The regulation of intercellular and interorgan communication is pivotal for cell fate decisions i... more The regulation of intercellular and interorgan communication is pivotal for cell fate decisions in plant development and probably plays a significant role in the systemic regulation of gene expression and in defense reactions against pathogens or other biotic and abiotic environmental factors. In plants, symplasmic cell-to-cell communication is provided by plasmodesmata (Pd), coaxial membranous tunnels that span cell walls interconnecting adjacent cytoplasms. Macromolecules, proteins, and RNA may be transported through Pd by passive diffusion or by a facilitated mechanism. A quantitative tool was developed to measure the coefficient of conductivity, C(Pd), for diffusion-driven transport via Pd and to assess changes in the coefficient induced by developmental, biotic and abiotic signals. GFP C(Pd), the coefficient of conductivity for cell-to-cell spread of green-fluorescent protein (GFP), a protein with a Stokes radius of 2.82 nm, was determined in epidermal cells of sink and source leaves of wild-type and transgenic Nicotiana benthamiana plants expressing the movement protein of tobacco mosaic virus (MP TMV) incubated both in dark and light and at 16 and 25 ЊC. Under all conditions, Pd in source leaves conducted macromolecules, with GFP C(Pd) sink Ͼ GFP C(Pd) source. Light down-regulated GFP C(Pd) (all conditions); down-regulation was stronger for sink cells. The effect of MP TMV on GFP C(Pd) between epidermal cells was dependent on temperature and leaf development; at 16 ЊC, MP TMV down-regulated GFP C(Pd) only in source leaves, while at 25 ЊC, MP TMV had no significant effect. This quantitative tool should be useful for investigating differences in Pd conductivity that are induced by mutations or silencing.
The worldwide demand for reduced and restricted use of pesticides in agriculture due to serious e... more The worldwide demand for reduced and restricted use of pesticides in agriculture due to serious environmental effects, health risks and the development of pathogen resistance calls for the discovery of new bioactive compounds. In the medical field, antibiotic‐resistant microorganisms have become a major threat to man, increasing mortality. Endophytes are endosymbiotic microorganisms that inhabit plant tissues without causing any visible damage to their host. Many endophytes secrete secondary metabolites with biological activity against a broad range of pathogens, making them potential candidates for novel drugs and alternative pesticides of natural origin. We isolated endophytes from wild plants in Israel, focusing on endophytes that secrete secondary metabolites with biological activity. We isolated 302 different endophytes from 30 different wild plants; 70 of them exhibited biological activity against phytopathogens. One biologically active fungal endophyte from the genus Penicill...
<p>Representative bright-field images of (A) eggs, (B) L1 and (C) L4 larvae on an <i>... more <p>Representative bright-field images of (A) eggs, (B) L1 and (C) L4 larvae on an <i>E</i>. <i>coli</i> lawn exposed to 4-heptanone (0.2 mL, 1.43 mmole) or SVM for 48 h. Scale bar = 100 μm.</p
<p>L4 larvae from strain TJ356 were treated with 4-heptanone or SVM for 24 h. Representativ... more <p>L4 larvae from strain TJ356 were treated with 4-heptanone or SVM for 24 h. Representative images of DAF-16::GFP expression in (A) control TJ356 worms and worms exposed to 4-heptanone, and SVM. Three independent experiments with at least 10 nematodes per group were performed. Scale bar = 100 μm. (B) Quantification of DAF- 16::GFP. The y-axis denotes the average number of pixels in TJ356 nematodes measured after the treatment. Mean expression ± SE from 10 nematodes is shown. Pairwise comparison of the means was performed by analysis of variance followed by post-hoc Tukey–Kramer multiple comparison test. Different letters indicate significantly different values (<i>P</i> ≤ 0.05). (C) Effect of 4-heptanone and SVM on sub cellular distribution of DAF16::GFP. Nuclear translocation of DAF16::GFP was examined in approximately 50 worms for each condition. The worms were scored as cytosolic, intermediate, and nuclear, on the basis of localization of the DAF-16::GFP. Data are presented as mean ±SEM of the percentages of each phenotype.</p
<p>J2s were exposed to 0–3 culture plates of <i>D</i>. <i>cf</i>. &... more <p>J2s were exposed to 0–3 culture plates of <i>D</i>. <i>cf</i>. <i>concentrica</i> for 48 h. The numbers on the x-axis represent the numbers of Petri plates (50 mm in diameter, containing 5 mL of growth medium, and the fungal culture) in each 1-L sealed box. The number 0 indicates control treatment in which the nematodes were not exposed to the fungal culture plate. The viable J2s were separated using 30 μm sieves, and the numbers on the y-axis represent the numbers (mean ± SE) of viable J2s counted following the incubation. There were 10 repetitions, each using 300 J2s. The results were subjected to analysis of variance followed by the Tukey-Kramer multiple comparison test; different letters above the bars indicate a significant difference between samples at <i>P ≤</i> 0.05. The experiment was independently repeated three times, each time with similar results.</p
<p>(A) Untreated wheat grains. (B) Wheat grains after exposure to Mixture A at 0.25 mL/L. (... more <p>(A) Untreated wheat grains. (B) Wheat grains after exposure to Mixture A at 0.25 mL/L. (C) Wheat grains after exposure to Mixture B at 0.25 mL/L.</p
<p>Effects of the volatile compounds of <i>D</i>. <i>cf</i>. <i&... more <p>Effects of the volatile compounds of <i>D</i>. <i>cf</i>. <i>concentrica</i> and artificial mixtures on tested plant pathogenic fungi and oomycete</p
<p>Susceptible tomato seedlings were planted in inoculated soil, with or without pretreatme... more <p>Susceptible tomato seedlings were planted in inoculated soil, with or without pretreatment with the synthetic volatile mixture (SVM). Four treatments were designated: Control NIR—soil inoculated with nematode-infected roots (NIR) infested by <i>M</i>. <i>javanica</i> in amounts equivalent to 4,000 eggs in root suspension in each pot; SVM NIR—soil inoculated with nematode-infected roots infested by <i>M</i>. <i>javanica</i> in amounts equivalent to 4,000 eggs in root suspension in each pot and supplemented with the synthetic volatile mixture; Control J2 –soil inoculated by direct insertion of 4,000 <i>M</i>. <i>javanica</i> J2s into each pot; and SVM J2 –soil inoculated by direct insertion of 4,000 <i>M</i>. <i>javanica</i> J2s and supplemented with the synthetic volatile mixture. (A) Galling index (mean of five repetitions). The results were subjected to nonparametric comparison using Wilcoxon method; different letters above the bars indicate a significant difference between samples significant at <i>P ≤</i> 0.05. (B) Mean numbers (± SE) of <i>M</i>. <i>javanica</i> eggs per gram of root. The results were subjected to analysis of variance followed by the Tukey-Kramer multiple comparison test; different letters above the bars indicate a significant difference between samples at <i>P ≤</i> 0.05. (C) Mean weights (± SE) of tomatoes roots. The results were subjected to analysis of variance. No significant differences (<i>P</i> > 0.01) between treatments in root weight were found. The experiment was independently repeated twice, each time with similar results.</p
<p>Upper panel–Mixture A: (A) Peanuts inoculated with <i>A</i>. <i>niger&... more <p>Upper panel–Mixture A: (A) Peanuts inoculated with <i>A</i>. <i>niger</i> in the presence of mixture at 1 mL/L. (B) Peanuts inoculated with <i>A</i>. <i>niger</i> in the absence of mixture. (C) Uninoculated peanuts in the presence of mixture at 1 mL/L. (D) Uninoculated peanuts in the absence of mixture. Lower panel–Mixture B: (A) Uninoculated peanuts in the absence of mixture. (B) Uninoculated peanuts in the presence of mixture at 0.05 mL/L. (C) Uninoculated peanuts in the presence of mixture at 0.5 mL/L. Arrows indicate the development of intrinsic <i>Aspergillus</i> sp.</p
<p>(A) Eggs, (B) L1, (C) L4 and (D) YA larval stages. Eggs or nematodes were exposed to 3-m... more <p>(A) Eggs, (B) L1, (C) L4 and (D) YA larval stages. Eggs or nematodes were exposed to 3-methyl-1-butanol (0.1 mL, 0.92 mmole), (±)-2-methyl-1-butanol (0.1 mL, 0.93 mmole), 4-heptanone (0.2 mL, 1.43 mmole), isoamyl acetate (0.1 mL, 0.68 mmole), or up to 0.5 mL of SVM. The viable nematodes were passed through sieves and counted. Data on the y-axis are presented as no. of eggs hatching (A) and as percent mortality relative to controls (mean ± SE) (B-D). Graphs are representative of three independent experiments. Pairwise comparison of the means was performed with analysis of variance followed by post-hoc Tukey–Kramer multiple comparison test. Different letters indicate significantly different values (<i>P</i> ≤ 0.05).</p
<p>Biological activity of each chemical component consisting 1 mL/L (air space) of Mixture ... more <p>Biological activity of each chemical component consisting 1 mL/L (air space) of Mixture A</p
<p>(A) Control swollen fruits. (B) Swollen fruits in the presence of one culture dish of &l... more <p>(A) Control swollen fruits. (B) Swollen fruits in the presence of one culture dish of <i>D</i>. <i>cf</i>. <i>concentrica</i>. (C) Swollen fruits in the presence of two culture dishes of <i>D</i>. <i>cf</i>. <i>concentrica</i>.</p
<p>In each treatment the nematodes were incubated for 24 or 48 h in the presence or absence... more <p>In each treatment the nematodes were incubated for 24 or 48 h in the presence or absence of 4-heptanone (0.4 mL, 2.86 mmole), before viable J2s were separated using 30 μm sieves. The numbers on the y-axis represent the numbers (mean ± SE) of viable J2s counted following the incubation. There were 8–10 repetitions, each using 300 J2s. In the treatment designated as "4-heptanone 24 h + recovery 24 h" the nematodes were exposed to the compound for 24 h and then removed to a new container with no VOCs for recovery. The results were subjected to analysis of variance followed by the Tukey-Kramer multiple comparison test; different letters above the bars indicate a significant difference between samples at <i>P ≤</i> 0.05. The experiment was independently repeated three times, each time with similar results.</p
<p>J2s were exposed for 48 h either to the SVM at 1 mL/L (V/V) [3-methyl-1-butanol (0.2 mL,... more <p>J2s were exposed for 48 h either to the SVM at 1 mL/L (V/V) [3-methyl-1-butanol (0.2 mL, 1.84 mmole), (±)-2-methyl-1-butanol (0.2 mL, 1.86 mmole), 4-heptanone (0.4 mL, 2.86 mmole), and isoamyl acetate (0.2 mL, 1.35 mmole)] or to three 50-mm-diameter fungal culture plates, each with 5 mL of growth medium, and then the viable J2s were separated using 30 μm sieves. The numbers on the y-axis represent the numbers (mean ± SE) of viable J2s counted following the incubation. Each treatment was composed of 10 technical repetitions using 300 J2s in each repetition. The results were subjected to analysis of variance followed by the Tukey-Kramer multiple comparison test; different letters above the bars indicate a significant difference between samples at <i>P ≤</i> 0.05. The experiment was independently repeated three times, each time with similar results.</p
The bionematicidal effect of a synthetic volatile mixture (SVM) of four volatile organic compound... more The bionematicidal effect of a synthetic volatile mixture (SVM) of four volatile organic compounds (VOCs) emitted by the endophytic fungus Daldinia cf. concentrica against the devastating plant-parasitic root-knot nematode Meloidogyne javanica has been recently demonstrated in both in vitro and greenhouse experiments. However, the mode of action governing the observed irreversible paralysis of J2 larvae upon exposure to SVM is unknown. To unravel the mechanism underlying the anthelmintic and nematicidal activities, we used the tractable model worm Caenorhabditis elegans. C. elegans was also susceptible to both the fungal VOCs and SVM. Among compounds comprising SVM, 3-methyl-1-butanol, (±)-2-methyl-1-butanol, and 4-heptanone showed significant nematicidal activity toward L1, L4 and young adult stages. Egg hatching was only negatively affected by 4-heptanone. To determine the mechanism underlying this activity, we examined the response of C. elegans mutants for glutamate-gated chloride channel and acetylcholine transporter, targets of the nematicidal drugs ivermectin and aldicarb, respectively, to 4-heptanone and SVM. These aldicarb-and ivermectin-resistant mutants retained susceptibility upon exposure to 4-heptanone and SVM. Next, we used C. elegans TJ356 strain zIs356 (daf-16::GFP+rol-6), LD1 ldIs7 [skn-1B/C::GFP + pRF4(rol-6(su1006))], LD1171 ldIs3 [gcs-1p::gfp; rol-6 (su1006))], CL2166 dvIs19 (gst-4p::GFP) and CF1553 muIs84 (sod-3p::GFP+rol-6), which have mutations in genes regulating multiple stress responses. Following exposure of L4 larvae to 4-heptanone or SVM, there was clear nuclear translocation of DAF-16::GFP, and SKN-1::GFP indicating that their susceptibility involves DAF-16 and SKN1 regulation. Application of 4-heptanone, but not SVM, induced increased expression of, gcs-1::GFP and gst-4::GFP compared to controls. In contrast, application of 4-heptanone or SVM to the sod-3:: GFP line elicited a significant decline in overall fluorescence intensity compared to controls,
BACKGROUNDSclerotium rolfsii is a soil‐borne phytopathogenic fungus that causes diseases in econo... more BACKGROUNDSclerotium rolfsii is a soil‐borne phytopathogenic fungus that causes diseases in economically important crops. Eradication of the fungus is hampered by its wide range of hosts, as well as its capacity to form sclerotia. Recently, we have shown that the endophytic fungus Daldinia cf. concentrica emits biologically active volatile organic compounds (VOCs); we also demonstrated that one VOC, trans‐2‐octenal, was the most effective against various phytopathogenic fungi. Thus, the aim of this study was to examine the potential of this compound to control hyphae and sclerotia of S. rolfsii, both in vitro and in soil.RESULTSWe found that in vitro exposure of S. rolfsii mycelium to trans‐2‐octenal in air fully inhibits and kills the fungus. Elimination of sclerotia viability occurred at the same concentration, but direct contact between the sclerotia and the compound was needed. trans‐2‐Octenal also affected the viability of both hyphae and sclerotia of S. rolfsii in small pots c...
Plant-parasitic nematodes form one of the largest sources of biotic stress imposed on plants, and... more Plant-parasitic nematodes form one of the largest sources of biotic stress imposed on plants, and are very difficult to control; among them are the obligate parasites, the sedentary root-knot nematodes (RKNs)-Meloidogyne spp.-which are extremely polyphagous and exploit a very wide range of hosts. Endophytic fungi are organisms that spend most of their life cycle within plant tissue without causing visible damage to the host plant. Many endophytes secrete specialized metabolites and/or emit volatile organic compounds (VOCs) that exhibit biological activity. Recently, we demonstrated that the endophytic fungus Daldinia cf. concentrica secrets biologically active VOCs. Here we examined the ability of the fungus and its VOCs to control the RKN M. javanica both in vitro and greenhouse experiments. The D. cf. concentrica VOCs showed bionematicidal activity against the second-stage juveniles (J2s) of M. javanica. We found that exposure of J2s to fungal volatiles caused 67% reduction in via...
Endophytic fungi are organisms that spend most of their life cycle within plant tissues without c... more Endophytic fungi are organisms that spend most of their life cycle within plant tissues without causing any visible damage to the host plant. Many endophytes were found to secrete specialized metabolites and/or emit volatile organic compounds (VOCs), which may be biologically active and assist fungal survival inside the plant as well as benefit their hosts. We report on the isolation and characterization of a VOCs-emitting endophytic fungus, isolated from an olive tree (Olea europaea L.) growing in Israel; the isolate was identified as Daldinia cf. concentrica. We found that the emitted VOCs were active against various fungi from diverse phyla. Results from postharvest experiments demonstrated that D. cf. concentrica prevented development of molds on organic dried fruits, and eliminated Aspergillus niger infection in peanuts. Gas chromatography-mass spectrometry analysis of the volatiles led to identification of 27 VOCs. On the basis of these VOCs we prepared two mixtures that displ...
The regulation of intercellular and interorgan communication is pivotal for cell fate decisions i... more The regulation of intercellular and interorgan communication is pivotal for cell fate decisions in plant development and probably plays a significant role in the systemic regulation of gene expression and in defense reactions against pathogens or other biotic and abiotic environmental factors. In plants, symplasmic cell-to-cell communication is provided by plasmodesmata (Pd), coaxial membranous tunnels that span cell walls interconnecting adjacent cytoplasms. Macromolecules, proteins, and RNA may be transported through Pd by passive diffusion or by a facilitated mechanism. A quantitative tool was developed to measure the coefficient of conductivity, C(Pd), for diffusion-driven transport via Pd and to assess changes in the coefficient induced by developmental, biotic and abiotic signals. GFP C(Pd), the coefficient of conductivity for cell-to-cell spread of green-fluorescent protein (GFP), a protein with a Stokes radius of 2.82 nm, was determined in epidermal cells of sink and source leaves of wild-type and transgenic Nicotiana benthamiana plants expressing the movement protein of tobacco mosaic virus (MP TMV) incubated both in dark and light and at 16 and 25 ЊC. Under all conditions, Pd in source leaves conducted macromolecules, with GFP C(Pd) sink Ͼ GFP C(Pd) source. Light down-regulated GFP C(Pd) (all conditions); down-regulation was stronger for sink cells. The effect of MP TMV on GFP C(Pd) between epidermal cells was dependent on temperature and leaf development; at 16 ЊC, MP TMV down-regulated GFP C(Pd) only in source leaves, while at 25 ЊC, MP TMV had no significant effect. This quantitative tool should be useful for investigating differences in Pd conductivity that are induced by mutations or silencing.
The worldwide demand for reduced and restricted use of pesticides in agriculture due to serious e... more The worldwide demand for reduced and restricted use of pesticides in agriculture due to serious environmental effects, health risks and the development of pathogen resistance calls for the discovery of new bioactive compounds. In the medical field, antibiotic‐resistant microorganisms have become a major threat to man, increasing mortality. Endophytes are endosymbiotic microorganisms that inhabit plant tissues without causing any visible damage to their host. Many endophytes secrete secondary metabolites with biological activity against a broad range of pathogens, making them potential candidates for novel drugs and alternative pesticides of natural origin. We isolated endophytes from wild plants in Israel, focusing on endophytes that secrete secondary metabolites with biological activity. We isolated 302 different endophytes from 30 different wild plants; 70 of them exhibited biological activity against phytopathogens. One biologically active fungal endophyte from the genus Penicill...
<p>Representative bright-field images of (A) eggs, (B) L1 and (C) L4 larvae on an <i>... more <p>Representative bright-field images of (A) eggs, (B) L1 and (C) L4 larvae on an <i>E</i>. <i>coli</i> lawn exposed to 4-heptanone (0.2 mL, 1.43 mmole) or SVM for 48 h. Scale bar = 100 μm.</p
<p>L4 larvae from strain TJ356 were treated with 4-heptanone or SVM for 24 h. Representativ... more <p>L4 larvae from strain TJ356 were treated with 4-heptanone or SVM for 24 h. Representative images of DAF-16::GFP expression in (A) control TJ356 worms and worms exposed to 4-heptanone, and SVM. Three independent experiments with at least 10 nematodes per group were performed. Scale bar = 100 μm. (B) Quantification of DAF- 16::GFP. The y-axis denotes the average number of pixels in TJ356 nematodes measured after the treatment. Mean expression ± SE from 10 nematodes is shown. Pairwise comparison of the means was performed by analysis of variance followed by post-hoc Tukey–Kramer multiple comparison test. Different letters indicate significantly different values (<i>P</i> ≤ 0.05). (C) Effect of 4-heptanone and SVM on sub cellular distribution of DAF16::GFP. Nuclear translocation of DAF16::GFP was examined in approximately 50 worms for each condition. The worms were scored as cytosolic, intermediate, and nuclear, on the basis of localization of the DAF-16::GFP. Data are presented as mean ±SEM of the percentages of each phenotype.</p
<p>J2s were exposed to 0–3 culture plates of <i>D</i>. <i>cf</i>. &... more <p>J2s were exposed to 0–3 culture plates of <i>D</i>. <i>cf</i>. <i>concentrica</i> for 48 h. The numbers on the x-axis represent the numbers of Petri plates (50 mm in diameter, containing 5 mL of growth medium, and the fungal culture) in each 1-L sealed box. The number 0 indicates control treatment in which the nematodes were not exposed to the fungal culture plate. The viable J2s were separated using 30 μm sieves, and the numbers on the y-axis represent the numbers (mean ± SE) of viable J2s counted following the incubation. There were 10 repetitions, each using 300 J2s. The results were subjected to analysis of variance followed by the Tukey-Kramer multiple comparison test; different letters above the bars indicate a significant difference between samples at <i>P ≤</i> 0.05. The experiment was independently repeated three times, each time with similar results.</p
<p>(A) Untreated wheat grains. (B) Wheat grains after exposure to Mixture A at 0.25 mL/L. (... more <p>(A) Untreated wheat grains. (B) Wheat grains after exposure to Mixture A at 0.25 mL/L. (C) Wheat grains after exposure to Mixture B at 0.25 mL/L.</p
<p>Effects of the volatile compounds of <i>D</i>. <i>cf</i>. <i&... more <p>Effects of the volatile compounds of <i>D</i>. <i>cf</i>. <i>concentrica</i> and artificial mixtures on tested plant pathogenic fungi and oomycete</p
<p>Susceptible tomato seedlings were planted in inoculated soil, with or without pretreatme... more <p>Susceptible tomato seedlings were planted in inoculated soil, with or without pretreatment with the synthetic volatile mixture (SVM). Four treatments were designated: Control NIR—soil inoculated with nematode-infected roots (NIR) infested by <i>M</i>. <i>javanica</i> in amounts equivalent to 4,000 eggs in root suspension in each pot; SVM NIR—soil inoculated with nematode-infected roots infested by <i>M</i>. <i>javanica</i> in amounts equivalent to 4,000 eggs in root suspension in each pot and supplemented with the synthetic volatile mixture; Control J2 –soil inoculated by direct insertion of 4,000 <i>M</i>. <i>javanica</i> J2s into each pot; and SVM J2 –soil inoculated by direct insertion of 4,000 <i>M</i>. <i>javanica</i> J2s and supplemented with the synthetic volatile mixture. (A) Galling index (mean of five repetitions). The results were subjected to nonparametric comparison using Wilcoxon method; different letters above the bars indicate a significant difference between samples significant at <i>P ≤</i> 0.05. (B) Mean numbers (± SE) of <i>M</i>. <i>javanica</i> eggs per gram of root. The results were subjected to analysis of variance followed by the Tukey-Kramer multiple comparison test; different letters above the bars indicate a significant difference between samples at <i>P ≤</i> 0.05. (C) Mean weights (± SE) of tomatoes roots. The results were subjected to analysis of variance. No significant differences (<i>P</i> > 0.01) between treatments in root weight were found. The experiment was independently repeated twice, each time with similar results.</p
<p>Upper panel–Mixture A: (A) Peanuts inoculated with <i>A</i>. <i>niger&... more <p>Upper panel–Mixture A: (A) Peanuts inoculated with <i>A</i>. <i>niger</i> in the presence of mixture at 1 mL/L. (B) Peanuts inoculated with <i>A</i>. <i>niger</i> in the absence of mixture. (C) Uninoculated peanuts in the presence of mixture at 1 mL/L. (D) Uninoculated peanuts in the absence of mixture. Lower panel–Mixture B: (A) Uninoculated peanuts in the absence of mixture. (B) Uninoculated peanuts in the presence of mixture at 0.05 mL/L. (C) Uninoculated peanuts in the presence of mixture at 0.5 mL/L. Arrows indicate the development of intrinsic <i>Aspergillus</i> sp.</p
<p>(A) Eggs, (B) L1, (C) L4 and (D) YA larval stages. Eggs or nematodes were exposed to 3-m... more <p>(A) Eggs, (B) L1, (C) L4 and (D) YA larval stages. Eggs or nematodes were exposed to 3-methyl-1-butanol (0.1 mL, 0.92 mmole), (±)-2-methyl-1-butanol (0.1 mL, 0.93 mmole), 4-heptanone (0.2 mL, 1.43 mmole), isoamyl acetate (0.1 mL, 0.68 mmole), or up to 0.5 mL of SVM. The viable nematodes were passed through sieves and counted. Data on the y-axis are presented as no. of eggs hatching (A) and as percent mortality relative to controls (mean ± SE) (B-D). Graphs are representative of three independent experiments. Pairwise comparison of the means was performed with analysis of variance followed by post-hoc Tukey–Kramer multiple comparison test. Different letters indicate significantly different values (<i>P</i> ≤ 0.05).</p
<p>Biological activity of each chemical component consisting 1 mL/L (air space) of Mixture ... more <p>Biological activity of each chemical component consisting 1 mL/L (air space) of Mixture A</p
<p>(A) Control swollen fruits. (B) Swollen fruits in the presence of one culture dish of &l... more <p>(A) Control swollen fruits. (B) Swollen fruits in the presence of one culture dish of <i>D</i>. <i>cf</i>. <i>concentrica</i>. (C) Swollen fruits in the presence of two culture dishes of <i>D</i>. <i>cf</i>. <i>concentrica</i>.</p
<p>In each treatment the nematodes were incubated for 24 or 48 h in the presence or absence... more <p>In each treatment the nematodes were incubated for 24 or 48 h in the presence or absence of 4-heptanone (0.4 mL, 2.86 mmole), before viable J2s were separated using 30 μm sieves. The numbers on the y-axis represent the numbers (mean ± SE) of viable J2s counted following the incubation. There were 8–10 repetitions, each using 300 J2s. In the treatment designated as "4-heptanone 24 h + recovery 24 h" the nematodes were exposed to the compound for 24 h and then removed to a new container with no VOCs for recovery. The results were subjected to analysis of variance followed by the Tukey-Kramer multiple comparison test; different letters above the bars indicate a significant difference between samples at <i>P ≤</i> 0.05. The experiment was independently repeated three times, each time with similar results.</p
<p>J2s were exposed for 48 h either to the SVM at 1 mL/L (V/V) [3-methyl-1-butanol (0.2 mL,... more <p>J2s were exposed for 48 h either to the SVM at 1 mL/L (V/V) [3-methyl-1-butanol (0.2 mL, 1.84 mmole), (±)-2-methyl-1-butanol (0.2 mL, 1.86 mmole), 4-heptanone (0.4 mL, 2.86 mmole), and isoamyl acetate (0.2 mL, 1.35 mmole)] or to three 50-mm-diameter fungal culture plates, each with 5 mL of growth medium, and then the viable J2s were separated using 30 μm sieves. The numbers on the y-axis represent the numbers (mean ± SE) of viable J2s counted following the incubation. Each treatment was composed of 10 technical repetitions using 300 J2s in each repetition. The results were subjected to analysis of variance followed by the Tukey-Kramer multiple comparison test; different letters above the bars indicate a significant difference between samples at <i>P ≤</i> 0.05. The experiment was independently repeated three times, each time with similar results.</p
The bionematicidal effect of a synthetic volatile mixture (SVM) of four volatile organic compound... more The bionematicidal effect of a synthetic volatile mixture (SVM) of four volatile organic compounds (VOCs) emitted by the endophytic fungus Daldinia cf. concentrica against the devastating plant-parasitic root-knot nematode Meloidogyne javanica has been recently demonstrated in both in vitro and greenhouse experiments. However, the mode of action governing the observed irreversible paralysis of J2 larvae upon exposure to SVM is unknown. To unravel the mechanism underlying the anthelmintic and nematicidal activities, we used the tractable model worm Caenorhabditis elegans. C. elegans was also susceptible to both the fungal VOCs and SVM. Among compounds comprising SVM, 3-methyl-1-butanol, (±)-2-methyl-1-butanol, and 4-heptanone showed significant nematicidal activity toward L1, L4 and young adult stages. Egg hatching was only negatively affected by 4-heptanone. To determine the mechanism underlying this activity, we examined the response of C. elegans mutants for glutamate-gated chloride channel and acetylcholine transporter, targets of the nematicidal drugs ivermectin and aldicarb, respectively, to 4-heptanone and SVM. These aldicarb-and ivermectin-resistant mutants retained susceptibility upon exposure to 4-heptanone and SVM. Next, we used C. elegans TJ356 strain zIs356 (daf-16::GFP+rol-6), LD1 ldIs7 [skn-1B/C::GFP + pRF4(rol-6(su1006))], LD1171 ldIs3 [gcs-1p::gfp; rol-6 (su1006))], CL2166 dvIs19 (gst-4p::GFP) and CF1553 muIs84 (sod-3p::GFP+rol-6), which have mutations in genes regulating multiple stress responses. Following exposure of L4 larvae to 4-heptanone or SVM, there was clear nuclear translocation of DAF-16::GFP, and SKN-1::GFP indicating that their susceptibility involves DAF-16 and SKN1 regulation. Application of 4-heptanone, but not SVM, induced increased expression of, gcs-1::GFP and gst-4::GFP compared to controls. In contrast, application of 4-heptanone or SVM to the sod-3:: GFP line elicited a significant decline in overall fluorescence intensity compared to controls,
BACKGROUNDSclerotium rolfsii is a soil‐borne phytopathogenic fungus that causes diseases in econo... more BACKGROUNDSclerotium rolfsii is a soil‐borne phytopathogenic fungus that causes diseases in economically important crops. Eradication of the fungus is hampered by its wide range of hosts, as well as its capacity to form sclerotia. Recently, we have shown that the endophytic fungus Daldinia cf. concentrica emits biologically active volatile organic compounds (VOCs); we also demonstrated that one VOC, trans‐2‐octenal, was the most effective against various phytopathogenic fungi. Thus, the aim of this study was to examine the potential of this compound to control hyphae and sclerotia of S. rolfsii, both in vitro and in soil.RESULTSWe found that in vitro exposure of S. rolfsii mycelium to trans‐2‐octenal in air fully inhibits and kills the fungus. Elimination of sclerotia viability occurred at the same concentration, but direct contact between the sclerotia and the compound was needed. trans‐2‐Octenal also affected the viability of both hyphae and sclerotia of S. rolfsii in small pots c...
Plant-parasitic nematodes form one of the largest sources of biotic stress imposed on plants, and... more Plant-parasitic nematodes form one of the largest sources of biotic stress imposed on plants, and are very difficult to control; among them are the obligate parasites, the sedentary root-knot nematodes (RKNs)-Meloidogyne spp.-which are extremely polyphagous and exploit a very wide range of hosts. Endophytic fungi are organisms that spend most of their life cycle within plant tissue without causing visible damage to the host plant. Many endophytes secrete specialized metabolites and/or emit volatile organic compounds (VOCs) that exhibit biological activity. Recently, we demonstrated that the endophytic fungus Daldinia cf. concentrica secrets biologically active VOCs. Here we examined the ability of the fungus and its VOCs to control the RKN M. javanica both in vitro and greenhouse experiments. The D. cf. concentrica VOCs showed bionematicidal activity against the second-stage juveniles (J2s) of M. javanica. We found that exposure of J2s to fungal volatiles caused 67% reduction in via...
Endophytic fungi are organisms that spend most of their life cycle within plant tissues without c... more Endophytic fungi are organisms that spend most of their life cycle within plant tissues without causing any visible damage to the host plant. Many endophytes were found to secrete specialized metabolites and/or emit volatile organic compounds (VOCs), which may be biologically active and assist fungal survival inside the plant as well as benefit their hosts. We report on the isolation and characterization of a VOCs-emitting endophytic fungus, isolated from an olive tree (Olea europaea L.) growing in Israel; the isolate was identified as Daldinia cf. concentrica. We found that the emitted VOCs were active against various fungi from diverse phyla. Results from postharvest experiments demonstrated that D. cf. concentrica prevented development of molds on organic dried fruits, and eliminated Aspergillus niger infection in peanuts. Gas chromatography-mass spectrometry analysis of the volatiles led to identification of 27 VOCs. On the basis of these VOCs we prepared two mixtures that displ...
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