibacteriana alternativa 1,17 . A PDI, sendo efetiva contra vírus, bactérias e fungos, pode ser usada como uma terapia para infecções localizadas.…”

“…Portanto, FS positivamente carregados são geralmente mais eficientes em bactérias e agem em concentrações menores que as moléculas de FS neutras e aniônicas 1 .…”

“…A Terapia Fotodinâmica (TFD) baseia-se na administração tópica ou sistêmica de um corante não tóxico sensível à luz seguida da irradiação em baixas doses com luz visível de comprimento de onda adequado 1 . Na presença de oxigênio encontrado nas células, o fotossensibilizador (FS) ativado pode reagir com moléculas na sua vizinhança por transferência de elétrons ou hidrogênio, levando à produção de radicais livres (reação do tipo I) ou por transferência de energia ao oxigênio (reação do tipo II), levando à produção de oxigênio singlete.…”

A comparative study of irradiation systems for photoinactivation of microorganisms

“…Photodynamic Inactivation (PDI) 1 or Photodynamic Antimicrobial Chemotherapy (PACT) 2 is the name of a process, derived from Photodynamic Therapy, that causes the death of microorganisms under the action of visible light and a specific drug, also called photosensitizer. 3 The electronic excitation of the photosensitizer in the presence of oxygen implies in the generation of a large number of reactive oxygen species (ROS), as for example, singlet oxygen, 4,5 in a process known as photosensitization.…”

In vitro reduction of pathogenic sporothrix schenckii fungus by photodynamic therapy

“…The process of photodynamic therapy is based on the topical or systemic administration of a non-toxic photosensitizer, followed by low-dose irradiation with visible light and an adequate wavelength 13 .…”