Beneficial Effects of HIV Peptidase Inhibitors on Fonsecaea pedrosoi: Promising Compounds to Arrest Key Fungal Biological Processes and Virulence

BACKGROUND: Fonsecaea pedrosoi is the principal etiologic agent of chromoblastomycosis, a fungal disease whose pathogenic events are poorly understood. Current therapy for chromoblastomycosis is suboptimal due to toxicity of the available therapeutic agents and the emergence of drug resistance. Compounding these problems is the fact that endemic countries and regions are economically poor. PURPOSE AND PRINCIPAL FINDINGS: In the present work, we have investigated the effect of human immunodeficiency virus (HIV) peptidase inhibitors (PIs) on the F. pedrosoi conidial secreted peptidase, growth, ultrastructure and interaction with different mammalian cells. All the PIs impaired the acidic conidial-derived peptidase activity in a dose-dependent fashion, in which nelfinavir produced the best inhibitory effect. F. pedrosoi growth was also significantly reduced upon exposure to PIs, especially nelfinavir and saquinavir. PIs treatment caused profound changes in the conidial ultrastructure as shown by transmission electron microscopy, including invaginations in the cytoplasmic membrane, disorder and detachment of the cell wall, enlargement of fungi cytoplasmic vacuoles, and abnormal cell division. The synergistic action on growth ability between nelfinavir and amphotericin B, when both were used at sub-inhibitory concentrations, was also observed. PIs reduced the adhesion and endocytic indexes during the interaction between conidia and epithelial cells (CHO), fibroblasts or macrophages, in a cell type-dependent manner. Moreover, PIs interfered with the conidia into mycelia transformation when in contact with CHO and with the susceptibility killing by macrophage cells. CONCLUSIONS/SIGNIFICANCE: Overall, by providing the first evidence that HIV PIs directly affects F. pedrosoi development and virulence, these data add new insights on the wide-spectrum efficacy of HIV PIs, further arguing for the potential chemotherapeutic targets for aspartyl-type peptidase produced by this human pathogen

Chemical and antimicrobial analysis of husk fiber aqueous extract from Cocos nucifera L.

Cocos nucifera L. (Arecaceae) is a widely distributed species around the tropical areas. Popular uses have been reported in the treatment of arthritis and diarrhea. This study evaluates the antimicrobial activity of husk fiber aqueous extract from C. nucifera and performed the identification of some biological active substances. The minimal inhibitory concentration (MIC) against human pathogen microorganisms was determined. Chromatographic and spectrometric procedures were also performed to isolate and identify the components present in the extract. In the MIC assay of crude aqueous extract, only the methicillin sensible and the resistant (MRSA) Staphylococcus aureus strains were susceptible at 156 μg/mL. The ethyl acetate partition taken from crude extract was more promising (MIC of 78 μg/mL). No fungal growth inhibition was observed. Catechin, epicatechin, two procyanidin dimers and condensed tannins were found in the organic phase. In addition, gallic and ellagic acids were detected for the first time in C. nucifera husk fiber. Gallic acid showed MIC of 39 μg/mL and minimal bactericidal concentration (MBC) at 78 μg/mL. Ellagic acid was not active against the tested strains, as well as catechin and epicatechin. Additionally catechin, epicatechin, two procyanidin dimers and condensed tannins were also detected. The antimicrobial activity observed was selective to S. aureus strains.Keywords: Antimicrobial analyses, Cocos nucifera, arecaceae, ellagic acid, gallic acid, procyanidins, Staphylococcus aureus strainsAfrican Journal of Biotechnology Vol. 12(18), pp. 2478-248

Rhizopus arrhizus ucp1295 como fonte econômica para produção de biopolímeros funcionais quitina e quitosana utilizando substratos renováveis / Rhizopus arrhizus ucp1295 as economic source for production of functional biopolymers chitin and chitosan using renewable substrates

Neste trabalho foi investigada a produção de quitina e quitosana por Rhizopus arrhizus UCP 1295 isolado do solo da Caatinga do Estado de Pernambuco, Brasil, utilizando o efluente industrial de doces e milhocina como substratos de baixo custo, considerando a versatilidade de aplicação das biomoléculas. O micro-organismo foi cultivado em diferentes concentrações dos substratos efluente da indústria de doces e milhocina (CSL) em diferentes valores de pH, de acordo com um planejamento fatorial completo 23. Após 96 h de fermentação, a biomassa produzida foi liofilizada e submetida ao tratamento com álcali- ácido-. Os polissacarídeos extraídos foram caracterizados por espectroscopia por transformada de Fourier (FTIR) na região do infravermelho. A maior produção de biomassa (14,11 g/L) foi obtida na condição 6 (8% de efluente industrial de doces, 5% de milhocina e pH 5), enquanto os maiores rendimentos de quitina (169,3 mg/g) e quitosana (239,1 mg/g) foram obtidos em meio contendo 4% de efluente da indústria de doces, sem milhocina, nas condições 3 (pH 7) e 1 (pH 5), respectivamente. A quitina apresentou grau de acetilação de 71,4% e a quitosana de 86,0%, de desacetilação, respectivamente. Além disso, foi demonstrado que o efluente industrial de balas e milhocina são substratos renováveis e alternativos na formulação de novos meios de produção de quitina e quitosana. A versatilidade das biomoléculas deve-se as suas propriedades bioquímicas únicas, como biocompatibilidade, biodegradabilidade, não toxicidade, capacidade de formar filmes e aplicações industriais promissoras

Does the essential oil of Lippia sidoides Cham. (pepper-rosmarin) affect its endophytic microbial community?

Background: Lippia sidoides Cham., also known as pepper-rosmarin, produces an essential oil in its leaves that is currently used by the pharmaceutical, perfumery and cosmetic industries for its antimicrobial and aromatic properties. Because of the antimicrobial compounds (mainly thymol and carvacrol) found in the essential oil, we believe that the endophytic microorganisms found in L. sidoides are selected to live in different parts of the plant. Results: In this study, the endophytic microbial communities from the stems and leaves of four L. sidoides genotypes were determined using cultivation-dependent and cultivation-independent approaches. In total, 145 endophytic bacterial strains were isolated and further grouped using either ERIC-PCR or BOX-PCR, resulting in 76 groups composed of different genera predominantly belonging to the Gammaproteobacteria. The endophytic microbial diversity was also analyzed by PCR-DGGE using 16S rRNA-based universal and group-specific primers for total bacteria, Alphaproteobacteria, Betaproteobacteria and Actinobacteria and 18S rRNA-based primers for fungi. PCR-DGGE profile analysis and principal component analysis showed that the total bacteria, Alphaproteobacteria, Betaproteobacteria and fungi were influenced not only by the location within the plant (leaf vs. stem) but also by the presence of the main components of the L. sidoides essential oil (thymol and/or carvacrol) in the leaves. However, the same could not be observed within the Actinobacteria. Conclusion: The data presented here are the first step to begin shedding light on the impact of the essential oil in the endophytic microorganisms in pepper-rosmarin

Synergism effect of the essential oil from Ocimum basilicum var. Maria Bonita and its major components with fluconazole and its influence on ergosterol biosynthesis

The aim of this study was to evaluate the activity of the EO and its major components of Ocimum basilicum var. Maria Bonita, a genetically improved cultivar, against the fluconazole sensitive and resistant strains of Candida albicans and Cryptococcus neoformans. Geraniol presented better results than the EO, with a low MIC (76 mg/mL against C. neoformans and 152 mg/mL against both Candida strains). The combination of EO, linalool, or geraniol with fluconazole enhanced their antifungal activity, especially against the resistant strain (MIC reduced to 156, 197, and 38 mg/mL, resp.). The ergosterol assay showed that subinhibitory concentrations of the substances were able to reduce the amount of sterol extracted. The substances tested were able to reduce the capsule size which suggests they have an important mechanism of action. Transmission electron microscopy demonstrated cell wall destruction of C. neoformans after treatment with subinhibitory concentrations. In C. albicans ultrastructure alterations such as irregularities in the membrane, presence of vesicles, and cell wall thickening were observed. The biofilm formation was inhibited in both C. albicans strains at MIC and twice MIC. These results provide further support for the use of O. basilicum EO and its major components as a potential source of antifungal agents

Fonsecaea pedrosoi Sclerotic Cells: Secretion of Aspartic-Type Peptidase and Susceptibility to Peptidase Inhibitors

Fonsecaea pedrosoi is a dematiaceous fungus and the main causative agent of chromoblastomycosis that is a chronic disease usually affecting the human skin and subcutaneous tissues, which causes deformations and incapacities, being frequently refractory to available therapies. A typical globe-shaped, multiseptated and pigmented cells, known as sclerotic cells, are found in the lesions of infected individuals. In the present work, we have investigated the production of aspartic-type peptidase in F. pedrosoi sclerotic cells as well as the effect of peptidase inhibitors (PIs) on its enzymatic activity and viability. Our data showed that sclerotic cells are able to secrete pepstatin A-sensible aspartic peptidase when grown under chemically defined conditions. In addition, aspartic PIs (ritonavir, nelfinavir, indinavir, and saquinavir), which are clinically used in the HIV chemotherapy, significantly decreased the fungal peptidase activity, varying from 55 to 99%. Moreover, sclerotic cell-derived aspartic peptidase hydrolyzed human albumin, an important serum protein, as well as laminin, an extracellular matrix component, but not immunoglobulin G and fibronectin. It is well-known that aspartic peptidases play important physiological roles in fungal cells. With this task in mind, the effect of pepstatin A, a classical aspartic peptidase inhibitor, on the F. pedrosoi proliferation was evaluated. Pepstatin A inhibited the fungal viability in both cellular density- and drug-concentration manners. Moreover, HIV-PIs at 10 μM powerfully inhibited the viability (>65%) of F. pedrosoi sclerotic cells. The detection of aspartic peptidase produced by sclerotic cells, the parasitic form of F. pedrosoi, may contribute to reveal new virulence markers and potential targets for chromoblastomycosis therapy