The inhibitory effect of photodynamic therapy on dual-species biofilms of Candida albicans and Candida krusei can be determined by Candida albicans/Candida krusei ratio.

Candida krusei and Candida albicans present the ability to form communities of microorganisms called biofilms. Biofilms can be composed of a single species or more and are an important virulence factor. The inhibition of C. albicans and C. krusei as well as of their dual-species biofilms by antimicrobial Photodynamic Therapy (aPDT) has been demonstrated. This study aimed to investigate the effect of aPDT, with TBO, on dual-species biofilms of C. albicans and C. krusei using different culture mediums, RPMI-1640 and Sabouraud-dextrose broth (SDB) to produce biofilms presenting different C. albicans/C. krusei ratio. Biofilms formed using RPMI-1640 presented a higher C. albicans/C. krusei ratio, however, biofilms formed using SDB presented a predominance of C. krusei. The metabolic activity of biofilms produced using RPMI-1640 was inhibited by aP (∼40%), while biofilms produced using SDB were not affected by aPDT. In addition, biofilm biomass was reduced in biofilms produced using RPMI-1640 and treated with aPDT (∼20%). The results demonstrated that aPDT reduces C. albicans development in dual-species biofilms with C. krusei. However, no effect could be observed on C. krusei, demonstrating that C. krusei, when present in the structure of dual-species biofilms can be resistant to aPDT.

Effect of antimicrobial photodynamic therapy using toluidine blue on dual-species biofilms of Candida albicans and Candida krusei.

BACKGROUND: Although Candida albicans is the most frequent etiological agent of candidiasis, it has been reported a sizable number of infections related to the non-albicans Candida (NAC) species, Candida krusei. In addition, dual biofilms (biofilms composed by two species) may easily occur in vivo, becoming even more challenging the treatment of an infection. The fungicide effect of Photodynamic Therapy (PDT), using toluidine blue O (TBO) on both C. albicans and C. krusei development has been demonstrated. Thus, the objective of this study was to investigate the effects of PDT on dual-species biofilms of Candida albicans and Candida krusei. METHODS: The effect of PDT was observed on the metabolic activity of mature dual-species biofilms of Candida albicans and Candida krusei by a metabolic assay based on the reduction of XTT (2,3-bis(2­methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide sodium salt) assay and the identification of Candida albicans and Candida krusei was performed on CHROMagar Candida medium. RESULTS: it was observed a reduction of ∼30% in the metabolic activity of a mature biofilm treated with PDT, using 0.05 mg·mL-1 TBO and during biofilm formation a predominance of C. albicans on C. krusei was observed. The inhibition observed was related to reduction in the number of Colony Forming Units (CFU) of Candida albicans from 31.33 ± 3.7 to 17.0 ± 1.5. The number of CFU of C. krusei was not significantly modified. CONCLUSIONS: These results demonstrated the efficiency of PDT in inhibiting the dual-species biofilms of Candida albicans and Candida krusei by reducing C. albicans development.

Diphenyl diselenide suppresses key virulence factors of Candida krusei, a neglected fungal pathogen.

Candida krusei is a candidiasis etiological agent of relevance in the clinical setting because of its intrinsic resistance to fluconazole. Also, it has opened up new paths in the area of alternative therapeutic techniques. This project demonstrated the effects of diphenyl diselenide (PhSe)2 and p-cloro diphenyl diselenide (pCl-PhSe)2, two organochalcogen compounds, on relevant virulence factors for the early stage of the C. krusei host interaction and infection process. Both compounds inhibited adherence of C. krusei to both polystyrene surfaces and cervical epithelial cells and biofilm formation; the structure of the biofilm was also changed in a dose-dependent manner. In addition, both compounds inhibited C. krusei growth, but (PhSe)2 significantly increased the time duration of the lag phase and delayed the start of the exponential phase in growth kinetics. (PhSe)2 has more potential antifungal activity than (pCl-PhSe)2 in inhibiting the adherence to epithelial cells, biofilm formation, and growth of C. krusei.

(MeOPhSe)2, a synthetic organic selenium compound, inhibits virulence factors of Candida krusei: Adherence to cervical epithelial cells and biofilm formation.

BACKGROUND: Systemic candidiasis is produced by Candida albicans or non-albicans Candida species, opportunistic fungi that produce both superficial and invasive infections. Despite the availability of a wide range of antifungal agents for the treatment of candidiasis, failure of therapy is observed frequently, which opens new avenues in the field of alternative therapeutic strategies. METHODS: The effects of p,p'-methoxyl-diphenyl diselenide [(MeOPhSe)2], a synthetic organic selenium (organochalcogen) compound, were investigated on virulence factors of C. krusei and compared with its antifungal effects on the virulence factors related to adhesion to cervical epithelial cell surfaces with C. albicans. RESULTS: (MeOPhSe)2, a compound non-toxic in epithelial (HeLa) and fibroblastic (Vero) cells, inhibited the growth in a dose-dependent manner and changed the kinetics parameters of C. krusei and, most importantly, extending the duration of lag phase of growth, inhibiting biofilm formation, and changing the structure of biofilm. Also, (MeOPhSe)2 reduced C. albicans and C. krusei adherence to cervical epithelial cells, an important factor for the early stage of the Candida-host interaction. The reduction was 37.24 ± 2.7 % in C. krusei (p = 0.00153) and 32.84 ± 3.2 % in C. albicans (p = 0.0072) at 20 µM (MeOPhSe)2, and the effect is in a concentration-dependent manner. Surprisingly, the antifungal potential on adhesion was similar between both species, indicating the potential of (MeOPhSe)2 as a promising antifungal drug against different Candida infections. CONCLUSION: Overall, we demonstrated the potential of (MeOPhSe)2 as an effective antifungal drug against the virulence factors of Candida species.

BSA nanoparticles loaded-methylene blue for photodynamic antimicrobial chemotherapy (PACT): effect on both growth and biofilm formation by Candida albicans.

It has been demonstrated an increase in resistance of Candida albicans to conventional therapies, probably, due the indiscriminate use of the conventional antifungal drugs. In this aspect, the nanotechnology generates the possibility of creating new therapeutic agents. Thus, the objective of this paper was to produce and characterize a bovine serum albumin (BSA) nanoparticle encapsulated with Methylene Blue (MB). In addition, the effect of BSA nanoparticles encapsulated with MB (BSA-MB) was evaluated on both growth and biofilm formation by C. albicans by Photodynamic Antimicrobial Chemotherapy (PACT) protocols. The BSA-MB nanoparticles were prepared by the desolvation process. The nanoparticulate system was studied by steady-state techniques, scanning electron microscopy and their biological activity was evaluated in vitro both growth and biofilm formation by C. albicans. The synthetized BSA-MB nanoparticles were spherical in shape exhibiting a 100-200 nm diameter with a low tendency to aggregate (PDI values < 0.2). MB photophysical properties were shown to be preserved after BSA encapsulation. A significant reduction in C. albicans growth, after PACT was observed, in a dependent manner on MB-loaded in BSA nanoparticles concentration used. It was observed an inhibition of 23, 65 and 83% in the presence of MB-loaded in BSA nanoparticles 0.1, 0.5 and 1.0 µg.mL-1, respectively. In addition, MB-loaded BSA nanoparticles 0.5 µg.mL-1 were able to reduce both biofilm formation (80%) and the transition from yeast to filamentous form by C. albicans. The results presented here demonstrated a potentiation of the phototoxic effect of MB after BSA encapsulation, since the concentrations of MB-loaded BSA nanoparticles necessary to inhibits ∼50% of C. albicans development was 10 times minor than that observed for free MB. Taken together, these results suggest the potential of PACT, using MB-loaded BSA nanoparticles in inhibiting C. albicans development. The synthesis and design of BSA nanoparticles can be successfully applied for MB encapsulation and offer the possibility to drive the toxicity effect to a specific target, as an evaluation on both growth and biofilm formation by Candida albicans.