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As a result of more restrictive legislation regarding the use of pesticides, over the last two decades, the demand for new soil fumigants has increased. These compounds can come from a variety of sources, including organic residues. In this study, we identified in the cassava wastewater volatiloma a molecule with potential to be developed as a new fumigant nematicide. Cassava wastewater (20 mL) releases volatile compounds toxic to Meloidogyne javanica second-stage juveniles (J2), causing J2 mortality up to 89%. Through gas chromatography-mass spectrometry, eight compounds were identified in the cassava wastewater volatiloma, with alcohols being the major class. The compounds ethyl butanoate and butyric acid identified in cassava wastewater volatiloma were selected for in vitro nematicidal activities and substrate fumigation tests. The lethal concentration predicted LC50-48 hr values (effective doses to kill 50% of M. javanica J2 population after 48 h of exposure) were 172.6 µg ∙ mL - 1 and 301.2 µg ∙ mL - 1 for butyric acid and ethyl butanoate, respectively. In a pot assay, the application of butyric acid and ethyl butanoate as a soil fumigant, at a dose of 0.5 mL ∙ kg - 1 substrate, significantly (P < 0.05) decreased M. javanica infectivity and reproduction compared to the negative control (water). However, ethyl butanoate proved to be a more efficient soil fumigant (P < 0.05) than butyric acid, as its application reduced the number of galls and eggs to the level of the commercial fumigant Dazomet. This study is the first to demonstrate the potential of ethyl butanoate as a soil fumigant against M. javanica.

RESUMEN

In this study, the full genome sequence of Bacillus velezensis strain UFLA258, a biological control agent of plant pathogens was obtained, assembled, and annotated. With a comparative genomics approach, in silico analyses of all complete genomes of B. velezensis and closely related species available in the database were performed. The genome of B. velezensis UFLA258 consisted of a single circular chromosome of 3.95 Mb in length, with a mean GC content of 46.69%. It contained 3,949 genes encoding proteins and 27 RNA genes. Analyses based on Average Nucleotide Identity and Digital DNA-DNA Hybridization and a phylogeny with complete sequences of the rpoB gene confirmed that 19 strains deposited in the database as Bacillus amyloliquefaciens were in fact B. velezensis. In total, 115 genomes were analyzed and taxonomically classified as follows: 105 were B. velezensis, 9 were B. amyloliquefaciens, and 1 was Bacillus siamensis. Although these species are phylogenetically close, the combined analyses of several genomic characteristics, such as the presence of biosynthetic genes encoding secondary metabolites, CRISPr/Cas arrays, Average Nucleotide Identity and Digital DNA-DNA Hybridization, and other information on the strains, including isolation source, allowed their unequivocal classification. This genomic analysis expands our knowledge about the closely related species, B. velezensis, B. amyloliquefaciens, and B. siamensis, with emphasis on their taxonomical status.

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Since volatiles do not leave residues in food and cause low toxicity to humans, they can act as a skeleton for new nematicidal molecules, once they have demonstrated activity against phytonematodes. The aim of this study was to evaluate the effect of volatile organic compounds (VOCs) released by wood-associated fungi on controling second-stage juveniles (J2) of Meloidogyne incognita. All 28 wood-associated fungi isolates produced VOCs which caused in excess of 76 % immobility in exposed J2 of M. incognita. The fungus isolate VOCs also caused significant mortality compared to control when J2 were exposed to them. After 6 h of exposure, the fungus species Epicoccum nigrum and Schizophyllum commune produced VOCs that immobilized exposed J2 compounds, in relation to the control. When J2 were inoculated into tomato under greenhouse conditions, the M. incognita infectivity and reproduction were reduced by exposure to the VOCs. Volatiles produced by most of the fungi isolates and analyzed by gas chromatograph coupled to a mass spectrometer (CG-MS), included alcohols, esters, terpenes, and ketones. Certain compounds appeared only in S. commune or in E. nigrum suggesting their involvement in the high level of damage caused to nematode reproduction on tomato. Volatiles emitted by wood fungi demonstrated another mode of action of these microorganisms in nature.

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Since volatiles do not leave residues in food and cause low toxicity to humans, they can act as a skeleton for new nematicidal molecules, once they have demonstrated activity against phytonematodes. The aim of this study was to evaluate the effect of volatile organic compounds (VOCs) released by wood-associated fungi on controling second-stage juveniles (J2) of Meloidogyne incognita. All 28 wood-associated fungi isolates produced VOCs which caused in excess of 76 % immobility in exposed J2 of M. incognita. The fungus isolate VOCs also caused significant mortality compared to control when J2 were exposed to them. After 6 h of exposure, the fungus species Epicoccum nigrum and Schizophyllum commune produced VOCs that immobilized exposed J2 compounds, in relation to the control. When J2 were inoculated into tomato under greenhouse conditions, the M. incognita infectivity and reproduction were reduced by exposure to the VOCs. Volatiles produced by most of the fungi isolates and analyzed by gas chromatograph coupled to a mass spectrometer (CG-MS), included alcohols, esters, terpenes, and ketones. Certain compounds appeared only in S. commune or in E. nigrum suggesting their involvement in the high level of damage caused to nematode reproduction on tomato. Volatiles emitted by wood fungi demonstrated another mode of action of these microorganisms in nature.(AU)

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Meloidogyne exigua is a parasitic nematode of plants that causes great losses to coffee farmers. In an effort to develop parasitic controls, 154 chalcones were synthesized and screened for activity against this nematode. The best results were obtained with (2E)-1-(4'-nitrophenyl)-3-(2,4,5-trimethoxyphenyl)prop-2-en-1-one (6) with a 50% lethal concentration (LC50) of 171 µg/ml against M. exigua second-stage juveniles, in comparison to the commercially-available nematicide carbofuran which had an LC50 of 260 µg/ml under the same conditions. When coffee plants were used, 6 reduced the nematode population to ~50% of that observed in control plants. To investigate the mechanism of action of 6, an in silico study was carried out, which indicated that 6 may act against M. exigua through inhibition of a putative caffeic acid 3-O-methyltransferase homodimer, the amino acid sequence of which was determined by examining the genome of Meloidogyne incognita.

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A infectividade e a reprodução de Meloidogyne javanica e de Heterodera glycines, em cultivar suscetível e resistente de soja, diferiram, de acordo com a temperatura do solo. A 28ºC o número de massas de ovos, total de ovos, de fêmeas e a porcentagem de sucesso de parasitismo dos juvenis do segundo estádio (J2) em cada cultivar (suscetível e resistentes) de soja infestada com M. javanica foram, significativamente maiores, comparados com as demais temperaturas (32º C, 26º C, 24º C e 20º C). Entretanto, a soma de juvenis do terceiro e quarto estádios (J3 e J4) foi maior em todas as cultivares, quando a temperatura submetida às plantas inoculadas com M. javanica foi de 20º C comparada com as demais temperaturas. A capacidade reprodutiva da fêmea (ovos por fêmea) de M. javanica foi sempre maior na cultivar suscetível em cada temperatura testada, porém, significativamente maior na cultivar suscetível colocada a 32º C comparada com as demais temperaturas. Nas relações H. glycines e soja, as temperaturas testadas proporcionaram valores semelhantes para a população total (fêmeas e cistos) na cultivar resistente. Na cultivar suscetível, a temperatura de 28º C aumentou significativamente o número de fêmeas, total de fêmeas e cistos e a porcentagem de parasitismo de J2 de H. glycines, comparada com todas as demais temperaturas.

The infectivity and reproduction of Meloidogyne javanica and Heterodera glycines in susceptible and resistant soybean cultivars differs according to soil temperatures. At 28ºC the egg-mass number, total eggs, and females, and percentage of parasitism success of second stage juveniles (J2) in each cultivar (susceptible and resistants) of infested soybean with M. javanica were significantly greater when compared to other temperatures (32º C, 26º C, 24º C and 20º C). However, the sum of third and fourth juvenile stages (J3 and J4) was greater in all cultivars when M. javanica inoculated plants were submitted to 20ºC. The female reproductive capacity (eggs per female) of M. javanica was always greater on susceptible cultivars in each tested temperature, but significantly greater in the susceptible cultivars submitted to 32ºC as compared to other temperatures. On the interaction between H. glycines and resistant soybean cultivars, the different temperatures resulted in similar values for the total population (females + cysts no.) on the resistant cultivars. On susceptible cultivars 28º C increased significantly the number of females, total females + cysts and the percentage of J2 parasitism of H. glycines as compared to all other temperatures.

RESUMEN

Com vista ao estudo de atração e penetração de Meloidogyne javanica (Treub) Chitwood e Heterodera glycines (Ichinoe) em soja (Glycine max L.), desenvolveu-se uma técnica empregando-se segmento de raiz com 2cm de comprimento. Nos segmentos de raiz de soja infectados, observou-se que a penetração de juvenis de segundo estádio (J2) de M. javanica ocorre pela coifa seguida de migração entre os feixes vasculares do cilindro central. Juvenis de H. glycines penetraram, aproximadamente, 15mm da coifa. A região seccionada da raiz de soja atraiu três vezes mais J2 de M. javanica do que a região da coifa, mas esta não foi tão atrativa para J2 de H. glycines. A obstrução conjunta da coifa e do local seccionado reduziu (83 por cento) a penetração de J2, tanto de M. javanica quanto de H. glycines. Quando apenas um desses locais foi obstruído, a outra extremidade livre compensou o processo atrativo. Portanto, as substâncias atrativas são liberadas por essas extremidades. A penetração de J2 de M. javanica foi maior no segmento de raiz quando comparada com a plântula intacta de soja. Entretanto, os J2 de H. glycines penetraram menos em segmentos de raiz e em plântulas sem folhas, quando comparados com plântulas intactas e com as seccionadas no colo. Portanto, na cultivar de soja "Embrapa 20", a atração e os locais de penetração de J2 de H. glycines e M. javanica são diferenciados. Esta técnica poderá ser útil nos estudos de atração e penetração de outros nematoides endoparasitas.

To study the attraction and penetration of Meloidogyne javanica (Treub) Chitwood and Heterodera glycines (Ichinoe) in soybean (Glycine max L.), a technique using 2-cm long root segments was developed. In infected soybean root segments penetration of second stage juveniles (J2) of M. javanica occured through the root cap following migration between the vascular bundles of the central cylinder. Juveniles of H. glycines penetrated about 15mm from the root cap. The cut region of the soybean root attracted three times more J2 of M. javanica when compared to the root cap, but was not as attractive to H. glycines J2. Sealing both root cap and cut region reduced (83 percent) the J2 penetration of M. javanica and H. glycines. When only one of these penetration sites was sealed the open site compensated attractiveness. These results show that the attractive substances are released by these two penetration sites. The M. javanica J2 penetration was higher in root segments when compared to whole soybean plants. However, H. glycines J2 penetrated less in root segments and in roots of seedlings without leaves when compared to roots of whole soybean plants and roots of plants without the aerial part. In short, attraction and infection sites of H. glycines and M. javanica J2s in soybean cultivar "Embrapa 20" are different and this technique may be useful in studies of attraction and penetration of other endoparasitic nematodes.

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O fracionamento do extrato metanólico das folhas de Gochnatia barrosii Cabrera (Asteraceae) resultou no isolamento do flavonol glicosídico trans-tilirosídeo [kaempferol 3-O- -D-(6''-O-E-p-cumaroil)-glicopiranosídeo], que nunca tinha sido identificado na referida espécie vegetal. Em teste realizado in vitro, observou-se que tal substância a 500 μg/mL, não tem efeito sobre a mortalidade de juvenis do segundo estágio do nematóide Meloidogyne exigua Goeldi.

Fractionation of the methanolic extract from Gochnatia barrosii Cabrera (Asteraceae) leaves resulted in the isolation of the flavonol glycoside trans-tiliroside [kaempferol 3-O- -D-(6''-O-E-p-coumaroyl)-glucopyranoside], which had never been found in such plant species. Such substance at 500 μg/mL caused no in vitro effect on the mortality of second-stage juveniles of the nematode Meloidogyne exigua Goeldi.

RESUMEN

The results of studies about interactions between microorganisms involving at least one plant pathogen are of interest to the areas of ethiology and control in Plant Pathology. Various aspects of these interactions have been studied over the years but the toxicity of volatile organic compounds (VOCs) has been emphasized only recently, developing techniques and procedures, and producing additional knowledge to those already obtained with water-soluble substances. This new facet of these interactions based on VOCs is discussed in this review involving mainly fungi, bacteria and nematodes pathogenic to plants. Also discussed is the role of VOCs produced by microorganisms, especially fungi and bacteria, in soil fungistasis and the effect of VOCs on fungal agents used in the control of plant parasitic nematodes. The evidence of VOCs broadens the research studies about these interactions. However, the scarcity of the research results in this area show up gaps which need to be filled and some research proposals are discussed. The present and future accumulated VOC knowledge will perhaps be beneficial to farmers, especially aspects related to increasing soil suppressiveness to plant disease and to the finding of analog molecules of VOCs highly effective against plant pathogens.

Os resultados dos estudos sobre interações entre microorganismos quando envolve, no mínimo, um patógeno de plantas são de interesse às áreas de etiologia e controle de fitodoenças. Vários aspectos dessas interações têm sido, por muitos anos, estudados, porém, a toxicidade de compostos orgânicos voláteis (COV) tem sido enfatizada apenas recentemente, desenvolvendo-se técnicas e rotinas, além de produzir conhecimentos adicionais àqueles já obtidos por meio de pesquisas com substâncias solúveis em água. Essa nova faceta dessas interações, envolvendo COVs, é discutida nesta revisão, enfatizando, principalmente, o antagonismo a fungos, bactérias e nematóides patogênicos de plantas. Também é discutido o papel dos COVs produzidos por microorganismos especialmente, fungos e bactérias envolvidos na fungistase do solo, bem como o efeito dos COVs nos agentes fúngicos usados no controle de fitonematóides. A evidência comprovada dos COVs nessas interações amplia esse campo de pesquisa. Entretanto, a escassez de pesquisas nessa área afloram lacunas no conhecimento científico que precisam ser preenchidas e, desta forma, algumas propostas de pesquisas são discutidas. O conhecimento científico presentemente acumulado nessa área de pesquisa, juntamente com aquele a ser produzido, serão, talvez, úteis aos produtores rurais, especialmente nos aspectos envolvendo a constatação de supressividade de solo a fitodoenças e aos testes com moléculas análogas dos COVs altamente tóxicos a patógenos de plantas.

RESUMEN

As nematodes cause great damage to Brazilian coffee production, effective methods to control these parasites are necessary. In a previous work Allium cepa L., Cajanus cajan (L.) Mill., Crotalaria juncea L., Ficus elastica Roxb., Ruta graveolens L., Stylosanthes guianensis Aubl., Leucaena leucocephala (Lam.) Dewit., Brachiaria decumbens Stapf., Catharanthus roseus G. Don, Tagetes minuta L., Ricinus communis L. and Coffea arabica L. produced active substances against Meloidogyne exigua Goeldi, a nematode widely disseminated through Brazilian coffee fields. Thus, aqueous extracts of such plants, collected in a different season from that of the previous work, as well as crude metabolites produced in liquid medium by Fusarium moniliforme Shelden and Cylindrocarpon magnusianum (Sacc.) Woll., were submitted to in vitro assays with M. exigua second-stage juveniles (J2). All plants and fungi produced active substances against J2. Therefore, their metabolites were applied to six-month-old coffee plants inoculated with M. exigua. After 90 days in a greenhouse, those samples obtained from A. cepa, L. leucocephala, R. graveolens and F. moniliforme inhibited the production of galls and eggs by M. exigua, demonstrating potential to control such parasite.

Os nematóides acarretam grandes perdas aos produtores brasileiros de café, sendo necessário o desenvolvimento de métodos eficientes para o seu controle. Em trabalho anterior, Allium cepa L., Cajanus cajan (L.) Mill., Crotalaria juncea L., Ficus elastica Roxb., Ruta graveolens L., Stylosanthes guianensis Aubl., Leucaena leucocephala (Lam.) Dewit., Brachiaria decumbens Stapf., Catharanthus roseus G. Don, Tagetes minuta L., Ricinus communis L. e Coffea arabica L. produziram substâncias ativas contra o nematóide Meloidogyne exigua Goeldi, que é amplamente disseminado pelos cafezais brasileiros. Dando continuidade a esse trabalho, extratos aquosos das plantas mencionadas, coletadas em época diferente daquela mencionada no trabalho anterior, bem como metabólitos na forma bruta de Fusarium moniliforme Shelden e Cylindrocarpon magnusianum (Sacc.) Woll., produzidos em meio de cultura líquido, foram submetidos a testes in vitro com juvenis do segundo estádio (J2) de M. exigua. Todas as plantas e fungos produziram substâncias ativas contra J2. Portanto, seus metabólitos foram aplicados em mudas de café de seis meses de idade, inoculadas com M. exigua. Após 90 dias em casa de vegetação, verificou-se que os materiais oriundos de A. cepa, L. leucocephala, R. graveolens e F. moniliforme inibiram a produção de ovos e de galhas por M. exigua, demonstrando potencial para o controle de tal parasita.

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A possibilidade de manejo de Meloidogyne exigua Goeldi, 1887, pela ativação de mecanismos de defesa no cafeeiro representa uma alternativa potencialmente útil no manejo desse patógeno. Com este trabalho, objetivou-se avaliar a eclosão e mortalidade de juvenis do segundo estádio (J2) de M. exigua na presença de produtos indutores de resistência e avaliar o efeito do acibenzolar-S-metil (ASM, Bion®) na indução de resistência do cafeeiro (Coffea arabica L.) 'Catuaí- 144' contra M. exigua. A eclosão e mortalidade do J2 foram avaliadas no ASM e ácido salicílico (AS) nas dosagens de 0,2; 0,35 e 0,5 g. i. a./L; e no fosfito de potássio (Hortifós® PK) e silicato de potássio (Supa-potássio®) nas dosagens 5,0; 7,5 e 10,0 mL/L, empregando água e aldicarbe como testemunhas. No segundo ensaio o ASM (0,2 g i.a./L) foi aplicado na quantidade de 125 mL por planta de 'Catuaí-144' com um ano de idade, via pulverização foliar e diretamente ao solo aos 7 dias antes da inoculação e aos 2 e 7 dias após a inoculação de aproximadamente 7000 ovos de M. exigua/planta. Foram utilizadas 8 plantas/tratamento/bloco, totalizando 6 tratamentos (3 épocas de aplicação do ASM), testemunhas absoluta e inoculada, em 4 blocos. Aos 90 dias da inoculação, foi feita a avaliação da população final (número de ovos e juvenis de M. exigua), número de galhas, fator de reprodução (população final/população inicial) e peso da matéria fresca da raiz. A dosagem dos produtos não influenciou a eclosão e mortalidade dos J2 de M. exigua. Menor eclosão dos J2 de M. exigua ocorreu igualmente no Supa-potássio® e ácido salicílico, enquanto que a eclosão no ASM e na água foi igual (P < 0,05) e significativamente menor que a eclosão dos J2 no Hortifós® PK. Maior mortalidade dos J2 ocorreu igualmente (P < 0,05) no ácido salicílico (AS) e no aldicarbe (500 ppm). Os demais produtos causaram mortalidade igual e inferior àquela observada na água. Aos 90 dias da inoculação de M. exigua nas mudas, não...

The parasitism of coffee roots by Meloidogyne exigua Goeldi, 1887, widespread nematode in the main producing regions, can provoke a series of modifications in the normal development of the plant. The induction of resistance for the activation of existing latent mechanisms of defense in plants represents a potentially useful alternative for the management of M. exigua in coffee plantations. In a first assay, the present work aimed at evaluating hatching and mortality of second stage juveniles (J2) of M. exigua in solutions of acibenzolar-S-metil (ASM, Bion®) and salicylic acid (SA) at the dosages of 0.2, 0.35 and 0.5 g. of a. i./L and Supa-potássio® (potassium silicate), Hortifós® PK (potassium phosphite) at the dosages 5.0; 7.5 and 10.0 mL/L, using water and aldicarb as control. The effect of ASM (0.2 g of a.i./L) was also evaluated by foliar and drench applications at 7 days before inoculation and at 2 and 7 days after inoculation of 7000 eggs of M. exigua/plant. The hatching of M. exigua J2 was inhibited in potassium silicate and in salicylic acid solutions independent of the used dosage, while J2 hatching in ASM and water was equal and significantly lower than J2 hatching verified in potassium phosphite. The mortality of J2 in SA was next to 100 percent and equal to the observed in aldicarb (500 ppm). The remaining products caused the same mortality of J2, however lesser than the observed in water. At ninety days of the inoculation of M. exigua on seedlings, the final population (number of eggs + J2), number of galls, reproduction factor (final population/initial population) and fresh weight of roots of plants treated with ASM did not differed significantly from control (inoculated plants).

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Root-knot nematodes of the genus Meloidogyne are important pathogens affecting vegetable crop production in Brazil and worldwide. The pepper species Capsicum annuum includes both hot and sweet peppers; very little emphasis has been placed on breeding sweet peppers for nematode resistance. We report on the inheritance of resistance to Meloidogyne incognita (Kofoid & White) Chitwood race 2 in the hot pepper cultivar Carolina Cayenne. The hot pepper cv. Carolina Cayenne was used as seed parent and the sweet pepper cv. Agronômico-8 was used as pollen parent to obtain the F1 and F2 generations and the backcross generations BC11 and BC12. The plants were inoculated with M. incognita race 2 at a rate of 60 eggs/ml of substrate and, after a suitable incubation period, the numbers of root galls and egg masses per root system were evaluated on each plant. Broad- (0.77 and 0.72) and narrow-sense (0.77 and 0.63) heritability estimates were high for both root galls and egg masses, respectively. The mean degree of dominance was estimated as 0.29 and 0.25 for numbers of galls and egg masses, respectively; these estimates were not significantly different from 0, indicating a predominantly additive gene action. The results were consistent with a hypothesis of monogenic resistance in Carolina Cayenne.

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RESUMEN

Root-knot nematodes of the genus Meloidogyne are important pathogens affecting vegetable crop production in Brazil and worldwide. The pepper species Capsicum annuum includes both hot and sweet peppers; very little emphasis has been placed on breeding sweet peppers for nematode resistance. We report on the inheritance of resistance to Meloidogyne incognita (Kofoid & White) Chitwood race 2 in the hot pepper cultivar Carolina Cayenne. The hot pepper cv. Carolina Cayenne was used as seed parent and the sweet pepper cv. Agronômico-8 was used as pollen parent to obtain the F(1) and F(2) generations and the backcross generations BC(11) and BC(12). The plants were inoculated with M. incognita race 2 at a rate of 60 eggs/ml of substrate and, after a suitable incubation period, the numbers of root galls and egg masses per root system were evaluated on each plant. Broad- (0.77 and 0.72) and narrow-sense (0.77 and 0.63) heritability estimates were high for both root galls and egg masses, respectively. The mean degree of dominance was estimated as 0.29 and 0.25 for numbers of galls and egg masses, respectively; these estimates were not significantly different from 0, indicating a predominantly additive gene action. The results were consistent with a hypothesis of monogenic resistance in Carolina Cayenne.

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