Potencial de biocatálise enantiosseletiva de lipases microbianas

Microbial lipases have a great potential for commercial applications due to their stability, selectivity and broad substrate specificity because many non-natural acids, alcohols or amines can be used as the substrate. Three microbial lipases isolated from Brazilian soil samples (Aspergillus niger; Geotrichum candidum; Penicillium solitum) were compared in terms of their stability and as biocatalysts in the enantioselective esterification using racemic substrates in organic medium. The lipase from Aspergillus niger showed the highest activity (18.2 U/mL) and was highly thermostable, retaining 90% and 60% activity at 50 ºC and 60 ºC after 1 hour, respectively. In organic medium, this lipase provided the best results in terms of enantiomeric excess of the (S)-active acid (ee = 6.1%) and conversion value (c = 20%) in the esterification of (R,S)-ibuprofen with 1-propanol in isooctane. The esterification reaction of the racemic mixture of (R,S)-2-octanol with decanoic acid proceeded with high enantioselectivity when lipase from Aspergillus niger (E = 13.2) and commercial lipase from Candida antarctica (E = 20) were employed.

Influência da temperatura, período de molhamento e concentração do inóculo de fungos na incidência de podridões pós-colheita em frutos de tomateiro

Foi avaliada a influência da temperatura (5, 15, 25 e 35 °C), da duração do período de molhamento (0, 3, 6, 12, 24, 48 e 72 h) e da concentração de inóculo (10¹, 10², 10(4), 10(6) e 10(8) esporos/ml) na incidência de podridões de pós-colheita em frutos de tomateiro (Lycopersicon esculentum) (cv. Santa Clara) causadas por Fusarium verticillioides, Geotrichum candidum e Rhizopus stolonifer. A temperatura influenciou significativamente (P=0,05) a incidência de podridões, ocorrendo maior incidência de lesões em frutos incubados a 25 °C. Não foi observado o desenvolvimento de podridão em frutos inoculados com F. verticillioides e incubados nas temperaturas de 5 e 35 °C, sendo nesta última verificada a menor incidência de podridões causadas por G. candidum e R. stolonifer. Nas temperaturas de 5, 15 e 25 °C, a incidência de podridões causadas por R. stolonifer variou entre 97,5 e 100%. A presença de água livre na superfície dos frutos de tomateiro foi desnecessária para a incidência de podridões causadas por F. verticillioides, G. candidum e R. stolonifer, embora os níveis de incidência tenham aumentado com o incremento no período de molhamento, exceto para R. stolonifer, que causou a incidência máxima de doença (100%), na ausência de molhamento. A incidência de podridões aumentou com o incremento na concentração de inóculo de 10¹ a 10(8) esporos/mL, atingindo o máximo (100%) na concentração de 10(4) esporos/ml para F. verticillioides e R. stolonifer, enquanto o mesmo nível foi atingido por G. candidum com 10(8) esporos/ml.

Caracterização dos danos pós-colheita em citros procedentes de "packinghouse"

As doenças pós-colheita representam um sério obstáculo à citricultura, uma vez que comprometem a qualidade e quantidade dos frutos colhidos. Este trabalho objetivou caracterizar as injúrias pós-colheita de frutos de laranja 'Pêra', 'Lima' e 'Natal' e de tangor 'Murcott', destinados ao mercado interno, após diferentes etapas do beneficiamento em "packinghouse". Foram coletados cem frutos na chegada ao "packinghouse", na banca de embalagem e no palete, após embalamento em caixas de madeira. Os frutos foram individualizados e submetidos à câmara úmida por 24 horas, permanecendo por mais 20 dias a 25ºC e 85% de umidade relativa. A incidência de podridões foi avaliada visualmente após a retirada da câmara úmida e a cada três dias. Os patógenos fúngicos encontrados tiveram a patogenicidade confirmada através da inoculação em frutos sadios. Não houve diferença significativa na incidência de doenças pós-colheita nas diferentes fases do processamento nas variedades Lima e Natal. Na variedade Pêra e no tangor 'Murcott', a incidência de doenças foi menor nas amostras coletadas na chegada ao "packinghouse". O bolor verde (Penicillium digitatum) foi a principal doença encontrada nos diferentes frutos cítricos. Outras doenças importantes foram a antracnose (Colletotrichum gloeosporioides), as podridões pedunculares (Lasiodiplodia theobromae e Phomopsis citri) e a podridão azeda (Geotrichum candidum).

Potencial de leveduras isoladas do solo e do filoplano de plantas cítricas no biocontrole de doenças de pós-colheito de citros

Pós-graduação em Microbiologia Agropecuária - FCAV

Estudo dos mecanismos de ação de leveduras envolvidos no biocontrole de doenças de pós-colheita em citros

Pós-graduação em Microbiologia Agropecuária - FCAV

Análise da susceptibilidade de leveduras isoladas de insetos sociais aos antifúngicos

A frequência de micoses invasivas causadas por fungos oportunistas, incluindo leveduriformes, vem aumentando significativamente nas últimas décadas e atinge principalmente indivíduos imunocomprometidos. A tribo Attini possui cerca de 230 espécies de formigas e todas possuem o hábito de cultivar e se alimentar de fungos. Algumas espécies cultivam o fungo mutualista em nódulos poligonais sólidos com 0,25 a 0,50mm de diâmetro, aproximadamente. No interior desses nódulos, encontram-se células na forma de leveduras, que quando semeadas em meio de cultura se desenvolvem como fungo filamentoso. Este tipo de fungicultura está restrito a algumas espécies do gênero Cyphomyrmex. Neste trabalho, isolamos 30 leveduras dos nódulos de oito ninhos de Cyphomyrmex transversus da região de Ilhéus (BA), e analisamos a susceptibilidade aos seguintes antifúngicos: Anfotericina B (AMB), Fluconazol (FCZ), Voriconazol (VCZ) e Itraconazol (ITZ). A caracterização das estirpes foi feita através de métodos morfológicos e moleculares, incluindo-se o sequenciamento dos domínios D1/D2 do rDNA. A avaliação da atividade antifúngica foi realizada através do método de microdiluição, de acordo com a norma M27-A2 (CLSI, 2002). Candida parapsilosis ATCC 22019 e Candida krusei ATCC 6258 foram utilizadas como controle. Os resultados foram interpretados segundo os pontos de corte (breakpoints) aplicados ao gênero Candida. As estirpes foram identificadas como Trichosporon asahii (18), T. coremiiforme (4), Candida carpophila (syn. C. guilliermondii) (4), Galactomyces candidum (syn. Geotrichum candidum) (3) e Cryptococcus laurentii (1). Todas as espécies são conhecidas por possuírem representantes que já foram relatados como patógenos oportunistas. Elas foram sensíveis aos azóis FCZ, VCZ e ITZ, com exceção de G. candidus, cujas estirpes apresentaram sensibilidade dose-dependente a FCZ e ITZ... (Resumo completo, clicar acesso eletrônico abaixo)

Surface microbial consortia from Livarot, a French smear-ripened cheese

The surface microflora (902 isolates) of Livarot cheeses from three dairies was investigated during ripening. Yeasts were mainly identified by Fourier transform infrared spectroscopy. Geotrichum candidum was the dominating yeast among 10 species. Bacteria were identified using Biotype 100 strips, dereplicated by repetitive extragenic palindromic PCR (rep-PCR); 156 representative strains were identified by either BOX-PCR or (GTG) 55-PCR, and when appropriate by 16S rDNA sequencing and SDS-PAGE analysis. Gram-positive bacteria accounted for 65% of the isolates and were mainly assigned to the genera Arthrobacter, Brevibacterium, Corynebacterium, and Staphylococcus. New taxa related to the genera Agrococcus and Leucobacter were found. Yeast and Gram-positive bacteria strains deliberately added as smearing agents were sometimes undetected during ripening. Thirty-two percent of the isolates were Gram-negative bacteria, which showed a high level of diversity and mainly included members of the genera Alcaligenes, Hafnia, Proteus, Pseudomonas, and Psychrobacter. Whatever the milk used (pasteurized or unpasteurized), similar levels of biodiversity were observed in the three dairies, all of which had efficient cleaning procedures and good manufacturing practices. It appears that some of the Gramnegative bacteria identified should now be regarded as potentially useful in some cheese technologies. The assessment of their positive versus negative role should be objectively examined.

Biodiversity of the surface microbial consortia from Limburger, Reblochon, Livarot, Tilsit, and Gubbeen cheeses

Comprehensive collaborative studies from our laboratories reveal the extensive biodiversity of the microflora of the surfaces of smear-ripened cheeses. Two thousand five hundred ninety-seven strains of bacteria and 2,446 strains of yeasts from the surface of the smear-ripened cheeses Limburger, Reblochon, Livarot, Tilsit, and Gubbeen, isolated at three or four times during ripening, were identified; 55 species of bacteria and 30 species of yeast were found. The microfloras of the five cheeses showed many similarities but also many differences and interbatch variation. Very few of the commercial smear microorganisms, deliberately inoculated onto the cheese surface, were reisolated and then mainly from the initial stages of ripening, implying that smear cheese production units must have an adventitious "house" flora. Limburger cheese had the simplest microflora, containing two yeasts, Debaryomyces hansenii and Geotrichum candidum, and two bacteria, Arthrobacter arilaitensis and Brevibacterium aurantiacum. The microflora of Livarot was the most complicated, comprising 10 yeasts and 38 bacteria, including many gram-negative organisms. Reblochon also had a very diverse microflora containing 8 yeasts and 13 bacteria (excluding gram-negative organisms which were not identified), while Gubbeen had 7 yeasts and 18 bacteria and Tilsit had 5 yeasts and 9 bacteria. D. hansenii was by far the dominant yeast, followed in order by G. candidum, Candida catenulata, and Kluyveromyces lactis. B. aurantiacum was the dominant bacterium and was found in every batch of the 5 cheeses. The next most common bacteria, in order, were Staphylococcus saprophyticus, A. arilaitensis, Corynebacterium casei, Corynebacterium variabile, and Microbacterium gubbeenense. S. saprophyticus was mainly found in Gubbeen, and A. arilaitensis was found in all cheeses but not in every batch. C. casei was found in most batches of Reblochon, Livarot, Tilsit, and Gubbeen. C. variabile was found in all batches of Gubbeen and Reblochon but in only one batch of Tilsit and in no batch of Limburger or Livarot. Other bacteria were isolated in low numbers from each of the cheeses, suggesting that each of the 5 cheeses has a unique microflora. In Gubbeen cheese, several different strains of the dominant bacteria were present, as determined by pulsed-field gel electrophoresis, and many of the less common bacteria were present as single clones. The culture-independent method, denaturing gradient gel electrophoresis, resulted in identification of several bacteria which were not found by the culture-dependent (isolation and rep-PCR identification) method. It was thus a useful complementary technique to identify other bacteria in the cheeses. The gross composition, the rate of increase in pH, and the indices of proteolysis were different in most of the cheeses.

Effet de la réduction et de la substitution du NaCl dans le Camembert sur la croissance de la microflore fongique d'affinage

Santé Canada encourage les transformateurs à réduire le contenu en NaCl dans plusieurs aliments, dont les fromages, puisque sa surconsommation est associée à plusieurs problèmes de santé. L’effet causé par la réduction du NaCl dans le Camembert, ou de sa substitution partielle par du KCl, sur la qualité globale de ce fromage n’est pas connu. Le but du projet visait à étudier la cinétique d’incorporation du sodium et du potassium lors du saumurage d’un Camembert commercial afin de produire des fromages réduits en NaCl et substitués par du KCl. Ces nouvelles conditions ont ensuite permis d’étudier le comportement des ferments fongiques d’affinage Penicillium camemberti et Geotrichum candidum. Les résultats ont démontré une modification du profil de croissance des deux espèces, plus accentuée chez G. candidum, en fonction des modifications salines apportées. L’impact causé par ces changements de comportement doit maintenant être évalué au niveau de l’activité spécifique des deux espèces.

Dipodascus capitatus (Geotrichum capitatum): infecção sistêmica fatal em paciente com leucemia mielocítica aguda

As infecções causadas por Dipodascus capitatus são raras e de difícil tratamento. Aqui se relata um caso em paciente com leucemia mielocítica aguda. O isolamento fúngico ocorreu a partir de hemocultura e a identificação fenotípica baseou-se em métodos micológicos clássicos; a identificação genotípica foi realizada através do sequenciamento da região D1/D2 do 26 rDNA. Os testes de suscetibilidade foram realizados através do Etest® e microdiluição em caldo. A antifungicoterapia foi ineficaz, registrando-se óbito da paciente no 17° dia após o diagnóstico. Os autores comparam o caso com relatos similares e discutem a emergência destas infecções bem como suas dificuldades diagnósticas e terapêuticas.

Gastrite espontânea do camondongo e cogumelos do gênero Geotrichum Link

Slight acute catarrhal gastritis associated with fungi of the genus Geotrichum LINK was found to occur in apparently healthy white mice. This is of practical importance considering that the mouse is extensively employed in experimental work on nutrition.

Production of volatile compounds by Geotrichum fragrans using cassava wastewater as substrate

The cassava processing industry generates wastewater named manipueira with a high organic content. Although considered a pollutant, manipueira can be used as substrate for fermentative processes including the cultivation of Geotrichum fragrans. This aerobic microorganism isolated from cassava wastewater has cyanide resistant respiration. Under cassava wastewater cultivation, G. fragrans produced fruit aroma volatile compounds. This study evaluated volatile compounds produced by G. fragrans in cassava liquid waste. The waste had a sugar composition composed of dextrin (2.6%), maltose (1.4%), sucrose (32.1%), glucose (38.3%), and fructose (25.6%). The average value of total sugars was 58.2 g l(-1), composed of 38.0 g l(-1) reducing and 20.2 g l(-1) non-reducing sugars. The chemical oxygen demand (COD) average value was 60 000 mg l(-1). G. fragrans used sugars (fructose and glucose) for energy generation reducing the COD value of the cassava wastewater by 40%. Biomass production of G. fragrans cultivated for 12 h in natural cassava liquid waste was 12.8 g l(-)1. The volatile compounds identified in the cassava liquid waste after 72 h cultivation were: 1-butanol, 3-methyl 1-butanol (isoamylic alcohol), 2-methyl 1-butanol, 1-3 butanodiol and phenylethanol; ethyl acetate, ethyl propionate, 2-methyl ethyl propionate and 2-methyl propanoic. The effect of substrate supplementation with glucose (50 g l(-1)), fructose (50 g l(-1)) and aqueous yeast extract (200 ml l(-1)) did not affect the qualitative and quantitative profiles of volatile compounds. These results indicate that the carbon (C) source utilized by microorganism was glucose or fructose, while nitrogen (N) supplementation was not necessary because the agent did not exhaust all the nitrogen of the wastewater. (C) 2003 Elsevier B.V. Ltd. All rights reserved.

Cypripedium candidum - Cypripedium villosum - Cypripedium calceolus

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Frauenschuh : Cypripedium candidum - Cypripedium villosum - Cypripedium calceolus

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Air fungal contamination in two elementary schools in Lisbon, Portugal

A descriptive study was developed to monitor air fungal contamination in two elementary schools in Lisbon, Portugal. Eight air samples of 250 litres through impaction method were collected in canteen, library, classrooms and also, outside premises as reference place. Simultaneously, were also monitored environmental parameters, including temperature, and humidity through the equipment Babouc, LSI Sistems and according to the International Standard ISO 7726 - 1998. Considering both schools, sixteen different species of fungi in air were identified, being the 2 most commonly isolated Cladosporium sp. (51,1%) and Penicillium sp. (27,5%). Besides these genera Trichoderma, Aspergillus, Alternaria, Chrysonilia, Botritys, Ulocladium, Athrium, Aureobasidium, Phoma, Scedosporium e Geotrichum were also isolated. Regarding yeasts, Candida sp., Cryptococcus sp. and Rhodotorula sp. were isolated. The youngest school, as well canteens in each school, presented the worst results concerning the air fungal contamination, maybe due to the higher number of occupants. There was no significant relationship (p>0,05) between fungal contamination and temperature and humidity.