Design, synthesis, and nematicidal activity of novel 1,2,4-oxadiazole derivatives containing amide fragments.

Plant-parasitic nematodes are seriously affecting agricultural production worldwide and there are few highly effective and low-risk nematicides to control nematode diseases. In order to discover new nematicides, a series of 1,2,4-oxadiazole derivatives containing amide fragments have been designed and synthesized with the principle of active substructure splicing. The nematicidal activity of the target compounds was evaluated in vitro and it indicated that compound C3 exhibited the most nematicidal activity against Bursaphelenchus xylophilus, Aphelenchoides besseyi, and Ditylenchus destructor with the LC50 values of 37.2, 36.6, and 43.4 µg/mL, respectively, which were superior to positive agent tioxazafen. The preliminary mechanism results revealed that compound C3 not only inhibited the reproduction of B. xylophilus populations, but also affected the production of ROS and the accumulation of lipofuscin and lipids. Furthermore, compound C3 showed good inhibition of succinate dehydrogenase (SDH) with the IC50 value of 45.5 µmol/L. Molecular docking indicated that compound C3 had excellent binding to amino acids around the SDH active pocket. This work indicated that 1,2,4-oxadiazole derivative containing amide fragment is a promising template for the discovery of new nematicides and compound C3 can be used as a potential nematicide candidate.

Deciphering Chemical Rules for Drug Penetration into Strongyloides.

Background: Strongyloidiasis, a parasitic infection, presents a significant public health challenge in tropical regions due to the limited repertoire of effective treatments. The screening of chemical libraries against the therapeutically relevant third-stage larvae (L3) of the model parasite Strongyloides venezuelensis yielded meager success rates. This situation is reminiscent of Gram-negative bacteria, where drug entry is a limiting factor. Methods: Here, we set out to determine whether similar barriers are in place and establish whether structural and property requirements exist for anti-strongyloides drug discovery. We focused on dyes as their uptake and effects on viability can be independently assessed in the multicellular parasite, thus providing a means to study the possibility of similar entry rules. We tested different dyes in in vitro assays on L3s. Results: We found that staining was necessary to reduce parasite viability, with some dyes achieving anti-strongyloides effects at concentrations similar to those of the reference drug, ivermectin (IV). Some dyes also showed activity against female adults at concentrations well below that of ivermectin. Unfortunately, the most potent dye, Methylene Blue, was unable to prevent the infection in a preliminary in vivo mouse model assay, presumably due to fast dye clearance. Structural analysis showed that positive charges facilitated the access of the compounds to the L3 tissue, thus providing a structural tool for the introduction of activity. For female adults, low globularity is additionally required. As a proof of concept, we added a positive charge to an inactive compound of one of our chemical libraries and re-determined the activity. Conclusions: These findings allow us to establish structural rules for parasite entry that could be of interest for future drug screening or drug development campaigns. These rules might also be applicable to other related parasites.

The New Nematicide Cyclobutrifluram Targets the Mitochondrial Succinate Dehydrogenase Complex in Bursaphelenchus xylophilus.

Bursaphelenchus xylophilus is a dangerous quarantine pest that causes extensive damage to pine ecosystems worldwide. Cyclobutrifluram, a succinate dehydrogenase inhibitor (SDHI), is a novel nematicide introduced by Syngenta in 2013. However, the nematocidal effect of cyclobutrifluram against plant-parasitic nematodes remains underexplored. Therefore, here, we aim to address this knowledge gap by evaluating the toxicity, effects, and mode of action of cyclobutrifluram on B. xylophilus. The result shows that cyclobutrifluram is the most effective agent, with an LC50 value of 0.1078 mg·L-1. At an LC20 dose, it significantly reduced the population size to 10.40 × 103 ± 737.56-approximately 1/23 that of the control group. This notable impact may stem from the agent's ability to diminish egg-laying and hatching rates, as well as to impede the nematodes' development. In addition, it has also performed well in the prevention of pine wilt disease, significantly reducing the incidence in greenhouses and in the field. SDH consists of a transmembrane assembly composed of four protein subunits (SDHA to SDHD). Four sdh genes were characterized and proved by RNAi to regulate the spawning capacity, locomotion ability, and body size of B. xylophilus. The mortality of nematodes treated with sdhc-dsRNA significantly decreased upon cyclobutrifluram application. Molecular docking further confirmed that SDHC, a cytochrome-binding protein, is the target. In conclusion, cyclobutrifluram has a good potential for trunk injection against B. xylophilus. This study provides valuable information for the screening and application of effective agents in controlling and preventing PWD in forests.

Assessing the In Vitro Individual and Combined Effect of Arthrobotrys oligospora and A. musiformis (Orbiliales) Liquid Culture Filtrates against Infective Larvae of the Sheep Blood-Feeding Nematode Haemonchus contortus (Trichostrongylidae).

BACKGROUND: Arthrobotrys species are nematophagous fungi that secrete extracellular nematocidal products (ECP). The individual and combined effects of ECP from Arthrobotrys oligospora (Ao) and A. musiformis (Am) growth in liquid media against Haemonchus contortus L3 (HcL3) were assessed. METHODS: The isolation, morphological (MI) and molecular identification (Mol-I), assessment of nematocidal activity (NA) of fungal liquid culture filtrates (LCF) in two liquid media alone and in combination and the myco-compound profile identification (MCP) were performed. RESULTS: The MI suggested that the fungi corresponded to the species Ao and Am. This result was confirmed by PCR analysis followed by sequencing, alignment and a phylogenetic analysis. Likewise, the highest Hc mortalities were 91.4% with individual LCF of Am and 86.2% with those of Ao at the highest concentration (100 mg/mL) in Czapek-Dox Broth. The combination of both LCF resulted in a similarly high larval mortality with no statistical differences in relation to individual activity (p > 0.05). The MCP showed the presence of alkaloids in both fungi. Coumarins, sterols and saponins were found only in Ao. MAIN CONCLUSIONS: Both fungi produced ECP with a high NA that could be identified and assessed in future studies as potential natural anthelmintic compounds.

New Thiosemicarbazide Derivatives with Multidirectional Biological Action.

Over the years, several new medicinal substances have been introduced for the treatment of diseases caused by bacteria and parasites. Unfortunately, due to the production of numerous defense mechanisms by microorganisms and parasites, they still pose a serious threat to humanity around the world. Therefore, laboratories all over the world are still working on finding new, effective methods of pharmacotherapy. This research work aimed to synthesize new compounds derived from 3-trifluoromethylbenzoic acid hydrazide and to determine their biological activity. The first stage of the research was to obtain seven new compounds, including six linear compounds and one derivative of 1,2,4-triazole. The PASS software was used to estimate the potential probabilities of biological activity of the newly obtained derivatives. Next, studies were carried out to determine the nematocidal potential of the compounds with the use of nematodes of the genus Rhabditis sp. and antibacterial activity using the ACCT standard strains. To determine the lack of cytotoxicity, tests were performed on two cell lines. Additionally, an antioxidant activity test was performed due to the importance of scavenging free radicals in infections with pathogenic microorganisms. The conducted research proved the anthelmintic and antibacterial potential of the newly obtained compounds. The most effective were two compounds with a 3-chlorophenyl substituent, both linear and cyclic derivatives. They demonstrated higher efficacy than the drugs used in treatment.

Comparative Analysis of Chemical Profile and Biological Activity of Juniperus communis L. Berry Extracts.

Researchers are looking for the most effective ways to extract the bioactive substances of Juniperus communis L. berries, which are capable of displaying the greatest range of biological activity, namely antimicrobial potential "against phytopathogens", antioxidant activity and nematocidal activity. This study provides detailed information on the chemical activity, group composition and biological activity of the extracts of juniper berries of 1- and 2-year maturity (JB1 and JB2), which were obtained by using different solvents (pentane, chloroform, acetone, methanol and 70% ethanol) under various extraction conditions (maceration and ultrasound-assisted maceration (US)). Seventy percent ethanol and acetone extracts of juniper berries were analyzed via gas chromatography-mass spectrometry, and they contained monoterpenes, sesquiterpenes, polysaccharides, steroids, fatty acid esters and bicyclic monoterpenes. The antimicrobial activity was higher in the berries of 1-year maturity, while the acetone extract obtained via ultrasound-assisted maceration was the most bioactive in relation to the phytopathogens. Depending on the extraction method and the choice of solvent, the antioxidant activity with the use of US decreased by 1.5-1.9 times compared to the extracts obtained via maceration. An analysis of the nematocidal activity showed that the sensitivity to the action of extracts in Caenorhabditis elegans was significantly higher than in Caenorhabditis briggsae, particularly for the acetone extract obtained from the juniper berries of 1-year maturity.

Nematocidal and Bactericidal Activities of Green Synthesized Silver Nanoparticles Mediated by Ficus sycomorus Leaf Extract.

Nanoparticles effectively control most plant pathogens, although research has focused more on their antimicrobial than their nematocidal properties. This study synthesized silver nanoparticles (Ag-NPs) through a green biosynthesis method using an aqueous extract of Ficus sycomorus leaves (FS-Ag-NPs). The nanoparticles were characterized using SEM, TEM, EDX, zeta sizer, and FTIR. The TEM results showed that the synthesized NPs were nanoscale and had an average particle size of 33 ± 1 nm. The elemental silver signal at 3 keV confirmed the formation of Ag-NPs from an aqueous leaf extract of F. sycomorus. The FTIR analysis revealed the existence of several functional groups in the prepared Ag-NPs. The strong-broad band detected at 3430 cm-1 indicated the stretching vibration of -OH (hydroxyl) and -NH2 (amine) groups. The nematocidal activity of biosynthesized FS-Ag-NPs has been evaluated in vitro against the root-knot nematode Meloidogyne incognita at 24, 48, and 72 h. The FS-Ag-NPs at a 200 µg/mL concentration applied for 48 h showed the highest effectiveness, with 57.62% nematode mortality. Moreover, the biosynthesized FS-Ag-NPs were also tested for their antibacterial activity against Pectobacterium carotovorum, P. atrosepticum, and Ralstonia solanacearum. With the application of nanoparticles, the reduction in bacterial growth gradually increased. The most potent activity at all concentrations was found in R. solanacearum, with values of 14.00 ± 2.16, 17.33 ± 2.05, 19.00 ± 1.41, 24.00 ± 1.41, and 26.00 ± 2.83 at concentrations of 5, 10, 15, 20, and 25 µg/mL, respectively, when compared with the positive control (Amoxicillin 25 µg) with a value of 16.33 ± 0.94. At the same time, the nanoparticles showed the lowest reduction values against P. atrosepticum when compared to the control. This study is the first report on the nematocidal activity of Ag-NPs using F. sycomorus aqueous extract, which could be a recommended treatment for managing plant-parasitic nematodes due to its simplicity, stability, cost-effectiveness, and environmentally safe nature.

Discovery of 1,2,4-Oxadiazole Derivatives Containing Haloalkyl as Potential Acetylcholine Receptor Nematicides.

Plant-parasitic nematodes pose a serious threat to crops and cause substantial financial losses due to control difficulties. Tioxazafen (3-phenyl-5-thiophen-2-yl-1,2,4-oxadiazole) is a novel broad-spectrum nematicide developed by the Monsanto Company, which shows good prevention effects on many kinds of nematodes. To discover compounds with high nematocidal activities, 48 derivatives of 1,2,4-oxadiazole were obtained by introducing haloalkyl at the 5-position of tioxazafen, and their nematocidal activities were systematically evaluated. The bioassays revealed that most of 1,2,4-oxadiazole derivatives showed remarkable nematocidal activities against Bursaphelenchus xylophilus, Aphelenchoides besseyi, and Ditylenchus dipsaci. Notably, compound A1 showed excellent nematocidal activity against B. xylophilus with LC50 values of 2.4 µg/mL, which was superior to that of avermectin (335.5 µg/mL), tioxazafen (>300 µg/mL), and fosthiazate (436.9 µg/mL). The transcriptome and enzyme activity results indicate that the nematocidal activity of compound A1 was mainly related to the compound which affected the acetylcholine receptor of B. xylophilus.

A new ß-class milbemycin derivative from a mutant of genetically engineered strain Streptomyces avermitilis AVE-H39.

A new ß-class milbemycin, 13α-hydroxy milbemycin ß6 (1), was isolated from the fermentation broth of a mutant of genetically engineered strain Streptomyces avermitilis AVE-H39. Its structure and absolute configuration were elucidated by extensive spectroscopic methods and confirmed by single crystal X-ray diffraction.

Microbial and Plant Derived Low Risk Pesticides Having Nematocidal Activity.

Microorganisms, virus, weeds, parasitic plants, insects, and nematodes are among the enemies that induce severe economic losses to agrarian production. Farmers have been forced to combat these enemies using different methods, including mechanical and agronomic strategies, since the beginning of agriculture. The development of agriculture, due to an increased request for food production, which is a consequence to the rapid and noteworthy growth of the world's population, requires the use of more efficient methods to strongly elevate the yield production. Thus, in the last five-to-six decades, a massive and extensive use of chemicals has occurred in agriculture, resulting in heavy negative consequences, such as the increase in environmental pollution and risks for human and animal health. These problems increased with the repetition of treatments, which is due to resistance that natural enemies developed against this massive use of pesticides. There are new control strategies under investigation to develop products, namely biopesticides, with high efficacy and selectivity but based on natural products which are not toxic, and which are biodegradable in a short time. This review is focused on the microbial and plant metabolites with nematocidal activity with potential applications in suitable formulations in greenhouses and fields.

Improvement of cell suspension cultures of transformed and untransformed Carica papaya cell lines, towards the development of an antiparasitic product against the gastrointestinal nematode Haemonchus contortus.

Parasitic diseases have a major impact on human and animal health worldwide. Despite the availability of effective anti-parasitic drugs, their excessive and uncontrolled use has promoted the emergence of drug resistance, severely affecting ecosystems and human health. Thus, developing environmentally friendly antiparasitic treatments is urgently needed. Carica papaya has shown promising effects against infectious diseases. C. papaya embryogenic calluses were genetically modified by our research team to insert immunogenic peptides with the goal of developing an oral anti-cysticercosis vaccine. Among these callus cell lines, one labeled as CF-23, which expresses the KETc7 immunogenic peptide, induced the highest protection levels against experimental cysticercosis. In the process of designing a natural antiparasitic product based on C. papaya that simultaneously induced immunity against cysticercosis, both transformed (SF-23) and untransformed (SF-WT) suspension cultures were produced and optimized. Our results showed a better duplication time (td) for SF-23 (6.9 days) than SF-WT (13.02 days); thus, the SF-23 line was selected for scale-up in a 2-L airlift bioreactor, reaching a td of 4.4 days. This is the first time that a transgenic line of C. papaya has been grown in an airlift bioreactor, highlighting its potential for scale-up cultivation in this type of reactor. Considering the previously reported nematocidal activity of C. papaya tissues, their activity against the nematode Haemonchus contortus of aqueous extracts of SF-WT and SF-23 was explored in this study, with promising results. The information herein reported will allow us to continue the cultivation of the transgenic cell suspension line of C. papaya under reproducible conditions, to develop a new anti-parasitic product.

Design, Synthesis, and Biological Activity of Novel Chalcone Derivatives Containing an 1,2,4-Oxadiazole Moiety.

To discover a lead compound for agricultural use, 34 novel chalcone derivatives containing an 1,2,4-oxadiazole moiety were designed and synthesized. Their nematocidal activities against Bursaphelenchus xylophilus, Aphelenchoides besseyi, and Ditylenchus dipsaci and their antiviral activities against tobacco mosaic virus (TMV), pepper mild mottle virus (PMMoV), and tomato spotted wilt virus (TSWV) were evaluated. Biological assay results indicate that compounds A13 and A14 showed good nematocidal activities against B. xylophilus, A. besseyi, and D. dipsaci, with LC50 values of 35.5, 44.7, and 30.2 µg/ml and 31.8, 47.4, and 36.5 µg/ml, respectively, which are better than tioxazafen, fosthiazate, and abamectin. Furthermore, compound A16 demonstrated excellent protective activity against TMV, PMMoV, and TSWV, with EC50 values of 210.4, 156.2, and 178.2 µg/ml, respectively, which are superior to ningnanmycin (242.6, 218.4, and 180.5 µg/ml).

Antibiotic and Nematocidal Metabolites from Two Lichen Species Collected on the Island of Lampedusa (Sicily).

The antibiotic and nematocidal activities of extracts from two coastal lichen species collected on Lampedusa Island (Sicily), Ramalina implexa Nyl. and Roccella phycopsis Ach., were tested. Methyl orsellinate, orcinol, (+)-montagnetol, and for the first time 4-chlororcinol were isolated from Roccella phycopsis. (+)-Usnic acid was obtained from Ramalina implexa. The crude organic extract of both lichen species showed strong antibiotic activity against some bacterial species and nematocidal activity. Among all the pure metabolites tested against the infective juveniles (J2) of the root-knot nematode (RKN) Meloydogine incognita, (+)-usnic acid, orcinol, and (+)-montagnetol had significant nematocidal activity, comparable with that of the commercial nematocide Velum® Prime, and thus they showed potential application in agriculture as a biopesticide. On the contrary, methyl orsellinate and 4-chlororcinol had no nematocidal effect. These results suggest that the substituent pattern at ortho-para-position in respect to both hydroxyl groups of resorcine moiety, which is present in all metabolites, seems very important for nematocidal activity. The organic extracts of both lichens were also tested against some Gram-positive and Gram-negative bacteria. Both extracts were active against Gram-positive species. The extract of Ramalina implexa showed, among Gram-negative species, activity against Escherichia coli and Acinetobacter baumannii, while that from Roccella phycopsis was effective towards all test strains, with the exception of Pseudomonas aeruginosa. The antimicrobial activity of (+)-usnic acid, methyl orsellinate, and (+)-montagnetol is already known, so tests were focused on orcinol and 4-chlororcinol. The former showed antibacterial activity against all Gram positive and Gram-negative test strains, with the exception of A. baumannii and K. pneumoniae, while the latter exhibited a potent antibacterial activity against Gram-positive test strains and among Gram-negative strains, was effective against A. baumannii and K. pneumonia. These results suggest, for orcinol and 4-chlororcinol, an interesting antibiotic potential against both Gram-positive and Gram-negative bacterial strains.

Pathogenicity and Metabolites of Purpureocillium lavendulum YMF1.00683 against Meloidogyne incognita.

Purpureocillium lavendulum is a biological control agent with several registered products that can parasitize the eggs and larvae of various pathogenic nematodes. In this study, the pathogenicity and secondary metabolites of the fungus P. lavendulum YMF1.00683 were investigated. The strain YMF1.00683 had infection efficiency against the plant root-knot nematode Meloidogyne incognita. The strain's process of infecting nematodes was observed under a microscope. Moreover, seven metabolites, including a new sterol (1), were isolated and identified from cultures of YMF1.0068 in Sabouraud's dextrose agar. A bioassay showed that 5-methoxymethyl-1H-pyrrole-2-carboxaldehyde (7) is toxic to M. incognita and affects the egg hatching. It caused 98.23% mortality in M. incognita and could inhibit 80.78% of the hatching eggs at 400 µg/mL over a period of 96 h. Furthermore, 5-methoxymethyl-1H-pyrrole-2-carboxaldehyde (7) showed a strong avoidance effect at 40 ppm, and its chemotactic index value was -0.37. The results indicate that P. lavendulum could produce active metabolites against M. incognita.

Gene sdaB Is Involved in the Nematocidal Activity of Enterobacter ludwigii AA4 Against the Pine Wood Nematode Bursaphelenchus xylophilus.

Bursaphelenchus xylophilus, a plant parasitic nematode, is the causal agent of pine wilt, a devastating forest tree disease. Essentially, no efficient methods for controlling B. xylophilus and pine wilt disease have yet been developed. Enterobacter ludwigii AA4, isolated from the root of maize, has powerful nematocidal activity against B. xylophilus in a new in vitro dye exclusion test. The corrected mortality of the B. xylophilus treated by E. ludwigii AA4 or its cell extract reached 98.3 and 98.6%, respectively. Morphological changes in B. xylophilus treated with a cell extract from strain AA4 suggested that the death of B. xylophilus might be caused by an increased number of vacuoles in non-apoptotic cell death and the damage to tissues of the nematodes. In a greenhouse test, the disease index of the seedlings of Scots pine (Pinus sylvestris) treated with the cells of strain AA4 plus B. xylophilus or those treated by AA4 cell extract plus B. xylophilus was 38.2 and 30.3, respectively, was significantly lower than 92.5 in the control plants treated with distilled water and B. xylophilus. We created a sdaB gene knockout in strain AA4 by deleting the gene that was putatively encoding the beta-subunit of L-serine dehydratase through Red homologous recombination. The nematocidal and disease-suppressing activities of the knockout strain were remarkably impaired. Finally, we revealed a robust colonization of P. sylvestris seedling needles by E. ludwigii AA4, which is supposed to contribute to the disease-controlling efficacy of strain AA4. Therefore, E. ludwigii AA4 has significant potential to serve as an agent for the biological control of pine wilt disease caused by B. xylophilus.

Bacillus thuringiensis toxins with nematocidal activity against the pinewood nematode Bursaphelenchus xylophilus.

The pine wilt disease is caused by the pinewood nematode Bursaphelenchus xylophilus and it results in serious ecological and economic losses. Therefore, effective prevention and control methods for the pinewood nematode are urgently required. Bacillus thuringiensis (Bt), a widely used microbial insecticide, produces toxins that are toxic to several species of parasitic nematodes, however, its effects on B. xylophilus have not been determined. In this study, Cry5Ba3, App6Aa2, Cry12Aa1, Cry13Aa1, Cry14Aa1, Cry21Aa3, Cry21Fa1, Xpp55Aa1, and Cyt8Aa1 toxins' nematocidal activity against B. xylophilus was evaluated, six toxins with high toxicity were identified: App6Aa2 (LC50 = 49.71 µg/mL), Cry13Aa1 (LC50 = 53.17 µg/mL), Cry12Aa1 (LC50 = 58.88 µg/mL), Cry5Ba3 (LC50 = 63.99 µg/mL), Xpp55Aa1 (LC50 = 65.14 µg/mL), and Cyt8Aa1 (LC50 = 96.50 µg/mL). The six toxins caused shrinkage and thinning of the intestinal cells, contraction of the intestine from the body wall, vacuolization, and degenerated appearance of the pinewood nematodes. The results of this study provide basic information to study the action mechanism of nematocidal toxins on the pinewood nematode and direction for the use of nematocidal toxins in the biological control of B. xylophilus.

A Nematode Crude Extract Acts as an Elicitor of the Nematocidal Activity of Nematophagous Fungi Liquid Culture Filtrates Against Haemonchus contortus (Nematoda: Trichostrongylidae).

AIM: This study was designed to investigate if culturing nematophagous fungi (NF) in the presence of a Haemonchus contortus larva crude extract (HcCE) enhances the nematocidal activity of nematophagous fungi liquid culture filtrates (NFCF). MATERIALS AND METHODS: Four NF Arthrobotrys oligospora, A. musiformis, Duddingtonia flagrans and Clonostachys rosea were cultured in flasks (n = 5) containing Czapek-Dox broth medium (CDB) in the presence or absence of HcCE. NFCF recovered by filtration of each fungus (200 mg/mL) were assessed on H. contortus infective larvae (L3) using 96-well micro-titer plates (n = 4). Additionally, CDB and water were considered negative controls, while Ivermectin acted as a positive control. After 48 h confrontation, ten 10-µL aliquots of each well were deposited on slides and observed under the microscope (40 ×). Dead and alive larvae in the aliquots were quantified, and a mortality rate (MR) was estimated. RESULTS: The MR of the different NFCF was greatly enhanced by the presence of HcCE. The four NF incubated in the absence of HcCE showed low mortality percentages from 8.2 to 25.8%; in contrast, when the assessed NF growth in the presence of HcCE showed a lethal activity ranging from 66.8 to 80.5%. Only C. rosea showed a moderate increase in the presence of the elicitor (42.7%). CONCLUSION: This study shows evidence about the HcCE enhances the production of nematocidal activity in NFCF. Future studies should be performed to elucidate the compounds responsible of the nematocidal activity that could have important implications in the control of sheep haemonchosis.

Activities and metabolomics of Cordyceps gunnii under different culture conditions.

Many active metabolites have been identified from various species of the fungal genus Cordyceps. A predominant species of this genus is Cordyceps gunnii, but there are limited reports on the active ingredients from this species. This study aimed to conduct activity assays and metabolome analysis on extracts of C. gunnii obtained under different culture conditions. Five different solid media were selected to culture the mycelium of C. gunnii and the metabolites were extracted with organic solvents; concurrently, the wild stroma and host complexes of C. gunnii were extracted by ethyl acetate. Extracts were subsequently assayed for various biological activities and were analyzed by untargeted metabolomics. There were significant differences in the activities and metabolites of C. gunnii extracts from different culture conditions and from wild stroma and host complexes. The extracts of stroma and host complexes and mycelia cultured on WGA medium for 21 days exhibited similar effective inhibitory activity against five cell lines. A total of 51 metabolites were annotated and included various structural types. The literatures indicate that most of the identified compounds have a variety of different biological activities. These findings provide the basis for further systematic excavation of C. gunnii and improved utilization of this fungal species.

Bacterial strains integrated with surfactin molecules of Bacillus subtilis MTCC441 enrich nematocidal activity against Meloidogyne incognita.

Bacteria secrete lipopeptide (surfactin) molecules, which are known to act as natural antibiotics. Recently, research on lipopeptide molecules has grown because of their role in increasing resistance to plant pathogens. Isolated surfactin molecules at several concentrations, viz. 35, 25, 15 and 5 ppm, were used in an in vitro study for analysis of egg hatching inhibition and second-stage juvenile (J2) mortality of the nematode Meloidogyne incognita. Cell suspensions (1.2 × 108  cfu·ml-1 ) of both Bacillus subtilis (MTCC-441) and Pseudomonas putida (MTCC-102) were used in J2 inoculated tomato plants. Root-dip treatment of tomato seedlings with the crude lipopeptide (35 ppm) was also performed to analyse biocontrol potential. MALDI-TOF MS analysis was conducted to determine specific lipopeptide molecules. Data showed nematode egg mortality of 85% in the 35 ppm surfactin concentration 96 h after exposure. The maximum ovicidal activity was 83.97% after incubation with 35 ppm surfactin for the same exposure period. Plant growth attributes and biochemical parameters were significantly improved when bacterial cultures were applied before J2 inoculation of tomato seedlings. We also recorded a reduction in egg masses, nematode population and root galling. The J2 penetration into tomato roots was effective in the root-dip experiments. Surfactin mass peak was determined at m/z 1058 [M+Na]+ using MALDI-TOF MS. These results indicate that bacterial cell suspensions can be used as a potent and versatile source to deal with nematode infection and provide a rich source of bioactive compounds with antinematode activity.

Two new 13-hydroxylated milbemycin metabolites from the genetically engineered strain Streptomyces avermitilis AVE-H39.

Two new milbemycin metabolites, 13α-hydroxymilbemycin ß13 (1) and 26-methyl-13α-hydroxymilbemycin ß13 (2), were isolated from the fermentation broth of a genetically engineered strain Streptomyces avermitilis AVE-H39. Their structures were determined by the comprehensive spectroscopic data, including 1 D, 2 D NMR, MS spectral analysis and the comparison with data from the literature. Compounds 1 and 2 not only exhibited potent acaricidal activities against Tetranychus cinnabarinus, but also had nematocidal activity against Bursaphelenchus xylophilus.