Use of polysaccharides from calophylum inophyllum for the preparation of antiviral agents for plants

Abstract

The application discloses application of a polysaccharide of Ossyra edulis in preparation of an anti-plant virus agent. The polysaccharide is obtained by alkali extraction and alcohol precipitation of dry sclerotia of Ossyra edulis. The application finds that the polysaccharide of Ossyra edulis shows significant anti-virus activity on various plant pathogenic viruses in a pot experiment, and its preparation also shows good activity in a field efficacy test, and is significantly better than a positive control agent. Compared with the prior art, the application develops an anti-virus agent pesticide from a biological source of natural substances, has good environmental compatibility, safety, high efficiency, green environmental protection and the like, and has a wide application prospect.

Description

Technical Field

[0001] This invention belongs to the field of fungi and their applications, specifically relating to the application of *Lycopodium clavatum* polysaccharide in the preparation of antiviral agents for plants. Background Technology

[0002] Plant viral diseases, also known as "plant cancer," are the second largest plant disease, causing economic losses of up to US$60 billion worldwide each year, of which food crops alone account for US$20 billion. Plant viral diseases have caused serious damage to my country's food production. The prevalence of wheat soil-borne mosaic virus and wheat dwarf virus has led to a 20% to 30% reduction in wheat yield in northern my country; the prevalence of rice viral diseases in southern China has led to a 20% to 30% reduction in rice yield; and the prevalence of cucumber mosaic virus and tobacco mosaic virus has led to a reduction in the yield of various fruits and vegetables (Xiao Qinzhi, Deng Bin, Zou Hailu, Teng Kai, Tang Qianjun, Zhou Zhicheng. Research progress on biological control of plant viral diseases [J]. Southern Agriculture, 2021, 15(34):64-69.).

[0003] Due to the absolute parasitism of plant viruses on their hosts, controlling plant viral diseases and developing plant virus inhibitors are extremely difficult (Fan HT, Song BA, Bhadury PS, et al. Antiviral Activity and Mechanism of Action of Novel Thiourea Containing Chiral Phosphonate on Tobacco Mosaic Virus. International Journal of Molecular Sciences, 2011, 12(7), 4522-4535.). Currently, no agent can completely protect plants from plant virus infection, nor can any agent eliminate plant pathogens from infected plant tissues under field conditions (Chen J, Yan XH, Dong JH, et al. Tobacco Mosaic Virus (TMV) Inhibitors from Picrasma quassioides Benn. Journal of Agricultural and Food Chemistry, 2009, 57(15), 6590-6595.). Summary of the Invention

[0004] This invention provides the application of *Lycopodium clavatum* polysaccharide in the preparation of antiviral agents for plants, wherein the structure of the *Lycopodium clavatum* polysaccharide is shown in Formula I:

[0005]

[0006] Where 10≤n≤700.

[0007] The polysaccharide from *O. lapidescens* described in this invention is isolated from the fungus *O. lapidescens*. An optional isolation method includes crushing the sclerotia of *O. lapidescens*, extracting with alkaline water, adding ethanol to the supernatant for precipitation, collecting the precipitate, and removing proteins to obtain the final product.

[0008] The present invention also provides an antiviral agent for plants, comprising the above-mentioned *Lycopodium clavatum* polysaccharide and adjuvants. Optionally, the content of *Lycopodium clavatum* polysaccharide is 1% to 80% by mass percentage. Preferably, it is 10%.

[0009] The antiviral agent of the present invention is an agriculturally acceptable formulation type, such as an aqueous solution, soluble liquid, microemulsion, soluble powder, water emulsion, suspension or water-dispersible granule.

[0010] Omphalia lapidescens Schroet., a fungus belonging to the genus Omphalia of the family Tricholomataceae, is a dried sclerotium. Its fruiting body has a short lifespan, and the most common form is the sclerotium, which typically grows near the roots of bamboo, paulownia, or pine trees, and has historically been found in the wild. Modern pharmacological studies have shown that it possesses tapeworm-expelling, anti-inflammatory, and immunomodulatory effects, and can be used to treat tumors and enhance the body's immunity. This invention further isolates and purifies the antiviral active ingredients of Omphalia lapidescens extract and prepares an antiviral agent for plants with Omphalia lapidescens polysaccharides as the active ingredient.

[0011] The polysaccharide and related antiviral agents of this invention exhibit significant antiviral activity and can be used on various food crops and cash crops, including but not limited to Tobacco mosaic virus (TMV), Cucumber mosaic virus (CMV), Pepper mild mottle virus (PMMoV), Potato virus Y (PVY), Wheat yellow mosaic virus (WYMV), and Rice black-streaked dwarf virus (RBSDV). These agents have advantages such as low pathogen resistance, a relatively broad control spectrum, and compatibility with chemical agents, making them a pesticide that meets the requirements of green pest control. Attached Figure Description

[0012] Figure 1 For the determination of the molecular weight of the polysaccharide from Leiwan as described in this invention;

[0013] Figure 2The infrared absorption spectrum of the *Leiwan* polysaccharide described in this invention;

[0014] Figure 3 The nuclear magnetic resonance hydrogen spectrum (500MHz, solvent: d6-DMSO) of the Leiwan polysaccharide described in this invention;

[0015] Figure 4 The nuclear magnetic resonance carbon spectrum (125MHz, solvent: d6-DMSO) of the polysaccharide of Leiwan described in this invention;

[0016] Figure 5 The polysaccharide of Leiwan described in this invention 1 H- 1 H COSY related spectra;

[0017] Figure 6 The HSQC correlation spectrum of the polysaccharide from Leiwan described in this invention;

[0018] Figure 7 This is a gas chromatography-mass spectrometry (GC-MS) spectrum of the acetylated derivative of the hydrolysis product of the *Lycopodium clavatum* polysaccharide described in this invention. Detailed Implementation

[0019] Unless otherwise specified, the scientific and technical terms used in this article are intended for understanding by those skilled in the art.

[0020] The technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0021] The *Leiwan* polysaccharide disclosed in this invention can be a commercially available product or obtained through various techniques, such as water extraction, acid extraction, alkali extraction, enzyme extraction, ultrasonic-assisted extraction, microwave-assisted extraction, and supercritical fluid extraction. Materials used to prepare the *Leiwan* polysaccharide can include *Leiwan* sclerotia and *Leiwan* fermentation broth. The method for preparing the *Leiwan* polysaccharide used in this disclosure involves crushing the sclerotia of the fungus *Leiwan*, extracting them with alkaline water, adding ethanol to the supernatant for precipitation, collecting the precipitate, removing proteins, and purifying the precipitate. The *Leiwan* polysaccharide used in the following examples was obtained using the following method:

[0022] 200g of dried sclerotia of *Leiwan* were pulverized using a pulverizer and extracted with 2L of 0.1mol / L NaOH aqueous solution under hot reflux for 2h. This process was repeated 6 times. The filtrates were combined and centrifuged at 12000G for 30min. The supernatant was collected. Ethanol was slowly added to the supernatant with stirring until it reached 20% of the total volume. The mixture was allowed to stand overnight at 4°C and then centrifuged to collect the precipitate. After removing protein from the precipitate using the TCA method, the sugar content was determined to be 92.4%. Gel chromatography analysis showed that it was a polysaccharide with a relatively concentrated molecular weight, and it was named *Leiwan* polysaccharide.

[0023] Molecular weight determination results showed that there were two component peaks in Leiwan polysaccharide ( Figure 1 The first component has a weight-average molecular weight Mw of 338827 Da, a number-average molecular weight Mn of 172916 Da, and a molecular weight distribution width (Mw / Mn) of 1.959. The second component has a weight-average molecular weight Mw of 10923 Da, which is significantly different from the first component. It is speculated that the second component may have been generated by the hydrolysis and breakage of the first component.

[0024] After hydrolysis, reduction, and acetylation, GC-MS analysis showed that a single acetylated product was mainly produced from the polysaccharide of *Lycopodium clavatum*. Figure 7 The corresponding monosaccharide is glucose, proving that it is a homogeneous polysaccharide composed of glucose.

[0025] Ultraviolet full-wavelength scanning of *Lycopodium clavatum* polysaccharide revealed no ultraviolet absorption in the 220-400 nm wavelength range, indicating the absence of protein. The infrared spectrum of *Lycopodium clavatum* polysaccharide is shown below. Figure 2 The infrared spectrum is shown at 3450.9 cm⁻¹. -1 The presence of a strong absorption peak at 1435.42 cm⁻¹ indicates the presence of intermolecular hydroxyl groups in the polysaccharide. This peak corresponds to the stretching vibration of the -OH group. -1 The absorption peak belongs to the CH2 symmetric stretching vibration peak and the -OH in-plane deformation vibration peak, 1336.88 cm⁻¹. -1 Belongs to the CH2 shear vibration peak, 1000-1200cm -1 The peaks, belonging to the CH deformation vibration and CO stretching vibration, are basically consistent with the standard pyranose absorption peaks, indicating that Leiwan polysaccharide is a typical pyranose; the peak at 877.30 cm⁻¹ appears. -1 The absorption peak at 1632.86 cm⁻¹ is the anomeric stretching vibration peak of the β-configuration polysaccharide. -1 The peak at this point is the characteristic absorption peak of a hemiacetal.

[0026] The nuclear magnetic resonance spectrum of *Leiwan* polysaccharide exhibits typical β-glucan characteristics. 1 H-NMR spectrum ( Figure 3 Two main hydrogen signals on the terminal carbons (δ) can be observed in the sample. H 4.06 and δ H(4.27), indicating that there are at least two ways of linking glucose in its structure. The proton signal on the non-terminal carbon is mainly located at δ H Range: 3.00-3.69. 13 C-NMR spectrum ( Figure 4 A coincident terminal carbon signal (δ) can be observed in the sample. C (103.57), further utilization 1 H- 1 H COSY( Figure 5 ) and HSQC ( Figure 6 The relevant spectra were used to assign the C2-C6 and C2-H-C6-H signals. It is worth noting that the carbon signals of Leiwan polysaccharide all showed broad peaks, and C-2, C-4, and C-5 also showed multiple sets of peaks, indicating that the glucose units in Leiwan polysaccharide are in different chemical environments. However, due to the limited clarity of the NMR spectra, more effective information could not be provided in this regard.

[0027] The periodic acid oxidation reaction results indicate that the Leiwan polysaccharide molecule contains (1-6) glycosidic bonds. The Smith degradation products do not contain erythritol, but contain a small amount of glycerol and a large amount of glucose. This proves that the Leiwan polysaccharide molecule contains (1-6) and (1-3) glycosidic bonds. The gas chromatography-mass spectrometry results of the fully methylated hydrolysis products show that the Leiwan polysaccharide contains 1-3, 1-6, and 1,3-6 glycosidic bonds, and the molar ratio of 2,4,6-Me3-Glcp, 2,3,4,6-Me4-Glcp, and 2,4-Me2-Glcp is approximately 1:2:2 (Table 1). This indicates that there are three types of glucose substitution in the basic repeating unit of the Leiwan polysaccharide, namely 1,3-substituted, 1-substituted, and 1,3,6-substituted, with a content ratio of 1:2:2.

[0028] Table 1. Methylation analysis results of *Lycopodium clavatum* polysaccharides

[0029] Methylated sugars Retention time Connection type Peak area ratio 2,4,6-Me3-Glcp 13.562 <![CDATA[→3)- Glcp -(1→]]> 1 2,3,4,6-Me4-Glcp 12.865 <![CDATA[ Glcp -(1→]]> 2 2,4-Me2-Glcp 15.549 <![CDATA[→3,6)- Glcp -(1→]]> 2

[0030] Comparing the above NMR data and glycoderivatization structure analysis data, and comparing them with the literature (K. Saito, M. Nishijima, T. Miyazaki, Further examination on the structure of an alkali-soluble glucan isolated from Omphalia lapidescens. Studies on fungal polysaccharide. XXXVI. Chemical and Pharmaceutical Bulletin, 1990, 38(6): 1745-1747), the following structure of Leiwan polysaccharide is inferred:

[0031]

[0032] The range of n is: 10≤n≤700.

[0033] Example 1: Determination of the antiviral activity of *Lycopodium clavatum* polysaccharide against plant viruses using a greenhouse pot culture method.

[0034] Pot experiments were conducted on tobacco to control tobacco mosaic virus (TMV, MN186255.1) with Leiwan polysaccharide; on rice to control Southern Rice Black-Streaked Dwarf Virus (SRBSDV, MN244978.1) with Leiwan polysaccharide inducers; on wheat to control wheat yellow mosaic virus (WYMV, MG678447.1) with Leiwan polysaccharide inducers; and on potatoes to control potato Y64 virus. Pot experiments were conducted on the prevention of soybean mosaic virus (SMV, MK868081.1) using a polysaccharide inducer on soybeans, cucumber mosaic virus (CMV, MN792888.1) using a polysaccharide inducer on cucumbers, and pepper mottle virus (PepMoV, MH476198.1) using a polysaccharide inducer on peppers. All viruses used in the experiments were provided by the Shaanxi Provincial Engineering Technology Research Center for Biological Pesticides, isolated from diseased plants in the field, and preserved in the biopharmaceutical laboratory.

[0035] Prepare aqueous solutions of 0.02 mg / mL, 0.1 mg / mL and 0.5 mg / mL of *Lycopodium clavatum* polysaccharide, and select healthy plants with uniform growth to conduct pot experiments on plant virus resistance, and determine the protective, in vitro passivation and disease-curing activities respectively.

[0036] The protective activity assay method was as follows: the potted plants were sprayed with the agent, and after 48 hours, they were inoculated with a virus solution diluted 2000 times (the concentration of the diluted virus solution was 10 μg / mL). The therapeutic activity assay method was the reverse: the potted plants were sprayed with the agent after inoculation with a virus solution diluted 2000 times, and after 48 hours, they were sprayed with the agent. The in vitro passivation activity assay was performed by preparing a solution with a concentration 1 times higher than the above agent concentration, mixing it evenly with a virus solution diluted 1000 times, passivating for 1 hour, and then inoculating. Each treatment was repeated 3 times. The blank control was water treatment, and the positive controls were 0.1 mg / mL lentinan and 0.1 mg / mL ningnanmycin.

[0037] After the onset of symptoms, the viral particle content in the leaves was measured using the ELISA method, and the inhibition rate was calculated.

[0038] Inhibition rate (%) = (Virus particle content in blank control - Virus particle content in treatment) / Virus particle content in blank control × 100

[0039] The results are shown in Table 2.

[0040] Table 2: Pot experiment on the control of plant viruses by polysaccharides from *Lycopodium clavatum*

[0041]

[0042]

[0043]

[0044] As shown in Table 1, Leiwan polysaccharide has a significant protective effect against viruses such as TMV. Its inactivation and inhibition of proliferation are relatively weaker, but they are all much higher than the positive control agents Lentinan and Ningnanmycin.

[0045] Example 2: Preparation of 1% Leiwan Polysaccharide Aqueous Solution

[0046] Weigh 0.1 kg of *Dalbergia latifolia* polysaccharide and dissolve it in 3 kg of 0.01 mol / L NaOH aqueous solution. Then add 1.5 kg of calcium dodecylbenzenesulfonate, 0.5 kg of polyoxyethylene styrene ether, and 0.5 kg of ethylene glycol. Add water to a final volume of 10 kg. Mix thoroughly and stir at 800-1000 rpm for 10-30 minutes to obtain 10 kg of 1% *Dalbergia latifolia* polysaccharide aqueous solution. The stability and appearance of the preparation meet the requirements for commercial formulations.

[0047] Example 3: Preparation of 10% Lycopodium clavatum polysaccharide soluble solution

[0048] Weigh 1 kg of *Lycopodium clavatum* polysaccharide and dissolve it in 3 kg of 0.1 mol / L NaOH aqueous solution. Then add 1.5 kg of calcium dodecylbenzenesulfonate and 0.5 kg of polyoxyethylene styrene ether, and bring the water to a final volume of 10 kg. Mix thoroughly and stir at 800-1000 rpm for 10-30 minutes to obtain 10 kg of 10% *Lycopodium clavatum* polysaccharide soluble concentrate. The stability and appearance of the preparation meet the requirements for commercial formulations.

[0049] Example 4: Preparation of 80% Lycopodium clavatum polysaccharide soluble powder

[0050] Weigh out 8 kg of *Lycopodium clavatum* polysaccharide, 1.7 kg of sodium sulfate, and 0.7 kg of calcium dodecylbenzenesulfonate, mix them thoroughly at room temperature, and dissolve them in a 0.01 mol / L NaOH aqueous solution at room temperature, stirring until fully dissolved. After standing at room temperature for 24 hours, dry in a box dryer at 50-70℃ for 10 days until dry, obtaining a solid. Crush the solid using a pulverizer and pass it through a 320-mesh sieve to obtain 10 kg of 80% *Lycopodium clavatum* polysaccharide soluble powder. The particle size and other appearance of the preparation meet the requirements of commercial preparations.

[0051] Example 5: Field plot efficacy test of three antiviral agents made from *Lycopodium clavatum* polysaccharides

[0052] Field plot efficacy trials were conducted on tobacco, rice, wheat, and cucumber using a polysaccharide antiviral agent to control tobacco mosaic virus (TMV); on rice, rice, and cucumber, rice, and cucumber, respectively, to control southern rice black-streaked dwarf virus (SRBSDV); on wheat, wheat yellow mosaic virus (WYMV); on potatoes, potatoes, and cucumbers, respectively, to control cucumber mosaic virus (CMV).

[0053] The cell experiment was designed with randomized permutations, repeated three times. The cell area was approximately 60m², depending on the actual situation. 2 The experimental sites were selected with uniform fertility, consistent crop planting and management levels, and relatively uniform disease occurrence and severity. Protective rows were set up around each treatment area and the experimental zone. The three *Lysimachia christinae* polysaccharide inducers prepared in Examples 2-4 were used for foliar spraying at constant volume. A 1000-fold dilution of 2% *Lysimachia christinae* aqueous solution and a 1000-fold dilution of 10% *Ningnanmycin* soluble powder were used as positive controls for foliar spraying at constant volume. A blank control of clear water was also included. All tested agents had to be diluted twice. Spraying began at the initial stage of disease, and was repeated every 7 days for a total of 3 times. The incidence rate and disease index were investigated before the first spray and 10 days after the last spray to calculate the control efficacy.

[0054] The disease grading standards are as follows:

[0055] Grade 0: The entire plant is disease-free;

[0056] Grade 1: The veins of the heart leaves are clear or there is slight mosaic or slight yellowing, and the plant is not obviously stunted;

[0057] Grade 2: 1 / 3 to 1 / 2 of the leaves turn yellow with mosaic or mottled patterns, or a few leaves become deformed; or the main vein turns black, and the plant is stunted to 2 / 3 to 3 / 4 of its normal height;

[0058] Grade 3: 1 / 2 to 2 / 3 of the leaves are yellowed or mottled, or deformed, or the main and lateral veins turn black or die, and the plant is dwarfed to 1 / 2 to 2 / 3 of the normal plant height;

[0059] Level 4: The leaves of the entire plant turn yellow with mosaic or mottled patterns, are severely deformed or necrotic, and the diseased plant is stunted to less than 1 / 2 the height of a normal plant.

[0060] The severity index is calculated based on the severity of the disease, and the effectiveness of prevention and control measures is used to measure the effectiveness of different treatments.

[0061] Disease index = ∑[(Number of infected plants × Representative value of severity level) / (Total number of plants surveyed × Representative value of the highest severity level)] × 100

[0062] Efficacy (%) = [(Mean disease index of blank control - Mean disease index of all treatment groups) / Mean disease index of blank control] × 100%

[0063] The results are shown in Table 3.

[0064] Table 3. Field plot efficacy test of *Leiwan* polysaccharide antiviral agent for the control of plant viral diseases.

[0065]

[0066]

[0067]

[0068] Note: - indicates inactive.

[0069] As shown in the table above, the antiviral agent Leiwan polysaccharide applied to farmland can effectively prevent and control plant viral diseases on various crops, with good control effect, significantly better than the positive control lentinan and Ningnanmycin.

[0070] Example 6: Activity-tracking isolation of active components from *Lycopodium clavatum* polysaccharide extract

[0071] In this embodiment, tobacco was used as the test plant, and tobacco mosaic virus (TMV, MN186255.1) was used as the test pathogen. Protective activity was used as the activity evaluation method (the evaluation method is the same as in Example 1). The activity tracking and separation method was used to identify the polysaccharide of *Lycopodium clavatum* as the effective antiviral component in the extract of *Lycopodium clavatum*. The specific steps are as follows:

[0072] 200g of dried sclerotia of *Lycopodium clavatum* were pulverized using a pulverizer and divided into four equal portions of 50g each. Each portion was then extracted with 300mL petroleum ether, 300mL acetone, 300mL methanol, 300mL water, and 300mL 0.1mol / L NaOH aqueous solution under reflux for 2 hours, repeated six times. The filtrates were combined and concentrated to obtain petroleum ether extract (F petroleum ether), acetone extract (F acetone), methanol extract (F methanol), water extract (F water), and alkaline water extract (F alkaline water). As shown in Table 4, the 0.1mol / L NaOH aqueous solution extraction showed the highest extraction rate and antiviral activity. The water extract exhibited some antiviral activity, while the low-polarity petroleum ether and acetone extractions showed very low extraction rates and no antiviral activity. This indicates that the antiviral active component of *Lycopodium clavatum* is a highly polar substance, and its solubility in alkaline water is the highest.

[0073] The product was then dissolved in alkaline water after hot reflux extraction with 0.1 mol / L NaOH aqueous solution, and then extracted with dichloromethane and n-butanol respectively to prepare dichloromethane phase (F alkaline water-1), n-butanol phase (F alkaline water-2) and aqueous phase (F alkaline water-3). The dichloromethane phase and n-butanol phase were present in low amounts. The dichloromethane phase had no antiviral activity, the n-butanol extract phase had weak antiviral activity, while the aqueous phase had a high content and significant antiviral activity, which is the effective fraction of the extract of *Lycopodium clavatum*.

[0074] Furthermore, the aqueous phase of the product extracted by hot reflux of 0.1 mol / L NaOH aqueous solution was purified in stages using a gradient alcohol precipitation method. The specific operation steps are as follows:

[0075] After adjusting the ethanol content in the aqueous phase supernatant to 20%, incubate overnight at 4 degrees Celsius, centrifuge (1200 rpm) and collect the precipitate (F alkaline water-3-1);

[0076] Adjust the ethanol content in the aqueous phase supernatant to 40% by mass, incubate at 4 degrees Celsius overnight, and centrifuge (1200 rpm) to collect the precipitate (F alkaline water-3-2);

[0077] Adjust the ethanol content in the aqueous phase supernatant to 60% by mass, incubate overnight at 4 degrees Celsius, and centrifuge (1200 rpm) to collect the precipitate (F alkaline water-3-3);

[0078] Adjust the ethanol content in the aqueous phase supernatant to 80% by mass, incubate overnight at 4 degrees Celsius, and centrifuge (1200 rpm) to collect the precipitate (F alkaline water-3-4);

[0079] The remaining supernatant of the treatment group with an ethanol mass percentage of 80% was concentrated to obtain F alkaline water-3-5.

[0080] Among the extracts, F-alkali-3-1 had the highest content and exhibited significantly better antiviral activity than F-alkali and F-alkali-3 at the same concentration (0.5 mg / mL). F-alkali-3-2 showed some antiviral activity, while F-alkali-3-3, F-alkali-3-4, and F-alkali-3-5 had low content and no antiviral activity. This indicates that the antiviral active ingredient of *Lycopodium clavatum* mainly exists in F-alkali-3-1, which is the 20% alcohol precipitation fraction of the aqueous phase in the hot reflux extraction product of 0.1 mol / L NaOH aqueous solution. After removing proteins using the TCA method, *Lycopodium clavatum* polysaccharide was prepared using the same method described above.

[0081] Table 4. Yields of different fractions of *Lycopodium clavatum* polysaccharide isolated from activity tracking and its anti-tobacco mosaic virus activity.

[0082]

[0083] Note: - indicates no activity; the concentration for antiviral activity determination of each fraction was 0.5 mg / mL.

[0084] Although the present invention has been described in detail above with general descriptions and specific embodiments, modifications or improvements can be made to it, which will be obvious to those skilled in the art. Therefore, all such modifications or improvements made without departing from the spirit of the present invention fall within the scope of protection claimed by the present invention.

Claims

1. Use of a polysaccharide of Ophiocordyceps sinensis for preparing an anti-plant virus agent, wherein the polysaccharide has a structure as shown in Formula I: ###0001### Formula I 10≤n≤700; and the plant virus is rice black-streaked dwarf virus, wheat yellow mosaic virus, soybean mosaic virus, cucumber mosaic virus, potato virus Y or pepper mottle virus. Ⅰ wherein O. lapidescens 2. Use according to claim 1, characterized in that, The said polysaccharide of the Leimaru is isolated from the Leimaru fungus of the genus Inonotus The isolation method comprises the following steps: crushing sclerotia of the fungus O. sinensis, extracting the crushed sclerotia with alkaline water, adding ethanol to the supernatant to precipitate, collecting the precipitate, and removing protein to obtain a product. ).

3. Use according to claim 2, characterized in that, The anti-plant virus agent comprises the polysaccharide of O. sinensis and an adjuvant.

4. Use according to claim 1, characterized in that, The content of the polysaccharide of O. sinensis is 1% to 80% by mass percentage.

5. Use according to claim 4, characterized in that, The anti-plant virus agent is an aqueous agent, a soluble liquid agent, a microemulsion, a soluble powder, an aqueous emulsion, a suspension agent or a water-dispersible granule.

6. Use according to claim 4, characterized in that, ​

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