Enhanced production of pinosylvin stilbene with aging of Pinus strobus callus and nematicidal activity of callus extracts against pinewood nematodes

AbstractPinosylvin stilbenes are phenolic compounds mainly occurring in the Pinaceae family. We previously reported that the accumulation of two pinosylvin stilbene compounds, dihydropinosylvin methyl ether (DPME) and pinosylvin monomethyl ether (PME), in Pinus strobus trees was highly enhanced by infection with pine wood nematodes (PWNs: Bursaphelenchus xylophilus), and these two compounds showed strong nematicidal activity against PWNs. In this work, we established a system of pinosylvin stilbene (DPME and PME) production via the in vitro culture of P. strobus calli, and we examined the nematicidal activity of callus extracts. Calli were induced from the culture of mature zygotic embryos of P. strobus. Optimized growth of calli was obtained in 1/2 Litvay medium with 1.0 mg/L 2,4-D and 0.5 mg/L BA. DPME and PME accumulation did not occur in nonaged (one-month-old) calli but increased greatly with prolonged callus culture. The concentrations of DPME and PME in three-month-old dark-brown calli were 6.4 mg/g DW and 0.28 mg/g DW, respectively. The effect of methyl jasmonate treatment on the accumulation of DPME and PME was evaluated in cell suspension culture of P. strobus. However, the treatment appeared to show slight increase of DPME accumulation compared to callus browning. A test solution prepared from crude ethanol extracts from aged calli (three months old) containing 120 µg/ml DPME and 5.16 µg/ml PME treated with PWNs resulted in 100% immobilization of the adult PWNs and 66.7% immobilization of the juvenile PWNs within 24 h. However, nonaged callus extracts did not show any nematicidal activity against juvenile PWNs and showed less than 20% nematicidal activity against adult PWNs. These results indicate that pinosylvin stilbenes can be effectively produced by prolonged culture of P. strobus calli, can be isolated using simple ethanolic extraction, and are applicable as beneficial eco-friendly compounds with nematicidal activity against PWNs.

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Enhanced Production of Pinosylvin Stilbene with Aging of Pinus strobus Callus and Nematicidal Activity of Callus Extracts against Pinewood Nematodes

10.21203/rs.3.rs-849585/v1 ◽

2021 ◽

Author(s):

Hyo Bin Koo ◽

Hwan-Su Hwang ◽

Jung Yeon Han ◽

Eun Ju Cheong ◽

Yong-Soo Kwon ◽

...

Keyword(s):

Pinus Strobus ◽

Monomethyl Ether ◽

Bursaphelenchus Xylophilus ◽

Nematicidal Activity ◽

Zygotic Embryos ◽

Enhanced Production ◽

Pinewood Nematodes ◽

Ethanolic Extraction ◽

Ethanol Extracts

Abstract Pinosylvin stilbenes are phenolic compounds mainly occurring in the Pinaceae family. We previously reported that the accumulation of two pinosylvin stilbene compounds, dihydropinosylvin methyl ether (DPME) and pinosylvin monomethyl ether (PME), in Pinus strobus trees was highly enhanced by infection with pine wood nematodes (PWNs: Bursaphelenchus xylophilus), and these two compounds showed strong nematicidal activity against PWNs. In this work, we established a system of pinosylvin stilbene (DPME and PME) production via the in vitro culture of P. strobus calli, and we examined the nematicidal activity of callus extracts. Calli were induced from the culture of mature zygotic embryos of P. strobus. Optimized growth of calli was obtained in 1/2 Litvay medium with 1.0 mg/L 2,4-D and 0.5 mg/L BA. DPME and PME accumulation did not occur in nonaged (one-month-old) calli but increased greatly with prolonged callus culture. The concentrations of DPME and PME in three-month-old dark-brown calli were 6.4 mg/g DW and 0.28 mg/g DW, respectively. A test solution prepared from crude ethanol extracts from aged calli (three months old) containing 120 µg/ml DPME and 5.16 µg/ml PME treated with PWNs resulted in 100% immobilization of the adult PWNs and 66.7% immobilization of the juvenile PWNs within 24 hr. However, nonaged callus extracts did not show any nematicidal activity against juvenile PWNs and showed less than 20% nematicidal activity against adult PWNs. These results indicate that pinosylvin stilbenes can be effectively produced by prolonged culture of P. strobus calli, can be isolated using simple ethanolic extraction, and are applicable as beneficial eco-friendly compounds with nematicidal activity against PWNs.

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Enhanced accumulation of pinosylvin stilbenes and related gene expression in Pinus strobus after infection of pine wood nematode

Tree Physiology ◽

10.1093/treephys/tpab053 ◽

2021 ◽

Author(s):

Hwan-Su Hwang ◽

Jung Yeon Han ◽

Yong Eui Choi

Keyword(s):

Transcription Factors ◽

Defense Mechanisms ◽

De Novo ◽

Pinus Strobus ◽

Pine Wilt Disease ◽

Monomethyl Ether ◽

Bursaphelenchus Xylophilus ◽

Nematicidal Activity ◽

Dependent Manner ◽

Pine Wood

Abstract Pine wood nematodes (PWNs: Bursaphelenchus xylophilus) infect pine trees and cause serious pine wilt disease. Eastern white pine (Pinus strobus) has resistance to PWN. However, the detailed defense mechanisms of P. strobus against PWN are not well known. When P. strobus plants were infected with PWNs, the accumulation of stilbenoids, dihydropinosylvin monomethyl ether (DPME) and pinosylvin monomethyl ether (PME), were increased remarkably. DPME and PME had the high nematicidal activity. Interestingly, the nematicidal activity of the two compounds was resulted in a developmental stage-dependent manner. PME was more toxic to adult PWNs than juveniles, whereas DPME was found more toxic to juvenile PWNs than the adults. The genes involved in PME and DPME biosynthesis such as phenylalanine ammonia-lyase (PAL), 4-coumarate-CoA ligase (4CL), pinosylvin synthase (STS), and pinosylvin O-methyltransferase (PMT) were isolated using de novo sequencing of the transcriptome in P. strobus. In addition, transcription factors (bHLH, MYB and WRKY) related to stilbene biosynthesis were isolated. qPCR analyses of the selected genes (PAL, 4CL, STS, and PMT) including transcription factors (bHLH, MYB and WRKY) revealed that the expression level of the selected genes highly enhanced after PWN infection. Our results suggest that pinosylvin-type stilbenoid biosynthesis is highly responsive to PWN infection and plays an important role in PWN resistance of P. strobus trees.

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Morphometric analysis of reserve substances and ultrastructural changes during caulogenic determination and loss of competence of Eastern White pine (Pinus strobus) cotyledons in vitro

Canadian Journal of Botany ◽

10.1139/b89-105 ◽

1989 ◽

Vol 67 (3) ◽

pp. 779-789 ◽

Cited By ~ 18

Author(s):

Barry S. Flinn ◽

David T. Webb ◽

William Newcomb

Keyword(s):

Basal Medium ◽

Pinus Strobus ◽

Adventitious Shoot ◽

Cell Types ◽

Nuclear Size ◽

Ultrastructural Changes ◽

Zygotic Embryos ◽

White Pine ◽

Mitochondrial Area

Lipid, starch, and protein reserves were monitored cytochemically and, along with certain ultrastructural changes, were quantified morphometrically during adventitious shoot determination or loss of competence in cotyledons from cultured Pinus strobus zygotic embryos. Initial expiants were rich in lipid, which declined during culture. There was more lipid retention in explants on cytokinin medium by day 5 than on basal medium. However, by day 7, five- and six-celled clusters showed greater lipid utilization than most other cells on cytokinin, which may have indicated a shift towards shoot determination in these cells. Except for an initially greater retention on cytokinin, a similar pattern for storage protein degradation was observed in both treatments. Starch levels increased during culture, but did not differ between treatments during the first 7 days. Vacuolation increased during culture and was greater on basal medium than on cytokinin. Relative nuclear size also increased, but was greater on cytokinin. Relative mitochondrial area increased during culture, except in noncluster cells on basal medium. While few differences were noted between cell types within each treatment, distinct differences in nuclear size, vacuolation, and lipid content existed between cells on cytokinin and basal medium. These coincided with the timing of caulogenic determination in cytokinin-treated expiants. The loss of lipid reserves, as well as ultrastructural changes associated with the maturation of cells on basal medium, was associated with the loss of competence.

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Nematicidal activity of compounds extracted from medicinal plants against the pine wood nematode Bursaphelenchus xylophilus

Nematology ◽

10.1163/156854109x424353 ◽

2009 ◽

Vol 11 (6) ◽

pp. 835-845 ◽

Cited By ~ 6

Author(s):

Dang-Minh-Chanh Nguyen ◽

Van-Nam Nguyen ◽

Dong-Jun Seo ◽

Ro-Dong Park ◽

Woo-Jin Jung

Keyword(s):

Medicinal Plants ◽

Plant Extracts ◽

Bursaphelenchus Xylophilus ◽

Nematicidal Activity ◽

Hexane Fraction ◽

Bark Extract ◽

Pine Wood Nematode ◽

Pine Wood ◽

Significant Difference

Abstract To investigate in vitro nematicidal activity against the pine wood nematode, Bursaphelenchus xylophilus, plant methanolic extracts were obtained from 26 medicinal plants and herbs in Vietnam. Of the plant extracts tested, a 10 mg ml−1 concentration of Cinnamomum cassia bark extract showed the highest level of nematicidal activity against juvenile and adult B. xylophilus after treatment for 7 h. The remainder of the plant extracts had little or no effect on juveniles or adults. Four solvents, n-hexane, chloroform, ethyl acetate and n-butanol, were used initially to partition the compounds obtained from C. cassia extracts. The compound (5 mg ml−1) in the n-hexane fraction showed the highest nematicidal activity at 60 min after treatment. Compounds III and IV were then separated from the n-hexane fraction using silica gel plates. These compounds showed very similar activity against the pine wood nematode. However, treatment with different concentrations of these compounds resulted in a significant difference in their effects on the mortality of the pine wood nematode, with greater concentrations leading to increased mortality after the same exposure time. Overall, treatment with 3 mg ml−1 of compounds III and IV resulted in greater than 95% mortality against juvenile and adult B. xylophilus at 50 min after treatment.

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A new strain of Volutella citrinella with nematode predation and nematicidal activity, isolated from the cysts of potato cyst nematodes in China

BMC Microbiology ◽

10.1186/s12866-021-02385-x ◽

2021 ◽

Vol 21 (1) ◽

Author(s):

Xinyue Zhang ◽

Hui Zhang ◽

Zhaochun Jiang ◽

Qing Bai ◽

Shishi Wu ◽

...

Keyword(s):

Fermentation Broth ◽

Bursaphelenchus Xylophilus ◽

Nematophagous Fungi ◽

Plant Diseases ◽

Nematicidal Activity ◽

Parasitic Nematodes ◽

Negative Effects ◽

Cyst Nematodes ◽

Highly Active

Abstract Background Plant parasitic nematodes (PPNs) are responsible for causing many plant diseases and are extremely difficult to control at present. Currently, due to the negative effects of chemical agents on the environment and human health, the development of new biological pesticides has become an important part of plant nematode control. Nematophagous fungi refers to a class of fungi that kill plant nematodes. Notably, a large number of nematophagous fungi resources remain to be studied. The objective of our study was to use in vitro screening to identify nematophagous fungi and select strains that were highly active against nematodes, providing a primary research for the development and utilization of new nematophagous fungi. Results A new nematophagous fungal strain (GUCC2219) was isolated from cysts of possibly Globodera spp. and Heterodera spp., identified as Volutella citrinella. The hyphae of V. citrinella produced ring structures of variable size and exhibited predatory and nematicidal activity. The hyphal predation rates (in vitro) against three species of nematodes, Aphelenchoides besseyi, Bursaphelenchus xylophilus, and Ditylenchus destructor, averaged 59.45, 33.35, and 50.95%, respectively, while the fermentation broth produced by the fungus exhibited mortality rates of 100, 100, and 55.63%, respectively, after 72 h. Conclusion V. citrinella is a new strain with nematophagous properties, which are a novel discovery. At the same time, this is the first report of nematicidal and nematode predation activity in the genus Volutella.

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Anti-amnesic activity screening of the seed ethanol extracts of Turkish Paeonia taxa by in vitro methods

Planta Medica ◽

10.1055/s-0031-1282924 ◽

2011 ◽

Vol 77 (12) ◽

Author(s):

D Sevim ◽

FS Senol ◽

I Orhan ◽

B Şener ◽

E Kaya

Keyword(s):

In Vitro Methods ◽

Activity Screening ◽

Ethanol Extracts

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Evaluation of Snake Venom’s PhospholipaseA2 Enzyme Inhibition Activity of Cyphostemma adenocoule

Current Chemical Biology ◽

10.2174/2212796814999200917114914 ◽

2020 ◽

Vol 14 (3) ◽

pp. 196-202

Author(s):

Atul Kaushik ◽

Teamrat S. Tesfai ◽

Daniel K. Barkh ◽

Furtuna K. Ghebremeskel ◽

Habtom G. Zerihun ◽

...

Keyword(s):

Enzyme Inhibition ◽

Snake Venom ◽

Ethanol Extract ◽

Egg Yolk ◽

Chloroform Extract ◽

Inhibition Activity ◽

Traditional Use ◽

Snake Bites ◽

Ethanol Extracts

Background: A snake bite is fundamentally an injury often resulting in puncture wounds meted out by the animal's fangs and occasionally resulting in envenomation. Rate of snake bites around 5,400,000 bites per year leads to over 2,500,000 envenomings and around 125,000 fatal cases annually. Snake venom enzymes are rich in metalloproteinases, phospholipaseA2, proteinases, acetylcholinesterases and hyaluronidases. Objective: Cyphostemma adenocoule is traditionally being used for the treatment of snake bites in Eritrea. The present research was aimed at evaluating the snake venom enzyme inhibition activity of C. adenocoule against puff adder venom and developing a base for the traditional use of the plant against snakebites in Eritrea. Methods: The anti-venom activity of C. adenocoule was assessed in-vitro through phospholipaseA2 enzyme inhibition assay using egg yolk as a cell. The ethanol and chloroform extracts of C. adenocoule showed in vitro anti phospholipase A2 activity, whereas the water extracts of the plant showed no activity. Results: Among the extracts of C. adenocoule, the highest percentage of inhibition was obtained from chloroform extract (95.55% at 100mg/ml). The extract showed prominent activity at different concentrations (34.7% at10mg/ml, 48.8% at 20mg/ml, 54.8% at 40mg/ml, 60.9% at 60mg/ml, 80.5% at 80mg /ml). The ethanol extract also showed certain activity at various concentrations (25.22% at10mg/ml, 14.78% at 20mg/ml, 2.6% at40mg/ml). The activity of the chloroform extracts increases as concentration increases, whereas the activity of the ethanol extracts decreases as concentration increases. The aqueous extract of C. adenocoule did not show any activity at all concentrations. Conclusion: In this study, the chloroform and ethanol extracts of the plant inhibited the enzyme of interest and thus proved the efficacy of anti-snake venom activity of the plant.

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Novel physiological properties of ethanol extracts from Eremurus chinensis Fedtsch. roots: in vitro antioxidant and anticancer activities

Food & Function ◽

10.1039/c2fo30152k ◽

2012 ◽

Vol 3 (12) ◽

pp. 1310 ◽

Cited By ~ 9

Author(s):

Haifang Xiao ◽

Yutang Wang ◽

Qisen Xiang ◽

Chunxia Xiao ◽

Li Yuan ◽

...

Keyword(s):

Anticancer Activities ◽

Physiological Properties ◽

Ethanol Extracts ◽

Roots In Vitro

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Characterization of Alternaria Species Associated with Heart Rot of Pomegranate Fruit

Journal of Fungi ◽

10.3390/jof7030172 ◽

2021 ◽

Vol 7 (3) ◽

pp. 172

Author(s):

Francesco Aloi ◽

Mario Riolo ◽

Simona Marianna Sanzani ◽

Annamaria Mincuzzi ◽

Antonio Ippolito ◽

...

Keyword(s):

Elongation Factor ◽

Monomethyl Ether ◽

Alternariol Monomethyl Ether ◽

Translation Elongation ◽

Microscopic Features ◽

Alternaria Species ◽

Rot Disease ◽

Multigene Phylogenetic Analysis ◽

Pomegranate Fruit

This study was aimed at identifying Alternaria species associated with heart rot disease of pomegranate fruit in southern Italy and characterizing their mycotoxigenic profile. A total of 42 Alternaria isolates were characterized. They were obtained from pomegranate fruits with symptoms of heart rot sampled in Apulia and Sicily and grouped into six distinct morphotypes based on macro- and microscopic features. According to multigene phylogenetic analysis, including internal transcribed spacer (ITS), translation elongation factor 1-α (EF-1α), glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and a SCAR marker (OPA10-2), 38 isolates of morphotypes 1 to 5 were identified as Alternaria alternata, while isolates of morphotype 6, all from Sicily, clustered within the Alternaria arborescens species complex. In particular, isolates of morphotype 1, the most numerous, clustered with the ex-type isolate of A. alternata, proving to belong to A. alternata. No difference in pathogenicity on pomegranate fruits was found between isolates of A. alternata and A. arborescens and among A. alternata isolates of different morphotypes. The toxigenic profile of isolates varied greatly: in vitro, all 42 isolates produced tenuazonic acid and most of them other mycotoxins, including alternariol, alternariol monomethyl ether, altenuene and tentoxin.

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Analytical Profiling of Proanthocyanidins from Acacia mearnsii Bark and In Vitro Assessment of Antioxidant and Antidiabetic Potential

Molecules ◽

10.3390/molecules23112891 ◽

2018 ◽

Vol 23 (11) ◽

pp. 2891 ◽

Cited By ~ 8

Author(s):

Xiao Chen ◽

Jia Xiong ◽

Shenlin Huang ◽

Xun Li ◽

Yu Zhang ◽

...

Keyword(s):

Antioxidant Activity ◽

Degree Of Polymerization ◽

Acacia Mearnsii ◽

Oligomeric Proanthocyanidins ◽

Normal Phase Hplc ◽

Inhibitory Activities ◽

Ethanol Extracts ◽

Alternative Drugs ◽

Tof Ms

The proanthocyanidins from ethanol extracts (80%, v/v) of Acacia mearnsii (A. mearnsii) bark on chemical-based and cellular antioxidant activity assays as well as carbolytic enzyme inhibitory activities were studied. About 77% of oligomeric proanthocyanidins in ethanol extracts of A. mearnsii bark were found by using normal-phase HPLC. In addition, HPLC-ESI-TOF/MS and MALDI-TOF/TOF MS analyses indicated that proanthocyanidins from A. mearnsii bark exhibited with a degree of polymerization ranging from 1 to 11. These results of combined antioxidant activity assays, as well as carbolytic enzyme inhibitory activities of proanthocyanidins from A. mearnsii bark, indicated an encouraging antioxidant capacity for the high polyphenol content and a potential for use as alternative drugs for lowering the glycemic response.

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