This study evaluated the effect of Pratlenchus brachyurus and Fusarium verticillioides, and their relationship, on growth and development of two mayze hybrids (30F80 and 30K73). The experiment was conducted in greenhouse conditions at Pioneer Hi-bred Research Center in Itumbiara, GO, Brazil. The experimental design was randomized blocks with 15 treatments and 10 replications. Treatments included a non-inoculated control; inoculation of Fusarium in the seed and in the soil; three populations levels of Pratylenchus at sowing and at 30 days after planting (50, 100 and 500 nematodes/plot), with and without the inoculation of Fusarium in the soil; and three populations leves of Pratylenchus inoculated 30 days after sowing with Fusarium inoculated in seed at sowing. At plant maturity the following variables were evaluated: plant height and ear length, stalk diameter, root fresh and dry matter, stalk fresh and dry matter, reproduction factor and fusariosis severity on the stalk. Fusarium stalk colonization was evaluated in a scale from 1 to 9. The 30F80 hybrid had lower plant height and ear length, stalk diameter, stalk and root fresh matter when the seeds were inoculated with the suspension of Fusarium and sowing immediately after inoculation. The maize hybrids 30F80 and 30K73, showed the lowest root length, stalk fresh and dry matter, and root fresh and dry matter mainly when the soil was inoculated with Fusarium and sown in the same day, and the inoculation with 500 juveniles and/or adults of Pratylenchus, was done 30 days after sowing. The maize hybrid 30K73 obtained lower plant height and ear length when seeds were inoculated with the suspension of Fusarium and sown in the same day. Maize seeds inoculated with Fusarium verticillioides at planting decreased plant height, ear length, and root and shoot fresh weight on both hybrids. Stalk diameter of 30K73 hybrid and root length of hybrid 30K73 were also reduced. Soil inoculation of Fusarium verticillioides at planting and Pratylenchus brachyurus inoculated 30 days after planting (500 nematodes/plot) decreased root elongation and stalk, and root dry weights on both hybrids. However, plant height was only reduced on 30K73 hybrid. Future research is needed to better understand the relationship between Pratylenchus brachyurus and Fusarium verticillioides on maize growth and development. / O objetivo deste estudo foi avaliar o efeito de Pratylenchus brachyurus e/ou Fusarium verticillioides e sua relação no desenvolvimento de dois híbridos de milho (30F80 e 30K73). O experimento foi conduzido em casa de vegetação no Centro de Pesquisa da Pioneer Sementes de Itumbiara-GO. O delineamento experimental foi inteiramente ao acaso com 15 tratamentos/por híbrido (totalizando 30 tratamentos) e 10 repetições. Os tratamentos foram: testemunha (sem inoculações), inoculação de fungo nas sementes e no solo; inoculação de três populações de nematóide na semeadura e 30 dias após a semeadura (50, 100 e 500 nematóides/vaso), com e sem inoculação do fungo no solo; e três níveis populacionais de nematóides inoculados 30 dias após a semeadura, com o fungo inoculado na semente. Após a maturação (estádio R7), foram avaliadas as características: altura de planta e de espiga, diâmetro de colmo, matéria fresca e seca de raiz, matéria fresca e seca de colmo, fator de reprodução do nematóide e severidade de fusariose no colmo do milho. Observou-se que o híbrido de milho 30F80 obteve uma menor altura de planta e de espiga, diâmetro de colmo, matéria fresca de colmo e de raíz, quando as sementes foram inoculadas com a suspensão de conídios de Fusarium verticillioides e semeadura realizada no mesmo dia dessa inoculação. Os híbridos de milho 30F80 e 30K73 apresentaram o menor comprimento de raiz, matéria fresca e seca de colmo, e matéria fresca e seca de raiz, principalmente quando o solo foi inoculado com Fusarium verticillioides, semeou-se no mesmo dia e a inoculação do nematóide na população de 500 juvenis e/ou adultos de Pratylenchus brachyurus realizada 30 dias após a semeadura. O híbrido de milho 30K73 obteve uma menor altura de planta e de espiga quando as sementes foram inoculadas com a suspensão de conídios de Fusarium verticillioides e semeadura realizada no mesmo dia dessa inoculação. Sementes de milho inoculadas com o fungo na semeadura diminuiram a altura de planta e de espiga e peso fresco de raiz e de parte aérea nos dois híbridos de milho. Diâmetro de colmo no híbrido 30F80 e comprimento de raiz no híbrido 30K73 também foram reduzidos. A inoculação do solo com o fungo na semeadura e nematóide inoculado 30 dias após (500) proporcionou menor comprimento de raiz e peso seco e fresco de raiz e colmo nos dois híbridos. Contudo, altura de planta foi reduzida somente no híbrido 30K73. Outros estudos de interações entre microrganismos, em especial de Pratylenchus brachyurus e Fusarium verticilliodes, deverão ser realizados, a fim de melhor entender o efeito desta combinação no prejuízo das plantas. / Mestre em Agronomia

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No. of bitstreams: 2 Dissertação - Maythsulene Inácio de Sousa Oliveira - 2015.pdf: 3135445 bytes, checksum: a08e11c7f2df635f3ff7726cfde58f49 (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Previous issue date: 2015-04-11 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES / Common bean (Phaseolus vulgaris L.) is grown in Brazil in three different cropping seasons, and in diverse agroecosystems. In such different environments, the crop is exposed to several constraints responsible for yield losses, such as pathogenic organisms. Among common bean relevant diseases, fusarium wilt (Fusarium oxysporum f. sp. phaseoli), dry root-rot (Fusarium solani) and Curtobacterium wilt (Curtobacterium flaccumfaciens pv. flaccumfaciens) have similar symptoms, hindering diagnosis in the field, and whose identification in seed health testing is also limited. In both cases, identification at species level is an important step to manage this root pathogen complex, whose detection can be improved by molecular biology tools. Therefore, this study aimed to: 1) to develop and validate a multiplex PCR (m-PCR) method for simultaneous identification of three common bean pathogens, F. oxysporum f. sp. phaseoli, F. solani and C. flaccumfaciens pv. flaccumfaciens; and 2) develop an isothermal amplification of DNA (LAMP) method to detect of F. oxysporum f. sp. phaseoli on seeds. M-PCR method was developed for identification of isolated colonies, as well as infected seeds. In seeds, total DNA was obtained by alkaline lysis method, which inactivates nucleases during the extraction process. M-PCR allowed the identification of all pathogens, with detection of C. flaccumfaciens pv. flaccumfaciens, F. oxysporum f. sp. phaseoli and F. solani amplicons in agarose gel with respectively 306, 609 and 143 base pairs. Furthermore, m-PCR also reduced costs and time to detect Fusarium oxysporum f. sp. phaseoli from 10 days to three hours. It was not possible to develop an optimized protocol for detection of F. oxysporum f. sp. phaseoli by the LAMP method, using only the tf1 gene for design of primers, since such primers were functional only for amplifying large amounts of target DNA. Based on the negative results with LAMP, it is suggested that further studies should be performed using other DNA sequences available in GenBank database. / O feijoeiro-comum (Phaseolus vulgaris L.) é cultivado durante todo o ano no território brasileiro, em três épocas distintas e em vários agroecosistemas. Nestes ambientes distintos, a cultura está exposta a diversos fatores que causam perdas de rendimento, como o ataque de patógenos. Dentre as doenças do feijoeiro-comum encontram-se a murcha-de-fusarium (Fusarium oxysporum f. sp. phaseoli), a podridão-radicular-seca (Fusarium solani) e a murcha-de-curtobacterium (Curtobacterium flaccumfaciens pv. flaccumfaciens) que apresentam sintomas semelhantes, dificultando seu diagnóstico no campo, e cuja identificação em testes de sanidade de sementes também é limitada. Em ambos os casos, a identificação em nível de espécie é uma importante etapa do manejo deste complexo de patógenos, cuja detecção pode ser aperfeiçoada com a adoção de ferramentas de biologia molecular. Portanto, este estudo teve como objetivos: 1) Desenvolver e validar um método de multiplex PCR (m-PCR) para identificação simultânea de três espécies de patógenos do feijoeiro-comum, F. oxysporum f. sp. phaseoli, F. solani e C. flaccumfaciens pv. flaccumfaciens; e 2) desenvolver a técnica de amplificação isotérmica de DNA (LAMP) para detecção de F. oxysporum f. sp. phaseoli em sementes. O método de m-PCR foi desenvolvido para identificação de colônias isoladas bem como sementes infectadas. Nas sementes, o DNA total foi obtido pela lise alcalina, método que inativa nucleases durante o processo de extração. A m-PCR possibilitou a identificação de todos os patógenos, com detecção de C. flaccumfaciens pv. flaccumfaciens, F. oxysporum f. sp. phaseoli e F. solani em bandas formadas em gel de agarose respectivamente com 306, 609 e 143 pares de base. Além disso, a extração do DNA total das sementes pela lise alcalina em combinação com a m-PCR também possibilitou redução de custos e tempo de realização do diagnóstico de Fusarium oxysporum f. sp. phaseoli, de 10 dias para três horas. Não foi possível estabelecer um protocolo otimizado para detecção de F. oxysporum f. sp. phaseoli pelo método LAMP, utilizando somente o gene tf1 para desenho dos iniciadores, uma vez que, os iniciadores revelaram-se funcionais apenas para a amplificação com grandes quantidades de DNA alvo. Diante dos resultados obtidos com LAMP, sugere-se que estudos posteriores sejam realizados empregando outras sequências de DNA disponíveis no banco de dados GenBank.

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES / The pepper (Capsicum annuum L.) has commercial importance, by having sources of vitamins, minerals and fiber. However, a post-harvest problem, excessive softening which reduces the life and favors action pathogens such as Pectobacterium caratovorum subsp. caratovorum- Pcc, causal agent of soft rot, a major disease of post-harvest chili that is favored by the reduction of firmness. One technique that has recently been used for firmly maintaining the application is pectin methyl esterase (PME) with the addition of a calcium solution prologando thus reducing its lifetime and pathogen attack. The objective of this study was to use methyl pectin esterase (PME) exogenous associated with calcium chloride in maintaining firmness and control of Pectobacterium caratovorum subsp. caratovorum about pepper. The first experiment was conducted in a completely randomized design in a 4x5 factorial scheme with three replications for 12 days, evaluated every three days. In the second experiment, the test was conducted in a completely randomized design in a factorial 5x5 with three replications. In the first study the fruits of pepper were subjected to vacuum infusion method with pressure of 200 mmHg for 5 minutes, and following evaluated the loss of weight (PMF), fruit firmness (FF), skin color ( CC), soluble solids (SS), pH, total acidity (TA) and activity of SMEs. In the second test the fruits of pepper were subjected to vacuum infusion method with pressure of 200 mmHg for five minutes, then the fruits were inoculated with the PCC then held the fruit firmness analysis (FF), SME activity and the severity of the disease in chili (SD). The fruits obtained in general a reduction of over time firmly in all treatments but was found significant effect in maintaining the fruit firmness when treated in vacuum infusion with calcium chloride, without altering the physicochemical characteristics, as soluble solids content, total acidity and activity of SMEs, slowing the fruit ripening process. The fruits when treated with vacuum infusion with calcium chloride associated with pectin methyl esterase was not favorable because it altered the physicochemical properties chili, highlighting the decline of firmness, thus deteriorating the quality of the fruit. Regarding inoculation Pcc in the fruit was observed inhibition of growth of this pathogen, prolongation of fruit and better firmness treated fruits infusion over calcium inoculation Pcc chloride. / O pimentão (Capsicum annuum L.) tem grande importância comercial, por possuir fontes de vitaminas, minerais e fibras. Contudo, possui problema pós-colheita, o amolecimento excessivo que reduz a vida útil e favorece ação de patógenos como a Pectobacterium caratovorum subsp. caratovorum- Pcc, agente causal da podridão-mole, uma das principais doenças da pós-colheita do pimentão que é favorecida pela redução da firmeza. Uma técnica que vem sendo utilizada recentemente para a manutenção da firmeza é aplicação da pectina metil esterase (PME) com a adição de solução de cálcio prologando, assim, a sua vida útil e diminuindo o ataque do patógeno. Assim, o objetivo do trabalho foi utilizar a pectina metil esterase (PME) exógena associada ao cloreto de cálcio na manutenção da firmeza e no controle da Pectobacterium caratovorum subsp. caratovorum sobre o pimentão. O primeiro experimento foi realizado em delineamento inteiramente casualizado em esquema fatorial 4x5 com três repetições, durante 12 dias, avaliados a cada 3 dias. No segundo experimento, o ensaio foi realizado em delineamento inteiramente casualizado no esquema fatorial 5x5 com três repetições. No primeiro trabalho os frutos de pimentão foram submetidos ao método de infusão a vácuo com pressão de 200 mmHg por 5 minutos, e por seguinte avaliou-se a perda de massa fresca (PMF), firmeza do fruto (FF), cor da casca (CC), teor de sólidos solúveis (SS), pH, acidez total, (AT) e atividade de PME. No segundo ensaio os frutos de pimentão foram submetidos ao método de infusão a vácuo com pressão de 200 mmHg por cinco minutos, posteriormente os frutos foram inoculados com a Pcc em seguida realizou-se as análises de firmeza do fruto (FF), atividade de PME e a severidade da doença no pimentão (SD). Os frutos obtiveram de forma geral uma redução da firmeza ao longo do tempo em todos os tratamentos, porém foi verificado o efeito significativo na manutenção da firmeza dos frutos quando tratados em infusão a vácuo com cloreto de cálcio, não alterando as características físico-químicas, como o teor de sólidos solúveis, acidez total e atividade de PME, retardando o processo de amadurecimento do fruto. Os frutos quando tratados com infusão a vácuo com cloreto de cálcio associado à pectina metil esterase não foi favorável, pois alterou as propriedades físico-químicas do pimentão, com destaque para o declínio da firmeza, deteriorando assim a qualidade do fruto. Em relação à inoculação da Pcc no fruto, observou-se uma inibição do crescimento desse patógeno, prolongamento do fruto e uma melhor firmeza nos frutos tratados com infusão de cloreto de cálcio mais a inoculação da Pcc.

O presente trabalho teve por objetivos avaliar métodos de inoculação de Nectria haematococca e Phytophthora parasitica e idades de Passiflora edulis f. flavicarpa suscetíveis à infecção; avaliar a ocorrência de damping-off e podridão de colo do maracujazeiro em solo infestado; avaliar o comportamento de diferentes Passsifloraceas e genótipos de maracujazeiro amarelo aos respectivos patógenos; realizar testes de controle químico in vitro, tratamento químico erradicante em solo infestado e tratamento químico curativo em P. edulis f. flavicarpa para os respectivos patógenos. Inoculações no colo das plantas de P. edulis f. flavicarpa proporcionaram maiores níveis de doença comparadas às inoculações no sistema radicular, previamente ferido. Os resultados sugerem que N. haematococca seja um patógeno que penetra através de ferimentos. A mortalidade foi maior quando a inoculação foi realizada em plantas mais jovens e quando os patógenos N. haematococca e P. parasitica estavam em associação. Dentre as 17 espécies de Passiflora avaliadas para resistência aos patógenos, as espécies P. nitida, P. laurifolia e P. alata apresentaram as menores médias de lesões de N. haematococca, enquanto que para P. parasitica foram as espécies P. suberosa, P. foetida e P. morifolia as menos afetadas. Passiflora sidaefolia, P. edulis f. flavicarpa e P. edulis f. edulis foram as mais suscetíveis a ambos os patógenos, com sintomas que culminaram com a morte de plantas. Os genótipos de P. edulis f. flavicarpa mais resistentes a N. haematococca foram os procedentes de Morretes (PR) e a variedade Maguari e de Sapucaí (SP), enquanto que para P. parasitica foram os genótipos de Morretes (PR), Jaboticabal (SP) e LE13P2 (IAC) os menos afetados. A variedade Sul-Brasil e o genótipo de Livramento (BA) foram altamente suscetíveis a ambos os patógenos, com sintomas que culminaram com a morte de plantas. No teste de fungitoxidade in vitro avaliou-se a eficiência dos fungicidas na inibição do crescimento micelial de N. haematococca e P. parasitica. Na dose 100 ppm somente prochloraz inibiu totalmente o crescimento micelial de N. haematococca e nenhum produto inibiu acima de 82 % o crescimento de P. parasitica. Os fungicidas prochloraz, thiabendazole, thiram+thiabendazole, carbendazim, triflumizole e captan exerceram controle erradicante em solo infestado com N. haematococca, inibindo a incidência da doença em plantas com seis semanas pós-germinação. O mesmo foi observado com os produtos kif, dimethomorph, metalaxyl+mancozeb, mancozeb, cymoxanil+maneb e oxicloreto de cobre para P. parasitica. Os fungicidas testados em tratamento curativo inibiram o desenvolvimento da doença com melhores resultados quando aplicados dois dias após a inoculação, comparado a sete dias. Os fungicidas prochloraz e carbendazim destacaram-se por evitar a morte de plantas inoculadas com N. haematococca e os fungicidas kif, dimethomorph, metalaxyl+mancozeb e cymoxanil+maneb apresentaram eficiência semelhante entre si e superior a fosetyl- Al no controle de P. parasitica. / The objectives of the present work were to evaluate methods of inoculation of Nectria haematococca and Phytophthora parasitica and ages of Passiflora edulis f. flavicarpa which are susceptible to infection; to evaluate the damping-off and collar rot of passion fruit plant in infested soil; to evaluate the behavior of different Passsifloraceas and yellow genotypes of passion fruit to the respective pathogens; to carry out tests of chemical control in vitro, eradicative chemical treatment in infested soil and curative chemical treatment in P. edulis f. flavicarpa for the respective pathogens. Inoculations in the collar zone of P. edulis f. flavicarpa plants provided higher levels of disease when compared to the inoculations in the radicular system previously wounded. The results suggest that N. haematococca is a pathogen that penetrates through wounds. Mortality was higher when the inoculation was carried out in younger plants and when both pathogens were together. Amongst the 17 species of Passiflora tested for resistance to the pathogens, P. nitida, P. laurifolia, and P. alata showed the lowest average of N. haematococca lesions, while P. suberosa, P. morifolia, and P. foetida were the least affected species by P. parasitica. Passiflora sidaefolia, P. edulis f. flavicarpa, and P. edulis f. edulis were the most susceptible to both pathogens, showing symptoms that culminated with the death of the plants. The most resistant genotypes of P. edulis f. flavicarpa to N. haematococca were those from Morretes (PR), Maguari variety, and those from Sapucaí (SP); with respect to P. parasitica, the genotypes from Morretes (PR), Jaboticabal (SP), and LE13P2 (IAC) were the least affected. The Sul- Brasil variety and the genotype from Livramento (BA) had been highly susceptible to both pathogens, having symptoms that culminated with the death of plants. The in vitro efficiency of the fungicides in the inhibition of the mycelial growth of N. haematococca and P. parasitica was evaluated. At 100 ppm, only prochloraz inhibited totally the mycelial growth of N. haematococca and no product inhibited over 82 % the growth of P. parasitica. Prochloraz, thiabendazole, thiram+thiabendazole, carbendazim, triflumizole, and captan controlled eradicatively the soil infested by N. haematococca, inhibiting the incidence of the disease in plants which were six weeks old. The same was observed for the products kif, dimethomorph, metalaxyl+mancozeb, mancozeb, cymoxanil+maneb, and copper oxychloride for P. parasitica. The tested fungicides in curative treatment inhibited the development of the disease with better results when applied two days after the inoculation, compared to seven days. Prochloraz and carbendazim were outstanding for preventing the death of plants inoculated with N. haematococca. Kif, dimethomorph, metalaxyl+mancozeb, and cymoxanil+maneb showed similar efficiency and were superior to fosetyl-Al in the control of P. parasitica.

La bioconversion en méthane de biomasses lignocellulosiques est l’une des alternatives les plus prometteuses pour la production de méthane issu de la digestion anaérobie. Toutefois, les biomasses lignocellulosiques présentent des caractéristiques bio-physico-chimiques très variables en raison leur composition biochimique et de l’organisation structurale très diverses. Par ailleurs, leur faible biodégradabilité en conditions anaérobie nécessite de les prétraiter avant méthanisation pour optimiser la production de méthane. Ce travail vise à évaluer l’influence des caractéristiques d’une large gamme de substrats lignocellulosiques sur leur biodégradabilité anaérobie et les corrélations entre leurs caractéristiques bio-physico-chimiques et le potentiel biométhanogène, et d’étudier les effets du prétraitement fongique en présence de Ceriporiopsis subvermispora sur le potentiel biométhanogène de biomasses lignocellulosiques sélectionnées dans la présente étude et de caractériser les changements de leurs caractéristiques après le prétraitement fongique. La caractérisation de 36 biomasses lignocellulosiques représentatives d’une large gamme de gisements potentiellement mobilisables a permis de mettre en évidence les corrélations linéaires entre le potentiel biométhanogène des biomasses et certaines de leur caractéristiques bio-physico-chimiques, dont la teneur en lignine et la demande biochimique en oxygène. Les biomasses sylvicoles et agricoles ont montré des caractéristiques distinctes de la biodégradabilité aérobie et anaérobie. Les résultats de prétraitement fongique sur les 5 biomasses ont permis de mettre en évidence que le champignon de pourriture blanche Ceriporiopsis subvermispora réagit distinctement selon la biomasse prétraitée. Pour certaines biomasses, le prétraitement fongique conduit à augmenter significativement la production de méthane et la vitesse de bioconversion en méthane. Cette espèce présente la capacité de dégrader sélectivement la lignine sur certaines biomasses et, sur d’autres, celle de dégrader de manière non-sélective des polysaccharides et des lignines. De plus, pour les deux souches de Ceriporiopsis subvermispora testées, des métabolismes différents ont été mis en évidence sur une même biomasse. Les résultats de compositions et ceux de l’analyse structurale des biomasses (initiales, autoclavées, contrôles, et prétraitées par Ceriporiopsis subvermispora) ont montré que leur structure peut être modifiée sans toutefois observer une transformation significative de leur composition biochimique. / Bioconversion to methane lignocellulosic biomass is one of the most promising alternatives for the production of methane from anaerobic digestion. However, lignocellulosic biomass has various bio-physicochemical characteristics due to their biochemical composition and diverse structural organization. Moreover, their low biodegradability in anaerobic condition requires pretreatment before methanation to optimize methane production. This work aims to evaluate the influence of the characteristics of a wide range of lignocellulosic substrates on their anaerobic biodegradability and correlations between their bio-physical-chemical characteristics and biomethane potential, and study the effects of fungal pretreatment in the presence of Ceriporiopsis subvermispora on the biogas potential of lignocellulosic biomass selected in this study and characterize their changes of their characteristics before and after the fungal pretreatment. The characterization of 36 representative lignocellulosic biomass of a wide range of potentially mobilized deposits allowed to highlight the linear correlations between biomethane potential of biomass and some of their bio-physical-chemical characteristics, of which the lignin content and biochemical oxygen demand. The forest and agricultural biomass exhibited distinct characteristics of the aerobic and anaerobic biodegradability. The results of fungal pretreatment of the 5 biomass indicated that the white rot fungus Ceriporiopsis subvermispora reacts distinctly depending on the pretreated biomass. For some biomass, fungal pretreatment leads to significant increase of methane production and the bioconversion rate of methane. This species presents the ability to selectively degrade lignin on some biomasses, in others, the ability to non-selectively degrade polysaccharides and lignins. In addition, for both strains of Ceriporiopsis subvermispora tested, different metabolisms were highlighted on the same biomass. The results of compositions and those of the structural analysis of biomass (initials, autoclaved, controls, and pretreated with Ceriporiopsis subvermispora) showed that their structure can be modified without observing a significant transformation of their biochemical composition.

La vanille, utilisée en alimentation et parfumerie, est une orchidée originaire d’Amérique centrale et maintenant cultivée dans l’océan indien (Vanilla planifolia, 97% de la production mondiale) et dans l’océan pacifique (Vanilla tahitensis, 3% de la production mondiale, essentiellement en Polynésie française).J’ai montré que Vanilla tahitensis était aussi sensible que Vanilla planifolia à la fusariose et représentait la maladie principale de la vanille en Polynésie française comme dans le reste des zones de production des vanilles aromatiques. J’ai participé à démontrer qu’il s’agissait d’une nouvelle espèce de Fusarium oxysporum « f. sp. radicis-vanillae » car elle ne s’attaque qu’au cortex racinaire et ne pénètre que peu dans les tissus vasculaires. J’ai suivi pendant 4 ans l’évolution qualitative et quantitative des dégâts dus à 6 pathogènes et ravageurs dans 51 plantations à Raiatea, principalement des ombrières. J’ai ainsi montré que, contrairement aux autres pathologies et ravageurs, la fusariose connaissait un développement presqu’exponentiel ces 3 dernières années, comme c’est également le cas sur Vanilla planifolia dans le reste du monde. Ayant, par ailleurs, conduit un travail d’enquête auprès des producteurs, j’ai indiqué (l’analyse statistique et la modélisation afférente ne sont pas terminées) que le soin apporté aux lianes par les producteurs était essentiel au bon contrôle de la fusariose. / Vanilla species, used for aroma and flagrances, are orchidaceae originated from central America. They are now mostly cultivated in the Indian ocean (Vanilla planifolia, 97%, mainly in Madagascar, Indonesia, India and smaller islands from the Indian ocean) and in the Pacific ocean (Vanilla tahitensis, 3%, mainly French Polynesia).I have shown that Vanilla tahitensis was as susceptible as Vanilla planifolia towards fusarium root rot. It does represent the major losses on Vanilla tahitensis in Raiatea like reported for all the other areas of vanilla production worldly. I have participated to show that this fungus was delimited to root cortex maceration and did not invade vascular tissues, reason for which we use a new name for this pathogen : Fusarium oxysporum f sp radicis-vanillae.I have followed the etiological and epidemiological characteristics of this disease and of 5 other pathogens and insects on 51 Vanilla plantations, mainly shade-houses, during 4 years, for each vine in cohort analysis manner. I have shown, among this 6 biological causes of loss, fusarium root rot was the only one to present an exponential trajectory within the last 3 years, while the other causes remain stable or display a year increase because of climatic reasons. I did interview all the producers owning these plantations and analyze statistically the answers to the very numerous questions. The statistical and modelling analysis is not yet finished. Nevertheless, I can already claim that regular vine cleaning is absolutely necessary to maintain the vanilla plantations healthy, something which, unfortunately is not a rule for everyone.

This diploma thesis focuses on the determination of sulfonamide antibiotics especially the possibility of elimination of these substances from the aquatic ecosystem. Nowadays, environmental contamination of the pharmaceuticals and their residues is a serious concern. Main sources of this contamination are wastewater treatment plants (WWTPs), where these compounds are not effectively removed by contemporary conventional technology. For this reason, new methods are being developed and tested that could eliminate the number of contaminants entering the environment in this way. There is a possibility to use the potential of the enzymatic system of wood-decay fungi, especially white rot fungi. Six representatives of sulfonamide antibiotics were selected and isolated from the aquatic matrix via solid phase extraction. The final identification and quantification method was high performance liquid chromatography with mass spectrometric detection. Monitoring of the concentration level of selected sulfonamide antibiotics at the inflow and effluent at the Brno-Modřice WWTP was carried out weekly. Moreover, the effectiveness of elimination of selected antibiotics from the aquatic ecosystem by the use of Trametes versicolor wood-decay fungi cultured on a suitable carrier was verified.

This work is focused on compilation of general diagnostic methods used for diagnostic of wooden structures specifically listed wooden roof truss of church st. Bartholomew in Ivan. The main impulse of diagnosis was extensive damage and attack of truss and effort on rehabilitation and construction of the entire building of the church with an emphasis on preserving historical values. Diagnosis of truss was based on the research used sense and instrumental methods. In the first step, identify all dimensions of the individual elements of the structure was determined, the type truss system was intended. Follow-up survey was focused on determining the extent and causes of damage. In design of the rehabilitation process emphasis is placed on structural rehabilitation provided by follow-up processes, such as preventive chemical protection.

L’aglio è coltivato a livello mondiale nelle regioni temperate, secondo la FAO nel 2016 1.5 milioni di ettari sono stati destinati a questa coltura. A partire dal 2002 Fusarium proliferatum è stato segnalato come principale agente causale del marciume secco dell’aglio in post raccolta. I sintomi sono identificabili come macchie necrotiche sui bulbilli e in presenza di forti infezioni è possibile osservare la presenza di micelio bianco. F. proliferatum è un patogeno in grado di produrre Fumonisina B1 e B2, micotossine che potrebbero accumularsi all’interno dei bulbi ed essere tossiche per il consumatore. In Italia nel 2012 il 30% del raccolto è stato perso a causa di questo patogeno. Essendo la Fusariosi una malattia emergente a livello mondiale in letteratura si trovano ancora scarse informazioni sul suo sviluppo a livello di campo e sulle strategie di contenimento. 1. Allo scopo di verificare la quantità di inoculo fungino presente nell’ambiente si è proceduto all’analisi dei suoli in presemina con specifica attenzione alla quantificazione e all’identificazione delle specie fungine presenti. A tale scopo è stata eseguita: la conta delle Unità Formanti Colonia per grammo di terreno (UFC/g) su terreni specifici per l’isolamento del genere Fusarium spp. per grammo di terreno. Le identificazioni sono state eseguite al microscopio ottico e confermate successivamente con metodi molecolari. 2. Per seguire l’avanzamento della malattia durante la stagione colturale, invece, si è proceduto al campionamento in tre fasi fenologiche (inizio formazione dei bulbilli BBCH 15, ingrossamento dei bulbilli BBCH 45, maturazione di raccolta BBCH 49) con caratterizzazione dei sintomi, isolamento e riconoscimento dei funghi associati al marciume. 3. Per verificare la correlazione tra andamento meteo e incidenza delle specie fungine associate al marciume secco, sono stati raccolti i dati di meteorologici relativi al totale delle piogge, ai gradi giorno, all’umidità relativa media e alla temperatura media nei quadrati corrispondenti alle aziende agricole oggetto di studio. I dati sono stati correlati attraverso il coefficiente di correlazione di Pearson con i valori di gravità e incidenza della malattia stimati a fine stagione colturale. 4. Per verificare l’insorgenza dalla malattia nella fase di post raccolta si è proceduto con campionamenti di bulbi in conservazione, posa in piastra di bulbilli sintomatici e asintomatici e calcolo dell’incidenza delle specie fungine associate ai sintomi del marciume. 5. Al fine di verificare la possibile presenza di fumonisine nei campioni analizzati durante la stagione colturale e nel post raccolta, si è proceduto all’analisi attraverso HPLC di estratti di aglio. 6. Per individuare possibili strategie di controllo della malattia durante la stagione colturale sono stati eseguiti test di efficacia di prodotti chimici e biologici in vitro e in campo. I prodotti chimici sono stati provati su PDA modificato inoculato centralmente con F. proliferatum, mentre per gli agenti di biocontrollo sono state allestite prove di coltura duale. La prova in campo, invece, è stata eseguita all’interno di un campo sperimentale a strip plot. L’aglio delle diverse tesi è stato conservato in cella frigorifera per 9 mesi, per valutare la persistenza dei prodotti utilizzati alla concia. I risultati ottenuti hanno dimostrato che F. proliferatum e F. oxysporum sono le specie maggiormente associate al marciume dell’aglio durante la stagione colturale. L’andamento delle due specie è complementare e varia a seconda dell’andamento meteorologico della stagione colturale. F. proliferatum è correlato positivamente con l’aumento della temperatura e delle piogge, mentre F. oxysporum sembra prevalere nelle stagioni meno piovose ed ha mostrato correlazione positiva con la gravità dei sintomi rilevati in campo sulle corone. La carica micotica di Fusarium nel terreno rimane costante negli anni di analisi, facendo presupporre un maggiore ruolo del seme nella trasmissione della malattia. Per quanto riguarda il post raccolta, invece, F. proliferatum risulta la specie isolata con maggiore frequenza dai bulbilli e si correla positivamente ai sintomi rilevati sugli spicchi, confermando il suo ruolo come agente causale del marciume secco durante lo stoccaggio. F. oxysporum, invece, colonizza in prevalenza le radici e la parte basale della pianta dividendo il patosistema in due subsistemi: F. proliferatum-bulbi; F. oxysporum-radici. F. proliferatum è stato isolato anche dagli spicchi asintomatici con frequenza del 25%, ed è stato possibile rilevare la presenza di fumonisine con l’avanzare del tempo di stoccaggio in cella. Essendo il fungo presente anche sugli spicchi asintomatici maggiori studi saranno necessari per garantire la sicurezza dei consumatori. Infine, dalle prove di concia in campo è emerso che il principio attivo Tebuconazolo, riduce la comparsa dei sintomi da Fusarium, ma non in modo risolutivo. Ciononostante, una volta che il prodotto viene riportato a temperatura ambiente dopo lo stoccaggio in cella refrigerata, l’incidenza di F. proliferatum aumenta nuovamente con possibilità di sviluppo di danni al prodotto da commercializzare. / Since 2002, Fusarium proliferatum has been reported as the main causal agent of garlic dry rot during the postharvest stage, but information on the development of the disease throughout the production chain was nearly absent. Dry rot has caused huge economic losses in the past few years (up to 30 % of the yield), symptoms are visible on bulbs during storage as necrotic spots and in the most severe attacks, white mycelium may become visible on cloves. Few pest management strategies were tested in the recent past, but none were satisfactory. Due to the economic effect that this pathogen can have on local productions, the thesis aimed to deeply investigate the pathosystem with a field to fork approach and to test new strategies to control fungal infections. First of all, the work focused on garlic (Allium sativum L.) cropping season, intending to clarify the role of F. proliferatum in bulb infection as well as the impact of crop growing conditions on the development of the pathogen. A 3-year study was conducted in Piacenza (northern Italy) by sampling six garlic farms with different dry rot history (three highly contaminated and three low contaminated). Soil samples were recovered at sowing time for the counting of fungal colony-forming units (CFU). Plant samples were collected at three relevant growth stages, from April to July, for which disease severity assessment and fungi isolations were performed. Fusarium was the most frequently isolated genus, and F. proliferatum and F. oxysporum the dominant species during the garlic cropping season. F. oxysporum was dominant in the first year of the study, but F. proliferatum registered the highest incidence in all the farms tested. F. oxysporum incidence was correlated with dry weather, whereas F. proliferatum was enhanced in rainy years. To conclude, F. proliferatum is confirmed to be associated with garlic bulbs, even at crop’s early growth stages and symptoms are visible mainly on roots and basal plates at the field stage, related to F. oxysporum. Then, the focus was made in detecting the presence of F. proliferatum on garlic bulbs during prolonged storage, and to identify other fungal species associated with garlic dry rot. Moreover, fumonisin contamination in symptomatic and asymptomatic cloves were detected. Samples of 100 plants were collected over three production seasons in six farms located in Northern Italy at three-time points (at harvest, processing, and 6 months storage at –4° C). Results obtained lead to think that Fusarium–garlic pathosystem is split into two parts: basal plate/root and bulb. F. proliferatum had the highest incidence in infected bulbs and was confirmed as the causal agent of postharvest dry rot in garlic (mean incidence: 35.4%). F. oxysporum co-occurred with F. proliferatum but symptoms were visible only on basal plate/root. Dry rot incidence slightly increased during cold storage (from 14.6% at processing to 18.4% at 6-month storage); although, F. proliferatum incidence was stable during cold storage, fumonisin were produced from harvest through storage. Cloves showing symptoms were more contaminated compared to those asymptomatic, both by the fungus (mean incidence 39% vs 25.3%) and the toxin (287.0 vs 24.4 µg kg-1). Therefore, cold storage limits garlic dry rot, but health concerns related to fumonisin should be seriously considered. Regarding disease management, garlic crop is commonly propagated by plant parts (cloves). To protect garlic crop from early growth stages it is important to find commercial products able to control the pathogen growth on seedlings. The experiment aimed to test in vitro and in vivo the efficacy of triazoles and biocontrol agents (BCAs) against F. proliferatum and F. oxysporum. In in vitro trials, the best performance was achieved by propiconazole+prochloraz (100%), followed by tebuconazole (88.9%). BCAs were less effective but still showed great capacity to control the pathogen with maximum growth inhibition of 80% (Trichoderma harzianum +T. gamsii). In both cases, temperature influenced the capacity to control the pathogen with minimum effect at 25°C compared to lower temperatures. In vivo bacterial BCAs showed a similar capacity to control Fusaria compared to chemical products (mean of severity index 18.6% and 11.7%, respectively) and did not show side effects on root length. In vitro and in vivo results are comparable, except for Trichoderma, with the worst performances in terms of disease severity on plants. Finally, a field trial was designed to verify the efficacy of chemical and biological active ingredients as seed coating both at crop stage and postharvest, simulating the entire production chain, by taking into account visible symptoms and incidence of fungi. All products tested reduced the severity of symptoms on basal plates at the field stage, but none of them was able to reduce Fusarium incidence. A postharvest analysis conducted on bulbs demonstrated the efficacy of Tebuconazole, B. subtilis, and Trichoderma+B. subtilis in reducing the number of cloves showing symptoms per bulb (mean 34.3% vs control 45.8%). Moreover, Tebuconazole was able to reduce the incidence of F. proliferatum by 48% with respect to untreated control. The trial highlighted also that the incidence of F. proliferatum increased by 37% when garlic bulbs were kept for 15 days at room temperature simulating storage at consumers houses. Results obtained in the trial are promising and seed coating had a positive effect on garlic dry rot postharvest; although further studies are needed to test the persistence of seed coating treatments after prolonged storage period, especially when the product is kept outside cold chambers.

Le principali anomalie evidenziate nelle nocciole avariate riguardano la presenza di macchie necrotiche, imbrunimenti interni rilevati inseguito al taglio dei frutti e la presenza di marciumi parziali o totali su di essi con conseguente riduzione del prodotto disponibile sul mercato e ingenti perdite economiche. Sulla base di quanto premesso, lo scopo principale di questa tesi è stato quello di identificare e caratterizzare l'agente causale/i dei difetti del nocciolo e di definire un protocollo di gestione del problema, secondo i principi IPM. È stata quantificata l'incidenza dei generi fungini isolati da nocciole provenienti da diverse aree geografiche (Caucaso e Turchia), raccolte in diversi anni e in diverse fasi di crescita della coltura. Diaporthe è stato identificato come il principale genere fungino associato ai difetti della nocciola, e la caratterizzazione molecolare ha confermato il ruolo predominante della specie D. eres. Sono state studiate le caratteristiche ecologiche di questo fungo e l'efficacia in vitro di fungicidi chimici e biologici per controllarne la crescita miceliale, lo sviluppo di picnidi e la formazione di cirri. I risultati ottenuti costituiscono una base per definire linee guida per la corretta gestione della coltura al fine di prevenire la comparsa del patogeno identificato e delle difettosità nelle nocciole / Defects on hazelnuts are responsible for a significant reduction in the availability of high-quality hazelnuts with profound economic impacts. A reduction in the occurrence of defects, based on the identification of causal agents and on their control, is critical to improve the quality of raw nut products and reduce yield losses. Based on this background, the main aim of this thesis was to identify and characterize the causal agent/s of kernel defects in hazelnuts and to define a management protocol, according to IPM principles. Defects affecting hazelnuts are cause of brown spots on the kernel surface and darkening/browning inside the kernel, visible after nuts are half-cut. The incidence of fungal genera in hazelnuts coming from different geographical areas (Caucasus and Turkey), collected in different years at different crop growth stages, was quantified. Diaporthe was identified as the main genus involved in hazelnut defects, and the molecular characterization confirmed the predominant role of D. eres. Its ecological needs were investigated, and the in vitro efficacy of chemical and biological fungicide were screened to control D. eres growth, pycnidial conidiomata development and cirrhi occurrence. This would be an essential tool for a sustainable crop protection approach in this economically important crop