J2 Hatching Research Articles

Phytochemical Analysis and Binding Interaction of Cotton Seed Cake Derived Compounds with Target Protein of Meloidogyne incognita for Nematicidal Evaluation.

The root-knot nematode Meloidogyne incognita is one of the most damaging plant-parasitic nematodes and is responsible for significant crop losses worldwide. Rising human health and environmental concerns have led to the withdrawal of commonly used chemical nematicides. There has been a tremendous demand for eco-friendly bio-nematicides with beneficial properties to the nematode hosting plants, which encourages the need for alternative nematode management practices. The current study was undertaken to determine the nematicidal potential of cotton seed cake (CSC) against second-stage juvenile (J2) hatching, J2 mortality, and J2 penetration of M. incognita in tomato plants in vitro. J2s and egg masses of M. incognita were exposed to four concentrations (250, 500, 750, and 1000 mg/L) of CSC extracts. The higher J2 mortality and inhibition of J2 hatching were found at 1000 mg/L, while the least effective result was observed at 250 mg/L of the CSC extract. The CSC extract applied with the concentrations mentioned above also showed inhibition of J2 penetration in tomato roots; 1000 mg/L showed the highest inhibition of penetration, while 250 mg/L displayed the least inhibition. Using gas chromatography-mass spectroscopy, we identified 11 compounds, out of which 9,12-Octadecadienoic acid, Hexadecanoic acid, and Tetradecanoic acid were found as major compounds. Subsequently, in silico molecular docking was conducted to confirm the nematicidal behavior of CSC based on binding interactions of the above three major compounds with the targeted protein acetylcholine esterase (AChE) of M. incognita. The values of binding free energy are -5.3, -4.5, and -4.9 kcal/mol, observed for 9,12-Octadecadienoic acid, n-Hexadecanoic acid, and Tetradecanoic acid, respectively, suggesting that 9,12-Octadecadienoic acid binds with the receptor AChE more efficiently than the other two ligands. This study indicates that CSC has nematicidal potential that can be used to control M. incognita for sustainable agriculture.

Nematicidal potential of Macrotyloma axillare 'Java' extract against Meloidogyne javanica and its metabolic profile

Plants produce chemical compounds with high nematicidal activity and potential for large-scale application in nematode control. The purpose of this study was to identify chemical compounds in crude extract of Macrotyloma axillare 'Java' and investigate their inhibitory effects on Meloidogyne javanica . Crude methanolic extract was fractionated into hexane, dichloromethane, ethyl acetate , and hydromethanol fractions. Chemical characterization of dichloromethane, ethyl acetate, and hydromethanol fractions was performed by dereplication using high-resolution mass spectrometry. The inhibitory activity of samples on the hatching of second-stage juveniles (J2) was tested at concentrations of 0%, 0.05%, 0.5%, and 5% (0, 500, 5000, 50000 ppm) and on nematode penetration and reproduction on soybean at 0.1% (1000 ppm). Distilled water was used as control in nematicidal assays. Several compounds belonging to the groups of organic acids, phenolic acids , fatty acids, flavonoids , pterocarpans, terpenoids, and sugars were identified. All fractions and concentrations reduced J2 hatching, reaching a plateau at 0.05%. Crude methanolic extract and dichloromethane and ethyl acetate fractions decreased M. javanica penetration at 10 days after inoculation, and all fractions reduced nematode penetration and female numbers at 30 days after inoculation. Extract fractions also reduced total nematode numbers, providing 49–95% control of the nematode. M. axillare 'Java' extract is rich in nematicidal compounds and has toxic effects on M. javanica , inhibiting J2 hatching and nematode penetration, development, and reproduction on soybean. • Macrotyloma axillare 'Java' is a plant that can be used to control nematodes. • 'Java' extract reduces penetration, development and reproduction of Meloidogyne javanica in soybean. • 135 compounds were identified in the 'Java' extract. • The chemical compounds of 'Java' show toxicity to Meloidogyne javanica .

Resistance induction and nematicidal activity of certain monoterpenes against tomato root-knot caused by Meloidogyne incognita.

This research was performed to evaluate the potential of carvone, cuminaldehyde, cineole, and linalool for the control of root-knot of tomato. The tested control agents were evaluated for their ability to stimulate systemic resistance to Meloidogyne incognita in tomato by monitoring the transcription levels of defense-related genes. Moreover, the ability of the tested agents to induce nematicidal activity concerning second-stage juveniles (J2) hatching and mortality was evaluated. Furthermore, the effect of the tested agents on certain tomato growth and yield parameters was assessed. The tested monoterpenes showed high nematicidal activity against M. incognita concerning J2 hatching inhibition and mortality. Carvone, cuminaldhyde, linalool, and cineole had LC50 values of 123.5, 172.2, 354.9, 466.4, and 952.3 μg/mL, respectively. Carvone was found to be the most efficient hatching inhibitor. The tested monoterpenes showed a high potential against root-knot under greenhouse and field conditions with respect to root-galling, egg masses, and the number of J2. Carvone was the most effective treatment. The growth and yield characters of treated tomato were significantly increased in monoterpenes treatments compared to untreated control. Treated tomato plants showed expression of defense-related genes (PR1 and PAL) 5-8 folds higher than the control. The results also showed that cuminaldhyde, followed by carvone, linalool, and cineole, had the greatest levels of expression in tomato plants. Taken together, the selected monoterpenes could be used as alternatives to control the root-knot of tomato.

Isolation and characterisation of nematicidal compound, leolorin C, from Leonotis leonurus acetone leaf extract

Ethnopharmacological relevanceLeonotis leonurus (L.) R.Br. (Lamiaceae) is a perennial shrub native to South Africa used to treat various diseases including digestive tract problems, intestinal worms and constipation. Aim of the studyThe aim was to isolate and characterise nematicidal compounds from leaves of L. leonurus. Materials and methodsBioassay-guided fractionation was carried out using the free-living nematode Caenorhabditis elegans as a model organism. Structural elucidation of the purified compound was carried out using NMR spectroscopic analyses and UPLC-QTOF-MS. The fractions and the isolated compound were tested for nematicidal activity on motility of plant-parasitic Meloidogyne incognita juveniles (J2s) and J2 hatch inhibition. Further screening was done to determine the minimum inhibitory concentration (MIC) of the fractions against bacterial phytopathogens and cytotoxicity against Vero kidney cells. ResultsLeoleorin C isolated from L. leonurus had moderate activity against C. elegans juveniles (34%) but was not active against J2 motility and J2 hatch of M. incognita. Thus, activity against the free-living C. elegans did not correspond with efficacy against plant-parasitic nematodes. Leoleorin C was not active against the tested bacterial phytopathogens, but some activity was observed in the bioautography assay against Clavibacter michiganensis subsp. michiganensis, the organism causing bacterial canker in tomatoes. The plant extract, fractions and leolorin C were relatively non-toxic to Vero cells with LC50 values greater than 0.01 mg/mL. ConclusionThe crude extract of L. leonurus and fractions may be useful in developing complementary treatments for controlling nematodes and phytopathogens. This study does not support the use of free-living nematodes as a model to isolate anti-parasitic compounds from plants.

Effect of salt stress on the parasitism of Meloidogyne enterolobii in cowpea

Cowpea, Vigna unguiculata (L.) Walp, is a very important Fabaceae in the diet of families in the northeast region of Brazil. This plant species is among the different cultures affected by the root-knot nematode. In this region, in addition to phytonematodes, another condition that affects cowpea productivity is salinity, one of the main abiotic limiting factors. The cultivar Pitiúba, however, has good adaptability to salt stress conditions. However, information on the behavior of root-knot nematodes in an environment with high levels of salinity for this crop is still scarce. Based on the above, the objective of this work was: 1) to evaluate the hatchability of J2 of Meloidogyne enterolobii in 100mM NaCl solution; 2) evaluate the motility and infectivity of J2 of M. enterolobii in saline solution at the levels of 0, 25, 50, 75, 100, 125, 150, 175, 200, 300, and 400mM; 3) to investigate the impairment of the development of M. enterolobii in cowpea Pitiúba under irrigation with NaCl solution at 100mM. It was found that J2 hatching, motility, and infectivity of juveniles of M. enterolobii were not affected by the levels of salinity tested in vitro. In Pitiúba, irrigation with 100 mM saline solution considerably affected the development of juveniles of the pathogen in the roots.

Can vermicompost and biochar control Meloidogyne javanica on eggplant?

Summary The root-knot nematode, Meloidogyne javanica, is the most damaging species of plant-parasitic nematodes in eggplant cultivation in Iran. We studied the effect of pistachio and date palm biochars, vermicompost and a mixture of each of these biochars with vermicompost on M. javanica. To investigate the effect of these organic materials on survival, hatching and attraction of the second-stage juvenile (J2) towards the root, bioassays were performed in the laboratory using extracts of organic material. Vermicompost extract increased J2 mortality and decreased hatching and the number of J2 attracted to the eggplant roots. However, either type of biochar alone or mixed with vermicompost did not cause J2 mortality, nor did they inhibit hatching and attraction of J2 towards the root. To determine the effect of organic matter on J2 invasion and reproduction, pot experiments were performed in a completely randomised design with four replications in the glasshouse. Vermicompost reduced the number of J2 that penetrated the roots, number of egg masses and the final population of M. javanica eggs and J2. The suppressing effect of vermicompost might be attributed to the release of toxic compounds such as ammonium and improved soil nutrient and plant growth, leading to plants more tolerant to nematode damage. Pistachio and date biochars alone or combined with vermicompost had no adverse effect on the nematode reproduction compared to non-treated soil. Both types of biochar reduced the suppressing effect of vermicompost, most probably due to the absorbance of suppressive compounds in vermicompost by the high surface area of the biochar.

Volatile fatty acids from whey volatilome as potential soil fumigants to control Meloidogyne incognita

Organic residues emit volatile organic compounds (VOCs) that are toxic to plant-parasitic nematodes (PPNs). In this study, we demonstrated that whey produced by cow milk emit VOCs that are toxic to Meloidogyne incognita second-stage juveniles (J2), causing immobility above 80% and significant (P < 0.05) mortality compared to controls. The analysis of the volatilome done by gas chromatography coupled with mass spectrometry revealed 28 compounds, predominantly carboxylic acids and esters. Four of those volatile fatty acids (VFAs), namely acetic acid, octanoic acid, ethyl octanoate, and isovaleric acid, were chosen to further studies. The four tested VFAs were toxic to M. incognita J2, with lethal concentration values required to kill 50% of the nematode population (LC50), ranging from 134.30 to 236.08 μg mL−1. However, on eggs, only acetic acid and ethyl octanoate were consistent with J2 hatch inhibition, reaching 80% at a concentration of 1000 μg mL−1. In a greenhouse assay, when the VFAs were applied as soil fumigant, M. incognita infectivity and reproduction were significantly (P < 0.05) reduced compared to the negative control (water). Among them, ethyl octanoate application reduced (P < 0.05) the number of eggs to the level of the commercial fumigant dazomet. Overall, the volatile compounds released by whey were toxic to M. incognita J2 and contained a great diversity of molecules. Among the VFAs, ethyl octanoate stood out and showed the potential to be used in future field studies.

Effects of temperature and duration of storage on the hatching behaviour ofHeteroderalatipons (Nematoda: Heteroderidae)

Mediterranean cereal cyst nematode (CCN) Heterodera latipons is considered as one of the main CCN that causes significant economic losses in the production of intolerant cereal crops in Turkey. Accordingly, this study aimed at evaluating the effect of different storing temperatures and durations on the emergence of second-stage juveniles (J2s) of the nematode populations obtained from the eastern Mediterranean (Adana and Hatay) and south-eastern (Gaziantep and Kilis) regions of Turkey. The obtained results revealed a variation in the J2s hatching profiles of the regional nematode populations. The eastern-Mediterranean and the south-eastern populations hatched at between 5 and 20 °C; however, considerable divergences in the J2 hatching were noticed. The highest emergence of J2s for the populations occurred at 10 °C, whereas the lowest level was observed at 5 °C. To simulate the preplanting soil temperatures in Turkey, the cysts were stored at 5 °C and 20 °C, before incubation at 10 and 15 °C, to stimulate hatching of the J2s. The highest cumulative hatching was obtained at a constant temperature of 10 °C, while the lowest cumulative hatching occurred at 5 °C. Moreover, storing cysts at 5 °C, before incubation at 10 and 15 °C, significantly stimulated hatching of the populations of H. latipons when compared to the control. However, storing the cysts at 20 °C, before incubation at 10 °C, substantially stimulated the emergence of J2s relative to the control. The obtained results about the hatching behaviour of H. latipons may help in the development of effective control strategies for this nematode in Turkey.

The Use of Essential Oil and Hydrosol Extracted from Cuminum cyminum Seeds for the Control of Meloidogyne incognita and Meloidogyne javanica

The essential oil (EO) and hydrosol (HL) isolated from Cuminum cyminum (cumin) seeds were evaluated against the root-knot nematodes Meloidogyne incognita and M. javanica. The efficacy of extracts on the motility, hatching, and survival in soil of second-stage juveniles (J2s), and the activity on egg differentiation were tested. All J2s were paralyzed after immersion in the EO at 62.5 μL/L concentration for 96 h. Encouraging results were recorded using HL equal to or higher than 10% concentration for both Meloidogyne species tested. More than 70% paralyzed J2s were recorded after immersion for 48 h, while the percentage was increased to higher than 90% after 96 h of immersion. A clear effect on egg differentiation was observed after immersion in EO or HL. A significant decrease in egg differentiation was revealed at even low concentrations of EO while an evident decrease in egg differentiation was recorded after immersion of eggs in 50% HL dilution. Decreased hatching of M. incognita and M. javanica J2s was observed with the increase in concentration. The lowest numbers of hatched J2s were recorded when EO was used at 1000 and 2000 μL/L concentrations. A constant reduction in root-knot nematode J2 hatching was observed upon increasing the concentration of HL from 5% to 50%. The EO of C. cyminum is characterized by the presence of γ-terpinene-7-al (34.95%), cumin aldehydes (26.48), and α-terpinene-7-al (12.77%). The above constituents were observed in HL following the same order as that observed in EO. The components γ-terpinene (11.09%) and ο-cymene (6.56%) were also recorded in EO while they were absent in HL.

Hatching of Globodera pallida Induced by Root Exudates Is Not Influenced by Soil Microbiota Composition.

Plant-parasitic nematodes are among the most harmful pests of cultivated crops causing important economic losses. The ban of chemical nematicides requires the development of alternative agroecological approaches to protect crops against nematodes. For cyst nematodes, egg hatching is stimulated by host plant root exudates. Inducing “suicide hatching” of nematode second-stage juveniles (J2), using root exudates in the absence of the host plant, may constitute an effective and innovative biocontrol method to control cyst nematodes. However, before considering the development of this approach, understanding the effect of soil biotic component on cyst nematode hatching by root exudates is a major issue. The effectiveness of this approach could be modulated by other soil organisms consuming root exudates for growth as soil microbiota, and this must be evaluated. To do that, four different native agricultural soils were selected based on their physicochemical properties and their microbiota composition were characterized by rDNA metabarcoding. To disentangle the effect of microbiota from that of soil on hatching, four recolonized artificial soils were obtained by inoculating a common sterile soil matrix with the microbiota proceeding from each agricultural soil. Each soil was then inoculated with cysts of the potato cyst nematode, Globodera pallida, and low or high doses of potato root exudates (PREs) were applied. After 40 days, viable J2 remaining in cysts were counted to determine the efficiency of root exudates to stimulate hatching in different soils. Results showed that (i) when physicochemical and microbiota compositions varied among native soils, the hatching rates remained very high albeit small differences were measured and no dose effect was detected and (ii) when only microbiota composition varied among recolonized soils, the hatching rates were also high at the highest dose of PREs, but a strong dose effect was highlighted. This study shows that abiotic and biotic factors may not compromise the development of methods based on suicide hatching of cyst nematodes, using root exudates, molecules inducing J2 hatch, or trap crops.

Application forms and mode of action of biocontroller in the management of Meloidogyne incognita in tomato

ABSTRACT Mixture of antagonist agents and form of application may influence in the control of root-knot nematodes. Thus, the aim of this work was to study the action of a biological product based on enzyme mixtures, Bacillus sp. and Trichoderma sp., at different concentrations on hatching, motility, mortality and reproduction of Meloidogyne incognita in tomato, considering two ways of application of the product. Eggs and second-stage juveniles (J2) of M. incognita were placed in biological product solutions at concentrations: 0, 1.25, 2.5, 5, 10, 20 and 40 g L-1. The same concentrations were applied to planting pits or to the soil surface. Afterwards, 3,692 eggs of M. incognita were inoculated in tomato crop. A significant reduction in J2 hatching of M. incognita J2 was observed in the highest concentrations and higher mortality of J2 from concentration of 5 g L-1. The interaction between concentrations and form of application of the product significantly influenced the infectivity and reproduction of M. incognita. Greater root system mass was obtained by applying the biological product to the surface, regardless of concentration.

Activity against Meloidogyne incognita of volatile compounds produced during amendment of soil with castor bean cake

Summary Incorporation of castor bean cake into the soil results in the emission of four main compounds: skatole, γ-decalactone, 4-methylphenol and phenol. The toxic effects of these compounds on the life cycle of Meloidogyne incognita were studied here. Only phenol did not reduce hatching of M. incognita second-stage juveniles (J2) at any of the concentrations tested. The other compounds reduced J2 hatching by 42.1 to 59.3% at the highest concentrations. When J2 were immersed in Tween 80® solutions of each compound, at their respective LC50 values and inoculated into soil with tomato plants, reductions in the numbers of galls (48.5-69.3%) and eggs (42.8-62.5%) were observed in comparison with the control; this result was similar to that of carbofuran. The highest reductions in the numbers of galls (21.8-88.5%) and eggs (49.6-98.9%) occurred with all compounds when the highest concentration (500 mg l−1) was applied to the substrate at the moment of J2 inoculation. Volatile compounds of γ-decalactone and skatole failed to cause J2 immobility or mortality. However, phenol caused 95.8% and 4-methylphenol caused 100% mortality. With fumigation, the highest concentration (1000 mg (l substrate)−1) of any tested compound reduced gall numbers at a similar rate to the nematicide dazomet (98.7-100%) and reduced egg numbers (87.1-99.7%). The volatile compounds skatole, γ-decalactone, 4-methylphenol and phenol have a nematicidal effect on the life cycle of M. incognita and alter its pathogenic behaviour on the plant.

Effects of yeast fermentation broths on the Meloidogyne incognita population in soybean

Root-knot nematodes are pathogenic to a wide range of plants worldwide and can reduce the yield potential of important crops. The objective of this study was to investigate the potential effects of yeast fermentation broths (YFBs) on the hatching of Meloidogyne incognita second-stage juveniles (J2s) in vitro. In addition, application methods of the yeasts for the control of the nematode in soybean were evaluated. Fermentation broths (FBs) of Pichia manshurica CCMA0762, Pichia caribbica CCMA0759, Lachancea thermotolerans CCMA0763, Hanseniaspora opuntiae CCMA0760 and Kodamaea (Pichia) ohmeri CCMA0758 were evaluated in the hatching test in vitro. The same treatments were separately applied to the soil, soil+shoot and shoots of plants inoculated with M. incognita, and the results were compared with those of the control and those from a yeast-based commercial product. L. thermotolerans CCMA0763 and P. caribbica CCMA0759 FBs were selected for evaluation of nematode infection. All YFBs inhibited J2 hatching. The L. thermotolerans FB reduced the number of nematodes in one experiment when applied to soil. The most effective application mode differed by FB type. L. thermotolerans and P. caribbica FBs, when applied to shoots, decreased the number of J2s, sausage-shaped juveniles, adult females and total nematodes at different evaluation times.

Microbes Attaching to Endoparasitic Phytonematodes in Soil Trigger Plant Defense Upon Root Penetration by the Nematode.

Root-knot nematodes (Meloidogyne spp.) are among the most aggressive phytonematodes. While moving through soil to reach the roots of their host, specific microbes attach to the cuticle of the infective second-stage juveniles (J2). Reportedly, the attached microorganisms affect nematodes and reduce their performance on the host plants. We have previously shown that some non-parasitic bacterial strains isolated from the cuticle of Meloidogyne hapla in different soils affected J2 mortality, motility, hatching, and root invasion. Here we tested whether cuticle-attached microbes trigger plant defenses upon penetration of J2. In in vitro assays, M. hapla J2-attached microbes from a suppressive soil induced pathogen-associated molecular pattern-triggered immunity (PTI) in tomato roots. All tested PTI-responsive defense genes were upregulated after root invasion of J2 with attached microbes, compared to surface-sterilized J2, particularly the jasmonic acid-mediated PTI marker genes TFT1 and GRAS4.1. The strain Microbacterium sp. K6, that was isolated from the cuticle, significantly reduced root invasion when attached to the J2. Attached K6 cells supported plant defense and counteracted suppression of plant basal defense in roots by invaded J2. The plant response to the J2-attached K6 cells was stronger in leaves than in roots, and it increased from 1 to 3 days post inoculation (dpi). At 1 dpi, the plant responded to J2-attached K6 cells by ameliorating the J2-triggered down-regulation of defense genes mostly in roots, while at 3 dpi this response was systemic and more pronounced in leaves. In a reactive oxygen species (ROS) assay, the compounds released from J2 with attached K6 cells triggered a stronger ROS burst in tomato roots than the compounds from nematodes without K6, or the metabolites released from strain K6 alone. Leaves showed a 100 times more sensitive response than roots, and the metabolites of K6 with or without J2 induced strong ROS bursts. In conclusion, our results suggest the importance of microorganisms that attach to M. hapla in suppressive soil, inducing early basal defenses in plants and suppressing nematode performance in roots.

Effect of Heterodera schachtii female age on susceptibility to three fungal hyperparasites in the genus Hyalorbilia.

Three closely related nematophagous fungi in the genus Hyalorbilia were compared for their ability to parasitize females and eggs of Heterodera schachtii at different developmental stages. DoUCR50, StM, and ARF were originally isolated from Heterodera schachtii, Meloidogyne incognita, and Heterodera glycines, respectively. Phylogenetic analysis and pairwise sequence analysis showed that DoUCR50 and StM are more closely related to each other than they are to ARF. DoUCR50 parasitism suppressed 100% of the J2 hatch from 3-week-old H. schachtii females and 75% of the hatch from 4-week-old females. Eggs within 5-week-old females were resistant to parasitism, and hatch of J2 was unaffected by exposure to DoUCR50. StM and ARF did not reduce the hatch of J2 from H. schachtii females of any age. Eggs removed from females and spread onto water agar cultures of the fungi were mostly resistant to parasitism. DoUCR50 parasitized only 16% of such eggs from 3-week-old females. Extracellular hydrolytic enzyme production by the three fungal strains grown on PDA or parasitized H. schachtii females was evaluated using API ZYM (bioMérieux) test strips. All three fungi produced extracellular hydrolytic enzymes when grown on PDA or H. schachtii females. Trypsin-like protease activity was uniquely detected in DoUCR50 grown on PDA and H. schachtii females, with the highest activity associated with the fungus grown on parasitized females.

Density-dependent growth response and sensitivity of Meloidogyne incognita to Maerua angolensis and Tabernaemontana elegans: Card model

ABSTRACTCurve-fitting Allelochemical Response Dosage (CARD) computer-based model has proven to be a valuable tool in the definition of dosage response relationships between organisms and various plant extracts. Its use in the explanation of the relationship between Maerua angolensis and Tabernaemontana elegans concentrations and Meloidogyne incognita second-stage juvenile (J2) hatch and mortality has not been reported. The objective of this study was to use the CARD model in the determination of M. incognita sensitivity, density-dependent growth (DDG) patterns and minimum inhibition concentrations of M. angolensis and T. elegans plant extracts on juvenile hatch and mortality. The CARD model quantified concentration ranges of the two plant extracts that could stimulate (Dm − Rh), saturate (Rh − D0) and inhibit (D0 − D100), J2 hatch and mortality. Meloidogyne incognita J2 hatch was moderately sensitive to M. angolensis and T. elegans plant extracts with average sensitivity ranking of 20 and 19 units, respectively. J2 mortalities were moderately sensitive to T. elegans average sensitivity ranking of 15 units, whereas highly sensitive to M. angolensis at 2 units. The minimum juvenile hatch inhibition and lethal concentrations for the two extracts were comparable and very low at 0.07, 0.06 and 0.13, 094 mg mL−1, respectively. In conclusion, the low minimum concentrations for M. incognita J2 hatch inhibition and lethal concentrations of M. angolensis and T. elegans plant extracts and moderate sensitivity values provide substantial evidence of high potency of the two extracts in the potential management of M. incognita.

Nematicidal screening of essential oils and potent toxicity of Dysphania ambrosioides essential oil against Meloidogyne incognita in vitro and in vivo

AbstractIt is known that some plant essential oils have pesticide activities. Among the 29 oils evaluated in this study, 14 showed nematicidal activities of 8 to 100% at the concentration of 1,000 μg/ml, compared with a control of 0.01 g/ml Tween 80®. At a lower concentration of 500 μg/ml, only Dysphania ambrosioides oil caused &gt;90% mortality of second‐stage juveniles (J2) of Meloidogyne incognita. The LC50 and LC95 values for D. ambrosioides oil were 307 μg/ml and 580 μg/ml, respectively. M. incognita eggs placed in D. ambrosioides oil solutions had a significant reduction in J2 hatching compared with controls. Therefore, the oil had a toxic effect on both eggs and J2 of M. incognita. This was in contrast to nematicides on the market that act efficiently only on J2. When J2 were placed in D. ambrosioides oil at its LC50 concentration and inoculated onto tomato plants, the reduction in numbers of galls and eggs was 99.5% and 100%, respectively. Dysphania ambrosioides oil applied to the potting substrate of plants at a concentration of 1,100 μg/ml significantly reduced the number of galls and eggs of M. incognita, whereas a concentration of 800 μg/ml only reduced the number of eggs compared with the controls (Tween 80® and water). The main components of the D. ambrosioides oil detected by gas chromatography–mass spectrometry were (Z)‐ascaridole (87.28%), E‐ascaridole (8.45%) and p‐cymene (3.35%), representing 99.08% of the total oil composition. Given its nematicidal activity, D. ambrosioides oil represents an exciting raw material in the search for new bioactive molecules for the pesticide industry.

Sensitivity of Meloidogyne incognita second-stage juvenile hatch, motility and viability to pure cucurbitacins and cucurbitacin-containing phytonematicides

Previous claims suggested that pure active ingredients from botanicals were less effective in pest management than their fermented crude-extracts. The objective of this study was to compare the toxicity of pure (98%) cucurbitacin A and B on nematode bioactivities with those of their fermented crude-extracts, the Nemarioc-AL and Nemafric-BL phytonematicides, respectively. Purified active ingredients were each diluted to 0.00, 0.25, 0.50, 0.75, 1.00, 1.25, 1.50, 1.75, 2.00, 2.25 and 2.50 μg ml‒1 and their fermented crude-extracts to 0.0%, 0.5%, 1.0%, 1.5%, 2.0%, 2.5%, 3.0%, 3.5%, 4.0%, 4.5% and 5.0% phytonematicide. The exposure time for second-stage juveniles (J2) hatch, motility and viability of the J2 to each concentration was 24, 48 and 72 h. The overall sensitivities (Σk) of J2 hatch to Nemarioc-AL, cucurbitacin A, Nemafric-BL and cucurbitacin B were 1, 30, 5 and 2 units, respectively, for motility 7, 12, 2 and 12 units, respectively, and for viability 2, 4, 1 and 4 units, respectively. Generally, nematodes displayed high sensitivities to cucurbitacin-containing phytonematicides than to their respective purified active ingredients (cucurbitacins). In conclusion, the cucurbitacin-containing phytonematicides were more effective in nematode suppression than their purified active ingredients.

Nematicidal Weeds, Solanum nigrum and Datura stramonium.

We investigated Solanum nigrum (seeds) and Datura stramonium (shoots) against root-knot nematodes in terms of J2 paralysis and egg hatch inhibition (methanol extract), as well as inhibition of nematode development in host roots (soil amending with either S. nigrum seeds' or D . stramonium shoots' meal). Datura stramonium was found equally effective at inhibiting motility of Meloidogyne incognita and Meloidogyne javanica (both EC 50 = 427 μg mL -1 at 3 day), but inhibition occurred more quickly for M. incognita (1 day). Solanum nigrum was faster and more effective at inhibiting motility of M. incognita than M. javanica (EC 50 = 481 and 954 μg mL -1 at 3 day, respectively). Datura stramonium was slower, but eventually more potent in decreasing egg hatch and cell division in M. incognita eggs, than S. nigrum . Specifically, D. stramonium significantly inhibited cell division in eggs immersed in at least 100 and 1 μg mL -1 at Day 6 and 10, respectively. Solanum nigrum impeded cell division in un-differentiated eggs immersed in not less than 10 and 100 μg mL -1 after days 2 and 6, respectively. Both extracts were similar in suppressing J2 exclosure but D. stramonium was effective in smaller test concentrations. Specifically, D. stramonium suppressed J2 emerging from eggs immersed in 10 μg mL -1 at day2, and in at least 1 μg mL -1 at day 6. Solanum nigrum significantly reduced J2 hatch from eggs immersed in a minimum of 100 μg mL -1 at day 2 and not less than 1,000 μg mL -1 at day 6. In pots, powdered S. nigrum seeds meal was more active than D. stramonium and the respective EC 50 females/ g values for M. incognita were 1.13 and 11.4 mg g -1 of soil, respectively. The chemical composition of active extracts was determined after derivatization by GC-MS. Chemical analysis of active extracts showed the presence of fatty acids with known nematicidal activity.

Extracts and semi-purified fractions of Tagetes patula flowers in the control of root-knot nematodes

The genus Tagetes constitutes a group of antagonistic plant species that are cytotoxic against plant pathogenic nematodes, with T. patula being particularly efficient. The aim of this study was to evaluate the in vitro effect of extracts and semi-purified fractions of T. patula flowers on eggs and second-stage juveniles (J2) of Meloidogyne incognita, M. javanica, and M. paranaensis, as well as to verify the nematicidal effect of the flavonoids present in T. patula flowers. Extracts and semi-purified fractions were obtained from dried T. patula flowers after maceration and liquidification, yielding crude aqueous (CAE) and crude ethanol-water (CEWE) extracts. Dried flowers were also treated with n-hexane to obtain a crude defatted aqueous extract (CDAE) and a crude ethanol-water defatted extract (CEWDE). Then, the CEWE was fractionated, and the ethyl acetate (EAF), ethanol (EF), methanol (MF), and ethanol:water (EWF) fractions were obtained. CAE, CEWE, CDAE, and CEWDE were tested to evaluate their effects on hatching, mobility, and mortality of J2 of M. incognita, M. javanica, and M. paranaensis. EAF, EF, MF, and EWF fractions were tested on the same variables of M. incognita. All extracts significantly reduced J2 hatching of M. incognita, M. javanica, and M. paranaensis when compared to water and water + DMSO. CEWE had nematicidal effects on the three evaluated species, whereas CEWDE demonstrated nematicidal effects against M. incognita and M. javanica, and nematostatic effects on M. paranaensis. This toxic effect showed by CEWE may be related to the high content of quercetin, a major substance present in this sample. It was also observed that EAF accentuated the nematicidal response on Meloidogyne spp., suggesting that other medium polarity (methoxylated) flavonoids act as nematotoxic substances. Thus, these results suggest that quercetin contributes significantly to the nematicidal activity of CEWE and EAF.