Nematicidal Activity of 20-Deoxyingenol-3-angelate from Euphorbia peplus Latex Through Protein Kinase C Isotype TPA-1.

The latex of Euphorbia peplus and its major component 20-deoxyingenol-3-angelate (DI3A) displayed significant nematicidal activity against Caenorhabditis elegans and Panagrellus redivivus. DI3A treatment inhibited the growth and development of nematodes and caused significantly negative effects on locomotion behavior, reproduction, and accumulation of reactive oxygen species. Transcriptome analysis indicated that differential expression genes in DI3A-treated C. elegans were mainly associated with the metabolism, growth, and development process, which were further confirmed by RT-qPCR experiments. The expression level of TPA-1 gene encoding a protein kinase C isotype was obviously upregulated by DI3A treatment, and knockdown of TPA-1 by RNAi technology in the nematode could relieve the growth-inhibitory effect of DI3A. Metabolic analysis indicated that DI3A was hardly metabolized by C. elegans, but a glycosylated indole derivative was specifically accumulated likely due to the activation of detoxification. Overall, our findings suggested that DI3A from E. peplus latex exerted a potent nematicidal effect through the gene TPA-1, which provides a potential target for the control of nematodes and also suggests the potential application value of E. peplus latex and DI3A as botanical nematicides.

Harnessing nature's arsenal: Ochrobactrum bacteria metabolites in the battle against root- knot nematode - Insights from in vitro and molecular docking studies.

Agricultural Productivity and plant health are threatened by the root-knot nematode. The use of biocontrol agents reduces the need for chemical nematicides and improves the general health of agricultural ecosystems by offering a more environmentally friendly and sustainable method of managing nematode infestations. Plant-parasitic nematodes can be efficiently managed with the use of entomopathogenic nematodes (EPNs), which are widely used biocontrol agents. This study focused on the nematicidal activity of the secondary metabolites present in the bacteria Ochrobactrum sp. identified in the EPN, Heterorhabditisindica against Root-Knot Nematode (Meloidogyne incognita). Its effect on egg hatching and survival of juveniles of root- knot nematode (RKN) was examined. The ethyl acetate component of the cell-free culture (CFC) filtrate of the Ochrobactrum sp. bacteria was tested at four different concentrations (25 %, 50 %, 75 % and 100 %) along with broth and distilled water as control. The bioactive compounds of Ochrobactrum sp. bacteria showed the highest suppression of M. incognita egg hatching (100 %) and juvenile mortality (100 %) at 100 % concentration within 24 h of incubation. In this study, unique metabolite compounds were identified through the Gas Chromatography- Mass Spectrometry (GC-MS) analysis, which were found to have anti- nematicidal activity. In light of this, molecular docking studies were conducted to determine the impact of biomolecules from Ochrobactrum sp. using significant proteins of M. incognita, such as calreticulin, sterol carrier protein 2, flavin-containing monooxygenase, pectate lyase, candidate secreted effector, oesophageal gland cell secretory protein and venom allergen-like protein. The results also showed that the biomolecules from Ochrobactrum sp. had a significant inhibitory effect on the different protein targets of M. incognita. 3-Epimacronine and Heraclenin were found to inhibit most of the chosen target protein. Among the targets, the docking analysis revealed that Heraclenin exhibited the highest binding affinity of -8.6 Kcal/mol with the target flavin- containing monooxygenase. Further, the in vitro evaluation of 3- Epimacronine confirmed their nematicidal activity against M. incognita at different concentrations. In light of this, the present study has raised awareness of the unique biomolecules of the bacterial symbiont Ochrobactrum sp. isolated from H. indica that have nematicidal properties.

Exploiting fly ash as an ecofriendly pesticide/nematicide on Abesmoschus esculuntus: Insights into soil amendment-induced antioxidant fight against nematode mediated ROS.

Conventional pest control measures, such as chemical pesticides and nematicides, have limited efficacy and raise environmental concerns, necessitating sustainable and eco-friendly alternatives for pest management. Therefore, to find a complementary eco-friendly pesticide/nematicide, this study investigated the role of fly ash (FA) in managing a notorious pest, Meloidogyne javanica and its impact on the growth and physiology of Abelmoschus esculentus. Molecular characterization using SSU and LSU rDNA gene markers confirmed the identity of Indian M. javanica as belonging to the same species. Biotic stress induced by nematode infection was significantly alleviated (P < 0.05) by FA application at a 20% w/v, regulating of ROS accumulation (44.1% reduction in superoxide anions and 39.7% reduction in hydrogen peroxide content) in the host plant. Moreover, FA enhanced antioxidant defence enzymes like superoxide dismutase (46.6%) and catalase (112%) to combat nematode induced ROS. Furthermore, the application of FA at a 20% concentration significantly improved the biomass and biochemical attributes of okra. Fly ash also upregulated the activity of the important osmo-protectant proline (11.5 µmol/g FW) to mitigate nematode stress in host cells. Suppression of disease indices like gall index and reproduction factor, combined with in-vitro experiments, revealed that FA exhibits strong nematode mortality capacity and thus can be used as a sustainable and eco-friendly control agent against root-knot nematodes.

Linking the protease activity to the nematicidal action of edible mushroom.

Biological control using edible mushrooms as natural enemies is a sustainable alternative for pest management. Despite the well-established literature on toxins and secondary metabolites produced by these fungi in the biochemical control of nematodes, the nematicidal activity of proteases from different Pleurotus species is yet to be investigated. Therefore, this study aimed to correlate protease to the nematicidal activity of different mushrooms, Pleurotus sp., P. ostreatus (SB), P. ostreatus (Pearl), and P. djamor. For such a purpose, we performed motility assays of Panagrellus sp. at different time intervals, 6, 12, and 24 h for each of the mushrooms. In addition, the protease activity was measured using different pH (5, 7, and 9) and fermentation time intervals (45 and 75 days). Furthermore, we also evaluated the effect of this cell-free extract on Panagrellus sp. In response to these experiments, all edible mushrooms showed a reduction over 82% for the nematode-feeding activity (p < 0.01). The cell-free crude extract of each of the fungi studied showed nematocidal activity (p < 0.01). For the 45-day fermentation, P. djamor exhibited statistical significance (p < 0.01) compared with the others, reaching a reduction percentage of 73%. For the 75-day fermentation, Pleurotus sp. and P. ostreatus (Pearl) showed significant differences compared with the other fungi (p < 0.01), with reduction percentages of 64 and 62%, respectively. Herein, protease activity was associated with the nematicidal action of different Pleurotus species in controlling Panagrellus sp.

Discovery and Mechanism of a Nematicide Candidate (W3): A Novel Amide Compound Containing a Cyclopropyl Moiety.

To find novel nematicides, we screened the nematicidal activity of compounds in our laboratory compound library. Interestingly, the compound N-((1R,2R)-2-(2-fluoro-4-(trifluoromethyl)phenyl)cyclopropyl)-2-(trifluoromethyl)benzamide (W3) showed a broad spectrum and excellent nematicidal activity. The LC50 values of compound W3 against second-stage juveniles of Bursaphelenchus xylophilus (B. xylophilus), Aphelenchoides besseyi, and Ditylenchus destructor are 1.30, 1.63, and 0.72 mg/L, respectively. Nematicidal activities of compound W3 against second-stage juveniles of Meloidogyne incognita were 87.66% at 100 mg/L. Meanwhile, compound W3 can not only observably inhibit the feeding, reproduction, and egg hatching of B. xylophilus but can also effectively promote the oxidative stress adverse reactions of nematodes and cause intestinal damage. Compound W3 can promote the production of MDA and inhibit the activities of defense enzymes SOD and GST in B. xylophilus. Compound W3 can affect the transcription of genes involved in regulating the tricarboxylic acid cycle in nematodes, resulting in weakened nematode respiration and reduced nematode activity and even death. In addition, compound W3 had good inhibitory activity against five pathogenic fungi. Among them, the EC50 of compound W3 against Fusarium graminearum was 8.4 mg/L. In the future, we will devote ourselves to the toxicological and structural optimization research of the candidate nematicide W3.

Rare NRPS Gene Cluster for Desferriferrichrome Biosynthesis Controls the Conflict between Trap Formation and Nematicidal Activity in Arthrobotrys oligospora.

The formation of the trapping device induced by nematodes has been assumed as an indicator for a switch from saprophytic to predacious lifestyles for nematode-trapping fungi. However, fungal nematocidal activity is not completely synonymous with fungal trap formation. We found that the predominant nematode-trapping fungus Arthrobotrys oligospora harbored a rare NRPS (Ao415) gene cluster that was mainly distributed in nematode-trapping fungi. The gene Ao415 putatively encodes a protein with a unique domain organization, distinct from other NRPSs in other fungi. Mutation of the two key biosynthetic genes Ao415 and Ao414 combined with nontarget metabolic analysis revealed that the Ao415 gene cluster was responsible for the biosynthesis of a hydroxamate siderophore, desferriferrichrome (1). Lack of desferriferrichrome (1) and its hydroxamate precursor (3) could lead to significantly increased Fe3+ content, which induced fungal trap formation without a nematode inducer. Furthermore, the addition of Fe3+ strongly improved fungal trap formation but deleteriously caused broken traps. The addition of 1 significantly attenuated trap formation but enhanced fungal nematicidal activity. Our findings indicate that iron is a key factor for trap formation and provide a new insight into the underlying mechanism of siderophores in nematode-trapping fungi.

Functional validation of novel levamisole resistance marker S168T in Haemonchus contortus.

Recently, a S168T variant in the acetylcholine receptor subunit ACR-8 was associated with levamisole resistance in the parasitic helminth Haemonchus contortus. Here, we used the Xenopus laevis oocyte expression system and two-electrode voltage-clamp electrophysiology to measure the functional impact of this S168T variant on the H. contortus levamisole-sensitive acetylcholine receptor, L-AChR-1.1. Expression of the ACR-8 S168T variant significantly reduced the current amplitude elicited by levamisole compared to acetylcholine, with levamisole changing from a full to partial agonist on the recombinant L-AChR. Functional validation of the S168T mutation on modulating levamisole activity at the receptor level highlights its critical importance as both a mechanism and a marker of levamisole resistance.

Chemo-profiling of Purpureocillium lilacinum and Paecilomyces variotii isolates using GC-MS analysis, and evaluation of their metabolites against M. incognita.

Nematophagous fungi are the best alternatives to chemical nematicides for managing nematodes considering environmental health. In the current study, activity of metabolites from ten isolates of Purpureocillium lilacinum (Thom) Luangsa-ard (Hypocreales: Ophiocordycipitaceae) and two isolates of Paecilomyces variotii Bainier (Eurotiales: Trichocomaceae), were examined to inhibit the hatching of Meloidogyne incognita (Kofoid & White) Chitwood (Tylenchida: Heteroderidae) eggs. At 100%, 50%, and 25% concentrations, respectively, the culture filtrate of the isolate P. lilacinum 6887 prevented 97.55%, 90.52%, and 62.97% of egg hatching. Out of all the isolates, Pl 6887, Pl 6553, and Pl 2362 showed the greatest results in the hatching inhibition experiment.Gas chromatography-mass spectrometry (GC-MS) analysis revealed a variety of nematicidal compounds from different isolates. A total of seven nematicidal compounds, including four very potent nematicidal fatty acids were found in the isolate Pl 6553. Secondary metabolites of the same isolate possess the highest M. incognita juvenile mortality, i.e., 43.33% and 92% after 48 hrs of treatment at 100 and 200 ppm concentrations, respectively. Significant difference was observed in juvenile mortality percentage among the isolate having highest and lowest nematicidal compounds. Nematicidal fatty acids like myristic and lauric acid were found for the first time in P. lilacinum. Multiple vacuole-like droplets were found inside the unhatched eggs inoculated with the culture filtrate of isolate Pl 6887, and also in the juveniles that perished in the ethyl acetate extract of isolate Pl 6553.

Nematicidal activity of paucimannose-type glycoconjugates from acacia honey.

Natural honey contains glycoconjugates as minor components. We characterized acacia honey glycoconjugates with molecular masses in the range of 2-5 kDa. The glycoconjugates were separated by RP-HPLC into three peaks (termed RP-2-5 k-I, RP-2-5 k-II, and RP-2-5 k-III) which demonstrated paralyzing effects on the model nematode C. elegans (ED50 of 50 ng glycoconjugates/µL). To examine molecular mechanisms underlying the nematicidal effects of honey glycoconjugates, expressional analyses of genes that are essential for the growth, development, reproduction, and movement of C. elegans were carried out. Quantitative PCR-based assays showed that these molecules moderately regulate the expression of genes involved in the citric acid cycle (mdh-1 and idhg-1) and cytoskeleton (act-1 and act-2). MALDI-ToF-MS/MS analysis of RP-HPLC peaks revealed the presence of paucimannose-like N-glycans which are known to play important roles in invertebrates e.g., worms and flies. These findings provided novel information regarding the structure and nematicidal function of honey glycoconjugates.

3-(Methylthio)Propionic Acid from Bacillus thuringiensis Berliner Exhibits High Nematicidal Activity against the Root Knot Nematode Meloidogyne incognita (Kofoid and White) Chitwood.

Root knot nematodes cause serious damage to global agricultural production annually. Given that traditional chemical fumigant nematicides are harmful to non-target organisms and the environment, the development of biocontrol strategies has attracted significant attention in recent years. In this study, it was found that the Bacillus thuringiensis Berliner strain NBIN-863 exhibits strong fumigant nematicidal activity and has a high attraction effect on Meloidogyne incognita (Kofoid and White) Chitwood. Four volatile organic compounds (VOCs) produced by NBIN-863 were identified using solid-phase microextraction and gas chromatography-mass spectrometry. The nematicidal activity of four VOCs, namely, N-methylformamide, propenamide, 3-(methylthio)propionic acid, and phenylmalonic acid, was detected. Among these compounds, 3-(methylthio)propionic acid exhibited the highest direct contact nematicidal activity against M. incognita, with an LC50 value of 6.27 µg/mL at 24 h. In the fumigant bioassay, the mortality rate of M. incognita treated with 1 mg/mL of 3-(methylthio)propionic acid for 24 h increased to 69.93%. Furthermore, 3-(methylthio)propionic acid also exhibited an inhibitory effect on the egg-hatching of M. incognita. Using chemotaxis assays, it was determined that 3-(methylthio)propionic acid was highly attractive to M. incognita. In pot experiments, the application of 3-(methylthio)propionic acid resulted in a reduction in gall numbers, decreasing the number of galls per gram of tomato root from 97.58 to 6.97. Additionally, the root length and plant height of the treated plants showed significant increases in comparison with the control group. The current study suggests that 3-(methylthio)propionic acid is a novel nematicidal virulence factor of B. thuringiensis. Our research provides evidence for the potential use of NBIN-863 or its VOCs in biocontrol against root knot nematodes.

Structure-activity relationship and target investigation of 2-aryl quinolines with nematocidal activity.

Within the context of our anthelmintic discovery program, we recently identified and evaluated a quinoline derivative, called ABX464 or obefazimod, as a nematocidal candidate; synthesised a series of analogues which were assessed for activity against the free-living nematode Caenorhabditis elegans; and predicted compound-target relationships by thermal proteome profiling (TPP) and in silico docking. Here, we logically extended this work and critically evaluated the anthelmintic activity of ABX464 analogues on Haemonchus contortus (barber's pole worm) - a highly pathogenic nematode of ruminant livestock. First, we tested a series of 44 analogues on H. contortus (larvae and adults) to investigate the nematocidal pharmacophore of ABX464, and identified one compound with greater potency than the parent compound and showed moderate activity against a select number of other parasitic nematodes (including Ancylostoma, Heligmosomoides and Strongyloides species). Using TPP and in silico modelling studies, we predicted protein HCON_00074590 (a predicted aldo-keto reductase) as a target candidate for ABX464 in H. contortus. Future work aims to optimise this compound as a nematocidal candidate and investigate its pharmacokinetic properties. Overall, this study presents a first step toward the development of a new nematocide.

Metabolites from Lysobacter gummosus YMF3.00690 Against Meloidogyne javanica.

The strains in Lysobacter spp. have the potential to control plant-parasitic nematodes. In our experiment, L. gummosus YMF3.00690 showed antagonistic effects against plant root-knot nematode. Nine metabolites were isolated and identified from cultures of L. gummosus YMF3.00690, of which compound 1 was identified as a new metabolite tetrahydro-4,4,6-trimethyl-6-[(tetrahydro-6,6-dimethyl-2-oxo-4(1H)-pyrimidinylidene) methyl]-2(1H)-pyrimidinone. The activity assay showed that two compounds, 5-(hydroxymethyl)-1H-pyrrole-2-carbaldehyde (2) and 1H-pyrrole-2-carboxylic acid (3), had nematicidal activities against Meloidogyne javanica with mortalities of 69.93 and 90.54% at 400 ppm for 96 h, respectively. These two compounds were further tested for the inhibition activity of eggs hatching, and compound 3 showed a significant inhibition rate of 63.36% at 50 ppm for 48 h. In the chemotactic activity assay, three compounds (1 to 3) were found to have concentration-dependent chemotactic activity, of which compound 1 showed attractive activity. This experiment explored the active metabolites of L. gummosus YMF3.00690 against M. javanica and laid the foundation for biopesticide development.

Characterization and Effect of a Nematophagous Fungus Talaromyces cystophila sp. nov. for the Biological Control of Corn Cyst Nematode.

The dynamic of plant-parasitic nematode populations in soil is closely related to soil microorganisms. Fungi from Heterodera zeae cysts were isolated to explore the phenomenon of decline in the H. zeae population in the soil. Phylogenetic study of partial ITS, BenA, CaM, and RPB2 gene sequences, in addition to morphological investigations, was utilized to identify a nematode-destroying fungus. The nematicidal activity of a novel strain GX1 against H. zeae was assessed in vitro and in the greenhouse. Our findings revealed that strain GX1 is a new species of Talaromyces, named Talaromyces cystophila. It has a strong parasitic and lethal effect on H. zeae cysts, with 91.11% parasitism on cysts at 3 days after treatment. The contents of second-stage juveniles (J2s) and eggs inside the cysts were degraded and formed dense vacuoles, and the damaged eggs could not hatch normally. The spore suspension exhibited high nematophagous activity against nematodes, and fermentation filtrate exhibited marked inhibition of egg hatching and nematicidal activities on J2s. The hatching inhibition rates of eggs exposed to 1 × 108 CFU/ml spore suspensions or 20% 1-week fermentation filtrate (1-WF) for 15 days were 98.56 and 100%, respectively. The mortality of J2s exposed to 1 × 108 CFU/ml spore suspension reached 100% at 24 h; exposure to 50% 2-WF was 98.65 and 100% at 24 and 48 h, respectively. Greenhouse experiments revealed that the spore suspension and fermentation broth considerably decreased H. zeae reproduction by 56.17 to 78.76%. T. cystophila is a potential biocontrol strain with nematophagous and nematicidal activity that deserves attention and application.

Volatile organic compounds produced by Paenibacillus polymyxa J2-4 exhibit toxic activity against Meloidogyne incognita.

BACKGROUND: Root knot nematodes cause great damage to crops worldwide. Due to the negative effects of the application of fumigant and old chemical nematicides, biological nematicides have drawn increasing attention in recent years. Here we tested the fumigant activity of the volatile organic compounds (VOCs) blends emitted from Paenibacillus polymyxa and pure commercial VOCs against M. incognita. RESULTS: In this study, we investigated whether P. polymyxa strain J2-4 could produce VOCs that exhibit nematicidal activity. In vitro assays indicated that J2-4 VOCs were highly toxic to second stage juveniles (J2s) and could inhibit egg hatching. Three-layered pot experiments showed that the number of nematodes that penetrating in cucumber roots was reduced by 69.27% after the application of J2-4 VOCs under greenhouse conditions. We identified 14 volatiles using solid-phase micro-extraction gas chromatography-mass spectrometry. The efficacy of six commercially available VOCs, namely 2-isobutyl-3-methylpyrazine, 2,4-dimethoxybenzaldoxime, 2-dodecanone, 2-tridecanol, 2-tridecanone, and 2-tetradecanol, against M. incognita were examined. Except for 2,4-dimethoxybenzaldoxime, the remaining five VOCs showed strong direct-contact nematicidal activity against J2s of M. incognita, and only 2-isobutyl-3-methylpyrazine showed strong fumigant activity against J2s of M. incognita. In pot experiments, 2-isobutyl-3-methylpyrazine and 2-dodecanone reduced the number of root galls by about 70%, and 2-tridecanone reduced the number of root galls and egg masses by about 63% compared with controls. CONCLUSION: Paenibacillus polymyxa strain J2-4 exhibited high fumigant activity against M. incognita. Our results provide evidence for the use of J2-4 and its VOCs as biocontrol agents in the management of root-knot nematodes. © 2023 Society of Chemical Industry.

Design, Synthesis, Nematicidal Activity, and Mechanism of Novel Amide Derivatives Containing an 1,2,4-Oxadiazole Moiety.

To discover new nematicides, a series of novel amide derivatives containing 1,2,4-oxadiazole were designed and synthesized. Several compounds showed excellent nematicidal activity. The LC50 values of compounds A7, A18, and A20-A22 against pine wood nematode (Bursaphelenchus xylophilus), rice stem nematode (Aphelenchoides besseyi), and sweet potato stem nematode (Ditylenchus destructor) were 1.39-3.09 mg/L, which were significantly better than the control nematicide tioxazafen (106, 49.0, and 75.0 mg/L, respectively). Compound A7 had an outstanding inhibitory effect on nematode feeding, reproductive ability, and egg hatching. Compound A7 effectively promoted the oxidative stress of nematodes and caused intestinal damage to nematodes. Compound A7 significantly inhibited the activity of succinate dehydrogenase (SDH) in nematodes, leading to blockage of electron transfer in the respiratory chain and thereby hindering the synthesis of adenosine triphosphate (ATP), which consequently affects the entire oxidative phosphorylation process to finally cause nematode death. Therefore, compound A7 can be used as a potential SDH inhibitor in nematicide applications.

The nematicide emamectin benzoate increases ROS accumulation in Pinus massoniana and poison Monochamus alternatus.

Pine wilt disease (PWD) is caused by the pine wood nematode (PWN, Bursaphelenchus xylophilus) and transmitted by a vector insect, the Monochamus alternatus. The PWN has caused much extensive damage to pine-dominated forest ecosystems. Trunk injection of emamectin benzoate (EB) has been found to be the most useful protective measure against the PWN, due to its low effective dose and long residence time in the field. However, the interactions between EB and the host or the environment remain largely unknown, which limits the efficacy and stability of EB in practical field settings. In this study, we investigated the impact on PWN from EB injection for both adult and young host plants (Pinus massoniana) by taking a multi-omics (phenomics, transcriptomics, microbiome, and metabolomics) approach. We found that EB injection can significantly reduce the amount of PWN in both living adult and young pine trees. Additionally, EB was able to activate the genetic response of P. massoniana against PWN, promotes P. massoniana growth and development and resistance to Pine wilt disease, which requires the presence of PWN. Further, the presence of EB greatly increased the accumulation of reactive oxygen species (ROS) in the host plant in a PWN-dependent manner, possibly by affecting ROS-related microbes and metabolites. Moreover, we uncovered the function of EB limiting the consumption of P. massoniana by the JPS. Based on biochemical and gut microbial data, we found that EB can significantly reduces cellulase activity in JPS, whose transcription factors, sugar metabolism, and the phosphotransferase system are also affected. These results document the impact of EB on the entire PWD transmission chain through multi-omics regarding the dominant pine (P. massoniana) in China and provide a novel perspective for controlling PWD outbreaks in the field.

Toxicity of fungal-derived volatile organic compounds against root-knot nematodes.

BACKGROUND: Root-knot nematodes (RKNs), including Meloidogyne species, are among the most destructive plant-parasites worldwide. Recent evidence suggests that entomopathogenic fungi (EPF) can antagonize RKNs. Such antagonistic effects are likely mediated by toxic metabolites, including volatile organic compounds (VOCs), produced by the fungi. However, how widespread these effects are across EPF species, and which VOCs mediate negative interactions between EPF and RKNs needs to be further elucidated. RESULTS: First, we evaluated the nematicidal effect of VOCs emitted by 46 EPF isolates against Meloidogyne incognita and found variable toxicity depending on the isolate. Second, we measured the nematicidal effect of highly toxic isolates, including species in the genus Talaromyces, Aspergillus, Clonostachys, and Purpureocillium and, third, we analyzed the nematicidal effect of major VOCs, including 2-methyl-1-propanol, 3-methyl-1-butanol, isopropyl alcohol and 2-methyl-3-pentanone. The mortality of M. incognita juveniles (J2s) was generally high (50%) either via airborne or in-solution contact with VOCs. Moreover, the tested VOCs significantly inhibited egg hatching, and repelled J2s away from the VOCs. CONCLUSION: This study not only provides insights into the ecological function of VOCs in the rhizosphere, but also provides new approaches for developing environmentally friendly control methods of RKNs in agroecosystems. © 2023 Society of Chemical Industry.

Effect of 4-Ethylbenzaldehyde from the Volatilome of Annona muricata on Meloidogyne incognita.

The demand for new soil fumigants has increased as a result of more restrictive legislation regarding the use of pesticides. In the present study, the potent nematicidal activity of volatile organic compounds released by the Annona muricata leaf macerate was demonstrated. In addition, we searched in the A. muricata volatilome for a molecule with potential to be developed as a new fumigant nematicide. In the greenhouse, even the lowest concentration of soursop leaf macerate tested (1.0%) as a biofumigant caused a significant (P < 0.05) reduction in Meloidogyne incognita infectivity and reproduction when compared with the nontreated control (0%). Forty-one compounds were identified through gas chromatography-mass spectrometry analysis, of which three (sabinene, caryophyllene oxide, and 4-ethylbenzaldehyde) were selected for studies against the nematode. Among these compounds, in in vitro trails, only 4-ethylbenzaldehyde showed nematicidal activity at 250 µg ml-1. The effective doses of 4-ethylbenzaldehyde predicted to kill 50 and 95% of the M. incognita second-stage juvenile population after 48 h of exposure were 35 and 88 µg ml-1, respectively. In in vitro tests, 4-ethylbenzaldehyde at 150 µg ml-1 reduced M. incognita egg hatching to values similar (P > 0.05) to those of the commercial nematicide fluensulfone at a concentration of 200 µg ml-1. In plant experiments, as a soil fumigant, 4-ethylbenzaldehyde at a dose of 1 ml/liter of substrate had an effect similar (P > 0.05) to that of the commercial fumigant Dazomet (250 µg ml-1). Therefore, 4-ethylbenzaldehyde shows potential for development as a new nematicide.

Antibacterial peptides from Monochamus alternatus induced oxidative stress and reproductive defects in pine wood nematode through the ERK/MAPK signaling pathway.

Pine wilt disease is a devastating disease of pine caused by the pine wood nematode (PWN) Bursaphelenchus xylophilus. Long-term use of chemical nematicides leads to the development of resistance in nematodes and harms the environment. Evaluations for green environmental protection agents, identified the antibacterial peptide, MaltDef1, from Monochamus alternatus which had nematicidal effect. We studied its nematicidal activity and action against PWN. In this study, the antibacterial peptide S-defensin was synthesized from M. alternatus. The results showed that S-defensin caused mortality to the PWN, causing shrinkage, pore, cell membrane dissolution and muscle atrophy. In addition, PWN reproduction was also affected by S-defensin; it decreased in a concentration dependent manner with increasing treatment concentration. By contrast, reactive oxygen species (ROS) in vivo increased in a concentration-dependent manner. We applied transcriptome to analyze the changes in gene expressions in S-defensin treated PWN, and found that the most significantly enriched pathway was the ERK/MAPK signaling pathway. RNAi was used to validate the functions of four differential genes (Let-23, Let-60, Mek-2 and Lin-1) in this pathway. The results showed that knockdown of these genes significantly decreased the survival rate and reproductive yield of, and also increased ROS in PWN. The antibacterial peptide S-defensin had a significant inhibitory effect on the survival and reproduction of PWN, shown by cell membrane damage and intracellular biological oxidative stress via regulating the ERK/MAPK signaling pathway. This indicates that S-defensin has a target in B. xylophilus, against which new green target pesticides can be developed.

Exploring the role of detoxification genes in the resistance of Bursaphelenchus xylophilus to different exogenous nematicidal substances using transcriptomic analyses.

Bursaphelenchus xylophilus (Pine wood nematode, PWN) has become a worldwide forest disease due to its rapid infection ability, high lethality and difficulty in control. The main means of countering B. xylophilus is currently chemical control, but nematicides can present problems such as environmental pollution and drug resistance. The development of novel environmentally-friendly nematicides has thus become a focus of recent research. In this study, BxUGT3 and BxUGT34, which might be related to detoxification, were investigated by comparing transcriptomic and WGCNA approaches. Three other genes with a similar expression pattern, BxUGT13, BxUGT14, and BxUGT16, were found by gene family analysis. Further bioassays and qPCR assays confirmed that these five genes showed significant changes in transcript levels upon exposure to α-pinene and carvone, demonstrating that they respond to exogenous nematicidal substances. Finally, RNAi and bioassays showed that B. xylophilus with silenced BxUGT16 had increased mortality in the face of α-pinene and carvone stress, suggesting that BxUGT16 plays an important role in detoxification. Taken together, this study used novel molecular research methods, explored the detoxification mechanism of B. xylophilus at a transcriptomic level, and revealed a molecular target for the development of novel biopesticides.