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Seed quality plays an important role in the establishment of healthy crop stands. The aim of this study was to identify the mycoflora associated with maize grain collected over two growing seasons, one experiencing severe drought, from smallholder farms in KwaZulu-Natal (KZN) and the Eastern Cape (EC). These are two adjacent provinces in South Africa with many maize-producing smallholder farmers. Asymptomatic maize ears were collected at harvest during the 2014/2015 and 2015/2016 maize growing seasons from farms located in Hlanganani (KZN), Ntabamhlophe (KZN), KwaNxamalala (KZN), Bizana (EC) and Tabankulu (EC). Maize grain was subjected to seed health tests using the agar plate method. The percentage incidence of fungal species isolated from maize grain was determined with species identities confirmed by ITS sequencing. Eleven fungal genera were identified with Fusarium species and Stenocarpella maydis the most prevalent. Fusarium verticillioides, Fusarium graminearum and S. maydis were isolated from all sites in both seasons. No fungal species exhibited a higher incidence in the drought season across all sites. F. graminearum and S. maydis had higher incidences in the wetter season at four and three sites, respectively. F. verticillioides had a greater incidence at the EC sites, particularly the coastal Bizana site. We conclude that local factors that affect the inoculum levels of each species, such as land preparation, previous crop, amount of debris from previous season, fertilizer application, and the micro-environment at field scale had a greater impact than the drought season on the population structure of ear-rot pathogens. The widespread presence of fungi that are potentially mycotoxin-producing in asymptomatic maize grain poses health risks to consumers and is worthy of further investigation.

Diplodia ear rot (DER), caused by the fungus Stenocarpella maydis, has become a prevalent corn disease in the Midwest. The survival of S. maydis on corn kernels and stalks was evaluated in Tippecanoe, Co., Indiana, over 17 months. Infested corn residue was either left on the soil surface or buried at 10 or 20 cm. Survival of S. maydis was determined by observing the presence of pycnidia per unit of corn residue under a stereomicroscope and by assessing conidia viability by evaluating germination. Months of environmental exposure and soil depth affected S. maydis survival. Pycnidia production and conidia germination were observed up to 11 months on corn kernels and 17 months on corn stalks when left on the soil surface. Minimal or no conidia germination of S. maydis was observed on residue recovered from 10 or 20 cm after 4 months on kernels and 7 months on stalks. These results demonstrate that surface corn residue can be a long-term source of inoculum and at least two years of rotation to a nonhost crop or residue removal through tillage may be necessary to reduce potential initial inoculum.

Although stalk and ear rot is a major disease affecting corn plants, little is known regarding the genetic variability of Stenocarpella maydis, its main causal agent. In this study, we evaluated inter-simple sequence repeat (ISSR) primers for analysis of genetic variability of the pathogen. For DNA analysis, six isolates of S. maydis were amplified using 42 ISSR primers. Polymorphic information content (PIC), marker index (MI), and resolving power (RP) were calculated for all primers. Of the 42 primers, 50 % showed high-quality amplification products. The average polymorphism was 90.4 %, and the average similarity among the isolates was 33 %. The PIC, MI, and RP values ranged from 0.31 to 0.45, 4.84 to 18.46, and 0.40 to 10.00, respectively. The primers UBC 848, 873, 808, 811, 807, 827, 861, 868, 809, and 835 were found to be the most informative for genetic variability analysis. Considerable genetic variation was observed in S. maydis, with a core set of 10 primers.

The causal agents of Diplodia ear rot are two species of the Stenocarpella genus, S. macrospora and S. maydis. In addition to ears, both pathogens can infect leaves and stalks, and both are present in most corn-production regions around the world. It is difficult to visually distinguish between the two pathogens based on plant symptoms and fungal signs. To facilitate accurate and rapid pathogen identification, polymerase chain reaction (PCR) assays were developed for identification of each species. Species-specific primers of 18 to 20 nucleotides in length were designed, targeting a portion of the internal transcribed spacer (ITS) region of the fungal genome for conventional and real-time PCR assays. The conventional PCR method successfully amplified a single 1.7-kb and 800-bp fragment for each S. maydis and S. macrospora isolate, respectively. The real-time method was performed using SYBR green dye, and detection of each specific target pathogen was successfully obtained. In total, 82 S. maydis and 15 S. macrospora isolates were tested to evaluate the reproducibility of these primers. Both methods provide a rapid and specific tool for the detection of Stenocarpella spp.

This study aimed to find an efficient method of inoculation of Stenocarpella maydis to produce white ear rot (WER) and estimate pathogen damage on maize grain yield components. Measured components were ear mass, grain mass per ear, and thousand grain weight. The experiments were performed in Ponta Grossa, PR, Brazil in a randomized block design with treatments arranged in a split plot with three replications. Three hybrids were studied. For each, five methods of inoculation in the ear at the soft dough stage were compared to un-inoculated controls. The ears were inoculated with 1 mL of the spore suspension (104 conidia/ml). Evaluations of the disease index (%), severity (%) and lesion area were performed in three (1st experiment) and four periods (2nd experiment), and the area under the disease progression curve was calculated for each of these periods. The area under the disease progression curve was calculated for each period. Inoculation at the center of the ear resulted in the best growth and development of the pathogen in both experiments. Inoculation at the base and center of the ears resulted in greater reductions in yield components, with degrees of damage varying from 27.8 to 43.1%. The inoculation of S. maydis in the center of the ear can be considered an appropriate method for resistance screening to WER in maize breeding programs. Key words: White ear rot, disease progress curve, severity, yield components.

Diplodia ear rot, a corn (Zea mays L.) disease caused by the fungus Stenocarpella maydis (Berk.) B. Sutton, has been a persistent ear rot across the United States. Management options are currently limited. Field trials conducted under inoculated and non-inoculated conditions were established at two locations in Indiana from 2011 to 2013 to test the fungicides azoxystrobin plus propiconazole and prothioconazole against Diplodia ear rot. Fungicides were applied at three individual growth stages during each year. Fungicides did not consistently reduce Diplodia ear rot compared to non-fungicide-treated controls in any year. Applications also did not consistently increase yield at any timing under inoculated and non-inoculated plots compared with the non-fungicidetreated control. Fungicides were tested in an in vitro assay to determine the effective fungicide concentration at which 50% of mycelial growth or conidial germination of S. maydis was inhibited (EC50). Propiconazole and prothioconazole EC50 values indicated efficacy in reducing fungal growth under controlled conditions; however, current fungicide application methods and plant barriers to fungicide contact with the pathogen may prevent these products from effectively reducing Diplodia ear rot in a field setting. Accepted for publication 23 August 2015. Published 1 September 2015

Given what is already known in regard to seed health and the availability of molecular methods for detection of the pathogens Stenocarpella maydis and Stenocarpella macrospora in maize seeds, Colletotrichum gossypii var. cephalosporioides in cotton seeds, and Corynespora cassiicola in soybean seeds, the aim of this study was to evaluate seed vigor according to different inoculum potentials. The fungus isolates were inoculated on seeds by the technique of water restriction, through which different inoculum potentials are obtained, corresponding to times of seed exposure of 0, 24, 48, and 96 hours for maize and cotton seeds, and 0, 36, 108, and 144 hours for soybean seeds. The seeds were subjected to germination, electrical conductivity, health, and qPCR tests. Results of the blotter test showed that in most pathosystems, there was a higher incidence of the fungi with an increase in inoculum potential. A decrease in germination percentage was observed in all species as inoculum potential increased, as well as further degradation of seed membranes. The qPCR test confirmed that the most damaged seeds in the tests had higher presence of the pathogens. Resumo: Diante do que já se conhece em sanidade de sementes e tendo-se em mãos métodos moleculares para a detecção dos fungos Stenocarpella maydis, Stenocarpella macrospora em sementes de milho, Colletotrichum gossypii var. cephalosporioides em algodão e Corynespora cassiicola em soja, objetivou-se neste estudo avaliar o vigor das sementes em função dos diferentes potenciais de inóculo. Os isolados dos fungos foram inoculados nas sementes por meio da técnica de restrição hídrica pela qual se obtém diferentes potenciais de inóculo, que foram representados por P0, P24, P48 e P96 e P0, P36, P108 e P144, que correspondem à exposição das sementes aos períodos de tempo de 0, 24, 48, 96 horas (milho e algodão) e nos tempos 0, 36, 108 e 144 horas (soja), respectivamente. As sementes foram submetidas aos testes de germinação, condutividade elétrica, sanidade e qPCR. Pelo blotter test, na maioria dos patossistemas, houve maior incidência do fungo com o aumento do potencial de inóculo. Foi observada uma queda na porcentagem de germinação de todas as espécies com o aumento do potencial de inóculo, assim como maior degradação das membranas das sementes. A qPCR confirmou que nas sementes mais prejudicadas havia maior quantidade de inóculo dos patógenos.

Stenocarpella macrospora and Stenocarpella maydis may result in the seedlings death or cause rotting at the corn stalk base and in all or part of the ear. In addition, S. macrospora can cause leaf spot. Double-haploid strains from corn hybrids resistant to S. macrospora and S. maydis were identified. Also the incidence of these pathogens in the Cerrado and in Southern Brazil localities was determined. One hundred and forty double-haploid maize hybrids, in addition to the controls, were inoculated with S. macrospora and S. maydis and evaluated for resistance reaction in three locations of the Cerrado and three locations of the South regions. The grains attacked by these fungi were collected and variable quantities of S. macrospora, S. maydis and other fungal species were registered. The results demonstrated the prevalence of S. macrospora in the Cerrado as well as other non-Stenocarpella sp. fungi in the South. The city of Abelardo Luz (Santa Catarina) was the only place where S. maydis was found to have a higher incidence than S. macrospora. Environmental effects influence the prevalence of fungi, causing grain rot. These results indicated genetic gains in the selection of hybrids resistant to this fungi for use as direct breeders in Stenocarpella-corn pathological system research. Stenocarpella macrospora e Stenocarpella maydis em milho, podem resultar na morte de plântulas ou causar apodrecimento na base do caule e da totalidade ou parte da espiga. Além disso, S. macrospora pode causar manchas foliares. Identificou-se linhagens duplo-haplóides de híbridos de milho resistentes a S. macrospora e S. maydis; determinou-se também a incidência desses patógenos no Cerrado e do Sul do Brasil. Cento e quarenta híbridos duplo-haplóides de milho além dos controles (testemunhas) foram inoculados com S. macrospora e S. maydis e avaliados quanto à resistência em três localidades do Cerrado e três de Sul do Brasil. Os grãos atacados pelos fungos foram colhidos e avaliados quanto à incidência dos dois patógenos. Foram estimadas as porcentagens (%) de S. Macrospora e de S. Maydis e também a ocorrência de outros fungos pelo método de blotter. Houve maior presença de S. macrospora do Cerrado. No Sul do Brasil, o município de Abelardo Luz foi o único local onde S. maydis foi encontrado em maior incidência do que S. macrospora. Os resultados mostraram efeitos ambientais sobre a prevalência de fungos que causam grãos ardidos. Estes resultados indicaram ganhos genéticos na seleção de híbridos resistentes ao fungo S. Macrospora e obtenção de híbridos resistentes em milho, tanto na região do Cerrado como no Sul do Brasil.

A produtividade e a qualidade do milho podem ser influenciadas pela ocorrência de doenças, cuja intensidade pode ser afetada por fatores ambientais e por práticas fitotécnicas. Dessa forma, o objetivo deste estudo foi avaliar o efeito do arranjo de plantas de milho e da densidade de semeadura no rendimento de grãos e na massa de mil grãos, bem como na incidência de podridões da base do colmo, grãos ardidos e de fungos associados aos grãos. O estudo foi realizado nas safras 2010/2011 e 2012/2013, em Chapecó, SC, Brasil. Os tratamentos foram constituídos por diferentes espaçamentos entre linhas (0,45 e 0,80 m) e densidades de plantas (50, 60, 70, 80 e 90 mil plantas ha-1), com delineamento de blocos casualizados em esquema de parcelas sub-subdivididas. Na parcela principal foram testados dois híbridos simples de milho de ciclo precoce (P30F53H e DKB 240H), nas subparcelas os espaçamentos e nas sub-subparcelas as populações de plantas. As unidades experimentais consistiram de seis linhas de 4 m de comprimento. O rendimento de grãos no espaçamento 0,45 m foi 29,65% superior ao obtido no espaçamento 0,80 m para os híbridos estudados. A massa de mil grãos e a incidência de grãos ardidos reduziram com o aumento da população de plantas. No entanto, a incidência de Fusarium verticillioides, Fusarium graminearum e Stenocarpella maydis não foi influenciada pelas populações de plantas nem pelos espaçamentos testados nas condições do presente estudo.

Stenocarpella maydis and Sporisorium reilianum are phytopathogenic fungi that cause white rot in corn cob and head smut in maize (Zea mays L.) respectively, diseases that are spread worldwide and cause many economic losses. In this chapter the characteristics of the above diseases, such as their life cycle, pathogenicity factors, control methods, as well as the biotechnological potential of the fungi involved in this processes are described, specifically in connection to their extracellular enzymes.

Diplodia ear rot (DER) has been a persistent corn disease across the Midwest, and in recent years it has become an annual problem. The objectives of this study were to i) develop a molecular assay for the identification of Stenocarpella maydis, causal agent of DER, ii) evaluate the effect of crop rotation and tillage on DER severity, iii) determine the survival period of S. maydis in corn residue at different soil depths in a corn field, and iv) identify genetic diversity among S. maydis isolates collected from the Midwestern and Southern United States. The genus Stenocarpella contains two species, S. maydis and S. macrospora, both able to infect corn plants. Previously, identification of the two species was based only on time consuming morphological methods. An accurate molecular assay was developed that can be performed under conventional or real-time PCR, and both are able to distinguish between S. maydis and S. macrospora from pure culture or infected tissue. Field experiments were conducted at the Agronomy Center for Research and Education (ACRE) in Tippecanoe County, Indiana. From 2013 to 2014, four corn production systems were evaluated for DER control under non-inoculated and inoculated conditions. Stenocarpella maydis survival was assessed over 12 months at the soil surface, and at 10 and 20 cm burial depths. Results indicate that crop rotation with soybean and tillage practices did not eliminate Diplodia ear rot, and in certain years did not reduce DER severity. Stenocarpella maydis was able to survive on the soil surface in corn residue for at least 11 months. However, S. maydis conidia recovered from samples were not able to infect corn ears under controlled conditions after 7 months. These results indicate that at least a one year rotation away from corn is necessary in order to eliminate inoculum. Tillage operations must reduce surface residue to a minimal percentage to reduce inoculum. High genotypic diversity was observed among S. maydis isolates in the US. Genotypic diversity was higher within sample regions and similar between the Midwestern and Southern US S. maydis populations. This indicates that recombination occurs among S. maydis isolates. Although no sexual reproduction has been reported, both mating type genes, Mat1-1 and Mat1-2 were identified and amplified across 60 selected isolates.

The British mycologist, I.B. Pole-Evans, was appointed as the first South African government mycologist in 1905 following the Anglo-Boer War (1899-1902). The Onderstepoort Veterinary Research Institute was founded in 1908 with the Swiss veterinarian, Arnold Theiler, as the first director. Thus, the stage was set for the commencement of mycotoxicological research when the Union of South Africa came into being in 1910. The first accounts of this pioneering research appeared in the 'Seventh and eight reports of the Director of Veterinary Research, Union of South Africa. 1918' in which D.T. Mitchell reported on the experimental reproduction of the neurotoxic syndrome, diplodiosis, in cattle with pure cultures of Stenocarpella maydis (= Diplodia zea) isolated by P.A. Van der Bijl and grown on sterile maize kernels. This is the first report of the experimental reproduction of a veterinary mycotoxicosis with a pure culture of a fungus in South Africa and possibly in the world. This seminal research was followed by a great deal of multidisciplinary research on veterinary mycotoxicoses as well as human syndromes in which fungal toxins are suspected to be involved, taxonomy of mycotoxigenic fungi and chemistry of mycotoxins in South Africa. The mycotoxicoses studied in South Africa include the following (more or less in chronological order): diplodiosis, Paspalum staggers, aflatoxicosis, human hepatocellular carcinoma, ochratoxicosis, lupinosis, facial eczema, tremorgenic mycotoxicosis, hyperoestrogenism, stachybotryotoxicosis, ergotism, leukoencephalomalacia and human oesophageal cancer. A major breakthrough in mycotoxicological research was made in South Africa in 1988 with the isolation and chemical characterisation of the carcinogenic fumonisins produced by Fusarium verticillioides in maize. Current research at the PROMEC Unit of the South African Medical Research Council on the risk assessment of fumonisins and intervention methods to reduce fumonisin intake by rural populations on a maize staple diet is highlighted. This paper concludes with a selected list of mycotoxicological publications by South African mycologists/plant pathologists, veterinarians and chemists/biochemists.

Diplodiosis is a neuromycotoxicosis of cattle and sheep caused by ingestion of maize infected with the ear-rot fungus Stenocarpella (= Diplodia ) maydis . Apart from ataxia, paresis, and paralysis, the toxin is responsible for stillbirths and neonatal losses characterized by the presence of spongiform degeneration in the white matter of the brain in the offspring of dams exposed to infected maize cobs. In the present study a toxin, named diplonine, which induced neurological signs in guinea pigs resembling some of those occurring in cattle and sheep, was isolated from S. maydis cultures. Purification of diplonine was achieved by methanol extraction followed by chromatographic separation on silica gel and RP-18 stationary phases. The structure and relative configuration of diplonine were defined by analysis of NMR and MS data as (S)-2-amino-2-[(1R,2S)-1-hydroxy-2-methylcyclopropyl]acetic acid or the (S)-2-amino-2-[(1S,2R)-diastereomer.

Research that focused on maize production on the central Highveld of South Africa was traditionally, since the early twentieth century, dealt with by the Department of Agriculture at Potchefstroom. During the 1970s the Highveld Region at Potchefstroom assigned a group of researchers with an interest in maize to a unit termed the Summer Grain Centre that in 1992 became part of the Agricultural Research Council’s Grain Crops Institute. Members of this group in particular gave research attention to various aspects of crop protection in maize, including the soil insect complex, the maize stem borer complex, maize streak disease, the ear rot complex and various leaf diseases. This paper deals with research progress on the African stem borer, Busseola fusca, and ear rot caused by Stenocarpella maydis. The emphasis is on research with a practical application rather than that of an academic nature.

In an attempt to obtain superior levels of resistance to Stenocarpella maydis ear rot, three elite maize inbred lines (DO620Y, E739 and B37) from different genetic backgrounds that had previously been shown to have useful levels of resistance were crossed in all combinations. Additionally, two susceptible experimental maize populations were crossed to a resistant experimental inbred line as donor parent. Progeny in each of the five populations was selfed and selected for resistance over five successive generations, following artificial inoculation of plants at the localities Potchefstroom, Cedara and Hazyview. The five populations progressed from the S1 during 1997/98 to the S5 during 2000/01. A progressive improvement in resistance occurred over seasons in all progenies. After three cycles of selection, approximately 50% of the selections showed less than 10% infected ears, with a total of 84 selections showing no infection. Selections displaying the lowest disease incidence at any one locality were not ...

Stenocarpella maydis is one of the most prevalent ear and stalk rot pathogens of maize globally, causing reductions of grain quality and yield. Various molecular methods, including polymerase chain reaction (PCR)-based techniques and nucleotide microarrays, have been developed for the identification of S. maydis infestation in maize grain. In addition to diplodiatoxin, new metabolites, namely dipmatol, diplonine and chaetoglobosins K and L, have been isolated from S. maydis infected cultures. S. maydis infected maize is also associated with intoxication in ruminants. Diplodiosis, a nervous disorder of cattle and sheep, results from ingestion of mouldy ears, kernels and maize stubble infected by S. maydis. Although this disease is most common in southern Africa, it has also been reported in Australia, Argentina and Brazil. Diplodiosis is characterised by reluctance of the animals to move, a wide-based stance, incoordination, paralysis and death. Myelin degeneration (status spongiosis) is the only histopathological change observed in affected animals, especially in cases of perinatal mortality. To date, none of the purified S. maydis metabolites has been administered to ruminants in order to reproduce diplodiosis. However, recent studies have focused on investigating the toxicity of the metabolites on cell cultures. Cytotoxicity studies where cultured cells were exposed to the S. maydis metabolites indicated that diplodiatoxin and dipmatol affected the activity of the mitochondrial succinate dehydrogenase enzyme and the overall viability of the cells. More detailed in vitro toxicity studies are still required to elucidate how the currently available S. maydis metabolites influence parameters such as the mechanism of cell death. Development of analytical test methods to quantify and establish the presence and distribution of these mycotoxins in infected maize commodities also needs investigation. It is also critical that the role of S. maydis stalk rot be evaluated as a potential source and cause of diplodiosis.

The objective of this study was to provide a better understanding of the interrelationship between methods of screening for resistance to ear rot, caused by Stenocarpella maydis. The inheritance of resistance as well as genetic and phenotypic correlation of various selection criteria were investigated. A complete diallel cross of 10 inbred lines, ranging from resistant to susceptible, was evaluated at each of three localities, representative of different environments within the South African maize production area. Plants were artificially inoculated twice during the vegetative stages of plant development. Resistance to ear rot was found to be controlled by additive gene effects, with low dominance and interaction effects. Significant genetic correlations existed between various characteristics relating to disease incidence, with high heritability and highly correlated responses. The incidence of rotted ears was the most practical and reliable method to select for resistance. However, the simultaneous use ...

Diplonine, a mycotoxin that induces neurotoxic clinical signs in the guinea pig, resembling those occurring in cattle and sheep with diplodiosis, was isolated previously from a Stenocarpella maydisculture. Knowledge of the chemical properties of the toxin, which was characterised as a substituted ß-cyclopropylamino acid, enabled amendments in the present study to the initial steps of the isolation procedure. Extraction with water and fractionation by cation exchange chromatography improved the efficiency of isolation, potentially allowing the preparation of larger amounts of the toxin.

Fifteen maize inbred lines were evaluated for resistance to ear rot, using artificial inoculation of plants with isolates of Stenocarpella maydis from 13 localities in the South African maize production area. Five inbreds and four isolates were common to the four seasons of evaluation. The inbreds B37, D0620Y and E739 showed levels of resistance superior to that of the standard Hill. Isolates provided inconsistent infection levels in different years. All interactions were significant, with seasonal effects predominating over the genotype x isolate interaction.

The cytotoxicity of three Stenocarpella maydis metabolites (diplodiatoxin, dipmatol and diplonine) was investigated on Neuro-2a, CHO-K1 and MDBK cell lines. Diplodiatoxin was the most cytotoxic followed by dipmatol. Conversely, diplonine was not cytotoxic. Diplodiatoxin and dipmatol affected mitochondrial succinate dehydrogenase (MTT assay) and the overall viability of cells as assessed in real-time (xCELLigence assay). The results obtained so far indicate that diplodiatoxin and dipmatol exert their toxicity possibly via the necrotic cell death pathway.

Stem and ear rot caused by Stenocarpella maydis are responsible for severe losses in maize production. Treatment of seeds with fungicides may induce environmental damage. Hence, this study aimed to evaluate the effects of essential oils extracted from Cymbopogon winterianus, Thymus vulgaris, Cymbopogon citratus, Corymbia citriodora, Cinnamomum zeylanicum, and Syzygium aromaticum on the development of in vitro S. maydis. In addition, maize seeds were treated with these essential oils to determine their possible mode of action and effects. The oils from S. aromaticum, C. zeylanicum, and T. vulgaris inhibited fungal development at concentrations higher than 0.025%. The oils from S. aromaticum and C. zeylanicum showed seed germination rates of 89.0% and 84.5%, which were higher than that of the control. The oils from S. aromaticum and C. zeylanicum reduced the pathogen incidence in the seeds to 39.0% and 28.0%, respectively. Further, these oils as well as that from T. vulgaris produced lower reduction of maize stand. Scanning electron microscopy examination revealed that essential oils from S. aromaticum and T. vulgaris acted directly on the conidia, impeding germination. The findings suggest that the oils from S. aromaticum, C. zeylanicum, and T. vulgaris are potential alternatives for maize seed treatment in the control of S. maydis. A podridão-do-colmo e espiga, causada por Stenocarpella maydis, é responsável por graves perdas na produção de milho. O tratamento de sementes com fungicidas pode provocar danos ambientais. Por isso, este estudo objetivou avaliar o efeito de óleos essenciais extraídos de Cymbopogon winterianus (citronela), Thymus vulgaris (tomilho), Cymbopogon citratus (capim-limão), Corymbia citriodora (eucalipto), Cinnamomum zeylanicum (canela) e Syzygium aromaticum (cravo-da-Índia) sobre o desenvolvimento de S. maydis in vitro. Além disso, sementes de milho foram tratadas com esses óleos essenciais para determinar seus possíveis modos de ação e efeitos. Os óleos de S. aromaticum, C. zeylanicum e T. vulgaris inibiram o desenvolvimento do fungo nas concentrações maiores que 0,025%. Os óleos de S. aromaticum e C. zeylanicum mostraram taxas de germinação de sementes de 89,0% e 84,5%, as quais foram maiores que a testemunha. Esses óleos reduziram a incidência do patógeno nas sementes para 39,0% e 28%, respectivamente. Além disso, esses óleos, bem como o óleo de T. vulgaris, produziram menor redução do estande de milho. O exame de microscopia eletrônica de varredura revelou que os óleos de S. aromaticum and T. vulgaris agiram diretamente sobre os conídios, impedindo a germinação. Os resultados deste trabalho sugerem que os óleos de S. aromaticum, C. zeylanicum e T. vulgaris são alternativas potenciais para o tratamento de sementes de milho no controle de S. maydis.

This study aimed to evaluate the production and germination of Stenocarpella maydis pycnidiospores under in vitro conditions as affected by substrate composition (sorghum, wheat, black oat or barley), light regime (continuous dark, 12-h light dark -1 or continuous light conditions), and incubation temperature (21, 24, 27, 30 or 33oC). Each substrate (20 g of grain) was soaked in 100 mL of water for 24h and sterilized twice for 20 min at 127oC. Three plugs (5 mm diameter) of a single-spored culture of S. maydis were used as inocula for each substrate. Assessments of pycnidiospore production per gram of grain and percent germination were made 14 days after inoculation. Barley, black oat or wheat grains were the best substrates for the mass production of S. maydis pycnidiospores and the maintenance of high germination rates. The highest pycnidiospore production (67,600 pycnidiospores g -1 ) was obtained using barley grain as a substrate with incubation at 27oC under a 12-h light dark -1 cycle.

Stenocarpella maydis is a fungal pathogen of major importance that causes a dry-rot of maize ears and is associated with a neuromycotoxicosis in cattle grazing harvested maize fields in southern Africa and Argentina. In an effort to investigate the potential roles of S. maydis metabolites in the fungal disease cycle, ethyl acetate extracts of solid-substrate fermentations of several S. maydis isolates from maize grown in the United States were found to exhibit significant phytotoxic, antifungal, and antiinsectan activity. Chemical investigations of extracts of S. maydis isolates from Illinois and Nebraska led to the isolation or detection of the known metabolites diplodiatoxin, chaetoglobosins K and L, and (all- E )-trideca-4,6,10,12-tetraene-2,8-diol as major components. A culture of Stenocarpella macrospora from maize grown in Zambia produced diplosporin and chaetoglobosins K and L as major components that were isolated. Diplodiatoxin produced significant lesions in a maize leaf puncture wound assay. Diplosporin and chaetoglobosin K displayed moderate antiinsectan activity in dietary assays against the fall armyworm Spodoptera frugiperda , while chaetoglobosin K exhibited significant antifungal activity against Aspergillus flavus and Fusarium verticillioides . Using LC-ESIMS and 1 H NMR data, diplodiatoxin was detected as a major component in S. maydis -rotted grain, stalks, and stalk residues. This constitutes the first report of chaetoglobosins K and L from S. maydis , of (all- E )-trideca-4,6,10,12-tetraene-2,8-diol from Stenocarpella , and the first reported detection of diplodiatoxin, or any other Stenocarpella metabolite, in diseased maize seeds and stalk tissues.

The optimum inoculation position on the maize plant, and optimum time of inoculation with Stenocarpella maydis for artificial induction of ear rot were determined in field trials. The placement of a conidial suspension on the shank at silking resulted in the highest incidence of S. maydis ear rot.

Survival of and pycnidium production by Stenocarpella maydis in artificially infected maize stalk stubble placed on, half buried into, and buried in the soil were determined in field trials over a nine month period. Survival was higher in stubble which remained on the soil surface than in buried stubble. Pycnidial development increased on surface and buried stubble from September onwards, but subsequently decreased on buried stubble. Conidia retrieved from the variously positioned stubble segments did not vary in viability. More pycnidia were produced over an eleven month period on naturally infected stubble retrieved from the soil surface than from in soil. Reisolation and conidium viability of S. maydis from the surface stubble were higher than from buried stubble. Zusammenfassung Der Zusammenhang zwischen der Lage des Maisstoppels im Boden und dem Uberleben von Stenocarpella maydis (Diplodia maydis) Uber einen Zeitraum von 9 Monaten wurde das Uberleben und die Pyknidienproduktion von Stenocarpella maydis in kunstlich inokulierten Maisstoppeln unter Feldbedingungen beobachtet. Die inokulierten Stoppeln wurden auf dem Boden, zur Halfte oder ganz im Boden plaziert. Die hochste Uberlebensrate wurde in den Stoppeln, die auf der Bodenoberflache lagen, festgestellt. Die Entwicklung von Pyknidien erhohte sich an den Stoppeln ab September, unabhangig von der Lagerung, bei den begrabenen Stoppeln wurde jedoch ein Ruckgang der Pykinidienentwicklung zu einem spateren Zeitpunkt beobachtet. Die Viabilitat der Konidien alier Stoppeln war gleich. Mit naturlich befallenen Stoppeln wurde eine hohere Anzahl von Pyknidien uber eine 11monatige Periode von den Stoppelstucken gewonnen, die von der Bodenoberflache aufgelesen worden waren, als von denen, die in der Erde begraben waren. Auserdem war die Reisolierung und Konidienviabilitat bei den von der Oberflache stammenden Stoppelstucken hoher als von den begrabenen.

A podridão do colmo e a podridão espigas do milho (Zea mays L.) causadas pelos fungos StenocarpeLZa macrospora (Earle) Sutton [Sino Diplodia macrospora Earle in BulI.] e Stenocarpella maydis (Berk.) Sutton [Sino Diplodia maydis (Berk.) Sacc.; D. Zeae (Schw.) Lev.] são de ocorrência ampla, abrangendo toda a região de plantio dessa cultura, principalmente nas regiões quentes e úmidas. Esses fitopatógenos afetam a germinação das sementes, causam colapso prematuramente das plantas infectadas na fase reprodutiva e paralisam o processo normal de enchimento de grãos, com conseqüênte queda da produtividade. A presença desses fungos nos grãos de milho reduz a qualidade nutricional das rações, além de poduzirem micotoxinas. Normalmente são necessários quatorze dias para detecção desses fungos, nos testes de sanidade com incubação direta da semente de milho. Neste sentido, os objetivos desta pesquisa foram: avaliar, em maiores detalhes, as relações do complexo Stenocarpella em associação com as sementes de milho; caracterizar por meio de técnicas moleculares os fungos S. macrospora e S. maydis e verificar a possibilidade do uso de marcadores moleculares e proteõmicos como parte de testes diagnósticos destes fungos em sementes. Observou-se o efeito de cinco isolados de S. mcrospora sobre os híbridos de milho DKB-212 e DKB-333-B e o efeito dos cinco isolados de S. maydis sobre os híbridos de milho DKB-214 e DKB-333-B, inoculados em sementes. Observou-se que todos os isolados de S. macrospora aumentaram o índice de severdiade de doença (ISD) e afetaram a germinação, a emergência de plântulas, o índice de velocidade de emergência (IVE), a sobrevivência de plântulas e o peso de matéria seca da parte aérea (PMSA) e de raiz (PMSR). Todos os isolados de S. maydis, exceto um (May43), proporcionaram o aumento do ISD das sementes dos híbridos DKB-214 e DKB-333-B inoculados, e afetaram negativamente a germinação, a sobrevivência de plântulas, o PMSA e o PMSR do híbrido DKB-333-B. Para as sementes do híbrido DKB-214, apenas dois isolados (May-57 e May-74) afetaram negativamente essas variáveis. A caracterização morfológica dos isolados de S. macrospora e de S. maydis foi determinada através do crescimento miceliano, o número de picnídioslcm2 em meio BDA, OA, EMA, CZA (meio Czapek), em dois regimes de temperatura, a 20°C e a 25°C. A caracterização molecular dos isolados de S. macrospora e de S. maydis foi realizada por meio do seqüenciamento dos produtos de PCR obtidos com oligonucleotídios desenhados para a região ITS do rDNA e para o gene da beta-tubulina. Ambos os métodos foram eficientes na diferenciação dos isolados de S. macrospora e de S. maydis. Entretanto, não foi possível distinguir, com consistência, a origem geográfica dos isolados, a partir das análises de caracterização morfológica e molecular dos isolados de S. macrospora e S. maydis. Os produtos da digestão tríptica do extrato de proteína total obtidos de culturas puras de Drechslera maydis, S. maydis, S. macrospora, F. verticillioides e de sementes de milho inoculadas com S. maydis e S. macrospora, pela técnica de restrição hídrica (RH) por 72 horas foram analisadas por RP-HPLC. A análise em espectrômetro de massa Q-TOF foi realizada com os produtos da digestão tríptica do extrato protéico total de sementes de milho não inoculadas, das sementes inoculadas com S. maydis e com S. macrospora, pela técnica de restrição hídrica e o extrato miceliano de culturas puras de S. maydis, S. macrospora e F. verticillioides. Os dados obtidos por RP-HPLC revelaram diferenças nos perfis cromatográficos de culturas puras de ambos fungos, no tempo de retenção de 25 minutos. Entretanto, diferenças marcantes nos perfis cromatográficos obtidos de sementes inoculadas com os dois fitopatógenos, não foram identificadas por essa técnica. A análise discriminante realizada com base nos espectros MS, dos intervalos de 20 a 30 minutos e 50 a 60 minutos, obtidos das diferentes amostras estudadas revelaram que os fungos S. maydis e S. macrospora podem ser detectados diferencialmente em sementes de milho e também possibilitaram a distinção das diferentes espécies fúngicas analisadas. Desta forma, os dados obtidos neste trabalho, indicam que existe grande potencial de utilização da espectrometria de massa no desenvolvimento de novos métodos de detecção de fitopatógenos em sementes. Made available in DSpace on 2018-04-04T00:36:46Z (GMT). No. of bitstreams: 1 VariabilidadedanosedeteccaodeStenocarpellamaydiseStenocarpellamacrosporaemsementesdemilho..pdf: 3690062 bytes, checksum: c2eddb33c447240add64b92a9cfcb63a (MD5) Previous issue date: 2006-11-28 Tese (Doutorado em Fitopatologia) - Universidade Federal de Lavras, Lavras, MG. Orientador: José da Cruz Machado, Co-Orientador: Alan Carvalho Andrade.

Para o controle integrado de doenças, o tratamento químico das sementes de milho (Zea mays) torna-se indispensável, sendo a sua eficácia e influenciada por vários fatores, entre eles o tamanho das sementes. O objetivo do trabalho foi avaliar a relação do tamanho da semente e doses do fungicida thiabendazole no controle de Stenocarpella maydis, considerando-se diferentes tamanhos de sementes e doses do fungicida. As sementes classificadas nas peneiras 19, 22 e 24 foram inoculadas com S. maydis e tratadas com o fungicida thiabendazole em três doses, sendo avaliadas a incidência do patógeno e desempenho das sementes por meio de testes de qualidade fisiológica. Constatou-se com o tratamento fungicida, aumento na germinação, maior peso de plântulas e redução de tombamento provocado por S. maydis, havendo comportamento diferenciado do tratamento químico em relação ao tamanho de sementes e doses do produto. To prevent disease dissemination chemical treatment of maize (Zea mays) seeds becomes indispensable. However, the efficacy of this treatment may be influenced by a number of factors, such as seed size. Thus, the objective of this work was to evaluate the efficacy of the fungicide thiabendazole in treatment of maize seed to control S. maydis, taking into account different seed sizes. The seeds classified according to sieves 19, 22 and 24 were inoculated with S. maydis and treated with the fungicide thiabendazole in three doses. To evaluate the effect of seed treatment, test of germination on paper roll substrate, health test, (blotter method), and the dry weight of seedlings were considered. Seed treatment with fungicide provided conditions for increased germination and weight of seedlings, and reduced damping-off, caused by S. maydis. The efficacy of the fungicide seed treatment varied according to seed size and doses of the fungicide used.