Phytochemical Analysis and Antimicrobial Activity of Myrcia tomentosa (Aubl.) DC. Leaves.

This work describes the isolation and structural elucidation of compounds from the leaves of Myrcia tomentosa (Aubl.) DC. (goiaba-brava) and evaluates the antimicrobial activity of the crude extract, fractions and isolated compounds against bacteria and fungi. Column chromatography was used to fractionate and purify the extract of the M. tomentosa leaves and the chemical structures of the compounds were determined using spectroscopic techniques. The antibacterial and antifungal activities were assessed using the broth microdilution method. The phytochemical investigation isolated 11 compounds: α-bisabolol, α-bisabolol oxide B, α-cadinol, ß-sitosterol, n-pentacosane, n-tetracosane, quercetin, kaempferol, avicularin, juglanin and guaijaverin. The crude ethanolic extract and its fractions were tested against 15 bacteria and 9 yeasts. The crude extract inhibited the in vitro growth of yeasts at concentration of 4 to 32 µg/mL. The hexane, dichloromethane, ethyl acetate and aqueous fractions inhibited Candida sp. at concentrations of 4 to 256 µg/mL, whereas the Cryptococcus sp. isolates were inhibited only by the hexane and dichloromethane fractions in minimal inhibitory concentrations (MICs) at 16 to 64 µg/mL. The flavonoid quercetin-3-O-α-arabinofuranose (avicularin) was the most active compound, inhibiting Candida species in concentrations of 2 to 32 µg/mL. The MIC values suggest potential activity of this plant species against yeast.

Drug to genome to drug: a computational large-scale chemogenomics screening for novel drug candidates against sporotrichosis.

Sporotrichosis is recognized as the predominant subcutaneous mycosis in South America, attributed to pathogenic species within the Sporothrix genus. Notably, in Brazil, Sporothrix brasiliensis emerges as the principal species, exhibiting significant sapronotic, zoonotic and enzootic epidemic potential. Consequently, the discovery of novel therapeutic agents for the treatment of sporotrichosis is imperative. The present study is dedicated to the repositioning of pharmaceuticals for sporotrichosis therapy. To achieve this goal, we designed a pipeline with the following steps: (a) compilation and preparation of Sporothrix genome data; (b) identification of orthologous proteins among the species; (c) identification of homologous proteins in publicly available drug-target databases; (d) selection of Sporothrix essential targets using validated genes from Saccharomyces cerevisiae; (e) molecular modeling studies; and (f) experimental validation of selected candidates. Based on this approach, we were able to prioritize eight drugs for in vitro experimental validation. Among the evaluated compounds, everolimus and bifonazole demonstrated minimum inhibitory concentration (MIC) values of 0.5 µg/mL and 4.0 µg/mL, respectively. Subsequently, molecular docking studies suggest that bifonazole and everolimus may target specific proteins within S. brasiliensis- namely, sterol 14-α-demethylase and serine/threonine-protein kinase TOR, respectively. These findings shed light on the potential binding affinities and binding modes of bifonazole and everolimus with their probable targets, providing a preliminary understanding of the antifungal mechanism of action of these compounds. In conclusion, our research advances the understanding of the therapeutic potential of bifonazole and everolimus, supporting their further investigation as antifungal agents for sporotrichosis in prospective hit-to-lead and preclinical investigations.

Antifungal Activity of the Essential Oils of Callitris neocaledonica and C. sulcata Heartwood (Cupressaceae).

Mortality due to fungal infections has increased substantially, becoming a worldwide problem in public health. As a contribution to the discovery of new antifungal agents, the properties of the heartwood essential oils of two trees growing in New Caledonia, Callitris neocaledonica and C. sulcata (Cupressaceae) were investigated. The essential oils extracted by hydrodistillation were characterized by GC-FID and GC/MS analyses. From C. neocaledonica oil, 31 constituents were identified, representing 97.0% of the total oil composition, which was mainly constituted by oxygenated sesquiterpenes (88%). Among them, guaiol (1; 30.2%), bulnesol (2; 12.5%), α-eudesmol (3; 10.5%), ß-eudesmol (4; 10.5%), γ-eudesmol (10.2%), and elemol (4.9%) predominated. The chemical composition of C. sulcata oil, from which 39 constituents were identified (96.8% of the total oil composition), showed some similarities with that of C. neocaledonica oil. The major constituents were also oxygenated sesquiterpenes, accounting for 78.5% of the oil, amongst them, mainly compounds 1 (16.1%), 3 and 4 (9.7% each), as well as 2 (7.4%). The antifungal activity of the oils against clinical isolates of four dermatophytic fungi (Trichophyton mentagrophytes, T. rubrum, Microsporum canis, and M. gypseum) and six yeasts (Candida albicans, C. parapsilosis, C. glabrata, C. krusei, Cryptococcus neoformans, and Cryptococcus gattii) was tested by determining minimum inhibitory concentrations (MICs) using the microdilution method. The best antifungal activities of the C. neocaledonica and C. sulcata oils were obtained against C. krusei (MICs of 3.9 and 0.975 µg/ml, resp.). These MIC values were similar to those of the reference drugs itraconazole and fluconazole (1.0 and 0.5 mg/ml, resp.). The oils were also subjected to a screening for their possible DPPH(.) (2,2-diphenyl-1-picrylhydrazyl) radical-scavenging activity. C. neocaledonica essential oil was more active than C. sulcata oil (93.3 vs. 32.2% DPPH(.) scavenged at 250 µg/ml).

Effects of Pimenta pseudocaryophyllus (Gomes) L. R. Landrum, on melanized and non-melanized Cryptococcus neoformans.

In the present study, the in vitro susceptibility and capsular width from both melanized and non-melanized Cryptococcus neoformans cells in the presence of Pimenta pseudocaryophyllus crude extract were determined. The results were compared with those obtained for voriconazole and amphotericin B. Melanization was obtained in minimal medium broth with the addition of L-dopa, and the antifungal susceptibility tests were performed using the broth microdilution method. Capsular width of 30 cells of each one of the isolates in medium with crude extracts of P. pseudocaryophyllus or voriconazole or amphotericin B at a concentration corresponding to 0.5 times the minimal inhibitory concentration (MIC) was measured, and the mean was calculated. The MICs and minimal fungicidal concentrations (MFCs) for plant extract and voriconazole were identical for both melanized and non-melanized C. neoformans isolates, but for amphotericin, the MFCs for melanized cells were up to 8 times higher than for non-melanized cells. The capsular width of C. neoformans cells was smaller (p < 0.001) in the presence crude extract of P. pseudocaryophyllus and of voriconazole regardless melanization. The findings of capsule alterations of C. neoformans verified in this study provide fertile ways for future research into the effects of antifungal agents on the pathogenesis of cryptococcosis.

Effects of antifungal agents in sap activity of Candida albicans isolates.

Some antifungal agents have shown to exert effects on expression of virulent factors of Candida as the production of secretory aspartyl proteinase (Sap). In this study, we sought to determine and to compare the influence of fluconazole and voriconazole in proteinase activity of this microorganism. Thirty-one isolates obtained from oral mucosa of human immunodeficiency virus positive (HIV) patients were used in this study. The minimal inhibitory concentrations (MIC) of fluconazole and voriconazole were determined using the broth microdilution method with RPMI 1640 medium and with yeast carbon base-bovine serum albumin (YCB-BSA) medium. The Sap activity following by digestion of BSA as substrate was determined for four Candida albicans strains arbitrarily chosen according to susceptibility (susceptible or resistant) to fluconazole or voriconazole. Besides, the SAP1 to SAP7 genes were screened by PCR for the same isolates that were determined by the Sap activity. In vitro susceptibility testing using the two media presented similar MIC values. Increased Sap activity was observed in resistant isolates on presence of drugs, but the Sap activity by susceptible isolates to azoles showed different behavior on the presence of drug. We detected the presence of SAP1 to SAP7 genes from all susceptible or resistant C. albicans isolates. The present study provides important data about the proteinase activity and the presence of genes of SAP family in fluconazole and voriconazole susceptible or resistant C. albicans isolates.

Antifungal activities of azole agents against the Malassezia species.

In this paper, we identified 95 Malassezia isolates by morphological and biochemical criteria and assessed the in vitro activity of fluconazole, itraconazole, ketoconazole and voriconazole by broth microdilution against these species using slightly modified Leeming-Notman medium. The Malassezia isolates were identified as M. furfur (74), M. sympodialis (11), M. obtusa (8) and M. globosa (2). The modified Leeming-Notman medium used for susceptibility testing allowed good growth of Malassezia spp. Visual reading of the minimal inhibitory concentration (MIC) was readily achieved until Day 5 of incubation at 32 degrees C. Although high MIC values of 16 microg/mL for fluconazole were observed in 9.5% of Malassezia isolates, in general these microorganisms were susceptible to all drugs studied. Interestingly, one M. globosa isolate showed high MIC values for voriconazole, itraconazole and fluconazole. For the 95 strains, the MIC ranges were <0.03-4 microg/mL for ketoconazole, <0.03 to >16 microg/mL for voriconazole, <0.125 to >64 microg/mL for fluconazole and <0.03-16 microg/mL for itraconazole. In summary, the good reproducibility and visual readings obtained using modified Leeming-Notman medium suggest that this medium should be proposed for antifungal testing of drugs against Malassezia spp.

Asymptomatic oral carriage of Candida species in HIV-infected patients in the highly active antiretroviral therapy era.

Oropharyngeal candidiasis is the most common opportunistic fungal infection in individuals infected with human immunodeficiency virus. CD4+ lymphocytes count and the quantification of viral RNA in blood plasma have been found to be the main markers of HIV disease progression. The present study was conducted to evaluate Candida sp. diversity in the oral cavity of HIV-infected patients and to determine whether there was association of CD4+ cell count and viral load with asymptomatic oral Candida carriage. Out of 99 HIV-positive patients studied, 62 (62.6%) had positive culture for Candida (oral carriage) and 37 patients (37.4%) had Candida negative culture (no oral carriage). The etiologic agents most common were C. albicans and C. tropicalis. The range of CD4+ was 6-2305 cells/mm3 in colonized patients and 3-839 cells/mm3 for non-colonized patients, while the viral load was 60-90016 copies/mL for colonized patients and 75-110488 copies/mL for non colonized patients. The viral load was undetectable in 15 colonized patients and in 12 non colonized patients. Our results showed that there was no significant difference of the variables CD4+ cell count and viral load between oral candida carriage and no oral candida carriage patients.

[Identification of Malassezia species in patients with pityriasis versicolor in Goiânia-GO]. / Identificação de espécies de Malassezia em pacientes com pitiríase versicolor em Goiânia-GO.

The objectives of this study were to investigate the frequency of pityriasis versicolor occurrence and to identify yeasts of the genus Malassezia in patients at the mycology laboratory of the Federal University of Goiás, in Goiânia, State of Goiás. Ninety-five cases of pityriasis versicolor were diagnosed, and four species of Malassezia were identified: Malassezia furfur, Malassezia sympodialis, Malassezia globosa and Malassezia obtusa.