阅读说明：本技术 雷公藤三萜酸在制备防治白斑综合征病毒药物的应用 (Application of tripterygium triterpenic acid in preparation of medicine for preventing and treating white spot syndrome virus ) 是由 王永杰 李凯 夏文旭 江录志 刘昱辰 覃冰 于 2019-09-19 设计创作，主要内容包括：本发明属于水产养殖的疾病防治领域,特别涉及雷公藤三萜酸在制备防治白斑综合征病毒药物的应用。尤其是在制备防治虾类白斑综合征病毒药物的应用中。雷公藤三萜酸经口投喂或经口注射的方式应用。本发明通过实验证明了雷公藤三萜酸对克氏原螯虾和南美白对虾的保护率均可以达到90％以上。雷公藤三萜酸可用于阻断WSSV通过经口方式感染宿主,作为生产防治虾白斑综合征的药物,可有效预防WSSV的感染和传播,成为虾养殖业中阻断WSSV感染的特效药物。(The invention belongs to the field of disease prevention and treatment of aquaculture, and particularly relates to application of tripterygium triterpenic acid in preparation of a medicine for preventing and treating white spot syndrome virus. In particular to the application of preparing the medicine for preventing and treating the shrimp white spot syndrome virus. The tripterygium triterpenic acid is applied by oral feeding or oral injection. The invention proves that the protection rate of the tripterygium triterpenic acid on procambarus clarkia and penaeus vannamei can reach more than 90 percent through experiments. Tripterygium wilfordii triterpenic acid can be used for blocking WSSV from infecting a host in an oral way, can be used for producing a medicament for preventing and treating shrimp white spot syndrome, can effectively prevent WSSV infection and transmission, and becomes a specific medicament for blocking WSSV infection in shrimp farming industry.)

1. Application of Tripterygium wilfordii triterpenic acid in preparing medicine for preventing and treating white spot syndrome virus is provided.

2. The use of claim 1, wherein the tripterygium triterpenic acid is used for preparing a medicament for preventing and treating shrimp white spot syndrome virus.

3. The use according to claim 2, wherein the shrimp is procambarus clarkii or penaeus vannamei.

4. The use of claim 1, 2 or 3, wherein the tripterygium triterpenic acid is administered by oral feeding or oral injection.

5. The use of claim 4, wherein the tripterygium triterpenic acid is orally administered by dissolving in 50% ethanol to obtain a solution of tripterygium triterpenic acid, wherein the effective concentration of tripterygium triterpenic acid in the solution is 0.5-150 μ g/mL, and the injection dosage is 0.002-0.6 μ g/g.

6. The use of claim 4, wherein the tripterygium triterpenic acid is orally injected into the solution for preventing and treating white spot syndrome of procambarus clarkia, and the effective concentration of the tripterygium triterpenic acid in the solution is 0.5-1.5 μ g/mL, and the injection dosage is 0.002-0.006 μ g/g.

7. The use of claim 4, wherein the tripterygium triterpenic acid is orally injected into the solution for preventing and treating white spot syndrome virus of penaeus vannamei, the effective concentration of the tripterygium triterpenic acid solution is 50-150 μ g/mL, and the injection dosage is 0.2-0.6 μ g/g.

8. The use of claim 4, wherein the amount of tripterygium triterpenic acid administered orally is 0.7-0.9 mg/prawn.

Technical Field

The invention belongs to the field of disease prevention and treatment of aquaculture, and particularly relates to application of tripterygium triterpenic acid in preparation of a medicine for preventing and treating white spot syndrome virus.

Background

White Spot Disease (WSSV), which is an acute Disease with high mortality and occurs in the early 90 th century, is caused by the White Spot Syndrome Virus (WSSV). Once infected, 100% of the shrimps die within 3-7 days. WSSV is a large double-stranded circular DNA virus in the form of a rod having a genome size of about 300kbp and is classified as a genus Whispovirus. WSSV has a wide range of crustacean hosts including cultured or wild shrimp, crab, lobster, freshwater cultured Macrobrachium rosenbergii, and the like. Although the susceptibility varies between hosts, WSSV is fatal to all shrimp species. After the prawn is infected with WSSV, the prawn shows the clinical symptoms of lethargy, reduced food intake, reduced day and night activity, loose skin and liver and pancreas discoloration, and is most obviously characterized in that white spots of 0.5-3.0mm are formed on the shell of the cephalothorax of the prawn.

For shrimp, one of the hosts of WSSV, it is highly beneficial for viral transmission due to the congeneric nature of feeding shrimp. The shrimp exposed in the water body containing the WSSV particles or orally ingest food infected by WSSV, which is very likely to cause WSSV infection. In addition, WSSV by physical contact is also sufficient to cause outbreaks of leukoderma in shrimp ponds. At present, WSSV is a great threat to the shrimp farming industry all over the world.

Morphologically, the WSSV particle has an oval or elliptical shape, 80-120X 250-380 nm, and has a tail-like structure at one end, and the function of the tail-like structure is unknown. The virus structure is composed of outermost envelope protein, bridging protein and nucleocapsid protein, and all the structural units are structural proteins.

The envelope protein is positioned at the outermost layer of the WSSV virus and plays an important role in the process of recognizing the virus and host cells. The function research of WSSV envelope protein has been advanced, and it is clear that WSSV envelope protein can participate in the interaction of host protein.

Since various envelope proteins of WSSV have the function of interacting with related proteins of a host, the following methods are commonly adopted in the prevention and treatment of WSSV: the WSSV envelope protein is subjected to recombinant protein vaccine protection, RNA interference gene silencing of corresponding receptor protein in shrimp bodies, preparation of WSSV envelope protein antibodies for neutralization experiments in shrimp bodies, and finding of drugs and compounds for neutralization of WSSV envelope protein. Specifically, the recombinant protein vaccine protection of WSSV envelope protein mainly comprises the steps of expressing and purifying a researched target protein through a eukaryotic or prokaryotic expression system to obtain a recombinant protein, and then adding the recombinant protein into feed for feeding experimental shrimps according to a certain proportion. After the WSSV virus solution is orally fed or injected, the death and virus infection conditions of the shrimps in the experimental group are observed after a period of time to determine the protection effect and the protection rate of the recombinant protein. The prevention methods in the prior art have little practical application value because they have the limitations of short timeliness, expensive production cost and difficult use in the culture environment.

Tripterygium triterpenic acid (CAS: 86632-20-4) belongs to pentacyclic triterpenoid compounds, and since the discovery in 1989, the tripterygium triterpenic acid is successively proved to have a plurality of isomers, and triterpenoids have wide anti-inflammatory action, immunoregulation and other functions, but the specific physiological research on the tripterygium triterpenic acid is still unclear at present.

In general, the specific physiological activity function and application range of the tripterygium triterpenic acid are not clear, and no report about the preparation of the medicine for preventing and treating the shrimp white spot syndrome by using the tripterygium triterpenic acid substance, especially the oral medicine, is found.

Disclosure of Invention

The invention aims to provide application of tripterygium triterpenic acid in preparation of a medicine for preventing and treating white spot syndrome virus, and experiments prove that the protection rate of the medicine of the tripterygium triterpenic acid prawn is 90-100%.

The purpose of the invention is realized by the following technical scheme:

tripterygium triterpenic acid (CAS: 86632-20-4, chemical formula C)₃₀H₄₈O₅) Molecular weight 488 g/mol. The molecular structural formula is as follows:

application of Tripterygium wilfordii triterpenic acid in preparing medicine for preventing and treating white spot syndrome virus is provided. Preferably, the tripterygium triterpenic acid is applied to the preparation of the medicine for preventing and treating the shrimp white spot syndrome virus. More preferably, the shrimp is procambarus clarkii or penaeus vannamei.

The tripterygium triterpenic acid is applied in a mode of oral feeding or oral injection.

When the tripterygium triterpenic acid is applied in an oral injection mode, ethanol with the concentration of 50% is dissolved into a tripterygium triterpenic acid solution, the effective concentration of the tripterygium triterpenic acid in the solution is 0.5-150 mu g/mL, and the injection dosage is 0.002-0.6 mu g/g.

Preferably, when the tripterygium triterpenic acid is applied to preventing and treating white spot syndrome virus of procambarus clarkia in an oral injection mode, 50% ethanol is used for dissolving the tripterygium triterpenic acid into a tripterygium triterpenic acid solution, the effective concentration of the tripterygium triterpenic acid in the solution is 0.5-1.5 mu g/mL, and the injection dosage is 0.002-0.006 mu g/g. Further preferably, the effective concentration of the tripterygium triterpenic acid solution is 1.0 mu g/mL, and the injection dosage is 0.004 mu g/g.

When the tripterygium triterpenic acid is applied to the prevention and treatment of the white spot syndrome virus of procambarus clarkii in an oral feeding mode, the oral feeding amount is 0.7-0.9 mg/million shrimps. Further preferably, when the tripterygium triterpenic acid is applied in an oral feeding mode, the oral feeding amount is 0.8 mg/kaleidoscope.

Preferably, when the tripterygium triterpenic acid is applied to preventing and treating white spot syndrome virus of penaeus vannamei through an oral injection mode, 50% ethanol is dissolved into a tripterygium triterpenic acid solution, the effective concentration of the tripterygium triterpenic acid in the solution is 50-150 mu g/mL, and the injection dosage is 0.2-0.6 mu g/g. Further preferably, the effective concentration of the tripterygium triterpenic acid solution is 100 mu g/mL, and the injection dosage is 0.4 mu g/g.

When the tripterygium triterpenic acid is applied in an oral feeding mode and is applied to the prevention and treatment of white spot syndrome virus of penaeus vannamei boone, the oral feeding amount is 70-90 mg/million shrimps. Further preferably, when the tripterygium triterpenic acid is applied in an oral feeding mode, the oral feeding amount is 80 mg/ten thousand shrimps.

The beneficial effects are that:

1. the tripterygium triterpenic acid provided by the invention can be used for blocking WSSV from infecting a host in an oral way, can be used for producing a medicament for preventing and treating shrimp white spot syndrome, can effectively prevent the infection and spread of WSSV, and becomes a specific medicament for blocking WSSV infection in the shrimp farming industry.

2. The experiments of the invention prove that the protection rate of the tripterygium triterpenic acid on procambarus clarkia (crayfish) and penaeus vannamei can reach more than 90 percent, and the tripterygium triterpenic acid can effectively block WSSV from infecting shrimps by mouth. Especially when procambarus clarkia is used as a living body, the injection amount of a drug experimental group with the effective concentration of the tripterygium triterpenic acid of 1.0 mug/mL is 0.004 mug/g, and the protection rate reaches 100 percent; when the penaeus vannamei boone is taken as a living body, the injection amount of a drug experiment group with the effective concentration of the triterpenic acid of the tripterygium wilfordii of 100 mu g/mL is 0.4 mu g/g, and the protection rate also reaches 100 percent.

Drawings

FIG. 1 is a comparison graph of the results of the in vivo experiments of the pharmaceutical tripterygium wilfordii triterpenic acid in procambarus clarkia.

FIG. 2 is a comparison graph of the results of in vivo experiments on Penaeus vannamei Boone with the medicine Tripterygium triterpenic acid.

Detailed Description

The invention will now be further illustrated by reference to the following examples:

tripterygium triterpenic acid (CAS: 86632-20-4, chemical formula C)₃₀H₄₈O₅). Molecular weight 488g/mol, purchased from Carbosynth China Ltd, Calbonson chemical technology, Suzhou;

the WSSV quantitative standard plasmid which is constructed in the laboratory and takes VP28 as a target gene. Marine animal tissue genome DNA extraction kit (TIANGEN), 2 XchamQTM UniversalqPCR Master Mix(Vazyme)，ddH₂O, WSSV specific primer VP28 (Forward sequence: AGGTGTGTG) for fluorescent quantitative detectionGAACAACACATCAAG, reverse sequence: TGCCAACTTCATCCTCATCA) was produced by Sangon.

Example 1

In vivo protocols are shown in the table below.

(1) WSSV-free Procambrus clarkii: 120 WSSV-free Procambrus clarkii (20-30 g each) were selected for challenge experiments. Ensuring the consistency of the body type, the health state and the vitality of each procambarus clarkii.

WSSV-TW suspension, copy number 8X 10⁷copies/. mu.L, purified by the laboratory.

(2) And the experimental process is as follows:

a. healthy active WSSV-free Procambrus clarkii were randomly distributed in 10 shrimp pots per group and the groups were labeled.

b. Diluting 400 μ L (concentration of 1mg/mL) of mother liquor of medicinal tripterygium triterpenic acid to obtain 4mL of medicinal liquid with concentration of 100 μ g/mL, and diluting according to gradient of 10 times to obtain medicinal liquid with four concentration gradients of medicinal tripterygium triterpenic acid. The four concentrations were 0.1. mu.g/mL, 1. mu.g/mL, 10. mu.g/mL, 100. mu.g/mL, respectively.

c. WSSV-TW virus suspension (copy number 8X 10)⁷copies/. mu.L) from-80 deg.C, thawing on ice for 2-3h, and diluting with TM buffer to obtain 8 × 10⁶copies/. mu.L WSSV-TW viral suspension.

d. The names of the experimental groups are respectively marked in 5 sterile centrifuge tubes of 15 mL. Placing on ice, and adding 1.2mL of 8X 10 solution into the test tubes of the four labeled drug groups A, B, C and D⁶copies/. mu.L WSSV-TW viral suspension. Subsequently, 1.2mL of 0.1. mu.g/mL liquid medicine is added into the medicine group A, 1.2mL of 1. mu.g/mL liquid medicine is added into the medicine group B, 1.2mL of 10ug/mL liquid medicine is added into the medicine group C, and 1.2mL of 100ug/mL liquid medicine is added into the medicine group D, and the mixture is respectively and evenly shaken. Finally, a WSSV-TW suspension of the positive control group and a TM buffer of the negative control group were prepared.

e. Placing the test tubes together in a constant temperature shaking table INNOVA 40R. The incubation conditions were: 25 ℃, 85rpm, 1.5 h.

f. And taking out the incubated test tubes of the drug group A, the drug group B, the drug group C, the drug group D, the drug control group, the negative control group and the positive control group from the constant-temperature shaking table, and placing at room temperature for later use. 200 μ L of the above liquid was separately aspirated, and each procambarus clarkii in each group was injected by oral injection. Note that the groups for oral injection were in order negative control, drug group a, drug group B, drug group C, drug group D, drug control (100ug/mL drug) and positive control, and the glove was replaced with a new one every time the group was completed.

g. After the injection was completed, the death of procambarus clarkii was checked every noon and recorded. Feeding the freeze-dried red worms at intervals of 2-3 days. The air conditioner was set to 26 ℃. The dead procambarus clarkii groups were collected, the groups and the death date were marked, and the dead shrimp samples were stored at-20 ℃.

(3) And detecting

Carrying out WSSV background detection on the procambarus clarkii one by one: after the experimental period lasted three weeks, all groups of procambarus clarkii were trimmed of 30mg of abdominal muscle with sterile scissors and placed in 1.5mL sterile centrifuge tubes. And extracting the total genomic DNA of the abdominal muscle tissue of the procambarus clarkia according to a marine animal tissue genomic DNA extraction kit. And (3) measuring the concentration of the DNA by using a multifunctional enzyme-labeling instrument, ensuring the concentration range to be 50-100 ng/mu L, subpackaging the extracted DNA and storing the DNA in a refrigerator at the temperature of-20 ℃. Finally, these total DNAs were used as templates to detect the WSSV content in each sample by qPCR (real-time fluorescent quantitative PCR technique).

Single 20 μ L qPCR reaction: 10 μ L of qPCR Master Mix, 0.6 μ L of forward and reverse primers, 6.8 μ L of ddH₂O, 2. mu.L of DNA template. Reaction program setup for fluorescent quantitative PCR instrument ABI7500 FAST: holding stage (95 ℃ 30s), Cycling stage (40 cycles, 95 ℃ 15s, 60 ℃ 30 s). Melt cut Stage (continues, default parameters).

(4) Conclusion

Through 22 days of experimental observation, after the experiment is finished, the survival conditions of procambarus clarkii (crayfish) in each group are counted, graph plotting is carried out by utilizing GraphPad Prism 8 software, and the in-vivo experimental result of the tripterygium triterpenic acid medicine is obtained, as shown in figure 1, the protection rate of the tripterygium triterpenic acid medicine is 90% -100%, and the WSSV infection of the crayfish can be effectively blocked.

To this end, we performed qPCR assays for WSSV on all crayfishes to further demonstrate whether the drug tripterygium triterpenic acid blocks viral infection of shrimp.

And (3) detecting the WSSV content in the crayfish body by qPCR: the crayfish of the negative control group TM buffer did not die in the whole experimental period, indicating that the experimental skill of oral injection has no misoperation, and all crayfish surviving in the negative control group are detected to have no WSSV through the subsequent q-PCR detection analysis. The crayfish in the drug tripterygium triterpenic acid (CAS: 86632-20-4) control group (100 mug/mL drug) survives completely, so the maximum dose of the drug can not kill the crayfish, and the 100 mug/mL tripterygium triterpenic acid (CAS: 86632-20-4) has no toxicity to the crayfish. The WSSV-TW positive control group died gradually from the third day through the entire experimental cycle until a cumulative death of 80% was reached at the end of the experimental cycle, and 2 crayfish remained alive at day 22. Q-PCR detection of 8 dead crayfishes in the positive control group shows that the WSSV order of magnitude is 10⁷-10¹⁰copies/. mu.L, 2 alive, 1 WSSV of the order of 10⁶copies/. mu.L, 1 is only 10⁹copies/. mu.L. It can be seen that the strain WSSV-TW is active and is capable of infecting crayfish.

For the four gradient groups of the drug experimental group: drug group A (drug 0.1. mu.g/mL + WSSV-TW) died 1 crayfish only on day 14, and the order of WSSV was found to be 7.5X 10 in the 1 crayfish died in group A by q-PCR detection analysis⁹copies/. mu.L, live crayfish did not contain WSSV. Drug group B (drug 1. mu.g/mL + WSSV-TW) died 1 crayfish on day 6, and no WSSV (not dead due to viral infection) was detected in the 1 died crayfish in group B as analyzed by q-PCR assay, and 9 live crayfish in group B did not contain WSSV. No dead shrimp were present in the drug group C (drug 10. mu.g/mL + WSSV-TW) during the entire experiment, and all crayfish did not contain WSS by q-PCR detection analysisAnd V. Drug group D (drug 100. mu.g/mL + WSSV-TW) died 1 crayfish on day 17, and the order of magnitude of WSSV was found to be 5.14X 10 in the died 1 crayfish by q-PCR detection analysis⁷copies/. mu.L, no WSSV was detected in 9 live crayfish in the same group D. Through in-vivo experiments and q-PCR detection results, after the medicine tripterygium triterpenic acid and WSSV virus particles are incubated together, the virus basically loses the capability of invading a host. Therefore, the tripterygium triterpenic acid has a good protection effect on procambarus clarkii (crayfish), and the protection rate is as high as 90-100%.

In conclusion, the medicine in vivo experiment shows that for four gradient medicine groups, the protection rate of the medicine tripterygium wilfordii triterpenic acid on procambarus clarkia (crayfish) reaches 90-100%, and WSSV oral infection of the crayfish can be effectively blocked. Especially, the protection rate of the medicine groups B and C reaches 100 percent.

Example 2

Because procambarus clarkii has certain resistance to WSSV and the penaeus vannamei has almost no resistance to WSSV, in order to further verify that the tripterygium wilfordii triterpenic acid (CAS: 86632-20-4) can still play a similar protective role in shrimps of different species, on the basis of the example 1, further verification and optimization of the test are carried out, the penaeus vannamei is selected as a test shrimp species, and oral toxicity counteracting test verification is carried out by drug-embedded WSSV.

In vivo protocols are shown in the table below.

(1) WSSV-free Penaeus vannamei: 50 WSSV-free Penaeus vannamei Boone (weight: 10.5-12g, length: 10.5-13.5cm) were selected for challenge experiments. Ensuring the consistency of the body type, the health state and the vitality of each penaeus vannamei.

WSSV-TW suspension, copy number 8X 10⁷copies/. mu.L, purified by the laboratory.

(2) And the experimental process is as follows:

a. healthy active WSSV-free Penaeus vannamei Boone was randomly distributed in 10 per group into each shrimp jar and the groups were labeled.

b. The two medicinal liquids of the tripterygium triterpenic acid in the embodiment 1 are selected, and the concentrations are respectively 10 mug/mL and 100 mug/mL.

c. Preparation according to example 1 gives 8X 10⁶copies/. mu.L WSSV-TW viral suspension.

d. The experimental group names were marked in 2 sterile 15mL centrifuge tubes, respectively. Placed on ice, to the two labeled test tubes of drug group A and drug group B, 2.4mL of 8X 10 was added to drug group A⁶After copies/mu L WSSV-TW virus suspension, 2.4mL of 100 mu g/mL liquid medicine is added (taking the drug group A as a preferred group, and a group of parallel test groups are added); drug group B was added 1.2mL of 8X 10⁶After copies/. mu.L of WSSV-TW virus suspension, 1.2mL of 10. mu.g/mL liquid medicine was added, and the mixture was gently shaken and mixed. Finally, a WSSV-TW suspension of the positive control group and a TM buffer of the negative control group were prepared.

e. Placing the test tubes together in a constant temperature shaking table INNOVA 40R. The incubation conditions were: 25 ℃, 85rpm, 1.5 h.

f. And taking the incubated test tubes of the drug group A, the drug group B, the negative control group and the positive control group out of the constant-temperature shaking table, and placing at room temperature for later use. Each penaeus vannamei in each group was injected by oral injection by pipetting 200 μ L of each of the above liquids. Note that the groups injected orally were in order negative control group, drug group a, drug group B, and positive control group, and the glove was replaced with a new one every time the group was completed.

g. After the injection is finished, whether the penaeus vannamei died or not is checked every noon, and a record is made. The feed is fed once in the morning and at night every day. The water temperature was controlled at 22 ℃. The dead penaeus vannamei boone groups were collected, the groups and the death date were marked, and the dead penaeus vannamei boone samples were stored at-20 ℃.

(3) And detecting

Carrying out WSSV background detection on the penaeus vannamei one by one: detection method and procedure the detection method in example 1 was followed.

(4) Conclusion

Through 14 days of experimental observation, after the experiment is finished, the survival conditions of the penaeus vannamei boone in each group are counted, graph Pad Prism 8 software is used for drawing, and the in-vivo experimental result of the tripterygium triterpenic acid is obtained, as shown in figure 2, the protection rate of the tripterygium triterpenic acid is 80% -100%, and the WSSV infection of the penaeus vannamei boone can be effectively blocked.

Therefore, we carried out WSSV qPCR detection on all the white shrimps to further prove whether the medicine Tripterygium wilfordii triterpenic acid can block virus infection of the white shrimps.

And (3) detecting the WSSV content in the white shrimp body by qPCR: the white shrimps of the TM buffer in the negative control group have no death in the whole experimental period, which indicates that the experimental skill of oral injection has no misoperation, and all the surviving white shrimps in the negative control group have no WSSV detected through the subsequent q-PCR detection analysis. The positive control group, WSSV-TW, died gradually from the third day through the entire experimental cycle until all died on the sixth day, accumulating to 100%. All the dead white shrimps in the positive control group are detected by q-PCR to find that the WSSV copy number is 3.46 multiplied by 10⁷-9.49×10⁷copies/. mu.L. As can be seen, the strain WSSV-TW is active and can infect white shrimp.

For both gradient sets of the drug experimental set: only one of the two drug groups A (drug 100. mu.g/mL + WSSV-TW) died 1 prawn at day 5, and the WSSV copy number was found to be 6.51X 10 in the 1 prawn that died by q-PCR detection and analysis³copies/. mu.L, none of the remaining live white shrimps contained WSSV.

The drug group B (drug 10. mu.g/mL + WSSV-TW) killed 1 prawn on day 4 and 5 respectively, and the copy number of WSSV in the 2 killed prawns in the group B was found to be 1.66X 10 respectively by q-PCR detection analysis⁴And 8.52X 10⁵And the rest of the live white shrimps do not contain WSSV. Through in-vivo experiments and q-PCR detection results, after the medicine tripterygium triterpenic acid and WSSV virus particles are incubated together, the virus basically loses the capability of invading a host. Therefore, the medicine tripterygium triterpenic acid has a good protection effect on the penaeus vannamei boone, and the protection rate is as high as 80-100%.

In conclusion, the medicine in vivo experiments show that for the two gradient medicine groups, the protection rate of the medicine tripterygium triterpenic acid on the penaeus vannamei boone reaches 80-100%, and the WSSV oral infection of the penaeus vannamei boone can be effectively blocked. Especially, the protection rate of the medicine group A can reach 90 to 100 percent.

In conclusion, according to the actual protection effect of a medicine in-vivo experiment, the medicine tripterygium triterpenic acid is proved to be capable of being combined with the cyst membrane protein on the surface of the WSSV after being incubated with the WSSV, so that the active site of the cyst membrane protein is closed, and the WSSV is effectively blocked from invading a shrimp host through the mouth, and on the basis, the tripterygium triterpenic acid can be used in an actual culture water body, so that the tripterygium triterpenic acid has an actual application value.

In addition, due to the following application of the medicine and the economic aspect, the preferable mass concentration is 50% ethanol as a solvent, the tripterygium triterpenic acid liquid medicine is sprinkled in the shrimp culture pond, when the effective concentration of the tripterygium triterpenic acid is 1 mu g/mL, the oral feeding amount is 0.8 mg/ten thousand of shrimps, and the protection rate of the tripterygium triterpenic acid on procambarus clarkii (crayfish) reaches 90%;

when the effective concentration of the tripterygium triterpenic acid is 100 mug/mL, the amount of the tripterygium triterpenic acid is 80 mg/million tail shrimps by oral feeding, the protection rate of the tripterygium triterpenic acid on the penaeus vannamei boone reaches 90%, and the WSSV can be effectively blocked from invading the shrimp host.

The above description is only for the purpose of illustrating the preferred embodiments of the present invention, and the present invention should not be limited by the disclosure of the preferred embodiments. Therefore, it is intended that all equivalents and modifications which do not depart from the spirit of the invention disclosed herein are deemed to be within the scope of the invention.