A Chinese medicinal compound preparation for preventing and treating Mycoplasma gallisepticum infection and its preparation method

Abstract

The invention discloses a Chinese medicine compound preparation for preventing and treating Mycoplasma gallisepticum infection and a preparation method thereof, and belongs to the technical field of Chinese veterinary medicine. The Chinese medicine compound preparation comprises 15-30 parts of Pueraria root, 15-30 parts of Scutellaria baicalensis, 15-30 parts of Polygonum cuspidatum, 5-20 parts of Houttuynia cordata, 5-20 parts of Chrysanthemum, 5-20 parts of Radix Isatidis, 3-10 parts of Platycodon grandiflorum, 3-10 parts of Bamboo Leaf, 3-10 parts of Mint and 3-10 parts of Licorice. The invention mixes the above raw materials, decocts them with water, filters them after cooling, and feeds 1 mL of the filtrate to chickens per kilogram of body weight every day or orally for 7 consecutive days. The product of the invention is a pure Chinese medicine preparation with reasonable combination and compatibility, which can not only inhibit the growth of Mycoplasma gallisepticum, but also has the effects of clearing away heat and detoxifying, moistening the lungs and relieving cough and anti-inflammatory, and has a significant effect on chronic respiratory diseases caused by Mycoplasma gallisepticum infection. The invention can improve and regulate the imbalance in the disease process, treat both the symptoms and the root causes, has a high cure rate, is safe and reliable, is not prone to drug resistance, ensures food safety, has a simple preparation method, is low-cost, and has broad application prospects.

Description

Technical Field

[0001] The invention relates to the technical field of traditional Chinese medicine, and in particular to a traditional Chinese medicine compound preparation for preventing and treating Mycoplasma gallisepticum infection and a preparation method thereof. Background Art

[0002] Mycoplasma gallisepticum (MG) is the pathogen of chronic respiratory disease (CRD) and infectious sinusitis in turkeys, causing clinical symptoms such as coughing, tearing, and respiratory rales in chickens. CRD is widely prevalent worldwide. Chickens of different breeds, ages, and genders can get sick in all seasons, and the incidence rate is increasing year by year. It is one of the important infectious diseases that endanger the poultry industry.

[0003] MG is mainly adsorbed on the host respiratory mucosa surface through the special terminal structure of lipid-related proteins on the membrane surface. Its adhesion proteins bind to the receptors on the surface of host cells, causing host cell denaturation, tracheal cilia damage, and destruction of the natural barrier of the mucosa and the immune system. If not prevented and treated in time, it will lead to multi-system and multi-organ lesions. MG invades chicken farms and is difficult to be completely eliminated. It exists in chickens for a long time, causing repeated infections and spread, resulting in reduced poultry feed utilization, slow growth, increased mortality, decreased egg production, and increased deformed egg rates. MG is often mixed or secondary infected with Escherichia coli, infectious bronchitis virus, Newcastle disease virus, etc., which is more harmful to chickens, with a mortality rate of more than 30%, causing huge economic losses to the poultry industry.

[0004] Currently, the most effective way to prevent MG infection in chickens is vaccination. Although chickens can produce a certain degree of immune protection after vaccination, it cannot prevent infection, and immune chickens can still get sick in large numbers under stress. In addition, MG is constantly mutating, and vaccines need to be developed in a timely manner to deal with mutant strains. In production, antibiotics such as doxycycline, tylosin, tylosin, tilmicosin and azithromycin are mainly used to treat CRD. Antibiotics can quickly kill Mycoplasma gallisepticum in the early stage of infection and alleviate the clinical symptoms of chickens, but they are prone to relapse after discontinuation of the drug, and long-term use of antibiotics will lead to increased drug resistance of MG, and toxic side effects and drug residues are prominent. Therefore, it is urgent to explore ideal drugs for the prevention and treatment of Mycoplasma gallisepticum infection.

[0005] Traditional Chinese medicine has the effects of dispelling evil and strengthening the body, clearing away heat and detoxifying, relieving cough and relieving asthma, and has the effects of antibacterial, antiviral, and enhancing the body's immunity. The ingredients of traditional Chinese medicine are all biological organic matter, with small toxic and side effects, low drug residue, and long-term safe and reliable use. Therefore, inventing a traditional Chinese medicine compound preparation that can effectively prevent and treat Mycoplasma gallisepticum infection is a problem that researchers in this field urgently need to solve. Summary of the invention

[0006] The invention provides a traditional Chinese medicine compound preparation for preventing and treating Mycoplasma gallisepticum infection and a preparation method thereof.

[0007] The present invention adopts the following technical scheme:

[0008] The medicine of the present invention is composed of the following raw materials in parts by weight: 15-30 parts of kudzu root, 15-30 parts of scutellaria, 15-30 parts of knotweed, 5-20 parts of houttuynia, 5-20 parts of chrysanthemum, 5-20 parts of isatis root, 3-10 parts of platycodon, 3-10 parts of bamboo leaf, 3-10 parts of mint and 3-10 parts of liquorice.

[0009] The present invention further preferably comprises the following components by weight: 17 parts of kudzu root, 22 parts of scutellaria, 16 parts of polygonum cuspidatum, 10 parts of houttuynia cordata, 10 parts of isatis root, 5 parts of platycodon, 5 parts of bamboo leaf, 5 parts of mint and 10 parts of liquorice.

[0010] In the above formula, the effects of each raw material are as follows:

[0011] Pueraria root: It enters the lung, spleen, and stomach meridians. It has a light Qi, is good at dispelling exterior pathogens and inducing sweat, and can raise the clear Yang Qi of the spleen and stomach - "Bencao Zhengyi".

[0012] Scutellaria baicalensis: It enters the lung and spleen meridians, and is mainly used to relieve the fire in the upper part of the lung and stomach. It is mainly used to treat adverse qi in the chest, block hot phlegm, and relieve asthma and cough. - "Pharmaceutical Chemistry".

[0013] Polygonum cuspidatum: It enters the liver, gallbladder, and lung meridians. It has the effects of dispelling wind and dampness, relieving cough and reducing phlegm. It can be used for cough caused by lung heat, and cough with excessive phlegm - "Chinese Pharmacopoeia".

[0014] Houttuynia cordata: enters the liver and lung meridians, clears away heat and detoxifies, discharges pus and eliminates carbuncle. Treats lung carbuncle cough, anti-inflammatory and swelling - "Chinese Materia Medica".

[0015] Chrysanthemum: enters the lung and liver meridians, dispels wind and clears heat, soothes the liver and improves eyesight, clears the depressed fire in the three burners, eliminates chest heat, soothes the intestines and stomach, benefits the five meridians, and regulates the limbs - "Bielu".

[0016] Isatis root: enters the liver and stomach meridians, clears away heat and detoxifies, cools the blood and relieves sore throat, prevents epidemics and kills insects - "Bencao Bian Du".

[0017] Platycodon grandiflorum: enters the lung meridian, opens and promotes lung qi, removes phlegm and discharges pus, leads other medicines upward, and serves as a vessel for other medicines - "Bencao Qiuzhen".

[0018] Bamboo leaves: sweet, mild and slightly cold, enter the heart, lung, liver and gallbladder meridians, have the effect of cooling the heart and spleen, clearing phlegm and relieving cough. Bamboo leaves grow on branches, can penetrate the medicine, lead the medicine upward, treat wind evil in the upper part of the body, irritability, cough, reverse breathing - "Compendium of Materia Medica".

[0019] Peppermint: enters the lung and liver meridians, dispels wind and heat, detoxifies and opens the orifices, is good at moving to the skin and surface, and guides other medicines into the Ying and Wei to relieve stagnant Qi - "Pharmaceutical Chemistry".

[0020] Licorice: enters the heart, lung, spleen, and stomach meridians, tonifies the spleen and replenishes qi, moistens the lungs, clears away heat and detoxifies, and harmonizes various medicines - "Bencao Jingjie".

[0021] Pueraria root is pungent, sweet and slightly cold in nature. When used with Scutellaria baicalensis and Polygonum cuspidatum, it can remove the fire in the upper part of the body, dispel the heat in the lungs, and achieve the effects of relieving cough, removing phlegm and relieving asthma. It plays a major role and is the main drug. Houttuynia cordata, chrysanthemum and Isatis root all clear away heat and detoxify, assisting the main drug in dispersing wind and clearing heat, calming the liver and improving eyesight, calming the mind and benefiting the orifices, and are the minister drugs. Platycodon grandiflorum opens and promotes lung qi and guides the medicine upward; bamboo leaves grow on branches and are light and floating, which can penetrate the medicine and guide the medicine to float upward; mint guides the medicine into the Weiying. These three can all carry the medicine and are adjuvant drugs. Licorice nourishes the spleen and replenishes qi, moistens the lungs and relieves coughs, and harmonizes the medicines. The combination of various medicines plays the effects of clearing heat and detoxifying, opening and promoting lung qi, removing phlegm and relieving coughs, calming the mind and improving eyesight.

[0022] The preparation method of the Chinese medicinal compound preparation of the present invention is as follows:

[0023] Weigh various raw materials according to the above compatibility ratio, except for houttuynia and mint, mix the rest evenly, put them in a decoction machine, soak them in cold water for 30 minutes, boil them over high heat, simmer them over low heat for 30 minutes, then add houttuynia and mint, and continue to simmer them over low heat for 15 minutes. After the decoction is completed, filter it after cooling, bag the liquid medicine, store it at 4℃ for later use, and preheat the liquid medicine in a 40℃ water bath before use.

[0024] The method for using the Chinese medicinal liquid of the present invention is as follows: 1 ml of the medicinal liquid is administered or drunk by the chicken per kilogram of body weight every day (each ml of the medicinal liquid is equivalent to 250 mg of the original medicinal material), and the method is used continuously for 7 days.

[0025] Beneficial effects of the present invention: (1) The Chinese medicine compound preparation provided by the present invention has significant efficacy on mycoplasma infection, can improve the body's immunity, and prevent the decline of production performance. (2) The compound preparation is a pure Chinese medicine preparation, without adding any antibiotics or other chemically synthesized antibiotics, avoiding the occurrence of drug resistance problems, and the compound preparation is harmless to the body, has no residue in livestock and poultry products, improves the safety of poultry products, and is beneficial to human health. (3) The compound preparation has low cost and simple production process, and is suitable for actual production applications. BRIEF DESCRIPTION OF THE DRAWINGS

[0026] Figure 1 It is the minimum inhibitory concentration of Chinese herbal compound preparation against Mycoplasma gallisepticum.

[0027] Figure 2 Trachea slices of chickens in each group

[0028] Figure 2-1 This is the trachea of ​​a normal chicken, with complete structure and tissue, and clearly visible cilia;

[0029] Figure 2-2The trachea of ​​the chickens treated with the compound Chinese medicine preparation alone had a complete tissue structure, and the cilia of the mucosal epithelial cells were continuous and clearly visible, with no difference from the normal group;

[0030] Figure 2-3 This is the trachea of ​​a chicken in the MG infection model group, with cilia loss, mucosal thickening, and local mucosal epithelial necrosis and shedding;

[0031] Figure 2-4 , 2-5 2-6 are the trachea of ​​chickens in the low-dose prevention group, the medium-dose prevention group and the high-dose prevention group, respectively. The tissue structure is intact, the cilia are clearly visible, and there is no loss, which is no different from the trachea of ​​the normal group;

[0032] Figure 2-7 In the chicken trachea of ​​the low-dose compound treatment group, the cilia were completely lost, the mucosal epithelium was degenerated, the mucosal layer was significantly thickened, and the number of mononuclear lymphocytes was significantly increased;

[0033] Figure 2-8 This is the trachea of ​​the chicken in the medium-dose compound treatment group. The tissue structure is relatively intact, the cilia are partially lost (see black arrows), and lymphoid follicles appear;

[0034] Figure 2-9 This is the trachea of ​​the chicken in the high-dose compound treatment group, with normal tissue structure and complete and clear cilia, which is no different from the normal group;

[0035] Figure 2-1 0 is the trachea of ​​chickens in the tylosin treatment group, with intact mucosal epithelium and visible but sparse cilia (see black arrows). DETAILED DESCRIPTION

[0036] In order to facilitate a better understanding of the present invention, the specific therapeutic effects of the present invention are described below by specific examples. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in the field without creative work are within the scope of protection of the present invention. The Chinese medicine raw materials described therein are all commercially available unless otherwise specified, and the instruments involved are all conventional instruments unless otherwise specified, and the experimental methods involved are all conventional methods unless otherwise specified.

[0037] Embodiment 1: Weigh 85g of kudzu root, 110g of scutellaria, 80g of knotweed, 50g of houttuynia, 50g of isatis root, 25g of platycodon, 25g of bamboo leaf, 25g of mint, and 50g of liquorice. Mix kudzu root, scutellaria, knotweed, isatis root, platycodon, bamboo leaf, and liquorice evenly, place in a decoction machine, soak in cold water for 30 minutes, boil over high heat, simmer over low heat for 30 minutes, then put in mint and houttuynia and decoct for 15 minutes. After the liquid medicine is cooled, filter, bag, store at 4°C for standby use, and preheat the liquid medicine with a 40°C water bath before use. The concentration of the liquid medicine prepared this time is 250mg / mL.

[0038] Example 2: In vitro antibacterial test of the compound Chinese medicine preparation of the present invention

[0039] The compound Chinese medicine preparation obtained in the embodiment of the present invention was concentrated to 500 mg / mL, filtered and sterilized, and the minimum inhibitory concentration of the compound Chinese medicine preparation was determined by the two-fold dilution method. The concentration of the Mycoplasma gallisepticum (MG-HS) culture was diluted to 1×10 4 CCU / mL. Take 13 2mL sterile EP tubes, the final concentration of the drug solution in each tube is 100, 50, 25, 12.5, 6.25, 3.13, 1.56, 0.78, 0.39, 0.195, 0.098, 0.048, 0.024mg / mL, and then add 0.5mL of Mycoplasma gallisepticum solution to each tube. Take another 2mL sterile EP tube and add 1mL of culture medium as a control. Culture at 37℃ for 14 days, observe the color change, and find that the minimum inhibitory concentration of the compound Chinese medicine preparation is 25mg / mL.

[0040] Example 3: Efficacy test of the compound Chinese medicine preparation of the present invention

[0041] 180 Qingjiao Ma chickens were raised to 11 days of age, weighed, and randomly divided into 10 groups, 18 in each group, 3 replicates in each group, and 6 in each replicate. The groups were A (normal group), B (compound Chinese medicine preparation group), C (MG infection model group), D (low-dose prevention group), E (medium-dose prevention group), F (high-dose prevention group), G (low-dose treatment group), H (medium-dose treatment group), I (high-dose treatment group), and J (tylosin treatment group). Among them, the chickens in group A were not treated during the entire experimental period. From the 12th day, the chickens in group B were gavaged with 0.5 mL of compound Chinese medicine preparation per kilogram of body weight per day, and the drugs were stopped after 21 consecutive days of use and then raised normally. On days 12-18, chicks in groups D, E, and F were gavaged with 0.25 mL, 0.5 mL, and 1 mL of the drug per kg body weight per day, respectively, while groups C, G, H, I, and J were not treated. On days 19-23, groups C, D, E, F, G, H, I, and J were artificially challenged with 1×10 9Each chicken was given 0.3 mL of MG-HS bacterial liquid with CCU / mL of nasal drops and eye drops and 0.7 mL of air bag injection every day. After the infection was completed, the infection was checked; from the 24th to the 30th day, groups D, E, F, G, H, and I were treated with traditional Chinese medicine, and the dosage was 0.25 mL / kg for groups D and G, 0.5 mL / kg for groups E and H, 1 mL / kg for groups F and I, and group J was given tylosin drinking water at a dosage of 350 mg / L; from the 31st to the 44th day, all groups of chickens were raised normally without any treatment. During the entire trial period, production performance indicators were recorded and clinical symptoms of chickens were observed. At the end of the trial on the 45th day, all chickens were blooded and slaughtered to observe the pathological changes of internal organs. The middle section of the trachea of ​​each group of chickens was fixed with 4% formaldehyde to prepare paraffin sections, and the degree of tracheal damage was observed by hematoxylin and eosin staining. The results are as follows:

[0042] (1) Clinical manifestations of chickens

[0043] When the disease is latently infected, its symptoms are not obvious or typical enough. If the disease is simply contracted, inflammation and swelling occur in the trachea and bronchi, causing the serous and viscous fluid in the respiratory tract to flow upward to form serous and viscous nasal fluid. Due to the swelling of the trachea and pharynx, the pharynx becomes itchy and painful, which in turn causes sneezing and coughing in chickens. When the infection worsens, sinusitis will occur; secretory and viscous serous fluid will flow out of the eyes, making it difficult to open them. In severe cases, the face will swell and the eyes will become blind. There are a lot of sputum in the respiratory tract of sick chickens, causing breathing difficulties, such as frequent rales, continuous open mouth breathing, coughing and mild or even severe panting. Severely ill chickens are extremely depressed, their feathers are fluffy and messy, they don't like to eat feed, they drink more water, their growth stagnates, and even negative growth occurs.

[0044] (2) Effects of compound Chinese medicine preparations on chicken production performance

[0045] As shown in Table 1 and Table 2, there were no significant differences in weight gain and feed-to-meat ratio between the group using the compound preparation alone and the normal group, indicating that the compound Chinese medicine had no effect on the production performance of chickens.

[0046] After the virus attack, the body weight gain of the MG infection model group was significantly lower than that of the other groups; there was no significant difference in body weight gain among the normal group, the compound preparation group, and the high and medium dose prevention groups, while the body weight gain of the low dose prevention group was slightly higher than that of the infection group, but there was no significant difference. In terms of feed-to-meat ratio, the infection group was significantly higher than the other groups, and the low dose prevention group was significantly lower than the infection group but higher than the other groups.

[0047] After 7 days of drug treatment, the weight gain and feed-to-meat ratio of each group are shown in Table 2 Treatment period. In terms of weight gain, the MG infection group was significantly lower than the other groups, but there was no significant difference with the G group (low-dose treatment group); there was no significant difference between the DEF group (preventive group), the IH group (high and medium dose treatment group) and the J group (Tiamulin group); there was no significant difference between the D group (preventive low dose group), the GHI group (treatment group) and the J group (Tiamulin group); there was no significant difference between the FE group (preventive high and medium dose group), the A group (normal group) and the B group (compound preparation group). In terms of feed-to-meat ratio, the MG infection group was significantly higher than the other groups, and there was no significant difference between the other groups.

[0048] After the treatment was completed and the drug was stopped for observation for 14 days, the weight gain and feed-to-meat ratio of each group were shown in Table 2. In terms of weight gain, the MG infection group was significantly lower than the other groups; there was no significant difference between the DEF group (prevention group), GHI group (treatment group), and J group (Tymi group), but it was significantly lower than the A group (normal group) and B group (compound preparation group). In terms of feed-to-meat ratio, as the chickens grew older, there was no significant difference among the groups.

[0049] In summary, the use of high- and medium-dose compound Chinese medicines to prevent MG infection can effectively prevent the decline in chicken production performance; after MG infection, if the production performance of chickens needs to be effectively improved in a short period of time, high- and medium-dose compound Chinese medicines can be used for treatment; and low-dose compound Chinese medicines can also improve the production performance of infected chickens for a longer period of time.

[0050] Table 1 Effect of compound preparation on chicken production performance in preventing MG infection

[0051]

[0052] Table 2 Effect of compound preparation on production performance in preventing and treating MG infection

[0053]

[0054] (3) Protective effect of compound Chinese medicine on air sac and trachea

[0055] After the experiment, all chickens were slaughtered and the air sac injury was scored. The air sac was thin and transparent with a glossy texture, which was scored as 0 points; the air sac had yellow spots, which was scored as 1 point; the yellow cheese on the air sac surface was less and foamy, which was scored as 2 points; the air sac surface had more yellow cheese, which was obviously thickened and became turbid, which was scored as 3 points; the air sac surface was attached with a large amount of yellow cheese-like exudate, which was scored as 4 points. Air sac injury reduction rate = (air sac score of the infected group - air sac score of the experimental group) / air sac score of the infected group. The scoring results are shown in Table 3.

[0056] The air sac score of the MG infection group was significantly higher than that of the other groups; there was no difference between the high-medium-dose group for prevention and the high-dose group for treatment and the normal group and the compound group, and it was significantly lower than the other groups; there was no significant difference between the low-dose group for prevention, the low-medium-dose group for treatment and the Thaimil group. It can be seen that after stopping the drug and continuing to observe for 14 days, the air sac injury reduction rate of chickens in the high-medium-dose group for prevention and the high-dose group for treatment was the highest, reaching 100%, followed by the low-dose group for prevention, and the effect was better than the Thaimil group; the air sac injury reduction rate of the low-medium-dose group for treatment was also more than 70%. This shows that the use of compound Chinese medicine for prevention and treatment can effectively protect the air sac of chickens.

[0057] Table 3 Air sac injury scores and air sac injury reduction rates of chickens in each group

[0058]

[0059] According to the slice results (Figure Figure 2 ), the cilia of the mucosal epithelial cells in the normal group were intact and clearly visible, while the cilia of the MG infection group were almost completely shed, the mucosal layer was significantly thickened, and local mucosal epithelial cells were necrotic and shed; the tissue structure of the compound preparation group, the high, medium, and low dose prevention groups, and the high dose treatment group were all intact, with dense, continuous, and clearly visible cilia, which were no different from the normal group; the tissue structure of the medium dose treatment group was relatively intact, with local cilia shed and lymphoid follicles; the cilia of the low dose treatment group were severely shed, the mucosal epithelium was degenerated, the mucosal layer was significantly thickened, and the number of mononuclear lymphocytes increased significantly; the mucosal epithelium of the Thai Miao group was intact, and the cilia were visible, but some were sparse. The above results show that MG infection can destroy the ciliary structure of the trachea, cause the thickening of the tracheal mucosa, and cause the necrosis and shedding of mucosal epithelial cells, and the use of compound Chinese medicine for prevention and treatment can effectively repair the inflammatory damage caused by MG to the trachea.

[0060] (4) Effects of compound Chinese medicine preparations on the prevention and treatment of Mycoplasma gallisepticum infection

[0061] The effect of compound Chinese medicine on the prevention and treatment of chronic respiratory disease in artificially infected chickens is shown in Table 4. The results show that after the virus attack, the infection group, prevention group, treatment group and tylosin group were all generally infected, but the morbidity of the prevention group was significantly lower, indicating that the prevention had a certain effect; among them, the mortality rate of the MG infection model group was the highest, and there was no MG mortality in the other groups; the cure rate of the high and medium dose prevention group was the highest, reaching more than 90%, and the effect was equivalent to tylosin; followed by the high and medium dose treatment group, with a cure rate of 87.5%; and then the prevention and treatment low dose group, but its cure rate also reached more than 80%.

[0062] Table 4 The effect of compound Chinese medicine on the prevention and treatment of chronic respiratory disease in artificially infected chickens

[0063]

[0064] In summary, the compound Chinese medicine preparation provided by the present invention has a high antibacterial effect and has a significant preventive and therapeutic effect on artificially modeled chronic respiratory disease chickens. In addition, the preparation process is simple, the dosage is small, the cost is low, it is not easy to produce drug resistance, the veterinary drug residues are reduced, the safety of livestock and poultry products is improved, and it can be used for the prevention and treatment of chronic respiratory diseases caused by Mycoplasma gallisepticum.

Claims

1. A Chinese medicinal compound preparation for preventing and treating Mycoplasma gallisepticum infection, characterized in that The medicine is prepared from the following raw materials in parts by weight: 15-30 parts of kudzu root, 15-30 parts of scutellaria, 15-30 parts of knotweed, 5-20 parts of houttuynia, 5-20 parts of chrysanthemum, 5-20 parts of isatis root, 3-10 parts of platycodon, 3-10 parts of bamboo leaf, 3-10 parts of mint and 3-10 parts of liquorice.

2. A method for preparing a Chinese medicinal compound preparation for preventing and treating Mycoplasma gallisepticum infection according to claim 1 is as follows: (1) Weighing each API according to the weight portion described in claim 1; (2) Mix kudzu root, scutellaria, knotweed, chrysanthemum, isatis root, platycodon, bamboo leaf and liquorice, put them into a decoction machine, and soak them in cold water for 30 minutes, with the water covering the medicine by 3 to 4 cm; (3) Bring to a boil over high heat, then simmer over low heat for 45 minutes; (4) After boiling for 30 minutes, add Houttuynia cordata and mint and continue to boil for 15 minutes; After cooling, filter, bag, and store at room temperature or in the refrigerator for later use; it can be stored at room temperature for 7 days and at 4℃ for 15 days.

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