The use of mice as animal model for testing acute toxicity (LD-50) of toxic shock syndrome toxin / Utilização de camundongos como modelo animal para a verificação da toxicidade aguda da toxina-1 da síndrome do choque tóxico

Acute toxicity test (LD-50) using toxic shock syndrome toxin (TSST-1) was tested in BALB/c, C57BL/6 and Swiss mice. Animals (n = 10) were intraperitoneally injected with TSST-1 (0.01-10.0µg/mouse) followed 4h later by potentiating dose of lipopolysaccharide (75.0µg of LPS - E. coli O111:B4) and cumulative mortality was recorded over 72h. Control animals received either TSST-1 or LPS alone. The data were submitted to qui-Square test and acute toxicity test was calculated by probit analysis (confidence limits expressed as µg toxin/kg). BALB/c mice was the most sensitive (20.0µg/kg, 95 percent confidence limits: 9.0-92.0) followed by C57BL/6 (38.5µg/kg, 95 percent confidence limits: 9.11- 401.6). Data from Swiss mice was not conclusive, indicating only low sensitivity. Selection of the animal model and standardization of the experiment are fundamental for the development of serum neutralization tests used for final quality control of vaccine production.

A toxicidade aguda (DL-50) da toxina da síndrome do choque tóxico (TSST-1) foi testada em linhagens de camundongos BALB/c, C57BL/6 e Suíça. Os animais (n=10) inoculados intraperitoneal com doses crescentes de toxina (0,01 - 10,0µg/animal) receberam 4h após 75µg de LPS (E. coli O111: B4). A toxicidade aguda (DL50) foi observada por um período de 72h e os dados submetidos ao teste de qui- quadrado. Os resultados e os limites de confiança foram expressos em µg de toxina/kg. A linhagem BALB/c apresentou maior sensibilidade (20µg/kg - limite de confiança a 95 por cento entre 9,0- 92,0), seguida da C57BL/6 (38,5µg/kg - limite de confiança a 95 por cento entre 9,11 - 401,6). A amplitude dos limites de confiança deve-se à natureza da toxina, ao mecanismo de ação, a via de inoculação e ao animal utilizado. A seleção do modelo animal e a padronização do experimento são fundamentais para o desenvolvimento de testes de soro neutralização para fins de controle de qualidade do processo de produção de vacinas.

Bioconversion of nitriles by Candida guilliermondii CCT 7207 cells immobilized in barium alginate.

Nitrile degradation by Candida guilliermondii CCT 7207 using free and immobilized cell systems was compared. Different specific growth rates were observed for immobilized (mumax=0.021 h(-1)) and the free cells (mumax=0.029 h(-1)). The maximum specific rate of acetic acid formation was 0.387 h(-1) and 0.266 h(-1) for free and immobilized cells, respectively. Cell adhesion to the support materials was confirmed by scanning electron microscopy. When immobilized, the yeast was able to use high nitrile and amide concentrations (aliphatic and aromatic) as nitrogen sources. The results suggest that C. guilliermondii CCT 7207 presents a physiological pattern potentially useful for the bioremediation of polluted environments or for the bioproduction of amides and organic acid of high commercial value.

Degradation of phenol by Trichosporon sp. LE3 cells immobilized in alginate.

The degradation of phenol by freely suspended cells of Trichosporon sp. LE3 and alginate-immobilized cells was studied in batch culture. The alginate concentration (2 or 4%) and the cross-linking salt used (BaCl2 or CaCl2) affected the rate and percentage of phenol degradation. The highest values were obtained for immobilized cells at 2% calcium alginate, although complete degradation of 15 and 18 mM phenol was not observed. When the cell concentrations in the assays were doubled, the 2% calcium alginate-immobilized cells were able to degrade up to 30 mM phenol in less than 120 hours, although the free cells did not completely degrade phenol at concentrations above 20 mM. The maximum phenol degradation rate was a strong function of initial phenol concentrations, being the highest values being observed for 20 mM phenol.

Application of factorial design to study of heavy metals biosorption by waste biomass from beverage distillery.

A full factorial design leading to 20 sets of sorption runs was conducted to study the influence of four variables (bleaching earth and biomass concentrations, pH, and sorption time) on the iron, nickel, and chromium removal from stainless steel effluent using waste biomass from a beverage industry. Similar factor effects and interactions were found for each metal involved in this biosorption study, and the main factors were pH (positive effect) and biomass concentration (negative effect). Response surface methodology was adopted and an empirical linear polynomial model constructed on the basis of the specific uptake (mg of metal/g of biomass as dryweight) for each metal species. Under optimized process conditions (pH 4.0, biomass concentration of 2.0 g/L, absence of Celite), uptake values of 155 mg of Fe/g, 38 mg of Cr/g, and 0.4 mg of Ni/g were achieved after 3 h. This corresponded to a reduction in heavy metals concentration of approx 94% for Cr, 57% for Fe, and 25% for Ni.

Yeast communities and genetic polymorphism of Saccharomyces cerevisiae strains associated with artisanal fermentation in Brazil.

Yeast communities and genetic polymorphism of prevalent Saccharomyces cerevisiae strains isolated from the spontaneous fermentation of the sugarcane juice during the production of aguardente in three distilleries in the state of Minas Gerais, Brazil, were studied. S. cerevisiae was the prevalent species during the process of aguardente production, but Schizosaccharomyces pombe was predominant in old fermentations in one distillery. Transient yeast species were found in a variable number, probably due to the daily addition of sugarcane juice, and they were different for each of the three distilleries studied. PFGE and PCR analysis of the predominant strains of S. cerevisiae isolated from the fermented must showed a high degree of genetic polymorphism among the three distilleries. A high molecular variability of S. cerevisae strains was also observed among strains isolated from the same vat at different fermentation ages. Our results showed that there was a succession of geneticly different strains of S. cerevisae during the process of aguardente production.

Enzymatic degradation of nitriles by a Candida guilliermondii UFMG-Y65.

Candida guilliermondii UFMG-Y65, isolated from a gold mine, was able to utilize different nitriles and the corresponding amides as sole source of nitrogen, at concentrations up to 2 M. Resting cells cultivated on YCB-acetonitrile medium showed nitrile hydrolyzing enzyme activities against acrylonitrile and benzonitrile. These enzymes were inducible and intracellular; the optimum pH was 7.0-8.0, and the optimum temperature 25 degrees C-30 degrees C. Liquid chromatographic analysis indicated that C. guilliermondii UFMG-Y65 metabolized 12 mM benzonitrile to 11 mM benzoic acid and 10 mM acrylonitrile to 7.9 mM acrylic acid. The results suggest that C. guilliermondii UFMG-Y65 may be useful for the bioproduction of amides and acids, and for the bioremediation of environments contaminated with nitriles.

Metabolism of benzonitrile by Cryptococcus sp. UFMG-Y28.

The yeast Cryptococcus sp. UFMG-Y28 can utilize benzonitrile as a nitrogen and possible additional carbon source. The kinetics of growth on Yeast Carbon Base (YCB) added of benzonitrile as sole nitrogen source showed that benzonitrile was metabolized to benzoic acid and ammonia. Liquid chromatography analysis indicated that Cryptococcus sp. UFMG-Y28 metabolized 12 mM benzonitrile to 10 mM benzoic acid. Resting cells cultivated on YCB-propionitrile medium showed nitrilase activity against benzonitrile. This strain appears to be promising for bioconversion of nitriles to high value acids and for bioremediation of sites contaminated with aliphatic and aromatic nitriles.

Screening of yeasts from Brazilian Amazon rain forest for extracellular proteinases production.

Eighty seven yeast strains representing 34 species isolated from Parahancornia amapa fruit and associated Drosophila flies collected in the Brazilian Amazon rain forest, were screened for proteinase production. Proteolytic activity was tested through casein hydrolysis in solid medium supplemented with 0.5% casein and glucose. Among 23 strains, 18 from genus Candida and 5 from Pichia were caseinolytic and produced proteinases in yeast carbon base liquid medium supplemented with casein 0.01%. The proteolytic activity was tested on pH ranging from 2.0 to 9.0 in correspondence to the pH of the cultures media in which the yeasts were grown. Six highly proteolytic strains: Candida parapsilosis AP153A, C. krusei AP176, C. sorbosa DR215, C. sorbosa AP259, C. valida AP209A and C. sorboxylosa AP287 were selected and the pH optima of production and the proteolytic activity were determined. In general the secretion of proteinase was maximum throughout the exponential and the stationary phases. Greater production occurred in acidic culture and high activity was observed at pH 3.0, 4.0 and 5.0.

Utilization of acetonitrile and other aliphatic nitriles by a Candida famata strain.

A variant of a yeast strain identified as Candida famata isolated from gold mine effluent was able to grow on acetonitrile, acrylonitrile, butyronitrile, isobutyronitrile, methacrylnitrile, propionitrile, succinonitrile, valeronitrile, acetamide, isobutyamide, and succinamide as sole nitrogen source, after acclimatization. The yeast grew on acetonitrile and acetamide at concentrations up to 4%. The utilisation of acetonitrile and acetamide by the C. famata strain probably involves hydrolysis in a two-step reaction mediated by both inducible and intracellular nitrile hydratase and amidase.

Cyanide degradation by an Escherichia coli strain.

Chemical formation of a glucose-cyanide complex was necessary for metabolic degradation of cyanide at concentrations up to 50.0 mg/L by a strain of Escherichia coli isolated from gold extraction circuit liquids. Ammonia accumulating during the culture log phase as the sole nitrogen by-product was further utilized for bacterial growth. Washed (intact) cells, harvested at different periods of bacterial growth on cyanide, consumed oxygen in presence of cyanide. These findings suggest that metabolism of cyanide involved a dioxygenase enzyme that converted cyanide directly to ammonia, without the formation of cyanate.

Utilization of ammonia, generated from abiotic cyanide degradation, by Rhodotorula rubra.

The viability of the yeast Rhodotorula rubra, isolated from liquid samples of gold-mine effluents, was not affected by the presence of 11.52 mM cyanide. The yeast was able to utilize ammonia, generated from abiotic cyanide degradation in the presence of reducing sugars, in aerobic culture at pH 9.0. These physiological characteristics encourage studies with mixed cultures of cyanide-degrading organisms, using this yeast as an assimilator of ammonia.

Intraspecific protoplast fusion of amylase-producing strains of Candida fennica.

The production of α-amylase was increased by protoplast fusion of auxotrophic mutants of Candida fennica FTPT-8903. One prototrophic fusant was 90% and 32% more efficient in producing α-amylase in semi-solid and liquid fermentation, respectively, than the parental strains. Protoplast fusion did not significantly stimulate the synthesis of glucoamylase in the fusants.

Screening of beta-galactosidase producing yeasts / Estudio de levaduras productoras de betagalactosidasa

Se evaluaron varias cepas de levaduras para determinar los rendimientos en ß galactosidasa. La mejor cepa productora resultó Criptococcus. La actividad enzimática de algunas cepas de Cr. albidus y Cr. luteolus fueron superiores a las cepas de levadura Kluyveromyces fragilis y K. lactis, que han sido reportadas en la literatura como las mejores fuentes de la enzima para su utilización en la industria alimenticia

Production of amylases by yeasts.

Yeasts (228) isolated for natural habitats were screened for their ability to produce amylases in semisolid medium of wheat bran. Strains of Aureobasidium pullulans, Candida famata, and Candida kefyr showed high enzymatic activity for α-amylase, glucoamylase, and debranching enzyme. Key words: Aureobasidium, Candida, amylolytic yeasts, α-amylase, glucoamylase.