1. Fusarium roseum �Graminearum� was the predominant fungus associated with stalk rot of maize in eastern Australia in the 1972, 1973 and 1974 growing seasons. All isolates of this pathogen were of the Group 2 type. Thus Group 2 contrasts with Group I which is normally isolated :Erora crown rot of wheat and grasses. Other fungi isolated in order of frequency were Diplodia maydis, F. rnoniliforme �Subglutinans�, Bipolaris sorokiniana, Nigrospora oryzac, F. roseum �Semitectum�, F. moniliforme, F. roseum �Equiseti�, F. roseum �Concolor�, Macrophomina phaseolina, Rhizoctonia sp., F. roseum �Acuminatum�, F. oxysporum, F. solani, F. tricinctum and F. roseum �Heterosporum�. The relative isolation frequencies of the fungi varied according to the seasonal conditions. Stalk rots were not of major importance in 1973, a relatively dry growing season. However, in 1974, a wet growing season, stalk rot diseases were common in all areas investigated. 2. Isolates of F. roseum �Graminearum�,derived mainly from wheat and maize but also from other sources and from various regions of eastern Australia, were examined for perithecia formation, colony characteristics, fertility, colony growth, conidia production and conidia size. The distribution of the fungus in field colonized maize and wheat plants was also studied. The Group 1 isolates did not produce perithecia, were heterothallic and very infertile, had a mean colony growth of 4.4 cm per 3 days (range, 3.9- 5.1) and produced relatively large numbers of conidia. In contrast, Group 2 isolates were homothallic and produced perithecia readily, had a mean colony growth of 5.4 cm per 3 days (range, 4.7�6.1) and produced relatively low numbers of conidia. Group 1 isolates were found to be commonly associated with crowns and roots of plants and Group 2 isolates were commonly associated with aerial plant parts. 3. The ability of a number of Group 1 and Group 2 isolates to produce the fungal hormone, zearalenone was assessed. Group I isolates produced three to four times more zearalenone than Group 2 isolates. In addition, a. culture which had previously produced perithecia but had lost that ability following numerous transfers, produced no detectable zearalenone. The results provided good evidence that the observed difference in perithecia formation was directly related to the ability to produce zearalenone. 4. The pathogenicity to wheat, maize and carnations of Group 1 isolates from crown rot affected wheat plants and Group 2 isolates from stalk rot affected maize plants was tested. Pathogenicity of 11 other isolates from teosinte, carnations, pearl millet, wheat and barley scab, banana, ginger and common wheat grass was also assessed. The results indicated that pathogenic specialization exists within F. roseum �Graminearum�. Wheat isolates were the most pathogenic to wheat, carnation isolates were the most pathogenic to carnations and all maize isolates were pathogenic to maize while those from wheat and common wheat grass were not as pathogenic to maize. Moreover, Group 2 isolates were more pathogenic when inoculated in aerial plant parts, and the Group I isolates were more pathogenic when inoculated in plant parts in soil. Inoculations on wheat seedlings in sterile field soil demonstrated that the inherent pathogenicity to wheat seedlings of isolates from wheat and maize were similar. 5. Some factors which could contribute to the observed pathogenic differences between isolates from wheat and maize to wheat seedlings in field soil were examined. Conidia volume, germination rate and inherent germinability in the soil were studied. The Group I isolates had the largest volume, the most rapid germination and the highest inherent germinability. Pathogenicity was positively correlated with conidium volume and inherent germinability. In addition, the inherent germinability and conidium volume were positively correlated. Thus, it was established that pathogenic behaviour of conidia of Group 1 and Group 2 reflected differences in conidia morphology.

1. Fusarium roseum �Graminearum� was the predominant fungus associated with stalk rot of maize in eastern Australia in the 1972, 1973 and 1974 growing seasons. All isolates of this pathogen were of the Group 2 type. Thus Group 2 contrasts with Group I which is normally isolated :Erora crown rot of wheat and grasses. Other fungi isolated in order of frequency were Diplodia maydis, F. rnoniliforme �Subglutinans�, Bipolaris sorokiniana, Nigrospora oryzac, F. roseum �Semitectum�, F. moniliforme, F. roseum �Equiseti�, F. roseum �Concolor�, Macrophomina phaseolina, Rhizoctonia sp., F. roseum �Acuminatum�, F. oxysporum, F. solani, F. tricinctum and F. roseum �Heterosporum�. The relative isolation frequencies of the fungi varied according to the seasonal conditions. Stalk rots were not of major importance in 1973, a relatively dry growing season. However, in 1974, a wet growing season, stalk rot diseases were common in all areas investigated. 2. Isolates of F. roseum �Graminearum�,derived mainly from wheat and maize but also from other sources and from various regions of eastern Australia, were examined for perithecia formation, colony characteristics, fertility, colony growth, conidia production and conidia size. The distribution of the fungus in field colonized maize and wheat plants was also studied. The Group 1 isolates did not produce perithecia, were heterothallic and very infertile, had a mean colony growth of 4.4 cm per 3 days (range, 3.9- 5.1) and produced relatively large numbers of conidia. In contrast, Group 2 isolates were homothallic and produced perithecia readily, had a mean colony growth of 5.4 cm per 3 days (range, 4.7�6.1) and produced relatively low numbers of conidia. Group 1 isolates were found to be commonly associated with crowns and roots of plants and Group 2 isolates were commonly associated with aerial plant parts. 3. The ability of a number of Group 1 and Group 2 isolates to produce the fungal hormone, zearalenone was assessed. Group I isolates produced three to four times more zearalenone than Group 2 isolates. In addition, a. culture which had previously produced perithecia but had lost that ability following numerous transfers, produced no detectable zearalenone. The results provided good evidence that the observed difference in perithecia formation was directly related to the ability to produce zearalenone. 4. The pathogenicity to wheat, maize and carnations of Group 1 isolates from crown rot affected wheat plants and Group 2 isolates from stalk rot affected maize plants was tested. Pathogenicity of 11 other isolates from teosinte, carnations, pearl millet, wheat and barley scab, banana, ginger and common wheat grass was also assessed. The results indicated that pathogenic specialization exists within F. roseum �Graminearum�. Wheat isolates were the most pathogenic to wheat, carnation isolates were the most pathogenic to carnations and all maize isolates were pathogenic to maize while those from wheat and common wheat grass were not as pathogenic to maize. Moreover, Group 2 isolates were more pathogenic when inoculated in aerial plant parts, and the Group I isolates were more pathogenic when inoculated in plant parts in soil. Inoculations on wheat seedlings in sterile field soil demonstrated that the inherent pathogenicity to wheat seedlings of isolates from wheat and maize were similar. 5. Some factors which could contribute to the observed pathogenic differences between isolates from wheat and maize to wheat seedlings in field soil were examined. Conidia volume, germination rate and inherent germinability in the soil were studied. The Group I isolates had the largest volume, the most rapid germination and the highest inherent germinability. Pathogenicity was positively correlated with conidium volume and inherent germinability. In addition, the inherent germinability and conidium volume were positively correlated. Thus, it was established that pathogenic behaviour of conidia of Group 1 and Group 2 reflected differences in conidia morphology.

Master of Science / Department of Agronomy / Tesfaye Tesso / Sorghum [Sorghum bicolor (L.) Moench] is listed as one of the potential feedstock sources for biofuel production. While sorghum grain can be fermented into ethanol in a similar way as maize, the greatest potential of the crop is based on its massive biomass and sugar rich juices. Thus development of the crop as alternative energy source requires improvement of these traits. The objectives of this study were (1) to determine the mode of inheritance of traits related to ethanol production and identify suitable genetic sources for use in breeding programs, and (2) to evaluate the potential of low lignin mutations for biomass feedstock production and assess biotic stress risks associated with deployment of the mutations. The study consisted of three related experiments: (i) estimating the combining ability of selected sweet and high biomass sorghum genotypes for biofuel traits and resistance to stalk lodging, (ii) determine the impact of brown mid-rib mutations on biofuel production and their reaction to infection by Macrophomina phaseolina and Fusarium thapsinum, and (iii) assess the reaction of low lignin mutants to green bug feeding. In the first experiment six sorghum genotypes of variable characteristics (PI193073, PI257602, PI185672, PI195754, SC382 and SC373) were crossed to three standard seed parent lines ATx3042, ATx623 and ATx399. The resulting hybrids and the parents were evaluated at four locations, three replications during 2009 and 2010 seasons. Data were collected on phenology, plant height, juice yield, °brix score and biomass production. In the second experiment, two brown mid-rib mutations (bmr6 and bmr12) and their normal versions were studied in four forage sorghum backgrounds (Atlas, Early Hegari, Kansas Collier and Rox Orange). The experiment was planted in four replications and at 14 d after flowering five plants in a plot were artificially infected with F. thapsinum and another five with M. phaseolina. The plants were harvested and rated for disease severity (lesion length and nodes crossed). Another five normal plants in each plot were harvested and used to determine biofuel traits (juice yield, ºbrix score and biomass). In the third experiment, a subset of entries evaluated in experiment II and three tolerant and susceptible checks were tested for greenbug feeding damage. Biotype K greenbug colony was inoculated to each genotype using double sticky foam cages. Feeding damage was assessed as percent chlorophyll loss using SPAD meter. There was significant general combining ability (GCA) effect among the male entries for juice yield, stem obrix and biomass production indicating that these traits are controlled by additive genes. Lines PI257602 and PI185672 in particular, had the highest GCA for all the traits and should serve as excellent breeding materials. There was no significant difference among the bmr mutants and between the bmr and normal genotypes for both stalk rot and greenbug damage. In conclusion, juice yield, °brix and biomass are largely controlled by additive genes and hence are amenable to genetic manipulation. The bmr mutations despite their impact on lignin content do not increase risk of attack by stalk rot pathogens and greenbugs and thus can be deployed for biofuel production without incurring losses to these factors.

Colleotrichum graminicola is the causal agent of anthracnose stalk rot, and is one of the most common and aggressive pathogens of maize. The goal of my Ph.D. project was to contribute to a better understanding of the biology of the interaction between C. graminicola and its host. C. graminicola produces two type of asexual spores: one is produced on the surface of infected tissues and is thought to be involved in the spread of the disease in the field. The second type of spore, oval in shape, is produced inside the infected plant tissues, and was believed to be involved in the movement of the pathogen inside the plant tissues via the vascular system. I tested this hypothesis with both cytological and molecular approaches. I used strains of C. graminicola expressing green fluorescent proteins (GFP) to inoculate wounded plants, and followed the development of the pathogen over time. This study revealed that C. graminicola is not a vascular pathogen. C. graminicola primarily moved through the rind and vascular fibers. Oval spores were produced in colonized parenchyma cells and remained dormant, and did not appear to be involved in the movement of the pathogen, at least during the early stages of the disease development. I also studied pathogen ingress in the absence of a wound. I inoculated unwounded plants with the GFP expressing strains. C. graminicola efficiently colonized the epidermis and, given enough time, penetrated and colonized the deeper parenchyma tissues, after first moving through the fibers. To further test the role of sporulation in colonization of maize tissues, I used targeted mutagenesis to disrupt a major gene known to regulate sporulation and vegetative growth in several other fungi. The gene Cgg1, orthologue of the A. nidulans fadA, was disrupted using the split marker method. The Cgg1 mutants were less pathogenic than the wildtype to wounded plants. This was associated with an apparent increase in production of spores and primary infection hyphae. This suggests that Cgg1 signaling pathway plays a role in maximizing colonization of host tissues, and that this involves negative regulation of sporulation and primary hyphae production in planta.

Doctor of Philosophy / Department of Agronomy / Tesfaye Tesso / Fusarium stalk rot and charcoal rot caused by Fusarium thapsinum and Macrophomina phaseolina respectively are devastating global diseases in sorghum that lead to severe quality and yield loss each year. In this study, three sets of interrelated experiments were conducted that will potentially lead to the development of resistance based control option to these diseases. The first experiment was aimed at identifying sources of resistance to infection by M. phaseolina and F. thapsinum in a diverse panel of 300 sorghum genotypes. The genotypes were evaluated in three environments following artificial inoculation. Out of a total of 300 genotypes evaluated, 95 genotypes were found to have resistance to M. phaseolina and 77 to F. thapsinum of which 53 genotypes were resistant to both pathogens. In the second experiment, a set of 79,132 single nucleotide polymorphisms (SNPs) markers were used in an association study to identify genomic regions underlying stalk rot resistance using a multi-locus mixed model association mapping approach. We identified 14 loci associated with stalk rot and a set of candidate genes that appear to be involved in connected functions controlling plant defense response to stalk rot resistance. The associated SNPs accounted for 19-30% of phenotypic variation observed within and across environments. An analysis of associated allele frequencies within the major sorghum subpopulations revealed enrichment for resistant alleles in the durra and caudatum subpopulations compared with other subpopulations. The findings suggest a complicated molecular mechanism of resistance to stalk rots. The objective of the third experiment was to determine the functional relationship between stay-green trait, leaf dhurrin and soluble sugar levels and resistance to stalk rot diseases. Fourteen genotypic groups derived from a Tx642 × Tx7000 RIL population carrying combinations of stay-green quantitative trait loci were evaluated under three environments in four replications. The stg QTL had variable effects on stalk rot disease. Genotypes carrying stg1, stg3, stg1,3 and stg1,2,3,4 expressed good levels of resistance to M. phaseolina but the combination of stg1 and stg3 was required to express the same level of resistance to F. thapsinum. Other stg QTL blocks such as stg2 and stg4 did not have any impact on stalk rot resistance caused by both pathogens. There were no significant correlations between leaf dhurrin, soluble sugar concentration, and resistance to any of the pathogens.

O gênero Fusarium é um grupo de fungos de importância mundial, não só por causar patologias em plantas, mas também porque abriga espécies toxigênicas. Dentre estas, Fusarium verticillioides, Fusarium proliferatum e Fusarium subglutinans, cujos teleomorfos se agrupam no complexo Gibberella fujikuroi, estão associadas a patologias em milho. O presente estudo caracterizou uma coleção de 100 isolados de Fusarium spp. tanto do ponto de vista morfológico como molecular com vistas a identificá-los em nível de espécie. Para isto, foram usados como marcadores morfológicos a presença/ausência de clamidósporos, o tipo de célula conidiogênica e a disposição dos microconídios sobre a célula conidiogênica. Os isolados foram também identificados em espécie baseado em informações disponíveis na literatura acerca de marcadores moleculares desenvolvidos a partir da reação de PCR e primers espécie-específicos. A ausência de clamidósporo permitiu alocar a totalidade dos isolados no complexo G. fujikuroi. Os demais critérios morfológicos permitiram identificar 77 isolados como F. verticillioides, 20 como F. proliferatum, 2 como F. subglutinans. Apenas 1 isolado não foi possível identificar em espécie. Análises moleculares concordaram em 100% dos casos em que os isolados foram identificados como F. verticillioides e F. subglutinans. Porem, no caso dos 20 isolados identificados morfologicamente como F. proliferatum apenas 4 foram confirmados na análise molecular; os demais foram identificados como F. verticillioides. A diversidade genética estudada por AFLP ratificou a separação das espécies F. verticillioides e F. proliferatum, com um índice de similaridade de 0,40. Marcadores AFLP também evidenciaram alta diversidade genética de F. verticillioides. Todos os isolados causaram podridão do colmo em três híbridos comerciais de milho e não variaram em agressividade, independente do nível de resistência dos híbridos. / The genus Fusarium is an important group of fungi worldwide, not only for its capability to cause disease but because it also contains species which produce toxins. Among these, Fusarium verticillioides, Fusarium proliferatum and Fusarium subglutinans, which teleomorphs are grouped in the Gibberella fujikuroi group are associated with diseases in maize. This study characterized a collection of 100 isolates of Fusarium spp. under morphological and molecular criteria to identify them at the species level. For this purpose, morphological markers such as presence/absence of chlamydospores, conidiogenou cell type and microconidia arrangement on conidiogenou cell were assessed. The isolates were also identified at the species level based upon available information about molecular markers developed from the PCR reaction using speciesspecific primers. The absence of chlamydospores in all of the isolates placed them within the G. fujikuroi complex, and based on the other morphological criteria, 77 isolates were identified as F. verticillioides, 20 as F. proliferatum, 2 as F. subglutinans and one isolate remained unidentified at the species level. The molecular analyses agreed with the morphological identification of all F. verticillioides and F. subglutinans. However, in the case of 20 isolates identified morphologically as F. proliferatum only 4 were confirmed, the rest being identified as F. verticillioides. The genetic diversity based on AFLP confirmed the separation of F. verticillioides and F. proliferatum in two groups, with a similarity index of 0.40. AFLP markers also showed high genetic diversity within F. verticillioides. All isolates were caused stalk rot on three commercial hybrids and did not vary in aggressiveness regardless of the resistance level of the hybrids.

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / The expansion of area under maize (Zea mays L.), and the use of no-till without a proper crop rotation have increased the incidence of stalk rot on this crop, due to the survival of this disease-causing pathogens (Fusarium spp., Diplodia spp., Colletotrichum graminicola, among others) in soil and crop residues. Fertilization has shown positive effect on the crop, but little is known about its relation to these pathogens and the organisms that control them. The objective of this study was to evaluate the effect of short and long-term of organic and mineral fertilizers and the effect of treatment of corn seeds with Trichoderma spp. Persson on the population of total fungi, Fusarium spp., and Trichoderma spp. in the soil and the production of dry matter (DM), accumulation of nitrogen (N), phosphorus (P) and potassium (K) in the tissue and grains, the incidence of stalk rot and corn yield. The experiment was conducted at the experimental area of the Soils Department, Federal University of Santa Maria, which has a history of six years application of organic and mineral fertilizers, and consisted in a factorial 5 (fertilization treatments) X 2 (seeds treatment) with split-plot in strips, using the randomized block design. The fertilization treatments (pig slurry; swine deep bedding; cattle slurry; mineral fertilizer; control treatment) were applied to the plots and the seeds treatment (with and without Trichoderma spp.) in the subplots (strips). It was used the hybrid Pioneer ® 30F53 Y, with 45 cm row spacing and final population of 78,000 plants per hectare. Soil samples were collected before application of treatments, for chemical analysis, and before and after, to characterize the soil fungal population. In the laboratory, dilution of samples and quantification of the fungi present was performed. At the flowering stage, three corn plants per subplot were collected for the assessment of DM production and nutrient accumulation in the tissue. At physiological maturity stage, the incidence of stalk rot was assessed, and the ears of corn harvested for productivity assessment. Nutrient content in corn kernels was also determined. The organic fertilizers pig slurry and swine deep bedding, in the long-term, increased population of Fusarium spp. in the soil. The organic and mineral fertilizers increased the production of DM and productivity of corn. Trichoderma spp. increased the production of DM of corn, but had no reflection on productivity. Both factors increased the incidence of stalk rot in corn. The swine deep bedding and cattle slurry increased the content of P and K in the tissue of corn plants. It is concluded that: the organic fertilizers can be used to increase the production of DM and productivity of corn and; Trichoderma spp. can be used if the objective is greater DM production. / A expansão da área cultivada com milho (Zea mays L.) e a utilização do plantio direto sem uma rotação de culturas adequada têm aumentado a incidência de podridões de colmo nessa cultura, devido à sobrevivência dos patógenos causadores da doença (Fusarium spp., Diplodia spp., Colletotrichum graminicola, entre outros) no solo e em restos culturais. A adubação tem apresentado efeitos positivos sobre a cultura, mas pouco se sabe sobre sua relação com esses patógenos e os organismos que promovem seu controle. O objetivo do presente trabalho foi avaliar o efeito a curto e longo prazos da adubação orgânica e o efeito do tratamento de sementes de milho com Trichoderma spp. sobre a população de fungos totais, Fusarium spp. e Trichoderma spp. no solo e a produção de matéria seca (MS), acúmulo de nitrogênio (N), fósforo (P) e potássio (K) no tecido e nos grãos, a incidência de podridões de colmo e a produtividade de milho. O experimento foi conduzido na área experimental do Departamento de Solos da Universidade Federal de Santa Maria, que possui histórico de seis anos de aplicação de adubos orgânicos e mineral, e constituiu-se em um fatorial 5 (tratamentos de adubação) X 2 (tratamento das sementes), com parcelas subdivididas em faixas, sendo utilizado o delineamento blocos casualizados. Os tratamentos de adubação (dejeto líquido de suínos; cama sobreposta de suínos; dejeto líquido de bovinos; adubação mineral; tratamento testemunha) foram aplicados nas parcelas e o tratamento das sementes (com e sem Trichoderma spp.) nas subparcelas (faixas). Foi utilizado o híbrido 30F53 Y, da empresa Pioneer®, espaçamento entre linhas de 45 cm e população final de 78000 plantas por hectare. Amostras de solo foram coletadas antes, para análise química, e antes e após a aplicação dos tratamentos para caracterização da população fúngica do solo. Em laboratório, foi realizada a diluição serial das amostras e quantificação dos fungos presentes. No estádio de florescimento foram coletadas três plantas de milho por subparcela para a avaliação da produção de matéria seca e acúmulo de nutrientes no tecido. No estádio de maturação fisiológica, foram realizadas a avaliação da incidência de podridões de colmo e a colheita das espigas para o cálculo da produtividade. Também foi determinado o teor de nutrientes nos grãos de milho. Os adubos orgânicos dejeto líquido de suínos e cama sobreposta de suínos, em longo prazo, aumentaram a população de Fusarium spp. no solo. Os adubos orgânicos e mineral aumentaram a produção de MS e a produtividade de milho. Trichoderma spp. aumentou a produção de MS de milho, porém não teve reflexo sobre a produtividade. Os adubos orgânicos aumentaram a incidência de podridões de colmo no milho. A cama sobreposta de suínos e o dejeto líquido de bovinos aumentaram o teor de P e K no tecido das plantas de milho. Conclui-se que: os adubos orgânicos podem ser utilizados para aumentar a produção de MS e a produtividade de milho e; Trichoderma spp. pode ser utilizado quando se objetiva maior produção de MS.

Made available in DSpace on 2014-06-11T19:35:00Z (GMT). No. of bitstreams: 0 Previous issue date: 2004-12-16Bitstream added on 2014-06-13T20:45:44Z : No. of bitstreams: 1 fortes_nlp_dr_botfca.pdf: 629205 bytes, checksum: 40d1e91cf18ab3678f66c246588008c7 (MD5) / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / Universidade Estadual Paulista (UNESP) / O trabalho objetivou verificar os fatores químicos e microbiológicos do lodo de esgoto e do solo responsáveis pela incidência da podridão do colmo do milho, causada por Fusarium em solo fertilizado com lodo, e foi desenvolvido em condições de campo, numa área que vem sendo tratada com lodo e cultivada com milho desde 1999 em latossolo vermelho distróférrico (textura argilosa), localizada no município de Jaguariúna/SP. Os dois lodos utilizados foram obtidos nas Estações de Tratamento de Esgoto de Franca e de Barueri/SP. Os lodos foram incorporados ao solo nas concentrações de 0, 1, 2, 4 e 8 vezes a dose recomendada com base no teor de nitrogênio, em dois cultivos sucessivos de milho (Savana 133S) nas safras 2001/2002 e 2002/2003, em parcelas de 200 m2. As doses de lodo foram comparadas com a adubação mineral recomendada para a cultura do milho. Os tratamentos foram distribuídos em delineamento experimental casualizado em blocos e com três repetições. Foram verificadas, nas safras de 2001/2002 e 2002/2003, diferenças significativas entre as doses de lodo de esgoto, sobre a variação de Fusarium spp. no solo, na rizosfera, na raiz, no colmo e na semente, bem como sobre a incidência de plantas com podridão no colmo e nas espigas. As análises de regressão mostraram que a porcentagem de plantas doentes foi positivamente correlacionada com a concentração dos lodos incorporados ao solo. Os coeficientes de determinação para o cultivo de 2001/2002 foram de R2=0,61 e R2=0,32, enquanto para o cultivo de 2002/2003 foram de R2=0,76 e R2=0,45, para os lodos de Franca e Barueri, respectivamente. As concentrações de lodos também apresentaram correlação positiva com a comunidade de Fusarium do solo e da rizosfera e com a incidência de Fusarium na raiz, no colmo e na semente. As correlações entre a comunidade de Fusarium do solo e da rizosfera e o percentual de plantas... . / This work aimed at verifying the chemical and microbiological sewage sludge and soil factors re 0sponsible for the incidence of the corn stalk rot caused by Fusarium in soil fertilized with sludge and has been developed in field conditions, in an area that has been treated with sludge and cultivated with corn since 1999, in red dystroferric latosol (clayey texture), in Jaguariúna/SP. Both sludges used were obtained in the sewage sludge treatment stations of Franca and Barueri/SP. Sludges were incorporated into the soil in concentrations of 0, 1, 2, 4 and 8 times in relation to the recommended rate based on the nitrogen contents, in two successive corn cultures (Savana 133S), in 2001/2002 and 2002/2003, in plots of 200 m2. Sludge doses were compared to the mineral fertilization recommended to the corn culture. The treatments were distributed in a randomized design experimental drawing in blocks and with three repetitions. Significant differences were verified in both 2001/2002 and the 2002/2003 crops among the sewage sludge doses about the variation of Fusarium spp. in soil, root, stalk and seed, as well as about the incidence of plants with stalk and corn rot. The regression analyses have showed that the percentage of diseased plants was positively correlated to the concentration of the sludges incorporated to the soil. The determination coefficients to the 2001/2002 culture were R2=0.61 and R2=0.32, while the 2001/2003 culture showed R2=0.76 and R2=0.45, to Franca and Barueri sludges, respectively. The sludges concentrations also presented positive correlation with the Fusarium community of the soil and the rhizosphere, and with the incidence of Fusarium in the root, stalk and seed. The correlations between the Fusarium community of the soil and rhizosphere and the percentage of diseased plants with chemical soil attributes were significant and positive, for both kinds of sludges, in the level of 1 and 5%, to phosphorus and calcium contents.

O gênero Fusarium é um grupo de fungos de importância mundial, não só por causar patologias em plantas, mas também porque abriga espécies toxigênicas. Dentre estas, Fusarium verticillioides, Fusarium proliferatum e Fusarium subglutinans, cujos teleomorfos se agrupam no complexo Gibberella fujikuroi, estão associadas a patologias em milho. O presente estudo caracterizou uma coleção de 100 isolados de Fusarium spp. tanto do ponto de vista morfológico como molecular com vistas a identificá-los em nível de espécie. Para isto, foram usados como marcadores morfológicos a presença/ausência de clamidósporos, o tipo de célula conidiogênica e a disposição dos microconídios sobre a célula conidiogênica. Os isolados foram também identificados em espécie baseado em informações disponíveis na literatura acerca de marcadores moleculares desenvolvidos a partir da reação de PCR e primers espécie-específicos. A ausência de clamidósporo permitiu alocar a totalidade dos isolados no complexo G. fujikuroi. Os demais critérios morfológicos permitiram identificar 77 isolados como F. verticillioides, 20 como F. proliferatum, 2 como F. subglutinans. Apenas 1 isolado não foi possível identificar em espécie. Análises moleculares concordaram em 100% dos casos em que os isolados foram identificados como F. verticillioides e F. subglutinans. Porem, no caso dos 20 isolados identificados morfologicamente como F. proliferatum apenas 4 foram confirmados na análise molecular; os demais foram identificados como F. verticillioides. A diversidade genética estudada por AFLP ratificou a separação das espécies F. verticillioides e F. proliferatum, com um índice de similaridade de 0,40. Marcadores AFLP também evidenciaram alta diversidade genética de F. verticillioides. Todos os isolados causaram podridão do colmo em três híbridos comerciais de milho e não variaram em agressividade, independente do nível de resistência dos híbridos. / The genus Fusarium is an important group of fungi worldwide, not only for its capability to cause disease but because it also contains species which produce toxins. Among these, Fusarium verticillioides, Fusarium proliferatum and Fusarium subglutinans, which teleomorphs are grouped in the Gibberella fujikuroi group are associated with diseases in maize. This study characterized a collection of 100 isolates of Fusarium spp. under morphological and molecular criteria to identify them at the species level. For this purpose, morphological markers such as presence/absence of chlamydospores, conidiogenou cell type and microconidia arrangement on conidiogenou cell were assessed. The isolates were also identified at the species level based upon available information about molecular markers developed from the PCR reaction using speciesspecific primers. The absence of chlamydospores in all of the isolates placed them within the G. fujikuroi complex, and based on the other morphological criteria, 77 isolates were identified as F. verticillioides, 20 as F. proliferatum, 2 as F. subglutinans and one isolate remained unidentified at the species level. The molecular analyses agreed with the morphological identification of all F. verticillioides and F. subglutinans. However, in the case of 20 isolates identified morphologically as F. proliferatum only 4 were confirmed, the rest being identified as F. verticillioides. The genetic diversity based on AFLP confirmed the separation of F. verticillioides and F. proliferatum in two groups, with a similarity index of 0.40. AFLP markers also showed high genetic diversity within F. verticillioides. All isolates were caused stalk rot on three commercial hybrids and did not vary in aggressiveness regardless of the resistance level of the hybrids.

Submitted by Claudia Rocha (claudia.rocha@udesc.br) on 2018-02-21T14:08:34Z No. of bitstreams: 1 PGPV15MA187.pdf: 174820 bytes, checksum: ee57afc9c4d2a1b5e1435af1a2438825 (MD5) / Made available in DSpace on 2018-02-21T14:08:34Z (GMT). No. of bitstreams: 1 PGPV15MA187.pdf: 174820 bytes, checksum: ee57afc9c4d2a1b5e1435af1a2438825 (MD5) Previous issue date: 2015-02-27 / Capes / The objective of this study was to evaluate the incidence and damage of stalk rot (SR) in corn hybrids grown in tillage rotation with beans and succession to oats; quantify the efficiency of detection and control of Fusarium verticillioides in seed and seedlings health test according to the time of incubation. The first study was carried out at Muitos Capões municipal district on Rio Grande do Sul State, during the agricultural seasons of 2012/13 and 2013/14.The hybrids tested were P30F53 Hx, P32R22 YHR, DKB 250 VTPRO2, AS1555 VTPRO2, AS 1656 VTPRO2, AG8025 VTPRO e AG 9045 VTPRO. A randomized block design with four replications was employed. The incidence of SR was determined during harvesting and the damage by threshing operation analyzed separately in healthy and unhealthy plants. The means were compared by Scott-Knott test with a significance level of 5% (P <0.05). Statistical interaction between corn hybrid and harvesting season was detected. For this reason, the data was analyzed by harvesting season. In the 2012/13 growing season the prevailing diseases were diplodia blight, Anthracnosis and Fusarium, and as opposed to 2013/14, when Gibberella, Diplodia blight and Fusarium wilting were the prevailing in 2013/14. The incidence of SR in 2012/13 and 2013/14 ranged from 7.3% to 100% (46.2% in average), and from 3.14% to 72.6% (34.8% in average). The average of damage in the yield of hybrids were 6,48% and 8,75%, ranging from 197 kg ha-1 to 1205 kg ha-1 and from 113 kg ha-1 to 2536 kg ha-1, with an average of 640 kg ha-1 and 768 kg ha-1, respectively in 2012/2013 and 2013/14. In both growing seasons the hybrids P32R22 YHR and AG9045 VTPRO showed the highest SR incidence. The lowest incidence of this disease was detected in the P30F53 Hx and AG8025 VTPRO hybrids in 2012/13 and AG8025 VTPRO and DKB 250 VTPRO2 in 2013/14. The highest damages were found in AS1555 VTPRO2 (14%) and AG9045 VTPRO (11.2%) in 2012/13, and AG9045 VTPRO (26.3%) in 2013/14. The yield ranged from 8.430 kg ha-1 to 10.667 kg ha-1 in 2012/13 and 7.175 kg ha-1 and 8.790 kg ha-1 in the 2013/14. Secondly, hybrid corn seeds Status TL TG treated with metalaxyl-m + fludioxonil fungicides and seeds without seed treatment were cultured in PDA Culture Media to obtain symptomatic seedlings (infected by F. verticullioides) and asymptomatic, with 7 days of age. The samples were separated and transferred to test tubes containing agar-water culture media and maintained in growth chamber at 25 °C under 12 hours of photoperiod until the completion of 20 days. The treatments were T1 - uninfected seedlings without TS; T2 - infected seedlings without TS; T3 - uninfected seedlings with TS; T4 - infected seedlings with TS. The fungus incidence was quantified in the mesocotyl, crown and remaining seed and shoot and root length and dry matter weight were measured. Randomized block experimental design with eight replications was used. The data was submitted to variance analysis and Tukey’s test at 5% of significance level. The incidence of F. verticillioides in the remaining seeds was 96% for T1 and T3, and 100% for T2 and T4, with no difference between treatments. The incidence of F. verticillioides was 88% and 93% for mesocotyl, and 88% and 95% in the crown in the T1 and T3 seedlings, respectively. The root length showed 14.1 cm and 10.7 cm for T3 and T4 respectively, which are higher than T1 (9.6 cm) and T2 (5.8 cm). Higher root weight was detected in T3 (0.25 g) and T4 (0.18 g) cultivars compared to T1 (0.12 g) and T2 (0.09 g). T3 showed the highest shoot length (14.2 cm) when compared to the other treatments. The corn seed treatment with metalaxyl-m + fludioxonil fungicides is not effective in controlling F. verticillioides. The detection of F. verticillioides in BDA media at the seventh day of incubation does not represent the real incidence of the fungus, once the pathogen was recovered from asymptomatic seedlings when they were re-evaluated at 20 days. In all treatments, the seed treatment enhanced the seedlings development / Os objetivos deste trabalho foram avaliar a incidência e os danos causados por podridões do colmo (PC) em híbridos de milho cultivados em sistema de semeadura direta com rotação ao feijão e sucessão à aveia; e quantificar a eficiência de detecção e controle de Fusarium verticillioides em meio batata-dextrose-ágar em sementes e plântulas de milho em função do tempo de incubação. Foram realizados dois estudos, sendo estes conduzidos no município de Muitos Capões, estado do Rio Grande do Sul, nas safras agrícolas 2012/13 e 2013/14. Foram avaliados os híbridos P30F53 Hx, P32R22 YHR, DKB250 VTPRO2, AS1555 VTPRO2, AS1656 VTPRO2, AG8025 VTPRO e AG9045 VTPRO. O delineamento experimental foi de blocos casualizados com quatro repetições. A incidência de PC foi determinada na colheita do milho, sendo o dano mensurado pela trilha em separado de espigas das plantas sadias e plantas doentes. A comparação das médias foi realizada pelo teste de Scott-Knott, em nível de significância de 5% (p<0,05). Houve interação entre híbridos e safras, sendo os dados analisados separadamente por safra. Em 2012/13, as doenças predominantes foram diplodia, antracnose e fusariose; em 2013/14, houve predomínio de giberela, diplodia e fusariose. A incidência de PC em 2012/13 e 2013/14 variou de 7,3% a 100% (média de 46,2%) e de 3,14% a 72,6% (média de 34,8%), respectivamente. O dano médio no rendimento de grãos dos híbridos foi de 6,8% e 8,7%, respectivamente para 2012/13 e 2013/14. Nas duas safras os híbridos P32R22 YHR e AG9045 VTPRO apresentaram maior incidência de PC. As menores incidências foram detectadas nos híbridos P30F53 Hx e AG8025 VTPRO na safra de 2012/13, e AG8025 VTPRO e DKB250 VTPRO2 na safra de 2013/14. Os maiores danos foram encontrados nos híbridos AS1555 VTPRO2 (14%) e AG9045 VTPRO (11,2%) em 2012/13, e AG9045 VTPRO (26,3%) em 2013/14. No segundo estudo, sementes de milho tratadas com fungicida metalaxil-m + fludioxonil e sem tratamento foram semeadas em meio de cultura de batata-dextrose-ágar para obtenção de plântulas sintomáticas (infectadas por F. verticillioides) e assintomáticas. Aos sete dias as plântulas foram separadas e transferidas para tubos de ensaio contendo meio de cultura ágar-água e mantidas em câmara de crescimento com temperatura de 25ºC e fotoperíodo de 12 h, onde permaneceram até completarem 20 dias. Obtiveram-se os tratamentos: T1 - plântulas assintomáticas sem fungicida; T2 - plântulas sintomáticas sem fungicida; T3 – plântulas assintomáticas com fungicida; T4 – plântulas sintomáticas com fungicida. A incidência foi quantificada no mesocótilo, coroa e semente remanescente das plântulas e o comprimento e peso seco da parte aérea e raiz. Os dados foram submetidos à análise de variância e teste de Tukey a 5%. A incidência de F. verticillioides em sementes remanescentes foi de 96% para T1 e T3, e 100% para T2 e T4, não diferindo os tratamentos entre si. Foi observada incidência de F. verticillioides de 88 e 93 % para mesocótilo, e 88 e 95% para coroa em plântulas provenientes de T1 e T3, respectivamente. O comprimento de raiz foi de 14,1 cm e 10,7 cm para T3 e T4 respectivamente, sendo estes superiores ao T1 (9,6 cm) e T2 (5,8 cm). Foram detectados maiores pesos de raiz nos tratamentos T3 (0,25 g) e T4 (0,18 g), quando comparados a T1 (0,12 g) e T2 (0,09 g). O T3 apresentou o maior comprimento de parte aérea,14,2 cm, em relação aos demais tratamentos. O tratamento de sementes com metalaxil-m + fludioxonil não foi eficiente no controle de F. verticillioides. A detecção de F. verticillioides em meio BDA aos sete dias de incubação não representou a real incidência do fungo, pois o patógeno foi recuperado de plântulas até então assintomáticas quando essas foram reavaliadas aos 20 dias. De qualquer forma o tratamento proporcionou maior comprimento e peso de raiz e parte área em plântulas de milho