Vicente Paulo Campos

Volatiles produced by interacting microorganisms potentially useful for the control of plant pathogens

Os resultados dos estudos sobre interacoes entre microorganismos quando envolve, no minimo, um patogeno de plantas sao de interesse as areas de etiologia e controle de fitodoencas. Varios aspectos dessas interacoes tem sido, por muitos anos, estudados, porem, a toxicidade de compostos orgânicos volateis (COV) tem sido enfatizada apenas recentemente, desenvolvendo-se tecnicas e rotinas, alem de produzir conhecimentos adicionais aqueles ja obtidos por meio de pesquisas com substâncias soluveis em agua. Essa nova faceta dessas interacoes, envolvendo COVs, e discutida nesta revisao, enfatizando, principalmente, o antagonismo a fungos, bacterias e nematoides patogenicos de plantas. Tambem e discutido o papel dos COVs produzidos por microorganismos especialmente, fungos e bacterias envolvidos na fungistase do solo, bem como o efeito dos COVs nos agentes fungicos usados no controle de fitonematoides. A evidencia comprovada dos COVs nessas interacoes amplia esse campo de pesquisa. Entretanto, a escassez de pesquisas nessa area afloram lacunas no conhecimento cientifico que precisam ser preenchidas e, desta forma, algumas propostas de pesquisas sao discutidas. O conhecimento cientifico presentemente acumulado nessa area de pesquisa, juntamente com aquele a ser produzido, serao, talvez, uteis aos produtores rurais, especialmente nos aspectos envolvendo a constatacao de supressividade de solo a fitodoencas e aos testes com moleculas analogas dos COVs altamente toxicos a patogenos de plantas.

Diversity of Meloidogyne exigua (Tylenchida: Meloidogynidae) populations from coffee and rubber tree

Isozymes (esterase and malate dehydrogenase), SCAR and RAPD-PCR were studied in 15 populations of three races of Meloidogyne exigua collected in coffee-producing areas in Brazil, Bolivia and Costa Rica and one population from rubber tree plantations in Brazil. This study revealed four esterase phenotypes (E1, E2, E2a, E3) and three malate dehydrogenase phenotypes (N1, N1a, N2) for M. exigua populations. The most common multi-enzyme phenotype was E2N1. The enzymatic phenotypes do not separate M. exigua races. Sixteen populations of M. exigua were tested in Multiplex PCR using SCAR primers ex-D15F/R that allowed the identification of all M. exigua populations. Phylogenetic analyses showed high intraspecific polymorphism (25.9-59.6%) for all M. exigua studied. However, all populations clustered together with 100% bootstrap support, thereby demonstrating the consistency of species identification. In general, no correlation was found between enzymatic profile, race and genetic polymorphism of the studied populations.

Reaction of coffee genotypes to different populations of Meloidogyne spp.: detection of a naturally virulent M. exigua population

The reaction of seven genotypes of Coffea arabica to 10 Meloidogyne spp. populations collected mainly from coffee plantations in Brazil and Costa Rica was evaluated under greenhouse conditions. The inoculum consisted of 10,000 eggs per plant. Evaluations were done 8 months after inoculations considering the root fresh weight, gall and egg mass indices, number of eggs per gram of root and reproduction factor (RF). The cultivars Obata IAC 1669-20, Sarchimor IAC 4361 and Tupi Amarelo IAC 5111 exhibited susceptibility to the four Brazilian M. exigua populations tested. However, cv. Tupi Vermelho IAC 1669-33 revealed resistance (RF value of 0.7) to the M. exigua population from Lavras, Minas Gerais State, Brazil. A population of M. exigua from Bom Jesus de Itabapoana, Rio de Janeiro State, Brazil, was highly virulent on cv. IAPAR 59 (RF= 165.7), bearing resistance gene Mex-1, and was also virulent on genotype Paraiso (H 419-5-4-5-2) (RF=396.2). A Meloidogyne sp. population on coffee from Garca, Sao Paulo State, Brazil, reproduced at low rates (RF ranging from 0.1 to 3.9) on all genotypes. All tested cultivars were susceptible to M. incognita and M. paranaensis. M. mayaguensis of guava from Parana State, Brazil, reproduced at low rates in all coffee genotypes; however, another population of coffee, from Costa Rica, was more aggressive and showed RF value that ranged from 0.8 to 12.4. Results of this study point for the first time to the ability of a naturally occurring M. exigua population to overcome the resistance conferred by the Mex-1 gene.

Influência de leguminosas no controle de fitonematóides no cultivo orgânico de alface americana e de repolho

A demanda por produtos orgânicos e crescente devido as restricoes ao uso dos agroquimicos sinteticos. Para verificar o efeito das plantas antagonistas no controle de fitonematoides na cultura da alface americana (Lactuca sativa) e do repolho (Brassica oleracea var. capitata), em cultivo orgânico, instalou-se um experimento na Universidade Federal de Lavras, no periodo de dezembro de 2001 a agosto de 2002. O delineamento experimental utilizado foi o de blocos casualizados em esquema fatorial 4 x 2, com tres repeticoes. O primeiro fator foi constituido pelas leguminosas mucuna-preta (Stizolobium aterrimum), feijao-de-porco (Canavalia ensiformis), crotalaria (Crotalaria juncea) em cultivo orgânico e por uma testemunha (vegetacao espontânea) em cultivo convencional. O segundo, pelas culturas de alface americana cultivar Raider e repolho, cultivar Kenzan. Os tratamentos foram constituidos por todos em cultivo orgânico e testemunha (vegetacao espontânea) em cultivo convencional. As populacoes de Meloidogyne spp. e de Helicothylenchus dihystera foram avaliadas empregando-se a tecnica de Jenkins. As amostras de solo utilizadas para quantificar os nematoides foram retiradas aos 45 dias apos o plantio das leguminosas; aos 30 e 60 dias apos o plantio da alface americana e aos 30, 60 e 90 dias apos o plantio do repolho. A incorporacao das leguminosas mucuna-preta e crotalaria, em cultivo orgânico, reduziu a populacao de Meloidogyne spp. em 42 e 51%, respectivamente, nessas hortalicas. A leguminosa feijao-de-porco causou reducao da populacao de nematoide apenas nas parcelas com repolho apos 90 dias.

Eclosão e mortalidade de Meloidogyne exigua em extratos e em produtos naturais

The hatching and mortality of Meloidogyne exigua second-stage juveniles (J2) were evaluated after 24 h in aqueous extracts of Bixa orellana, Cinnamomum zeylanicum, Petroselium crispum, Piper nigrum, Syzygium aromaticum, Zingiber officinale whey of milk, hidroponic solution, sodium chloride (NaCl) and saccharose aqueous solution, yeast and probiotic (Controlmix®) solutions. The whey of milk, C. zeylanicum, yeast and sodium chloride (NaCl) solutions caused 100% mortality of J2 (P<0,05). Extracts of S. aromaticum and Controlmix® probiotic caused more than 50% mortality of J2 (P<0,05). Hatching was evaluated by placing eggs of M. exigua in those different extracts and products for 16 days. The area under the curve of hatching progress, evaluated at 48 h intervals, showed that all extracts and products caused inhibition as compared to water (P<0,01), except for saccharose and hidroponic solutions. Whey of milk, probiotic, C. zeylanicum and S. aromaticum accounted for greater inhibition, among all the tested extracts and products (P<0,01). As shown, whey of milk and C. zeylanicum extract also caused 100% mortality of J2.

Manejo do solo e da irrigação como nova tática de controle de Meloidogyne incognita em feijoeiro

Population of Meloidogyne incognita were evaluated in the soil and by biotesting. The population fluctuation of M. incognita was also studied during the beans (Phaseolus vulgaris) life cycle. Two days after the treatments in the field, plowing, without irrigation was found to be the most efficient in reducing J2 population (P<0,01), followed by plowed-irrigated plots. The soil inoculum infectivity, however, was lower (P<0,01) in plowed-irrigated plots than in plowed soil only. At 14 days, the lowest (P<0,01) J2 population was observed in plowed-irrigated plots, followed by plowed soil. The infectivity at 14 days was different (P<0,01) for all treatments, but lowest (P<0,01) in plowed-irrigated plots. At 45 days after bean planting in the field, the J2 population in soil was, also, different (P<0,01) for all treatments, but still the lowest (P<0,01) in plowed-irrigated soil 14 days before seeding. At 90 days, in the field, the numbers of eggs and egg-masses per bean plant, and the numbers of J2 in the soil were also the lowest (P<0,01) in plowed-irrigated plots. The root and shoot weight of field bean in plowed and plowed-irrigated plots were equally (P<0,01) high, but higher (P<0,01) than in control and in only irrigated plots. Crop yield was different in all treatments (P<0,01). The greatest (P<0,01) bean yield occurred in plowed-irrigated plots 14 days before seeding with an increased productivity of four times compared to the control. Irrigation as a way of increasing effectiveness against plant parasitic nematodes has been previously postulated but never proved experimentally, as it was accomplished in this work.

Purification of two substances from bulbs of onion ( Allium cepa L.) with nematicidal activity against Meloidogyne exigua Goeldi

To contribute to the development of a new method for Meloidogyne exigua Goeldi control, a nematode widely disseminated through Brazilian coffee fields, this work aimed to purify the nematicidal substances produced by onion (Allium cepa L.). Thus, the methanolic extract of onion bulbs was concentrated under reduced pressure and submitted to a purification process guided by in vitro assays with second-stage juveniles (J2) of M. exigua. After several steps, two nematicidal substances that seemed to be interconverting when in solution were isolated. When exposed to 500 ppm aqueous solutions of substances 1 and 2, 55 and 38% of the J2, respectively, were dead after 24 h. Very preliminary nuclear magnetic resonance spectroscopy studies suggested that these substances correspond to a pair of carbohydrates.

Toxicidade de filtrados fúngicos a meloidogyne incognita

In the search for new nematicidal molecules, 18 fungal species filtrates and seven fungus mycelium extracts were studied in vitro to see their influence on hatching, mobility and mortality of second stage juveniles (J2) of Meloidogyne incognita. Fungal filtrates were obtained from fifteen day Czapek cultures. Methanol was used as solvent to obtain fungus mycelium extracts. Paecilomyces lilacinus, Fusarium moniliforme and Fusarium oxysporum filtrates reduced J2 motility and hatching, and increased J2 mortality (P < 0,05) similarly to Aldicarb. Some fungus filtrates reduced J2 mobility without leading to death. Aspergillus flavus, Cylindrocarpon magnusianum, Fusarium solani and Mortierella sp. filtrates reduced (P < 0,05) only J2 hatching. Production of toxic fungus metabolites was not dependent on the amount of mycelium produced. P. lilacinus produced toxic filtrates after one day culturing. Since then, toxicity increased steadily, reaching 100% J2 mortality on the thirteenth day.

Filtrados de culturas bacterianas endofíticas na motilidade, mortalidade e eclosão de juvenis de segundo estádio de Meloidogyne javanica

To obtaining the filtrates, bacteria were cultivated in liquid trypic soy broth medium for seven days at 28 oC under constant stirring, centrifugated at 10,000 g for 15 min and the supernatant was filtrated by 0,22 µm Millipore. Nonmotile Meloidogyne javanica and dead ones were counted 24 and 48 h later and hatched J2 were counted 15 days after setting up the experiment. Seven of 40 bacterial isolates tested immobilized the juveniles within 24 h, and they could not recover even in water. This effect was similar to the aldicarb treatment used as control. The filtrates of isolates also inhibited egg hatching. Two isolates killed more than 90% of J2 after 48 h of exposure. Filtrate dilutions of the most efficient isolate in 1:1 and 1:2 (bacterial filtrate: water) also showed to have effective action on nematode behaviors.

Occurrence of Meloidogyne paranaensis in coffee plantations in the Alto Paranaíba region of Minas Gerais, Brazil

Occurrence of Meloidogyne paranaensis in coffee plantations in the Alto Paranaiba region of Minas Gerais, Brazil Coffee (Coffea arabica) plantations showing symptoms of decline, with peeled and rough roots and fewer side rootlets were observed in the State of Minas Gerais, Brazil. Meloidogyne spp. females were obtained from the thicker roots and processed for perineal patterns and α -esterase electrophoresis analysis. Meloidogyne paranaensis was determined to be the main cause of the decline. The incidende and field symptom patterns found were similar to those observed on coffee plantations in the State of Sao Paulo and Parana, where most damage caused by M. paranaensis has been reported. O parasitismo dos fitonematoides constitui-se num dos principais problemas da cultura do cafeeiro (Coffea arabica L.), e o genero Meloidogyne Goeldi e responsavel pelos maiores danos [Campos, V.P. Cafe, Doencas causadas por nematoides. In: Ribeiro do Vale & Zambolim (Eds.) Controle de Doencas de Plantas: Grandes Culturas, v.1, p.141, 1997]. Este autor relatou que M. exigua esta amplamente disseminada em Minas Gerais, contudo M. incognita (Kofoid & White) Chitwood e M. coffeicola Lordello & Zamith tem sido constatadas em poucas regioes cafeeiras daquele Estado. Nos ultimos quatro anos, levantamentos de fitonematoides associados a cultura do cafeeiro em Minas Gerais vem sendo feitos de modo efetivo e com apoio da EMBRAPA-FUNCAFE. Recentemente, em algumas lavouras nos municipios de Serra do Salitre e de Patrocinio, regiao do Alto Paranaiba em Minas Gerais, observaram-se plantas desfolhadas, com sistemas radiculares reduzidos, raizes grossas apresentando superficie macia, com escamacoes, e lesoes com aspecto de cancro (Figuras 1 A e B). Para diagnose parasitologica e taxo nomica, coletaram-se amostras de solo e raizes em torno de plantas afetadas. Em raizes, caracterizando-se associacao constante, femeas de Meloidogyne sp. foram encontradas e retiradas para exame da configuracao perineal e extracao de proteinas, que foram submetidas a eletroforese em gel de poliacrilamida, para identificacao da especie segundo o fenotipo de α-esterase [Barker, K.R., Carter, C.C. & Sasser, J.N. (Eds.) An Advanced Treatise on Meloidogyne, v. 2, p.69, 1985]. As observacoes revelaram configuracoes perineais com arco dorsal elevado, trapezoidal e estrias lisas a onduladas. A analise de α-esterase revelou o fenotipo F1 (Figura 1C), tipico de M. paranaensis (Carneiro et al., Journal of Nematol., 28:179, 1996). Nas areas estudadas, o padrao sintomatologico observado coincidiu com o observado em cafezais dos Estados de Sao Paulo e Parana. Urgem medidas excludentes para evitaremse disseminacoes. FIG. 1 Sintomas do parasitismo de Meloidogyne paranaensis em cafeeiro (Coffea arabica). (A) Parte aerea e (B) raizes. C) F1, fenotipo de αesterase, tipico de M. paranaensis , J3, fenotipo de M. javanica, utilizado como padrao de comparacao. 02104 A B C NOTAS FITOPATOLOGICAS / PHYTOPATHOLOGICAL NOTES