Laying hen metritis molding agent as well as preparation method and application thereof

Abstract

The invention discloses a laying hen metritis molding agent as well as a preparation method and application thereof. The molding agent comprises the following components: a lipopolysaccharide solution and a phenol solution in a volume ratio of (0.5-1.5): (0.5-1.5). According to the method, lipopolysaccharide and a phenol solution are used as a molding reagent, and the lipopolysaccharide and the phenol solution act together, so that inflammation can be caused while the mucosal barrier of the uterus part of the fallopian tube of the laying hen is damaged, and the problems that a pathogenic microorganism molding agent has no effect and a chemical reagent molding agent is separated from actual production are solved; according to the method, inflammatory reaction can be rapidly generated, typical symptoms of laying hen hysteritis appear, the molding dosage is accurately mastered, the repeatability is improved, the method is not easily influenced by microbial toxicity and organism immunity, pollution caused by pathogenic bacteria is reduced, and the requirement for a test site is lowered.

Description

Technical Field

[0001] The invention relates to the field of animal pathology modeling, in particular to a laying hen metritis modeling agent and a preparation method and application thereof. Background Art

[0002] In laying hen farming, metritis is a common and frequently occurring disease. Laying hens of different ages are susceptible, especially during the peak egg-laying period. Common pathogens that cause metritis in laying hens are Gram-negative bacteria, such as Escherichia coli and Salmonella. When the protective power of the mucosal barrier of the oviduct of laying hens is weakened, pathogenic bacteria can easily invade the uterine part of the oviduct through the cloaca, causing varying degrees of edema and bleeding in the uterus of laying hens, and then fibrin, pus, and cheesy exudates appear in the uterus, causing a decrease in the egg production rate of laying hens and an increase in unqualified eggs. In severe cases, it can lead to peritonitis or even death. Laying hens suffering from metritis will lead to a decline in animal welfare, reduced egg production performance, and affected reproductive performance, causing great economic losses and affecting the vigorous development of the laying hen farming industry.

[0003] At present, most of the research on modeling of uterine inflammation in laying hens at home and abroad uses pathogenic microorganism modeling or chemical reagent modeling. Pathogenic microorganism modeling mainly uses Salmonella, Escherichia coli or mixed bacteria inoculation, which has the advantage of being consistent with actual clinical symptoms, but this method has a slow reaction, the inflammatory reaction is not obvious, there is a risk of pathogenic bacteria contaminating the environment, and the success rate of modeling is easily affected by the virulence of microorganisms. When the mucosal barrier is in good condition, this method cannot achieve obvious results, resulting in the failure of establishing a uterine inflammation model. Escherichia coli has weak resistance to the outside world and can usually be killed at 60°C for 15 minutes or 55°C for 60 minutes. The products of Escherichia coli freeze-dried powder vary greatly from batch to batch, the strain is not stable, easy to mutate, and the quality cannot be kept consistent. After freeze-drying, Escherichia coli freeze-dried powder needs to be activated and cultured, and the growth delay period is long. It requires two consecutive subcultures to grow normally. During the operation, protective measures need to be taken throughout the process to prevent the strain from being contaminated. The spread of pathogens may also occur during the activation process, leading to environmental pollution. In recent years, lipopolysaccharide (LPS) has been used alone to replace Salmonella and Escherichia coli for metritis modeling. As the main virulence factor of the cell wall of Gram-negative bacteria, it has strong biological activity and can cause a strong systemic inflammatory response in the body. However, the use of LPS alone will lead to modeling failure, which may be related to the integrity of the uterine mucosal barrier and normal structure of laying hens. Some researchers used phenol to corrode the uterine mucosa of laying hens, thereby causing metritis in laying hens, but the dosage is difficult to control and can easily lead to stress death of laying hens. Summary of the invention

[0004] The purpose of the present invention is to provide a modeling agent for laying hen metritis and a preparation method and application thereof. The modeling agent can quickly cause inflammatory reactions in the vagina and uterus, with typical inflammatory symptoms and high susceptibility rate.

[0005] In order to solve the problems existing in the prior art, the technical solution adopted by the present invention is: In a first aspect, the present invention provides a laying hen metritis modeling agent, comprising the following components and composed in the following volume ratio: lipopolysaccharide solution: phenol solution = (0.5-1.5): (0.5-1.5), preferably lipopolysaccharide solution: phenol solution = 1: 1. Furthermore, the volume concentration of liquefied phenol in the phenol solution is 20-30%, preferably 25%.

[0006] Furthermore, the phenol solution is composed of the following components by volume percentage: 20-30% liquefied phenol, 2-3% Tween 20, 2-3% Span 20, 9-11% glucose, 55-65% phosphate buffer solution.

[0007] Preferably, the phenol solution consists of the following components by volume percentage: 25% liquefied phenol, 2.5% Tween 20, 2.5% Span 20, 10% glucose, 60% phosphate buffer solution.

[0008] Furthermore, the lipopolysaccharide solution is prepared from lipopolysaccharide and phosphate buffered saline (PBS), and the concentration of the lipopolysaccharide solution is 3.5-4 mg / ml, preferably 3.7 mg / ml.

[0009] A large number of tests have confirmed that the modeling agent has a better effect when the lipopolysaccharide solution with a concentration of 3.5-4 mg / ml is combined with the phenol solution. The modeling agent has the best effect when the lipopolysaccharide solution concentration is 3.7 mg / ml. If the concentration of the lipopolysaccharide solution is too low, the success rate of modeling will be reduced, and modeling failure will occur; if the concentration of the lipopolysaccharide solution is too high, it will not only increase the cost of modeling, but the effect will be reduced compared to the 3.5-4 mg / ml lipopolysaccharide solution.

[0010] Furthermore, the concentration of the phosphate buffer solution is 0.01 M, and the pH value is 7.0-7.4, preferably pH 7.2.

[0011] In a second aspect, the present invention provides a method for preparing the laying hen metritis modeling agent described in the first aspect, comprising the following steps: ①Prepare lipopolysaccharide solution: Heat the phosphate buffer solution, add lipopolysaccharide and stir until it is completely dissolved; ②Preparing phenol solution: After heating phenol until it is completely melted into liquefied phenol, heat the phosphate buffer solution, add the liquefied phenol, Tween 20, Span 20, and glucose, and stir until they are completely dissolved; ③ Evenly mix the lipopolysaccharide solution prepared in step ① and the phenol solution prepared in step ② according to the proportion.

[0012] In a third aspect, the present invention provides use of the laying hen metritis modeling agent described in the first aspect in preparing a laying hen metritis model.

[0013] In a fourth aspect, the present invention provides a method for modeling metritis in laying hens, comprising the following steps: The oviduct opening of the laying hen is exposed, and then the laying hen metritis modeling agent described in the first aspect is placed into the uterus of the laying hen oviduct, and observed for 24-72 hours, preferably 48 hours.

[0014] Furthermore, the dosage of the laying hen metritis modeling agent is 1-3 ml / laying hen, preferably 2 ml / laying hen.

[0015] The advantages and beneficial effects of the present invention are: Lipopolysaccharide is an endotoxin extracted from Escherichia coli, and phenol is a corrosive chemical reagent. The present invention utilizes lipopolysaccharide solution and phenol solution as active ingredients of laying hen metritis modeling reagent, combines the advantages of pathogenic microorganism modeling and chemical reagent modeling, and reduces environmental pollution caused by pathogenic bacteria. Because both lipopolysaccharide solution and phenol solution have a synergistic effect, the typical inflammatory reaction of metritis in the uterus of laying hens is rapidly generated, and a strong systemic inflammatory reaction is avoided, and the side effect is small. The present invention can accurately grasp the modeling dosage, has high repeatability, is not easily affected by microbial toxicity and body immunity, and reduces the requirements for the test site. The laying hen metritis modeling method provided by the present invention is safe and reliable, has good repeatability, is simple to operate, is convenient to control, and reduces costs. BRIEF DESCRIPTION OF THE DRAWINGS

[0016] Figure 1 The photos showing the effects of different modeling agents on the cloaca of laying hens 48 hours after modeling in the test example are shown; wherein, A is a photo of the cloaca of laying hens in the blank control group. B is a photo of the cloaca of laying hens in the lipopolysaccharide solution group. C is a photo of the cloaca of laying hens in the phenol solution group. D is a photo of the cloaca of laying hens in Example 1. E is a photo of the cloaca of laying hens in Example 2. F is a photo of the cloaca of laying hens in group 3 of Example, G is a photo of the cloaca of laying hens in group 1 of comparative example. H is a photo of the cloaca of laying hens in group 2 of the comparative example. I is a photo of the cloaca of laying hens in group 3 of the comparative example, J is a photo of the cloaca of laying hens in group 4 of the comparative example; Figure 2The effect of different modeling agents on the appearance of eggs laid by laying hens after 48 hours of modeling in the test example is shown; wherein, A is a photo of the appearance of eggs laid by laying hens in the blank control group. B is a photo of the appearance of eggs laid by laying hens in the lipopolysaccharide solution group. C is a photo of the appearance of eggs laid by laying hens in the phenol solution group. D is a photo of the appearance of eggs laid by the laying hens in Example 1. E is a photo of the appearance of eggs laid by laying hens in group 2 of Example 2. F is a photo of the appearance of eggs laid by the laying hens in group 3 of Example, G is a photo of the appearance of eggs laid by the laying hens in group 1 of comparative example. H is a photo of the appearance of eggs laid by the laying hens in group 2 of the comparative example. I is a photo of the appearance of eggs laid by 3 groups of laying hens in the comparative example, J is a photo of the appearance of eggs laid by the laying hens in group 4 of the comparative example; Figure 3 The anatomical diagram of the uterus and vagina of the oviduct of laying hens by different modeling reagents after 48 hours of modeling in the test example is shown; wherein, A is the anatomical diagram of the uterus and vagina of laying hens in the blank control group. B is the anatomical diagram of the uterus and vagina of laying hens in the lipopolysaccharide solution group. C is the anatomical diagram of the uterus and vagina of laying hens in the phenol solution group. D is the anatomical diagram of the uterus and vagina of the laying hen in Example 1. E is a photo of the uterus and vagina of the laying hen in Example 2. F is a photo of the uterus and vagina of laying hens in Example 3. G is a photo of the uterus and vagina of laying hens in group 1 of comparative example. H is a photo of the uterus and vagina of laying hens in group 2 of the comparative example. I is a photo of the uterus and vagina of 3 groups of laying hens in comparative example, J is a photo of the uterus and vagina of laying hens in group 4 of the comparative example; Figure 4 The pathological diagrams of the blank group and Example 1 group on the oviduct uterus of laying hens after 48 hours of modeling are shown; wherein, A is the pathological picture of the uterus of laying hens in the blank control group. B is the anatomical diagram of the uterus and vagina of laying hens in the lipopolysaccharide solution group. C is the anatomical diagram of the uterus and vagina of laying hens in the phenol solution group. D is the pathological picture of the uterus of laying hens in Example 1. E is the pathological picture of the uterus of laying hens in Example 2. F is the pathological picture of the uterus of laying hens in group 3 of Example, G is the pathological picture of the uterus of laying hens in group 1 of comparative example. H is the pathological picture of the uterus of laying hens in group 2 of the comparative example. 1 is the pathological picture of uterus of laying hens in group 3 of comparative example, J is the pathological picture of the uterus of laying hens in group 4 of the comparative example. DETAILED DESCRIPTION

[0017] The following will be combined with the drawings in the embodiments of the present application to clearly describe the technical solutions in the embodiments of the present application. Obviously, the described embodiments are part of the embodiments of the present application, rather than all the embodiments. All other embodiments obtained by ordinary technicians in this field based on the embodiments in the present application belong to the scope of protection of this application.

[0018] The experimental methods in the following examples are conventional methods unless otherwise specified, and are carried out according to the techniques or conditions described in the literature in the art or according to the product instructions. The materials, reagents, etc. used in the following examples are commercially available unless otherwise specified. Any range described in the present invention includes any end value and any value between the end values ​​and any sub-range consisting of any value between the end values ​​or the end values. The percentages are all volume percentages unless otherwise specified, and the solutions are all aqueous solutions unless otherwise specified. Embodiment 1:

[0019] The present embodiment provides a laying hen metritis modeling agent, comprising the following components and composed in the following volume ratio: 3.7 mg / ml lipopolysaccharide solution: 25% phenol solution by volume = 1:1.

[0020] The phenol solution is composed of the following components by volume percentage: 25% liquefied phenol, 2.5% Tween 20, 2.5% Span 20, 10% glucose, 60% pH 7.2 phosphate buffer solution, wherein the concentration of the phosphate buffer solution is 0.01M.

[0021] The 3.7 mg / ml lipopolysaccharide solution is prepared by lipopolysaccharide and pH 7.2 phosphate buffer solution (PBS), and the concentration of the phosphate buffer solution is 0.01 M. The lipopolysaccharide is endotoxin extracted from Escherichia coli. The lipopolysaccharide in this example was purchased from Sigma-Aldrich (Shanghai) Trading Co., Ltd. and is derived from Escherichia coli 055:B5.

[0022] The preparation method of the laying hen metritis modeling agent of the present embodiment comprises the following steps: ①Prepare lipopolysaccharide solution: Heat the phosphate buffer solution to above 20°C, add lipopolysaccharide and stir until completely dissolved; ②Prepare phenol solution: Heat phenol to above 65°C. After it is completely melted into liquefied phenol, heat phosphate buffer solution to above 65°C, add liquefied phenol, Tween 20, Span 20, and glucose and stir until they are completely dissolved. ③ Mix the lipopolysaccharide solution prepared in step ① and the phenol solution prepared in step ② according to the proportion.

[0023] The method for modeling laying hen metritis using the laying hen metritis modeling agent described in this embodiment comprises the following steps: Before modeling, they were fed normally at 8:00 a.m. and 3:00 p.m. every day. When the egg production rate of laying hens stabilized at more than 88%, modeling began at 3:00 p.m. One person was responsible for fixing the laying hens, and they were inverted with the cloaca facing up and the head facing down. One hand gently pressed down both sides of the cloaca, and the other hand pushed up the hen's chest at the same time to turn the oviduct opening out and expose the oviduct opening of the laying hen. Another person put the modeling reagent into the oviduct uterus of the laying hen at a dosage of 2 ml per laying hen and observed for 48 hours. Embodiment 2:

[0024] This embodiment provides a modeling agent for uterine inflammation in laying hens, comprising the following components and in the following volume ratio: 3.7 mg / ml lipopolysaccharide solution: 25% phenol solution by volume = 1:1.5. The rest is the same as in embodiment 1. Embodiment 3:

[0025] This embodiment provides a modeling agent for uterine inflammation in laying hens, comprising the following components and in the following volume ratio: 3.7 mg / ml lipopolysaccharide solution: 25% phenol solution by volume = 1:0.5. The rest is the same as in embodiment 1. Comparative Example 1:

[0026] This comparative example provides a laying hen metritis modeling agent. The difference between this comparative example and Example 1 is that lipopolysaccharide is used instead of lipopolysaccharide solution, and the rest is the same as Example 1. Comparative Example 2:

[0027] This comparative example provides a laying hen metritis modeling agent, comprising the following components and in the following volume ratio: 3.7 mg / ml lipopolysaccharide solution: 25% phenol solution by volume = 1:5. The rest is the same as in Example 1. Comparative Example 3:

[0028] This comparative example provides a laying hen metritis modeling agent, comprising the following components and in the following volume ratio: 3.7 mg / ml lipopolysaccharide solution: 25% phenol solution by volume = 5:1. The rest is the same as in Example 1. Comparative Example 4:

[0029] This comparative example provides a laying hen metritis modeling agent. The difference between this comparative example and Example 1 is that the phenol solution is prepared by only liquefied phenol and PBS, and the volume concentration of the liquefied phenol is 25%. The rest is the same as Example 1.

[0030] Experimental Example 1: 1. Experimental animals: This experiment was conducted in the Ya'an Teaching and Research Park of Sichuan Agricultural University. 80 51-week-old Hy-Line laying hens were selected, each weighing about 1.8 kg.

[0031] 2. Grouping: 80 Hy-Line grey laying hens were randomly divided into 10 groups, each with 4 replicates and 2 hens in each replicate, including 8 hens in the blank control group (PBS), 8 hens in the lipopolysaccharide solution group, 8 hens in the phenol solution group, 8 hens in the Example 1-3 group and 8 hens in the Comparative Example 1-4 group, all of which were modeled using the operating method for modeling laying hen metritis described in Example 1 of the present invention.

[0032] Blank control group: Phosphate buffer solution with a pH value of 7.2 and a concentration of 0.01 M was used as a modeling agent for modeling; Lipopolysaccharide solution group: A 1.85 mg / ml lipopolysaccharide solution was used as a modeling agent for modeling; the 1.85 mg / ml lipopolysaccharide solution was prepared by lipopolysaccharide and pH 7.2 phosphate buffer solution (PBS), and the concentration of the phosphate buffer solution was 0.01M.

[0033] Phenol solution group: the phenol solution with a volume concentration of 25% described in Example 1 was used as a modeling agent for modeling; Example 1 group: Modeling was performed using the modeling agent prepared in Example 1; Example 2 group: Modeling was performed using the modeling agent prepared in Example 2; Example 3 group: Modeling was performed using the modeling agent prepared in Example 3; Comparative Example 1: Modeling was performed using the modeling agent prepared in Comparative Example 1; Comparative Example 2: The modeling agent prepared in Comparative Example 2 was used for modeling; Comparative Example 3: The modeling agent prepared in Comparative Example 3 was used for modeling; Comparative Example 4: The modeling agent prepared in Comparative Example 4 was used for modeling.

[0034] 48 hours after modeling, the cloaca, egg appearance, vaginal part and uterine part of the laying hens were photographed, and the clinical manifestations were compared among the 10 treatment groups.

[0035] 48 hours after modeling, one laying hen was selected from each replicate, and a total of 10 laying hens were killed by jugular vein bleeding, and the oviduct and uterus were taken for pathological analysis. The results are shown in Tables 1 and 2.

[0036] 48 hours after modeling, blood was collected from the jugular vein of laying hens in each group to prepare serum for detecting the contents of cytokines IL-1β, IL-6 and TNF-α. The test results are shown in Table 3.

[0037] Results Analysis 1. Cloaca Depend on Figure 1 It can be seen that: 48 hours after modeling, the laying hens in the blank control group had no secretions at the cloaca, and the surrounding feathers were clean and free of feces; the laying hens in the lipopolysaccharide solution group had no secretions at the cloaca; the laying hens in the phenol solution group had no secretions near the cloaca; the laying hens in the Example 1-3 groups all had a large amount of secretions at the cloaca and were stained with feces, among which the group in Example 1 had the most secretions and was stained with feces, polluting the surrounding feathers.

[0038] In the comparative example 1 group, only a small amount of feces was stained on the feathers near the cloaca of the laying hens. After the lipopolysaccharide solution was replaced by lipopolysaccharide, not only the cost was greatly increased, but also the modeling effect of the modeling agent was reduced. This is because the lipopolysaccharide solution with a concentration of 3.5-4 mg / ml and the phenol solution have a synergistic effect. After the lipopolysaccharide solution with a concentration of 3.5-4 mg / ml was replaced by lipopolysaccharide, the synergistic effect disappeared, resulting in a decrease in the modeling effect of the modeling agent.

[0039] In the comparison groups 2 and 3, there was only a small amount of feces on the feathers near the cloaca opening of the laying hens. This is because when the proportion of phenol solution is too high and the proportion of lipopolysaccharide solution is reduced, the laying hens will be over-stressed and the mortality rate will increase; if the phenol solution is too low and the lipopolysaccharide solution increases, the incidence of salpingitis in laying hens will be reduced, resulting in modeling failure.

[0040] Only a small amount of feces was stained on the feathers near the cloaca of the laying hens in the comparative example 4 group. This is because the phenol solution prepared by Tween 20, Span 20, glucose, and liquefied phenol and the lipopolysaccharide solution have a synergistic effect. Tween 20, Span 20, glucose and lipopolysaccharide solution can alleviate the corrosive effect of liquefied phenol, evenly disperse the phenol, and prevent the local phenol concentration from being too high and the corrosive effect from being too strong. It can also prevent the modeling preparation from flowing back. When Tween 20, Span 20 and glucose are removed, the synergistic effect disappears, resulting in a decrease in the modeling effect of the modeling agent.

[0041] 2. Egg appearance Depend on Figure 2It can be seen that 48 hours after modeling, the eggs in the blank control group had a smooth and clean appearance; the eggs in the lipopolysaccharide solution group had a smooth and clean appearance; the eggs in the phenol solution group had clean shells; a large amount of blood stains appeared in the shells of the eggs in the Example 1-3 groups, especially in the Example 1 group. Among the eggs in the Comparative Examples 1-4 groups, the eggs in the Comparative Examples 1 and 2 had a little blood stain on their shells, and the eggs in the Comparative Examples 3 and 4 had a clean appearance.

[0042] 3. Anatomy of the uterus and vagina Depend on Figure 3 It can be seen that: 48 hours after modeling, the mucosa of the uterus and vagina of the laying hens in the blank control group was normal, without symptoms such as bleeding, congestion, and necrosis; the laying hens in the lipopolysaccharide solution group had a strong systemic inflammatory response, the mucosa of the vagina was normal, and the mucosa of the uterus showed congestion symptoms; the laying hens in the phenol solution group had a strong systemic inflammatory response, protein denaturation appeared in the mucosa of the vagina, and the mucosa of the uterus was normal; the groups of Examples 1-3 all had typical uterine inflammatory reactions only in the uterus, and no strong systemic inflammatory reactions occurred. Among them, the laying hens in the Example 1 group had serious lesions in their reproductive organs, protein denaturation appeared in the vaginal mucosa, severe bleeding and edema, the uterine mucosa was completely covered with white calcified particles, and there were sticky white secretions and cheese-like masses in the uterus, and the inflammatory reaction was obvious. The laying hens in the Example 2 and 3 groups had lesions in their reproductive organs, protein denaturation, bleeding and edema appeared in the vaginal mucosa, and the uterine mucosa showed symptoms of congestion and redness. The vaginal mucosa of comparison groups 1 and 2 showed protein denaturation, while the uterine mucosa was normal without congestion and necrosis symptoms; the vaginal mucosa of comparison group 3 was normal, while the uterine mucosa had congestion symptoms; the vaginal mucosa of comparison group 4 was normal, while the uterine mucosa showed congestion symptoms.

[0043] 4. Uterine pathology score Table 1 Analysis of pathological scores of laying hens' uterus by different modeling reagents

[0044] Data in the same column with different letters on their shoulders indicate significant differences (P<0.05).

[0045] The uterine pathology scoring standards for laying hens in Table 1 are shown in Table 2: Table 2:

[0046] As shown in Tables 1 and 2, different modeling agents have significant differences in the pathological scores of the uterus of laying hens. As shown in Tables 1 and 2, the modeling agent of Example 1 has the best effect.

[0047] 5. Cytokines in serum As shown in Table 3, 48 hours after modeling, compared with the blank control group, the content of cytokine TNF-α in the serum of laying hens in the lipopolysaccharide solution group increased significantly, the content of cytokines IL-1β, IL-6 and TNF-α in the serum of laying hens in the phenol solution group increased significantly, and the content of cytokines IL-1β, IL-6 and TNF-α in the serum of laying hens in the Example 1-3 groups increased significantly, especially the content of cytokines IL-1β, IL-6 and TNF-α in the serum of laying hens in the Example 1 group was the highest. The content of cytokines IL-1β, IL-6 and TNF-α in the serum of laying hens in the Comparative Examples 1-4 groups also increased, but the increase was much lower than that in the examples of the present application.

[0048] 6. Cytokines in the Uterus As shown in Table 4, 48 hours after modeling, compared with the blank control group, the content of cytokine TNF-α in the uterus of laying hens in the lipopolysaccharide solution group increased significantly, the content of cytokine TNF-α in the uterus of laying hens in the phenol solution group increased significantly, and the content of cytokines IL-1β, IL-6 and TNF-α in the uterus of laying hens in Example 1-3 groups increased significantly, especially the content of cytokines IL-1β, IL-6 and TNF-α in the uterus of laying hens in Example 1 group was the highest. The content of cytokines IL-1β, IL-6 and TNF-α in the uterus of laying hens in Comparative Examples 1-4 groups also increased, but the increase was much lower than that in the examples of the present application.

[0049] 6. Uterine pathology Depend on Figure 4 It can be seen that: 48 hours after modeling, the uterine mucosal structure of the blank control group was normal; the uterine mucosal structure of the lipopolysaccharide solution group was normal, with a small amount of glandular atrophy; the uterine mucosal structure of the phenol solution group was slightly damaged, with a small amount of vacuoles; the uterine mucosal structure of the Example 1 group was largely destroyed, the mucosal top of the uterine part narrowed, the glands atrophied, and a large number of vacuoles appeared. The uterine mucosal structure of the Example 2 and 3 groups was destroyed, the mucosal top of the uterine part narrowed, some glands atrophied, and some vacuoles appeared; the uterine mucosal structure of the comparative example 1 and 2 groups was slightly destroyed, with a small amount of vacuoles and a small amount of glandular atrophy; the uterine mucosal structure of the comparative example 3 and 4 groups was normal, with some vacuoles.

[0050] 7. Conclusion Based on the appearance of the cloaca, eggs, uterus and vagina, as well as the results of serum cytokines and uterus cytokines, the modeling reagent of Example 1, which is composed of 3.7 mg / ml lipopolysaccharide and 25% phenol mixed in a volume ratio of 1:1, caused uterine inflammation in laying hens, and the inflammatory symptoms were as follows: 1) A large amount of secretions appeared at the cloaca of laying hens, contaminating the surrounding feathers, and blood stains appeared on the egg shells; 2) Protein denaturation, bleeding and edema appeared in the vaginal mucosa of laying hens, and there were sticky white secretions and cheese-like masses in the uterus, with obvious inflammatory reactions; 3) The levels of inflammatory factors IL-1β, IL-6 and TNF-α in serum and uterus increased significantly. The inflammatory cytokine content of Example 1 was the highest and the inflammatory effect was the most obvious, which was presumably related to the composition of the phenol solution and the lipopolysaccharide solution and the ratio of the two. Phenol solution and lipopolysaccharide solution are used in combination in a specific ratio. Phenol solution can destroy the mucous membrane of the oviduct to help the lipopolysaccharide solution enter the blood. Once the lipopolysaccharide solution enters the body's blood, it can pass through multiple signal transduction pathways, ultimately causing the release of inflammatory cytokines such as IL-1β, IL-6, TNF-α, etc., causing an inflammatory response in the oviduct and increasing the success rate of typical inflammatory symptoms in the oviduct of laying hens.

[0051] Table 3 Serum cytokine levels of laying hens in each group 48 hours after modeling:

[0052] Data in the same column with different letters on their shoulders indicate significant differences (P<0.05).

[0053] Table 4 Test example 48 hours after modeling, the content of cytokines in the oviduct uterus of laying hens in each group:

[0054] Data in the same column with different letters on their shoulders indicate significant differences (P<0.05).

[0055] The embodiments of the present application are described above in conjunction with the accompanying drawings, but the present application is not limited to the above-mentioned specific implementation methods. The above-mentioned specific implementation methods are merely illustrative and not restrictive. Under the guidance of the present application, ordinary technicians in this field can also make many forms without departing from the purpose of the present application and the scope of protection of the claims, all of which are within the protection of the present application.

Claims

1. A laying hen metritis modeling agent, characterized in that: The composition includes the following components and is composed according to the following volume ratio: lipopolysaccharide solution: phenol solution = (0.5-1.5): (0.5-1.5).

2. The laying hen metritis modeling agent according to claim 1, characterized in that: The volume concentration of liquefied phenol in the phenol solution is 20-30%, preferably 25%.

3. The laying hen metritis modeling agent according to claim 2, characterized in that: The phenol solution consists of the following components by volume percentage: 20-30% liquefied phenol, 2-3% Tween 20, 2-3% Span 20, 9-11% glucose, and 55-65% phosphate buffer solution.

4. The laying hen metritis modeling agent according to claim 3, characterized in that: The phenol solution is composed of the following components by volume percentage: 25% liquefied phenol, 2.5% Tween 20, 2.5% Span 20, 10% glucose, 60% phosphate buffer solution.

5. The laying hen metritis modeling agent according to claim 1, characterized in that: The lipopolysaccharide solution is prepared from lipopolysaccharide and phosphate buffer solution, and the concentration of the lipopolysaccharide is 3.5-4 mg / ml, preferably 3.7 mg / ml.

6. The laying hen metritis modeling agent according to any one of claims 3 to 5, characterized in that: The pH value of the phosphate buffer solution is 7.0-7.4, preferably pH 7.

2.

7. The method for preparing the laying hen metritis modeling agent according to claims 1-6, characterized in that: The following steps are involved: ①Prepare lipopolysaccharide solution: Heat the phosphate buffer solution, add lipopolysaccharide and stir until it is completely dissolved; ②Preparing phenol solution: After heating phenol until it is completely melted into liquefied phenol, heat the phosphate buffer solution, add the liquefied phenol, Tween 20, Span 20, and glucose, and stir until they are completely dissolved; ③ Evenly mix the lipopolysaccharide solution prepared in step ① and the phenol solution prepared in step ② according to the proportion.

8. Use of the laying hen metritis modeling agent according to any one of claims 1 to 6 in preparing a laying hen metritis model.

9. A method for modeling metritis in laying hens, characterized in that: The following steps are involved: The oviduct opening of the laying hen is exposed, and the laying hen metritis modeling agent according to any one of claims 1 to 6 is placed into the uterus of the oviduct of the laying hen.

10. The method for modeling of metritis in laying hens according to claim 9, characterized in that: The dosage of the laying hen metritis modeling agent is 1-3 ml / laying hen, preferably 2 ml / laying hen.

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