Efficacy of Purpureocillium lilacinum AUMC 10149 as biocontrol agent against root-knot nematode Meloidogyne incognita infecting tomato plant

Abstract Root-knot nematode Meloidogyne incognita is among the biotic factors which has greatly affected both the yield and the quality of the tomato crop. The egg parasitic nematode, Purpureocillium lilacinum (Pl) is considered as one of the most promising agents in controlling and overcoming this plant pathogen. The nematicidal effect of the native isolate Pl AUMC 10149 on second stage juveniles survival and egg hatching of M. incognita at different times of exposure was tested in vitro. The obtained data showed that Pl gave a maximum percentage of J2 mortality (97.6%) and egg hatching inhibition (79.8%) after 72 hours of exposure. The potentiality of Pl as well as Bio-Nematon to control M. incognita infecting tomato was conducted using different times of application in vivo. Nine treatments with five replicates were used for such bioagents compared with the nematicide Oxamyl. Each seedling was inoculated with 1000 J2s of nematode/pot and 10 mL of Pl (1x1010 CFU/mL) or Bio-Nematon spore suspension (1x108 CFU/mL) 10mL/pot. The results indicated that the most effective treatments in reducing nematode population, number of galls and egg masses of M. incognita in plant roots was performed with treatment by Pl pre-planting and post-infection with Pl (Rf 1.9) giving a significant enhancement in plant length (64.9%), fresh weight (72.52%) and shoot dry weight (163.41%) without negatively impacting environment. Therefore, the present study confirmed that using P. lilacinum AUMC 10149 can be used as a practical supplement to environmentally friendly disease management of root-knot nematodes in Egypt.

Resumo O nematoide-das-galhas Meloidogyne incognita está entre os fatores bióticos que afetaram enormemente a produção e a qualidade da cultura do tomate. O nematoide parasita de ovos, Purpureocillium lilacinum (Pl), é considerado um dos mais promissores agentes no controle e superação desse fitopatógeno. O efeito nematicida do isolado nativo Pl AUMC 10149 na sobrevivência de juvenis de segundo estágio e na eclosão dos ovos de M. incognita em diferentes momentos de exposição foi testado in vitro. Os dados obtidos mostraram que o Pl deu um percentual máximo de mortalidade de J2 (97.6%) e inibição da eclosão dos ovos (79.8%) após 72 horas de exposição. A potencialidade de Pl e de Bio-Nematon para controlar M. incognita infectando tomate foi conduzida em diferentes tempos de aplicação in vivo. Nove tratamentos com cinco repetições foram usados para tais bioagentes em comparação com o nematicida Oxamyl. Cada muda foi inoculada com 1.000 J2s de nematoide / vaso e 10 mL de Pl (1×1010 CFU/mL). Ou suspensão de esporos Bio-Nematon (1×108 CFU/mL) 10mL/pot. Os resultados indicaram que os tratamentos mais eficazes na redução da população de nematoides, número de galhas e desovas de M. incognita nas raízes das plantas foram realizados com Pl pré-plantio e pós-infecção com Pl (Rf 1.9), dando um aumento significativo no comprimento da planta (64.9%), massa fresca (72.52%) e massa seca da parte aérea (163.41%) sem impactar negativamente o meio ambiente. Portanto, o presente estudo confirmou que o uso de P. lilacinum AUMC 10149 pode ser usado como um suplemento prático para o manejo ecologicamente correto de nematoides-das-galhas no Egito.

Efficacy of Purpureocillium lilacinum AUMC 10149 as biocontrol agent against root-knot nematode Meloidogyne incognita infecting tomato plant / Eficácia de Purpureocillium lilacinum AUMC 10149 como agente de biocontrole contra o nematoide Meloidogyne incógnita que infecta plantas de tomate

Abstract Root-knot nematode Meloidogyne incognita is among the biotic factors which has greatly affected both the yield and the quality of the tomato crop. The egg parasitic nematode, Purpureocillium lilacinum (Pl) is considered as one of the most promising agents in controlling and overcoming this plant pathogen. The nematicidal effect of the native isolate Pl AUMC 10149 on second stage juvenile's survival and egg hatching of M. incognita at different times of exposure was tested in vitro. The obtained data showed that Pl gave a maximum percentage of J2 mortality (97.6%) and egg hatching inhibition (79.8%) after 72 hours of exposure. The potentiality of Pl as well as Bio-Nematon to control M. incognita infecting tomato was conducted using different times of application in vivo. Nine treatments with five replicates were used for such bioagents compared with the nematicide Oxamyl. Each seedling was inoculated with 1000 J2s of nematode/pot and 10 mL of Pl (1x1010 CFU/mL) or Bio-Nematon spore suspension (1x108 CFU/mL) 10mL/pot. The results indicated that the most effective treatments in reducing nematode population, number of galls and egg masses of M. incognita in plant roots was performed with treatment by Pl pre-planting and post-infection with Pl (Rf 1.9) giving a significant enhancement in plant length (64.9%), fresh weight (72.52%) and shoot dry weight (163.41%) without negatively impacting environment. Therefore, the present study confirmed that using P. lilacinum AUMC 10149 can be used as a practical supplement to environmentally friendly disease management of root-knot nematodes in Egypt.

Resumo O nematoide-das-galhas Meloidogyne incognita está entre os fatores bióticos que afetaram enormemente a produção e a qualidade da cultura do tomate. O nematoide parasita de ovos, Purpureocillium lilacinum (Pl), é considerado um dos mais promissores agentes no controle e superação desse fitopatógeno. O efeito nematicida do isolado nativo Pl AUMC 10149 na sobrevivência de juvenis de segundo estágio e na eclosão dos ovos de M. incognita em diferentes momentos de exposição foi testado in vitro. Os dados obtidos mostraram que o Pl deu um percentual máximo de mortalidade de J2 (97.6%) e inibição da eclosão dos ovos (79.8%) após 72 horas de exposição. A potencialidade de Pl e de Bio-Nematon para controlar M. incognita infectando tomate foi conduzida em diferentes tempos de aplicação in vivo. Nove tratamentos com cinco repetições foram usados ​​para tais bioagentes em comparação com o nematicida Oxamyl. Cada muda foi inoculada com 1.000 J2s de nematoide / vaso e 10 mL de Pl (1×1010 CFU/mL). Ou suspensão de esporos Bio-Nematon (1×108 CFU/mL) 10mL/pot. Os resultados indicaram que os tratamentos mais eficazes na redução da população de nematoides, número de galhas e desovas de M. incognita nas raízes das plantas foram realizados com Pl pré-plantio e pós-infecção com Pl (Rf 1.9), dando um aumento significativo no comprimento da planta (64.9%), massa fresca (72.52%) e massa seca da parte aérea (163.41%) sem impactar negativamente o meio ambiente. Portanto, o presente estudo confirmou que o uso de P. lilacinum AUMC 10149 pode ser usado como um suplemento prático para o manejo ecologicamente correto de nematoides-das-galhas no Egito.

Suppression and management of Meloidogyne incognita in soil using Trichoderma harzianum NFCF160 and Trichoderma virens Isf-77

Aims@#Meloidogyne incognita adversely affects numerous crop plants worldwide. Therefore, the modern world has been moving towards biocontrol methods to prevent nematode attacks. This study was aimed to (i) investigate the potential use of Trichoderma harzianum NFCF160 and T. virens Isf-77 in managing M. incognita in soil and (ii) identify trapping mechanisms employed by both Trichoderma strains to suppress M. incognita.@*Methodology and results@#Three weeks old, Basella alba L. plants were subjected to five different treatments. The above and below ground growth parameters and the galling indices of these plants were measured every four weeks for three sampling times. Trapping mechanisms employed by Trichoderma strains were examined following plate assays. Plants treated with T. harzianum NFCF160 and T. virens Isf-77 had significantly higher values for the total number of leaves (34 ± 2.84) and (27 ± 2.61), fresh weight of the shoot (81 ± 9.51 g) and (91 ± 9.70 g), dry weight of the shoot (71 ± 5.24 g) and (62 ± 5.81 g), respectively eight weeks after inoculation of M. incognita. Significantly low galling indices (2 and 2) were recorded in B. alba treated with Trichoderma strains. Both Trichoderma strains exhibited various nematode-trapping mechanisms, such as non-constricting rings and adhesive spores.@*Conclusion, significance and impact of study@#This investigation highlighted the potential of both Trichoderma strains as biocontrol agents to control M. incognita effect in sustainable agriculture.

Phytochemical management of root knot nematode (Meloidogyne incognita) kofoid and white chitwood by Artemisia spp. in tomato (Lycopersicon esculentum L.) / Manejo fitoquímico do nematoide do nódulo de raiz (Meloidogyne incognita) kofoid e do chitwood branco por Artemisia spp. em tomate (Lycopersicon esculentum L.)

Abstract In vitro and screen house experiments were conducted to investigate the effectiveness of thirteen phytochemicals from Artemisia elegantissimia and A. incisa on root knot nematode, Meloidogyne incognita in tomato (Lycopersicon esculentum L.) cv. Rio Grande. A positive control (Carbofuran) and negative control (H2O) were also used for comparison. Effectiveness of phytochemicals against juveniles (J2s) mortality and egg hatch inhibition were evaluated after 24, 48 and 72 hours of incubation at three concentrations viz; 0.1, 0.2 and 0.3 mg/mL in vitro conditions. Amongst thirteen phytochemicals, Isoscopletin (Coumarin), Carbofuran and Apigenin (Flavonoid) showed the highest mortality and egg hatch inhibition of M. incognita at all intervals. Inhibition of eggs and J2s mortality were the greatest (90.0%) and (96.0%) at 0.3 mg/mL concentration. Application of phytochemicals caused reduction in number of galls, galling index, and egg masses on tomato plant and enhanced plant growth parameters under screen house conditions. Gall numbers (1.50), galling index (1.00), number of juveniles (4.83) and egg masses (4.00) were greatly reduced and plant growth parameters such as; plant height (28.48 cm), fresh (72.13 g) and dry shoot weights (35.99 g), and root fresh (6.58 g) and dry weights (1.43 g) were increased significantly by using Isoscopletin. In structure activity relationship, juveniles of M. incognita, exhibited variations in their shape and postures upon death when exposed to different concentrations of phytochemicals of Artemisia spp. The present study suggests that Artemisia based phytochemicals possess strong nematicidal effects and can be used effectively in an integrated disease management program against root knot nematodes.

Resumo Experimentos in vitro e de triagem foram conduzidos para investigar a eficácia de treze constituintes fitoquímicos de Artemisia elegantissimia e A. incisa no nematóide de galhas, Meloidogyne incognita em tomateiro (Lycopersicon esculentum L.) cv. Rio Grande. Um controle positivo (carbofuran) e controle negativo (H2O) também foram utilizados para comparação. A eficácia dos fitoquímicos contra a mortalidade juvenil (J2s) e a inibição da eclosão de ovos foram avaliadas após 24, 48 e 72 horas de incubação em três concentrações, tais como: 0,1; 0,2 e 0,3 mg/mL em condições in vitro. Dentre os treze fitoquímicos, isoscopletina (cumarina), carbofurano e apigenina (flavonoide) apresentaram a maior mortalidade e a inibição da eclosão de ovos de M. incognita em todos os intervalos. A inibição da mortalidade dos ovos e J2s foi a maior (90,0%) e (96,0%) na concentração de 0,3 mg/mL. A aplicação de fitoquímicos causou redução no número de galhas, índice de fricção e massa de ovos no tomateiro e melhorou os parâmetros de crescimento das plantas em condições de triagem. Números de galhas (1,50), índice de insetos galhadores (1,00), número de juvenis (4,83) e massas de ovos (4,00) foram bastante reduzidos e os parâmetros de crescimento das plantas, como altura da planta (28,48 cm), peso fresco (72,13 g) e seco (35,99 g), raiz fresca (6,58 g) e peso seco (1,43 g) foram significativamente aumentados usando isoscopletina. Na relação atividade estrutura, juvenis de M. incognita, exibiram variações em sua forma e posturas após a morte quando expostos a diferentes concentrações de fitoquímicos de Artemisia spp. O presente estudo sugere que os fitoquímicos à base de artemísia possuem fortes efeitos nematicidas e podem ser usados ​​eficazmente em um programa integrado de controle de doenças contra nematóides de galhas.

Response of tuberose genotypes to root knot nematode, Meloidogyne incognita: Biochemical, histological and nutritional characterization of host-pathogen interaction

Aim: The study was conducted to screen the tuberose genotypes for nematode resistance and to establish histopathological, biochemical and nutritional changes in resistant/tolerant genotype against root knot nematode, Meloidogyne incognita. Methodology: Twenty seven tuberose genotypes comprising of two types viz., single (18 genotypes) and double (9 genotypes) were evaluated for root knot nematode resistance. Tuberose bulbs of uniform size were surface sterilized and planted in 9 inch plastic pots and inoculated with nematode juveniles (J2) 15 days after planting. Gall Index was recorded on a 0-5 scale after 120 days of inoculation. Histopathological, biochemical (total phenols, peroxidase, polyphenol oxidase and phenylalanine ammonia lyase) and nutrient estimation studies were conducted on the roots of resistant/tolerant genotypes against root knot nematode, Meloidogyne incognita. Results: Among the twenty seven tuberose genotypes screened for nematode resistance, Suarna Rekha was categorized as resistant genotype and Arka Shirngar, IIHR-2, IIHR-4, Variegated, IIHR-12, GK-T-C4, Calcutta Double, Bidhan Rajani-2 and Bidhan Rajani-3 were categorized as tolerant genotypes. Histological characterization of resistant/tolerant genotypes revealed that giant cell formation was disrupted with higher number of phenolic cells and cells with lignified cell wall that prevented the successful establishment of nematode feeding site. The resistant and tolerant genotypes significantly recorded higher accumulation of phenols and defence enzymes viz., polyphenol oxidase (PPO), peroxidase (PO) and phenyl alanine ammonia lyase (PAL) and macro and micro nutrients as compared to susceptible genotypes. Interpretation: Screening of tuberose genotypes for root knot nematode resistance in this study has led to the identification of several nematode resistant/ tolerant genotypes. This would serve as a valuable information for the breeders to utilize resistant/ tolerant genotypes as parents in their breeding programmes.

Role of Synadenium grantii latex proteases in nematicidal activity on Meloidogyne incognita and Panagrellus redivivus / Papel das proteases do látex de Synadenium grantii na atividade nematicida sobre Meloidogyne incognita e Panagrellus redivivus

Abstract Synadenium grantii is a Euphorbiaceae plant commonly found in Brazil, known as Janaúba or Leitosinha, whose latex is traditionally used for several purposes. However, it is not known whether the nematicidal action of this plant latex occurs due to the action of proteases. The present work aims to evaluate the nematicidal activity of proteases from Synadenium grantii latex on Meloidogyne incognita and Panagrellus redivivus. S. grantii latex used in the present study was collected from specimens found in Universidade Federal de Viçosa, Viçosa, Minas Gerais, Brazil. The drained latex was collected in Eppendorf microtubes and immediately stored on ice at 4 °C. After this extraction, the latex was frozen (-20 °C) during 2 hours, thawed at room temperature (25 °C) and centrifuged at 10,000 g at 4 °C for 30 minutes to remove larger particles and concentrate the proteases. After the centrifugation, assays of enzymatic activity were performed in order to know in which of the phases the enzymes were found. S. grantii latex presented protease, but no chitinase activity. The results show that there was a significant difference (p <0.01) between the treated and control groups, with 100% mortality of Meloidogyne incognita and 72% average mortality of Panagrellus redivivus. In addition, it was demonstrated that the nematicidal action occurred due to the action of the proteases, since the control was only differentiated from the treatment by the presence of the enzymes with biological activity.

Resumo Synadenium grantii é uma planta Euphorbiaceae comumente encontrada no Brasil, conhecida como Janaúba ou Leitosinha, e tem seu látex usado tradicionalmente para diferentes propósitos. Entretanto, não se conhece se a atividade nematicida da planta ocorre devido à ação de proteases. O presente trabalho tem como objetivo avaliar a atividade nematicida das proteases do látex de Synadenium grantii sobre Meloidogyne incognita e Panagrellus redivivus. O látex de S. grantii utilizado no presente trabalho foi coletado a partir de espécimes encontradas na Universidade Federal de Viçosa, Viçosa, Minas Gerais, Brazil. O látex foi coletado em microtubos Eppendorf e imediatamente armazenado em gelo a 4 °C. Após esta extração, o látex foi congelado (-20 °C) durante 2 horas, descongelado à temperatura ambiente (25 °C) e centrifugado a 10000 g a 4 °C durante 30 minutos para a remoção de partículas e concentração das proteases. Após a centrifugação, foram realizados ensaios de atividade enzimática para saber em qual das fases as enzimas foram encontradas. O látex de S. grantii apresentou atividade de protease, mas nenhuma atividade de quitinase. Os resultados mostram que houve diferença significativa (p <0,01) entre os grupos tratados e controle, com 100% de mortalidade de Meloidogyne incognita e 72% de mortalidade média de Panagrellus redivivus. Além disso, foi demonstrado que a ação nematicida ocorreu devido à ação das proteases, uma vez que o grupo controle só foi diferenciado do tratamento pela presença das enzimas com atividade biológica.

Supplanting resistance of the Mi gene by root-knot nematode in industrial tomato in the Cerrado in Goiás State of Brazil / Suplantação de resistência do gene Mi por nematoide-das-galhas em tomate industrial no Cerrado de Goiás, Brasil

ABSTRACT: Problems with root-knot nematodes, caused by genus Meloidogyne, can be controlled through the introduction of resistance genes in commercial varieties of tomatoes. However, plants of the hybrid 'Heinz 9992' for industrial processing, carrying the Mi gene, were reported with their roots severely infected in experimental cultivation in Goiás State of Brazil with symptoms typical of galls caused by Meloidogyne spp. After dissecting the galls present in the root samples, the females of the nematodes were removed and afterwards, performed the analysis of the perineal pattern and the biochemical characterization by electrophoresis of isoenzymes, besides the pathogenicity test. The pathogen analyzed was identified as Meloidogyne incognita. The pathogenicity test confirmed the supplanting resistance of the tomato hybrid with the Mi gene by this virulent population of M. incognita.

RESUMO: Problemas causados pelos nematoides-das-galhas, pertencentes ao gênero Meloidogyne podem ser controlados com a introdução de genes de resistência em cultivares comerciais de tomateiro. No entanto, plantas do híbrido 'Heinz 9992' adequadas ao processamento industrial, portadoras do gene Mi, foram encontradas com suas raízes severamente infectadas, em um cultivo experimental no município de Morrinhos (GO) com sintomas típicos de galhas causadas por Meloidogyne spp. Após dissecar as galhas presentes nas amostras de raízes, as fêmeas do nematoide foram retiradas para a análise do padrão perineal e a caracterização bioquímica por eletroforese de isoenzimas, além do teste de patogenicidade. O patógeno em estudo foi identificado como Meloidogyne incognita. O teste de patogenicidade confirmou que ocorreu a suplantação de resistência do híbrido de tomateiro, portador do gene Mi, por esta população virulenta de M. incognita.

Identifying resistance to root-knot nematodes in Capsicum genotypes / Identificação de genótipos de Capsicum resistentes a nematoides de galha

The present study aimed to evaluate Capsicum accessions for resistance to Meloidogyne incognita race 3, Meloidogyne javanica and Meloidogyne enterolobii. Two experiments with different genotypes of hot and sweet peppers were carried out in a completely randomized design. The first experiment was conducted in a 31 x 3 factorial scheme with 27 genotypes of Capsicum annuum, two cultivars of hot pepper, one line of Capsicum frutescens and tomato 'Santa Cruz Kada', and three species of nematodes (M. incognita race 3, M. javanica and M. enterolobii). In the second experiment, we used a factorial scheme 39 x 3 with 36 accessions of C. annuum, two hot pepper cultivars and the 'Santa Cruz Kada' tomato and three nematodes species mentioned earlier. The total number of eggs and second-stage juveniles (TNEJ), number of eggs and second-stage juveniles per gram of root (NEJGR), reproduction index (RI) and reproduction factor (RF) were evaluated. Based on RI and RF, the genotypes CNPH 185, CNPH 187 and CNPH 680 were resistant and very resistant to M. incognita race 3 and M. javanica, simultaneously. The C. frutescens line presented resistance to the three root-knot nematode species.

O presente trabalho teve por objetivo avaliar acessos de Capsicum quanto à resistência a Meloidogyne incognita raça 3, Meloidogyne javanica e Meloidogyne enterolobii. Foram realizados dois experimentos, com diferentes genótipos de pimentas e pimentões, em delineamento inteiramente casualizado sendo o primeiro em esquema fatorial 31 x 3 com 27 genótipos de Capsicum annuum, duas cultivares de pimenta, uma linhagem de Capsicum frutescens, o tomateiro 'Santa Cruz Kada' e três espécies de nematoides (M. incognita raça 3, M. javanica e M. enterolobii). No segundo experimento foi utilizado esquema fatorial 39 x 3 com 36 acessos de C. annuum, duas cultivares de pimenta, o tomateiro 'Santa Cruz Kada' e três espécies de nematoides mencionadas anteriormente. Avaliou-se o número total de ovos e juvenis de segundo estádio (NTOJ), número de ovos e juvenis de segundo estádio por grama de raízes (NOJGR), índice de reprodução (IR) e fator de reprodução (FR). Com base no FR e IR os genótipos CNPH 185, CNPH 187 e CNPH 680 foram resistentes e muito resistentes a M. incognita raça 3 e M. javanica, simultaneamente. A linhagem de C. frutescens apresentou resistência às três espécies de nematoides de galha.

Nematicidal Compounds from the Leaves of Schinus terebinthifolius Against Root-knot Nematode, Meloidogyne incognita Infecting Tomato

The root-knot nematode, Meloidogyne incognita caused a serious damage to many plants. The phenolic components of the leaves of Schinus terebinthifolius were investigated as potential nematicidal agents for M. incognita. Nine compounds were isolated and characterized as viz., 1,2,3,4,6-pentagalloyl glucose (1), kaempferol-3-O-α-L-rhamnoside (Afzelin) (2), quercetin-3-O-α-L-rhamnoside (Quercetrin) (3), myricetin (4), myricetin-3-O-α-L-rhamnoside (Myricetrin) (5), methylgallate (6), protocatechuic acid (7), quercetin (8), and gallic acid (9) using nuclear magnetic resonance (NMR) spectroscopy. Compound 1 showed pronounced nematicidal activity compared to Oxamyl as a positive control. It showed the lowest eggs-hatchability (34%) and the highest mortality in nematode population (21% after 72 hours of treatment) at a concentration of 200 µg/mL. It exhibited the best suppressed total nematode population, root galling and number of eggmasses in infected tomato plants. The total carbohydrates and proteins were also significantly induced by 1 with reduction in total phenolics and increase in defense-related proteins. Thus, compound 1 could be a promising, more safe and effective natural nematicidal agent for the control of root-knot nematodes.

Role of Synadenium grantii latex proteases in nematicidal activity on Meloidogyne incognita and Panagrellus redivivus

Abstract Synadenium grantii is a Euphorbiaceae plant commonly found in Brazil, known as Janaúba or Leitosinha, whose latex is traditionally used for several purposes. However, it is not known whether the nematicidal action of this plant latex occurs due to the action of proteases. The present work aims to evaluate the nematicidal activity of proteases from Synadenium grantii latex on Meloidogyne incognita and Panagrellus redivivus. S. grantii latex used in the present study was collected from specimens found in Universidade Federal de Viçosa, Viçosa, Minas Gerais, Brazil. The drained latex was collected in Eppendorf microtubes and immediately stored on ice at 4 °C. After this extraction, the latex was frozen (-20 °C) during 2 hours, thawed at room temperature (25 °C) and centrifuged at 10,000 g at 4 °C for 30 minutes to remove larger particles and concentrate the proteases. After the centrifugation, assays of enzymatic activity were performed in order to know in which of the phases the enzymes were found. S. grantii latex presented protease, but no chitinase activity. The results show that there was a significant difference (p 0.01) between the treated and control groups, with 100% mortality of Meloidogyne incognita and 72% average mortality of Panagrellus redivivus. In addition, it was demonstrated that the nematicidal action occurred due to the action of the proteases, since the control was only differentiated from the treatment by the presence of the enzymes with biological activity.

Resumo Synadenium grantii é uma planta Euphorbiaceae comumente encontrada no Brasil, conhecida como Janaúba ou Leitosinha, e tem seu látex usado tradicionalmente para diferentes propósitos. Entretanto, não se conhece se a atividade nematicida da planta ocorre devido à ação de proteases. O presente trabalho tem como objetivo avaliar a atividade nematicida das proteases do látex de Synadenium grantii sobre Meloidogyne incognita e Panagrellus redivivus. O látex de S. grantii utilizado no presente trabalho foi coletado a partir de espécimes encontradas na Universidade Federal de Viçosa, Viçosa, Minas Gerais, Brazil. O látex foi coletado em microtubos Eppendorf e imediatamente armazenado em gelo a 4 °C. Após esta extração, o látex foi congelado (-20 °C) durante 2 horas, descongelado à temperatura ambiente (25 °C) e centrifugado a 10000 g a 4 °C durante 30 minutos para a remoção de partículas e concentração das proteases. Após a centrifugação, foram realizados ensaios de atividade enzimática para saber em qual das fases as enzimas foram encontradas. O látex de S. grantii apresentou atividade de protease, mas nenhuma atividade de quitinase. Os resultados mostram que houve diferença significativa (p 0,01) entre os grupos tratados e controle, com 100% de mortalidade de Meloidogyne incognita e 72% de mortalidade média de Panagrellus redivivus. Além disso, foi demonstrado que a ação nematicida ocorreu devido à ação das proteases, uma vez que o grupo controle só foi diferenciado do tratamento pela presença das enzimas com atividade biológica.

Phytochemical management of root knot nematode (Meloidogyne incognita) kofoid and white chitwood by Artemisia spp. in tomato (Lycopersicon esculentum L.)

Abstract In vitro and screen house experiments were conducted to investigate the effectiveness of thirteen phytochemicals from Artemisia elegantissimia and A. incisa on root knot nematode, Meloidogyne incognita in tomato (Lycopersicon esculentum L.) cv. Rio Grande. A positive control (Carbofuran) and negative control (H2O) were also used for comparison. Effectiveness of phytochemicals against juveniles (J2s) mortality and egg hatch inhibition were evaluated after 24, 48 and 72 hours of incubation at three concentrations viz; 0.1, 0.2 and 0.3 mg/mL in vitro conditions. Amongst thirteen phytochemicals, Isoscopletin (Coumarin), Carbofuran and Apigenin (Flavonoid) showed the highest mortality and egg hatch inhibition of M. incognita at all intervals. Inhibition of eggs and J2s mortality were the greatest (90.0%) and (96.0%) at 0.3 mg/mL concentration. Application of phytochemicals caused reduction in number of galls, galling index, and egg masses on tomato plant and enhanced plant growth parameters under screen house conditions. Gall numbers (1.50), galling index (1.00), number of juveniles (4.83) and egg masses (4.00) were greatly reduced and plant growth parameters such as; plant height (28.48 cm), fresh (72.13 g) and dry shoot weights (35.99 g), and root fresh (6.58 g) and dry weights (1.43 g) were increased significantly by using Isoscopletin. In structure activity relationship, juveniles of M. incognita, exhibited variations in their shape and postures upon death when exposed to different concentrations of phytochemicals of Artemisia spp. The present study suggests that Artemisia based phytochemicals possess strong nematicidal effects and can be used effectively in an integrated disease management program against root knot nematodes.

Resumo Experimentos in vitro e de triagem foram conduzidos para investigar a eficácia de treze constituintes fitoquímicos de Artemisia elegantissimia e A. incisa no nematóide de galhas, Meloidogyne incognita em tomateiro (Lycopersicon esculentum L.) cv. Rio Grande. Um controle positivo (carbofuran) e controle negativo (H2O) também foram utilizados para comparação. A eficácia dos fitoquímicos contra a mortalidade juvenil (J2s) e a inibição da eclosão de ovos foram avaliadas após 24, 48 e 72 horas de incubação em três concentrações, tais como: 0,1; 0,2 e 0,3 mg/mL em condições in vitro. Dentre os treze fitoquímicos, isoscopletina (cumarina), carbofurano e apigenina (flavonoide) apresentaram a maior mortalidade e a inibição da eclosão de ovos de M. incognita em todos os intervalos. A inibição da mortalidade dos ovos e J2s foi a maior (90,0%) e (96,0%) na concentração de 0,3 mg/mL. A aplicação de fitoquímicos causou redução no número de galhas, índice de fricção e massa de ovos no tomateiro e melhorou os parâmetros de crescimento das plantas em condições de triagem. Números de galhas (1,50), índice de insetos galhadores (1,00), número de juvenis (4,83) e massas de ovos (4,00) foram bastante reduzidos e os parâmetros de crescimento das plantas, como altura da planta (28,48 cm), peso fresco (72,13 g) e seco (35,99 g), raiz fresca (6,58 g) e peso seco (1,43 g) foram significativamente aumentados usando isoscopletina. Na relação atividade estrutura, juvenis de M. incognita, exibiram variações em sua forma e posturas após a morte quando expostos a diferentes concentrações de fitoquímicos de Artemisia spp. O presente estudo sugere que os fitoquímicos à base de artemísia possuem fortes efeitos nematicidas e podem ser usados eficazmente em um programa integrado de controle de doenças contra nematóides de galhas.

Efecto del filtrado crudo de Purpureocillium sp. (Cepa UdeA0106), sobre la eclosión de huevos y movilidad de juveniles de Meloidogyne incognita-javanica / Effect of the raw filtrate of Purpureocillium sp. (UdeA0106 strain) on the eclosion of eggs and the motility of Meloidogyne incognitajavanica juveniles

Meloidogyne incognita-javanica representa una limitación de los cultivos agrícolas y es necesario buscar alternativas para su manejo. El presente trabajo tuvo como objetivo evaluar el efecto in vitro del filtrado crudo de Purpureocillium sp. (Cepa UdeA0106) sobre laeclosión de huevos y los estadios juveniles de Meloidogyne spp. Adicionalmente, se evaluaron los medios: caldo extracto de malta (CEM), caldo extracto de levadura (CEL) y caldo papa glucosa (CPG), escogiéndose aquel que no ejerciera daño sobre la morfología de los huevos y permitiera la eclosión de los estadios juveniles. El hongo se multiplicó durante 7 días y fue filtrado. Para establecer su efecto sobre los huevos y estadios J2 de Meloidogyne spp., se probaron concentraciones del filtrado al 100 %, 90 %, 70 %, 50 %, 25 % y 10 %; comparándolas con un control compuesto de estadios J2 en agua destilada estéril (ADE) y CPG. Las variables a evaluar fueron el porcentaje de eclosión e inmovilidad en los J2. La unidad experimental fue la caja de Petri, cada una con 200 huevos/J2, con 5 réplicas de cada tratamiento. Se utilizó un diseño experimental completamente aleatorizado con ANOVA (P = 0,05) y análisis de comparación múltiple de Tukey. Se encontraron diferencias significativas entre todos los tratamientos (P < 0,05) pero el tratamiento correspondiente al filtrado puro (100 %) logró mantener el porcentaje más bajo de eclosión y mayor porcentaje de inmovilidad de los J2 (96,8 %). Las concentraciones al 90 % y 70 % igualmente lograron efectos en los J2, con porcentajes altos de inmovilidad.

Meloidogyne incognita-javanica represents an important threat to the agricultural crops, and it is necessary to seek alternatives for its control. The aim of this research it was to evaluate a crude filtrate from the strain Purpureocillium sp. UdeA0106 in order to establish the in vitro effect on the egg hatching and the immobility of juvenile stages of Meloidogyne spp. The mediums evaluated were malt extract broth (MEB), yeast extract broth (YEB), and potato dextrose broth (PDB), after which it was chosen the one that didn't produce any damages on nematode eggs and allowed the emergence of the larval stages. The fungus was multiplied for 7 days, and then it was filtrated. To evaluate its effect on the eggs and the J2 of Meloidogyne spp., filtrate concentrations were made at 100 %, 90 %, 70 %, 50 %, 25 % and 10 %, contrasting them with a control composed of the J2 stages in sterile distilled water and PDB. The variables evaluated were the percentage of altered eggs and the immobility observed in J2 stages. The experimental unit was the Petri dish, each one containing 200 eggs /J2. Each treatment was replicated 5 times. It was used a completely randomized design, and an ANOVA was performed (P = 0,05) as well as a multiple comparison test (Tukey). Significant differences were found between all treatments (P < 0,05), although the most effective in inhibiting both egg hatching and J2 mobility (96,8 %) were the full strength filtering (100 %). The dilutions at 90 % and 70 %, were able to produce effects with high percentages in the immobility.

Management of Root-Knot Disease on Tomato with Bioformulated Paecilomyces lilacinus and Leaf Extract of Lantana camara

Glasshouse experiments were conducted to evaluate the efficacy of application frequency of a bioformulated Paecilomyces lilacinus in combination with five concentration of Lantana camara crude aqueous leaf extract against Meloidogyne incognita race I on tomato. The experiment was a 3x5 factorial laid out in a completely randomized design (CRD) with four replications. Each seedling was inoculated with 5000 eggs of M. incognita. Application of the bionematicide and L. camara leaf extract alone significantly (P≤ 0.05) inhibited root galling and egg production compared with their respective control. However, the severity of root galling and egg mass production was more significantly (P< 0.05) suppressed with the application of P. lilacinus than L. camara leaf extract. Double inoculation with P. lilacinus in combination with 0.80 g mL-1 of the L. camara leaf extract changed the susceptibility of the tomato cultivar with gall index (GI=4.00) to GI=1.50. Application of P. lilacinus twice (at transplanting and two weeks after transplanting) in combination with 0.80g mL-1 of L. camara leaf extract was the most effective treatment in gall and egg mass inhibition, growth enhancement and dry matter accumulation. This environment-friendly approach could be incorporated into integrated root-knot disease management in tomato.

Correlação entre densidade populacional de nematoides e produtividade de algodoeiro / Correlation between density population of nematodes and productivity of cotton

O sistema de produção do algodoeiro em Mato Grosso tem tido sua produtividade reduzida e seus custos de produção aumentados em função da presença dos nematoides. Entre os nematoides associados à cultura, estão Pratylenchus brachyurus, Meloidogyne incognita e Rotylenchulus reniformis, podendo ocorrer isoladamente ou de forma simultânea. Este trabalho teve por objetivo avaliar efeitos de diferentes densidades populacionais de nematoides em duas cultivares de algodoeiro (FMT 701 e IMACD 6001LL). A presença simultânea dos referidos nematoides contribuíram para a redução da produtividade. Comparando a produção nessas áreas foi possível observar perdas em média, 23,1% e 35% no rendimento de algodão em caroço para as cultivares IMACD 6001 LL e FMT 701, respectivamente, em áreas infestadas.

The cotton crop is of great commercial value, main for its fiber and are adding each year the area under cultivation in the state of Mato Grosso, thus increasing its importance to the State. The system of cotton production in Mato Grosso has had its productivity and reduced production costs increased due to the presence of nematodes. Among the nematodes associated to the culture are Pratylenchus brachyurus, Meloidogyne incognita and Rotylenchulus reniformis, which may occur alone or in combination with one or more nematodes. This study aimed to evaluate effects of different population densities of nematodes in two varieties of cotton (FMT 701 and IMACD 6001LL). The simultaneous occurrence of Pratylenchus brachyurus, Meloidogyne incognita and Rotylenchulus reniformis, contributed to reduced productivity and increased costs. Comparing the production in these areas was possible to observe losses on average, 23.1% and 35% in seed cotton yield, for cultivars IMACD 6001 LL and FMT 701, respectively, in infested areas

Biomanagement of root knot nematode through non edible oil seed cakes infesting cicer arietinum L.

A pot experiment was conducted to evaluate the efficacy of different oil cakes against the root knot nematode. Root knot nematodes are the major agricultural pest of wide range of crops worldwide. Chickpea, Cicer arietinum L., family Fabaceae, is a high yielding proteinaceous pulse crop with India being the leading producer. Root knot nematodes are one of the major biological constraint that reduces per capita yield of this pulse crops. In the present investigation, studies were made to determine the efficacy of different oil cakes viz. Azadirachta indica (Neem), Brassica campestris (Mustard) and Gossypium hirsutum (Cotton). Two doses (50 and 100 g) of each neem, mustard and cotton oil cakes were applied to reduce nematode infestation in chickpea. Results revealed that root gall development due to Meloidogyne incognita were significantly reduced in all the treatments and enhance all the plant growth characters of C.arietinum. Higher dose of neem oil cake was found to be most effective as compared to other treatments. Hence, it may be concluded that oil cakes are better substitute against nematicide for the effective control of root-knot nematode and reduce environmental hazards for ecologically safe environment.

The nematicidal effect of some bacterial biofertilizers on Meloidogyne incognita in sandy soil

In a greenhouse experiment, the nematicidal effect of some bacterial biofertilizers including the nitrogen fixing bacteria (NFB) Paenibacillus polymyxa (four strains), the phosphate solubilizing bacteria (PSB) Bacillus megaterium (three strains) and the potassium solubilizing bacteria (KSB) B. circulans (three strains) were evaluated individually on tomato plants infested with the root-knot nematode Meloidogyne incognita in potted sandy soil. Comparing with the uninoculated nematode-infested control, the inoculation with P. polymyxa NFB7, B. megaterium PSB2 and B. circulans KSB2, increased the counts of total bacteria and total bacterial spores in plants potted soil from 1.2 to 2.6 folds estimated 60 days post-inoculation. Consequently, the inoculation with P. polymyxa NFB7 increased significantly the shoot length (cm), number of leaves / plant, shoot dry weight (g) / plant and root dry weight (g) / plant by 32.6 percent, 30.8 percent, 70.3 percent and 14.2 percent, respectively. Generally, the majority treatments significantly reduced the nematode multiplication which was more obvious after 60 days of inoculation. Among the applied strains, P. polymyxa NFB7, B. megaterium PSB2 and B. circulans KSB2 inoculations resulted in the highest reduction in nematode population comparing with the uninoculated nematode-infested control. They recorded the highest reduction in numbers of hatched juveniles/root by 95.8 percent, females/root by 63.75 percent and juveniles/1kg soil by 57.8 percent. These results indicated that these bacterial biofertilizers are promising double purpose microorganisms for mobilizing of soil nutrients (nitrogen, phosphate and potassium) and for the biological control of M. incognita.

A New Technique for Single Spore Isolation of Two Predacious Fungi Forming Constricting Ring

A new technique for single spore isolation was developed for predacious fungi forming constricting rings directly on the spores using Dactylaria brochopaga and Arthrobotrys dactyloides. Constricting rings were induced directly on the spores by transferring the spores in 25 ppm solution of DL-Valine in sterile distilled water. Freshly hatched and thoroughly washed second stage juveniles of Meloidogyne incognita were transferred into cavity blocks containing induced rings for trapping and killing of nematodes. The killed nematodes were surface sterilized with streptomycin and inoculated into petri dishes containing maize meal agar media with 100 ppm streptomycin. The petri dishes were incubated at 29+/-1degrees C for few days which yielded axenic culture of these fungi.