José da Cruz Machado

Biological control of mycotoxin-producing molds

Mycotoxins are produced by the secondary metabolism of many fungi and can be found in almost 25% of the worlds agricultural commodities. These compounds are toxic to humans, animals, and plants and therefore, efforts should be made to avoid mycotoxin contamination in food and feed. Besides, up to 25% of all harvested fruits and vegetables are lost due to storage molds and/or mycotoxin contamination and many methods have been applied to mitigate these issues, but most of them rely on the use of fungicides. Although chemicals are often the first defensive line against mycotoxigenic fungi, the indiscriminate use of fungicides are awakening the public perception due to their noxious effects on the environment and human/animal health. Thus, there is an increasing public pressure for a safer and eco-friendly alternative to control these organisms. In this background, biological control using microbial antagonists such as bacteria, fungi and yeasts have been shown to be a feasible substitute to reduce the use of chemical compounds. Despite of the positive findings using the biocontrol agents only a few products have been registered and are commercially available to control mycotoxin-producing fungi. This review brings about the up-to-date biological control strategies to prevent or reduce harvested commodity damages caused by storage fungi and the contamination of food and feed by mycotoxins.

Controle da germinação de sementes de soja em testes de sanidade pelo uso da restrição hídrica

The inhibition of seed germination in health testing is important because germinated seeds may cause contamination and make microscopic assessment difficult. To inhibit seed germination of some dicotyledons species the use of 2,4-D is commonly recommended and for monocotyledons the use of deep freezing is used. In both cases, there are serious inconveniences for several reasons. In the present study the use of the water restriction technique was investigated as mean to inhibit soybean seeds in substitution to 2,4-D in the blotter test. Manitol, NaCl and KCl were the osmotic compounds tested. The effects of the water restriction were also evaluated on the occurrence of the fungi associated with seeds. Water restriction in the range of -0,6 to -1,0 MPa was effective in the inhibition of the seed germination without affecting the occurrence of the fungi in the samples used.

Uso da restrição hídrica na inoculação de fungos em sementes de algodoeiro (Gossypium hirsutum)

Availability of seeds infected by fungi or other transmissible pathogens by this way is extremely important in seed pathology for several purposes. The present research was conducted to evaluate the use of the water restriction technique in the inoculation of cotton seeds in order to obtain seeds infected by Colletotrichum gossypii, Colletotrichum gossypii var. cephalosporioides, Botryodiplodia theobromae and Fusarium oxysporum f. sp. vasinfectum. The premise was based on the fact that seed infection is better ensured and controlled by keeping the seeds in contact with the developing fungal colonies on agar media containing mannitol at osmotic potentials that would arrest seed germination for a longer period of time. In this study mannitol was tested at the potentials of 0, -0.4, -0.6, -0.8 e -1 MPa. Seeds on the fungal colonies were placed in a growth room at 25oC, with a photoperiod of 12 hours, where they remained for different periods of time, according to the water potential used. After each incubation period seeds were taken out of the media and dried out on paper towels in a lab environment and submitted to the germination, emergence and health tests. Incubation of seeds at water potential ranging from -0.8 to -1 MPa, provided conditions to impede seeds from germinating for a period of up to 4 and 5 days, respectively, and then presenting a greater level of infection and higher percentages of diseased seedlings.

Inoculação de Fusarium oxysporum f. sp. phaseoli em sementes de feijoeiro através de restrição hídrica

Em diversos estudos sobre a associacao de Fusarium oxysporum f. sp. phaseoli com sementes de feijoeiro, ha necessidade de obtencao de sementes infectadas, inclusive com graus de incidencia diferenciados. Assim, objetivou-se com este trabalho testar a metodologia de inoculacao de Fusariumoxysporum f. sp. phaseoli em sementes de feijoeiro utilizando meio batata-sacarose-agar (BSA) adicionado com solutos que proporcionam restricao hidrica ao meio, permitindo o crescimento do fungo e inibindo a germinacao das sementes. Foram utilizados os solutos sacarose, cloreto de potassio e manitol, adicionados ao meio BSA para obter os potenciais osmoticos de 0,8, -1,0, -1,2 MPa, e diferentes tempos de exposicao das sementes ao fungo (36, 72, 108 e 144 h). Para avaliar os efeitos da colonizacao fungica nas sementes, utilizaram-se testes de germinacao e sanidade (blotter-test). O crescimento micelial do fungo foi avaliado in vitro, nao havendo inibicao em nenhum dos potenciais hidricos e solutos utilizados, observando-se maior crescimento nos potenciais osmoticos de 0,8 e -1,0 MPa. Nos maiores periodos de tempo, 108 e 144 h, o crescimento micelial foi maior, afetando o desempenho das sementes. Dos solutos utilizados, o KCl proporcionou a maior incidencia media (64%) de sementes com Fusariumoxysporum f. sp. phaseoli. Quando as sementes permaneceram por 144 h sobre o mesmo soluto, a incidencia foi de 70%.

Use of semi-seletive media for detection of Sclerotinia sclerotiorum on bean and soybean seeds

O objetivo deste trabalho foi avaliar a possibilidade do uso do azul de bromofenol como um indicador para determinar a presenca of Sclerotinia sclerotiorum em sementes de feijao (Phaseolus vulgaris) e soja (Glycine max), quando da incubacao dessas em meios agar e papel de filtro. As sementes foram inoculadas artificialmente com quatro isolados de S. sclerotiorum, plaqueadas em meio agar, denominado Neon, e em meio Neon modificado, sendo incubadas sob 14 e 20 oC por sete dias no escuro. Metade das sementes inoculadas foi desinfestada superficialmente, previamente ao plaqueamento nos meios em teste. As sementes que apresentaram mudanca de coloracao do meio ao seu redor, de azul para amarelo com formacao de micelio tipico ou que produziram esclerodios, em alguns casos, foram consideradas infetadas ou contaminadas por S. sclerotiorum. Para a maioria dos isolados testados, nao houve diferencas significativas entre as duas temperaturas de incubacao comparadas (P<0.05) nos diferentes meios. De acordo com os resultados obtidos neste estudo, o emprego do meio Neon em condicoes de incubacao as temperaturas de 14 ou 20 oC, pode ser considerado adequado por proporcionar uma deteccao rapida e segura de S. sclerotiorum em sementes de feijao e soja naturalmente infetadas ou contaminadas

Performance of common bean seeds infected by the fungus Sclerotinia sclerotiorum

The fungus Sclerotinia sclerotiorum, causal agent of white mold disease on several economically important crops, such as bean, soybean, and cotton, is commonly disseminated through seeds and can cause high losses on their quality and in productivity of these species. The aim of this study was assessing the effects of different initial inoculums potentials of this fungus on common bean seeds using two different strains of the fungus and two genotypes of common beans (Perola and Ouro Negro) artificially inoculated. Seeds were sown on soil and the cultivation was performed under controlled environmental conditions favorable to development of the white mold disease. Variables assessed were: germination; seed health; emergence speed index; initial and final seedling number; and dry weight of aerial parts and roots. It was found that with the gradual increase in inoculum potential in the seeds also occurred gradual reduction in the values of: germination; emergence speed index; seedling stand; and length and dry mass of aerial parts and roots. These results show the importance of initial inoculum potential of S. sclerotiorum present in common bean seeds, as much in disseminating the pathogen as on direct damages caused in field by reducing productive potential of the emerged plants.

Incorporação de microrganismos, aminoácidos, micronutrientes e reguladores de crescimento em sementes de alface pela técnica de peliculização

Seed coating is a technology that allows incorporation of additive to the seeds, without change its size or form. The coating technique is a strategy that guarantees high seedling emergence in the field, gain in productivity and better aggregation of agricultural inputs to the seeds. Thus, this study had the objective of evaluating the effect of the incorporation of antagonistic microorganisms, mycorrhiza, amino acids, micronutrients and growth regulators through the seed coating technique on the quality of lettuce seeds. The seeds were coated with two types of polymer that were applied together with antagonistic microorganisms (Trichoderma viride, Trichoderma polysporhum, Trichoderma stromaticum, Beauveria bassiana, Metarhizium anisopliae), mycorrhiza, amino acids, micronutrients and growth regulators according to the methodology proposed by the manufacturer. The germination percentage and emergence, the emergence speed index, the number of plants, the dry mass of aerial and root parts and the height of the seedlings were evaluated. Microbiolization of the seeds with the mixture of microorganisms Trichoderma viride, T. polysporhum, T. stromaticum, Beauveria bassiana, Metarhizium anisopliae and mycorrhiza negatively affected the quality of the lettuce seeds. The inoculation of lettuce seeds with Trichoderma viride and growth regulator increased the percentage and the seedling emergence speed index.

Transmissibility and effect of Acremonium strictum in maize seeds

Acremonium strictum is pathogenic to seeds, stalk and seedlings of maize. It is a fungus that can interfere in the vigor, stand and final productivity of plants. In addition, it can be disseminated and transmitted through seeds to the progeny, although the real proportions of that are not well known. This study reports the effects of A. strictum on maize seeds and seedlings quality (germination, vigor, healthness), some vegetative growth parameters and also sanity (infection and transmission rates) of young maize plants. The transmission rate of the fungus was evaluated using the water restriction technique. The fungus formulation was cultivated on malt extract agar medium amended with mannitol to a water potential of -1,4 MPa. The maize seeds germination was reduced by seeds exposition to A. strictum for periods longer than 72 hours. The potentials vigor and viability were not affected by A. strictum. Theincidence of this fungus reduced the vegetative growth of maize plants measured 28 days after sowing. The fungus originating from inoculated seeds was transmitted to young maize plants.

Variabilidade genética de isolados do complexo Colletotrichum associados a sementes de algodoeiro, por meio de técnicas moleculares e inoculação em plantas

Anthracnosis and ramulosis are important diseases of cotton (Gossypium hirsutum), caused by Colletotrichum gossypii and C. gossypii var. cephalosporioides, respectively; both fungi seed transmitted. Ramulosis, also known as witchs broom, is considered to be more devastating, causing severe losses in several regions of Brazil where cotton is cultivated in large scale. It is difficult to detect both fungi in seed samples by conventional methods since the morphological variability between isolates within the same species makes detection unreliable. The goal of the presente work was to investigate the variation of 10 isolates of both fungi using DNA/RAPD markers. Part of the isolates was classified as C. gossypii and part as C. gossypii var. cephalosporioides, using the pathogenic test. The results showed that the biochemical markers used were not able to distinguish the isolates of anthracnosis from the isolates of ramulosis. On the other hand, the genetic similarity between the isolates demonstrated by RAPD analysis, was 51,78%, indicating the potential of this technique.

Fungicidas aplicados via água de irrigação no controle do mofo-branco no feijoeiro e incidência do patógeno na semente

The effectiveness of four fungicides applied through irrigation water to control white mould of common beans (Phaseolus vulgaris) was evaluated in Vicosa, MG. The trial was installed on April in a naturally infested field with sclerotia of Sclerotinia sclerotiorum. The following fungicides and rates of active ingredient per hectare were tested: benomyl (1.0 kg), iprodione (0.75 kg), procymidone (0.5 kg), and fluazinam (0.5 l). Simulating sprinkler irrigation, fungicides were applied with watering-cans over the plants at 39 and 52 days after emergence (DAE) with water volume of 35,000 l/ha. At these two periods, fluazinam was also applied by a backpack sprayer (667 l/ha) or with watering-cans (35,000 l/ha) between the rows and near the soil surface. The last treatment was to verify the fluazinam effect on white mould control when applied on the soil surface. An untreated plot was also included. After harvest, seed transmission of the pathogen was evaluated. Fluazinam (applied either by backpack sprayer or through irrigation water onto plants), benomyl, and procymidone were the most efficient fungicides for white mould control. Among them, only procymidone did not provide yield higher than that of control. Fluazinam applied only on soil reduced white mould incidence and the amount of sclerotia produced. Yields varied from 1,406 kg/ha (control) to 2,054 kg/ha (fluazinam applied by backpack sprayer). There was no effect of treatments on the pathogen seed transmission, which varied from 0.25% (procymidone) to 1.08% (fluazinam applied only on the soil).