Title from PDF of title page (University of Missouri--Columbia, viewed on Feb 24, 2010). The entire thesis text is included in the research.pdf file; the official abstract appears in the short.pdf file; a non-technical public abstract appears in the public.pdf file. Dissertation advisor: Dr. Georgia Davis. Vita. Includes bibliographical references.

Aspergillus flavus is a pathogenic fungus causing maize ear rot disease and producing aflatoxins that are carcinogenic to humans and animals. Characterizing maize host resistance mechanisms and prioritizing candidate resistance genes are important to the development of resistant maize germplasm. This study investigated transcriptomics approaches and statistics methods on the identification of maize host resistance genes. Full-length cDNA libraries were also constructed and evaluated for gene cloning and functional analysis. This study established important sources for functional studies on differentially expressed genes and for identification of allelic gene forms to characterize gene polymorphisms and facilitate the development of DNA markers.

Aflatoxins are a secondary metabolite produced by the fungus Aspergillus flavus. A. flavus has been known to infect several crops including tree nuts, peanuts, rice, cotton and maize. Aflatoxins have been found to cause tumors with aflatoxin B1 being the most carcinogenic biologically produced substance known to man. Therefore, the FDA has restricted the amount of aflatoxin in maize for human consumption to 20 ppb (ng/g). An estimated $225 million are lost each year in the United States due to aflatoxin contamination in maize crops alone. Agriculture is a vital part of Mississippi’s economy, and maize is one of its largest crops. The purpose of this research is to track the correlations between aflatoxin accumulation and Aspergillus flavus fungal biomass for the first several weeks after inoculation, as well as the spreading of the fungus and the aflatoxin throughout the inoculated ear of maize. This will allow for better understanding of the pathogen-host interactions and how the fungus progresses over time. GA209 x T173 is the aflatoxin accumulation susceptible maize hybrid, GA209 x Mp313E is the susceptible and resistant hybrid, and Mp717 x Mp313E is the resistant maize hybrid to aflatoxin accumulation. These maize hybrids were each inoculated with toxin producing Aspergillus flavus NRRL 3357 and water as a control 21 days after silk maturation. Collections of the inoculated maize cobs were made 3, 7, 14, 21, 28, 35, and 60 days after inoculation. Maize samples were collected and analyzed for aflatoxin and DNA concentration. The extracted aflatoxin was analyzed using an LC/MS. The fungal biomass was determined by performing quantitative real time polymerase chain reaction (PCR). GA209xT173 and Mp717xMp313E showed no aflatoxin production two days after inoculation. The resistant maize hybrid lead in aflatoxin accumulation the last two years but had the least amount of fungal biomass for second and third years of the experiment The production of aflatoxin seems to begin decelerating after 21 days after inoculation. Resistance characteristics are more to prevent fungal infection. Fungal biomass was significantly higher in the susceptible hybrid GA209xT173 compared to the other hybrids. However, fungal spread was significantly higher in Mp313ExT173 and Mp717xMp313E.

No description available.

Muitos fungos filamentosos podem produzir metabólitos secundários, denominados micotoxinas, podendo ser encontradas em produtos alimentícios e produtos agrícolas. Um dos principais gêneros de fungos micotoxigênicos relacionados à cadeia alimentar é o Aspergillus spp. Existem mais de 400 micotoxinas descritas na literatura, sendo as mais comuns as aflatoxinas B1, B2, G1 e G2. As micotoxinas são frequentemente encontradas em alimentos e são consideradas como um dos mais perigosos contaminantes, sendo a aflatoxina B1 classificada no Grupo 1 pela International Agency of Research on Cancer. As aflatoxinas são termorresistentes, resistindo por mais de uma hora em autoclave, fazendo-se necessário outro meio de degradação dessas toxinas. Este trabalho teve como objetivo observar os efeitos da radiação gama de 60Co e de feixes de elétrons na degradação das aflatoxinas e comparar os danos causados na morfologia do fungo Aspergillus flavus. O fungo foi cultivado em agar batata dextrose (PDA) por 10 dias e posteriormente foi transferido para o meio agar coco, sendo mantido por 14 dias à 25 °C. Após esta etapa o agar coco foi triturado até se tornar um meio pastoso homogêneo e foi irradiado com doses de 2,5, 5,0, 10 e 20 kGy. As amostras utilizadas na microscopia eletrônica de varredura foram irradiadas com as doses de 0, 2,5, 5,0, 10 e 20 kGy com fontes de 60Co e de feixes de elétrons. A irradiação com acelerador de elétrons apresentou uma degradação ligeiramente superior à radiação gama, reduzindo 29,93 %, 34,50 %, 52,63 % e 72,30 % para as doses de 2,5, 5,0, 10 e 20 kGy, respectivamente. A microscopia eletrônica de varredura demonstrou que as doses de 2,5 até 10 kGy não causaram danos no fungo, porém com a dose de 20 kGy pode-se observar danos nas estruturas fúngicas. / Many filamentous fungi can produce secondary metabolites, called mycotoxins, which can be found in food and agricultural products. One of the main genera of mycotoxigenic fungi related to the food chain is the Aspergillus spp. There are over 400 mycotoxins described in the literature, the most common the aflatoxins B1, B2, G1 and G2. The mycotoxins are commonly found in foods and are considered one of the most dangerous contaminants. The aflatoxin B1 is classified in group one by the International Agency of Research on Cancer. Aflatoxins resisting for more than one hour in autoclave making it necessary to other means of degradation of these toxins. This work aimed to observe the effects of gamma radiation of 60Co and electron beams in the degradation of aflatoxins and compare the damage caused on the morphology of the Aspergillus flavus. The fungus was grown on potato dextrose agar (PDA) for 10 days and was subsequently transferred to coconut agar medium, and maintained for 14 days at 25 °C. After this step the coconut agar was ground to become a homogeneous pasty and was irradiated with doses of 2.5, 5.0, 10 and 20 kGy. The samples used in scanning electron microscopy were irradiated with doses of 0, 2.5, 5.0, 10 and 20 kGy with sources of 60Co and electron beams. Irradiation with electron accelerator showed a slightly higher degradation to gamma radiation, reducing 29.93 %, 34.50 %, 52.63 % and 72.30 % for doses of 2.5, 5.0, 10 and 20 kGy, respectively. The Scanning Electron Microscopy showed that doses of 2.5 to 10 kGy did not cause damage to the fungus, but with a dose of 20 kGy it can be observed fungal damage to structures. SUMÁRIO

Muitos fungos filamentosos podem produzir metabólitos secundários, denominados micotoxinas, podendo ser encontradas em produtos alimentícios e produtos agrícolas. Um dos principais gêneros de fungos micotoxigênicos relacionados à cadeia alimentar é o Aspergillus spp. Existem mais de 400 micotoxinas descritas na literatura, sendo as mais comuns as aflatoxinas B1, B2, G1 e G2. As micotoxinas são frequentemente encontradas em alimentos e são consideradas como um dos mais perigosos contaminantes, sendo a aflatoxina B1 classificada no Grupo 1 pela International Agency of Research on Cancer. As aflatoxinas são termorresistentes, resistindo por mais de uma hora em autoclave, fazendo-se necessário outro meio de degradação dessas toxinas. Este trabalho teve como objetivo observar os efeitos da radiação gama de 60Co e de feixes de elétrons na degradação das aflatoxinas e comparar os danos causados na morfologia do fungo Aspergillus flavus. O fungo foi cultivado em agar batata dextrose (PDA) por 10 dias e posteriormente foi transferido para o meio agar coco, sendo mantido por 14 dias à 25 °C. Após esta etapa o agar coco foi triturado até se tornar um meio pastoso homogêneo e foi irradiado com doses de 2,5, 5,0, 10 e 20 kGy. As amostras utilizadas na microscopia eletrônica de varredura foram irradiadas com as doses de 0, 2,5, 5,0, 10 e 20 kGy com fontes de 60Co e de feixes de elétrons. A irradiação com acelerador de elétrons apresentou uma degradação ligeiramente superior à radiação gama, reduzindo 29,93 %, 34,50 %, 52,63 % e 72,30 % para as doses de 2,5, 5,0, 10 e 20 kGy, respectivamente. A microscopia eletrônica de varredura demonstrou que as doses de 2,5 até 10 kGy não causaram danos no fungo, porém com a dose de 20 kGy pode-se observar danos nas estruturas fúngicas. / Many filamentous fungi can produce secondary metabolites, called mycotoxins, which can be found in food and agricultural products. One of the main genera of mycotoxigenic fungi related to the food chain is the Aspergillus spp. There are over 400 mycotoxins described in the literature, the most common the aflatoxins B1, B2, G1 and G2. The mycotoxins are commonly found in foods and are considered one of the most dangerous contaminants. The aflatoxin B1 is classified in group one by the International Agency of Research on Cancer. Aflatoxins resisting for more than one hour in autoclave making it necessary to other means of degradation of these toxins. This work aimed to observe the effects of gamma radiation of 60Co and electron beams in the degradation of aflatoxins and compare the damage caused on the morphology of the Aspergillus flavus. The fungus was grown on potato dextrose agar (PDA) for 10 days and was subsequently transferred to coconut agar medium, and maintained for 14 days at 25 °C. After this step the coconut agar was ground to become a homogeneous pasty and was irradiated with doses of 2.5, 5.0, 10 and 20 kGy. The samples used in scanning electron microscopy were irradiated with doses of 0, 2.5, 5.0, 10 and 20 kGy with sources of 60Co and electron beams. Irradiation with electron accelerator showed a slightly higher degradation to gamma radiation, reducing 29.93 %, 34.50 %, 52.63 % and 72.30 % for doses of 2.5, 5.0, 10 and 20 kGy, respectively. The Scanning Electron Microscopy showed that doses of 2.5 to 10 kGy did not cause damage to the fungus, but with a dose of 20 kGy it can be observed fungal damage to structures. SUMÁRIO

An efficient approach to combat the accumulation of aflatoxin is the development of germplasm resistant to infection and spread of A. flavus in maize, one of the most important cereal grains in the world. Lipoxygenases (LOXs) are a group enzymes that catalyze oxygenation of polyunsaturated fatty acids (PUFAs). LOX derived oxilipins play critical roles in plant defense against pathogens such as A. flavus. The objectives of this study were to report sequence diversity and expression patterns for all LOX genes, and map their effect on aflatoxin accumulation via linkage and association mapping. Genes GRMZM2G102760 (ZmLOX 5) and GRMZM2G104843 (ZmLOX 8) fell under previously published QTL in one of four mapping populations and appear to have a measurable effect on the reduction of aflatoxin in maize grains. The association mapping result shows 19 of the total 215 SNPs found within the sequence of the ZmLOXs were associated with reduced aflatoxin levels.

Understanding the role of phytohormone auxin in defense responses is one of the vital tools for plant breeders to develop maize germplasm lines that exhibit high resistance to Aspergillus flavus and subsequent aflatoxin accumulation. Besides its critical role in different developmental processes throughout the life cycle of plants, auxin is also involved in the network of plant-pathogen interaction as demonstrated in previous studies. However, the actual mechanism for the auxin signaling pathway leading to resistance is unknown. Therefore, the critical gap in the knowledge base is a lack of understanding of the role of auxin signaling in pathogen resistance in maize. Continuation of this gap is an important problem because fungal resistance is a highly quantitative trait and breeding for resistance is a challenge. A complete understanding of the auxin mechanism in resistance could lead the production of corn hybrids with resistance to A. flavus and aflatoxin accumulation. The focus of this research was to determine the effect of exogenous auxin on A. flavus growth and production of aflatoxin in growth media. In addition, auxin levels, the amount of aflatoxin, and fungal growth in three resistant (Mp313E, Mp715, and Mp719) and one susceptible (B73) germplasm line were determined. As a result, auxin significantly increased mycelium growth and significantly decreased aflatoxin at a high concentration in potato dextrose broth under the lab conditions. Under the field conditions, auxin levels were low in resistant lines but did not change in response to A. flavus infection. Susceptible line had high auxin levels and auxin levels significantly decreased in response to A. flavus infection.

No description available.

Submitted by FERNANDA DA SILVA VON PORSTER (fdsvporster@univates.br) on 2018-01-05T11:09:04Z No. of bitstreams: 2 license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) 2017LucianeRosadaSilvaMohr.pdf: 3699054 bytes, checksum: fb9b0620e212eea8c37ccd1f068a1697 (MD5) / Approved for entry into archive by Ana Paula Lisboa Monteiro (monteiro@univates.br) on 2018-01-05T19:19:04Z (GMT) No. of bitstreams: 2 license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) 2017LucianeRosadaSilvaMohr.pdf: 3699054 bytes, checksum: fb9b0620e212eea8c37ccd1f068a1697 (MD5) / Made available in DSpace on 2018-01-05T19:19:04Z (GMT). No. of bitstreams: 2 license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) 2017LucianeRosadaSilvaMohr.pdf: 3699054 bytes, checksum: fb9b0620e212eea8c37ccd1f068a1697 (MD5) Previous issue date: 2018-01 / CAPES / PROSUP / A degradação e perda de habitats são as maiores ameaças à biodiversidade. O veste-amarela (Xanthopsar flavus) (Gmelin, 1788) é uma ave globalmente vulnerável à extinção. Habita áreas de campo e áreas abertas, sendo uma espécie prioritária para conservação e pesquisas. Esta tese apresenta informações sobre tamanho populacional, hábitos alimentares, uso de habitat, comportamento, biologia reprodutiva e sucesso reprodutivo de X. flavus em duas unidades de conservação em Viamão, RS, Brasil: a Área de Proteção Ambiental do Banhado Grande e o Refúgio de Vida Silvestre Banhado dos Pachecos, classificado como uma “Important Bird Area”. Além disso, apresenta um mapa da distribuição potencial da espécie, usando a modelagem de nicho ecológico. A procura por X. flavus foi realizada principalmente em áreas abertas e áreas úmidas. Fora das estações reprodutivas o comportamento dos indivíduos foi registrado através do método ad libitum. Durante as estações reprodutivas, pares de X. flavus foram observados através dos métodos ad libitum e de amostragem por sequências. As áreas foram visitadas mensalmente, totalizando cerca de 530h em 84 dias de observações (setembro/2014 a junho/2016), que incluíram duas estações reprodutivas. A espécie foi observada em todos os meses (não necessariamente no mesmo ano) e vários bandos de X. flavus foram registrados, alguns com mais de cem indivíduos (2-137). Duas colônias reprodutivas foram encontradas e 11 ninhos foram monitorados. O sucesso reprodutivo foi estimado em 10% na colônia 1 e em zero na colônia 2, onde todos os ovos e ninhegos foram predados. A predação parece ser o principal fator associado ao baixo sucesso reprodutivo da espécie. Baseado nessa informação, um experimento com ninhos artificiais foi realizado para verificar as taxas de predação em locais com ocorrência de X. flavus. Embora não esperado, por ser uma área de proteção ambiental, as maiores taxas de predação destes ninhos foram registradas dentro do refúgio, onde também foi detectada a menor taxa de sobrevivência diária dos ninhos. Estes fatores, associados à altura da vegetação nesta área, podem explicar a ausência da nidificação de X. flavus nesses locais, apesar da ocorrência da espécie. Outros experimentos podem ser realizados para verificar como os predadores podem impactar X. flavus e utilizar estas informações no manejo de áreas de nidificação. Além disso, baseado em registros de X. flavus no Brasil, Uruguai, Argentina e Paraguai, um mapa de distribuição potencial foi produzido usando o algoritmo Maxent, aplicando os parâmetros sugeridos pelo programa, para analisar as áreas de maior adequabilidade ambiental para a ocorrência de X. flavus. A maioria dessas áreas se encontra no Brasil, reforçando a necessidade deste país desenvolver estratégias para a conservação desta espécie. Os resultados também reforçam a importância das unidades de conservação para a manutenção de populações de X. flavus e a necessidade da preservação de áreas úmidas com gravatás (Eryngium sp.). A manutenção de áreas adequadas à reprodução e alimentação da espécie é necessária e urgente. As informações desta pesquisa estão sendo utilizadas na elaboração do plano de manejo do RVS Banhado dos Pachecos, no qual X. flavus é uma das espécies-alvo de conservação. / Habitat loss and degradation are major threats to biodiversity. The Saffron-cowled Blackbird (Xanthopsar flavus) (Gmelin, 1788) is a globally vulnerable icterid. The species inhabits grasslands and open areas, and is a priority species for research and conservation programs. This thesis provides information on the population size, feeding aspects, habitat, behavior, breeding biology and nest success of X. flavus in two conservation units in Viamão, state of Rio Grande do Sul, Brazil: the Environmental Protection Area Banhado Grande, and the Wildlife Refuge Banhado dos Pachecos, classified as an “Important Bird Area”. Also, provides a map of the species’ potential distribution using ecological niche modeling. Searches for X. flavus were carried out mainly in open areas and wetlands. Out of the breeding season individual behavior was recorded by the ad libitum method; during the breeding season, selected X. flavus pairs were observed following the sequence sampling and ad libitum method. Areas were visited once a month, totalizing approximately 530 h of observations (September 2014 to June 2016) over 84 days, which included two breeding seasons. The species was observed throughout all months (not necessarily within the same year) and several X. flavus flocks were encountered, some with more than one hundred individuals (range = 2 - 137). Two breeding colonies were found, and eleven nests were monitored. The estimated nesting success was 10% in Colony 1, but zero in Colony 2, where all eggs and nestlings were predated. Predation was found to be the main factor associated with low reproductive success of the species. Based on this information experiments were carried out with artificial nests to test predation rates of such nests in three areas within X. flavus’ range. Higher predation levels were recorded within the wildlife refuge, although this is not expected, as it is a protected area. Moreover, there, was recorded the lowest daily survival rate in the artificial nests. This, allied with the height of the vegetation within this area, may explain the absence of nesting of X. flavus at this site, despite the species being recorded there. Other experiments should be done to verify which predators could impact X. flavus, as such information can be used for management of nesting sites. In addition, with the records of X. flavus in Brazil, Uruguay, Argentina and Paraguay, a potential distribution map was produced using the Maxent algorithm, by applying the basic parameters suggested by the program, to analyze areas with environmental suitability for the occurrence of X. flavus. Most of these areas are in Brazil, which reinforces the need for strategies for the conservation of the species. The results highlight the importance of conservation units for the maintenance of X. flavus populations. They also highlight the need to preserve wetlands with Eryngium sp. as well as supporting actions to protect such areas. Maintenance of proper areas for feeding and breeding is necessary and urgent. Information from current research is being employed in the management plan of the Wildlife Refuge Banhado dos Pachecos in which X. flavus is one of the conservation target-species.