Production of 2-phenethyl alcohol and 2-phenyllactic acid in Candida species

2-Phenethyl alcohol (2-PEA) and 2-phenyllactic acid (2-PLA) were isolated from the culture filtrates of Candida species grown in media containing peptone or phenylalanine as nitrogen source. These compounds were characterized by comparing their UV, IR, and NMR spectral properties with authentic samples. Candida species differed markedly in their production of 2-PEA and 2-PLA. Experiments using [14C]-phenylalanine indicated that both 2-PEA and 2-PLA are synthesised from L-phenylalanine. A pathway for the biosynthesis of 2-PEA from L-phenylalanine has been proposed.

Stable hybrid containing haploid genomes of two obligate diploid Candida species

Candida albicans and Candida dubliniensis are diploid, predominantly asexual human-pathogenic yeasts. In this study, we constructed tetraploid (4n) strains of C. albicans of the same or different lineages by spheroplast fusion. Induction of chromosome loss in the tetraploid C. albicans generated diploid or near-diploid progeny strains but did not produce any haploid progeny. We also constructed stable heterotetraploid somatic hybrid strains (2n + 2n) of C. albicans and C. dubliniensis by spheroplast fusion. Heterodiploid (n + n) progeny hybrids were obtained after inducing chromosome loss in a stable heterotetraploid hybrid. To identify a subset of hybrid heterodiploid progeny strains carrying at least one copy of all chromosomes of both species, unique centromere sequences of various chromosomes of each species were used as markers in PCR analysis. The reduction of chromosome content was confirmed by a comparative genome hybridization (CGH) assay. The hybrid strains were found to be stably propagated. Chromatin immunoprecipitation (ChIP) assays with antibodies against centromere-specific histones (C. albicans Cse4/C. dubliniensis Cse4) revealed that the centromere identity of chromosomes of each species is maintained in the hybrid genomes of the heterotetraploid and heterodiploid strains. Thus, our results suggest that the diploid genome content is not obligatory for the survival of either C. albicans or C. dubliniensis. In keeping with the recent discovery of the existence of haploid C. albicans strains, the heterodiploid strains of our study can be excellent tools for further species-specific genome elimination, yielding true haploid progeny of C. albicans or C. dubliniensis in future.

In vitro activities of natural products against oral Candida isolates from denture wearers

Background: Candida-associated denture stomatitis is a frequent infectious disease. Treatment of this oral condition is difficult because failures and recurrences are common. The aim of this study was to test the in vitro antifungal activity of pure constituents of essentials oils. -- Methods: Eight terpenic derivatives (carvacrol, farnesol, geraniol, linalool, menthol, menthone, terpinen-4-ol, and aterpineol), a phenylpropanoid (eugenol), a phenethyl alcohol (tyrosol) and fluconazole were evaluated against 38 Candida isolated from denture-wearers and 10 collection Candida strains by the CLSI M27-A3 broth microdilution method. -- Results: Almost all the tested compounds showed antifungal activity with MIC ranges of 0.03-0.25% for eugenol and linalool, 0.03-0.12% for geraniol, 0.06-0.5% for menthol, a-terpineol and terpinen-4-ol, 0.03-0.5% for carvacrol, and 0.06-4% for menthone. These compounds, with the exception of farnesol, menthone and tyrosol, showed important in vitro activities against the fluconazole-resistant and susceptible-dose dependent Candida isolates. -- Conclusions: Carvacrol, eugenol, geraniol, linalool and terpinen-4-ol were very active in vitro against oral Candida isolates. Their fungistatic and fungicidal activities might convert them into promising alternatives for the topic treatment of oral candidiasis and denture stomatitis.

Prior colonisation with Candida species fails to guide empirical therapy for candidaemia in critically ill adults

Objectives Pre-emptive fluconazole (fcz) anti-fungal therapy is often based upon Candida colonisation of at least 2 non-contiguous non-sterile sites. The aim of this study was to evaluate the relationship between candidaemia and prior colonisation of non-sterile sites. Methods A retrospective observational study was performed in the intensive care unit/high dependency unit (ICU/HDU) of a University hospital on alternate years from 1999–2007, where a pre-emptive anti-fungal therapy policy was introduced in 2005. Results A higher proportion of blood isolates were Candida glabrata compared with non-sterile isolates (16/46 vs 106/1062; p

Correlation Between Etest (R), Disk Diffusion, and Microdilution Methods for Antifungal Susceptibility Testing of Candida Species From Infection and Colonization

The correlation between the microdilution (MD), Etest (R) (ET), and disk diffusion (DD) methods was determined for amphotericin B, itraconazole and fluconazole. The minimal inhibitory concentration (MIC) of those antifungal agents was established for a total of 70 Candida spp. isolates from colonization and infection. The species distribution was: Candida albicans (n = 27), C. tropicalis (n = 17), C. glabrata (n = 16), C. parapsilosis (n = 8), and C. lusitaniae (n = 2). Non-Candida albicans Candida species showed higher MICs for the three antifungal agents when compared with C. albicans isolates. The overall concordance (based on the MIC value obtained within two dilutions) between the ET and the MD method was 83% for amphotericin B, 63% for itraconazole, and 64% for fluconazole. Considering the breakpoint, the agreement between the DD and MD methods was 71% for itraconazole and 67% for fluconazole. The DD zone diameters are highly reproducible and correlate well with the MD method, making agar-based methods a viable alternative to MD for susceptibility testing. However, data on agar-based tests for itraconazole and amphotericin B are yet scarce. Thus, further research must still be carded out to ensure the standardization to other antifungal agents. J. Clin. Lab. Anal. 23:324-330, 2009. (C) 2009 Wiley-Liss, Inc.

Antifungal activity of tri- and tetra-thioureido amino derivatives against different Candida species

The in vitro antifungal activity of six thioureido substituted amines (P1-P6) was evaluated against Candida species, including Candida albicans, C. glabrata, C. krusei and C. parapsilosis. These tri- and tetra-thioureido amino derivatives with different methylation levels were synthesised through easy synthetic routes to evaluate their antifungal properties against Candida species. Among all studied derivatives, the tri-(2-thioureido-ethyl)-amine (P1) was the most active compound inhibiting C. albicans and C. glabrata at a concentration of 0.49 mu g ml(-1); P3, the N,N `,N ``,N ```-hexamethyl-derivative, also showed inhibitory activity against C. albicans and C. glabrata, but in higher concentrations (250 mu g ml(-1)). The N,N `,N ``,N ```-tetramethylated amine (P5) only inhibited the growth of C. glabrata, but its corresponding N,N `,N ``,N ```-octamethyl derivative (P6) was also active against C. glabrata (125 mu g ml(-1)) and it was the only compound active against C. parapsilosis. P2 and P4 showed no significant antifungal activity. The structure-activity relationship of the thioureido-substituted derivatives indicates that the molecular branching and the alkylation levels can influence the antifungal activity. This study demonstrated that thioureido derivatives exhibited significant antifungal activity against Candida species and that they can be considered as a very promising bioactive lead compound to develop novel antifungal agents.

Photodynamic inactivation of four Candida species induced by photogem®

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Catalase activity of different Candida species after exposition to specific antiserum

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Electrophoretic variants of intracellular catalase of different Candida species

Intracellular and extracellular catalases of different species of Candida were investigated using different culture media. All the Candida strains produced intracellular catalase, whose enzymatic activity was detected by non-denaturating polyacrylamide gradient (4-30%) gel electrophoresis. The cell extracts presented a major 230 kDa catalase band and in some strains variants of catalase with different molecular weights were detected. Candida catalase activity was not affected by heating at 50degreesC and incubation with beta-mercaptoethanol, but treatment with sodium dodecyl sulphate inhibited or reduced enzymatic activity. Extracellular enzyme activity was not detected in any of the culture filtrate extracts tested.

Studies on hyaluronidase, chondroitin sulphatase, proteinase and phospholipase secreted by Candida species

We studied the ability of different Candida species to produce,at the same time, hyaluronidase, chondroitin sulphatase, proteinase, and phospholipase to assess whether they could be related to Candida pathogenicity. Only C. albicans was able to produce the four enzymes tested (73%) and was highly virulent to mice. Strains, that lack the capacity to produce one or more of the enzymes assayed, seemed less virulent or avirulent, similarly to the spontaneous hyaluronidase, chondroitin sulphatase, phospholipase and proteinase-deficient C. albicans strain FCF 14, 1 which was non-pathogenic to mice. Among the other Candida species tested, none of them produced the four enzymes simultaneously, being less virulent in intravenously inoculated mice.

Photosensitization of different Candida species by low power laser light

The aim of this study was to evaluate the effects of the laser radiation (685 nm) associated with photosensitizers on viability of different species of Candida genus. Suspensions of Candida albicans, Candida dubliniensis, Candida krusei and Candida tropicalis, containing 106 viable cells per milliliter were obtained with the aid of a Neubauer's chamber. From each species, 10 samples of the cell suspension were irradiated with diode laser (685 nm) with 28 J/cm(2) in the presence of methylene blue (0.1 mg/ml), 10 samples were only treated with methylene blue, 10 samples were irradiated with laser in the absence of the dye, 10 samples were treated with the dye and irradiated with laser light and 10 samples were exposed to neither the laser light nor to the methylene blue dye. From each sample, serial dilutions of 10(-2) and 10(-3) were obtained and aliquots of 0.1 ml of each dilution were plated in duplicate on Sabouraud dextrose agar. After incubation at 37 degrees C for 48 h, the number of colony-forming units (CFU/ml) was obtained and data were submitted to ANOVA and Tukey's test (p < 0.05). Laser radiation in the presence of methylene blue reduced the number of CFU/ml in 88.6% for C. albicans, 84.8% for C. dubliniensis, 91.6% for C krusei and 82.3% for C tropicalis. Despite of this, only laser radiation or methylene blue did not reduce significantly the number of CFU/ml of Candida samples, except for C tropicalis. It could be concluded that the photo activation of methylene blue by the red laser radiation at 685 nm presented fungicide effect on all Candida species studied. (c) 2006 Elsevier B.V. All rights reserved.

rDNA-RFLP identification of Candida species in immunocompromised and seriously diseased patients

PCR was used to amplify a targeted region of the ribosomal DNA of 76 Candida spp. isolates from immunocompromised and seriously diseased patients. Thirty-seven strains isolated from different anatomical sites of 11 patients infected with HIV (Vitória, ES, Brazil), 26 isolates from patients under treatment at Odilon Behrens Hospital and 13 isolates from skin and urine samples from São Marcos Clinical Analysis Laboratory (Belo Horizonte, Brazil) were scored. Fragments of rDNA were amplified using primer pairs ITS1-ITS4, for the amplification of ITS1 and ITS2 regions, including the gene for the 5.8s subunit. Amplification resulted in fragments ranging in size from 350 to 950 bp. Amplicons were digested with eight restriction enzymes. A pattern of species-specificity among the different medically important Candida species could be identified following restriction digestion of the PCR products. Candida albicans was the species most frequently observed, except for the group of newborns under treatment at the Odilon Behrens Hospital and for the isolates from the clinical analysis laboratory. C. parapsilosis was the species most frequently observed in these two groups.