Giant peptiformis and its application in the prevention and treatment of intestinal inflammation or liver injury

Abstract

This invention discloses a *Macrococcus escherichiae* strain and its application in the prevention and treatment of intestinal inflammation and liver injury. The microbial preservation number of *Macrococcus escherichiae* provided by this invention is CGMCC No. 46926. The *Macrococcus escherichiae* strain provided by this invention exhibits good biocompatibility, acid and bile salt tolerance, and the ability to produce short-chain fatty acids. Through multiple mechanisms such as regulating intestinal flora structure, enhancing intestinal barrier function, inhibiting pro-inflammatory factor expression, promoting anti-inflammatory factor production, and alleviating oxidative stress, it significantly improves intestinal inflammatory symptoms such as DSS-induced weight loss and diarrhea, reduces the disease activity index, restores colon length, regulates inflammatory factor expression, and has a protective effect on the gut-liver axis. As a microbial preparation or feed additive, it shows promise for promoting animal growth and preventing or improving intestinal inflammation, hepatitis, liver injury, or spleen injury.

Description

Technical Field

[0001] This invention relates to a strain of intestinal probiotic and its application, particularly to a strain of Giant Escherichia coli and its application in the preparation of drugs or feed additives for the prevention and treatment of intestinal inflammation, hepatitis or liver damage, belonging to the field of Giant Escherichia coli ME1 and its application. Background Technology

[0002] In recent years, with the deepening of research on gut microbiota, the role of probiotics in maintaining gut health and preventing inflammatory diseases has received increasing attention. Probiotics have shown application potential in improving gut inflammation through multiple mechanisms, such as regulating gut microbiota structure, enhancing intestinal barrier function, and modulating immune responses.

[0003] Giant cocci (E. esculenta) Megasphaera elsdenii *Macrococcus escherichiae*, belonging to the Firmicutes phylum, is a Gram-negative anaerobic coccus naturally found in the intestines of ruminants and humans, and possesses the ability to produce metabolites such as short-chain fatty acids. Chinese patent CN110327375A discloses the application of *Macrococcus escherichiae* in the preparation of preparations that lower total cholesterol and / or low-density lipoprotein cholesterol, revealing the bacterium's function in lipid metabolism regulation. However, this patent only focuses on the lipid-lowering effect of *Macrococcus escherichiae* and does not address its application in the prevention and treatment of intestinal inflammation.

[0004] Chinese patent CN115873763A discloses a bacterial agent that alleviates intestinal inflammation by regulating intestinal serotonin homeostasis. This agent includes *Lactobacillus mucosa*, and may further include *Megacoccus echocerata* or *Lactobacillus amyloliquefaciens*. While this patent mentions the application of *Megacoccus echocerata* in a compound bacterial agent, it primarily focuses on the serotonin homeostasis regulation mechanism, with *Megacoccus echocerata* only as an optional component. It lacks systematic research on the efficacy of a single strain of *Megacoccus echocerata* in preventing and treating intestinal inflammation, and a clear elucidation of its mechanism of action.

[0005] While existing research has focused on the application of probiotics in the prevention and treatment of intestinal inflammation, the following shortcomings remain: First, there is a lack of systematic research on the prevention and treatment of intestinal inflammation using a single strain of *M. esculenta*. Existing patents either use *M. esculenta* as an optional component of compound bacterial agents or only focus on its applications in other areas such as lipid metabolism, failing to fully reveal its independent role and application value in the prevention and treatment of intestinal inflammation. Second, existing technologies have not revealed the protective effect of *M. esculenta* on the gut-liver axis, while intestinal inflammation is often accompanied by impaired intestinal barrier function, leading to the entry of harmful substances such as endotoxins into the bloodstream and causing secondary liver damage. There is a lack of systematic research and prevention and treatment strategies for this pathological mechanism. Third, existing technologies lack a comprehensive explanation of the mechanism of action of *M. esculenta* in preventing and treating intestinal inflammation, including insufficient systematic research data supporting multi-dimensional mechanisms such as gut microbiota regulation, enhanced intestinal barrier function, regulation of inflammatory factor expression, and oxidative stress relief.

[0006] Therefore, it is necessary to provide a strain of *Gastrococcus escherichiae* and systematically study its application in the prevention and treatment of intestinal inflammation, clarify its mechanism of action, and reveal its protective effect on the gut-liver axis, so as to provide a new biological therapy strategy for the prevention and treatment of intestinal inflammatory diseases. Summary of the Invention

[0007] Given the lack of systematic research on the prevention and treatment of intestinal inflammation by a single strain of *Gastrococcus escherichiae* in the existing technology, the failure to reveal the protective effect of *Gastrococcus escherichiae* on the gut-liver axis, and the lack of a comprehensive explanation of its mechanism of action in preventing and treating intestinal inflammation, the main objective of this invention is to provide *Gastrococcus escherichiae* and its application in the preparation of drugs or feed additives for the prevention and treatment of intestinal inflammation, hepatitis, or liver damage.

[0008] To achieve the above objectives, the present invention adopts the following technical solution: This invention first provides a strain of *Gastrococcus ehrlich* ME1, with accession number CGMCC No. 46926, and its classification name is: *Gastrococcus ehrlich*. Megasphaera elsdenii The deposit date is February 6, 2026; the depositary institution is the China General Microbiological Culture Collection Center; the depositary address is Institute of Microbiology, Chinese Academy of Sciences, No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing.

[0009] The 16S rDNA sequence of *M. esculenta* ME1 described in this invention has ≥99% homology with the *M. esculenta* reference sequence in the NCBI database. The accession number of the reference sequence is selected from any one of CP027569.1, CP027570.1, AP031433.1, HE576794.1, and NR_102980.1. The *M. esculenta* ME1 is γ-hemolytic and has good biocompatibility.

[0010] The logarithmic growth phase of *Gastrococcus ehrlich* ME1 described in this invention is 6-35 h after inoculation, and the plateau phase is 30-72 h after inoculation; *Gastrococcus ehrlich* ME1 has the ability to produce short-chain fatty acids, wherein the main products of the short-chain fatty acids include propionic acid and butyric acid.

[0011] The survival rate of *Gastrococcus ehrlich* ME1 described in this invention is ≥80% under pH 3.0-5.0 conditions; and ≥70% under a bile salt concentration of 0.3%-0.5%.

[0012] The *Gastrococcus ehrlich* ME1 strain described in this invention can significantly reduce the expression of pro-inflammatory factors IL-6, IL-17, IL-1β, and TNF-α, and increase the expression of anti-inflammatory factor IL-10.

[0013] The giant cocci ME1 provided by this invention mainly possesses the following pharmacological activities or functions: (1) Promotes animal growth or improves diarrhea: In the DSS-induced mouse intestinal inflammation model, Escherichia coli ME1 significantly improved weight loss and diarrhea symptoms. The DAI score of the M+D group was significantly lower than that of the DSS group (P<0.05); the colon length was significantly longer than that of the DSS group (P<0.05), recovering to more than 85% of the normal level; the colon pathology score was significantly lower than that of the DSS group (P<0.05), and the integrity of the crypt structure was well preserved.

[0014] (2) Enhanced intestinal barrier function: ME1 of giant cocci Ehrlich significantly upregulated the expression of colonic tight junction proteins ZO-1, Occludin, and Claudin-1 (P<0.01); serum LPS and DAO levels in the M+D group were significantly lower than those in the DSS group (P<0.01), indicating that intestinal permeability was reduced and barrier function was enhanced.

[0015] (3) Regulation of inflammatory factor expression: Reduce the expression levels of pro-inflammatory factors IL-6, IL-17, IL-1β and TNF-α by more than 60%; increase the expression level of anti-inflammatory factor IL-10 by more than 80%.

[0016] (4) Relief of oxidative stress: The levels of MDA in the colon and serum of the M+D group were significantly lower than those in the DSS group, while the SOD enzyme activity was significantly higher than that in the DSS group (P<0.05), indicating that the oxidative stress was effectively relieved.

[0017] (5) Improves the structure of intestinal flora and increases the relative abundance of beneficial bacteria: ME1 of giant cocci Eubacterium significantly increases the relative abundance of beneficial bacteria such as g_Lactobacillus, g_Bifidobacterium, g_Butyricimonas, g_Faecalibaculum, and g_Eubacterium_B, while decreasing the relative abundance of potentially pathogenic bacteria such as Proteobacteria and g_Bilophila.

[0018] (6) Protective effect of gut-liver axis: Reduced endotoxin entry into the blood, effective protection of gut-liver axis, reduction of liver index, improvement of liver pathology score, and reduction of liver pro-inflammatory factor level.

[0019] (7) Improve spleen inflammation: Escherichia coli ME1 significantly improves the pathological structural disorder of the spleen caused by DSS.

[0020] Therefore, the *Gastrococcus ehrlich* ME1 provided by this invention mainly has the following applications: (1) It is added to the diet of livestock and poultry as a feed additive to promote the growth of livestock and poultry.

[0021] (2) Used as a microbial preparation for the prevention and treatment of diarrhea or as a feed additive to prevent or alleviate diarrhea in livestock and poultry.

[0022] (3) As a microbial preparation for the prevention and treatment of enteritis or colitis in livestock and poultry, or as a feed additive added to the diet of livestock and poultry to prevent and treat enteritis or colitis in livestock and poultry; the prevention and treatment or relief of enteritis or colitis in livestock and poultry is achieved through at least one of the following mechanisms: regulating the structure of intestinal flora, enhancing intestinal barrier function, inhibiting the expression of pro-inflammatory factors, promoting anti-inflammatory factors, relieving oxidative stress, and protecting the integrity of intestinal mucosa.

[0023] (4) As a microbial preparation for the prevention and treatment of hepatitis or liver injury, or as a feed additive added to the diet of livestock and poultry to prevent and treat hepatitis or liver injury in livestock and poultry; the above is achieved through at least one of the following mechanisms: improving intestinal inflammation, reducing intestinal permeability, reducing endotoxin entry into the blood, inhibiting the expression of liver inflammatory factors, reducing liver oxidative stress, and protecting the integrity of liver tissue structure.

[0024] (5) Microbial preparations for improving intestinal flora or increasing the types or number of beneficial microorganisms in the intestine, or added as feed additives to the diets of livestock and poultry to improve intestinal flora or increase the types or number of beneficial microorganisms in the intestines of livestock and poultry; the beneficial microorganisms are selected from one or more of Lactobacillus, Bifidobacterium, Vibrio butyricum, Faecalibacterium or Eubacterium.

[0025] (6) Used as a microbial preparation to protect the spleen or added as a feed additive to the diet of livestock and poultry to improve spleen inflammation or damage.

[0026] Those skilled in the art can prepare clinically suitable microbial preparations from *Gastrococcus ehrlich* ME1 using conventional methods for preparing microbial preparations in the art; preferably, the microbial preparation can be one or more of the following: oral liquid, capsules, tablets, granules, enema, or microcapsule formulation. The administration route of the microbial preparation can be oral, enema, or enterotargeted delivery.

[0027] The viable concentration of *Gastrococcus eherae* ME1 in the microbial preparation or feed additive can be 1 × 10⁻⁶. 7 -1×10 9 CFU / mL, preferably 1×10⁻⁶ 8 CFU / mL.

[0028] The administration method of the microbial preparation is determined based on factors such as body weight, disease severity, and age. For reference, the daily dose of *Gastrococcus ehrlich* ME1 is 1 × 10⁻⁶. 7 -1×10 9 CFU / kg body weight, more preferably 1×108 CFU / kg body weight; preferably, the course of treatment is 7-21 days.

[0029] The amount of *Gastrococcus ehrlich* ME1 added as a feed additive to livestock and poultry diets can be determined according to the circumstances. For reference, it can be added to livestock and poultry diets at a rate of 0.1-5 wt%.

[0030] The giant cocci ME1 provided by this invention has good acid and bile salt resistance, with a survival rate of ≥80% under pH 3.0-5.0 conditions and a survival rate of ≥70% under 0.3%-0.5% bile salt concentration. This ensures that after oral administration, it can pass through the gastric acid and bile salt environment to reach the intestine and exert its effects, thereby improving bioavailability and practical application value.

[0031] This invention systematically studies the application of Giant Ehrlich aspergillus ME1 in the prevention and treatment of intestinal inflammation. Based on the DSS-induced mouse intestinal inflammation model, it is demonstrated that this strain can significantly improve weight loss and diarrhea symptoms, reduce the Disease Activity Index (DAI) score by more than 50%, and restore the colon length to more than 85% of the normal level.

[0032] This invention reveals the multidimensional mechanism of action of *Gastrococcus ehrlich* ME1 in preventing and treating intestinal inflammation, including regulating the intestinal flora structure (significantly increasing the relative abundance of beneficial bacteria such as *Lactobacillus* and *Bifidobacterium*), enhancing intestinal barrier function (significantly upregulating the expression of tight junction proteins ZO-1, Occludin, and Claudin-1, and reducing serum LPS and DAO levels), regulating the expression of inflammatory factors (significantly reducing the levels of pro-inflammatory factors IL-6, IL-17, IL-1β, and TNF-α by more than 60%, and significantly increasing the level of anti-inflammatory factor IL-10 by more than 80%), and alleviating oxidative stress (significantly reducing MDA levels and increasing SOD enzyme activity).

[0033] This invention reveals for the first time the protective effect of *Gastrococcus ehrlich* ME1 on the gut-liver axis, demonstrating that this strain can effectively reduce secondary liver injury caused by intestinal inflammation, reduce the liver histological activity index (HAI) score by more than 70%, significantly reduce the level of liver pro-inflammatory factors, and protect the integrity of liver tissue structure, providing a new prevention and treatment strategy for liver injury associated with intestinal inflammatory diseases.

[0034] In summary, the *Gastrococcus ehrlich* ME1 provided by this invention has good biocompatibility, acid and bile salt tolerance, and the ability to produce short-chain fatty acids. Through multiple mechanisms such as regulating intestinal flora structure, enhancing intestinal barrier function, inhibiting pro-inflammatory factor expression, promoting anti-inflammatory factor production, and alleviating oxidative stress, it has promising applications as a microbial preparation or feed additive in promoting animal growth and preventing or improving intestinal inflammation, hepatitis, liver injury, or spleen injury. Attached Figure Description

[0035] Figure 1 This is a growth curve of Ehrlich giant cocci ME1.

[0036] Figure 2 The image shows the results of the hemolytic characteristics test of Ehrlich giant cocci ME1.

[0037] Figure 3 The figure shows the effect of Ehrlich giant cocci ME1 on the growth performance of DSS-induced mice.

[0038] Figure 4 The figure shows the effect of Ehrlich giant cocci ME1 on DSS-induced fecal occult blood in mice.

[0039] Figure 5 The figure shows the effect of Ehrlich giant cocci ME1 on DSS-induced organ indices in mice.

[0040] Figure 6 The figure shows the effect of Escherichia coli ME1 on the colon length measurement results of DSS-treated mice.

[0041] Figure 7 The effect of Escherichia coli ME1 on the pathological observation and scoring of the colon in DSS-treated mice is shown in the figure. The red arrows in the figure represent atrophy of colonic mucosal epithelial cells; the yellow arrows represent destruction or absence of colonic mucosal crypts; the green arrows represent extensive inflammatory cell infiltration and fibrous tissue hyperplasia in the lamina propria of the colonic mucosa; and the brown arrows represent inflammatory cell infiltration and fibrous tissue hyperplasia in the submucosa of the colon.

[0042] Figure 8 The effect of Ehrlich giant cocci ME1 on the morphological observation and measurement results of DSS-induced mouse jejunum.

[0043] Figure 9 The effect of Ehrlich giant cocci ME1 on DSS-induced colonic inflammatory factors in mice.

[0044] Figure 10 The effects of Ehrlich giant cocci ME1 on the intestinal barrier and permeability of DSS-induced mice.

[0045] Figure 11 The effects of Ehrlich giant cocci ME1 on DSS-induced oxidative stress markers in the colon and serum of mice.

[0046] Figure 12 The effect of Ehrlich giant cocci ME1 on DSS-induced cecal microbial diversity in mice.

[0047] Figure 13 The figure shows the effect of Ehrlich giant cocci ME1 on the composition of the cecal microbiota in mice.

[0048] Figure 14 The effect of Ehrlich's giant cocci ME1 on DSS-induced liver pathological sections, liver tissue necrosis, and inflammation scores in mice is shown. Black arrows indicate inflammatory cell infiltration around the portal area; red arrows indicate portal enlargement with localized debris-like necrosis; yellow arrows indicate sinusoidal dilatation with inflammatory cell infiltration within the sinusoidal spaces.

[0049] Figure 15 The effect of Ehrlich giant cocci ME1 on inflammatory factors in the liver of DSS-induced mice.

[0050] Figure 16 The effect of Ehrlich's giant cocci ME1 on the histopathology of spleen tissue in DSS-induced mice. Black arrows represent splenic corpuscles; yellow arrows represent the central artery; green arrows represent the periarterial lymphatic sheath; orange arrows represent splenic cord lymphocytes; red arrows represent numerous neutrophils in the medullary sinus; and brown arrows represent a small number of neutrophils visible in the medullary sinus. Detailed Implementation

[0051] The present invention will be further described below with reference to specific embodiments, and the advantages and features of the present invention will become clearer with the description. However, it should be understood that the embodiments described are merely exemplary and do not constitute any limitation on the scope of the present invention. Those skilled in the art should understand that modifications or substitutions can be made to the details and form of the technical solutions of the present invention without departing from the spirit and scope of the present invention, but such modifications or substitutions all fall within the protection scope of the present invention.

[0052] Example 1: Isolation, purification, identification, and evaluation of biological characteristics of *Gastrococcus ehrlich* ME1 1. Isolation and purification of strains Using sterile forceps, take 1 g of feces from weaned piglets and add it to 9 mL of sterile physiological saline buffer. Mix well and perform serial dilution. Take samples of each dilution factor of 10... -5 10 -6 10 -7 100 μL of each bacterial culture was spread onto RCM (Reinforced Clostridial Medium) solid medium and incubated at 37 ℃ for 24 h in an anaerobic workstation. Colonies with larger size and stronger reproductive capacity were selected and streaked onto RCM solid medium at least three times to obtain purified strains.

[0053] 2. Molecular biological identification DNA from the bacterial strain was extracted and purified according to the instructions of the bacterial genomic DNA extraction kit. PCR amplification was performed using universal primers for bacterial 16S rDNA: the upstream primer 27F sequence was 5'-AGAGTTTGATCCTGGCTCAG-3', and the downstream primer 1492R sequence was 5'-GGTTACCTTGTTACGACTT-3'.

[0054] The PCR amplification system consisted of 50 μL, including: 25 μL of 2×Taq PCR Master Mix, 2 μL of upstream primer (10 μM), 2 μL of downstream primer (10 μM), 2 μL of DNA template, and 19 μL of ddH2O. The PCR amplification program was as follows: 94 ℃ pre-denaturation for 5 min; 94 ℃ denaturation for 30 s, 55 ℃ annealing for 30 s, and 72 ℃ extension for 90 s, for a total of 35 cycles; and a final extension at 72 ℃ for 10 min.

[0055] The PCR products were identified by 2% agarose gel electrophoresis, yielding a band of approximately 1500 bp. Sequencing of the PCR products and BLAST alignment of the sequencing results with the NCBI database showed that the 16S rDNA sequence of this strain shared over 99% homology with the reference sequence of *Megasphaera elsdenii* in the NCBI database. The accession numbers for similar sequences were CP027569.1, CP027570.1, AP031433.1, HE576794.1, and NR_102980.1. Therefore, this strain was identified as *Megasphaera elsdenii* and named *Megasphaera elsdenii* ME1.

[0056] 3. Growth curve determination Anaerobic RCM liquid medium was added to each well of a 96-well plate at a rate of 180 μL. The experimental group was inoculated with 20 μL of *Gastrococcus ehrlich* ME1 culture activated for 24 h per well. A blank control group containing only 200 μL of medium was also included. Each group had three replicates. After mixing under anaerobic conditions, 100 μL of paraffin oil was added for sealing. The 96-well plate was then placed in a growth curve analyzer and incubated at 37 ℃. OD was measured every 2 h. 600 Value, total culture time 72 h.

[0057] Growth curve measurement results are as follows Figure 1 As shown, the strain continued to grow after inoculation, with the fastest growth rate after 8 hours, entering the logarithmic growth phase. After 30 hours, the growth of the strain slowed down, entering the plateau phase. According to the growth curve data, the logarithmic growth phase of *Gastrococcus ehrlich* ME1 was 8-30 hours after inoculation, and the plateau phase was 30-72 hours after inoculation.

[0058] 4. Hemolysis test The activated *Gastrococcus ehrlich* ME1 strain, after two generations of incubation, was streaked onto Columbia blood agar medium and incubated at 37°C for 48 h in an anaerobic workstation; the hemolysis test results were as follows: Figure 2 As shown, no hemolytic zone appeared around the colony of *Gastrococcus ehrlich* ME1, indicating that it is a γ-hemolytic type, which means that this strain does not have hemolytic activity and has good biosafety.

[0059] 5. Viable Bacterial Count Determination Inside the anaerobic workstation, primary strains of *Gastrococcus ehrlich* ME1 were inoculated into RCM liquid anaerobic medium and anaerobic incubated for 48 h, followed by secondary subculturing for 24 h. The secondary bacterial culture was then serially diluted, with 10-10 fractions taken from each fraction. -5 10 -6 10 -7 100 μL of each bacterial culture was spread onto RCM solid medium, with three replicates for each gradient. After anaerobic incubation, plates with colony counts between 30 and 300 were selected for statistical analysis to determine the average colony count.

[0060] In 10 -5 On gradient plates, the number of single colonies of *Gastrococcus ehrlich* ME1 was approximately 100. Based on the dilution factor and plating volume, the final viable cell concentration of the bacterial solution was determined to be 1 × 10⁻⁶. 8 CFU / mL.

[0061] 6. Preparation of bacterial culture Giant Ehrlich aspergillus ME1 was inoculated onto RCM anaerobic medium and cultured in an anaerobic incubator for 48 h. After subculturing for 24 h, the bacterial suspension was collected. The suspension was washed twice with anaerobic sterile distilled water to obtain a viable bacterial concentration of 1 × 10⁻⁶. 8 A bacterial suspension of CFU / mL was used for subsequent animal experiments.

[0062] 7. Acid resistance test Giant Ehrlich. megaterium ME1 bacterial suspension was adjusted to pH 3.0, 3.5, 4.0, 4.5, and 5.0 with sterile HCl solution, with three replicates for each pH gradient. After anaerobic incubation at 37 °C for 2 h, the viable cell count was determined using the plate count method, and the survival rate was calculated.

[0063] Survival rate (%) = (number of viable bacteria after treatment / number of viable bacteria before treatment) × 100%.

[0064] The experimental results showed that the survival rate of *Macrococcus eherii* ME1 was 82.5±3.2% at pH 3.0, 85.7±2.8% at pH 3.5, 88.3±2.1% at pH 4.0, 90.6±1.9% at pH 4.5, and 92.4±1.5% at pH 5.0. The survival rate of *Macrococcus eherii* ME1 was ≥80% at pH 3.0–5.0, indicating that this ME1 strain has good acid resistance and can survive the acidic environment of the stomach.

[0065] 8. Bile salt tolerance test Oxgall salts (0.3%, 0.4%, and 0.5%) were added to RCM liquid medium, with three replicates for each concentration gradient. After inoculation with *Gastrococcus ehrlich* ME1, the culture was anaerobic at 37 °C for 24 h. Viable cell counts were determined using the plate count method, with the medium without added oxgall salts serving as a control. Survival rates were calculated.

[0066] Survival rate (%) = (number of viable bacteria in the bile salt treatment group / number of viable bacteria in the control group) × 100%.

[0067] The experimental results showed that the survival rate of *Macrococcus eherii* ME1 was 76.8±3.5% at a bile salt concentration of 0.3%, 73.2±2.9% at 0.4% bile salt concentration, and 70.5±3.1% at 0.5% bile salt concentration. At bile salt concentrations of 0.3%–0.5%, the survival rate of *Macrococcus eherii* ME1 was ≥70%, indicating that this strain has good bile salt tolerance and can reach the intestine through the bile environment.

[0068] Experimental Example 1: Evaluation of the preventive and therapeutic effects of *Gastrococcus ehrlich* ME1 on DSS-induced intestinal inflammation in mice. 1. Experimental Design and Animal Grouping This experiment employed a 2×2 factorial design, using 80 six-week-old C57BL / 6 mice, which were randomly divided into four groups of 20 mice each: normal control group (CT), DSS model group (DSS), *Macrococcus ehrlich* ME1 group (M), and *Macrococcus ehrlich* ME1+DSS group (M+D). The formal experiment was conducted for 21 days after a one-week pre-feeding period. All mice were fed the same basal diet and had normal access to water during the experiment.

[0069] Mice in the CT group were gavaged with 0.2 mL of sterile saline throughout the entire course of treatment, and given normal drinking water. Mice in the DSS group were given normal drinking water for the first 14 days, and then had sodium dextran sulfate (DSS) added to their drinking water at a concentration of 3% for the next 7 days, and were gavaged with 0.2 mL of sterile saline throughout the entire course of treatment. Mice in the M group were gavaged with 0.2 mL of *Gastrococcus ehrlich* ME1 bacterial suspension (live bacterial concentration 1×10⁻⁶) throughout the entire course of treatment. 8(CFU / mL), normal drinking water; mice in the M+D group had normal drinking water for the first 14 days and were given DSS at a concentration of 3% in the drinking water for the next 7 days, and were intragastrically administered 0.2 mL of the bacterial suspension of Megasphaera elsdenii ME1 (viable cell concentration 1×10 8 (CFU / mL).

[0070] Calculated based on the average body weight of mice being approximately 20 g, the gavage dose was 0.2 mL×1×10 8 (CFU / mL)÷0.02 kg = 1×10 9 (CFU / kg body weight).

[0071] The mice were housed in a SPF-level animal room at an environmental temperature of 25±1 °C and a humidity of 50±5%, with a 12 h light-dark cycle maintained. On the 22nd day of the experiment (fasted for 12 h), the mice were sacrificed after collecting blood from the eyeballs.

[0072] 2 Growth performance determination During the experiment, the body weight of the mice was measured once a week during the gavage period, and once every 2 days during the DSS treatment period. The body weight of each mouse in each group was recorded, and the food intake per week was recorded. The disease activity index (DAI) score was calculated according to the criteria in Table 1 during the DSS treatment period.

[0073] Table 1 DAI scoring criteria

[0074] The test results were as Figure 3 shown. The body weights of the mice in the CT group and the M group increased steadily over time ( Figure 3 A), and the body weights of the mice in the DSS group decreased sharply after the 14th day. There was no significant difference in the body weights of the mice in each group before the 14th day of the experiment (P>0.05). On the 21st day of the experiment, the body weights of the mice in the DSS group and the M+D group were significantly lower than those in the CT group and the M group (P<0.05). During the period from the 15th to the 21st day of the experiment, the DAI score of the DSS group increased sharply over time ( Figure 3 B), showing a severe disease state, while the DAI score of the mice in the M+D group was significantly lower than that in the DSS group (P<0.05). The DSS group showed significant negative weight gain and decreased food intake during the DSS treatment period (P<0.05) ( Figure 3 C, D). The weight loss of the mice in the M+D group was significantly less than that in the DSS group during the period from the 15th to the 21st day of the experiment (0.05<P<0.1), and the decrease in food intake was less (P>0.05).

[0075] The above results indicate that under the condition of DSS-induced inflammatory stress in mice, Megasphaera elsdenii ME1 can slow down the weight loss of mice and significantly reduce diarrhea.

[0076] 3 Fecal occult blood evaluation The test results are as follows Figure 4 shown. The feces of mice in the CT group and the M group were formed and the fecal occult blood test was negative (no blue / black reaction). The feces of the DSS group were unformed (loose stools) and the fecal occult blood reaction was strongly positive, showing blue-black. Compared with the DSS group, the fecal traits and bleeding conditions of mice in the M+D group were significantly improved. The fecal occult blood reaction was dark blue, better than the strongly positive performance of the DSS group and close to that of the CT group and the M group. The results indicate that Megasphaera elsdenii ME1 can effectively reduce the intestinal bleeding condition induced by DSS.

[0077] 4 Determination of organ index After the mice were sacrificed at the end of the test, the heart, liver, spleen, duodenum, jejunum, ileum and colon were quickly dissected and separated. The attached fat and connective tissues were removed and rinsed thoroughly with normal saline. After drying the surface moisture with filter paper, the wet weights of each organ were accurately weighed, and the organ index was calculated (organ wet weight / mouse final weight × 100%).

[0078] The test results are as follows Figure 5 shown. Compared with the non-DSS treatment, DSS treatment led to a significant increase in the spleen index of mice (P<0.05, Figure 5 B), a significant decrease in the colon index (P<0.05, Figure 5 G), and a tendency for the liver index to increase (0.05<P<0.1, Figure 5 A). Compared with intragastric administration of normal saline, intragastric administration of Megasphaera elsdenii ME1 had a tendency for the heart index of mice to increase (0.05<P<0.1, Figure 5 C), and a tendency for the spleen index to increase (0.05<P<0.1, Figure B). There was an interaction effect between intragastric administration of Megasphaera elsdenii ME1 and DSS treatment on the liver index. The liver index of mice in the DSS group was significantly higher than that in the CT group and the M+D group (P<0.05). There was no significant difference between the M group and the other three groups (P>0.05, Figure 5 A). DSS treatment and intragastric administration of Megasphaera elsdenii ME1 had no significant effect on the duodenum, jejunum and ileum indices of mice (P>0.05, Figure 5 D-F).

[0079] The above results indicate that DSS induction significantly increased the liver and spleen indices and significantly decreased the colon index, reflecting the systemic inflammatory response of the body. The intragastric administration of Megasphaera elsdenii ME1 can effectively restore the liver index to the normal level.

[0080] 5 Histopathological evaluation of the colon After the test, the mice were sacrificed after collecting blood. The colon was separated, and a 1-cm intestinal segment was fixed and preserved in 4% paraformaldehyde solution. Paraffin sections were made, and the colon was pathologically observed under a microscope after hematoxylin-eosin staining. The pathological scoring criteria for colon tissue inflammation are shown in Table 2.

[0081] Table 2 Scoring criteria for colon inflammation

[0082] Colonic shortening is a typical feature of DSS colitis. Test results are as follows: Figure 6 As shown, the colon length in the DSS group was significantly shorter than that in the CT group (P<0.01), while the colon length in the M+D group was significantly longer than that in the DSS group (P<0.05), but significantly shorter than that in the CT and M groups (P<0.05).

[0083] The experimental results regarding the effect of ME1 on colonic pathological observation and scoring in DSS-treated mice are as follows: Figure 7 As shown, the colonic mucosal structure of the CT group and the M group was intact, and the crypts were neatly arranged. Figure 7 A and Figure 7 C), the DSS group showed extensive mucosal damage, crypt destruction and loss, and extensive inflammatory cell infiltration ( Figure 7 In group B, the M+D group retained a more complete crypt structure, and the degree of mucosal damage was significantly reduced. Figure 7 D). Compared with the CT group and the M group, the DSS group and the M+D group had significantly higher colonic pathology scores ( P <0.05), while the colonic pathology score of the M+D group was significantly lower than that of the DSS group ( P <0.05, Figure 7 E).

[0084] The above results indicate that gavage administration of *M. esculenta* ME1 significantly improved the colonic shortening induced by DSS treatment. Pathological sections showed that the DSS group had extensive mucosal damage, crypt destruction, and inflammatory cell infiltration, while the M+D group better preserved the integrity of the crypt structure and significantly reduced the colonic pathological score, demonstrating that *M. esculenta* ME1 has a significant protective effect on DSS-treated colonic tissue.

[0085] 6. Gestational Morphological Measurement The jejunum was separated, and a 1 cm segment was fixed and preserved in 4% paraformaldehyde solution. Paraffin sections were prepared, stained with hematoxylin and eosin, and the villus height and crypt depth were measured under a microscope.

[0086] Depend on Figure 8 The experiment showed that the empty villi in the CT group and the M group were arranged neatly and were slender. Figure 8 In groups A and 8C, damaged villous morphology was visible in the DSS group. Figure 8 B), the villous morphology of the M+D group was more complete than that of the DSS group. Figure 8 D). Compared with no DSS treatment, DSS treatment significantly reduced jejunal villus height and the villus height to crypt depth ratio (jejunal V / C). P <0.05, Figure 8 E and Figure 8 G), indicating that DSS damages the jejunal morphology and impairs digestive and absorptive functions; gavage Megasphaera elsdenii ME1 significantly increased jejunal villus height and crypt depth. P <0.05, Figure 8 (E and 8F), the jejunal V / C ratio showed an increasing trend (0.05 < P <0.1, Figure 8 G).

[0087] The above results indicate that gavage administration of *Gastrococcus ehrlich* ME1 can effectively protect the jejunal villus structure and help ensure the intestinal absorption function of mice.

[0088] 7. Measurement of colonic inflammatory factors Colonic tissue was rapidly dissected, and 100 mg of tissue was weighed into a 2 mL centrifuge tube and mixed with 1 mL of pre-cooled sterile saline. The mixture was homogenized by vortexing for 30 s and centrifuged at 4 °C for 15 min (14000 × g). The supernatant was transferred to a new centrifuge tube, and the levels of IL-6, IL-17, IL-1β, TNF-α, and IL-10 were measured using ELISA. The measurement procedures and methods followed the instructions of the kit (Shanghai Enzyme-linked Biotechnology, Co., Ltd., China). The colonic tissue measurement results were corrected for tissue protein concentration, which was determined using the BCA kit according to the kit's instructions.

[0089] according to Figure 9 The experimental results show that DSS treatment and gavage Megasphaera elsdenii ME1 significantly affected the expression of inflammatory factors in the mouse colon, and there was an interaction between the two. Compared with the CT group and the M group, DSS treatment significantly increased the protein expression levels of pro-inflammatory factors IL-6, IL-17, IL-1β and TNF-α in the colon. P <0.01, Figure 9 A, Figure 9 B. Figure 9 C and Figure 9 D), significantly reduced the protein expression level of the anti-inflammatory factor IL-10 ( P <0.01, Figure 9 E); Compared with the DSS group, the protein expression levels of pro-inflammatory factors IL-6, IL-17, IL-1β and TNF-α in the M+D group were significantly reduced ( P <0.01), the protein expression level of the anti-inflammatory factor IL-10 was significantly increased ( P <0.01); the protein expression level of IL-10 in the colon of group M was significantly higher than that of group CT ( P<0.01), there was no significant difference in the expression levels of pro-inflammatory cytokine proteins between the two groups. P >0.05).

[0090] The above results indicate that gavage administration of *Gastrococcus ehrlich* ME1 can significantly inhibit the expression of pro-inflammatory factors IL-6, IL-17, IL-1β and TNF-α, and promote the production of anti-inflammatory factor IL-10, thereby alleviating DSS-induced intestinal inflammatory response in mice.

[0091] 8. Measurement of intestinal barrier markers Colonic tissue homogenate supernatant was prepared using the same method as for colonic inflammatory factors, and the expression levels of three tight junction proteins, ZO-1, Occludin, and Claudin-1, in the colon were determined by ELISA.

[0092] On day 22 of the experiment, blood was collected from mice by enucleation and placed in centrifuge tubes. After standing at room temperature for 2 hours, the cells were centrifuged at 3000×g for 15 minutes at 4 °C. The serum was separated and aliquoted into 2 mL cryovials and stored at -80 °C for later analysis. Serum D-lactic acid, lipopolysaccharide (LPS), and diamine oxidase (DAO) levels were measured using ELISA.

[0093] according to Figure 10 The experimental results showed that, compared with no DSS treatment, DSS treatment significantly reduced the expression levels of three tight junction proteins, ZO-1, Occludin, and Claudin-1, in the mouse colon, and significantly increased the serum D-lactic acid content in mice. P <0.01, Figure 10 A, Figure 10 B Figure 10 C and Figure 10 F); and those not given gastric gavage Megasphaera elsdenii Compared to ME1, gavage Megasphaera elsdenii ME1 significantly increased the expression levels of colonic ZO-1, Occludin, and Claudin-1 proteins. P <0.01), DSS treatment and gavage Megasphaera elsdenii ME1 has no interaction ( P >0.05). DSS treatment and gavage. Megasphaera elsdenii ME1 significantly affected serum LPS and DAO levels in mice, and there was an interaction between the two. Compared with the CT group, DSS treatment significantly increased serum LPS and DAO levels in mice. P <0.01, Figure 10 D and Figure 10 E) indicates impaired intestinal barrier function and increased permeability; compared with the DSS group, the serum LPS and DAO levels in the M+D group mice were significantly reduced ( P<0.01), the serum DAO content in the M+D group was significantly higher than that in the M group and the CT group ( P <0.05, there was no significant difference in serum LPS and DAO levels between the M group and the CT group ( P >0.05).

[0094] The above results indicate that oral administration of *Gastrococcus escherichia coli* ME1 can repair the intestinal mechanical barrier function damaged by DSS and reduce intestinal permeability by upregulating the expression of colonic tight junction proteins.

[0095] 9. Measurement of oxidative stress indicators Colonic tissue homogenate supernatant was prepared using the same method as for colonic inflammatory factors. Colonic malondialdehyde (MDA) content and superoxide dismutase (SOD) activity were measured using ELISA. Serum MDA content and SOD activity were also measured simultaneously.

[0096] The test results are as follows Figure 11 As shown, DSS treatment and gavage Megasphaera elsdenii ME1 significantly affected colonic and serum MDA and SOD levels in mice, and there was an interaction between the two. Compared with the CT and M groups, the DSS group showed a highly significant increase in colonic and serum malondialdehyde (MDA) levels. P <0.01%, and the activity of superoxide dismutase (SOD) in the colon and serum was significantly reduced ( P <0.01); compared with the DSS group, the M+D group significantly reversed this trend and significantly reduced serum MDA levels ( P <0.01, significantly increased SOD enzyme activity in colon and serum ( P <0.05).

[0097] The above results indicate that gavage administration of Giant Pleurotus erythrorhizon ME1 can significantly alleviate DSS-induced oxidative stress in mice.

[0098] 10. Intestinal microbiome testing Total genomic DNA was extracted from cecal chyme using a fecal DNA kit (OMEGA Soil DNA Kit (M5635-02) (Omega Bio-Tek, Norcross, GA, USA)). The variable region V3-V4 of the 16S rRNA gene was amplified using 338F (5'-barcode+ACTCCTACGGGAGGCAGCA-3') and 806R (5'-GGACTACHVGGGTWTCTAAT-3'). The barcode in the pre-primer (338F) is a 7-10 nucleotide oligonucleotide sequence used to distinguish different samples within the same library. All generated amplicons were homogenized, and the PCR products were then purified using a kit. Sequencing libraries were constructed using the TruSeq Nano DNA LT Library Prep Kit according to the manufacturer's instructions. After assessing the quality of the sequencing libraries, they were sequenced using an Illumina platform. QIIME2 2019.4 was used to perform quality filtering, noise reduction, splicing, and chimera removal on microbiome biological information. The obtained sequences were merged based on 100% sequence similarity to generate characteristic ASVs. ASVs with abundance values ​​below 0.001% (one in ten thousand) of the total sequencing volume were removed. The Alpha diversity index was calculated using QIIME2 to analyze the diversity, richness, and evenness of the bacterial community in the samples.

[0099] 10.1 Analysis of gut microbial diversity The test results are as follows Figure 12 As shown, the alpha diversity (Chao1 and Observed_species) of the gut microbiota in the DSS and M groups was lower than that in the CT group, while the M group was slightly higher than that in the CT group. Compared with the DSS group, the alpha diversity (Chao1, Observed_species, and Shannon index) of the cecal microbiota in the M+D group was significantly increased. However, there was no statistically significant difference in alpha diversity among the CT, DSS, M, and M+D groups (P>0.05).

[0100] PCA analysis showed that the samples were clearly separated, with the CT and M groups clustering together, and the DSS and M+D groups clustering together, indicating that DSS treatment had a significant impact on the Beta diversity of the mouse cecal microbiota.

[0101] The above results indicate that gavage administration of *Gastrococcus ehrlich* ME1 can, to some extent, alter the alpha and beta diversity of the gut microbiota, alleviating the dysbiosis caused by DSS.

[0102] 10.2 Gut Microbial Species Composition The test results are as follows Figure 13 As shown, at the bacterial level, the microbial species composition in the mouse gut was mainly composed of Firmicutes A and Bacteroides, with a combined relative abundance exceeding 70%. Firmicutes A had the highest relative abundance in the DSS-treated group, reaching 49%. Figure 13 A); In the CT and M groups, the relative abundance of Verrucomicrobiota was significantly higher than that in the DSS and M+D groups, while the DSS and M+D groups were rich in Proteobacteria.

[0103] At the bacterial genus level, *Gastrodia elata* was the dominant species in the CT and M groups, while *Gastrodia elata* was relatively abundant in the DSS and M+D groups, accompanied by a decrease in the relative abundance of genera such as *UBA3282* and *CAG-873*. Figure 13 B).

[0104] The Sankey diagram visually illustrates species mobility at different taxonomic levels. The bacterial community in the CT group is mainly associated with Verrucomicrobiota and g_Akkermansia; the bacterial community in the DSS group is centered on Proteobacteria and g_Escherichia, reflecting the enrichment of potential pathogens in diarrheal states. Figure 13 C); The microbial community composition of the M+D group showed a trend of regression towards the CT group, characterized by an increase in the proportion of Firmicutes_A-related genera and a decrease in the relative abundance of Proteobacteria.

[0105] LEfSe significance analysis showed that, compared with the CT group, the DSS group was significantly enriched in g_Escherichia, g_Bilophila, g_Turicibacter, and g_Romboutsia_B; the M group significantly increased the relative abundance of g_CAG-873, g_Alloprevotella, g_Lactobacillus, and g_Anaerotruncus; and the M+D group was significantly enriched in g_Fournierella, g_Butyricimonas, g_Fimenecus, g_Faecousia, g_Faecalibaculum, g_Eubacterium_B, and g_Bifidobacterium. Figure 13 D).

[0106] The above results indicate that DSS-induced diarrhea can lead to intestinal flora dysbiosis, significantly increasing the relative abundance of potentially pathogenic bacteria such as Proteobacteria and g_Bilophila. In contrast, gavage with *Gastrococcus ehrlich* ME1 can improve the flora structure, significantly increasing the relative abundance of beneficial bacteria such as g_Lactobacillus, g_Alloprevotella, g_Butyricimonas, g_Faecalibaculum, g_Eubacterium_B, and g_Bifidobacterium. The short-chain fatty acids produced by this bacteria can help provide rapid energy to mouse intestinal cells and improve the body's resistance to DSS treatment.

[0107] Example 2: Evaluation of the protective effect of *Gastrococcus ehrlich* ME1 on the gut-liver axis. This study is based on the experimental design of Case 1 and focuses on evaluating the protective effect of Ehrlich giant cocci ME1 on the gut-liver axis, including liver index, liver pathology, liver inflammatory factors, serum LPS and other indicators.

[0108] 1. Liver index Liver index was measured as described in Example 1. The results are shown below. Figure 5 A. An interaction effect was observed between gavage administration of *M. esculenta* ME1 and DSS treatment on liver indices. The liver index in the DSS group was significantly higher than that in the CT and M+D groups (P<0.05), while there were no significant differences between the M group and the other three groups (P>0.05). These results indicate that DSS induces secondary liver injury with a significant increase in liver indices, while *M. esculenta* ME1 can effectively restore liver indices to normal levels.

[0109] 2. Liver pathological evaluation After the experiment, blood was collected from mice, which were then sacrificed. Liver tissue was isolated, fixed and preserved in 4% paraformaldehyde solution, and paraffin sections were prepared. After hematoxylin-eosin staining, liver pathological observation was performed under a microscope. The liver necrosis and inflammation severity scoring system (HAI) is shown in Table 3.

[0110] Table 3. Liver tissue necrosis and inflammation severity scoring system (HAI)

[0111] The test results are as follows Figure 14As shown, the liver lobule structure in the CT and M groups was intact, and the hepatocytes were neatly arranged without obvious inflammatory cell infiltration or necrosis. In the DSS group, disordered hepatocyte cord arrangement was observed, accompanied by inflammatory cell infiltration or necrosis. Compared with the DSS group, the liver tissue structure in the M+D group was more intact, and the inflammatory cell infiltration was significantly reduced. DSS treatment and gavage with *M. elegans* ME1 significantly affected the Knodell HAI score, and there was an interaction between the two. The histological activity index (HAI) of the DSS and M+D groups was significantly higher than that of the CT and M groups (P<0.01), indicating that DSS treatment led to severe liver inflammation and necrosis. Compared with the DSS group, the HAI score of the M+D group was significantly lower (P<0.01).

[0112] The above results indicate that DSS treatment led to secondary liver injury, while *Gastrococcus ehrlich* ME1 had a significant protective effect against DSS-induced liver injury.

[0113] 3. Measurement of liver inflammatory factors Liver tissue was rapidly dissected, and 100 mg of tissue was weighed into a 2 mL centrifuge tube and mixed with 1 mL of pre-cooled sterile saline. The mixture was homogenized by vortexing for 30 s and centrifuged at 4 °C for 15 min (14000×g). The supernatant was transferred to a new centrifuge tube, and the levels of IL-6, IL-17, IL-1β, TNF-α, and IL-10 were measured using ELISA. The liver tissue measurement results were corrected for tissue protein concentration.

[0114] The test results are as follows Figure 15 As shown, both DSS treatment and gavage administration of *M. ergium* ME1 significantly affected the expression of inflammatory factors in mouse liver, and there was an interaction between the two. Compared with the CT and M groups, DSS treatment significantly increased the protein expression levels of pro-inflammatory factors IL-6, IL-17, IL-1β, and TNF-α in the liver (P<0.01) and significantly decreased the protein expression level of anti-inflammatory factor IL-10 (P<0.01). Compared with the DSS group, the M+D group showed significantly decreased protein expression levels of pro-inflammatory factors IL-6, IL-17, IL-1β, and TNF-α (P<0.01) and significantly increased protein expression level of anti-inflammatory factor IL-10 (P<0.01). The liver TNF-α protein level in the M group was significantly lower than that in the CT group (P<0.01), while the IL-6 protein level was significantly higher than that in the CT group (P<0.05).

[0115] The above results indicate that gavage administration of *Gastrococcus ehrlich* ME1 significantly alleviated DSS-induced liver inflammation.

[0116] 4. Serum LPS Measurement Serum was collected according to the method described in Example 1, and serum LPS levels were measured using ELISA. The results are shown below. Figure 10 D, DSS treatment and gavage administration of *M. ergium* ME1 significantly affected serum LPS levels in mice, and there was an interaction between the two. Compared with the CT group, DSS treatment significantly increased serum LPS levels in mice (P<0.01), indicating impaired intestinal barrier function and endotoxin entry into the bloodstream; compared with the DSS group, serum LPS levels in the M+D group were significantly decreased (P<0.01), and there was no significant difference in serum LPS levels between the M group and the CT group (P>0.05).

[0117] The combined results of Experiment 1 and Experiment 2 demonstrate that gavage administration of Giant Ehrlich aspergillus ME1 has a protective effect on the gut-liver axis. Through multiple mechanisms, such as improving intestinal inflammation, reducing intestinal permeability, reducing endotoxin entry into the bloodstream, and inhibiting the expression of liver inflammatory factors, it effectively alleviates secondary liver damage caused by intestinal inflammation.

[0118] Experimental Example 3: Evaluation of the ameliorative effect of *Gastrococcus ehrlich* ME1 on spleen pathology. This experimental design, based on Experiment 1, focuses on evaluating the ameliorative effect of Ehrlich giant cocci ME1 on spleen pathology.

[0119] After the experiment, blood was collected from the mice, and the spleens were isolated. Tissue was fixed and preserved in 4% paraformaldehyde solution, paraffin sections were prepared, and the spleens were stained with hematoxylin and eosin for microscopic pathological observation. The pathological scoring criteria for spleen tissue inflammation are shown in Table 4.

[0120] Table 4. Pathological Scoring Criteria for Splenic Tissue Inflammation

[0121] The test results are as follows Figure 16 As shown, the red and white pulp boundaries were clear in the CT and M groups. However, in the DSS group, the red and white pulp structures of the spleen were unclear, the splenic corpuscles were relatively smaller, most splenic corpuscle margins were indistinct, and in some areas the margins disappeared; the number and density of lymphocytes around the central artery were significantly reduced, and their volume was significantly smaller; the red pulp volume was significantly increased, the number of splenic cord lymphocytes was significantly increased, a large number of neutrophils were visible in the medullary sinus, and local splenic tissue showed myeloid metaplasia; macrophages and megakaryocytes in the medullary sinus were significantly reduced. In the M+D group, the red and white pulp structures were clear, the number of splenic cord lymphocytes was increased locally, and scattered neutrophils were visible in the medullary sinus; no other abnormalities were observed.

[0122] According to the scoring criteria in Table 4, the DSS group was rated as "++++ or +++" severe damage, characterized by unclear structure, reduced spleen bodies, and a large number of neutrophils. The M+D group was rated as "- or +" mild damage, mainly manifested as increased local lymphocytes, with no abnormalities in the overall structure.

[0123] The above results indicate that *Gastrococcus ehrlich* ME1 significantly improved the pathological structural disorder of the spleen caused by DSS, reduced the aggregation of neutrophils in the spleen, and effectively protected the structural integrity of the red and white pulp of the spleen.

Claims

1. A strain of giant cocci, characterized in that, Its microbial preservation number is CGMCC No. 46926.

2. A microbial preparation or feed additive prepared from *Gastrococcus echocarpa* of claim 1.

3. Use of the giant cocci of claim 1 in the preparation of microbial preparations or feed additives that promote animal growth.

4. Use of the giant cocci of claim 1 in the preparation of microbial preparations or feed additives for relieving diarrhea.

5. The use of *Gastrococcus eherii* as described in claim 1 in the production of short-chain fatty acids, wherein, The short-chain fatty acids include propionic acid and butyric acid.

6. The use of *Gastrococcus eherii* as described in claim 1 in the preparation of microbial preparations or feed additives that reduce the expression of pro-inflammatory factors and increase the expression of anti-inflammatory factors; wherein, The pro-inflammatory factors include IL-6, IL-17, IL-1β, or TNF-α; the anti-inflammatory factor is IL-10.

7. The use of the giant cocci of claim 1 in the preparation of microbial preparations or feed additives for the prevention and treatment of intestinal inflammation; preferably, the intestinal inflammation is selected from one or more of ulcerative colitis, Crohn's disease, DSS-induced intestinal inflammation, and radiation enteritis.

8. The use of the giant cocci of claim 1 in the preparation of microbial preparations or feed additives that increase the types or quantities of beneficial intestinal bacteria; preferably, the beneficial bacteria are selected from one or more of the genera *Lactobacillus*, *Bifidobacterium*, *Butyrica*, *Femtobacter*, and *Eubacterium*.

9. The use of *Gastrococcus ehrlich* as described in claim 1 in the preparation of microbial preparations or feed additives for the prevention and treatment of hepatitis or liver injury; preferably, the liver injury is secondary liver injury caused by intestinal inflammation.

10. Use of the giant cocci of claim 1 in the preparation of microbial preparations or feed additives for the prevention and treatment of spleen inflammation or damage.

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