Dentre os perigos relacionados com a produção de café verde, a ocratoxina A (OTA) é considerada significativa nesse produto. Com o objetivo de estudar a origem da OTA no processamento de café verde e avaliar os possíveis fatores de risco (região, etapas do processamento, fungos, insetos e ácaros) que poderiam contribuir na presença de OTA, foram amostradas três fazendas nos seguintes municípios: Bom Jardim (RJ), Espírito Santo do Pinhal (SP) e Cornélio Procópio (PR), onde havia a produção de café verde pela via seca e úmida. Foram coletadas 109 amostras de grãos de café, em diversas etapas do processamento, no período de maio a agosto de 2002. As amostras foram colhidas de acordo com fluxograma estabelecido e as etapas estudadas foram: cereja (campo), via seca, via úmida, armazém e café de varrição. Dessas amostras, 10 apresentaram OTA, que variou na sua quantificação de 3 a 101 ng.g-1. A maior prevalência de OTA foi a do café de varrição 57%, a via úmida apresentou uma prevalência de 10,3% e a via seca 7%. Na etapa cereja o fungo potencialmente toxigênico mais encontrado foi Penicillium spp, e quanto aos insetos, as larvas de Diptera e os Homoptera; na via úmida foram encontradas linhagens do Aspergillus seção Nigri e Circundatti e Penicillium spp, a infestação predominante foi a broca do café e os ácaros da família Acaridae; na etapa via seca registrou-se o isolamento de fungos do gênero Penicillium spp e Aspergillus seção Nigri , a broca do café foi a infestação predominante junto com os ácaros da família Acaridae, na etapa de armazém foram isolados Penicillium spp e Aspergillus seção Nigri, o inseto predominante foi a broca do café e os ácaros da família Acaridae; no café de varrição foram isolados Penicillium spp e Aspergillus seção Nigri e a broca do café estava presente junto com insetos da Ordem Diptera. Os resultados de cada etapa foram comparados por Análise de Variância em um nível de significância de 95%. Na análise de fatores de risco o café de varrição foi o mais significativo (p< 0,001). / The ochratoxin A (OTA) is considered to be among the most significant coffee contaminants with related hazards. With the purpose to study the origin of the OTA in the processing of green coffee and to evaluate the possible risk factors (region, step of processing, fungi, insects and mites) that could contribute in the OTA presence, had been selected three regions (states) of Brazil: Bom Jardim/RJ, Espírito Santo do Pinhal/SP and Cornélio Procópio/PR. A total of 109 samples of coffee beans were collected in conformity with flow chart established at different stages of ripening and processing: cherry (field), dry method, wet method, and warehouse and varrição coffee, from May to August of 2002. Of these samples 10 had presented OTA, which varied in its quantification from 3 to 101 ng.g-1. The biggest prevalence of OTA was the varrição coffee 57%, the wet method presented a prevalence of 10,3% and dry way 7%. In the stage cherry fungi potentially toxigenic was more found Penicillium spp, the insects had been the larvae of Diptera and the Homoptera; in the wet way it was found Aspergillus section Nigri and Circundatti and Penicillium spp, the predominant infestation was the coffee berry borer and the mite of the Acaridae family; in the stage wet methods spp was registered the isolation the Penicillium spp and Aspergillus section Nigri, the predominant infestation was the coffee berry borer together with the mites of the Acaridae family; in the stage of warehouse had been isolated Penicillium spp and Aspergillus section Nigri, and the predominant insect was the coffee berry borer and mites of the family Acaridae; in the varrição coffee had been isolated Penicillium spp and Aspergillus section Nigri the coffee berry borer was the most present insects together of Diptera. The results of each step had been with compared by Analysis of Variance. In the analysis of risk factors the varrição coffee\" was the factor most strongly related to the risk of the presence of OTA (p<0,001 ).

No description available.

No description available.

Les trichothécènes de Fusarium appartiennent au groupe des sesquiterpènes qui sont des inhibiteurs la synthèse des protéines des eucaryotes. Les trichothécènes causent d’une part de sérieux problèmes de santé aux humains et aux animaux qui ont consommé des aliments infectés par le champignon et de l’autre part, elles sont des facteurs importants de la virulence chez plantes. Dans cette étude, nous avons isolé et caractérisé seize isolats de Fusarium de la pomme de terre infectée naturellement dans un champs. Les tests de pathogénicité ont été réalisés pour évaluer la virulence des isolats sur la pomme de terre ainsi que leur capacité à produire des trichothécènes. Nous avons choisi F. sambucinum souche T5 comme un modèle pour cette étude parce qu’il était le plus agressif sur la pomme de terre en serre en induisant un flétrissement rapide, un jaunissement suivi de la mort des plantes. Cette souche produit le 4,15-diacétoxyscirpénol (4,15-DAS) lorsqu’elle est cultivée en milieu liquide. Nous avons amplifié et caractérisé cinq gènes de biosynthèse trichothécènes (TRI5, TRI4, TRI3, TRI11, et TRI101) impliqués dans la production du 4,15-DAS. La comparaison des séquences avec les bases de données a montré 98% et 97% d'identité de séquence avec les gènes de la biosynthèse des trichothécènes chez F. sporotrichioides et Gibberella zeae, respectivement. Nous avons confrenté F. sambucinum avec le champignon mycorhizien à arbuscule Glomus irregulare en culture in vitro. Les racines de carotte et F. sambucinum seul, ont été utilisés comme témoins. Nous avons observé que la croissance de F. sambucinum a été significativement réduite avec la présence de G. irregulare par rapport aux témoins. Nous avons remarqué que l'inhibition de la croissance F. sambucinum a été associée avec des changements morphologiques, qui ont été observés lorsque les hyphes de G. irregulare ont atteint le mycélium de F. sambucinum. Ceci suggère que G. irregulare pourrait produire des composés qui inhibent la croissance de F. sambucinum. Nous avons étudié les patrons d’expression des gènes de biosynthèse de trichothécènes de F. sambucinum en présence ou non de G. irregulare, en utilisant le PCR en temps-réel. Nous avons observé que TRI5 et TRI6 étaient sur-exprimés, tandis que TRI4, TRI13 et TRI101 étaient en sous-exprimés en présence de G. irregulare. Des analyses par chromatographie en phase-gazeuse (GC-MS) montrent clairement que la présence de G. irregulare réduit significativement la production des trichothécènes par F. sambucinum. Le dosage du 4,15-DAS a été réduit à 39 μg/ml milieu GYEP par G. irregulare, comparativement à 144 μg/ml milieu GYEP quand F. sambucinum est cultivé sans G. irregulare. Nous avons testé la capacité de G. irregulare à induire la défense des plants de pomme de terre contre l'infection de F. sambucinum. Des essais en chambre de croissance montrent que G. irregulare réduit significativement l’incidence de la maladie causée par F. sambucinum. Nous avons aussi observé que G. irregulare augmente la biomasse des racines, des feuilles et des tubercules. En utilisant le PCR en temps-réel, nous avons étudié les niveaux d’expression des gènes impliqué dans la défense des plants de pommes de terre tels que : chitinase class II (ChtA3), 1,3-β-glucanase (Glub), peroxidase (CEVI16), osmotin-like protéin (OSM-8e) et pathogenèses-related protein (PR-1). Nous avons observé que G. irregulare a induit une sur-expression de tous ces gènes dans les racines après 72 heures de l'infection avec F. sambucinum. Nous avons également trové que la baisse provoquée par F. sambucinum des gènes Glub et CEVI16 dans les feuilles pourrait etre bloquée par le traitement AMF. Ceci montre que l’inoculation avec G. irregulare constitut un bio-inducteur systémique même dans les parties non infectées par F. sambucinum. En conclusion, cette étude apporte de nouvelles connaissances importantes sur les interactions entre les plants et les microbes, d’une part sur les effets directs des champignons mycorhiziens sur l’inhibition de la croissance et la diminution de la production des mycotoxines chez Fusarium et d’autre part, l’atténuation de la sévérité de la maladie dans des plantes par stimulation leur défense. Les données présentées ouvrent de nouvelles perspectives de bio-contrôle contre les pathogènes mycotoxinogènes des plantes. / Fusarium trichothecenes are a large group of sesquiterpenes that are inhibitors of eukaryotic protein synthesis. They cause health problems for humans and animals that consume fungus-infected agricultural products. In addition some of Fusarium trichothecenes are virulence factors of plant pathogenesis. In this study, sixteen Fusarium strains were isolated and characterized from naturally infected potato plants. Pathogenicity tests were carried out to evaluate the virulence of these isolates on potato plants and their trichothecene production capacity. We chose F. sambucinum strain T5 as a model for this study because it was the most aggressive strain when tested on potato plants. It induces a rapid wilting and yellowing resulting in plant death. This strain produced 4,15-diacetoxyscirpenol (4,15-DAS) when grown in liquid culture. We amplified and characterized five trichothecene genes (TRI5, TRI4, TRI3, TRI11, and TRI101) involved in the production of 4,15-DAS. Nucleotide BLAST search showed 98% and 97% sequence identity with trichothecene biosynthetic genes of F. sporotrichioides and Gibberella zeae, respectively. We used F. sambucinum to determine if trichothecene gene expression was affected by the symbiotic arbuscular mycorrhizal fungus (AMF) Glomus irregulare. We found that the growth of F. sambucinum was significantly reduced in the presence of G. irregulare isolate DAOM-197198 compared with controls that consisted of carrot roots without G. irregulare or F. sambucinum alone. Furthermore, inhibition of the growth F. sambucinum was associated with morphological changes, which were observed when G. irregulare hyphae reached F. sambucinum mycelium, suggesting that G. irregulare may produce compounds that interfere with the growth of F. sambucinum. Using real-time qRT-PCR assays, we assessed the relative expression of trichothecene genes of F. sambucinum confronted or not with G. irregulare. When G. irregulare was confronted with F. sambucinum, TRI5 and TRI6 genes were up-regulated, while TRI4, TRI13 and TRI101 were down-regulated. We therefore used GC-MS analysis to determine whether G. irregulare affects trichothecene production by F. sambucinum. We found that the production of 4,15-DAS trichothecene was significantly reduced in the presence of G. irregulare compared with controls that consisted of carrot roots without G. irregulare or F. sambucinum alone. Interestingly, 4,15-DAS pattern was reduced to 39 μg/ml GYEP medium by G. irregulare compared to 144 μg/ml GYEP with F. sambucinum grown with carrot roots or F. sambucinum alone respectively. We tested the AMF capacity to induce defense responses of potato plants following infection with F. sambucinum. The response of AMF-colonized potatoes to F. sambucinum was investigated by tracking the expression of genes homologous with pathogenesis-related proteins chitinase class II (ChtA3), 1,3-β-glucanase (gluB), peroxidase (CEVI16), osmotin-like protein (OSM-8e) and pathogenesis-related protein (PR-1). We found that the AMF treatment up-regulated the expression of all defense genes in roots at 72 hours post-infection (hpi) with F. sambucinum. We also found that a decrease provoked by F. sambucinum in gluB and CEVI16 expression in shoots could be blocked by AMF treatment. Overall, a differential regulation of PR homologues genes in shoots indicates that AMF are a systemic bio-inducer and their effects could extend into non-infected parts. In conclusion, this study provides new insight into on the interactions between plants and microbes, in particular the effects of AMF on the growth and the reduction of mycotoxins in Fusarium. It also shows that AMF are able to reduce the disease severity in plants by stimulating their defense. The data presented provide new opportunities for bio-control against mycotoxin-producing pathogens in plants.

Dentre os perigos relacionados com a produção de café verde, a ocratoxina A (OTA) é considerada significativa nesse produto. Com o objetivo de estudar a origem da OTA no processamento de café verde e avaliar os possíveis fatores de risco (região, etapas do processamento, fungos, insetos e ácaros) que poderiam contribuir na presença de OTA, foram amostradas três fazendas nos seguintes municípios: Bom Jardim (RJ), Espírito Santo do Pinhal (SP) e Cornélio Procópio (PR), onde havia a produção de café verde pela via seca e úmida. Foram coletadas 109 amostras de grãos de café, em diversas etapas do processamento, no período de maio a agosto de 2002. As amostras foram colhidas de acordo com fluxograma estabelecido e as etapas estudadas foram: cereja (campo), via seca, via úmida, armazém e café de varrição. Dessas amostras, 10 apresentaram OTA, que variou na sua quantificação de 3 a 101 ng.g-1. A maior prevalência de OTA foi a do café de varrição 57%, a via úmida apresentou uma prevalência de 10,3% e a via seca 7%. Na etapa cereja o fungo potencialmente toxigênico mais encontrado foi Penicillium spp, e quanto aos insetos, as larvas de Diptera e os Homoptera; na via úmida foram encontradas linhagens do Aspergillus seção Nigri e Circundatti e Penicillium spp, a infestação predominante foi a broca do café e os ácaros da família Acaridae; na etapa via seca registrou-se o isolamento de fungos do gênero Penicillium spp e Aspergillus seção Nigri , a broca do café foi a infestação predominante junto com os ácaros da família Acaridae, na etapa de armazém foram isolados Penicillium spp e Aspergillus seção Nigri, o inseto predominante foi a broca do café e os ácaros da família Acaridae; no café de varrição foram isolados Penicillium spp e Aspergillus seção Nigri e a broca do café estava presente junto com insetos da Ordem Diptera. Os resultados de cada etapa foram comparados por Análise de Variância em um nível de significância de 95%. Na análise de fatores de risco o café de varrição foi o mais significativo (p< 0,001). / The ochratoxin A (OTA) is considered to be among the most significant coffee contaminants with related hazards. With the purpose to study the origin of the OTA in the processing of green coffee and to evaluate the possible risk factors (region, step of processing, fungi, insects and mites) that could contribute in the OTA presence, had been selected three regions (states) of Brazil: Bom Jardim/RJ, Espírito Santo do Pinhal/SP and Cornélio Procópio/PR. A total of 109 samples of coffee beans were collected in conformity with flow chart established at different stages of ripening and processing: cherry (field), dry method, wet method, and warehouse and varrição coffee, from May to August of 2002. Of these samples 10 had presented OTA, which varied in its quantification from 3 to 101 ng.g-1. The biggest prevalence of OTA was the varrição coffee 57%, the wet method presented a prevalence of 10,3% and dry way 7%. In the stage cherry fungi potentially toxigenic was more found Penicillium spp, the insects had been the larvae of Diptera and the Homoptera; in the wet way it was found Aspergillus section Nigri and Circundatti and Penicillium spp, the predominant infestation was the coffee berry borer and the mite of the Acaridae family; in the stage wet methods spp was registered the isolation the Penicillium spp and Aspergillus section Nigri, the predominant infestation was the coffee berry borer together with the mites of the Acaridae family; in the stage of warehouse had been isolated Penicillium spp and Aspergillus section Nigri, and the predominant insect was the coffee berry borer and mites of the family Acaridae; in the varrição coffee had been isolated Penicillium spp and Aspergillus section Nigri the coffee berry borer was the most present insects together of Diptera. The results of each step had been with compared by Analysis of Variance. In the analysis of risk factors the varrição coffee\" was the factor most strongly related to the risk of the presence of OTA (p<0,001 ).