The Effectiveness of Voriconazole in Therapy of Candida glabrata’s Biofilms Oral Infections and Its Influence on the Matrix Composition and Gene Expression

Abstract

Candida glabrata is one of most prevalent yeast in fungal infections, especially in immunocompromised patients. Its azole resistance results in a low therapeutic response, particularly when associated with biofilms. The main goal of this work was to study the effectiveness of voriconazole (Vcz) against C. glabrata biofilms oral pathologies, as esophageal or oropharyngeal candidiasis. Antifungal susceptibilities were determined in pre-formed 24-h-biofilms and ERG genes expression was determined by qRT-PCR. Protein quantification was performed using BCA Kit, carbohydrate was estimated according to the Dubois assay and β-1,3 glucans concentration were determined using Glucatell kit. Finally, ergosterol, Vcz, and fluconazole (Flu) concentrations within the biofilm matrices were determined by RP-HPLC. Results showed that C. glabrata biofilms were more susceptible to Vcz than to Flu and that ERG genes expression evidenced an overexpression of the three ERG genes in the presence of both azoles. The matrix content presented a remarked decrease in proteins and an increase in carbohydrates, namely β-1,3 glucans. Ergosterol was successfully detected and quantified in the biofilm matrices, with no differences in all the considered conditions. Vcz demonstrated better diffusion through the biofilms and better cell penetration capacities, than Flu, indicating that the structure of the drug molecule fully influences its dissemination through the biofilm matrices. This work showed that Vcz is notably more effective than Flu for the treatment of resistant C. glabrata oral biofilms, which demonstrates a clinical relevance in its future use for the treatment of oropharyngeal/esophageal candidiasis caused by this species.

Liposomal and Deoxycholate Amphotericin B Formulations: Effectiveness against Biofilm Infections of Candida spp.

“…Agar plates were incubated for 24 h at 37 °C, and the total number of CFUs was determined. The results were calculated by Log 10 CFU per cm 2 (Log 10 CFUs/cm 2 ) and presented by mg/L [ 101 ].…”

“…Serial decimal dilutions in PBS were plated on SDA and incubated for 24 h at 37 °C. The results were calculated as a total of CFUs per unit area (Log 10 CFUs/cm 2 ), and presented by mg/L [ 101 ].…”

Susceptibility ofCandida glabratabiofilms to echinocandins: alterations in the matrix composition

“…The biofilm matrices of Candida species have a strong net of exopolymers, providing protection against physical and chemical environmental attack, such as by drugs. These polymers make it difficult for drugs to diffuse into the biofilm cells, which make the biofilms recalcitrant to antifungals (Al-Fattani and Douglas 2004; Rodrigues, Gonçalves et al 2017;Dominguez et al 2018). Hence, the goal of this work was to evaluate seven C. glabrata isolates, by comparison with C. albicans, C. parapsilosis and C. tropicalis, regarding their susceptibility in the planktonic and biofilm form to Csf and Mcf, and the biochemical variations induced in the composition of the matrices after the drug exposure, in order to relate these matrix variations with drug effectiveness.…”

“…Serial decimal dilutions in PBS were plated on SDA and incubated for 24 h at 37°C. The results were presented as total CFUs per unit area (Log 10 CFU cm −2 ) (Rodrigues, Gonçalves et al 2017).…”

“…Extraction method: biofilms were formed in 24-well polystyrene microtitre plates (Orange Scientific) (Rodrigues, Gonçalves et al 2017). For this, 1,000 μl of yeast cell suspension (1 × 10 5 cells ml −1 in RPMI-1640 or RPMI-1640 with Csf and Mcf at concentrations corresponding to the MBEC of each species/strain) were added to each well and the biofilms were treated as described previously (Rodrigues, Gonçalves et al 2017). After 24 h, 500 μl of RPMI-1640 medium were removed and an equal volume of fresh RPMI-1640 with or without the antifungal agents was added.…”