Susceptibilidad a Fusarium verticillioides, Gibberella zeae y Diplodia maydis en líneas e híbridos pre-comerciales de maíces templados y tropicales

Las podredumbres de espiga (causada por Fusarium verticillioides y Gibberella zeae) y tallo (causada por F.verticillioides) provocan importantes pérdidas de rendimiento y calidad del grano en maíz (Zea mays L.), por lo que es importante mantener actualizado el conocimiento sobre la variabilidad genotípica de la especie para la tolerancia a las mismas así como la posible relación entre la presencia de una y otra podredumbre. Con estos objetivos se evaluaron (i) 438 líneas y 84 híbridos precomerciales de origen templado para resistencia a ambas podredumbres, y (ii) 84 líneas de origen templado y tropical para resistencia a podredumbre de la espiga por Diplodia maydis. Ambos ensayos fueron conducidos en un DBCA en Fontezuela (33.9 ºS, 60.47º O), Argentina, durante 2009-2010. Los tratamientos empleados fueron inoculación artificial e infección natural. Se encontraron diferencias significativas entre ambas para incidencia (I: porcentaje de plantas afectadas en el surco), Severidad (S: porcentaje promedio de podredumbre de las espigas sobre el total de espigas afectadas) y S×I, tanto para híbridos como para líneas. Existió una correlación positiva entre el tratamiento control sin inocular y el inoculado para todas las enfermedades. Se detectó una pequeña influencia de los caracteres morfológicos de la espiga y el grano sobre la resistencia a podredumbre de espiga por F.verticillioides y D. maydis, respectivamente. No se encontró G. zeae en el control, por lo que no pudieron realizarse los análisis correspondientes para esta enfermedad. La asociación entre el tratamiento inoculado con fusariosis de espiga y el control para fusariosis de tallo, y viceversa, nunca fue consistente, por lo tanto puede emplearse un tratamiento como control del otro. Los materiales de origen tropical fueron más resistentes a D. maydis que los templados, aunque las diferencias entre grupos heteróticos no resultaron altamente significativas. Se determinó que la inoculación artificial empleando infección natural como control es una práctica recomendable en el mejoramiento de maíz por resistencia a enfermedades. La inoculación artificial puede generar mayor variación en las respuestas, permitiendo observar y comparar el comportamiento de distintos materiales frente a la infección de patógenos.

Relative populations and toxin production by Aspergillus flavus and Fusarium verticillioides in artificially inoculated corn at various stages of development under field conditions

The presence, development and production of mycotoxins by Aspergillus flavus and Fusarium verticillioides were studied in corn ears under field conditions after artificial contamination of corn silks. The planted area was divided into five treatments: T1, inoculated with A.flavus solution containing 1 x 10(8) spores, ears covered; T2, inoculated with F. verticillioides solution containing 1 X 10(8) spores, ears covered; T3, inoculated with E verticillioides plus A. flavus solution containing 1 x 10(8) spores of each, ears covered; T4, sprayed with sterile phosphate-buffered saline, ears covered; TS, non-sprayed silks, uncovered ears. Soil and air samples were also collected and analysed for the occurrence of fungi. Water activity, relative air humidity, rainfall and temperature were determined to assess the correlation between abiotic factors and the presence of fungi in the samples. Contamination with the inoculated fungus predominated in T1 and T2. In the other treatments, F. verticillioides was the most frequently isolated contaminant irrespective of treatment. Considering the production of mycotoxins, a positive relation between the production of fumonisins B-1 and B-2 and the frequency of F. verticillioides was statistically verified in all treatments. (C) 2007 Society of Chemical Industry.

Molecular characterization and fumonisin production by Fusarium verticillioides isolated from corn grains of different geographic origins in Brazil

Gibberella moniliformis is most commonly associated with maize worldwide and produces high levels of fumonisins, some of the most agriculturally important mycotoxins. Studies demonstrate that molecular methods can be helpful for a rapid identification of Fusarium species and their levels of toxin production. The purpose of this research was to apply molecular methods (AFLP, TEF-1 alpha partial gene sequencing and PCR based on MAT alleles) for the identification of Fusarium species isolated from Brazilian corn and to verify if real time RT-PCR technique based on FUM1 and FUM19 genes is appropriated to estimate fumonisins B(1) and B(2) production levels. Among the isolated strains, 96 were identified as Fusarium verricillioides, and four as other Fusarium species. Concordant phylogenies were obtained by AFLP and TEF-1 alpha sequencing, permitting the classification of the different species into distinct clades. Concerning MAT alleles, 70% of the F. verricillioides isolates carried the MAT-1 and 30% MAT-2. A significant correlation was observed between the expression of the genes and toxin production r=0.95 and r=0.79 (correlation of FUM1 with FB(1) and FB(2), respectively, P < 0.0001): r=0.93 and r =0.78 (correlation of FUM19 with FB(1) and FB(2). respectively, P < 0.0001). Molecular methods used in this study were found to be useful for the rapid identification of Fusarium species. The high and significant correlation between FUM1 and FUM19 expression and fumonisins production suggests that real time RT-PCR is suitable for studies considering the influence of abiotic and biotic factors on expression of these genes. This is the first report concerning the expression of fumonisin biosynthetic genes in Fusarium strains isolated from Brazilian agricultural commodity. (c) 2010 Elsevier B.V. All rights reserved.

Toxicokinetics and toxicological effects of single oral dose of fumonisin B1 containing Fusarium verticillioides culture material in weaned piglets

Toxicokinetics and the toxicological effects of culture material containing fumonisin B(1) (FB(1)) were studied in male weaned piglets by clinical, pathological, biochemical and sphingolipid analyses. The animals received a single oral dose of 5 mg FB(1)/kg of body weight. obtained from Fusarium verticillioides culture material. FB(1) was detected by H PLC in plasma collected at 1-h intervals up to 6 h and at 12-h intervals up to 96 h. FB(1) eliminated in feces and urine was quantified over a 96-h period and in liver samples collected 96 h post-intoxication. Blood samples were obtained at the beginning and end of the experiment to determine serum enzyme activity, total bilirubin, cholesterol, sphinganine (Sa), sphingosine (So) and the Sa/So ratio. FB(1) was detected in plasma between 30 min and 36 h after administration. The highest concentration of FB(1) was observed after 2 h, with a mean concentration of 282 mu g/ml. Only 0.93% of the total FB(1) was detected in urine between 75 min and 41 h after administration, the highest mean concentration (561 mu g/ml) was observed during the interval after 8 at 24 h. Approximately 76.5% of FB(1) was detected in feces eliminated between 8 and 84 h after administration, with the highest levels observed between 8 and 24 h. Considering the biochemical parameters, a significant increase only occurred in cholesterol, alkaline phosphatase and aspartate aminotransferase activities. In plasma and urine, the highest Sa and Sa/So ratios were obtained at 12 and 48 h, respectively. (C) 2010 Elsevier Ireland Ltd. All rights reserved.

Infecção por Fusarium graminearum e Fusarium verticillioides em sementes de milho

The previous knowledge of the infection process and pathogens behavior, for evaluating the physiological potential of maize seeds, is essential for decision making on the final destination of lots that can endanger sowing. This research was carried out in order to study the minimum period required for maize seeds contamination by Fusarium graminearum Schwabe and Fusarium verticillioides (Sacc.) Nirenberg, as well as these pathogens influence on seed germination and vigor, by using the cold test. Three maize seeds hybrids, kept in contact with the pathogens for different periods, were evaluated with and without surface disinfection. After determining the most suitable period, new samples were contaminated by F. graminearum and F. verticillioides, under different infection levels, and subjected to germination tests in sand. The cold test was conducted with healthy and contaminated seeds, at different periods, in a cold chamber. The contact of maize seeds with F. graminearum and F. verticillioides for 16 hours was enough to cause infection. F. graminearum and F. verticillioides did not affect the maize seeds germination, however, F. graminearum reduced the vigor of seeds lots.

Produção, imobilização e aplicação de lipase de Fusarium verticillioides

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

First report of Fusarium verticillioides causing stalk and root rot of sorghum in Spain

Podredumbre del tallo y la raíz del sorgo causada por Fusarium verticillioides en España

Seleção de estirpes de Trichoderma ssp. com atividade antagônica a Fusarium verticillioides produtores de fumonisina isolados de milho e de sorgo.

2016

Neuro- and immunotoxic effects of fumonisin B1 in cells

Fumonisin B1 (FB1) is a mycotoxin produced by the fungus Fusarium verticillioides, which commonly infects corn and other agricultural products. Fusarium species can also be found in moisture-damaged buildings, and therefore there may also be human exposure to Fusarium mycotoxins, including FB1. FB1 affects the metabolism of sphingolipids by inhibiting the enzyme ceramide synthase. It is neuro-, hepato- and nephrotoxic, and it is classified as possibly carcinogenic to humans. This study aimed to clarify the mechanisms behind FB1-induced neuro- and immunotoxicity. Four neural and glial cell lines of human, rat and mouse origin were exposed to graded doses of FB1 and the effects on the production of reactive oxygen species, lipid peroxidation, intracellular glutathione levels, cell viability and apoptosis were investigated. Furthermore, the effects of FB1, alone or together with lipopolysaccharide (LPS), on the mRNA and protein expression levels of different cytokines and chemokines were studied in human dendritic cells (DC). FB1 induced oxidative stress and cell death in all cell lines studied. Generally, the effects were only seen after prolonged exposure at 10 and 100 µM of FB1. Signs of apoptosis were also seen in all four cell lines. The sensitivities of the cell lines used in this study towards FB1 may be classified as human U-118MG glioblastoma > mouse GT1-7 hypothalamic > rat C6 glioblastoma > human SH-SY5Y neuroblastoma cells. When comparing cell lines of human origin, it can be concluded that glial cells seem to be more sensitive towards FB1 toxicity than those of neural origin. After exposure to FB1, significantly increased levels of the cytokine interferon-γ (IFNγ) were detected in human DC. This observation was further confirmed by FB1-induced levels of the chemokine CXCL9, which is known to be regulated by IFNγ. During co-exposure of DC to both LPS and FB1, significant inhibitions of the LPS-induced levels of the pro-inflammatory cytokines interleukin-6 (IL-6) and IL-1β, and their regulatory chemokines CCL3 and CCL5 were observed. FB1 can thus affect immune responses in DC, and therefore, it is rather likely that it also affects other types of cells participating in the immune defence system. When evaluating the toxicity potential of FB1, it is important to consider the effects on different cell types and cell-cell interactions. The results of this study represent new information, especially about the mechanisms behind FB1-induced oxidative stress, apoptosis and immunotoxicity, as well as the varying sensitivities of different cell types towards FB1.

Influence of macro- and micronutrient fertilization on fungal contamination and fumonisin production in corn grains

The aim of this study was to investigate the effects of nutrients (nitrogen, zinc and boron) on fungal growth and fumonisins production in corn samples obtained at the beginning of grain formation and at harvest. Three nitrogen doses were applied to the corn plants through soil in combination with three zinc doses and two boron doses during sowing. Mycological analysis of grains, using Dichloran Rose-Bengal Chloramphenicol Agar, collected at the beginning of formation demonstrated a fungal population predominantly of yeasts. Analysis of freshly harvested corn revealed a higher frequency of Penicillium spp. (72%) and F verticillioides (27%). High Performance Liquid Chromatography analysis revealed that 100% of grains were contaminated with fumonisins B, at levels ranging from 0.3 to 24.3 mg/kg and 93% contaminated with fumonisin B(2) at levels ranging from 0.05 to 5.42 mg/kg. Nitrogen (50 kg/ha) in combination with boron (0.5 kg/ha) resulted in an increased fumonisin B2 production. (c) 2007 Elsevier Ltd. All rights reserved.

Mycoflora and fumonisin contamination in Brazilian sorghum from sowing to harvest

BACKGROUND: The aim of this study was to characterise the mycoflora and the presence of fumonisin in sorghum grains, correlating the results with the environment and abiotic factors. RESULTS: Fifty samples (five collections of ten samples each) of sorghum were analysed. All samples were found to be contaminated with fungi, with higher frequencies of Cladosporium spp. (61.8%) and Helminthosporium spp. (33.4%). Fusarium verticillioides was isolated from 15.1% of the samples, with 38% of them being contaminated with fumonisin B(1) (FB(1)) at levels ranging from 50 to 368.78 ng g(-1). Regarding abiotic factors, temperature, water activity and rainfall showed a positive correlation with the frequency of F. verticillioides and FB(1) production. There was a significant positive correlation between relative air humidity and FB(1) production. The results obtained from sexual crosses between standard F mating tester strains and the isolated strains confirmed that the strains isolated were F. verticillioides. CONCLUSION: It can be concluded that the decrease in F. verticillioides and fumonisin contamination occurred owing to atypical climatic factors during the period of sorghum cultivation, when there was any occurrence of rain and the level of water activity of grains did not reach 0.58. (C) 2010 Society of Chemical Industry

Growth and enzymatic responses of phytopathogenic fungi to glucose in culture media and soil

The effect of inoculation of Aspergillus flavus, Fusarium verticillioides, and Penicillium sp. in Dystrophic Red Latosol (DRL) and Eutroferric Red Latosol (ERL) soils with or without glucose on the total carbohydrate content and the dehydrogenase and amylase activities was studied. The fungal growth and spore production in culture medium with and without glucose were also evaluated. A completely randomized design with factorial arrangement was used. The addition of glucose in the culture medium increased the growth rate of A. flavus and Penicillium sp. but not of F. verticillioides. The number of spores increased 1.2 for F. verticillioides and 8.2 times for A. flavus in the medium with glucose, but was reduced 3.5 times for Penicillium sp. The total carbohydrates contents reduced significantly according to first and second degree equations. The consumption of total carbohydrates by A. flavus and Penicillium sp. was higher than the control or soil inoculated with F. verticillioides. The addition of glucose to soils benefited the use of carbohydrates, probably due to the stimulation of fungal growth. Dehydrogenase activity increased between 1.5 to 1.8 times (p <0.05) in soils with glucose and inoculated with the fungi (except F. verticillioides), in relation to soil without glucose. Amylase activity increased 1.3 to 1.5 times due to the addition of glucose in the soil. Increased amylase activity was observed in the DRL soil with glucose and inoculated with A. flavus and Penicillium sp. when compared to control.

Leucoencefalomalácia em equídeos da região Leste de Mato Grosso

Background: Fumonisins produced by Fusarium verticillioides are among the most important medical mycotoxins known. The intake of concentrate based on corn and corn by-products contaminated with fumonisins can cause severe poisoning in horses. The injuries are observed mainly in the white matter of the brain, and the disease is known as Equine Leukoencephalomalacia (ELEM). This study aims to describe and discuss the epidemiological, clinical and diagnostic aspects of an outbreak of ELEM occurred in three farms in the municipalities of Canarana and Agua Boa, in the eastern region of Mato Grosso, Brazil.Materials, Methods & Results: The outbreak occurred between May and August 2010. The disease affected six horses and four mules of different ages and sex. Clinical examination was only possible in animals with chronic evolution of the disease. All the affected animals showed neurological clinical signs such as ataxia and recumbency, which progressed to death or sudden death. Histopathological analysis showed foci of necrosis that predominantly affected the white matter, and the presence of gitter cells. Degenerative lesions were observed in the liver of the animals. Mortality rate ranged from 12.5 to 71%, and lethality reached 100%. The cases were preceded by sudden drops in the weather temperature. Fumonisins levels of 6.6 ppm were detected in the feed of the animals.Discussion: The presumptive diagnosis of leukoencephalomalacia was consistent and based on clinical and epidemiological studies. However, the definitive diagnosis was based upon the histological features of the brain including the presence of extensive areas of malacia. Moreover, the animals were being fed with corn or corn by-products contaminated with fumonisins levels considered to be toxic to equids. The mortality and lethality rates are in agreement with outbreaks described in previous studies. The animals showed neurological signs as the predominant clinical manifestation, with gait ataxia followed by recumbency, prostration and death between 24 h and 29 days. Similarly to other reports, the disease was more frequent in adult animals, which succumbed in 24-48 h. Conversely, the evolution of the disease in young animals was of 10 to 29 days. Sudden death was more prevalent in the mules. Previous studies have shown a predominance of cerebral and brainstem lesions in horses, whereas in mules the clinical signs are related to brainstem lesions. Corn and corn by-products are commonly used as energy supplementation to horses in the southern of Mato Grosso state, but outbreaks of the disease are uncommon. This may be influenced by the prevalence of hot climate conditions, which does not favor the production of toxin by the fungus. The atypical low weather temperatures (9-11 degrees C) observed prior to the outbreak could have contributed to the mycotoxin production by F. verticillioides, which requires temperatures between 8-12 degrees C to produce toxins. However, the disease in the region may be underestimated, considering that the practice of necropsies is not common among field technicians, mainly in the occurrence of sporadic deaths among horses intended for work. Preventive measures include avoiding the use of corn and corn by-products for horses after periods of sudden drops in temperature in the region. Furthermore, clinical and epidemiological surveys and post-mortem and histopathological analyses are undoubtedly important for appropriate differential diagnosis, especially in equids with neurological signs.

3-Hydroxypropionic Acid as an Antibacterial Agent from Endophytic Fungi Diaporthe phaseolorum

Endophytic fungi are considered a rich source of active compounds resulting from their secondary metabolism. Fungi from marine environment grow in a habitat with unique conditions that can contribute to the activation of metabolic pathways of synthesis of different unknown molecules. The production of these compounds may support the adaptation and survival of the fungi in the marine ecosystem. Mangroves are ecosystems situated between land and sea. They are frequently found in tropical and subtropical areas and enclose approximately 18.1 million hectares of the planet. The great biodiversity found in these ecosystems shows the importance of researching them, including studies regarding new compounds derived from the endophytic fungi that inhabit these ecosystems. 3-hydroxypropionic acid (3-HPA) has been isolated from the mangrove endophytic fungus Diaporthe phaseolorum, which was obtained from branches of Laguncularia racemosa. The structure of this compound was elucidated by spectroscopic methods, mainly 1D and 2D NMR. In bioassays, 3-HPA showed antimicrobial activities against both Staphylococcus aureus and Salmonella typhi. The structure of this antibiotic was modified by the chemical reaction of Fischer-Speier esterification to evaluate the biologic activity of its chemical analog. The esterified product, 3-hydroxypropanoic ethyl ester, did not exhibit antibiotic activity, suggesting that the free carboxylic acid group is important to the pharmacological activity. The antibiotic-producing strain was identified with internal transcribed spacer sequence data. To the best of our knowledge, this is the first report of antibacterial activity by 3-HPA against the growth of medically important pathogens.

Interaction between toxigenic fungi and weevils in corn grain samples

This study investigated the ability of weevils to transmit Aspergillus flavus and Fusarium verticillioides fungal spores and the consequent production of mycotoxins. For this purpose, corn grain samples were stored in flasks connected to a hose to form a closed system (flasks A and B). Flasks A were inoculated with the following groups: group 1 (corn + weevil); group 2 (corn + A. flavus); group 3 (corn + A. flavus + weevil); group 4 (corn + F. verticillioides); group 5 (corn + E verticillioides + weevil), and group 6 (corn + A. flavus + E verticillioides + weevil). Flasks B contained sterile grains. The samples were incubated for 10, 20 and 30 days, posteriorly, weight, water activity, mycoflora, aflatoxins and fumonisins. The corn grain samples were also submitted to scanning electron microscopy. Our results showed that weevils could enhance corn grains contamination by these fungi, hence, could increase mycotoxins production. These findings demonstrate the importance of weevils as fungal vectors and the need for good manipulation and storage practices of grains. (C) 2012 Elsevier Ltd. All rights reserved.