Application of wogonin in resisting siniperca chuatsi and frog iridovirus

Abstract

The invention belongs to the technical field of biology, and discloses application of wogonin in resisting siniperca chuatsi and frog iridovirus, the wogonin can effectively inhibit over-generation of ROS caused by MRV virus infection, and oxidative stress damage of host cells is relieved in a dose-dependent mode. The wogonin can be used for preventing and treating the siniperca chuatsi and frog iridovirus, the high expression state of a proinflammatory gene can be reduced, intestinal tissue damage induced by MRV infection can be relieved, proliferation of MRV in the micropterus salmoides is effectively inhibited, the MRV challenge survival rate of the micropterus salmoides is increased, a basis is provided for preparing a product for resisting the siniperca chuatsi and frog iridovirus from wogonin, and the wogonin has important application value. Meanwhile, the invention also discloses a medicine, a fish feed and a fish feed additive of the wogonin.

Description

Technical Field

[0001] This invention relates to the field of biomedical technology, specifically the use of baicalin in the treatment of mandarin frog iridovirus. Background Technology

[0002] Mandarin fish ranavirus (MRV) belongs to the family Iridoviridae and the genus Ranavirus. MRV can infect mandarin fish (Siniperca chuatsi) and largemouth bass (Micropterus salmoides). After infection, the virus causes systemic inflammatory damage, which in turn leads to dysfunction in multiple aspects of the body, including metabolism and immunity, and has an extremely high mortality rate.

[0003] Baicalein (7-dihydroxy-8-methoxyflavonoid) is a natural flavonoid compound isolated from the root of the traditional Chinese medicine Scutellaria baicalensis. Existing research shows that baicalein possesses diverse pharmacological activities, including anti-inflammatory, antiviral, anticancer, antioxidant, and immunomodulatory effects. It exhibits broad antiviral activity against various influenza viruses, rotaviruses, Coxsackieviruses, coronaviruses, and herpes zoster viruses. For example, prior art CN116019805A discloses the application of baicalein in the treatment of porcine epidemic diarrhea virus (PEDV). By targeting the PEDV-3c-like protease (PEDV-3CLpro), baicalein can significantly inhibit the internalization, intracellular replication, and release of new mature viral particles of PEDV. It can also directly inactivate PEDV in vitro. Prior art CN108836962B discloses the application of baicalein in the preparation of anti-rotavirus drugs.

[0004] Porcine epidemic diarrhea virus (PEDV) and rotavirus infect mammals. Rotavirus infects intestinal epithelial cells, causing cell vacuolization, apoptosis, and shedding, thereby disrupting the intestinal mucosal barrier function. PEDV causes intestinal villi atrophy and fusion, reducing the absorptive surface area and inducing osmotic diarrhea. It also inhibits the function of sodium-glucose cotransporter 1 (SGLT1), exacerbating electrolyte imbalance. Furthermore, PEDV is a single-stranded RNA virus, while rotavirus is a double-stranded RNA virus. Current research only supports the inhibitory effect of baicalin on these two viruses. Summary of the Invention

[0005] The purpose of this invention is to provide the use of baicalin in the preparation of products for the prevention and / or treatment of fish infected with mandarin frog iridovirus.

[0006] Meanwhile, the present invention also provides drugs, fish feeds and fish feed additives containing baicalin.

[0007] To achieve the above objectives, the present invention provides the following technical solution:

[0008] Use of baicalin in the preparation of products for the prevention and / or treatment of fish infected with mandarin frog iridovirus.

[0009] The genome of the mandarin frog iridovirus is double-stranded DNA. The mandarin frog iridovirus infects fish such as mandarin fish and largemouth bass, and causes systemic inflammation, multi-organ inflammatory damage, ascites, and body surface ulceration in fish.

[0010] Based on existing research on the antiviral mechanism of baicalin, it can be found that the mandarin frog iridovirus and the related viruses that are effective with traditional baicalin have different viral genome types, significantly different species that they act on, and completely different viral sites and symptoms.

[0011] This invention has shown that when baicalin is used in cell and live fish research subjects, it can effectively inhibit the excessive ROS production caused by MRV virus infection and reduce oxidative stress damage in host cells in a dose-dependent manner. Furthermore, it can reduce the high expression of pro-inflammatory genes, increase the expression of anti-inflammatory genes, alleviate MRV infection-induced intestinal tissue damage, effectively inhibit the proliferation of MRV in largemouth bass, and improve the survival rate of largemouth bass after MRV challenge.

[0012] This discovery has positive implications for expanding the application of baicalin as a medicine and for the prevention and treatment of mandarin frog iridovirus.

[0013] Preferably, the chemical formula of the baicalein is C 16 H 12 O5 has a molecular weight of 284.26.

[0014] Preferably, the product is a drug whose active ingredient is baicalein.

[0015] More preferably, the dosage form of the drug is an oral dosage form or an injection.

[0016] Preferably, the product is a fish feed or fish feed additive containing baicalin.

[0017] In addition, this invention discloses a drug for treating mandarin frog iridovirus, which contains baicalein.

[0018] In addition, the present invention also discloses a fish feed that resists the iridovirus of mandarin frogs, the fish feed containing baicalin.

[0019] Finally, this invention also discloses a fish feed additive that resists mandarin frog iridovirus, the fish feed additive containing baicalin.

[0020] Compared with the prior art, the beneficial effects of the present invention are:

[0021] This invention discloses the use of baicalin in the treatment of mandarin frog iridovirus. It can effectively inhibit excessive ROS production caused by MRV virus infection and reduce oxidative stress damage to host cells in a dose-dependent manner. Furthermore, it can reduce the high expression of inflammation-related genes, alleviate MRV infection-induced intestinal tissue damage, effectively inhibit MRV proliferation in largemouth bass, and improve the survival rate of largemouth bass after MRV challenge. This provides a foundation for the preparation of products using baicalin to treat mandarin frog iridovirus, fills the gap in the application of baicalin against mandarin frog iridovirus, and has significant economic value. Attached Figure Description

[0022] Figure 1 The figure shows the effect of baicalin on cell morphology.

[0023] Figure 2 Figure showing the effect of different concentrations of baicalin on cell viability;

[0024] Figure 3 CC of baicalein 50 Value analysis chart;

[0025] Figure 4 Figure showing the results of different administration routes of baicalein in alleviating MRV-induced cytopathic effects;

[0026] Figure 5 Figure showing the effect of different concentrations of baicalin on CPE in MRV-infected cells;

[0027] Figure 6 The figure shows the results of the inhibitory effect of baicalin on MRV replication.

[0028] Figure 7 EC of baicalein 50 Analysis chart;

[0029] Figure 8 A graph showing the survival rate of largemouth bass challenged with MRV;

[0030] Figure 9 The figure shows the results of the inhibitory effect of baicalin on viral load in tissues of surviving fish infected with MRV.

[0031] Figure 10 This is a diagram showing the results of single-cell isolation from the intestine.

[0032] Figure 11 Figure showing the effect of baicalin on ROS levels in MRV-infected intestinal cells;

[0033] Figure 12 The figure shows the effect of baicalin on the expression of inflammation-related genes.

[0034] Figure 13Figure showing the results of baicalin alleviating MRV-induced intestinal tissue damage. Detailed Implementation

[0035] The technical solution of the present invention will be clearly and completely described below with reference to the embodiments of the present invention. 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 of ordinary skill in the art without creative effort are within the scope of protection of the present invention.

[0036] Materials and product information:

[0037] The mandarin frog iridovirus (MRV) was isolated and preserved in our laboratory from mandarin frogs infected with MRV;

[0038] The commercial Chinese herbal monomer baicalein was purchased from Chengdu Manster Biotechnology Co., Ltd. (Wogonin, CAS: 632-85-9). It was dissolved in DMSO, stored in an ultra-low temperature freezer, and diluted to a safe concentration with L-15 medium before use.

[0039] Mandarin fish brain cell line (SCB3): Cells were cultured in L-15 medium at a temperature of 25°C.

[0040] Part 1: Cell Experiments

[0041] 1. Determination of safe concentration of baicalin and its concentration-lowering effect (CC) 50 calculate

[0042] Logarithmic growth phase SCB3 cells were seeded at a density of approximately 5,000 cells / well into 96-well cell culture plates, with 100 μL of cell suspension added to each well. The plates were then incubated at 25°C for 24 h until the cells were fully adherent. The culture medium was removed from the 96-well plates. 100 μL of serially diluted baicalein solution (160 μM, 80 μM, 40 μM, 20 μM, 10 μM, 5 μM, and 2.5 μM) in L-15 medium (containing 5% fetal bovine serum) was added to each well of the experimental group, while the control group received 100 μL of L-15 medium (containing 5% FBS) with 1% DMSO. Three replicates were set up for each group. The treated cells were then returned to the 25°C incubator for further culture. After 96 h, cell morphology was observed using an optical microscope. Subsequently, the culture medium was discarded, and the cells were gently washed once with PBS buffer. Add 100 μL of diluted CCK-8 solution (CCK-8 and PBS were diluted at a ratio of 1:10) to each well and incubate at 28°C in the dark for 6 hours. After incubation, the absorbance (OD value) of each well was measured at 450 nm using a microplate reader. Cell viability was calculated using the following formula: Cell viability (%) = (OD experimental group - OD blank group) / (OD control group - OD blank group) × 100%. Finally, the experimental data were processed using GraphPad Prism 9.5 software, and the half-maximal cytotoxic concentration (CMC) of baicalin was calculated. 50 (Value); Test results reference Figure 1 , Figure 2 , Figure 3 .

[0043] like Figure 1 As shown, when the concentration of baicalin was in the range of 2.50–10 μM, the cell morphology was similar to that of the untreated control group, and no significant morphological changes were observed. When the concentration of baicalin exceeded 10 μM, some cells began to show morphological changes, including nuclear pyknosis, cytoplasmic vacuolization, increased intracellular granular material, and decreased cell refractive index. (CCK-8 assay results) Figure 2 This further supports the above observations. Within the concentration range of 2.5–10 μM, there was no significant difference in cell viability between the drug-treated groups and the untreated control group (p>0.05). Statistical analysis of the fitted curves ( Figure 3 The CC of baicalein was calculated. 50 The value is 46.70 μM.

[0044] Figure 1 The figure shows the effect of baicalin on cell morphology.

[0045] Figure 2The figure shows the effect of different concentrations of baicalein on cell viability; the same letter in the figure indicates no significant difference between groups, and different letters indicate significant differences (p<0.05).

[0046] Figure 3 CC of baicalein 50 Value analysis chart.

[0047] 2. Study on the anti-MRV mechanism of baicalin

[0048] Three administration strategies were employed: prophylactic administration, combined administration, and therapeutic administration. The antiviral effects of baicalin under these three administration methods were compared, and its antiviral mechanism was analyzed.

[0049] SCB3 cells in the logarithmic growth phase were added to 6-well plates at a rate of 2 mL per well and then cultured at 25°C until the cells grew into a monolayer.

[0050] Prophylactic administration group: 2 mL of L-15 medium containing 10 μM baicalein was added, and the mixture was incubated at 25°C for 4 h. The solution was then removed, and 100 TCID50 was added. 50 The MRV virus solution was incubated at 25°C for 4 hours. After removing the virus solution, the sample was washed once with PBS, and then 2 mL of L-15 medium containing 5% FBS was added, and the sample was incubated for another 96 hours.

[0051] Mixed administration group: 100 TCID 50 MRV virus solution was mixed with an equal volume of 10 μM baicalein solution, and the solution was added to the cells and cultured at 25°C for 4 h. The mixture was then aspirated, the cells were washed once with PBS, and 2 mL of L-15 medium containing 5% FBS was added, followed by further culture for 96 h.

[0052] Treatment group: 100 TCID was added to the cells. 50 The MRV virus solution was incubated at 25°C for 4 hours. The virus suspension was aspirated, the cells were washed once with PBS, and 2 mL of L-15 medium containing 10 μM baicalein was added. The cells were then cultured for another 96 hours.

[0053] Two control groups were set up: a virus-infected control group (inoculated with virus only, without drugs) and a cell control group (neither inoculated with virus nor drugs). Cellular pathogenesis was observed regularly during culture, and cell morphology changes were recorded using an optical microscope after 96 hours. Cells were subjected to three freeze-thaw cycles, and a suitable amount of the freeze-thawed cell solution was used for nucleic acid extraction. Viral copy number was determined using qPCR. Results are referenced below. Figure 4 .

[0054] Optical microscopy observation showed ( Figure 4-a) Baicalein effectively reduced MRV-induced cytopathic effects through different administration strategies (preventive administration, combined administration, and therapeutic administration), with preventive and therapeutic administration showing particularly significant effects, resulting in more intact cell morphology and milder cytopathic effects. Figure 4 As shown in -b, compared with the virus-infected control group, all three administration methods significantly reduced the intracellular MRV viral copy number, with prophylactic administration showing the best effect, resulting in a significantly lower viral copy number than the mixed administration group and the treatment administration group. These results indicate that baicalin can inhibit MRV replication through prophylactic administration.

[0055] Figure 4 Figure showing the results of different administration routes of baicalein in alleviating MRV-induced cytopathic effects.

[0056] 3. Baicalein's anti-MRV EC 50 calculate

[0057] SCB3 cells in logarithmic growth phase were harvested, adjusted to a suitable cell density, and seeded at 200 μL per well into 48-well plates. The plates were then incubated at 25°C until the cells were fully attached to the bottom. For the experimental group, 200 μL of serially diluted baicalein solution (20 μM, 10 μM, 5 μM, 2.5 μM, and 1.25 μM) in L-15 medium containing 5% FBS was added, and the plates were incubated at 25°C for 4 h. The solution was then removed, and 100 TCID50 was added. 50 MRV virus solution was cultured at 25°C for 4 hours. After removing the virus solution, the cells were washed once with PBS, and then 200 μL of L-15 medium containing 5% FBS was added, followed by culturing for another 96 hours. Two control groups were set up: a virus-infected control group (inoculated with virus only, without drugs) and a cell control group (neither inoculated with virus nor drugs). The culture plates were returned to the incubator and cultured for another 96 hours to observe the development of cellular pathogenesis. Subsequently, the cells were repeatedly frozen and thawed three times to release intracellular viral particles. 400 μL of the frozen and thawed cell solution was used for nucleic acid extraction and virus copy number determination. The inhibition rate was calculated using the formula: Inhibition rate (%) = (1 - Virus copy number in the experimental group / Virus copy number in the virus-infected group) × 100%. The median effective concentration (EC50) of baicalin was calculated using GraphPad Prism 9.5. 50 ); Results Reference Figure 5 , Figure 6 , Figure 7 .

[0058] Optical microscopy revealed that the pathological severity of MRV-infected cells gradually decreased with increasing baicalin concentration. Figure 5qPCR results showed that as the concentration of baicalin gradually increased, the intracellular MRV viral copy number gradually decreased. Figure 6 The effect was dose-dependent. Based on the data on the viral load inhibition rate of different concentrations of baicalin, combined with the analysis of the fitted curve, the EC50 of baicalin was calculated. 50 5.428 μM ( Figure 7 ).

[0059] Figure 5 Figure showing the effect of different concentrations of baicalin on CPE in MRV-infected cells;

[0060] Figure 6 The figure shows the inhibitory effect of baicalin on MRV replication; in the figure, the same letter indicates no significant difference between groups, and different letters indicate significant differences, p<0.05;

[0061] Figure 7 EC of baicalein 50 Analysis diagram.

[0062] Part Two: Protective Experiment and Analysis of Baicalein against Largemouth Bass Challenge

[0063] Healthy largemouth bass were randomly divided into 5 groups, with 30 fish in each group. (1) Blank control group: administered PBS solution containing 1% DMSO by gavage, 4 times at 48-hour intervals. (2) MRV-infected control group: administered PBS solution containing 1% DMSO by gavage, 4 times at 48-hour intervals. (3) Low-dose baicalein group (7 mg / kg): administered 7 mg / kg baicalein solution by gavage, 4 times at 48-hour intervals. (4) Medium-dose baicalein group (21 mg / kg): administered 21 mg / kg baicalein solution by gavage, 4 times at 48-hour intervals. (5) High-dose baicalein group (63 mg / kg): administered 63 mg / kg baicalein solution by gavage, 4 times at 48-hour intervals. On day 8 of the experiment, the MRV-infected control group and the different doses of baicalein groups were intraperitoneally injected with 100 μL of 1×10 7 TCID 50 The supernatant of MRV cell virus solution was injected at a concentration of / mL, and the blank control group was injected with the same volume of L-15 medium as a negative control. The challenge experiment was analyzed as follows:

[0064] (1) Statistical analysis of survival rate after virus challenge

[0065] During the experiment, the survival status of each group of fish was recorded daily until no deaths occurred for 7 consecutive days, at which point the experiment was terminated, and the survival rate was analyzed using Kaplan-Meier analysis.

[0066] (2) After the experiment ended, five fish were randomly selected from the surviving fish in the above five groups of experiments for sampling and the following tests and analyses were performed:

[0067] ① Collect partial tissues from the liver, spleen, head kidney, pyloric sac, and intestine, extract total DNA from the tissues, and detect the viral load in the tissues using qPCR.

[0068] ② Collect partial tissues from the foregut, midgut, and hindgut, prepare single-cell tissue samples, and perform reactive oxygen species (ROS) detection.

[0069] Intestinal tissue was isolated, washed in ice-pretreated PBS buffer, and then cut into small pieces in Hank's balanced salt solution. The chopped intestinal tissue was transferred to centrifuge tubes containing digestion solution and digested in a 28°C shaking incubator (100 rpm). The digestion solution consisted of collagenase II, collagenase IV (both 1 mg / mL), and DNase I (0.1 mg / mL). The digested tissue suspension was centrifuged at 500 g for 8 min at 4°C. The supernatant was discarded, and the pellet was resuspended in L-15 medium containing 5% FBS. The pellet was filtered through a 75 μm cell sieve, and the filtrate was collected. The cell pellet was resuspended in L-15 medium containing 5% FBS, and 20 μL of single-cell suspension was mixed thoroughly with 0.4% trypan blue solution (9:1 v / v) and stained at room temperature for 3 min. Take 10 μL of stained cell sample, add it to a hemocytometer, and observe and count it under a microscope. The viable cell rate is calculated using the formula: (Total cell count - Blue cell count) / Total cell count × 100%. Centrifuge the single-cell suspension of intestinal tissue, add 1:1000 diluted DCFH-DA working solution, and incubate at 37℃ in the dark for 20–30 min. After incubation, centrifuge (500g, 8 min, 4℃) and discard the supernatant. Wash with pre-chilled L-15 solution and resuspend the cells for flow cytometry analysis.

[0070] ③ Collect intestinal tissue, extract total RNA using the Trizol method, and reverse transcribe it into cDNA. β-actin was used as an internal control. -ΔΔCt The expression changes of il-1β, il-8, tnf-α, ncf1, tgf-β, and il-10 genes were detected and analyzed using relative quantitative methods.

[0071] ④ Intestinal tissue was collected, fixed overnight with 4% paraformaldehyde, dehydrated by ethanol gradient, cleared with xylene, embedded in paraffin, and sectioned (5 μm). After drying, dewaxing, and hydration, the sections were analyzed for tissue damage and inflammatory infiltration using H&E staining.

[0072] Results Analysis

[0073] 1. Baicalein improves the survival rate of largemouth bass after MRV challenge.

[0074] To evaluate the protective effect of baicalin against MRV infection in largemouth bass, survival curves were analyzed to investigate the effect of different doses of oral baicalin on the survival rate of infected fish. Figure 8 As shown, the survival rate of the MRV-infected control group was 40.00%, while oral administration of baicalein improved the survival rate of the attacked fish: 50.00% in the 7 mg / kg dose group, 56.67% in the 21 mg / kg dose group, and 73.33% in the 63 mg / kg dose group. Compared with the MRV-infected control group, the survival rate of the 63 mg / kg dose baicalein oral administration group was significantly improved (*p<0.05).

[0075] Figure 8 A graph showing the survival rate of largemouth bass challenged with MRV.

[0076] 2. Baicalein effectively inhibits the proliferation of MRV in largemouth bass.

[0077] The viral load in various tissues of surviving fish after oral administration of different doses of baicalein (7 mg / kg, 21 mg / kg, and 63 mg / kg) in the MRV-infected control group was quantitatively analyzed using qPCR. The results showed that ( Figure 9 The viral load in various tissues of the MRV-infected control group of dead fish was 7.71 × 10⁻⁶. 7 ~5.82×10 8 The viral load in various tissues of surviving fish in the MRV-infected control group was 3.85 × 10⁻⁶ copies / ng total DNA. 5 ~9.58×10 6 The viral load in the surviving fish in the baicalein oral administration group was 1.76 × 10⁻⁶ copies / ng total DNA. 4 ~8.08×10 5 The viral load of baicalein was within the range of copies / ng total DNA. As shown in Table 1, compared with the viral load in fish that died after MRV infection (untreated), the in vivo inhibition rate of baicalein against MRV was 99.35%–99.99%; compared with the viral load in fish that survived MRV infection (untreated), the in vivo inhibition rate of baicalein against MRV was 88.68%–99.11%. The above results indicate that oral administration of baicalein can effectively inhibit the replication of MRV in largemouth bass.

[0078] Table 1. Inhibition rate of baicalin on MRV proliferation in largemouth bass

[0079]

[0080] Figure 9The figure shows the results of the inhibitory effect of baicalin on viral load in tissues of surviving fish infected with MRV.

[0081] 5. Baicalein reduces intestinal ROS after MRV infection.

[0082] Intestinal single-cell suspensions isolated by enzymatic digestion were assessed for cell viability using trypan blue staining. After counting and calculation, the cell viability was greater than 90%, indicating that the isolation process had minimal impact on cell viability and that the suspensions could be used for subsequent experiments. Figure 10 ).

[0083] To evaluate the inhibitory effect of baicalin on ROS production, the ROS levels in intestinal single cells were quantitatively analyzed using the DCFH-DA fluorescent probe combined with flow cytometry. Results are as follows: Figure 11 As shown, compared with the 24.6% reactive oxygen species (ROS) level in the blank control group, the ROS level in the intestinal cells of surviving fish in the MRV-infected control group increased to 57.5% (p<0.05). Compared with the 57.5% ROS level in the MRV-infected control group, baicalin significantly inhibited the increase of ROS in a dose-dependent manner: the ROS level in the 7 mg / kg dose group decreased to 51.0% compared with the infected group (p<0.05), the ROS level in the 21 mg / kg dose group decreased to 46.3% compared with the infected control group (p<0.05), and the ROS level in the 63 mg / kg dose group decreased to 36.5% compared with the infected group (p<0.05). This result indicates that baicalin can effectively inhibit the excessive ROS production caused by viral infection and alleviate the oxidative stress damage of host cells in a dose-dependent manner.

[0084] Figure 10 This is a diagram showing the results of single-cell isolation from the intestine.

[0085] 6. Effects of baicalin on the expression of inflammation-related genes

[0086] The effects of baicalin on the expression levels of inflammation-related genes (IL-1β, IL-8, TNF-α, NCF1, TGF-β, IL-10, etc.) after MRV infection were detected using RT-qPCR. The results showed that... Figure 12The intestinal tissue of surviving fish in the MRV-infected control group exhibited significant pro-inflammatory characteristics: compared with the blank control group, the mRNA expression levels of il-1β, il-8, tnf-α, and ncf1 in the intestines of surviving fish in the MRV-infected control group were upregulated by approximately 3.02-fold, 1.74-fold, 1.85-fold, and 1.74-fold, respectively. Oral administration of baicalein significantly reversed the MRV-induced high expression of inflammatory factors; the expression levels of the above genes in surviving fish in the baicalein oral administration group were reduced by approximately 0.20–0.76-fold, 0.60–0.80-fold, 0.46–0.79-fold, and 0.47–0.6-fold, respectively, compared with those in the MRV-infected control group. The mRNA expression levels of anti-inflammatory factors tgf-β and il-10 in the intestines of surviving fish in the baicalein oral administration group were approximately 1.56–3.33-fold and 1.09–2.24-fold, respectively, compared with those in the MRV-infected control group.

[0087] Figure 11 Figure showing the effect of baicalin on ROS levels in MRV-infected intestinal cells;

[0088] Figure 12 The figure shows the effect of baicalin on the expression of inflammation-related genes.

[0089] 7. Baicalein reduces MRV infection-induced intestinal tissue damage.

[0090] H&E staining results showed ( Figure 13 Compared with the blank control group, the intestinal tissue of fish infected with MRV showed obvious pathological damage, specifically manifested as disordered villus structure, partial villus loss, submucosal diffusion, and loss of intestinal structural integrity. Oral administration of different doses of baicalin significantly reduced the shedding of intestinal villus epithelial cells and the degree of submucosal diffusion. This indicates that baicalin can improve intestinal tissue damage caused by MRV infection.

[0091] Figure 13 Figure showing the results of baicalin alleviating MRV-induced intestinal tissue damage.

[0092] In summary, baicalin can effectively inhibit excessive ROS production caused by MRV virus infection and alleviate oxidative stress damage to host cells in a dose-dependent manner. Furthermore, it can reduce the high expression of pro-inflammatory genes such as il-1β, il-8, tnf-α, and ncf1, while increasing the expression of anti-inflammatory genes il-10 and tgf-β, thus mitigating MRV-induced intestinal tissue damage, effectively inhibiting MRV proliferation in largemouth bass, and improving the survival rate of largemouth bass after MRV challenge.

[0093] It will be apparent to those skilled in the art that the present invention is not limited to the details of the exemplary embodiments described above, and that the invention can be implemented in other specific forms without departing from the spirit or essential characteristics of the invention. Therefore, the embodiments should be considered in all respects as exemplary and non-limiting, and the scope of the invention is defined by the appended claims rather than the foregoing description. Thus, it is intended that all variations falling within the meaning and scope of equivalents of the claims be included within the present invention.

Claims

1. Use of baicalin in the preparation of products for the prevention and / or treatment of fish infected with mandarin frog iridovirus.

2. The use according to claim 1, characterized in that, The chemical formula of the baicalein mentioned is C 16 H 12 O5 has a molecular weight of 284.

26.

3. The use according to claim 1, characterized in that, The product in question is a drug whose active ingredient is baicalein.

4. The use according to claim 3, characterized in that, The drug is available in oral or injectable form.

5. The use according to claim 1, characterized in that, The product in question is fish feed or fish feed additive containing baicalein.

6. A drug for treating mandarin frog iridovirus, characterized in that, The medicine contains baicalein.

7. A fish feed resistant to mandarin frog iridovirus, characterized in that, Fish feed contains baicalein.

8. A fish feed additive resistant to mandarin frog iridovirus, characterized in that, Fish feed additives contain baicalin.

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