O estado de Tocantins tem despontado no cenário nacional como um grande produtor de soja. Entretanto, várias são os fatores que contribuem para a redução da produtividade da soja e dentre os principais estão as perdas por doenças e diminuição do estande de plantio. A utilização do controle biológico constitui uma estratégia de grande interesse e importância para viabilizar a redução ou substituição do uso de defensivos. Fungos como o Trichoderma e o Purpureocillium são importantes antagonistas no solo contra doenças radiculares e nematoides fitopatogênicos. Diversos produtos à base de Trichoderma e Purpureocillium são comercializados em todo mundo. Desse modo, objetivou se com o presente trabalhar avaliar os efeitos da inoculação de Trichoderma e Purpureocillium na cultura da soja em diferentes regiões (Gurupi, Porto Nacional, Alvorada e Crixas) em Tocantins. Os experimentos foram conduzidos em condições de campo nas diferentes cidades produtoras de soja no Tocantins, utilizando os produtos TrichoPlus e TrichoMix formulados a base de Trichoderma (Trichoderma asperellum UFT 201) e Purpureocillium lilacinum e com diferentes doses, visando o biocontrole para manutenção de estande e consequentemente aumento de produtividade. A inoculação de Trichoderma e Purpureocillium presentes nos produtos TrichoPlus e TrichoMix influenciaram positivamente a manutenção de estande, a produtividade da soja e auxiliou na promoção de crescimento em todos os locais avaliados. Em Gurupi, no experimento I e II, tanto a aplicação isolada de Trichoderma (TrichoPlus) como a aplicação conjunta de Trichoderma e Purpureocillium (TrichoMix) promoveram incrementos na manutenção de estandes e maiores valores de produtividade em relação aos demais tratamentos. Em Porto Nacional, a inoculação dos produtos TrichoPlus e TrichoMix influenciaram positivamente a manutenção de estande e a produtividade das cultivares de soja de ciclos precoce, intermediária e tardio. Em Alvorada, a inoculação de Trichoderma (TrichoPlus) em soja de primeiro e segundo ano aumentaram o índice de sobrevivência de plantas, embora não tenha tido efeito significativo na produtividade. Em Crixas, a inoculação de Trichoderma (TrichoPlus) promoveu incrementos na manutenção de estandes e maiores médias de produtividade. O uso de microrganismos como agentes de biocontrole de doenças de plantas, apesar de muito estudado ultimamente, ainda precisa ser melhor compreendido, para alcançarmos sua eficiência ideal. / The state of Tocantins has been standing out in Brazil as a big soybean producer. However, there are many factors that contribute for the reduction of soybean productivity; the main ones are: losses due to diseases and decrease of planting booth. The utilization of biological control constitutes an interesting and important strategy to enable the reduction or substitution of the use of pesticides. Fungi such as Trichoderma and Purpureocillium are important antagonists, in soil, against root diseases and phytopathogenic nematodes. Several products based on Trichoderma and Purpureocillium are worldwide commercialized. Thus, this study aimed to evaluate the effects of the inoculation of Trichoderma and Purpureocillium on soybean crop at different regions (Gurupi, Porto Nacional, Alvorada and Crixás) in Tocantins. The experiments were performed under field conditions at different cities, which produce soybean in Tocantins, using the products TrichoPlus and TrichoMix formulated based on Trichoderma (Trichoderma asperellum UFT 201) and Purpureocillium lilacinum at different dosages, aiming biocontrol for booth maintenance, and thus, also aiming increase in productivity. The inoculation of Trichoderma and Purpureocillium present in the products TrichoPlus and TrichoMix influenced positively the booth maintenance, soybean productivity and helped the growth promotion at all places assessed. In Gurupi, in experiments I and II, not only the isolated application of Trichoderma (TrichoPlus) but also the joint application of Trichoderma and Purpureocillium (TrichoMix) promoted increases in booth maintenance and higher productivity values in relation to other treatments. In Porto Nacional, the inoculation of products TrichoPlus and TrichoMix influenced positively the booth maintenance and productivity of precocious, intermediate and late soybean cultivars. In Alvorada, the inoculation of Trichoderma (Trichoplus) in soybean of first and second year increased the survival index of plants, but did not affect significantly the productivity. In Crixás, the inoculation of Trichoderma (Trichoplus) promoted increases in booth maintenance and also higher productivity values. The use of microorganisms as biocontrol agents of plant diseases, despite being more studied lately, still need to be better comprehended, in order to achieve its optimal efficiency.

Submitted by Luciana Ferreira (lucgeral@gmail.com) on 2018-03-26T14:57:06Z No. of bitstreams: 2 Dissertação - Thuana Marcolino Mota Rodrigues - 2018.pdf: 1461884 bytes, checksum: 1101273815dddbd06af73157343c0ccb (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) / Approved for entry into archive by Luciana Ferreira (lucgeral@gmail.com) on 2018-03-26T14:57:32Z (GMT) No. of bitstreams: 2 Dissertação - Thuana Marcolino Mota Rodrigues - 2018.pdf: 1461884 bytes, checksum: 1101273815dddbd06af73157343c0ccb (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) / Made available in DSpace on 2018-03-26T14:57:32Z (GMT). No. of bitstreams: 2 Dissertação - Thuana Marcolino Mota Rodrigues - 2018.pdf: 1461884 bytes, checksum: 1101273815dddbd06af73157343c0ccb (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Previous issue date: 2018-02-16 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES / The genus Trichoderma ssp, is today the group of species most commonly studied and used to act in agriculture as biological control against phytopathogens. Its rapid mycelial growth, associated with high production of conidia, synthesis of several antibiotics and ability to live in different forms (saprotrophic, symbiont or mycoparasite) are characteristics that make it attractive for this purpose. The relation of mycoparasitism, as well as oscillations in environmental conditions, naturally causes cell stress and consequent response to the stressing agent. This response occurs through changes in cellular metabolism, activating its defense mechanisms that include the performance of heat shock proteins (Hsp). The objective of this work was to identify the Hsp70 family of Trichoderma asperellum and to analyze the expression of three genes encoding proteins of this family during mycoparasitism and in situations of thermal and ethanol stress. The identification of the T. asperellum Hsp70 proteins was possible from the database analysis (JGI) of the T. asperellum genome, available, but not annotated. We identified a total of 12 proteins from the Hsp70 family in T. asperellum, three of which were selected for gene expression assays. Paired cultivation was carried out between T. asperellum and phytopathogens: Sclerotinia sclerotiorum and Fusarium oxysporum in three phases of mycoparasitism: pre-contact, contact and post-contact. We verified that the expression of Tahsp70a, Tahsp70b and Tahsp70c genes varies according to the phases of the mycoparasitism and the phytopathogen studied. Expression of hsp70 under thermal stress was evaluated at the 38 °C condition for 30 minutes, 1, 2 and 4 hours and at the conditions of 4, 10 and 32 °C for 1 hour. Ethanol stress was also performed for 1 hour. During the thermal stress Tahsp70a and Tahsp70b presented higher induction and the Tahsp70c gene had its highest induction in the cold shock at 4 °C. In ethanol stress the analyzed genes did not present significant expression. / O gênero Trichoderma ssp, constitui hoje o grupo de espécies de fungos mais estudadas e comumente usadas para atuar na agricultura como controle biológico contra fitopatógenos. Seu rápido crescimento micelial, associado a alta produção de conídios, síntese de diversos antibióticos e capacidade de viver de diversas formas (saprotrófica, simbionte ou micoparasita) são características que o tornam atraente para esse fim. A relação de micoparasitismo, bem como as oscilações nas condições ambientais, geram naturalmente estresse sobre as células desse fungo e consequente resposta ante o agente estressante. Esta resposta ocorre através de mudanças no metabolismo celular, ativando seus mecanismos de defesa que incluem o desempenho de proteínas de choque térmico (Hsp). O objetivo deste trabalho foi identificar a família Hsp70 de Trichoderma asperellum e analisar a expressão de três genes codificando proteínas desta família durante o micoparasitismo e em situações de estresse térmico e estresse por etanol. A identificação das proteínas Hsp70 de T. asperellum foi possível a partir de análises em banco de dados (JGI) do genoma de Trichoderma asperellum, disponível, mas não anotado. Identificamos no total 12 proteínas da família Hsp70 em T. asperellum, sendo três selecionadas para ensaios de expressão gênica. Foi realizado cultivo pareado entre T. asperellum e os fitopatógenos: Sclerotinia sclerotiorum e Fusarium oxysporum em três fases do micoparasitismo: pré-contato, contato e pós-contato. Verificamos que a expressão dos genes Tahsp70a, Tahsp70b e Tahsp70c varia de acordo com as fases do micoparasitismo e do fitopatógeno estudado. A expressão dos genes hsp70 em estresse térmico, foi avaliada na condição de 38 °C durante 30 minutos, 1, 2 e 4 horas e nas condições de 4, 10 e 32 °C por 1 hora, O estresse por etanol também foi realizado por 1 hora. Durante o estresse térmico, Tahsp70a e Tahsp70b apresentaram maior indução sendo que o gene Tahsp70c teve sua maior indução no choque frio à 4 °C. No estresse por etanol os genes analisados não apresentaram expressão significativa.

Fungos da espécie Trichoderma harzianum são estudados como agentes de controle biológico de fitopatógenos que habitam o solo, sendo sua estratégia um processo complexo que pode ocorrer por diferentes mecanismos ou uma combinação deles, incluindo micoparasitismo. Neste trabalho, foram analisadas consequências da deleção do gene que codifica a proteína Epl-1, descrita como elicitora de resistência em plantas nos processos de: modulação da expressão de genes envolvidos no processo de micoparasitismo de T. harzianum contra o fungo fitopatogênico Sclerotinia sclerotiorum; estabelecimento do processo de micoparasitismo; interação entre T. harzianum e plantas de feijão comum. Os resultados mostram que a deleção do gene epl-1 alterou a modulação da expressão de todos os genes relacionados ao processo de micoparasitismo analisados. Além disso, Epl-1 apresentou um papel no processo de modulação de expressão desses genes em T. harzianum, no reconhecimento da própria parede celular e da parede de diferentes hospedeiros, na interação com fitopatógenos hospedeiros, no processo de formação de estruturas de micoparasitismo, bem como na indução de resistência local e sistêmica em plantas de feijão comum / Fungal species Trichoderma harzianum, are studied as soilborne plant pathogens biological control agent and its strategy is a complex process, which can occur by different mechanisms or a combination thereof, including mycoparasitism. In this work, the consequences of epl-1 gene deletion, which encoding for Epl-1 protein, described as plants resistance elicitor, were analyzed on: modulation of gene expression involved in T. harzianum mycoparasitism process against phytopathogenic fungus Sclerotinia sclerotiorum; mycoparasitism establishment process and T. harzianum interaction with bean plants. The results showed that epl-1 encoding gene deletion altered the expression modulation of all analyzed genes related to mycoparasitism process. Furthermore, Epl-1 presented a role in modulating expression of these genes in the process of self and host cell wall recognition; the interaction with host plant pathogens in formation of mycoparasitism structures as well as local and systemic resistance in beans plants

The soil-borne filamentous fungus Trichoderma virens is a biocontrol agent with a well known ability to produce antibiotics, parasitize pathogenic fungi and induce systemic resistance in plants. Here we report the identification, purification and characterization of an elicitor secreted by T. virens; a small protein designated Sm1 (small protein 1). Confrontation and disk assays demonstrated that Sm1 lacks toxic activity against plants and microbes. Native, purified Sm1 triggers production of reactive oxygen species in rice (Oryza sativa) and cotton (Gossypium hirsutum), and induces the expression of defense related genes both locally and systemically in cotton. Gene expression analysis revealed that SM1 is expressed throughout fungal development and is transcriptionally regulated by nutrient conditions and the presence of a host plant. When T. virens was co-cultured with cotton in an axenic hydroponic system, SM1 expression and secretion of the protein was significantly higher than when the fungus was grown alone. These results indicate that Sm1 is involved in plant-Trichoderma recognition and the induction of resistance by activation of plant defense mechanisms. Following the cloning of SM1, strains disrupted in or over-expressing SM1 were generated. Targeted gene disruption revealed that SM1 was not involved in fungal development. Expression of defense related genes in cotton and maize (Zea mays) was induced locally and systemically following colonization by T. virens in the hydroponic system. Low levels of expression of cotton or maize defense genes were found when seedlings were grown with a T. virens strain disrupted in SM1, ssupporting the Sm1-elicitor hypothesis. Additionally, unique proteins in T.virens-cotton/maize interaction were identified. Thus, the induction of defense responses in two agriculturally important crops appears to be microbially mediated. Functional analysis of a cell wall degrading enzyme, beta-1,6-glucananse (Tv-bgn3) from T. virens, demonstrated involvement of this enzyme indirectly in mycoparasitic activity of T. virens. Protein extracts from the strain disrupted in TV-BGN3 displayed reduced capability to inhibit growth of Pythium ultimum as compared to the wild-type. Additionally, protein extracts from the strains co-expressed with TV-BGN2 (beta-1,3-glucananse) from T. virens showed a significantly increased capability to inhibit growth of P. ultimum and Rhizoctonia solani hyphae.

The soil-borne filamentous fungus Trichoderma virens is a biocontrol agent with a well known ability to produce antibiotics, parasitize pathogenic fungi and induce systemic resistance in plants. Here we report the identification, purification and characterization of an elicitor secreted by T. virens; a small protein designated Sm1 (small protein 1). Confrontation and disk assays demonstrated that Sm1 lacks toxic activity against plants and microbes. Native, purified Sm1 triggers production of reactive oxygen species in rice (Oryza sativa) and cotton (Gossypium hirsutum), and induces the expression of defense related genes both locally and systemically in cotton. Gene expression analysis revealed that SM1 is expressed throughout fungal development and is transcriptionally regulated by nutrient conditions and the presence of a host plant. When T. virens was co-cultured with cotton in an axenic hydroponic system, SM1 expression and secretion of the protein was significantly higher than when the fungus was grown alone. These results indicate that Sm1 is involved in plant-Trichoderma recognition and the induction of resistance by activation of plant defense mechanisms. Following the cloning of SM1, strains disrupted in or over-expressing SM1 were generated. Targeted gene disruption revealed that SM1 was not involved in fungal development. Expression of defense related genes in cotton and maize (Zea mays) was induced locally and systemically following colonization by T. virens in the hydroponic system. Low levels of expression of cotton or maize defense genes were found when seedlings were grown with a T. virens strain disrupted in SM1, ssupporting the Sm1-elicitor hypothesis. Additionally, unique proteins in T.virens-cotton/maize interaction were identified. Thus, the induction of defense responses in two agriculturally important crops appears to be microbially mediated. Functional analysis of a cell wall degrading enzyme, beta-1,6-glucananse (Tv-bgn3) from T. virens, demonstrated involvement of this enzyme indirectly in mycoparasitic activity of T. virens. Protein extracts from the strain disrupted in TV-BGN3 displayed reduced capability to inhibit growth of Pythium ultimum as compared to the wild-type. Additionally, protein extracts from the strains co-expressed with TV-BGN2 (beta-1,3-glucananse) from T. virens showed a significantly increased capability to inhibit growth of P. ultimum and Rhizoctonia solani hyphae.

Fungos da espécie Trichoderma harzianum são estudados como agentes de controle biológico de fitopatógenos que habitam o solo, sendo sua estratégia um processo complexo que pode ocorrer por diferentes mecanismos ou uma combinação deles, incluindo micoparasitismo. Neste trabalho, foram analisadas consequências da deleção do gene que codifica a proteína Epl-1, descrita como elicitora de resistência em plantas nos processos de: modulação da expressão de genes envolvidos no processo de micoparasitismo de T. harzianum contra o fungo fitopatogênico Sclerotinia sclerotiorum; estabelecimento do processo de micoparasitismo; interação entre T. harzianum e plantas de feijão comum. Os resultados mostram que a deleção do gene epl-1 alterou a modulação da expressão de todos os genes relacionados ao processo de micoparasitismo analisados. Além disso, Epl-1 apresentou um papel no processo de modulação de expressão desses genes em T. harzianum, no reconhecimento da própria parede celular e da parede de diferentes hospedeiros, na interação com fitopatógenos hospedeiros, no processo de formação de estruturas de micoparasitismo, bem como na indução de resistência local e sistêmica em plantas de feijão comum / Fungal species Trichoderma harzianum, are studied as soilborne plant pathogens biological control agent and its strategy is a complex process, which can occur by different mechanisms or a combination thereof, including mycoparasitism. In this work, the consequences of epl-1 gene deletion, which encoding for Epl-1 protein, described as plants resistance elicitor, were analyzed on: modulation of gene expression involved in T. harzianum mycoparasitism process against phytopathogenic fungus Sclerotinia sclerotiorum; mycoparasitism establishment process and T. harzianum interaction with bean plants. The results showed that epl-1 encoding gene deletion altered the expression modulation of all analyzed genes related to mycoparasitism process. Furthermore, Epl-1 presented a role in modulating expression of these genes in the process of self and host cell wall recognition; the interaction with host plant pathogens in formation of mycoparasitism structures as well as local and systemic resistance in beans plants

Soybean crops are vulnerable to a wide range of pathogens that reduce yield and cause extensive losses worldwide. In the United States, the soilborne pathogens Pythium spp., causing soybean damping-off, and Fusarium virguliforme, causing sudden death syndrome (SDS) of soybean, have been among the top diseases that most reduced soybean yields. This study demonstrated that biological control using native fungal antagonists could be a powerful tool to integrate with current management strategies for more efficient control of Pythium damping-off and SDS in soybean. Trichoderma spp. and Clonostachys rosea demonstrated the ability to mycoparasitize and antagonize the pathogens using different mechanisms and exhibited a protective effect on soybean in field conditions. The development of an efficient biological control program for disease management relies on a deep understanding of the BCA-pathogen interaction’s biology. This research also uncovered the molecular mechanisms involved in the F. virguliforme-T. afroharzianum interaction by using a dual RNAseq approach. Significant changes in both fungal organisms’ transcriptomes were discovered at different stages in their interaction. The results provided here can contribute to the future implementation of effective biological control programs for soybean. The benefits may also extend to other crops.

Pucciniomycotina is a subphylum with a high diversity in terms of habitat and life history strategies that include plant parasites, animal associates (including opportunistic human pathogens), saprobes and antagonists of other fungi. The class Cystobasidiomycetes within this subphylum is a representative of such diversity and remains understudied. Their role in nature and the associations they establish with their hosts for most of the species is still unknown. In this study we used taxonomic and phylogenetic methods to present an inventory of strains in this class collected for more than 20 years from all over the world and preserved in the Aime Lab Culture Collection. Molecular and morphological data for six new species in the genera <i>Bannoa, Buckleyzyma</i>, <i>Halobasidium</i> and <i>Sakaguchia </i>were also presented. Additionally, mating experiments were performed by pairing strains of <i>Bannoa</i>; this is the second time in the genus that clamp connections and basidia are observed. We also found that newly described species of <i>Bannoa</i> can be co-inhabitants of sori of rust fungi. In general, antagonistic interactions can occur through: 1) direct physical contact between two fungi, i.e., mycoparasitism; or, 2) the production of antimicrobial compounds. In the Cystobasidiomycetes, direct physical antagonistic interaction which is associated with sexual states has been reported in species of Cystobasidium, Naohidea, Cyphobasidium and Occultifur. On the other hand, the production of antimicrobial compounds which mainly occurs between the yeast stage of the fungi and other organisms has only been reported in Cystobasidium pallidumand Hasegawazyma lactosa. We also hypothesize that the common ancestor to Cystobasidiomycetes is a mycoparasite due to the fact that this life strategy is present in most of the lineages in the class. To test this hypothesis, we grouped into five categories the host association or substrate from which strains of extant species in the Cystobasidiomycetes were isolated, i.e.,animals, plants, fungi, aquatic or decaying organic matter. We constructed a resolved phylogeny for the class based on seven locito study the evolutionary origins of mycoparasitism through ancestral character reconstruction with representation of all described species. Our analysis suggests that the most likely hypothesis is that the most recent common ancestor of the Cystobasidiomycetes was associated to fungi.<br>

Submitted by Marlene Santos (marlene.bc.ufg@gmail.com) on 2014-10-10T21:54:56Z No. of bitstreams: 2 Tese - Saulo José Linhares de Siqueira - 2012.pdf: 3918109 bytes, checksum: cfb7931590a50ed74838d6c8cefab1ee (MD5) license_rdf: 23148 bytes, checksum: 9da0b6dfac957114c6a7714714b86306 (MD5) / Approved for entry into archive by Jaqueline Silva (jtas29@gmail.com) on 2014-10-13T20:44:05Z (GMT) No. of bitstreams: 2 Tese - Saulo José Linhares de Siqueira - 2012.pdf: 3918109 bytes, checksum: cfb7931590a50ed74838d6c8cefab1ee (MD5) license_rdf: 23148 bytes, checksum: 9da0b6dfac957114c6a7714714b86306 (MD5) / Made available in DSpace on 2014-10-13T20:44:05Z (GMT). No. of bitstreams: 2 Tese - Saulo José Linhares de Siqueira - 2012.pdf: 3918109 bytes, checksum: cfb7931590a50ed74838d6c8cefab1ee (MD5) license_rdf: 23148 bytes, checksum: 9da0b6dfac957114c6a7714714b86306 (MD5) Previous issue date: 2012-03-30 / Species of Trichoderma are commercially applied as biological control agents and are antagonists of important plant pathogenic fungi (as Rhizoctonia, Sclerotinia and Fusarium species) due to its mycoparasitic characteristics. Research has been performed to have a better comprehension of molecular aspects of the biocontrol mechanisms performed by Trichoderma and to find isolates with high antagonistic potential against plant pathogens. In the present study the expression of mycoparasitism-related genes was performed T. asperellum T00 and T. harzianum ALL42 (Enzimologia group ICB/UFG fungal collection) that have great potential as biocontrol agent. Each chapter of this work refers to one of the species studied. In Chapter 1 T. asperellum isolate T00, known to produce high levels of cell wall degrading enzymes, has its β-1,3-glucanases enzymes and genes (tag83 and tag27) studied. The gene tag27 was cloned and characterized and codes to an 27kDa endo-β-1,3glucanase with and 285 aminoacids and 96% similar to a glucanases from T.atroviride. The enzyme was detected when T. asperellum was grown in Rhizoctonia solani or Sclerotinia sclerotiorum cell wall-containing media but not in Fusarium oxysporum cell wall-containing media. The tag83 and tag27 genes was repressed in media containing glucose as carbon source and upregulated in cell wall containing media and during plate confrontation tests with pathogenic fungi. Chapter 2 shows T. harzianum isolate ALL42 genes involved in mycoparasitism against R. solani or S. sclerotiorum detected using subtractive hybridization approach. T. harzianum was grown with R. solani or S. sclerotiorum cell wall as carbon source and the RNA used both as tester and driver in each of two subtractive library constructed. Sequencing analysis resulted in 47 genes related with growth in R. solani cell wall media and 114 genes related with growth in S. sclerotiorum cell wall media. To confirm the obtained data, 18 genes were tested by quantitative real time RT-PCR and 9 were differentially expressed in the same condition of the library they were detected. Five of these genes were also differentially expressed during plate confrontation assay with the respective pathogen, two of them expressed during contact with R. solani (cutinase and alginate lyase) and 3 during contact with S. sclerotiorum (hsp98, serin endopeptidase and a hypotetic gene). The results presented in this study provides additional information about the role of 1,3glucanase genes in mycoparasitism and of other genes related to antagonism against specific pathogens, providing helpful insights in the mechanism of biocontrol performed by Trichoderma. / Espécies do gênero Trichoderma são eficientes antagonistas de fungos fitopatogênicos, como as espécies Rhizoctonia, Sclerotinia e Fusarium, e são comercializados como agentes de controle biológico principalmente por sua característica de micoparasita. Muitos estudos têm sido feitos para compreender as bases moleculares dos mecanismos de biocontrole de Trichoderma e também para encontrar espécies com alto potencial de antagonismo contra fitopatógenos. O objetivo deste trabalho foi analisar a expressão de genes relacionados ao micoparasitismo de T. asperellum T00 e T. harzianum ALL42 (Enzimologia ICB/UFG) que possuem potencial para uso como agente de biocontrole. Este trabalho foi dividido em dois capítulos. O Capítulo 1 se refere ao fungo T. asperellum T00 e suas β-1,3-glicanases (TAG83 e TAG27) que degradam componentes da parede celular de fungos fitopatógenos. O gene tag27 codifica para uma endo-β-1,3glicanase de 27kDa que possui 285 resíduos de aminoácidos e apresentou 96% de similaridade com uma enzima de T. atroviride. A enzima foi secretada em meio de cultura contendo parede celular de Rhizoctonia solani e Sclerotinia sclerotiorum, mas não em meio com parede de Fusarium oxysporum. A expressão dos genes tag83 e tag27 foi reprimida na presença de glicose e ativada tanto na presença de parede celular dos fitopatógenos quanto durante o contato entre os fungos em placa. O Capítulo 2 trata do fungo T. harzianum isolado ALL42 e de genes identificados pela técnica de hibridização subtrativa relacionados especificamente ao biocontrole contra R. solani e S. sclerotiorum. Foram construídas duas bibliotecas subtrativas sendo que os RNAs utilizados como condição teste e controle da subtração foram obtidos de T. harzianum crescido em meio contendo parede celular de R. solani ou S. sclerotiorum. As bibliotecas foram sequenciadas resultando em 47 genes relacionados ao crescimento em meio com R. solani e 114 genes relacionados ao crescimento em meio com S. sclerotiorum. Dos 18 genes escolhidos para validação da biblioteca por RT-PCR em tempo real, 9 se mostraram diferencialmente expressos na condição correspondente à biblioteca em que foram identificados. Dentre estes, 5 genes se mostraram também diferencialmente expressos em experimento de confronto em placa, 2 deles mais expressos contra R. solani (cutinase e alginato liase) e 3 contra S. sclerotiorum (hsp98, serina endopeptidase e um de função hipotética). Os resultados obtidos com as duas linhagens fornecem informações adicionais sobre genes de β-1,3-glicanases, conhecidamente envolvidos no processo de micoparasitismo, e sobre genes relacionados ao antagonismo de patógenos específicos, além de contribuir para o conhecimento relacionado ao biocontrole realizado por fungos do gênero Trichoderma.

Made available in DSpace on 2014-06-11T19:32:55Z (GMT). No. of bitstreams: 0 Previous issue date: 2010-08-25Bitstream added on 2014-06-13T20:04:39Z : No. of bitstreams: 1 simoes_mlg_dr_rcla.pdf: 7383499 bytes, checksum: 3bdb988e95e0edaf3bc9450a229443f5 (MD5) / Diferentes isolados de Trichoderma (T. stromaticum, T. pseudokoningii, T. harzianum, e T. viride) e um isolado de T. atroviride, T. longibrachiatum, T. virens e T. pilulliferum foram estudados para uso no biocontrole de quatro subgrupos genéticos de Moniliophthora perniciosa, designados como Mp1441, Mp1445, Mp1893 e Mp1916. A associação das características dos isolados de Trichoderma avaliados quanto à velocidade de crescimento micelial, germinação, produção de esporos e produtividade massal de esporos em arroz, além das suas capacidades de antibiose e micoparasitismo a cada subgrupo do patógeno, resultou na determinação do índice denominado Potencial para uso no biocontrole (%PCB) a cada subgrupo de M. perniciosa. Os %PCBs variaram, demonstrando haver variabilidade genética entre os isolados do antagonista quanto às características avaliadas, como também nos níveis de resistência do patógeno. T. harzianum 911 apresentou o melhor %PCB para todos os subgrupos de M. perniciosa, sendo o pior apresentado por T. reesei 1612. Mp1445 foi o subgrupo que apresentou a menor capacidade de supressão aos antagonistas, sendo a maior apresentada por Mp1916. T. harzianum 906 inibiu o crescimento de Mp1916 por metabólitos não voláteis e todos os isolados de Trichoderma avaliados inibiram todos os subgrupos do patógeno pela ação de metabólitos voláteis, variando, porém, os percentuais de inibição entre as espécies e entre isolados da mesma espécie. Mp1445 inibiu o crescimento de T. virens 2007 por metabólitos não voláteis, sendo comprovada, portanto, a possibilidade do patógeno inibir o antagonista. T. harzianum 911 apresentou capacidade de micoparasitismo em placa e em vassouras infectadas individualmente pelos subgrupos do patógeno, além de produzir as enzimas β-1,3-glucanase, quitinase, protease, FPase, CMCase e amilase, tanto em substratos específicos como em cultivos... / Different isolates of Trichoderma (T. stromaticum, T. pseudokoningii, T. harzianum and T. viride) and one isolate of T. atroviride, T. longibrachiatum, T. virens e T. pilulliferum were studied for their usage in biocontrol of four genetic subgroups of Moniliophthora perniciosa named as Mp1441, Mp1445, Mp1893 e Mp1916. The association of the characteristics of the Trichoderma isolates evaluated regarding to velocity of mycelial growth, germination, spores production in Petri dish and mass production of spores in rice, besides their capacities of antibiosis and mycoparasitism to each subgroup of the pathogen, resulted in the determination of the index named Potential to be used in Biocontrol (%PCB) to each subgroup of M. perniciosa. The %PCBs varied, pointing out that there is a genetic variability among the antagonistic isolates regarding to the evaluated characteristics, as well as the levels of the pathogen resistance. T. harzianum 911 showed the best %PCB for all subgroups of M. perniciosa, being the worst presented by T. reesei 1612. Mp1445 was the subgroup that presented the lowest antagonistic suppression capacity, being the highest presented by Mp1916. T. harzianum 906 inhibited the growth of Mp1916 by non-volatile metabolites and all the evaluated isolates of Thrichoderma inhibited all the subgroups through volatile metabolites, varying, although, the percentages of inhibition among the species and among the isolates of the same species. Mp1445 inhibited the growth of T. virens 2007 by non-volatile metabolites, being proved, therefore, the possibility of the pathogen to inhibit the antagonist. T. harzianum 911 presented the mycoparasitism capacity in Petri dishes and in brooms infected indiviadually by the pathogen, besides the production of β-1,3-glucanase, chitinase, protease, FPase, CMCase and amilase, in both specific substrates and in cultivations with dried mycelia... (Complete abstract click electronic access below)