ABSTRACT
Photodynamic inactivation (PDI) or antibacterial photodynamic therapy (aPDT) is a method based on the use of a photosensitizer, light of a proper wavelength and oxygen, which combined together leads to an oxidative stress and killing of target cells. PDI can be applied towards various pathogenic bacteria independently on their antibiotic resistance profile. Optimization of photodynamic treatment to eradicate the widest range of human pathogens remains challenging despite the availability of numerous photosensitizing compounds. Therefore, a search for molecules that could act as adjuvants potentiating antibacterial photoinactivation is of high scientific and clinical importance. Here we propose farnesol (FRN), a well described sesquiterpene, as a potent adjuvant of PDI, which specifically sensitizes Staphylococcus aureus to 5,10,15,20-tetrakis(1-methylpyridinium-4-yl)porphyrin tetratosylate (TMPyP) upon red light irradiation. Interestingly, the observed potentiation strongly depends on the presence of light. Analysis of this combined action of FRN and TMPyP, however, showed no influence of farnesol on TMPyP photochemical properties, i.e. the amount of reactive oxygen species that were produced by TMPyP in the presence of FRN. The accumulation rate of TMPyP in Staphylococcus aureus cells did not change, as well as the influence of staphyloxanthin inhibition. The precise mechanism of observed sensitization is unclear and probably involves specific molecular targets.
ABSTRACT
Photoantimicrobial chemotherapy (PACT) constitutes a particular type of stress condition, in which bacterial cells induce a pleiotropic and as yet unexplored effect. In light of this, the key master regulators are of putative significance to the overall phototoxic outcome. In Staphylococcus aureus, the alternative sigma factor σ(B) controls the expression of genes involved in the response to environmental stress. We show that aberration of any sigB operon genes in S. aureus USA300 isogenic mutants causes a pronounced sensitization (>5 log10 reduction in CFU drop) to PACT with selected photosensitizers, namely protoporphyrin diarginate, zinc phthalocyanine and rose bengal. This effect is partly due to aberration-coupled staphyloxanthin synthesis inhibition. We identified frequent mutations in RsbU, a σ(B) activator, in PACT-vulnerable clinical isolates of S. aureus, resulting in σ(B) activity impairment. Locations of significant changes in protein structure (IS256 insertion, early STOP codon occurrence, substitutions A230T and A276D) were shown in a theoretical model of S. aureus RsbU. As a phenotypic hallmark of PACT-vulnerable S. aureus strains, we observed an increased fluidity of bacterial cell membrane, which is a result of staphyloxanthin content and other yet unidentified factors. Our research indicates σ(B) as a promising target of adjunctive antimicrobial therapy and suggests that enhanced cell membrane fluidity may be an adjuvant strategy in PACT.
ABSTRACT
Light- and photosensitiser-based antimicrobial photodynamic therapy is a very promising approach to the control of microbial infections. How the phenotypic features of a microorganism affect its response to photosensitiser-based photokilling represents an area of substantial research interest. To understand the mechanisms governing the phenomenon of a strain-dependent response to photodynamic inactivation (PDI), we analysed the possible role of the membrane-located haem transporter HrtA in Staphylococcus aureus. We used a S. aureus strains with an inactivated component of the haem-regulated transporter, HrtA, along with its wild-type counterpart to determine differences in PDI outcome and photosensitiser uptake between the studied isogenic strains. We observed that a lack of HrtA protein potentiates the phototoxic effect towards S. aureus but only when extracellular protoporphyrin IX is used. The observed effect may depend on the function of the HrtA transporter but is likely to result from changed membrane properties following the absence of the protein in the membrane. This indicates that disturbing the membrane properties is an attractive method for improving the efficacy of the photodynamic inactivation of microorganisms.
