Seleção de micro-organismos celulolíticos comparados com o Trichoderma reesei CCT 2768

As celulases atualmente são enzimas extensivamente estudadas para a hidrólise de resíduos lignocelulósicos para obtenção de açúcares fermentescíveis utilizáveis em diferentes processos biotecnológicos, como a produção de bioetanol. O objetivo deste trabalho foi encontrar fungos celulolíticos que sejam eficientes nos processos de degradação da biomassa lignocelulósica. No presente trabalho foram selecionados 4 fungos celulolíticos previamente isolados pelo Laboratório de Biotecnologia Industrial (UNESP – Assis). Estes fungos foram cultivados em bagaço de cana-de-açúcar como substrato e para seus extratos enzimáticos foram testados suas atividades para celulases (FPase) e endoglucanases (CMCase). Os fungos FS09 (0,054 FPU/mL e 1,79 FPU/g de substrato, 0,874 U/mL e 29,1 U/g de substrato) e M51 (0,049 FPU/mL e 1,62 FPU/g de substrato, 1,094 U/mL e 36,46 U/g de substrato), mostrando valores superiores ao fungo T.reesei CCT 2768 que é referência em estudos e produção de celulases.

Produção de enzimas celulolíticas pelos fungos Trichoderma reesei e Myceliophthora thermophila e aplicação na sacarificação do bagaço de cana-de-açucar

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Avaliação de isolados de Trichoderma spp. para controle de Phytophthora nicotianae

Pós-graduação em Microbiologia Agropecuária - FCAV

Unraveling Trichoderma species in attine ant environment: description of three new taxa

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Marine Fungi Aspergillus sydowii and Trichoderma sp Catalyze the Hydrolysis of Benzyl Glycidyl Ether

Whole cells of the marine fungi Aspergillus sydowii Gc12, Penicillium raistrickii Ce16, P. miczynskii Gc5, and Trichoderma sp. Gc1, isolated from marine sponges of the South Atlantic Ocean (Brazil), have been screened for the enzymatic resolution of (+/-)-2-(benzyloxymethyl)oxirane (benzyl glycidyl ether; 1). Whole cells of A. sydowii Gc12 catalyzed the enzymatic hydrolysis of (R,S)-1 to yield (R)-1 with an enantiomeric excess (ee) of 24-46% and 3-(benzyloxy)propane-1,2-diol (2) with ee values < 10%. In contrast, whole cells of Trichoderma sp. Gc1 afforded (S)-1 with ee values up to 60% and yields up to 39%, together with (R)-2 in 25% yield and an ee of 32%. This is the first published example of the hydrolysis of 1 by whole cells of marine fungi isolated from the South Atlantic Ocean. The hydrolases from the two studied fungi exhibited complementary regioselectivity in opening the epoxide ring of racemic 1, with those of A. sydowii Gc12 showing an (S) preference and those of Trichoderma sp. Gc1 presenting an (R) preference for the substrate.

RNA Interference of Endochitinases in the Sugarcane Endophyte Trichoderma virens 223 Reduces Its Fitness as a Biocontrol Agent of Pineapple Disease

The sugarcane root endophyte Trichoderma virens 223 holds enormous potential as a sustainable alternative to chemical pesticides in the control of sugarcane diseases. Its efficacy as a biocontrol agent is thought to be associated with its production of chitinase enzymes, including N-acetyl-beta-D-glucosaminidases, chitobiosidases and endochitinases. We used targeted gene deletion and RNA-dependent gene silencing strategies to disrupt N-acetyl-beta-D-glucosaminidase and endochitinase activities of the fungus, and to determine their roles in the biocontrol of soil-borne plant pathogens. The loss of N-acetyl-beta-D-glucosaminidase activities was dispensable for biocontrol of the plurivorous damping-off pathogens Rhizoctonia solani and Sclerotinia sclerotiorum, and of the sugarcane pathogen Ceratocystis paradoxa, the causal agent of pineapple disease. Similarly, suppression of endochitinase activities had no effect on R. solani and S. sclerotiorum disease control, but had a pronounced effect on the ability of T. virens 223 to control pineapple disease. Our work demonstrates a critical requirement for T. virens 223 endochitinase activity in the biocontrol of C. paradoxa sugarcane disease, but not for general antagonism of other soil pathogens. This may reflect its lifestyle as a sugarcane root endophyte.

Conformational stability of recombinant manganese superoxide dismutase from the filamentous fungus Trichoderma reesei

Superoxide dismutases (SODS; EC 1.15.1.1) are part of the antioxidant system of aerobic organisms and are used as a defense against oxidative injury caused by reactive oxygen species (ROS). The cloning and sequencing of the 788-bp genomic DNA from Trichoderma reesei strain QM9414 (anamorph of Hypocrea jecorina) revealed an open reading frame encoding a protein of 212 amino acid residues, with 65-90% similarity to manganese superoxide dismutase from other filamentous fungi. The TrMnSOD was purified and shown to be stable from 20 to 90 degrees C for 1 h at pH from 8 to 11.5, while maintaining its biological activity. (C) 2011 Elsevier B.V. All rights reserved.

Control biológico de Fusarium graminearum : utilización de Trichoderma spp. y biofumigación con parte aérea de Brassica juncea

Los objetivos de este trabajo fueron: determinar la factibilidad de la utilización combinada de dos métodos de control biológico: la aplicación del hongo antagonista Trichoderma spp. y la biofumigación con la parte aérea de Brassica juncea en el estadio de fin de fructificación; evaluar su efecto sobre el crecimiento del patógeno Fusarium graminearum. Se trituraron plantas de B. juncea y se colocaron en recipientes de plástico en dosis de 5 y 10 g. Sobre el material triturado se apoyó una caja de Petri con agar papa glucosado al 2%, que contenía un disco con micelio de F. graminearum o Trichoderma spp. o ambos hongos. Los recipientes de plástico se cerraron e incubaron a 25±2°C en oscuridad durante 7 días. Finalizado este período, se midió el diámetro de las colonias. Se obtuvieron los siguientes resultados: i) cuando se biofumigaron por separado, no se observó efecto fungistático de B. juncea sobre Trichoderma spp. ni sobre F. graminearum; ii) en ausencia del biofumigante, Trichoderma spp. inhibió significativamente el crecimiento de las colonias de F. graminearum, iii) la combinación de Trichoderma spp. y la biofumigación con B. juncea mostró un efecto sinérgico sobre el control del crecimiento miceliar de F. graminearum. Los resultados in vitro sugieren que el crecimiento de Trichoderma spp. y su potencial efecto de biocontrol sobre F. graminearum, no son afectados por la biofumigación con B. juncea. La utilización combinada de Trichoderma spp. y la biofumigación con B. juncea, tendría un efecto sinérgico sobre el control del crecimiento de F. graminearum.

The cellulose-binding domain of the major cellobiohydrolase of Trichoderma reesei exhibits true reversibility and a high exchange rate on crystalline cellulose.

Cellulose-binding domains (CBDs) bind specifically to cellulose, and form distinct domains of most cellulose degrading enzymes. The CBD-mediated binding of the enzyme has a fundamental role in the hydrolysis of the solid cellulose substrate. In this work we have investigated the reversibility and kinetics of the binding of the CBD from Trichoderma reesei cellobiohydrolase I on microcrystalline cellulose. The CBD was produced in Escherichia coli, purified, and radioactively labeled by reductive alkylation with 3H. Sensitive detection of the labeled CBD allowed more detailed analysis of its behavior than has been possible before, and important novel features were resolved. Binding of the CBD was found to be temperature sensitive, with an increased affinity at lower temperatures. The interaction of the CBD with cellulose was shown to be fully reversible and the CBD could be eluted from cellulose by simple dilution. The rate of exchange measured for the CBD-cellulose interaction compares well with the hydrolysis rate of cellobiohydrolase I, which is consistent with its proposed mode of action as a processive exoglucanase.

Molecular evidence that the asexual industrial fungus Trichoderma reesei is a clonal derivative of the ascomycete Hypocrea jecorina.

The relationship of the important cellulase producing asexual fungus Trichoderma reesei to its putative teleomorphic (sexual) ancestor Hypocrea jecorina and other species of the Trichoderma sect. Longibrachiatum was studied by PCR-fingerprinting and sequence analyses of the nuclear ribosomal DNA region containing the internal transcribed spacers (ITS-1 and ITS-2) and the 5.8S rRNA gene. The differences in the corresponding ITS sequences allowed a grouping of anamorphic (asexual) species of Trichoderma sect. Longibrachiatum into Trichoderma longibrachiatum, Trichoderma pseudokoningii, and Trichoderma reesei. The sexual species Hypocrea schweinitzii and H. jecorina were also clearly separated from each other. H. jecorina and T. reesei exhibited identical sequences, suggesting close relatedness or even species identity. Intraspecific and interspecific variation in the PCR-fingerprinting patterns supported the differentiation of species based on ITS sequences, the grouping of the strains, and the assignment of these strains to individual species. The variations between T. reesei and H. jecorina were at the same order of magnitude as found between all strains of H. jecorina, but much lower than the observed interspecific variations. Identical ITS sequences and the high similarity of PCR-fingerprinting patterns indicate a very close relationship between T. reesei and H. jecorina, whereas differences of the ITS sequences and the PCR-fingerprinting patterns show a clear phylogenetic distance between T. reesei/H. jecorina and T. longibrachiatum. T. reesei is considered to be an asexual, clonal line derived from a population of the tropical ascomycete H. jecorina.

Secretome analysis of Trichoderma atroviride T17 biocontrol of Guignardia citricarpa

The fungal species Guignardia citricarpa is an important pathogen in citriculture. Members of the fungal genus Trichoderma are recognized as biocontrol agents but studies on the interactions between both fungi are scarce. This study aimed to identify extracellular proteins secreted by Trichoderma atroviride T17 that are related to the control of G. citricarpa. Two-dimensional gel electrophoresis (2D) was used to study the patterns of proteins secreted by T. atroviride T17 in medium containing glucose (control) and in medium containing G. citricarpa GC3 inactivated mycelium. We identified 59 of the 116 spots differentially expressed (50.86%) by LC–MS/MS. Of these, we highlight the presence of glycoside hydrolases (CAZy families 3, 43, 54, 76 and 93), chitinase, mutanase, a-1,3-glucanase, a-1,2-mannosidase, carboxylic hydrolase ester, carbohydrate-binding module family 13, glucan 1,3-b-glucosidase, a-galactosidase and Neutral protease 2. These proteins are related to mycoparasitism processes, stimuli and therefore to the biological control of pathogens. The results obtained are in agreement with reports describing an increase in the secretion of proteins related to mycoparasitism and biological control and a reduction in the secretion of proteins related to the metabolism of Trichoderma species grown in the presence of the pathogen. Moreover, these results are pioneer in understanding T. atroviride interaction with G. citricarpa. For the first time, we identified potential candidate proteins that may have a role in the antagonism mechanism of G. citricarpa by T. atroviride T17. Thus our results shed a light into the molecular mechanisms that T. atroviride use to control G. citricarpa.

Caracterización del agente causal de la producción radicular del aguacate y Trichoderma sp., como potencial agente de control biológico en Nicaragua.

En el cultivo de aguacate (Persea americana Mill.) se presentan problemas fitosanitarios importantes dentro de los cuales sobresalen por su relevancia las enfermedades de la raíz. Un fitopatógeno limitante de este cultivo es el oomicete Phytophthora cinnamomi Rands, que puede causar pérdidas hasta del 90%. Por tal razón el principal objetivo del estudio fue generar información acerca de la etiología del agente causal de la pudrición radicular del aguacate utilizando marcadores morfológicos y moleculares, además de proponer alternativas de manejo de carácter biológico que estén enmarcadas dentro de un programa de manejo integrado de la enfermedad. Se realizaron colectas de muestras de suelo en cuatro localidades del departamento de Masaya. La identificación morfológica del patógeno se realizó mediante claves taxonómicas y se confirmó a través de la técnica PCR-RFLP. Se identificó a P. cinnamomi como el principal agente causal de la pudrición radicular del aguacate. Los aislados de P. cinnamomi fueron enfrentados con Trichoderma sp por el método de cultivo dual en cajas Petri con medio PDA. Se determinó el porcentaje de inhibición de crecimiento radial (PICR) a las 72 horas, así como el grado de antagonismo de cada una de las cepas de Trichoderma sp utilizadas en el estudio. Las cepas de Trichoderma al enfrentarlas a aislados del patógeno P. cinnamomi se ubicaron en las Clases 1 y 2 de la escala de evaluación, por lo tanto se consideraron altamente antagonistas. Existe la posibilidad de manejo biológico de las poblaciones de P. cinnamomi con microorganismos antagonistas del género Trichoderma no solamente en agroecosistemas de aguacate, sino también en otros sistemas agrícolas y forestales donde el patógeno esté presente.

Trichoderma in Brazil: history, research, commercialization and perspectives.

2009

Control of Fusarium in chrysanthemum with sewage sludge, biofertilizer, hydrolyzed fish, chitosan and Trichoderma.

2009