Dissertations / Theses on the topic 'Anthracnose disease'

The first confirmation of the presence of Colletotrichum gloeosporioides Penz. on cashew in Mozambique was based on a combination of observed symptoms, isolation and identification using basic morphological and molecular techniques. Anthracnose is now the second most important in the country, after powdery mildew caused by Oidium anacardii Noack. The present thesis represents a broad overview of the disease in Mozambique. The main focus of this study was thus to gather scientific information on the relevance of this disease in the country and through experimentation, generate recommendations that help farmers and decision makers to mitigate the disease pressure. The specific objectives of this study were as follows: - Provide a distinctive description of anthracnose symptoms on leaves through hostpathogen interaction studies in the laboratory. - Enhance current knowledge on the identity of Mozambican pathogen isolates, using DNA tools. - Assess the current anthracnose management practices, both at nursery and field level with a view to formulate timely, local and adequate management strategies. - Conduct experimental trials to select economically effective fungicides spraying programs for anthracnose disease management. ii - Search for variability and germplasm tolerance among dwarf and common cashew plant populations in Mozambique. By analyzing and integrating existing published literature on the subject, we successfully separated issues that concerned previously inaccessible information from those that reflect insufficient scientific knowledge. A survey was initiated to determine, the status of cashew anthracnose disease management practices in Mozambique. Subsequently, the information obtained was used to develop a national strategic framework for research and extension in the country. Areas identified as gaps were aligned with the main goals of this thesis and include: - Areas where scientific information lacked were identified. - The symptoms of the disease on leaves were successfully and distinctively distinguished from other common leaf diseases that simultaneously occur in orchards. - The pathogen isolates were identified using PCR techniques. The presence of Colletotrichum acutatum Simmonds was not confirmed at least not among the suspected and tested isolates. - Knowledge on the epidemiology of the disease was generated and its application for more effective disease management was successfully applied. - Effective fungicide applications and disease control programmes were developed for Colletotrichum gloeosporioides Penz.. - Appropriate nursery management strategies that reduce anthracnose disease development were developed. - Variability in germplasm reaction to the disease was demonstrated and therefore tolerant and susceptible genotypes were identified. - A technique for rapid and accurate evaluation of leaf anthracnose symptom grades was developed.
Thesis (PhD)--University of Pretoria, 2013.
gm2014
Microbiology and Plant Pathology
Unrestricted

Anthracnose is one of the serious diseases of cowpea (Vigna unguiculata L. Walp) and common bean (Phaseolus vulgaris L.) caused by the Colletotrichum fungi. The disease is prevalent is small holder farmers’ fields due to the scarcity and high cost of the synthetic fungicides. This study was conducted with the main aim of improving food security and income of the smallholder farmers by increasing legumes, P. vulgaris and V. unguiculata thereby increasing production and improve food security and income of smallholder farmers. Investigations involved in vitro bioassaying for antifungal activities of the crude extracts on Colletotrichum lindemuthianum (Sacc. and Magn.) Bri. and Cav. and Colletotrichum dematium (Fr.) Grove var. truncata field isolates and evaluating the effect of crude plant extracts seed treatments on seed germination, emergence and control of anthracnose disease of common bean and cowpea. Furthermore, ultra-structural changes of plant extracts treated and efficacy of foliar application of extracts. The in vitro study showed that Allium sativum L., Agapanthus caulescens Spreng., Carica papaya L. and Syzygium cordatum Hochst.ex Krauss extracts have good antifungal activities against both C. lindemuthianum and C. dematium. The low concentrations (5 mg.ml-1) of Syzygium and Agapanthus water extracts and acetone extracts of Agapanthus and Carica gave a high percentage of bean seed germination, emergence, short mean emergence time (MET) and were effective in controlling the anthracnose disease. The treatment of Agapanthus (both water and acetone) extracts also increased the shoot length and dry weight of the seedlings. The Allium acetone extracts (5 mg.ml-1) was the only treatment that gave good results with respect to germination percentages, MET, shoot length, leaf area and dry mass of cowpea. Five mg.ml-1 concentrations of Syzygium and Agapanthus water extracts and acetone extracts of Agapanthus and Carica have potential as seed treatments on bean. Allium acetone extract (5 mg.ml-1) was the only potential cowpea seed treatment that could be recommended to farmers as an alternative to the synthetic fungicide. Electron microscopy revealed that principle differences were observed in the cotyledon-embryo connecting tissues of seeds treated with Agapanthus, which had few cristae in their mitochondria than the cells from other treatments. The embryonic root cells of bean seeds treated with Agapanthus had coalescing protein bodies. The embryonic root cells of cowpea and bean treated with Syzygium had fewer lipid bodies as compared to the control and the Agapanthus treated seeds. Bean plants that were foliar treated with the 15 mg.ml-1 concentrations of Allium water, Agapanthus water, Carica water, Agapanthus acetone, Carica 5 and 15 mg.ml-1 acetone, Syzygium 5 mg.ml-1 acetone extracts and the combinations (2.5 mg.ml-1 + 2.5 mg.ml-1) of Allium + Agapanthus, Allium + Carica, Agapanthus + Syzygium and Carica + Syzygium extracts registered low anthracnose (C. lindemuthianum,) disease severity and high leaf area. The cowpea plants treated with 15 mg.ml-1 water extracts of Agapanthus and the combinations of Allium + Agapanthus, Agapanthus + Carica and Agapanthus + Syzygium extracts recorded low cowpea anthracnose (C. dematium) disease severity, highest leaf area and dry mass. The study revealed that A. sativum, Agapanthus, C. papaya and S. cordatum plant extracts have antifungal activities and can be used as alternative seed treatments and foliar fungicides against the anthracnose diseases of legumes (cowpea and common bean) instead of synthetic fungicides without causing any negative effect on seed germination, emergence, ultra-structure of seeds and plant growth. Copyright
Dissertation (MSc)--University of Pretoria, 2012.
Microbiology and Plant Pathology
Unrestricted

Duas espécies de Colletotrichum podem causar antracnose em goiabas: C. gloeosporioides e C. acutatum. Apesar de ser a principal doença pós-colheita da cultura, a influência de variáveis ambientais no seu desenvolvimento é desconhecida. O objetivo do presente trabalho foi determinar a influência das variáveis ambientais no desenvolvimento in vitro e nos processos de infecção e colonização dos fungos Colletotrichum gloeosporioides e C. acutatum em goiabas. A germinação e a formação de apressórios foram determinadas sob temperaturas de 10, 15, 20, 25, 30, 35 e 40 ºC, com períodos de molhamento de 6, 12 e 24 horas, sob escuro contínuo. Nos experimentos in vivo, goiabas \"Kumagai\" e \"Pedro Sato\" foram inoculadas, por ferimento, com suspensão de conídios das duas espécies e incubadas em câmaras de crescimento a 15, 20, 25 e 30 ºC e períodos de molhamento de 6 e 24 horas. Avaliou-se a incidência de frutos doentes, o diâmetro das lesões, a taxa de progresso da doença e os períodos de incubação e latência. Nas goiabas \"Kumagai\" também foi avaliada a influência dos estádios de maturação dos frutos no progresso da doença. Não houve germinação a 40 ºC em nenhuma das duas espécies. A faixa favorável à germinação e à formação de apressórios in vitro foi de 15 a 30 ºC para C. gloeosporioides, com máximo a 25 ºC e de 20 a 25 ºC para C. acutatum, com máximo a 20 ºC. Para C. acutatum, a germinação foi mais sensível a variações no período de molhamento, sendo significativamente menor com 6 horas em relação a 12 e 24 horas. Nos experimentos in vivo, temperaturas de 25 e 30 ºC e 24 horas de molhamento foram mais favoráveis para as variáveis analisadas em goiaba \"Kumagai\". Os diâmetros máximos de lesão foram de 4,0 cm para C. gloeosporioides e 4,1 cm para C. acutatum, em frutos em ponto de colheita, incubados sob temperatura de 25 ºC. A maior incidência da doença (100%) ocorreu 10 dias após a inoculação, a 30ºC e 24 horas de molhamento. O menor período de incubação foi de 7 dias para as duas espécies, observado a 30 ºC e o menor período de latência foi de 10 e 9 dias para C. gloeosporioides e C. acutatum, respectivamente, sob temperaturas de 25 ou 30 ºC. Em goiabas \"Pedro Sato\", as temperaturas entre 20 e 30 ºC e 24 horas de molhamento foram mais favoráveis. Os diâmetros máximos de lesão foram de 3,3 cm para C. gloeosporioides e 3,2 cm para C. acutatum sob temperatura de 25 ºC. A maior incidência da doença (100%) ocorreu 10 dias após a inoculação, a 25 e 30ºC sob 6 horas de molhamento. O período de incubação foi de 7 dias para as duas espécies entre 20 e 30 ºC e o período de latência foi de 8 dias para C. gloeosporioides e 9 dias para C. acutatum sob temperaturas de 25 e 30 ºC. As condições requeridas para as duas espécies fúngicas foram semelhantes, embora o intervalo de favorabilidade seja mais amplo na goiaba \"Pedro Sato\".
The main causal agents of Anthracnose in guava are Colletotrichum gloeosporioides and C. acutatum. Although anthracnose is the main postharvest disease affecting guava, little is known about the influence of environmental variables on its development. Consequently, the objective of the present study was to determine the influence of environmental factors on in vitro development and on colonization and infection processes of C. gloeosporioides and C. acutatum fungi in guava. The germination and apressorium formation were determined at temperatures of 10, 15, 20, 25, 30, 35 and 40 °C, with wetness durations of 6, 12 or 24 hours under continuous darkness. The in vivo experiments involved puncturing the skin of the Kumagai and Pedro Sato varieties of guava with a needle followed by inoculation with conidial suspensions of C. gloeosporioides and C. acutatum. Fruits were then incubated in growth chambers at temperatures of 15, 20, 25 and 30 °C with wetness duration of 6 and 24 hours. Assessments were made of the following: incidence of disease, lesion diameter, rate of disease progress, as well as incubation and latency periods. In the Kumagai variety, the influence of maturity on disease progression was also evaluated. There was no germination at 40 oC in any of the species. The germination and apressorium formation rate were rather high in the range of 15 to 30 ºC for C. gloeosporioides, with a maximum at 25 ºC and of 20 to 25 ºC for C. acutatum, with a maximum at 20 ºC. For the species C. acutatum, germination rate was more sensitive to variations in wetting periods, thus significantly smaller with 6 hours on 12 and 24 hours. Temperatures of 25 and 30 °C were found to be more favorable for the variables analyzed in the in vivo experiments of Kumagai variety. The maximum lesion diameter recorded in this variety was 4.0 cm for C. gloeosporioides and 4.1 cm for C. acutatum in harvest ready fruit that had been incubated at temperatures lower than 25 °C. The highest incidence of the disease (100%) occurred 10 days after inoculation, at 30 º C and 24 hours of wetting. The lowest incubation period for both species was 7 days at 30 °C and the lowest latency period of 9 days for C. gloeosporioides and 10 days for C. acutatum at temperatures between 25 and 30 °C. For the Pedro Sato variety, temperatures between 20 and 30 °C with a 24 hour wetness period were found to be the most favorable conditions. The maximum lesion diameter was 3.3 cm for C. gloeosporioides and 3.2 cm for C. acutatum at temperatures below 25 °C. The highest incidence of the disease (100%) occurred 10 days after inoculation, at 25 and 30 º C and 6 hours of wetting. The lowest incubation period for both species was 7 days at temperatures between 20 and 30 °C and the lowest latency period of 8 days for C. gloeosporioides and 9 days for C. acutatum at temperatures between 25 and 30 °C. In conclusion, development conditions for Colletotrichum gloeosporioides and Colletotrichum acutatum were similar, although the range of conditions favorable for the Pedro Sato variety was wider than that of the Kumagai cultivar.

Este trabalho teve como principais objetivos avaliar os efeitos dos agentes bióticos (Saccharomyces cerevisiae, Bacillus thuringiensis, Lentinula edodes e Agaricus blazei), e abióticos (UV-C, irradiação gama, acibenzolar-S-metil, quitosana, ácidos acético e salicílico) na proteção de mamões contra C. gloeosporioides, bem como estudar os mecanismos bioquímicos de resistência ativados no tecido vegetal, em resposta ao tratamento com os agentes de maior eficiência, além de investigar os efeitos destes sobre o desenvolvimento in vitro do fungo. Para tanto, mamões cv. Golden foram inoculados com C. gloeosporioides através de injeção subcuticular de 15 µL da suspensão de esporos e, após 10 h, tratados com os diferentes agentes bióticos e abióticos. Para avaliar a possibilidade de indução de resistência pelos agentes, mamões foram também inoculados após 24, 48 e 72 h dos tratamentos. Os frutos foram armazenados a 25 ºC / 80 %UR por 7 dias e, avaliados diariamente quanto a incidência e severidade da podridão. Ao final do período de armazenamento, efetuou-se a avaliação dos parâmetros físico-químicos (cor de casca e de polpa, firmeza, sólidos solúveis, pH e acidez total). Quando de interesse, as atividades de peroxidase, β-1,3-glucanase e quitinase foram também investigadas. In vitro, avaliou-se o crescimento micelial, a germinação de conídios e a esporulação do fungo em resposta aos diferentes tratamentos. Os resultados mostraram que a irradiação gama (0,75 e 1 kGy) reduziu a incidência e a severidade da antracnose. Não houve efeito da UV-C no controle da podridão e todas as doses testadas causaram danos na casca dos frutos. O acibenzolar-S-metil reduziu em mais de 50 % a incidência e a severidade da podridão, além de induzir a maior atividade das enzimas peroxidase, quitinase e β-1,3-glucanase, e não alterar as características físico-químicas dos frutos. O ácido acético, a 2,5 µL L-1, reduziu a severidade e a incidência das lesões nos frutos. A quitosana (1, 2 e 4 %) reduziu significativamente a severidade da antracnose, e a 4 % foi também eficiente em reduzir a incidência da podridão. Concentrações acima de 0,25 % suprimiram a esporulação de C. gloeosporioides nas lesões. No entanto, os frutos tratados com 2 e 4 % de quitosana não amadureceram normalmente, permanecendo com a coloração da casca verde até o final do período de armazenamento. S. cerevisiae (20 mg mL-1) e B. thuringiensis (7,5 mg mL-1), aplicadas 24 h antes da inoculação do patógeno, reduziram a incidência da antracnose nos frutos, mas não o acúmulo de proteínas relacionadas à patogênese. Os cogumelos (A. blazei e L. edodes) e o ácido salicílico não foram eficientes em reduzir a incidência e a severidade da antracnose. In vitro, a irradiação gama, a UV-C, os ácidos acético e salicílico, a quitosana, S. cerevisiae e L. edodes inibiram o crescimento micelial do fungo. A germinação de conídios foi reduzida pelas irradiações gama e UV-C, pelos ácidos acético e salicílico e pela quitosana. Esses resultados demonstram a possibilidade dos agentes estudados serem utilizados no manejo da antracnose, bem como na redução da utilização ou da dosagem de fungicidas empregados no controle da doença.
This work had as main objectives evaluate the effect of biotic and abiotic agents (Saccharomyces cerevisiae, Bacillus thuringiensis, Lentinula edodes and Agaricus blazei), and abiotic (UV-C, gamma irradiation, acibenzolar-S-methyl, chitosan, acetic and salicylic acids) on the protection of papaya fruits against C. gloeosporioides, and study the biochemical mechanisms of resistance activated in the tissues in response to the treatment with the agents exhibiting better efficiency. The effects of the agents on the in vitro development of the fungus were also investigated. For this, papaya fruits cv. Golden were inoculated with C. gloeosporioides through subcuticular injection of 15 µL of the spore suspension and after 10 h treated with the different biotic and abiotic agents. To evaluate the possibility of resistance induction by the different agents, fruits were also inoculated 24, 48 and 72 h after treatments. The fruits were stored at 25 ºC / 80 %RH for 7 days and evaluated daily for the incidence and severity of the anthracnose. At the end of the storage period, the evaluation of the physical-chemical parameters (skin and flesh color, firmness, total soluble solids, pH and tritatable acidity) was carried out. The peroxidase, β- 1,3-glucanase and chitinase activities were also investigated when need. In vitro, mycelial growth, conidium germination and sporulation of the fungus in response to the different treatments were also evaluated. The results showed that the gamma irradiation (0.75 and 1 kGy) reduced the anthracnose incidence and severity. The UV-C did not have effect on the control of the rot and all the doses caused damages in the skin of the fruits. The acibenzolar-S-methyl reduced in more than 50 % anthracnose incidence and severity, and induced the highest activity of peroxidase, chitinase and β-1,3-glucanase, and did not modify the physical-chemical characteristics of the fruits. The acetic acid at 2.5 µL L-1 reduced rot severity and incidence. The chitosan (1, 2 and 4 %) significantly reduced the rot severity, and at 4 % was also efficient in reducing anthracnose incidence. Chitosan concentrations above 0.25 % suppressed the sporulation of C. gloeosporioides in the lesions. However, the fruits treated with chitosan at 2 and 4 % did not ripen normally, remaining with green skin until the end of the storage period. S. cerevisiae (20 mg mL-1) and B. thuringiensis (7.5 mg mL-1), applied 24 h before the pathogen inoculation, reduced anthracnose incidence, but did not change the activities of pathogenesis related proteins. The mushrooms (A. blazei and L. edodes) and the salicylic acid were not efficient in reducing the incidence and the severity of anthracnose. In vitro, gamma irradiation, UV-C, acetic and salicylic acids, chitosan, S. cerevisiae and L. edodes inhibited the mycelial growth. The conidium germination was reduced by gamma and UV-C irradiation, acetic and salicylic acids and chitosan. These results show that these agents can be utilized for anthracnose management, and on the reduction in the use or dosage of fungicides utilized on the anthracnose control.

Os objetivos do presente trabalho foram estudar a herança da resistência à antracnose foliar em milho, estimar os parâmetros genéticos e identificar marcadores moleculares ligados a genes de resistência a esta doença. Parâmetros genéticos foram estimados com base na análise de modelos de herança mista de seis gerações de quatro cruzamentos entre duas linhagens resistentes (DAS4 e DAS3) e duas linhagens suscetíveis (DAS6 e DAS22). O delineamento experimental foi o de blocos casualizados com parcelas subdivididas, com três repetições, sendo as parcelas constituídas pelos cruzamentos e as subparcelas, pelas gerações. As plantas foram inoculadas artificialmente e avaliadas em dois experimentos através de uma escala de notas de 1 a 6. Os testes de hipóteses para selecionar o modelo de herança genética e as estimativas dos parâmetros foram realizados pelo método da máxima verossimilhança. Os resultados da análise de modelos mistos indicaram que a resistência é controlada por um gene de efeito maior em todos os cruzamentos e experimentos avaliados e também por poligenes, em pelo menos um dos experimentos. A ação gênica é aditiva e dominante, com predominância de efeitos genéticos aditivos. O mapeamento de QRLs foi realizado utilizando 141 indivíduos F1RC1 do cruzamento (DAS6 x DAS4) x DAS6, com base na avaliação fenotípica das suas famílias, em dois experimentos. O delineamento experimental foi o látice 12 x 12, incluindo as famílias, genitores e híbrido, com 3 repetições. As plantas foram inoculadas artificialmente e avaliadas através de uma escala de notas de 1 a 6. O mapeamento de QRLs, utilizando marcadores microssatélites e AFLPs e análise de bulks segregantes para detecção de marcadores candidatos, foi realizado pela análise de regressão linear múltipla (RLM) e pelo mapeamento por intervalo composto (MIC). Ambas metodologias de análise identificaram pelo menos um QRL no cromossomo 10 em cada experimento. Também foram detectados QRLs nos cromossomos 2, 3 e 5, apenas pela RLM. Os QRLs identificados pela RLM explicaram 25,7%, 23,3% e 24,5% da variação fenotípica no experimento 1, no experimento 2 e na análise conjunta, respectivamente. Já os QRLs identificados pelo MIC explicaram 28,9%, 32,3% e 31,0% da variação fenotípica no experimento 1, no experimento 2 e na análise conjunta, respectivamente. A análise de bulks segregantes permitiu a detecção apenas dos QRLs de efeitos fenotípicos mais expressivos localizados no cromossomo 10. Na maioria dos QRLs detectados, os alelos de resistência provieram do genitor resistente. A identificação destes QRLs oferece uma significativa contribuição para o entendimento da resistência de milho à antracnose foliar, podendo levar à identificação de genes e elucidação dos mecanismos envolvidos na expressão da resistência.
The objectives of this work were to study the inheritance of resistance to anthracnose leaf blight, estimate the genetic parameters, and identify molecular markers associated with resistance genes to this disease. Genetic parameters were estimated based on the analysis of mixed inheritance models in six generations of four crosses between two resistant (DAS4 and DAS3) and two susceptible inbred lines (DAS6 and DAS22). The experimental design consisted of randomized blocks containing split-plots, with three replicates, where the plots were represented by crosses and the subplots were the generations. The plants were inoculated artificially and evaluated in two experiments by means of a rating scale from 1 to 6. The hypothesis testing to select the genetic inheritance model and parameter estimates were obtained by the maximum likelihood method. The results from the mixed models analysis indicated that resistance is controlled by a major gene in all crosses and experiments evaluated, and also by polygenes in at least one experiment. The genetic action is additive and dominant, with predominance of additive genetic effects. QRL mapping was performed using 141 F1RC1 individuals from the (DAS6 × DAS4) × DAS6 cross, based on the phenotypic evaluation of their families, in two experiments. The experimental design was a 12 × 12 lattice which included the families, parents, and the hybrid, with 3 replicates. The plants were inoculated artificially and evaluated by means of a rating scale from 1 to 6. QRL mapping, using microsatellites and AFLPs markers, and bulked segregant analysis to detect candidate markers was performed by multiple linear regression analysis (MLR) and by composite interval mapping (CIM). Both methodologies of analysis identified at least one QRL in chromosome 10 in each experiment. QRLs were also detected, by MLR only, in chromosomes 2, 3 and 5. The QRLs identified by MLR explained 25.7%, 23.3%, and 24.5% of the phenotypic variation in the experiment 1, in the experiment 2 and in the joint analysis, respectively. The QRLs identified by CIM, however, explained 28.9%, 32.3%, and 31.0% of the phenotypic variation in the experiment 1, in the experiment 2 and in the joint analysis, respectively. The bulked segregant analysis only allowed the detection of QRLs that showed the more expressive phenotypic effects located in chromosome 10. In most detected QRLs, the resistance alleles came from the resistant parent. The identification of these QRLs offers a significant contribution to an understanding of resistance to anthracnose leaf blight in maize, and could lead to the identification of genes and to an elucidation of the mechanisms involved in the expression of resistance.

O Brasil é considerado o maior produtor de citros e o maior exportador de suco de laranja. Doenças de pós-colheita representam uma grande perda na citricultura, sendo que para muitos frutos a serem exportados, existe uma exigência para que os mesmos estejam isentos de resíduos químicos. Em relação a alguns patógenos de importância em pós-colheita podemos destacar, Guignardia citricarpa (Mancha-pretados- citros), Penicillium digitatum (Bolor-verde) e Colletotrichum gloeosporioides (Antracnose). Dada a importância econômica que representa esse complexo de doenças dos frutos cítricos, tanto em termos de comprometimento da qualidade dos frutos, limitações às exportações e dificuldade de controle, a busca de alternativas adicionais que possam viabilizar a capacidade produtiva dos produtores e garantir a obtenção de frutos com excelentes padrões de qualidade torna-se imprescindível. Nesse contexto, pode-se inserir o emprego de medidas de controle alternativas que não englobam o controle químico clássico. Sob esse ponto de vista inclui-se o controle através do uso de agentes bióticos e abióticos e a indução de resistência em plantas. Portanto neste trabalho, foi estudada a viabilidade do controle de doenças pós-colheita em citros, envolvendo a ação direta sobre os patógenos através do uso dos extratos etanólicos de albedo (mesocarpo) de laranja doce (Citrus sinensis var. Valência) e flavedo (exocarpo ou epicarpo) de limão-Tahiti (Citrus aurantifolia Swing var. Tahiti). O extrato do albedo apresentou efeito antifúngico sobre G. citricarpa e o flavedo do limão “Tahiti” sobre C. gloeosporioides, além de se demonstrar a existência de compostos voláteis com efeito tóxico. A segunda parte envolveu o controle e a indução de resistência em frutos, através do uso dos agentes bióticos Lentinula edodes e Agaricus blazei e do agente abiótico ácido jasmônico. Foi possível observar que o extrato aquoso do albedo (C. sinensis), flavedo (C. aurantifolia), L. edodes e A. blazei diminuíram o aparecimento de novas lesões causadas por G. citricarpa, porém não apresentaram efeitos sobre P. digitatum e C. gloeosporioides em frutos de C. sinensis var. Valência quando tratados em pós-colheita. Dessa maneira, no presente trabalho demonstrou-se a viabilidade de um possível controle alternativo de doenças pós-colheita em citros, buscando-se novos agentes que atuem como indutores de resistência ou de controle direto sobre os fitopatógenos.
Brazil is considered the biggest citrus producer and the biggest orange juice exporter. Post-harvest diseases represent a great loss in the citriculture, and for many fruits to be exported they should be free of chemical residues. In relation to some pathogens of importance in post-harvest it can be mentioned Guignardia citricarpa (black-spot-ofcitrus), Penicillium digitatum (green-mold) and Colletotrichum gloeosporioides (anthracnose). Because of the economical importance that represents this disease complex in citric fruits, in terms of compromising fruit quality, limitations to the exports and control difficulties, the search for alternative control measures that can make possible improve the producing capacity of the producers and the obtaining of fruits with excellent quality are indispensable. Thus, in this context it can be included measures of alternative control that do not include the chemical control. Under this point of view, control include the use of biotic and abiotic agents and the resistance induction in plants. Therefore, it was studied the viability of the control of post-harvest diseases in citrus, involving the direct action on the patogens by using ethanolic extracts of albedo (mesocarp) of sweet orange (Citrus sinensis var. Valência) and flavedo (exocarp or epicarp) of lemon-Tahiti (Citrus aurantifolia Swing var. Tahiti). The results showed that the extract of the albedo exhibited antifungal activity on G. citricarpa and the flavedo of the "Tahiti" lemon on C. gloeosporioides, and it was also demonstrated the existence of volatile compounds toxic to the fungus. The second part involved the control and resistance induction in the fruits, by using the biotic agents Lentinula edodes and Agaricus blazei and the abiotic agent jasmonic acid. It was possible to observe that the aqueous extracts from the albedo (C. sinensis), flavedo (C. aurantifolia), L. edodes and A. blazei reduced the formation of new lesions caused by G. citricarpa, however they did not exhibit effects on P. digitatum and C. gloeosporioides in fruits of C. sinensis var. Valência when treated in post-harvest. Thus, in the present work it was demonstrated the viability of possible alternative control measures of diseases in post-harvest of citrus, indicating the need of looking for new agents to act as resistance inducers or agents to directly control on the phytopathogens.

Industry demand for local sources of grain for animal feed has increased sorghum production in the mid-Atlantic region of the U.S. Sorghum anthracnose (causal agent Colletotrichum sublineola) and the grain mold complex, which includes mycotoxin-producing Fusarium spp., limit the yield and quality of grain sorghum in humid climates worldwide. A majority of U.S. grain sorghum production is in arid regions, and management strategies have not been developed for the mid-Atlantic U.S. where warm, wet conditions favor disease. The specific objectives of this research were to: (1) determine the effectiveness of fungicides and their application timing for the management of sorghum foliar anthracnose, (2) compare five grain sorghum hybrids for their susceptibility to foliar anthracnose, grain mold and mycotoxin contamination under field conditions, (3) integrate host resistance and fungicide application to manage anthracnose and grain mold, and (4) identify Fusarium spp. associated with grain mold and mycotoxin contamination of sorghum in the mid-Atlantic U.S. For Objective 1, it was determined that a single application of pyraclostrobin-containing fungicide no later than flowering reduced anthrancose, protected yield and maximized farm income. Objective 2 focused on sorghum hybrid selection as a disease management tactic, and it was determined that hybrids with high yield potential and moderate disease resistance should be selected for mid-Atlantic sorghum production in order to maximize grain yield and quality while minimizing the need for fungicide inputs. Objective 3 focused on integrated management and demonstrated that under moderate disease pressure, a high-yielding susceptible hybrid required a single application of pyraclostrobin-based fungicide to minimize fungal diseases and maintain acceptable yields, whereas under high disease pressure it was necessary to integrate hybrid resistance and judicous applications of fungicides. The aim of Objective 4 was to characterize potential causal agents of mycotoxin contamination in mid-Atlantic sorghum, and thirteen phylogenetically distinct Fusarium species (F. lacertarum, F. graminearum. F. armeniacum, F. proliferatum, F. fujikuroi, F. verticillioides, F. thapsinum and several in Fusarium incarnatum-equiseti species complex) were found to be associated with grain mold and fumonisin and/or deoxynivalenol contamination of sorghum grain. This work has provided insights into the impacts of fungal diseases on grain sorghum yield and quality in the mid-Atlantic and has aided in development of best management practices for the region.
Doctor of Philosophy

Thesis (Ph. D.)--Ohio State University, 2005.
Title from first page of PDF file. Document formatted into pages; contains xiii, 118 p.; also includes graphics Includes bibliographical references (p. 114-118). Available online via OhioLINK's ETD Center

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Coordenação de Aperfeiçoamento de Pessoal de Nível Superior
The concern with the environment and the human health has stimulated the search for methods that reduce the use of fungicides in the control of phytopathogens. Considering many positive results already reached by using induced resistance, and the great demand for new elicitors of resistance to pathogens, this work had the objective to evaluate the fungitoxic effect in vitro of the aqueous extract (EA) of Pycnoporus sanguineus basidiocarp, against Colletotrichum lindemuthianum and evaluate under greenhouse conditions, the control of anthracnose in beans, as well as the induction of resistance by the determination of peroxidases activity. For the assays in vitro were used EAs, in the concentrations of 1, 5, 10, 15 and 20 %, autoiclavated or not. And evaluating the micelial growth and spores germination of C. lindemuthianum. For the in vivo assays, EAs not autoclavated at 5, 10 and 20% were sprayed in 7th´s leaves of beans plants three days before the inoculation of the pathogen (5x104 conidia/mL) that was occurred in 7th´s and 8th´s leaves. Water and fungicide (azoxystrobin 0,6 g. p. c./L) were used as control tratments. For peroxidase, 7th´s and 8th´s leaves were sampled at the moment of treatments and after three, six, nine and twelve days. The results showed direct antimicrobial activity of the EAs of P. sanguineus, with inhibition of up to 96% of the germination in vitro of the spores of C. lindemuthianum, independent of the autoclavation of the extracts. The EAs stimulated in up to 34 % the micelial growth. In the plants, eA 20% controlled the anthracnose with reduction of 70% severity in 7th´s leaf (treated and inoculated), while for the fungicide the reduction was of 73%. For 8 th´s leaf (only inoculated), the EAs at 5, 10 and 20% reduced the severity in 58, 64 and 68%, respectively, while for the fungicide the reduction was of 74%. This reduction in severity can be associated with the activity of peroxidase, which was presented high, in both 7th´s and 8th´s leaves in the moment of the inoculation, it means, three days after the treatment. These results indicate the P. sanguineus extracts potential for control of anthracnose in beans plants, that can occurr by direct antimicrobial activity and/or local and systemic induction.
A preocupação com o ambiente e a saúde humana tem incentivado a busca por métodos que reduzam o uso de fungicidas no controle de fitopatógenos. Considerando-se os vários resultados positivos já alcançados pela utilização da resistência induzida, bem como a grande demanda de novos eliciadores de resistência a fitopatógenos, este trabalho teve como objetivos avaliar o efeito fungitóxico in vitro do extrato aquoso (EA) de basidiocarpos de Pycnoporus sanguineus, sobre Colletotrichum lindemuthianum; e avaliar em de casa de vegetação, o controle da antracnose em feijoeiro por esses extratos, bem como a indução de resistência pela determinação da atividade de peroxidase. Para os ensaios in vitro foram utilizados EAs, nas concentrações de 1, 5, 10, 15 e 20 %, autoclavados ou esterilizados por filtração, avaliando-se o crescimento micelial e a germinação de esporos de C. lindemuthianum. Para os ensaios in vivo, EAs não autoclavados a 5, 10 e 20% foram aplicados nas 7ªs folhas de plantas de feijoeiro três dias antes da inoculação do patógeno (5x104 conídios /mL) que ocorreu nas 7ªs e 8ªs folhas. Água e fungicida azoxystrobin (0,6 g. p. c. / L) foram utilizados como testemunhas. Para a dosagem de peroxidase foram amostradas as 7ªs e 8ªs folhas no momento dos tratamentos e após três, seis, nove e doze dias. Os resultados indicaram atividade antimicrobiana direta dos EAs de P. sanguineus, com inibição de até 96% da germinação in vitro dos conídios de C. lindemuthianum, independente da autoclavagem ou não dos extratos. Os EAs estimularam em até 34 % o crescimento micelial. Nas plantas, apenas o EA a 20% controlou a antracnose, com redução de 70% na severidade na 7ª folha (tratada e inoculada), enquanto que para o fungicida a redução foi de 73%. Para a 8ª folha (apenas inoculada), os EAs a 5, 10 e 20% reduziram a severidade em 58, 64 e 68%, respectivamente, enquanto que para o fungicida a redução foi de 74%. Essa redução na severidade pode estar associada com a atividade de peroxidase, a qual se apresentava alta, tanto na 7ª quanto na 8ª folha, no momento da inoculação, ou seja, três dias após o tratamento. Estes resultados indicam o potencial de controle da antracnose em feijoeiro pelo extrato de P. sanguineus, que pode ocorrer por atividade antimicrobiana direta e/ou indução de resistência local e sistêmica.

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The increase in diseases in maize crops, with emphasis on the region of Campos Gerais in Paraná, has been relevant due to the commitment quantitative and qualitative of the grain. Because of this, the purposes of this work were: to evaluate the incidence of leaf diseases and their impact on productivity; to evaluate the effect of different fungicides to control pathogens that cause damaged grains; to relate the incidence of the main leaf diseases in corn crop and its interference with the production components; to evaluate the performance of corn inoculated with Colletotrichum graminicola to treatments with different mixtures of fungicides of strobilurins and triazoles groups; to verify the effect of various essential oils and fungicides on mycelial growth of C. graminicola; to develop a diagrammatic scale to assess the severity of anthracnose leaf blight in maize crop and to verify the best concentration of essential oils in the control of leaf anthracnose and to estimate possible effects of phytotoxicity of the essential oils on the corn crop. For that were installed 14 experimental fields with several hybrids of corn during harvest 2011/12, one in the 2012/13 crop, and one in the 2013/14 crop; in laboratory were installed a 2012 experiment, and another between 2014/15; It was developed a diagrammatic scale in 2015 and carried out an experiment in a greenhouse in 2016. The results brought information such as: between the late material and early ones, the results were specific to each location where Puccinia sorghi, Phaeosphaeria maydis, Exserohilum turcicum had greater severity of disease in 6 of the 9 experiments, and was also the most sensitive in the experiment installed in late materials; there was difference of fungicide control depending on the studied pathogen; regarding to yields higher than 10 tons, the components production that directly interfered were: length and diameter of the ear, number of kernels per row and weight of 1000 grains, on the other hand the most affected yield components in those hybrid with productions around 8.5 tons were: diameter of the ear, number of kernels per row and the 1000 grains; None of the Colletotrichum graminicola application times obtained results regarding the incidence of leaf anthracnose. The treatments with fungicides scored lower percentage of damaged kernels; regarding the use of essential oils, clove, oregano and cinnamon there were inhibitory effect on mycelial growth of C.graminicola at all concentrations tested, citronella, lime and orange, showed fungistatic effects at concentrations of 5 and 10%, while lemongrass and thyme showed better results in concentrations of 15 and 20%; the scale was presented as an assertive tool to quantify the severity of anthracnose leaf blight; it was observed that independent of concentration, all essential oils showed phytotoxicity on corn plants, none of the essential oils in the used concentrations totally controlled C. graminicola.
O aumento de doenças na cultura do milho (Zea mays L.), na Região dos Campos Gerais do Paraná, tem sido relevante em virtude do comprometimento quantitativo e qualitativo dos grãos. Em função disso, foram objetivos deste trabalho: verificar o efeito de diferentes óleos essenciais e fungicidas sobre o crescimento micelial de Colletotrichum graminicola; verificar a melhor concentração dos óleos essenciais no controle da antracnose da folha e estimar possíveis efeitos de fitotoxicidade dos óleos essenciais; elaborar uma escala diagramática para estimar a severidade da antracnose foliar na cultura do milho; avaliar o efeito de diferentes fungicidas no controle de patógenos causadores de grãos ardidos, avaliar a ocorrência de doenças foliares e seu impacto na produtividade; relacionar a ocorrência das principais doenças foliares na cultura do milho e a sua interferência sobre os componentes de produção; avaliar o desempenho da cultura do milho inoculada com C. graminicola a tratamentos com diferentes misturas de fungicidas dos grupos de estrobilurinas e triazóis; sobre a cultura do milho. Para isso em laboratório foram instalados experimentos; também foi elaborada uma escala diagramática em 2015 e realizado um experimento em casa de vegetação em 2016; além da instalação de 14 campos experimentais com diversos híbridos de milho, durante as safras agrícolas 2011/2012, 2012/2013 e 2013/2014. Nenhuma das épocas de aplicação de Colletotrichum graminicola obtiveram resultados quanto à ocorrência da antracnose foliar. Os tratamentos que receberam fungicidas obtiveram as menores porcentagens de grãos ardidos; quanto ao uso de óleos essenciais, cravo da índia, canela e orégano tiveram efeito inibitório no crescimento micelial de C. graminicola em todas as concentrações testadas, citronela, laranja e lima, apresentaram efeito fungistático nas concentrações de 5 e 10%, enquanto erva cidreira e tomilho apresentaram melhores resultados nas concentrações de 15 e 20%; a escala apresentou-se como uma ferramenta assertiva para a quantificação da severidade da antracnose foliar. Constatou-se que independente da concentração, todos osóleos essenciais apresentaram fitotoxicidade sobre as plantas de milho, nenhum dos óleos essenciais nas concentrações utilizadas controlaram totalmente C. graminicola. Entre os materiais tardios e os precoces, os resultados foram específicos para cada localidade onde a ferrugem comum (Puccinia sorghi), mancha branca (Phaeosphaeria maydis) e mancha de turcicum (Exserohilum turcicum) tiveram maior severidade da doença em 6 dos 9 experimentos, e também foi o mais suscetível no experimento instalado com materiais tardios. Houve diferença de controle fungicida dependendo do patógeno estudado; com relação às produtividades maiores que 10 toneladas, os componentes de produção que interferiram diretamente foram: comprimento e diâmetro da espiga, número de grãos por fileira e massa de 1000 grãos, em contrapartida os componentes de produção mais afetados nos híbridos com produções em torno de 8,5 toneladas foram: diâmetro da espiga, número de grãos por fileira e massa de 1000 grãos.

碩士
國立臺灣大學
植物醫學碩士學位學程
107
Strawberry is an important economic crop in Taiwan, the total planting area has reached 550 hectares, and the annual output value is nearly 1 billion NTD. However, due to the high temperature and humidity in Taiwan, strawberry cannot be planted continuously. Instead, strawberry farmers replant with strawberry runners annually. Anthracnose is one of the most severe diseases in the strawberry nursery industry, leading to a great loss on strawberry seedling. The lack of healthy seedlings has become a huge obstacle in the strawberry industry in Taiwan. Currently, farmers mainly rely on fungicides to prevent anthracnose, as a consequence, the massive use of fungicides has given rise to the anthracnose drug resistance. As a result, it is an emerging need of new methods for controlling this disease, in order to reduce the use of fungicides. Numerous studies have shown different light wavelengths capable of suppressing both plant pathogen growth and plant diseases. Here, in this study, we illuminated different light wavelengths (white light, red light, 2/3red+1/3blue, 1/3red+2/3blue, blue light) at 200 PPFD on Colletotrichum siamense on PDA (Potato dextrose agar) , inoculated strawberry leaves and seedlings, and co-treatment with difenoconazole, to obtain the optimal wavelength for controlling anthracnose disease, to promote strawberry seedlings growth, and to implement with difenoconazole, respectively. Our results showed that, in III compared to white light, 1/3red+2/3blue could inhibit 14% C. siamese mycelial growth, 5% of the sporulation and 11% of spore germination. In addition to C.siamense growth inhibition, 1/3red+2/3blue light could also augment the strawberry defense gene expressions, such as Chitinase-2 gene. The 1/3red+2/3blue light was also found to increase leave number, runner number, and overall fresh weight of seedlings than white light does. Furthermore, all light wavelengths would not impair difenoconazole’s effectiveness or enhance phytotoxicity. In conclusion, our study indicates that 1/3red+2/3blue light shows the characteristics of the growth inhibition of C. siamense, defense genes upregulation, and the growth of strawberry. Therefore, we recommend 1/3red+2/3blue light condition during the nursery period can be applied for suppressing anthracnose disease and promoting strawberry seedling growth.

The aim of the project was to develop a strategy towards anthracnose resistance in lupin using molecular techniques. Colletotrichum species are considered to be major plant pathogens of cereals and legumes around the world, causing significant crop losses. Colletotrichum acutatum causes anthracnose disease on lupin. Sweet white lupin (Lupinus albus) is a high protein grain crop that could alleviate protein shortage in South Africa, since it has the highest protein levels (34-45%) compared to Lupinus angustifolius. In an effort to combat the lupin anthracnose threat to the South African lupin industry, which has an annual turnover of approximately 60 million rands, a project was embarked upon to introduce defense genes into a white lupin and a narrow leaf lupin cultivar. Bean polygalacturonase inhibiting protein (PvPGIP), either extracted from bean or from transgenic tomato expressing the bean pgip1 gene (Pvpgip1), inhibited the C. acutatum polygalacturonase (PG) activity (isolate SHK 788) only by 18-25%, compared to apple PGIP (MdPGIP) that inhibited the C. acutatum PG activity by 70%. These results led to the Mdpgip1 gene, rather than the Pvpgip1 gene, being chosen for genetic engineering of lupin towards anthracnose resistance. However, since plants express more than one PGIP, the protein in the extract prepared from the fruit of apple cv. Granny Smith, could be encoded by any one of at least two closely related copies of pgip genes found in apple. Screening of eight putative first generation Mdpgip1 transformed tobacco plants using PCR, showed that all eight plants contained the Mdpgip1 gene. Inhibition studies, using the C. acutatum PGs, were performed which identified Mdpgip1 transgenic tobacco plant #8 as being the highest expresser of the MdPGIP1, since the MdPGIP1 extract from this plant exhibited the highest level of C. acutatum PG inhibition. The PGIP extract from the non-transgenic tobacco plant, as well as heat denatured MdPGIP1 extracts from the Mdpgip1 transgenic tobacco plants, resulted in no inhibition of C. acutatum PG activity. Mdpgip1 transgenic tobacco plant #8 was chosen for the purification of MdPGIP1. The protein was purified to apparent homogeneity using anion and cation exchange chromatography. N-terminal sequencing deduced the first 15 amino acids, which aligned 100% to the sequence of a pgip gene (called Mdpgip) from Golden Delicious apples (Genbank: accession no. MDU 77041), confirming isolation of MdPGIP1. The protein had a molecular mass of approximately 46kDa as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and an isoelectric point of 8.0. Purified MdPGIP1 inhibited the PGs produced by C. acutatum and the PGs produced by two apple pathogens, B. obtusa and D. ambigua. Results indicated that much less MdPGIP1 is required for effective inhibition of the B. obtusa and D. ambigua PGs, compared to the C. acutatum PGs. However, at higher MdPGIP1 concentrations all three fungal PGs were inhibited equally well. A purified endo-PG from Aspergillus niger was not inhibited by MdPGIP1. This constitutes the first report on the inhibitory activity of MdPGIP1 towards the PGs from C. acutatum, and the two apple pathogens B. obtusa and D. ambigua. As part of a multigene approach to the production of anthracnose resistant lupin, the use of a yeast exo--1,3-glucanase (EXG1) as an antifungal agent towards C. acutatum was investigated. The exo--1,3-glucanase (exg1) gene had been isolated from Saccharomyces cerevisiae. Yeast cultures transformed with the exg1 gene, as well as untransformed yeast cultures, were obtained from the Institute for Wine Biotechnology, South Africa. Fungal spore suspensions, from isolate SHK 788, were prepared and used in inhibition studies with spore concentrations ranging from 2.5.103 spores to 80.103 spores per flask. Inhibition of C. acutatum mycelial growth ranged from 41%, at a fungal spore concentration of 2.5.103 spores, to 20%, at a fungal spore concentration of 80.103 spores. Ammonium sulphate concentrated yeast extracts containing the glucanase enzyme did not result in increased inhibition of C. acutatum mycelial growth. As an added control, an inhibition study using Botrytis cinerea spores yielded similar results to those obtained for the C. acutatum inhibition studies. An inhibition of at least 50% for all spore concentrations was set as the criterium to decide that the exg1 gene is potent enough for genetic engineering of disease resistance. This extent of inhibition was not obtained and the use of the exg1 gene for protection of lupin against C. acutatum was therefore not considered a worthwhile commercial option. The defense gene plant transformation vectors prepared for lupin transformation, pCAMBIA 3300-virG, pCAMBIA 3301-virG, pCAMBIA 3300-virG-applePGIP and pCAMBIA 0390:applePGIP were successfully transformed into the A. tumefaciens strains LBA 4404 and AGL1. Lupin transformation was performed by the transformation group at CSIR Bio/Chemtek using A. tumefaciens-mediated transformation of shoot apical meristems. This group showed that the inclusion of the supervirulence virG gene enhanced the levels of transient GUS expression in L. angustifolius by more than two fold. However, transformation efficiency was low, and regeneration of the lupin plant proved to be even more difficult. To overcome the difficulties with plant tissue culture-based transformation systems, an A. tumefaciens seed vacuum infiltration transformation method was utilised. Extracts obtained from Mdpgip1 transgenic tobacco plants produced at CSIR Bio/Chemtek (pCAMBIA 3300-virG-applePGIP as well as pCAMBIA 3300-virG/pCAMBIA 0390:applePGIP transformants) inhibited the C. acutatum PGs. The Mdpgip1 gene thus codes for an active protein in the transgenic tobacco plants, and the defense gene constructs prepared for lupin transformation are functional in planta. The shpx6a peroxidase gene was isolated from Stylosanthes humulis, as the second defense gene to be used in the strategy towards anthracnose resistance in lupin, and substitute for the yeast exg1 gene. Sequencing data confirmed the successful isolation of the shpx6a peroxidase gene, which was subsequently cloned into pCAMBIA 0390:applePGIP upstream from the NOS terminator to produce pCAMBIA 0390:applePGIP:peroxidase. Seeing that the constitutive CaMV 35S promoter was going to be used upstream from the selection gene (bar), the Mdpgip1 gene and the additional shpx6a peroxidase gene, there was a concern that one type of gene silencing could occur. Use of one promoter can block expression of another gene being expressed from the same promoter on account of methylation of the promoter DNA. A 4.2kb fragment containing the inducible class-III chitinase (if3) promoter was isolated from L. albus, using the GenomeWalkerTM kit, for use in the pCAMBIA 0390:applePGIP:peroxidase defense gene construct, i.e. upstream from the shpx6a peroxidase gene. The 4.2kb fragment was successfully cloned into the pGEM-T Easy vector and sequenced. The sequence was compared to known sequences in the Genbank database but exhibited no significant homology. Using bioinformatic tools, five possible eukaryotic promoter-containing sites, including the TATA boxes, were identified within the isolated 4.2kb fragment. Deletion studies were performed in order to test for the minimal sequence needed for retaining of promoter activity. The 1.818kb, 1.512kb and 1.138kb if3 promoter-containing fragments were each cloned separately into the pDM327 vector upstream from the bar-gus fusion gene to produce pDM327:Prom1.8, pDM327:Prom1.5 and pDM327:Prom1.1 and used in the BiolisticTM transformation of plant tissue. BiolisticTM transformation of Ornithogalum and bean callus tissue, as well as maize and lupin immature embryos all demonstrated that the if3 DNA fragment isolated from L. albus contains promoter activity, indicated by the efficient stimulation of the expression of the gus reporter gene. Based on these results a provisional patent was filed [Application number: 2003/2405, and entitled “Plant Promoter”]. Bioinformatic analysis indicated the presence of various putative cis-acting regulatory elements, that could be important in controlling the expression of the 1.8kb if3 promoter-containing fragment. A single putative MBS regulatory cis-acting element was present in the 1.13kb promoter-containing fragment. It acts as a Myb transcription factor binding site that regulates transcription of several plant genes in response to various environmental factors, including elicitors and wounding. Several CAAT boxes were also identified within the 1.81kb promoter-containing fragment which play an important role in the determination of promoter efficiency. Most of the putative fungal elicitor activated (Box-W1 and ELI-box3) and wound-inducible [WUN-motif and ERE (ethylene responsive element)] cis-acting elements were present in the 1.13kb promoter-containing fragment. This supports the hypothesis that all regulatory elements needed for the activation of the if3 gene promoter are located within the first 1.13kb fragment upstream from the initiation codon of the if3 gene. The final evaluation of the main hypothesis that the combinatorial approach, by using two defense genes, will be much more effective than one gene or natural resistance in the suppression of anthracnose in lupin will need to be evaluated once successful transformation and regeneration of lupin has been obtained.
Prof. Ian Dubery

碩士
國立臺灣大學
植物病理與微生物學研究所
104
Water caltrops (Trapa taiwanensis Nakai) is a common annual aquatic crop in Guan Tian, Tainan. It adapts well in wetland like swamp or paddy areas. The most common diseases affecting water caltrops are anthracnose and sclerotium rot. The anthracnose disease of water caltrops is caused by Colletotrichum gloeosporioides Penzig. On water caltrops, the anthracnose can cause black spot symptoms that can coalesce into big lesions on leaves, petiole and fruits. This study is aimed to confirm the pathogenicity of anthracnose fungi on water caltrops and to investigate the epidemiology of this disease in Guan Tian area. Some herbal material extracts and antagonistic microorganisms are also evaluated for their potential for controlling this disease. Through the pathogenicity tools and molecular identification, we accomplish the Koch’s postulates of this disease. Culturing this pathogen at different temperatures showed that the pathogen grows best at 25℃. The epidemiological study from 2014 to 2015 showed that the disease occurred after a long period of raining summer season and may continue to the winter time. The correlation coefficient between last-monthly rainfall and disease severity in 2014 and 2015 are 0.9371 and 0.9297, respectively. If we combine the favorable rainfall, temperature and wind speed in together, the correlation coefficient between favorable hours and the disease severity in 2014 and 2015 can be as high as 0.9485 and 0.9271, respectively. Results indicated that the disease severity is positively correlated with favorable temperature, high wind speed and high rainfall. The non-pesticide control study showed that antagonistic microorganism had better effectiveness than herbal extracts. Water extracts of all herbal material showed low control rate on PDA. Although ethanol extracts of all nine herbal material showed significant inhibition rate in spore germination and mycelial growth tests, they didn’t have good effectiveness in the pot plant test. Both Bacillus subtilis and Streptomyces sp. YU01 has about 45 % control rate in pot plant test, when applied 3 days before inoculation with pathogen. Whereas Trichoderma asperillum expressed only about 30% control rate in pot plant test. Besides non-pesticide materials, difenoconazole and carbendazim showed best satisfactory to control this disease. We also found that anthracnose isolates from mango, strawberry and pomelo, cannot cause anthracnose disease on the water caltops.

碩士
國立臺灣大學
生物產業機電工程學研究所
101
The growth and appearance of plant are affected and destroyed when infected by pathogens. Plant diseases not only decrease the production rate, but also cause economic losses. The thesis aims to evaluate the potential of strawberry foliar Anthracnose disease and Bok Choy black spot disease identification by using hyperspectral images. Hyperspectral imaging is a non-destructive measurement technique that combines digital imaging and spectroscopy in visible and near infrared wavelength regions. The technique can provide physical and chemical information of leaves simultaneously, such as the change of chlorophyll and water in leaves. In this study, two feature selection algorithms, stepwise discriminant analysis (SDA) and elastic net (EN), were applied to define the significant wavelengths for discriminating plant diseases, and then employed the defined to establish a linear discriminant analysis classifier. Moreover, pseudo color image were also used to represent the infected locations and regions on the leaf. In this approach, the reflectivity of the selected wavelengths, and the first and second derivative of the reflectivity were employed as the features. The recognition performance of the features was compared. The accuracy of classifying healthy Bok Choy leaves and infected leaves is 98.3%, and the classification model was the built with 12 wavelengths selected by SDA. To discriminate healthy, incubation and symptomatic of Bok Choy leaves, the accuracy is 77.4% with 4 selected wavelengths. For strawberry, the accuracy of the classifying healthy and infected leaves reaches to 98.5%, and the classification model was established with 2 wavelengths selected by SDA according to the first derivative. For classifying three different Anthracnose infection status (healthy, incubation and symptomatic), the accuracy is 89.3%, and 16 wavelengths defined by SDA were applied to build the model. The experimental results imply that the use of machine learning algorithm is able to discriminate plant disease by few and significant wavelengths. Furthermore, the proposed method and procedure can be applied to establish an automatic, non-destructive plant disease detection system using multispectral imaging for monitoring plant disease.

碩士
國立中興大學
植物病理學系所
103
Mango is one of the most important fruit crops and is the highest value of exported fresh fruits in Taiwan. Colletotrichum gloeosporioides causes mango anthracnose and is one of the major pathogens in mango. It can infect mango leaves, flowers, branches and fruit, and causes considerable fruit disqualified during postharvest period. Currently, the disease management strategies majorly rely on fruit bagging and chemical fungicides application. However, fungicide application can result in fungicide resistance and excessive chemical residues. The purpose of this study is to screen and apply microbe- and plant-derived materials to control mango anthracnose. Since mango fruit is seasonally available, a stable inoculation platform for using mango leaf as inoculation material was established in the beginning in my thesis study. Mango (cv. Irwin) leaf with age within 8-days is highly susceptible to the infection of C. gloeosporioides TYC-2 and the infection is stable and repeatable. In addition, anthracnose lesions appeared earlier on the abaxial than the adaxial surface of a leaf, which is not related to the ability of germination and appressorial formation of the pathogen on both surfaces. Among 17 tested plants, ethanol extracts from 4 different plants showed nearly 50% of inhibition on mycelial growth of TYC-2. Plant A ethanol-extracts showed complete inhibition on TYC-2 spore germination at 0.08 mg/ml in vitro. The anthracnose lesion was not appeared when mango leaf and fruit were co-inoculated with TYC-2 and plant A ethanol-extracts (0.08 mg/ml). Total 45 yeast isolates and 83 bacterial isolates were isolated from the soil, leaves or flowers of mango and showed inhibitory ability to the mycelial growth of TYC-2. Among all yeast isolates, isolate 3H-4 showed strong competition against the growth of C. gloeosporioides in PDA medium. Among all bacterial isolates, strain A and B has significant inhibitory effect to 15 Colletotrichum isolates which cause the anthracnose of mango, Chinese cabbage and chili pepper. Strain A and B were identified as Bacillus amyloliquefaciens based on the comparison of 16S rDNA sequence and Biolog analysis. The 100-fold dilution of 4-day-old culture liquid of strain A or B cultured in culture medium (strain A-SSM or strain B-SSM) could completely inhibit anthracnose lesion production on detached leaves and fruit. Moreover, it showed 80% lesion area reduction when treated with 600-fold dilution of strain B-SSM on detached leaf. The bacteria population of strain B -SSM could increase slightly from 1.48 × 109cfu/ml to 2.6 × 109cfu/ml after stored for 20 weeks under room temperature. In addition, the antagonistic activity of strain B -SSM was resistant to heat treatment, 100℃ for 20 minutes. It indicates that strain B -SSM had great stability during short-time storage assay. To improve the antagonistic activity of strain B, various plant oils were added into SSM to culture this bacterial strain. The results showed that 0.5% (v/v) plant oil A amended SSM could increase the antagonistic activity of strain B by increasing the lesion area reduction from 43% to 100% when 200-fold diluted bacterial culture was applied. The active ingredient of strain B -SSM remained in the culture filtrate but not the culture pellets after the bioactivity assay on the detached leaf. Culture filtrate of strain B -SSM could inhibit spore germination and cause abnormal swelling of germ tubes of TYC-2 in vitro and in planta under the examination of light microscopy as well as scanning electron microscopy. There were many vacuole-like structures formed in hyphal 32 h after treated with strain B -SSM, and no necrosis lesion was observed on fruit 72 h after treatment. Thin layer chromatographic (TLC) analysis revealed that two regions (Rf 0.045 and 0.38) with antifungal activity were identified in strain B -SSM and strain B -SSM with 0.5% plant oil A. The bioactive components were recovered from the TLC plate and analyzed by high-performance liquid chromatography (HPLC). The data revealed that the bioactive components contain iturin A. Based on the results presented in this study, the ethanol-extracts of plant A and bacterial strain B has great potential for further development of biological control agents in field applications to control mango anthracnose.

碩士
國立臺灣大學
植物病理與微生物學研究所
102
Plants are sessile organisms and unable to avoid pathogen attack by moving ability. Plants may protect themselves by generating biochemical products with antimicrobial activity. LsGRP1 is a glycine-rich defense-related protein of Lilium cv. Stargazer, with a C-terminal portion (LsGRP1C) comprised of 38 amino acids rich in cysteine residue. Colletotrichum higginsianum causing crucifer anthracnose is a hemibiotrophic fungal pathogen. In vitro assays with the synthetic peptide of LsGRP1C showed inhibition of spore germination, appressorium formation and mycelial growth of C. higginsianum. Immunofluorescence staining showed that LsGRP1C bound to the outer layer of C. higginsianum mycelia. SYTOX Green staining assay demonstrated that LsGRP1C caused membrane permeabilization of C. higginsianum and its suppression by cations. Accordingly, disruption of fungal membrane integrity by LsGRP1C due to its net positive charge was presumed. On the other hand, DAPI, H2DCFDA and TUNEL staining assays showed that LsGRP1C caused apoptosis-like programmed cell death phenomena in C. higginsianum, including cytoplasm shrinkage, intracellular accumulation of reactive oxygen species, chromatin condensation and nuclear DNA cleavage etc. Transient expression of LsGRP1, LsGRP1ΔNΔG, LsGRP1ΔC or LsGRP1C in the leaves of Arabidopsis thaliana followed by inoculation with C. higginsianum demonstrated that expression of LsGRP1, LsGRP1ΔNΔG, and LsGRP1C could suppress symptom development and sporulation, and the secondary hyphae were not present as indicated by staining and microscopic examinations. Thus, the primary hyphal cell death of C. higginsianum caused by the expression of LsGRP1, LsGRP1ΔNΔG and LsGRP1C in planta was presumed.

The common bean (Phaseolus vulgaris L.) is an important crop grown widely in Uganda. It is also an important source of income for smallholder farmers particularly women. Despite its importance, production in the cool highland regions is constrained by anthracnose disease which causes losses in both the quantity and the quality of beans produced. The principal aim of this research was to elucidate on the status of dry bean anthracnose and the genetics governing its resistance. A participatory rural appraisal study was conducted to explore farmers' knowledge, experience, problems and cultivar preferences in association with managing dry bean anthracnose disease. This study revealed that anthracnose is an important constraint to production which is not controlled in any way. Although farmers have varying cultivar preferences, they use mostly home saved seed and only 1% could access improved seed. The study suggested the need for practical approaches in the provision of quality anthracnose resistant seed in consideration of farmers' preferences and the dynamics of their rural livelihoods. A study was conducted to determine the variability of the anthracnose (Colletotrichum lindemuthianum) pathogen in some of the major bean growing regions of Uganda. Use was made of a set of 12 internationally accepted anthracnose differential cultivars to identify the physiological races present. The results obtained indicated the presence of eight races with one race (767) being dominant and most aggressive. Differential cultivars AB 136 and G2333 were resistant to all the eight races, and can be utilised as potential sources of resistant genes. A germplasm collection of mostly Ugandan accessions was screened for anthracnose resistance. Using the area under disease progression curve as the tool for assessing disease severity, eleven accessions were identified that posses good levels of anthracnose resistance. The yield loss attributed to the anthracnose disease was determined on three susceptible Ugandan market-class dry bean cultivars and two resistant cultivars. The results showed that the yield of susceptible cultivars was reduced by about 40% and an almost equivalent yield was lost due to poor quality seed. In comparison, the yield lost by the resistant cultivars was not significant. The study suggested the use of resistant cultivars as the best solution in combating anthracnose resistance. Three susceptible Ugandan market class dry bean cultivars and six resistant cultivars were used for the study of the inheritance of resistance to the anthracnose pathotype 767 in a complete 9x9 diallel design. The results clearly indicated that the resistance was predominately conditioned by additive gene action. It was also established that epistatic gene action was important. More than one pair of genes displaying partial dominance were responsible for determining resistance and the maternal effect did not have an influence on resistance. Additionally, the result showed that some of susceptible cultivars combined very well with the resistant cultivars and that anthracnose resistance heritability estimates in both the narrow and broad sense were high. These results suggested that the use of simple pedigree breeding procedures such as backcross selection could be useful in improving anthracnose resistance levels in the Ugandan market class varieties.
Thesis (Ph.D.)-University of KwaZulu-Natal, Pietermaritzburg, 2006.

碩士
國立中興大學
植物病理學系所
103
Tea (Camellia sinensis) and passion fruit (Passiflora edulis) are the most consumed beverages in the world. Brown blight of tea and anthracnose of passion fruit caused by Colletotrichum gloeosporioides are both devastating diseases hindering their production. Extensive use of fungicides has led to serious development of resistance in pathogen populations and caused negative consequences for human health and the environment. Biological control has been taken as an alternative to disease control by synthetic pesticide. Bacillus and Streptomyces species are representative genera of plant growth promoting rhizobacteria which not only promote plant growth but could also act as biocontrol agents by producing antibiotics, triggering induced systemic resistance. The main objectives of my study are to select and identify native antagonistic microorganisms against pathogens of brown blight of tea and anthracnose of passion fruit, and to investigate their potential application in disease control and growth promotion and to investigate the putative control mechanisms. Among seventy isolates which were isolated from rhizospere soils and collected from Nantou county, Kaohsiung city and Taichung city in this study, strains AM4-1, CA-1, 151B1 and 151B4 were classified as Bacillus subtilis group based on the analysis of 16S rRNA sequences, DNA polymorphism, physiology and biochemistry tests and the analysis by Biolog System III. Strains UN3S2 and PES4 were classified as Streptomyces species based on the analysis of 16S rRNA sequences. Eight strains including B. subtilis TKS1-1, Streptomyces griseobrunneus S3 and strains B. subtilis AM4-1, CA-1, 151B1, 151B4 and Streptomyces sp. strain UN3S2 and PES4 showed antagonistic effect on mycelial growth and conidial germination of C. gloeosporioides TE04. Application of B. subtilis 151B1 culture broth increased survival rates of passion fruit cuttings with or without the challenging of anthracnose fungi. Culture filtrates from strains 151B1 and TKS1-1 resulted in cell death and chromatin fragmentation of C. karstii N-PF1. The treatment of culture filtrates from strains 151B1 and TKS1-1 were also found to cause reduction in mitochondrial membrane potential and energy metabolism of Colletotrichum karstii N-PF1 compared to the medium control, suggesting its function in triggering apoptotic-like cell death. In addition, the aberrant hyphal morphology, non germinated conidia were observed for C. karstii N-PF1 on leaves of passion fruit while 12hrs and 24hrs post-challenging with the culture filtrates. Among four strains, TKS1-1, 151B1, S3 and PES4, B. subtilis 151B1 was found to show the superior enhance the leaf length and thickness (for 2-fold increase) of Chin-Shin Oolong seedlings 8 weeks post-treatment, and in numbers of leaves (2.6-fold increase), nodes (2-fold increase), leaf width (2-fold increase), and shoot fresh weight (1.5-fold increase) of passion fruit seedlings. In addition, Streptomyces sp. PES4 showed the greatest effect on promotion of plant height (2-fold increase) of Chin-Shin Oolong seedlings compared to the other treatments. In conclusion, B. subtilis strain 151B1 showed superior antagonistic activity against anthracnose fungi and enhancing the survival rates of passion fruit cuttings and with or without challenging with anthracnose fungi, and exhibited the greatest growth promotion of seedlings of tea and passion fruit. Suggesting it’s potential as a biocontrol agent. Our findings also suggest that culture filtrates of B. subtilis TKS1-1 and 151B1 caused aberrant hyphal morphology and inhibit germination of anthracnose fungi which may part attribute to their ability in triggering program cell death of fungi.

碩士
國立屏東科技大學
植物保護系
94
Six strobilurin susceptible isolates of Colletotrichum gloeosporioides, the causal agent of chilli anthracnose, were collected from the central and southern part of Taiwan. The EC50 of six isolates were from 0.08 to 0.54 µg/ml. Besides, two azoxystrobin resistance isolates, SF02 and SN01, were collected from Pingtung area. The EC50 of two resistance isolates were 41.99 and 56.20 µg/ml and resistance ratios were 262.4 and 351.3, respectively. Azoxystrobin at the rate of 10 µg/ml may be suggested as a discrimination rate for resistance monitoring. A field resistance monitoring was done from 2003 to 2004 in the central and southern Taiwan. The result showed that 38 out of 85 isolates collected from the chilli field affected by C. gloeosporioides had fungicide resistance reaction to azoxystrobin. There was no significant fitness penalty of resistant pathogens that affected the activity of the mycelial growth, production and germination of conidial spores. There was no known point mutation, G143A, found after the PCR-RFLP and DNA sequence analysis. In vitro sensitivities of different C. gloeosporioides isolates to azoxystrobin in the presence of SHAM suggested that other alternative resistance mechanism still existed. The result from the cross-resistance test showed that there was no cross-resistance between azoxystrobin and difenoconazole or cholorthalonil, but cross-resistance between azoxystrobin and trifloxystrobin or kresoxim-methyl was confirmed. The mixture of azoxystrobin with difenoconazloe or with cholorthalonil was suggested as a good partner for the resistance management in the QoI fungicide; furthermore, a better chilli anthracnose disease control approach could be obtained by applying the azoxystrobin mixture instead of azoxystrobin alone.

碩士
國立虎尾科技大學
生物科技系碩士在職專班
107
In order to cope with the possible shortage of food caused by the increase in population, all countries in the world are working hard to increase food production. However, how to achieve a balance between food production and environmental and ecological systems is the goal of national agricultural policy efforts, such as promoting organic agriculture is one of its policies. Biological control is recognized as one of the important means to replace or reduce chemical pesticide use in the future. Therefore, this thesis is conducted to isolate antagonistic microorganisms from organic tea garden and test their disease control ability, in order to develop bio-pesticide in the future. The experimental fields were selected in the Fenghuang Nature Education Areas of the Experimental Forest National Taiwan University to explore the differences in the composition of soil microbial phase structure in tea gardens under the management of organic farming methods and conventional farming methods, and to screen for beneficial microorganisms from their soils and to detect anthracnose disease control of chinese cabbage caused by Colletotrichum higginsianum and the pathogen growth. According to soil microbial survey, the average number of fungi in organic tea gardens (59.82×103 cfu/g) is generally higher than that of conventional tea gardens (18.86×10^3 cfu/g); similarly, the average number of Trichoderma spp. is in organic tea gardens (23.72×10^3 cfu/g) is also about 4 times higher than the conventional tea garden (5.78×10^3 cfu/g). For bacteria and actinomycetes number investigations, the results showed no significant difference between organic tea gardens and conventional tea gardens. Comparing the number of microbial populations with the annual rainfall and temperature, it was found that the number of fungi and actinomycetes was positively correlated with the amount of rainfall in the area, but the number of all microorganisms was not significantly correlated with temperature. For dual culture of pathogen and antagonists on medium, 8 strains of Trichoderma spp. which had better growth inhibition of C. higginsianum were screened out, and to control anthracnose disease of chinese cabbage was detected in the greenhouse. The results showed that the 8 selected strains of antagonists were effective to reduce the anthracnose disease of chinese cabbage about 32~53% compared with the untreated control, among which strain O1-5-3 was the best. This strain O1-5-3 was identified by gene sequencing and assigned as Trichoderma virens.

碩士
中興大學
植物病理學系所
99
Mango anthracnose, caused by Colletotrichum gloeosporioides, is not only an important postharvest disease but also one of the reasons causing crop losses. Up to now, the use of fungicides is the main approach to control mango anthracnose, however, due to the emergence of fungicide-resistant pathogens and the rise of environmental awareness, the search for alternative methods is important. The main purpose of this study is to search and formulate a medium for antagonistic bacteria to control mango anthracnose, and in turn develop it into a biopesticide to control the disease. The antagonistic bacteria obtained from Nantou and Yunlin were identified and evaluated by detached leaf bioassay, hemolysis test and molecular identification – 16S rDNA. Results indicated that five strains, including Paenibacillus macerans EC-21-02, Bacillus licheniformis EC-31-02 and EC-34-01, Brevibacillus aqri NA-27-01 and NN-13-01 were able to reduce the severity of mango anthracnose from 100% to 0%, did not produce hemolysis reactions and were not opportunistic pathogens for humans. Twenty-five cultural media containing different components were designed. The fermented broths of EC-34-01 cultured in SY+raffinose pentahydrate, NA-27-01 in SY+molasses and NN-13-01 in Yeast+NaCl did show to be the most effective in inhibiting mycelial growth of the pathogen. Then fermented broths of EC-34-01, NA-27-01 and NN-13-01 cultured in Surimi-Molasses (SM) and Soybean meal-Surimi-Molasses (SSM) media were analyzed by bioassay method of detached mango leaf. One hundredfold dilution of fermented broth of EC-34-01 cultured in SSM medium could reduce 86% anthracnose severity compared to the control, and found SSM at pH 8.0 was the most optimal for culturing EC-34-01 in controlling mango anthracnose. B. licheniformis EC-34-01 was reconfirmed via physiological and biochemical characteristics assays, Biolog MicrologTM computer software and the analysis of fatty acid methyl esters microbial identification system. In addition, twenty-seven specific primers were used to detect gene sequences in Bacillus licheniformis EC-34-01 with fourteen kinds of ability for producing 4’-phosphopantetheinyl transferase、Acyl-homoserine lactonasem、Bacillomycin A、Bacillomycin D、Iturin C、Iturin A、Bacilysin、Bacillaene、Difficidin、Bacillibactin、Fengycin、Macrolactin、Surfactin and Pleiotropic regulator. After fermentation, five hundredfold dilution of EC-34-01 fermented broth with 5% ethanol was effective in reducing disease severity level from 100% to 10%. Analysis of the crude extracts of EC-34-01 fermented broths by inhibitory activity assay showed that the crude extract obtained from the condition at pH 2 did not only show inhibitory zone for C. gloeosporioides MG-2 but also inhibited its mycelial growth and conidial germination. The Rf value of bioactive compounds of crude extract of EC-34-01 fermented broth was approximately 0.78, indicating the compounds belong to the low polarity and heat-tolerance of peptides.

Soil application of a known activator of Systemic Acquired Resistance (SAR), benzo(1,2,3)thiadiazole-7-carbothioic acid-S-methyl ester (BTH), and Harmonizer, a polychlorinated copper (II) phthalocyanine pigment, reduced severity of Colletotrichum orbiculare in Nicotiana benthamiana by 99% and 38%, respectively. BTH induced expression of nine SAR/progammed cell death-related genes and primed expression of two Induced Systemic Resistance (ISR)-related genes, while Harmonizer induced expression of only one SAR-related gene. Soil application of Harmonizer also reduced severity of Sclerotinia homoeocarpa in Agrostis stolonifera up to 39%, whereas BTH was ineffective. Next generation sequencing identified over 1000 genes in A. stolonifera with two-fold or higher increased expression following Harmonizer treatment relative to a water control, and induced expression of three defense-related genes was confirmed by relative RT-PCR. These results demonstrate that Harmonizer can activate systemic resistance in a dicot and a monocot, but changes in expression of genes indicated that it differed from BTH-activated SAR.
Petro-Canada, Natural Sciences and Engineering Research Council of Canada, Ontario Turfgrass Research Foundation

Avocados are one of the major food sources in tropical and subtropical regions and are an important horticultural crop in South Africa. Avocados are exported over long distances and may have storage times of up to 30 or more days at temperatures of about 5.5oC. This procedure increases the risk of poor fruit quality, including physiological disorders, early softening and postharvest disease incidence. A major component of the postharvest diseases is Anthracnose caused by Colletotrichum gloeosporioides. Anthracnose infects unripe fruit and once infected, the fungus remains dormant in the fruit until ripening begins. This leads to a problem for producers and packers, as the presence of the disease cannot be detected on the pack line, and fruit is not removed. Anthracnose control is normally done through pre-harvest treatment with copper-based fungicides. While effective such treatment needs to be repeated frequently, resulting in copper residues on the avocados. The study was conducted to investigate the effects of phosphoric acid and potassium silicate on known antifungal compounds and critical enzymes of the pathways elemental for systemic resistance inducers, so as to evaluate the potential for using them as alternatives to or in conjunction with, copper fungicides in the control of Anthracnose in avocado fruit. The study included storage temperature and time variations, to take account of the logistics in shipping avocado fruit to distant markets. Pre- and postharvest applications of phosphoric acid and potassium silicate were used, and after harvest, fruit were either ripened at room temperature (22oC) without storage or stored for 28 days at temperatures of 5.5oC or 2oC before analysis. Concentrations of phenolics, activity of the enzyme phenylalanine ammonia lyase (PAL) and a known antifungal diene were determined in the fruit exocarp. Pre-harvest treatments of phosphoric acid showed that the highest phenolic concentration was found in fruit harvested 14 days after application for fruit stored at room temperature. For fruit stored at 5.5°C it was seen that as fruit softened, phenolic concentrations increased compared with hard fruit immediately after storage, with the highest increase noted for fruit harvested 7 days after application. When comparing the three storage temperatures, phenolic concentrations were enhanced most when fruit was stored at 2°C. Postharvest treatments showed a significant increase in phenolic concentrations for potassium silicate treated fruit stored at room temperature and 2°C when determined immediately after storage. Fruit stored at 5.5°C showed an increase in phenolic concentrations as it became softer. When considering PAL enzyme activity, it was found that postharvest treatments of both potassium silicate and phosphoric acid influenced enzyme activity, with potassium silicate having greater effects. Similarly, an increase in PAL activity was noted in the pre-harvest phosphoric acid treatment harvested 14 days after application for fruit ripened immediately as well as fruit stored at 5.5°C. Fruit stored at 2°C showed the highest PAL activity for fruit harvested 7 days after application. No results were obtained in the analysis of antifungal compounds for both pre- and postharvest treatments. However, it is suggested that the antifungal diene could follow similar trends to those found for phenolics. It is concluded that applications of both phosphoric acid and potassium silicate do create changes in phenolic concentrations and the activity of the enzyme PAL which is involved in the synthesis of phenolic compounds known to possess antifungal properties. It is therefore possible that phosphoric acid and potassium silicate may be used as part of an integrated programme for Anthracnose control, and should be tested as potential alternatives for high volume copper-based fungicides.
Thesis (M.Sc.)-University of KwaZulu-Natal, Pietermaritzburg, 2012.