Molecular mechanisms of azole resistance in Candida glabrata isolated from oropharyngeal candidiasis in head and neck cancer patients.

OBJECTIVE: The aim of the current work was to assess the molecular mechanisms of fluconazole-resistant Candida glabrata strains isolated from oropharyngeal candidiasis (OPC) in head and neck patients, as well as evaluation of virulence factors. DESIGN: Antifungal susceptibility pattern of sixty six clinical isolates of C. glabrata were evaluated by broth-microdilution method. The expression of ERG11, CDR1, CDR2, PDR1 genes as well as ERG11 gene capable of possible mutations was also detected in 21 fluconazol-resistant C. glabrata isolates. Phospholipase and proteinase activity of these isolates was estimated, too. The correlation between the virulence factors, antifungal susceptibility patterns and cancer type was also analyzed. RESULTS: Seven synonymous and four non-synonymous mutations were found in 21 fluconazole-resistant C. glabrata isolates; subsequently, four amino acid substitutions including H257P, Q47H, S487Y and I285N were then reported for the first time. High expression of CDR1 and PDR1 in related to other gene findings were tested in these isolates. Additionally, there was no significant difference between stage of cancer and MIC of all antimicrobial drugs. Significant differences between MIC of fluconazole, voriconazole and cancer types were also, found. The proteinase activity (92.4%) was higher than phospholipase activity in the isolates. Further, no significant difference between proteinase (rs: 0.003), phospholipase (rs: -0.107) activity and fluconazole MICs was observed. CONCLUSION: C. glabrata isolated from OPC in head and neck patients represented high capacities for proteolytic enzymes activity and high mRNA level of CDR1 and PDR1 gene and ERG11 mutations play an important role in azole drug resistance.

Shortening the sulfur cell cycle by a green approach for bio-production of extracellular metalloid-sulfide nanoparticles.

In the present study, a new approach was introduced regarding the extracellular synthesis of selenium sulfide micro/nano-particles using Saccharomyces cerevisiae in different ammonium sulfate supplementation and in the presence of sodium selenosulfate precursors (S1) and a blend of selenous acid and sodium sulfite (S2). In S1, only cell supernatant exposed to ammonium sulfate was able to reduce sodium selenosulfate. Whereas, in S2, cell supernatant in both pre-conditions of with or without ammonium sulfate (S2 + or S2-) were able to reduce selenous acid and sodium sulfite. Electron microscopy, also indicated that selenium sulfide NPs were successfully synthesized with average size of 288 and 332 nm for S2 + and S2- in SEM and 268 and 305 nm in TEM. Additionally, elemental mapping by energy-dispersive x-ray analysis confirmed the presence of sulfur/selenium elements in the particles in a proportion of 24.50 and 23.31 for S2- and S2 + , respectively. The mass spectrometry indicated the probability of Se2S2, SeS1.1, Se2, Se, SeS5, SeS3, Se3S5/Se5, Se3/SeS5, Se6, Se4/SeS7, Se2.57S5.43/Se2S2 and Se4S/Se2S6 molecules for S2 + and of Se, Se2, Se3S5/Se5, Se6 and Se4 species for S2-. In FTIR spectra, primary (i.e. 1090-1020 and 1650-1580 cm-1) and secondary (1580-1490 cm-1) amine bands duly confirmed the protein corona around the NPs.

Population Kinetics and Mechanistic Aspects of Saccharomyces cerevisiae Growth in Relation to Selenium Sulfide Nanoparticle Synthesis.

Biosynthesis of nanoparticles (NPs) by microorganisms is a cost- and energy-effective approach. However, how the production of NPs affects the population of producing organism remains as an unresolved question. The present study aimed to evaluate the kinetics of Saccharomyces cerevisiae growth in relation to synthesis of selenium sulfide nanoparticles by using a population model. To this end, the population of S. cerevisiae cells was investigated in terms of colony forming units (CFU) in the presence of the substrate in different time points. Fluctuation of sulfite reductase (SiR) activity, expression of MET5 and MET10 genes, and concentrations of sulfite and selenium were evaluated to support the population findings. CFU values in the test groups were lower than those in the control counterparts. The rise and fall of the SiR activity and MET5 and MET10 gene expression conformed to the variations of CFU values. The rate of reduction in the selenium and sulfite concentrations tended to decrease over the time. In conclusion, the cells population was negatively and positively affected by selenium and sulfite concentrations, respectively. The indirect relationship of the selenium ions concentration in the path analysis revealed that the product, selenium sulfide nanoparticles, caused this drop in S. cerevisiae cells population.

Physicochemical properties, antifungal activity and cytotoxicity of selenium sulfide nanoparticles green synthesized by Saccharomyces cerevisiae.

Selenium sulfide is a well-known bioactive chemical whose biosynthesis as a nanoparticle (NP) is a controversial issue. In the present study, we employed Saccharomyces cerevisiae to generate a novel synthetic process of selenium sulfide NPs. The addition of selenium/sulfur precursors to S. cerevisiae culture produced NPs, which we isolated and characterized the physicochemical properties, toxicity, and antifungal activity. Transmission electron microscopy indicated the presence of the NPs inside the cells. Selenium sulfide NPs were successfully synthesized with average size of 6.0 and 153 nm with scanning electron micrographs and 360 and 289 nm in Zeta sizer using different precursors. The presence of sulfur/selenium in the particles was confirmed by energy-dispersive X-ray spectroscopy and elemental mapping. Fourier-transform infrared spectroscopy supported the production of selenium sulfide NPs. X-ray diffractograms showed the presence of characteristic peaks of selenium sulfide NPs which were further confirmed by mass spectrometry. The obtained NPs strongly inhibited the growth of pathogenic fungi that belonged to the genera Aspergillus, Candida, Alternaria and the dermatophytes, while no cytotoxicity was observed in MTT assay. In conclusion, efficient green synthesis of selenium sulfide NPs with appropriate physicochemical properties is possible in bio-systems like S. cerevisiae.

α-Bisabolol inhibits Aspergillus fumigatus Af239 growth via affecting microsomal ∆24-sterol methyltransferase as a crucial enzyme in ergosterol biosynthesis pathway.

Finding new compounds with antifungal properties is an important task due to the side effects of common antifungal drugs and emerging antifungal resistance in fungal strains. ∆24-sterol methyltransferase (24-SMT) is a crucial enzyme that plays important roles in fungal ergosterol biosynthesis pathway and is not found in humans. In the present study, the effects of α-bisabolol on Aspergillus fumigatus Af239 growth and ergosterol synthesis on the base of 24-SMT enzyme activity were studied; in addition, the expression of erg6, the gene encoded 24-SMT, was considered. To our knowledge, this is the first report demonstrating that α-bisabolol inhibits A. fumigatus growth specifically via suppressing fungal 24-SMT. Since this enzyme is a specific fungal enzyme not reported to exist in mammalian cells, α-bisabolol may serve as a lead compound in the development of new antifungal drugs. Fungi were cultured in presence of serial concentrations of α-bisabolol (0.281-9 mM) for 3 days at 35 °C. Mycelia dry weight was determined as an index of fungal growth and ergosterol content was assessed. Microsomal 24-SMT activity was assayed in presence of α-bisabolol as an inhibitor, lanosterol as a substrate and [methyl-H3] AdoMet (S-Adenosyl methionin). In addition, the expression of erg 6 gene (24-SMT encoding gene) was determined after treatments with various concentrations of α-bisabolol. Our results demonstrated that α-bisabolol strongly inhibited A. fumigatus growth (35.53-77.17%) and ergosterol synthesis (26.31-73.77%) dose-dependently and suppressed the expression of erg 6 gene by 76.14% at the highest concentration of 9 mM. α-bisabolol inhibited the activity of 24-SMT by 99% at the concentration of 5 mM. Taken together, these results provides an evidence for the first time that α-bisabolol inhibits A. fumigatus Af239 growth via affecting microsomal ∆24-sterol methyltransferase as a crucial enzyme in ergosterol biosynthetic pathway.