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Orientador: Edson Luiz Furtado / Banca: Cesar Junior Bueno / Banca: Marli de Fátima Stradioto Papa / Resumo: A cultura da seringueira [Hevea brasiliensis (Willd. ex Adr. Jussieu) Muell. Arg.] vem sendo atacada por várias doenças de importância econômica, dentre as quais está a antracnose, causada pelo fungo Colletotrichum sp. (teleomorfo: Glomerella sp.). Este fungo causa vários danos na planta como lesões nos folíolos, nos ponteiros, nos ramos, nos frutos e cancros no painel de sangria. Somente Colletotrichum gloeosporioides foi relatado como agente causal desta doença no Brasil. O presente trabalho teve como objetivo identificar isolados de Colletotrichum spp. de seringueira de diversas regiões de plantio do Estado de São Paulo. O trabalho foi conduzido em cinco fases: a) caracterização cultural, na qual foram observadas a coloração e o aspecto das culturas in vitro. Produção de conídios e taxa de crescimento em seis temperaturas (10, 15, 20, 25, 30 e 35 °C); b) caracterização morfológica, na qual foi medido comprimento, largura e observado o formato dos conídios; c) teste de patogenicidade em folíolos destacados e em discos de folíolos, com quatro isolados de seringueira e dois de citros; d) crescimento em benomyl em quatro concentrações de princípio ativo; e) Identificação molecular para culturas monospóricas e multispóricas com primers específicos para as espécies de Colletotrichum gloeosporioides e Colletotrichum acutatum e os primers ITS1 e ITS4 os quais amplificaram uma pequena região (18S) e uma grande região (28S), e estes também permitiram a amplificação da região 5.8S do rDNA e os espaçadores internos transcritos (ITS1 e ITS2), e f) testes de crescimento em meio de cultivo acrescido com fungicidas: flutriafol, tebuconazol, epoxiconazol + piradostrobina, clorotalonil + tiofonato-metílico, captana, mancozebe, carbendazim, azoxistrobina + ciproconazol e propiconazol. Neste teste foram utilizados quatro isolados de diferentes órgãos da planta... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: The rubber cultivation [Hevea brasiliensis (Willd. ex Adr. Jussieu) Muell. Arg.] is being attacked by several diseases of economic importance, among which is the anthracnose; this is caused by the fungus Colletotrichum spp. (teleomorph: Glomerella spp). This fungus causes several damages in the plant such as injuries in the leaves, branches, fruits and cankers in the taping panel. Only Colletotrichum gloeosporioides was reported as the causal agent of this disease in Brazil. The aim of this study was to identify isolates of the fungus Colletotrichum spp. from rubber trees, localized in different regions of Sao Paulo state. The study was carried out by six phases: a) culture characterization, in which the color and the culture appearance were observed in vitro, conidial production and growth rate at six temperatures (10, 15, 20, 25, 30, 35 °C); b) morphological characterization, which consist of measuring the length and width, and observed the shape of the conidia; c) pathogenicity test on selected leaves and disks of leaves, with four isolates from rubber and two isolates from citrus; d) growth in fungicide benomyl at four concentrations of active ingredient; e) molecular identification for monosporic and multisporic cultures with specific primers to the species of Colletotrichum gloeosporioides and Colletotrichum acutatum and ITS1 and ITS4 primers which amplified a small region (18S) and a large region (28S), and these also allowed the amplification of 5.8S rDNA and internal transcribed spacers (ITS1 and ITS2); f) Growth tests in culture medium supplemented with fungicides: flutriafol, tebuconazole, epoxiconazole + piradostrobina, chlorothalonil + tiofonato-methyl, captan, mancozeb, carbendazim, azoxystrobin + cyproconazole and propiconazole. In this test, four isolates were used from... (Complete abstract click electronic access below) / Mestre

Submitted by (lucia.rodrigues@ufrpe.br) on 2017-02-21T13:02:56Z No. of bitstreams: 1 Beatriz Meireles Barguil.pdf: 866270 bytes, checksum: c8c6c11db8a2c4bc261affc072520e23 (MD5) / Made available in DSpace on 2017-02-21T13:02:56Z (GMT). No. of bitstreams: 1 Beatriz Meireles Barguil.pdf: 866270 bytes, checksum: c8c6c11db8a2c4bc261affc072520e23 (MD5) Previous issue date: 2006-11-23 / Conselho Nacional de Pesquisa e Desenvolvimento Científico e Tecnológico - CNPq / The tropical flower crop has increased in several states of the Northeast region of Brazil based on the high acceptance of the product and it potencial as a profitable activity. The anthracnose caused by Colletotrichum spp. can affects many of these species influencing negatively the quantity and quality of the final product. The present work aimed: a) to characterize Colletotrichum isolates from tropical flowers by morphological, exoenzimes production, pathogenicity and molecular methods; b) to develop a diagrammatic key for anthracnose on torch ginger; c) to evaluate the response of anthurium cultivars, the effect of inoculation point quantity, and the different development stages on anthracnose development. All isolates presented Colletotrichum gloeosporioides characteristics, by morphological and molecular methods, excepted the DNA from C 23 and C 35 isolates that weren`t amplified with the specific primer. The conidia from the different isolates were hyaline, unicellular and cilindrical, and apressoria varied in shape and size, with dark brown color. The colony color varied from white to dark gray. All isolates showed enzymes activity on specific mediums. Thepathogenicity was variably between isolates. Six anthurium`s isolates and four heliconia`s isolates weren`t pathogenic when inoculated on their hosts. All isolates from torch ginger were pathogenic. Some of them showed cross-infection capacity. Genetic variability was verified by ap-PCR with three different primers. The diagrammatic key elaborated to evaluate anthracnose on torch ginger includ the levels 1, 2, 4, 8, 16, 32, 64, 82 and 92% of diseased bracts area. On the first evaluation without the key raters overestimated and underestimate the disease severity. With the diagrammatic key raters obtained better levels of accuracy and precision, with absolute errors concentrated around 10%. Raters showed good repeatability and high reproducibility of estimative by using the key compared to the no use of it. The lesion size development was influenced by espathe age on anthurium. On espathes on stage 1, where the esphathes still closed, the lesions were significantly bigger them espathes on 4 and 5 stages. Bigger lesions were observed with five points per wound on the two cultivars (cvs. Tropical and Cananéia) and isolates tested. Inoculation time not influenced the lesion size oncultivars tested. The incubation period (IP) was variable with cultivars and isolate. The smaller IP for Cg 1 were observed on Astral, Tropical, Netuno and Farao. On Sonate,for Cg 1 and on Farao and Midori for Cg . / O cultivo de flores tropicais tem-se expandido em vários estados do Nordeste do Brasil pela grande aceitação desses produtos pelo mercado consumidor e a perspectiva de um agronegócio de elevado retorno econômico. A antracnose causada por Colletotrichum spp. pode afetar várias espécies tropicais, comprometendo a quantidade e qualidade do produto final. Devido à carência de informações nesses patossistemas, o presente trabalho objetivou: a) Caracterizar através de morfologia, produção de exoenzimas, patogenicidade e análise molecular de isolados de Colletotrichum de flores tropicais; b) Elaborar uma escala diagramática para avaliação da severidade da antracnose em bastão do imperador; e c) Avaliar o comportamento de cultivares de antúrio, em diferentes fases de desenvolvimento e o efeito da quantidade de pontos de inoculação na severidade da antracnose quando infectadas com C. gloeosporioides. Todos os 37 isolados testados produziram conídios hialinos, unicelulares, retos e cilíndricos e apressórios de coloração marrom escuro e formato variável. A coloração das colônias variou de branco a cinza escuro. O DNA da maioria dos isolados foi amplificado com o oligonucleotídeo específico para a espécie C. gloeosporioides, confirmando os dados obtidos através das características morfológicas. O DNA dos isolados C 23 e C 35foi amplificado com os primers espécie-específicos para a confirmação da espécie. Todos os isolados produziram enzimas amilolíticas, lipolíticas e proteolíticas em meios de cultura específicos. A patogenicidade dos isolados foi variável. Seis isolados obtidos de antúrio e quatro isolados de helicônia não foram patogênicos quando inoculados nos respectivos hospedeiros, sendo que todos os isolados provenientes de bastão do imperador foram patogênicos. Alguns isolados de antúrio, bastão do imperador, helicônia e musa foram patogênicos quando em inoculações cruzadas. A variabilidade genética dos 37 isolados de Colletotrichum foi determinada com três oligonucleotídeos arbitrários. A escala diagramática elaborada para a antracnose em bastão do imperador apresenta níveis de 1, 2, 4, 8, 16, 32, 64, 82 e 92% de área lesionada da bráctea. A avaliação da severidade da antracnose em bastão do imperador sem a utilização dessa escala gerou resultados contraditórios onde os avaliadores tanto superestimaram como subestimaram a intensidade da doença. Com o emprego da escala, os avaliadores obtiveram melhores níveis de acurácia e precisão, com os erros absolutos concentrandose na faixa de 10%. Os avaliadores apresentaram boa repetibilidade e elevada reprodutibilidade das estimativas com a utilização da escala, o mesmo não sendo verificado sem a utilização dessa. Em antúrios, a idade da espata influenciou o desenvolvimento da lesão para os dois isolados avaliados. Em espatas no estádio 1, sem a completa abertura, as lesões formadas foram significativamente maiores do que aquelas formadas em espatas nos estádios 4 e 5. As maiores lesões causadas pelos isolados do patógeno foram formadas com cinco pontos de ferimento nas duas cultivares de antúrio (cvs. Tropical e Cananéia). A época de inoculação não influenciou no tamanho da lesão, nas duas cultivares avaliadas. Os menores períodos de incubação (PI) para o isolado Cg 1 foram observados nas cultivares Astral, Tropical, Netuno e Farao, diferindo significativamente nas demais cultivares. Nas cultivares Sonate, Astral, Tropical, Netuno e Farao foram observados os menores PI para o isolado Cg 2. A menor AACPD resultante da inoculação com Cg 1 foi observada em Sonate, que diferiu das demais cultivares. As menores AACPD ocasionadas pelo isolado Cg 2 foram observadas em Farao e Midori, que diferiram das cultivares Cananéia, Sonate, Tropical e Netuno.

Orientador: Ângela Regina Araújo. / Banca: Dulce Helena Siqueira Silva / Banca:Lourdes Campaner dos Santos / Banca:Jairo Kennup Bastos / Banca:Luce Maria Brandão Torres / Resumo: Os fungos, diferente dos vegetais são heterotróficos, mais especificamente quimiorganotróficos, por suas características específicas pertencem a um reino separado. Alguns dos fungos através de sua evolução se especializaram em invadir os espaços intercelulares das plantas passando a sobreviver dentro dos seus hospedeiros, não provocando nenhuma patologia. Estes microrganismos são denominados de fungos endofíticos. A disputa por alimentos entre os fungos no interior das espécies vegetais, gerou a necessidade dos fungos produzirem metabolitos secundários com atividades contra outros microrganismos, podendo assim demarcar seu território. Todo este processo deve acontecer de forma a não causar patologias na planta que é a fonte de seu alimento, em alguns casos essas relações de mutualismo e comensalismo se aprimoraram sendo muito comum alguns vegetais possuirem fungos endofíticos protegendo contra pragas insetos e patógenos. Este trabalho descreve o isolamento e a purificação dos fungos endofíticos associados às folhas, caules, raízes e sementes da espécie vegetal Michela champaca. Foram isolados 8 fungos, sendo 3 dos caules, 4 das folhas, 1 das raízes, nenhum fungo foi isolado das sementes. Os extratos brutos produzidos por estes fungos em caldo de dextrose e batata foram avaliados quimicamente (CLAE-DAD e RMN1H) e submetidos à bioensaios para avaliação da potencialidade anticancerígena, antifúngica e anticolinesterásica. Todos os extratos apresentaram atividade contra os fungos fitopatogênicos Cladosporium cladosporioides e Cladosporium sphaerospermum e 40% apresentaram atividade frente às linhagens mutantes de Saccharomyces cerevisiae. Após esta triagem, os fungos endofíticos Mc-8R (ainda não identificado) e Colletotrichum gloeosporioides (Mc-7F) foram selecionados para estudo químico/biológico. / Abstract: Some fungal species, through their evolution, have specialized in invading plants and surviving inside their hosts causing no warm them. These microorganisms have been called endophytic which in the invasion process, produce chemical substances that inhibit competitors to invade their neighborhood and thus survive in harmony with the host. This work seeks to evaluate the chemical diversity of substances from endophytic fungi associated to Michelia champaca L. that present biological activity by isolating and identification of their secondary metabolites. This work resulted in the isolation of eight fungi species and selection of 2 strains which presented strong antifungal activity against Cladosporium cladosporioides and C. sphaerospermum, both of them pathogenic fungus; and a weak antitumor activity against mutant strains of Saccharomyces cerevisiae. The first one was codified as Mc-8R and chromatographic fractionation of its EtOAc extract allowed the identification of the Pirenocines A (1), E (2) and B (3) and the Pirenochaetic acids, C (4), A (5) and B (6), and in addition to the novel (7) and (8). Ergosterol (9), was found in the hexanic extract obtained from growing Mc-8R in rice, and the Ergosterol peroxide (10), which was isolated from the hexanic extract in corn. The other selected fungi species was identified as Colletotrichum gloeosporioides. The fractioning of its EtOAc extract led to the isolation of the known substances as Uracil (11), cyclo-(S*-Pro-S*-Tyr) (12), cyclo-(S*-Pro-S*-Val) (13), 2-Aminophenyl acetic acid (2-APA), (14), p-Hydroxyphenylacetic acid (15), 4-Hydroxybenzamide (16) and (2-Hydroxy-phenyl)- acetic acid (17), in addition to the novel compounds (18) and (19). 2-Hexylidene-3-methylsuccinic acid (20) was also identified as major compound from the fungus Mc-C3, (Xylaria sp.). / Doutor

No description available.

No description available.

The fungus <i>Colletotrichum truncatum</i> is a potential biocontrol agent (BA) against the noxious weed scentless chamomile (<i>Metricaria perforata</i> Mérat; syn.: <i>Tripleurospermum perforatum</i> (Mérat) Lainz) in western Canada. This potential BA, however, is taxonomically related to the anthracnose pathogen on lentil, raising questions about crop safety. Ribosomal DNA (rDNA) internal transcribed space (ITS) regions of <i>C. truncatum</i> isolates collected from different plant hosts were examined, and compared with additional Colletotrichum species. Sequences were amplified with the universal primers its4 and its5, and <i>C. truncatum</i> isolates from scentless chamomile and selected legume crops were differentiated consistently. All scentless chamomile isolates fell within a single cluster in phylogenetic trees, regardless of their geographic origins. These isolates were more closely related to lentil isolates of <i>C. truncatum</i> than to isolates from the other host species. Soybean isolates, with more falcate and slender conidia and slightly bigger appressoria, were distinguishable from other <i>C. truncatum</i> isolates, while the isolates from scentless chamomile, lentil and pea were morphologically more similar. Based on sequence information, strain-specific PCR primers were designed for <i>C. truncatum</i> isolates from these hosts and used to amplify specific DNA bands (markers) from isolates of <i>C. truncatum</i>. This technique may be used for rapid detection and differentiation of <i>C. truncatum,</i> from scentless chamomile and designated legume species, as well as for tracking the BA after release. Inoculation trials were conducted using detached leaves and whole plants to determine potential cross infection of these <i>C. truncatum</i> isolates. Isolates from scentless chamomile caused disease only on their original host, but not on lentil, pea, soybean or alfalfa. In contrast, lentil isolates caused severe disease on lentil and pea, light symptoms on alfalfa, but no disease on the other hosts tested. Potential penetration of lentil leaves by scentless chamomile isolates was tested, with 2-23% incidence of the fungus from inoculated detached, senescence leaves but disease symptoms were not observed on either detached leaves or whole plants. Examination of the infection process revealed that scentless chamomile and lentil isolates had a similar pattern of infection and disease development on their respective hosts; infection vesicles were produced 24 h after inoculation, both primary and secondary infection hyphae were present, and the onset of disease symptoms tended to coincide with the development of secondary hyphae. The current study provided molecular and pathological evidence that differentiates the potential BA of scentless chamomile from <i>C. truncatum</i> isolates from lentil, pea and soybean.

A plant's response to pathogen infection is tailored dependent on infection strategy. Successful plant pathogens employ various infection strategies to avoid or reduce plant defense responses for the establishment of host compatibility. Autophagy is a non-selective degradation pathway conserved in eukaryotic organisms, which has been implicated in the regulation of cell survival or cell death, depending on cell type and stimulus. In <i>Arabidopsis thaliana</i>, an autophagic response has been reported to be activated during nutrient deprivation. Cellular contents, such as cytoplasm and organelles, are sequestered into double-membraned autophagosomes and delivered to the vacuole for degradation; degradative products, such as amino acids, are released back into the cell and reutilized to maintain cellular function. In this study, the response of the autophagy pathway was investigated in <i>A. thaliana</i> leaf tissues upon biotrophic <i>Erysiphe cichoracearum</i> and hemibiotrophic <i>Colletotrichum higginsianum</i> infections. Expression of some autophagy genes was induced in <i>A. thaliana</i> at 9 days post infection with <i>E. cichoracearum</i> and, 3 and 5 days post infection with <i>C. higginsianum</i>. Using a transgenic <i>A. thaliana</i> plant line over expressing autophagosome associated protein autophagy-8e (<i>ATG8e</i>) conjugated to green fluorescent protein (GFP) (<i>ATG8e-GFP</i>), confocal analysis revealed that autophagosomes specifically accumulated at the infection sites during <i>E. cichoracearum</i> and <i>C. higginsianum</i> invasions. These results indicate that the plant autophagic pathway responds to an interaction between <i>A. thaliana</i> and fungal pathogens. None of the defense signaling molecules including salicylic acid, jasmonic acid, ethylene, hydrogen peroxide and nitric oxide consistently triggered expression of autophagy genes. The insensitivity to defense signaling molecules and the delayed induction of autophagy genes compared to expression of pathogenesis-related genes suggest that the activation of this pathway does not contribute to host resistance responses during the infection process. In <i>A. thaliana</i> mutants, <i>atg4a/b, atg5-1, atg9-1</i> and <i>atg9-6</i> deficient for the autophagic response, virulence of <i>E. cichoracearum</i> was retarded whereas pathogenesis of <i>C. higginsianum</i> was accelerated. Taken together, these data suggest that the autophagy pathway is a potential host susceptibility factor for pathogen infection, possibly involved in establishing/facilitating biotrophy in <i>A. thaliana</i>.

The fungus <i>Colletotrichum truncatum</i> is a potential biocontrol agent (BA) against the noxious weed scentless chamomile (<i>Metricaria perforata</i> Mérat; syn.: <i>Tripleurospermum perforatum</i> (Mérat) Lainz) in western Canada. This potential BA, however, is taxonomically related to the anthracnose pathogen on lentil, raising questions about crop safety. Ribosomal DNA (rDNA) internal transcribed space (ITS) regions of <i>C. truncatum</i> isolates collected from different plant hosts were examined, and compared with additional Colletotrichum species. Sequences were amplified with the universal primers its4 and its5, and <i>C. truncatum</i> isolates from scentless chamomile and selected legume crops were differentiated consistently. All scentless chamomile isolates fell within a single cluster in phylogenetic trees, regardless of their geographic origins. These isolates were more closely related to lentil isolates of <i>C. truncatum</i> than to isolates from the other host species. Soybean isolates, with more falcate and slender conidia and slightly bigger appressoria, were distinguishable from other <i>C. truncatum</i> isolates, while the isolates from scentless chamomile, lentil and pea were morphologically more similar. Based on sequence information, strain-specific PCR primers were designed for <i>C. truncatum</i> isolates from these hosts and used to amplify specific DNA bands (markers) from isolates of <i>C. truncatum</i>. This technique may be used for rapid detection and differentiation of <i>C. truncatum,</i> from scentless chamomile and designated legume species, as well as for tracking the BA after release. Inoculation trials were conducted using detached leaves and whole plants to determine potential cross infection of these <i>C. truncatum</i> isolates. Isolates from scentless chamomile caused disease only on their original host, but not on lentil, pea, soybean or alfalfa. In contrast, lentil isolates caused severe disease on lentil and pea, light symptoms on alfalfa, but no disease on the other hosts tested. Potential penetration of lentil leaves by scentless chamomile isolates was tested, with 2-23% incidence of the fungus from inoculated detached, senescence leaves but disease symptoms were not observed on either detached leaves or whole plants. Examination of the infection process revealed that scentless chamomile and lentil isolates had a similar pattern of infection and disease development on their respective hosts; infection vesicles were produced 24 h after inoculation, both primary and secondary infection hyphae were present, and the onset of disease symptoms tended to coincide with the development of secondary hyphae. The current study provided molecular and pathological evidence that differentiates the potential BA of scentless chamomile from <i>C. truncatum</i> isolates from lentil, pea and soybean.

A plant's response to pathogen infection is tailored dependent on infection strategy. Successful plant pathogens employ various infection strategies to avoid or reduce plant defense responses for the establishment of host compatibility. Autophagy is a non-selective degradation pathway conserved in eukaryotic organisms, which has been implicated in the regulation of cell survival or cell death, depending on cell type and stimulus. In <i>Arabidopsis thaliana</i>, an autophagic response has been reported to be activated during nutrient deprivation. Cellular contents, such as cytoplasm and organelles, are sequestered into double-membraned autophagosomes and delivered to the vacuole for degradation; degradative products, such as amino acids, are released back into the cell and reutilized to maintain cellular function. In this study, the response of the autophagy pathway was investigated in <i>A. thaliana</i> leaf tissues upon biotrophic <i>Erysiphe cichoracearum</i> and hemibiotrophic <i>Colletotrichum higginsianum</i> infections. Expression of some autophagy genes was induced in <i>A. thaliana</i> at 9 days post infection with <i>E. cichoracearum</i> and, 3 and 5 days post infection with <i>C. higginsianum</i>. Using a transgenic <i>A. thaliana</i> plant line over expressing autophagosome associated protein autophagy-8e (<i>ATG8e</i>) conjugated to green fluorescent protein (GFP) (<i>ATG8e-GFP</i>), confocal analysis revealed that autophagosomes specifically accumulated at the infection sites during <i>E. cichoracearum</i> and <i>C. higginsianum</i> invasions. These results indicate that the plant autophagic pathway responds to an interaction between <i>A. thaliana</i> and fungal pathogens. None of the defense signaling molecules including salicylic acid, jasmonic acid, ethylene, hydrogen peroxide and nitric oxide consistently triggered expression of autophagy genes. The insensitivity to defense signaling molecules and the delayed induction of autophagy genes compared to expression of pathogenesis-related genes suggest that the activation of this pathway does not contribute to host resistance responses during the infection process. In <i>A. thaliana</i> mutants, <i>atg4a/b, atg5-1, atg9-1</i> and <i>atg9-6</i> deficient for the autophagic response, virulence of <i>E. cichoracearum</i> was retarded whereas pathogenesis of <i>C. higginsianum</i> was accelerated. Taken together, these data suggest that the autophagy pathway is a potential host susceptibility factor for pathogen infection, possibly involved in establishing/facilitating biotrophy in <i>A. thaliana</i>.