RESUMO
Natural enzyme inhibitors have been widely described in literature because of its pharmacological and cosmetic applications. Fungi found in caves represent a promising source of bioactive substances that are still little explored scientifically. Thus, the present work evaluated the presence of enzymatic modulators in a filtrate obtained from the cultivation of the cave fungus Lecanicillium aphanocladii (Family: Cordycipitaceae). Snake venoms from Bothrops alternatus and Bothrops atrox were used as an enzymatic source for the induction of the phospholipase, proteolytic, thrombolytic, cytotoxic and coagulant activities. Compounds present in the fungal filtrate inhibited 50, 23·8, 26·6, 50·9 and 52·5% of the proteolytic, phospholipase, haemolytic, thrombolytic and coagulant activities respectively. The filtrate was not cytotoxic on erythrocytes, but induced partial dissolution of thrombi. Fungal enzyme inhibitors that have low or no toxicity and can be obtained on a large scale and at low cost are relevant in the medical-scientific context. Therefore, the inhibition of phospholipases A2 and proteases observed in the present work highlights the potential of fungal metabolites for the development of drugs that can be used in the treatment of haemostasis and inflammation-related disorders. SIGNIFICANCE AND IMPACT OF THE STUDY: In this study, secondary metabolites synthesized by Lecanicillium aphanocladii, a fungus isolated from caves, demonstrated modulating action on proteases and phospholipases A2 present in snake venoms of the Bothrops genus, widely used as tools for the study of pathophysiology processes related to haemostasis and inflammation. The results suggest the possibility of future applications for these metabolites in the development of pharmaceuticals of medical-scientific interest.
Assuntos
Ascomicetos/química , Bothrops/metabolismo , Venenos de Crotalídeos/enzimologia , Peptídeo Hidrolases/metabolismo , Inibidores de Fosfolipase A2/farmacologia , Fosfolipases A2/metabolismo , Inibidores de Proteases/farmacologia , Animais , Ascomicetos/metabolismo , Coagulação Sanguínea/efeitos dos fármacos , Eritrócitos/efeitos dos fármacos , Hemostasia/efeitos dos fármacos , Humanos , Inflamação/tratamento farmacológico , Proteólise/efeitos dos fármacosRESUMO
Biochemical and molecular analysis was used for identification of different kefir yeasts species from Brazil, Canada and the United States of America. The sugar/ethanol-resistant activity of the yeasts was evaluated. Saccharomyces cerevisiae and Kluyveromyces marxianus had the highest growth rates, suggesting biotechnological applications possible for these strains.
RESUMO
Biochemical and molecular analysis was used for identification of different kefir yeasts species from Brazil, Canada and the United States of America. The sugar/ethanol-resistant activity of the yeasts was evaluated. Saccharomyces cerevisiae and Kluyveromyces marxianus had the highest growth rates, suggesting biotechnological applications possible for these strains.
Assuntos
Sequência de Bases , Produtos Fermentados do Leite , Etanol/análise , Genoma Bacteriano , Técnicas In Vitro , Leveduras/genética , Leveduras/isolamento & purificação , Fenótipo , Reação em Cadeia da Polimerase/métodos , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/isolamento & purificação , Eletroforese , Variação Genética , Genótipo , MétodosRESUMO
Biochemical and molecular analysis was used for identification of different kefir yeasts species from Brazil, Canada and the United States of America. The sugar/ethanol-resistant activity of the yeasts was evaluated. Saccharomyces cerevisiae and Kluyveromyces marxianus had the highest growth rates, suggesting biotechnological applications possible for these strains.(AU)
Assuntos
Bioquímica , Etanol/análise , Açúcares , Biotecnologia/tendências , Leveduras/ultraestrutura , Saccharomyces cerevisiaeRESUMO
The goals of this study were to develop a PCR technique to detect ascV and aopB genes from the type III secretion system (T3SS), to evaluate the frequency of these genes in Aeromonas hydrophila strains isolated from diseased fish and from aquaculture environments, and to determine the relationship between the presence of these genes and virulence of A. hydrophila in Nile tilapia. The PCR assay developed here successfully detected the target genes, showing three different profiles for the strains ascV+/aopB+, ascV+/aopB-, and ascV-/aopB-. A higher frequency of ascV+/aopB+ was verified in isolates from diseased fish compared to those from aquaculture environments (P<0.05). Among 64 isolates from diseased fish, ascV+/aopB+ (62.5%) was the most frequent profile (P<0.05) and caused more intensive mortality rates. Environmental strains containing the ascV+/aopB+ profile were less virulent than isolates from clinical cases. These results suggest that the presence of a functional T3SS probably increases the virulence of A. hydrophila. The PCR technique was shown to be a specific and efficient tool for detection of T3SS, and this technique can be used for virulence typing of A. hydrophila isolates.
Assuntos
Aeromonas hydrophila/genética , Aeromonas hydrophila/patogenicidade , Ciclídeos , Doenças dos Peixes/microbiologia , Infecções por Bactérias Gram-Negativas/veterinária , Aeromonas hydrophila/metabolismo , Animais , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Regulação Bacteriana da Expressão Gênica/fisiologia , Infecções por Bactérias Gram-Negativas/microbiologia , VirulênciaRESUMO
We developed an improved protocol for the electrotransformation of Corynebacterium pseudotuberculosis, testing variations of parameters in the procedures that are routinely used for the preparation of electrocompetent cells of this species, including (i) culture conditions, (ii) cell growth phase, (iii) electroporation solutions and (iv) quantity of plasmid DNA. We obtained the greatest efficiency of transformation when the cells were grown until the stationary phase and then washed with 10% glycerol electroporation solution. The transformation efficiency was inversely proportional to the quantity of plasmid DNA. The transformation efficiency reached 10(5) colony-forming units (cfu)/mug plasmid DNA. This protocol would be useful for genetic studies of C. pseudotuberculosis.