Application of bacteroides simplex in preparation of medicine for preventing and / or treating ulcerative colitis and microbial preparation

Abstract

The invention belongs to the technical field of microbial medicine, and particularly relates to application of bacteroides simplex in medicine for preventing and / or treating ulcerative colitis and a microbial preparation. The invention provides an application of Bacteroides simplex in a medicine for preventing and / or treating ulcerative colitis. The Bacteroides simplex disclosed by the invention can be used for effectively treating the ulcerative colitis. In the specific embodiment of the invention, the bacteroides simplex can relieve weight loss of mice suffering from ulcerative colitis and relieve fecal occult blood of the mice suffering from ulcerative colitis; the disease activity index of a mouse with ulcerative colitis is reduced; reversing the colon length of the mouse with ulcerative colitis; the mouse inflammatory factor expression level of the ulcerative colitis is reduced, and the ulcerative colitis colon barrier is repaired. Therefore, the Bacteroides simplex disclosed by the invention can be used for remarkably relieving the symptoms of mouse colitis induced by DSS and has the effect of treating ulcerative colitis.

Description

Technical Field

[0001] This invention belongs to the field of microbial pharmaceutical technology, specifically relating to the application of Bacteroides monomorpha in the preparation of drugs for the prevention and / or treatment of ulcerative colitis and microbial preparations. Background Technology

[0002] Ulcerative colitis (UC) is a lifelong inflammatory disease affecting the rectum and colon. Epidemiological surveys show that there were 5 million UC patients worldwide in 2023, and its prevalence is increasing year by year. Clinical studies have shown that the risk of developing colon cancer increases with the duration of the disease.

[0003] UC is a specific type of colitis, but the two are not entirely equivalent. Colitis refers to inflammation of the colon in general, with a wide range of causes (such as infection, ischemia, and medications), while UC belongs to chronic autoimmune inflammatory bowel disease (IBD), with causes related to genetics and immune abnormalities. The lesions are limited to the colon and rectum, presenting as continuous mucosal ulcers. Compared with other types of colitis, UC has unique characteristics: pathologically, it presents as diffuse inflammation of the mucosal layer and crypt abscesses, requiring long-term clinical management; while infectious colitis can be cured with antibiotics, and ischemic colitis often has acute exacerbations due to blood flow obstruction. Diagnosis of UC requires colonoscopy, biopsy, and exclusion of other types (such as Crohn's disease and infectious colitis). In terms of treatment, UC relies on immunomodulators, while other types of colitis may be relieved through etiological treatment (such as anti-infection and improved circulation). UC is a lifelong disease with a high risk of relapse, while some types of colitis (such as acute infectious colitis) can be completely cured.

[0004] To date, there is no effective treatment for ulcerative colitis (UC). Treatment primarily focuses on relieving symptoms, promoting mucosal healing, and reducing complications. Common clinical treatments include salicylates, hormones, immunosuppressants, and anti-tumor necrosis factor-α (TNF-α) monoclonal antibodies. However, long-term use of immunosuppressants often leads to adverse reactions, and biologics are expensive with limited domestic availability, while also carrying a high risk of complications. Therefore, in-depth research into its pathophysiological mechanisms and the exploration of effective treatment strategies and drugs are of great significance in efforts to reduce the social and economic impact of this disease.

[0005] Currently, probiotic preparations for treating ulcerative colitis often employ mixed bacterial flora, which presents challenges such as complex strain interactions and unclear efficacy. The single-strain probiotics currently used to treat ulcerative colitis include *Lactobacillus reuteri* JYLB-291, *Lactobacillus casei* L. Casei21, *Bifidobacterium longum* JBLC-141, *Lactobacillus paracasei* JLPF-176, *Lactobacillus acidophilus* JYLA-191, and *Bifidobacterium adolescentis* BBF-06. These are all common lactic acid bacteria and require combination therapy with conventional Western medicines for ulcerative colitis; otherwise, the effect is poor. Summary of the Invention

[0006] The purpose of this invention is to provide an application of Bacteroides monomorphosum in the preparation of drugs for the prevention and / or treatment of ulcerative colitis and a microbial preparation. This invention utilizes specific Bacteroides monomorphosum strains to precisely regulate the pathological process of UC through single-strain targeted intervention, without the need for combination with Western medicines for the treatment of ulcerative colitis, thus overcoming the shortcomings of existing technologies and improving the treatment effect of ulcerative colitis.

[0007] To address the aforementioned technical problems, the present invention proposes the following technical solution:

[0008] This invention provides the use of Bacteroides uniformis in the preparation of medicaments for the prevention and / or treatment of ulcerative colitis.

[0009] Preferably, the Bacteroides monomorpha includes the Bacteroides monomorpha strain numbered BNCC139204.

[0010] Preferably, the application of the drug includes any one or more of the following:

[0011] 1) Relieves fecal occult blood in ulcerative colitis;

[0012] 2) Reduce the disease activity index of ulcerative colitis;

[0013] 3) Reverse the length of the colon in ulcerative colitis;

[0014] 4) Reduce the expression levels of inflammatory factors in ulcerative colitis;

[0015] 5) Relieves weight loss in patients with ulcerative colitis;

[0016] 6) Pathological indicators for relieving ulcerative colitis.

[0017] Preferably, the inflammatory factors include one or more of IL-18, IL-1β, and TNF-α.

[0018] This invention provides a microbial preparation for the prevention and / or treatment of ulcerative colitis, comprising Bacteroides uniformis and pharmaceutically acceptable excipients.

[0019] Preferably, the application of the monomorphic Bacteroides includes bacterial suspension.

[0020] Preferably, the method for preparing the bacterial solution includes: inoculating the Bacteroides monomorpha into Columbia blood agar medium for culture to obtain the bacterial solution.

[0021] Preferably, the culture includes anaerobic culture.

[0022] Preferably, the number of viable Bacteroides monomorpha in the microbial preparation is ≥1×10⁻⁶. 8 CFU / mL or ≥1×10 8 CFU / g.

[0023] Preferably, the dosage form of the microbial preparation includes one or more of oral dosage forms, injections, and suppositories.

[0024] The beneficial effects of this invention are as follows: This invention provides the application of Bacteroides uniformis in the preparation of drugs for the prevention and / or treatment of ulcerative colitis. The Bacteroides uniformis of this invention can significantly increase the abundance of butyrate-producing bacteria such as *Faecalibacterium prausnitzii* in the intestine; it can downregulate the NLRP3 inflammasome pathway (NLRP3 / ASC / Caspase-1), and reduce the levels of IL-1β, IL-18, and TNF-α; it can upregulate the expression of tight junction proteins (Claudin-1 and ZO-1), reducing intestinal mucosal damage, thereby effectively treating ulcerative colitis. In specific embodiments of this invention, the Bacteroides uniformis can alleviate fecal occult blood in mice with ulcerative colitis; reduce the disease activity index in mice with ulcerative colitis; reverse the colon length in mice with ulcerative colitis; reduce the expression levels of inflammatory factors in mice with ulcerative colitis; and simultaneously alleviate weight loss and pathological indicators of ulcerative colitis in mice. It is evident that the monomorphobacterium of the present invention can significantly alleviate the symptoms of DSS-induced colitis in mice and has the effect of treating ulcerative colitis. Attached Figure Description

[0025] Figure 1 Flowcharts for each group of experimental animals;

[0026] Figure 2 Disease Activity Index (DAI) score charts for each group of mice;

[0027] Figure 3 The graph shows the changes in body weight of mice in each group;

[0028] Figure 4 Images showing rectal bleeding around the anus in mice of each group;

[0029] Figure 5 Images showing fecal occult blood in each group of mice;

[0030] Figure 6 A comparative image of typical colon lengths in mice from each group;

[0031] Figure 7Representative H&E staining images of cross-sections of the large intestine of mice in the ABX-Control group (scale bar = 50 μm);

[0032] Figure 8 Representative H&E staining images of cross-sections of the large intestine of ABX-DSS group mice (scale bar = 50 μm);

[0033] Figure 9 Representative H&E staining images of cross-sections of the large intestine of ABX-B. uniformis mice (scale bar = 50 μm);

[0034] Figure 10 The graph shows the expression levels of TNF-α, IL-18, and IL-1β in the large intestine tissue of mice in each group, as detected by ELISA.

[0035] Figure 11 The mRNA expression maps of core targets TNF-α, IL-18, and IL-1β in the large intestine tissue of mice in each group were obtained by qRT-PCR. Figures 10-11 middle, ## P<0.01 compared with the control group, **P<0.01 compared with the model group;

[0036] Figure 12 AB-PAS staining images of colon tissue from each group of mice;

[0037] Figure 13 Immunofluorescence images of TNF-α in the colon tissue of mice in each group;

[0038] Figure 14 Immunofluorescence images of occludin in colon tissues of mice in each group.

[0039] Figure 15 Cluster heatmaps for quantifying short-chain fatty acids in the colon tissue of mice in each group. Detailed Implementation

[0040] This invention provides the use of Bacteroides uniformis in the preparation of medicaments for the prevention and / or treatment of ulcerative colitis.

[0041] This invention does not specifically limit the source or strain number of *Bacteroides uniformis*, which can be routinely purchased by those skilled in the art. In this embodiment, the *Bacteroides uniformis* mentioned in this invention includes strain BNCC139204. The *Bacteroides uniformis* strain BNCC139204 of this invention was purchased from the Henan Provincial Industrial Microbial Strains Engineering Technology Research Center of Beina Biotechnology.

[0042] Bacteroides monomorphosum is one of the core symbiotic bacteria of the gut. Existing technologies have reported that Bacteroides monomorphosum plays a role in gut microbiota regulation: competitively inhibiting the colonization of pathogenic bacteria (such as Escherichia coli) and promoting the proliferation of beneficial bacteria (such as Bifidobacterium). In this invention, it is discovered for the first time that Bacteroides monomorphosum has an anti-inflammatory effect: Bacteroides monomorphosum inhibits the expression of pro-inflammatory factors (TNF-α, IL-1β, IL-6) through metabolites (such as short-chain fatty acids) and activates anti-inflammatory pathways (such as IL-10); at the same time, Bacteroides monomorphosum also has a barrier repair function, which can upregulate the expression of tight junction proteins (Occludin, ZO-1) and reduce intestinal permeability. The results of the embodiments of the present invention show that: Oral administration of *Bacteroides monomorphosus* significantly alleviates DSS-induced colitis symptoms in mice, relieves fecal occult blood in mice with ulcerative colitis, reduces the disease activity index (DAI) in mice with ulcerative colitis (i.e., reduces the DAI score), reverses colon length in mice with ulcerative colitis, reduces the expression level of inflammatory factors in mice with ulcerative colitis, and simultaneously alleviates weight loss in mice with ulcerative colitis, alleviates pathological indicators of ulcerative colitis, restores the integrity of the colonic mucosa, gradually normalizes gland distribution, significantly alleviates inflammatory cell infiltration, and effectively controls crypt abscesses and ulcer formation. Therefore, *Bacteroides monomorphosus* of the present invention can effectively treat ulcerative colitis.

[0043] As an optional implementation, the application of the drug of the present invention preferably includes any one or more of the following: 1) effectively relieving fecal occult blood in ulcerative colitis; 2) reducing the disease activity index of ulcerative colitis; 3) reversing colon length in ulcerative colitis; 4) reducing the expression level of inflammatory factors in ulcerative colitis; 5) relieving weight loss in patients with ulcerative colitis; 6) alleviating pathological indicators of ulcerative colitis. As an optional implementation, the pathological indicators of alleviating ulcerative colitis include one or more of the following: restoring the integrity of colonic mucosa; gradually normalizing gland distribution; significantly alleviating inflammatory cell infiltration; effectively controlling crypt abscesses; promoting tissue repair and inhibiting ulcer progression; and promoting healing. As an optional implementation, the inflammatory factors of the present invention include one or more of IL-18, IL-1β, and TNF-α, more preferably IL-18, IL-1β, and TNF-α. The colon length of mice with ulcerative colitis is shortened, while the colon length of mice with ulcerative colitis is lengthened after treatment with Bacteroides monomorpha.

[0044] This invention provides a microbial preparation for the prevention and / or treatment of ulcerative colitis, comprising Bacteroides uniformis and pharmaceutically acceptable excipients.

[0045] This invention does not impose any special limitations on the pharmaceutically acceptable excipients; conventional products can be used.

[0046] As an optional implementation, the application form of *Bacteroides monomorphosus* of the present invention includes bacterial suspension. The preparation method of the bacterial suspension of the present invention includes: inoculating the *Bacteroides monomorphosus* into Columbia blood agar medium for culture to obtain a bacterial suspension. As an optional implementation, the present invention involves streaking the *Bacteroides monomorphosus* into Columbia blood agar medium for culture to obtain single colonies. As an optional implementation, the present invention does not have a particular limitation on the streaking method; conventional methods can be used. As an optional implementation, the culture temperature of the present invention is 35℃~38℃, or it can be 37℃; in specific embodiments of the present invention, the culture temperature is 35, 36, 37, or 38℃. The culture time of the present invention is 48~72h, or it can be 48~60h, more preferably 48h. In specific embodiments of the present invention, the culture time is 48, 50, 54, 58, 60, 64, or 72h. The culture of the present invention is anaerobic culture.

[0047] As an optional implementation, to obtain single colonies, the present invention mixes the single colonies with sterile water to obtain a bacterial solution. As an optional implementation, the viable count of the bacterial solution is ≥9 × 10⁻⁶. 8 CFU / mL, or ≥9×10 8 CFU / g ~ 10×10 10 CFU / mL, more preferably 9×10⁻⁶ 8 CFU / mL. As an optional embodiment, the Columbia blood culture medium of the present invention comprises 23.0 g / L of special peptone, 1.0 g / L of soluble starch, 5.0 g / L of sodium chloride and 10.0 g / L of agar, and the pH of the Columbia blood culture medium is 7.3 ± 0.2.

[0048] As an optional implementation, the number of viable Bacteroides monomorpha in the microbial preparation of the present invention can be ≥1×10⁻⁶. 8 CFU / mL, more preferably 9×10⁻⁶ 8 CFU / mL or ≥9×10 8 CFU / g ~ 10×10 10 CFU / mL. In a specific embodiment of the present invention, the concentration of the Bacteroides monomorpha BNCC139204 bacterial culture purchased by the present invention is 9 × 10⁻⁶ CFU / mL. 8 CFU / mL. The preferred dosing frequency for the microbial preparation described in this invention is once daily, and the dosage is [missing value]. This invention is the first to verify the intervention effect of Bacteroides monomorphosum BNCC139204 in the prevention and treatment of ulcerative colitis. A DSS-induced mouse model was established by intervention with Bacteroides monomorphosum BNCC139204 bacterial suspension, clarifying the therapeutic effect of Bacteroides monomorphosum BNCC139204 on ulcerative colitis.

[0049] As an optional embodiment, the dosage form of the microbial preparation of the present invention includes one or more of oral dosage forms, injections, and suppositories, more preferably oral dosage forms, injections, and suppositories. As an optional embodiment, the oral dosage form of the present invention is a lyophilized powder or enteric-coated capsule.

[0050] The etiology and specific pathogenesis of ulcerative colitis (UC) are not fully understood, but it is currently believed to be related to multiple factors, including gut microbiota dysbiosis, intestinal mucosal barrier disruption, intestinal mucosal immune abnormalities, genetic susceptibility, diet, psychological factors, and environmental factors. The Bacteroides monomorpha BNCC139204 of this invention can significantly alleviate DSS-induced colitis symptoms in mice and has a therapeutic effect on ulcerative colitis.

[0051] To further illustrate the present invention, the technical solutions provided by the present invention will be described in detail below with reference to the accompanying drawings and embodiments, but these should not be construed as limiting the scope of protection of the present invention.

[0052] Example 1: Source of the strain

[0053] Bacteroides uniformis bacterial suspension was purchased from the Henan Provincial Industrial Microbial Strains Engineering Technology Research Center of Beina Biotechnology. The strain number is BNCC139204, and it is designated as Bacteroides uniformis BNCC139204. The bacterial suspension concentration is 9 × 10⁻⁶. 8 CFU / mL. Bacteroides monomorpha BNCC139204 is an intestinal symbiotic bacterium preserved in the BeNa Culture Collection (BNCC) with a safety level of 2. This strain is provided in culture form and stored at 4°C for 7 days.

[0054] Bacteroides monomorpha BNCC139204 colonies are grayish-white, round, raised structures with a diameter of 0.5–1 mm, irregular edges, and Gram-negative (red bacillus).

[0055] The Columbia blood agar ready-to-use plate consists of: 23.0 g / L special peptone, 1.0 g / L soluble starch, 5.0 g / L sodium chloride and 10.0 g / L agar, with a pH of 7.3 ± 0.2.

[0056] Mechanism of action:

[0057] Microbial remodeling: Bacteroides monomorpha BNCC139204 can significantly increase the abundance of butyrate-producing bacteria in the gut, such as Clostridium prausnitzii.

[0058] Inflammatory inhibition: Bacteroides uniformis BNCC139204 can downregulate the activation of the NLRP3 inflammasome pathway (NLRP3 / ASC / Caspase-1) and reduce the levels of IL-1β, IL-18, and TNF-α.

[0059] Barrier repair: Bacteroides uniformis BNCC139204 can upregulate the expression of tight junction proteins (Occludin, ZO-1) and reduce intestinal mucosal damage.

[0060] Example 2 Pharmacodynamic verification (mouse model experiment)

[0061] 2.1 Experimental reagents.

[0062] Human urine and feces occult blood test kit (benzidine method), purchased from Nanjing Jiancheng Bioengineering Institute (batch number C027-1-1); Universal tissue fixative (Wuhan Sevier Biotechnology Co., Ltd., batch number G1101-3ML); ELISA kit purchased from Wuhan Elabscience Biotechnology Co., Ltd.; Normal saline; High-efficiency RIPA lysis buffer (Beijing Solarbio Science & Technology Co., Ltd. R0010-100ml), Prestained Protein Marker VIII (Wuhan Sevier Biotechnology Co., Ltd., product number MPC2311006); β-actin, Claudin-1 antibody, and Occludin antibody were all purchased from Wuhan Sanying Biotechnology Co., Ltd., batch numbers were 66009-1-AP, 28674-1-AP, 27260-1-AP respectively); Horseradish peroxidase (HRP)-labeled goat anti-rabbit immunoglobulin G (IgG) (Wuhan Sanying Biotechnology Co., Ltd., batch number: 20000858); PVDF membrane (cytiva company, batch number: 10600023); Super-sensitive ECL chemiluminescence kit (brand Xinsaimi, batch number: P10100).

[0063] 2.2 Experimental grouping, model establishment, and drug administration

[0064] (1) Experimental animals

[0065] 100 male C57BL / 6 mice aged 6 - 8 weeks (purchased from Zhejiang Vital River Laboratory Animal Technology Co., Ltd., license number: SCXK(Zhe)2020-0002), weighing 20 - 25 g, were raised in the SPF-level environment of the Animal Experiment Center of Henan University of Traditional Chinese Medicine. Raising environment: room temperature 24 ± 2 °C, humidity 50% - 60%, 12h light / 12h dark cycle. The mice were given free access to water and food, and after 1 week of adaptive feeding, they were grouped and modeled. All animal experiments in this invention were carried out under the guidance of the "Regulations on the Administration of Laboratory Animals" and approved by the Animal Center of Henan University of Traditional Chinese Medicine. (Animal ethics approval number: 202312043).

[0066] (2) The efficacy of Bacteroides monomorpha was observed in in vivo experiments on mice. Given that previous animal experiments by our research group showed that 3% DSS aqueous solution was the optimal concentration for establishing ulcerative colitis in mice, we also selected dextran sulfate sodium (DSS), a relatively classic colitis model, for drug intervention and efficacy evaluation.

[0067] After adaptive feeding, the mice were weighed and numbered. Following the principle of balanced weight distribution, 30 mice were selected from 100 mice and divided into three groups (n=10): a control group (ABX-Control), a model group (ABX-Model), and a Bacteroides uniformis group (ABX-B. uniformis). A bacterial depletion model was first established by gavage administration of a quadruple antibiotic cocktail for 5 days. Then, except for the control group, the other two groups were given 3% (w / v) DSS solution to induce acute UC for 7 days. Detailed experimental procedures are as follows... Figure 1 As shown in the figure, the body weight and DAI level of the constructed model group mice were measured. At the same time, HE staining was used to observe colonic pathological damage. The mice showed diarrhea, bloody stools, weight loss, and increased DAI. The colon showed pathological damage and increased levels of inflammatory factors, confirming the successful construction of the mouse UC model.

[0068] Seven days after establishing the mouse UC model, all three groups of mice had free access to pure water. In the Bacteroides monomorpha group, each mouse was administered Bacteroides monomorpha bacterial solution by gavage daily at a dose of 0.2 mL / 10g (based on mouse body weight), with a viable bacterial count of 9 × 10⁻⁶. 8 CFU / mL.

[0069] Experimental data were analyzed using Prism software. Quantitative data were expressed as mean ± standard deviation. One-way ANOVA was used when variances were homogeneous, and nonparametric tests were performed when variances were unequal. A p-value < 0.05 was considered statistically significant.

[0070] 2.3 Experimental Methods and Results

[0071] (1) Mouse disease activity index

[0072] The Disease Activity Index (DAI) consists of three indicators: weight change rate, stool characteristics, and fecal occult blood.

[0073] The DAI scoring criteria are shown in Table 1. The severity of clinical symptoms in mice was scored according to Table 1.

[0074] Table 1 DAI Scoring Criteria

[0075] Scoring Weight loss / % fecal state Blood in stool 0 0 normal Negative(-) 1 0～5 Between the two Between the two 2 5～10 semi-loose stool Occult blood (+) 3 10～15 Between the two Between the two 4 ≥15 loose stool Naked bloody stool

[0076] Note: Normal stool characteristics refer to formed stool; loose stool refers to pasty or semi-formed stool that does not stick to the anus; loose stool refers to watery stool that can stick to the anus.

[0077] The three evaluation indicators for DAI (Defecation Injury Assessment) were: weight change, stool shape, and fecal bleeding. The DAI score was calculated as the sum of the three scores divided by 3. DAI scores were assessed daily for 14 days, from the start of DSS induction to the end of the experiment. Results are shown below. Figure 2 .according to Figure 2 It was found that, through a comprehensive assessment of the disease activity index (DAI) of mice (including weight loss, stool characteristics, and rectal bleeding), the DAI score of the ABX-B. uniformis group was significantly lower than that of the ABX-DSS group (P<0.01). *Bacteroides monomorphosum* can reduce the disease activity index of ulcerative colitis.

[0078] (2) Weight change rate: From the start of DSS induction to the end of the experiment, a total of 14 days, the weight of mice in each group was monitored at fixed time points every day. Body weight (%) = weight on the day of the experiment / initial weight * 100%; percentage of weight loss = (initial weight - weight on the day of the experiment) / initial weight * 100%; see Figure 3 The experimental results showed that the body weight of mice in the ABX-Control group exhibited a physiological growth trend. Compared with the ABX-Control group, the body weight of mice in the ABX-DSS group decreased significantly over time (P < 0.01). Compared with the ABX-DSS group, the body weight decrease in the ABX-B. uniformis group was significantly smaller (P < 0.05). Bacteroides monomorpha BNCC139204 can alleviate the body weight loss in mice with ulcerative colitis.

[0079] (3) Fecal occult blood test method: Aniline method for detecting fecal occult blood in mice. The method utilizes the peroxidase activity of heme in hemoglobin, which catalyzes the release of nascent oxygen from hydrogen peroxide, resulting in color development.

[0080] The detection method was as follows: mouse feces were collected daily and tested using a human urine-fecal occult blood test kit (benzidine method). The results are shown in the table below. Figures 4-5It was observed that mice in the ABX-Control group were active, had glossy fur, moved nimbly, gained weight slowly, had dry, brownish-red granules in their feces, had clean anal sphincter without redness or swelling, and consistently negative occult blood tests. Their food and water intake remained stable. Mice in the ABX-DSS group exhibited decreased activity, lethargy, anal sphincter redness and swelling, consistently positive occult blood tests, dull fur, and decreased appetite. Compared to the ABX-DSS group, mice in the ABX-B. uniformis group showed increased activity, improved mental state, improved fur luster, reduced anal sphincter redness and swelling, a significantly lower positive rate in occult blood tests, and increased food and water intake. The improvement in the ABX-B. uniformis group was significant.

[0081] (4) Colon examination

[0082] The first day after the termination of the drug intervention experiment, i.e., 24 days after the end of the drug intervention. h Mice in each group were anesthetized with urethane via intraperitoneal injection. Blood was collected by enucleation. Whole blood was collected in 1.5 mL centrifuge tubes, allowed to stand for 2 hours, and then centrifuged at 1200 rpm, 4℃ for 15 minutes. The serum was separated and stored at -80℃. After euthanasia by cervical dislocation, the mice were dissected, and the entire colon from the anus to the cecum was removed. The length of the colon was measured, recorded, and photographed. Figure 6 The differences in colon morphology among the different groups of mice were visually demonstrated. It was found that shortened colon length, positive fecal occult blood test, and perianal hematoma were typical pathological manifestations in UC mice. In the ABX-Control group, there was no significant change in colon length, negative fecal occult blood test, and no perianal hematoma. In the ABX-DSS group, colon length was significantly shortened (P<0.01), the positive rate of fecal occult blood test was high, and perianal hematoma was severe. The ABX-B. uniformis group showed significant improvement in colon morphology (P<0.01).

[0083] 2.3 Observation and scoring of colon tissue pathological sections

[0084] On the first day after the experiment, approximately 1 cm of the distal colon of mice was harvested, rinsed with physiological saline to remove contents, and placed in tissue fixative overnight. After thorough tissue fixation, it was dehydrated, embedded, sectioned, and stained according to standard procedures. The degree and depth of colonic inflammation, the area of ​​inflammatory infiltration, and crypt damage were observed and statistically scored. Figures 7-9Microscopic examination revealed that the colon tissue of mice in the ABX-Control group was structurally intact, with a smooth colonic mucosa, intact and regularly arranged intestinal glands in the lamina propria, abundant goblet cells with normal distribution and morphology, and no inflammatory cell infiltration. In contrast, the colon tissue of mice in the ABX-DSS group showed significant congestion and thickening, accompanied by ulceration. The mucosa and submucosa showed extensive inflammatory cell infiltration, atrophy and disordered arrangement of glands in the lamina propria, partial gland absence, reduced and morphologically altered goblet cells, and inflammatory cell infiltration. Compared to the ABX-DSS group, the ABX-B. uniformis group showed varying degrees of improvement in the colon tissue lesions of mice. In conclusion, Bacteroides monomorpha BNCC139204 can restore the integrity of the colonic mucosa, gradually normalize gland distribution, significantly alleviate inflammatory cell infiltration, and effectively control crypt abscesses and ulcer formation.

[0085] 2.4 ELISA detection of peripheral blood interleukin-18 (IL-18), interleukin-1β (IL-1β), and α-tumor necrosis factor (TNF-α) levels

[0086] On the first day after the experiment, blood was collected from each group of mice and centrifuged at 3500 rpm for 15 min at 4°C. The supernatant serum was collected, and the levels of IL-18, IL-1β, and TNF-α in serum and colon tissue were detected according to the ELISA kit instructions. TNF-α, IL-1β, and IL-18 are key pro-inflammatory factors in ulcerative colitis (UC), and their expression levels directly reflect the severity of intestinal inflammation. The expression levels of pro-inflammatory factors IL-1β, IL-18, and TNF-α in the serum of each group of mice were detected using ELISA. Figure 10 The results showed that, compared with the ABX-Control group, the levels of TNF-α, IL-1β, and IL-18 in the colon tissue of mice in the ABX-DSS group were significantly increased (P<0.01), indicating a significantly enhanced inflammatory response in the UC mouse model. Compared with the ABX-DSS group, the expression levels of TNF-α, IL-1β, IL-6, and IL-18 in the colon tissue of mice in the ABX-B. uniformis group were significantly decreased (P<0.01).

[0087] The expression levels of TNF-α, IL-1β, and IL-18 inflammatory cytokine genes in the colon tissue of UC mice were detected by qRT-PCR. Results are shown below. Figure 11It was found that, compared with the ABX-Control group, the expression levels of TNF-α, IL-1β, and IL-18 in the colon of mice in the ABX-DSS group were significantly increased, while the expression levels of TNF-α, IL-1β, and IL-18 in the ABX-B. uniformis group were decreased (P < 0.01). This indicates that Bacteroides monomorpha BNCC139204 can reduce the levels of IL-1β, IL-18, and TNF-α in mouse serum and colon tissue, and has an anti-inflammatory effect.

[0088] AB-PAS staining was performed on mouse colon tissue, and immunofluorescence PCR was used to detect the expression levels of genes and proteins related to the intestinal barrier Occludin and NLRP3 pathways. Figures 12-14 AB-PAS staining is a histological staining method used to detect mucus secretion and the integrity of the mucus layer in colonic tissue. In the Control group, goblet cells were densely distributed in the colonic crypts, and the mucus layer was intact and uniformly covered the epithelial surface, exhibiting a normal blue color, indicating that the mucus layer structure and function were intact. Compared with the Control group, the DSS group mice showed a significant reduction in the number of goblet cells and a decrease in mucus layer thickness in their colonic tissue. After intervention with Bacteroides monomorpha BNCC139204, the number of goblet cells in each treatment group showed a recovery trend.

[0089] It is known that Bacteroides monomorpha BNCC139204 can downregulate the activation of the NLRP3 inflammasome pathway (NLRP3 / ASC / Caspase-1) and reduce the expression of TNF-α in mouse colon tissue, thus having an anti-inflammatory effect.

[0090] Bacteroides monomorpha BNCC139204 can upregulate the expression of tight junction proteins (Occludin, ZO-1), reduce intestinal mucosal damage, and repair the intestinal barrier.

[0091] The levels of short-chain fatty acids in feces were detected using targeted metabolomics methods. The results are shown in [Figure number missing]. Figure 15 Studies have shown that Bacteroides monomorpha BNCC139204 can significantly increase the abundance of butyric acid-producing bacteria in the gut and upregulate butyric acid levels. Previous research has confirmed that butyric acid in the gut may alleviate colitis in mice.

[0092] In summary, the Bacteroides monomorpha BNCC139204 of the present invention can significantly alleviate the symptoms of DSS-induced colitis in mice and has the effect of treating ulcerative colitis.

[0093] Although the above embodiments have provided a detailed description of the present invention, they are only some embodiments of the present invention, and not all embodiments. People can obtain other embodiments based on these embodiments without creative effort, and these embodiments all fall within the protection scope of the present invention.

Claims

1. The use of Bacteroides uniformis in the preparation of drugs for the prevention and / or treatment of ulcerative colitis.

2. The application according to claim 1, characterized in that, The monomorphobacteria includes the monomorphobacteria strain numbered BNCC139204.

3. The application according to claim 1, characterized in that, The application of the drug includes one or more of the following: 1) Relieves fecal occult blood in ulcerative colitis; 2) Reduce the disease activity index of ulcerative colitis; 3) Reverse the length of the colon in ulcerative colitis; 4) Reduce the expression levels of inflammatory factors in ulcerative colitis; 5) Relieves weight loss in patients with ulcerative colitis; 6) Pathological indicators for relieving ulcerative colitis.

4. The application according to claim 3, characterized in that, The inflammatory factors include one or more of IL-18, IL-1β, and TNF-α.

5. A microbial preparation for the prevention and / or treatment of ulcerative colitis, characterized in that, This includes Bacteroides uniformis and pharmaceutically acceptable excipients.

6. The microbial preparation according to claim 5, characterized in that, The application of the monomorphic Bacteroides includes bacterial suspension.

7. The microbial preparation according to claim 6, characterized in that, The method for preparing the bacterial solution includes: inoculating the Bacteroides monomorpha into Columbia blood agar medium and culturing it to obtain the bacterial solution.

8. The microbial preparation according to claim 7, characterized in that, The culture includes anaerobic culture.

9. The microbial preparation according to claim 5, characterized in that, The viable count of Bacteroides monomorpha in the microbial preparation is ≥1×10⁻⁶. 8 CFU / mL or ≥1×10 8 CFU / g.

10. The microbial preparation according to claim 5, characterized in that, The dosage forms of the microbial preparations include one or more of the following: oral dosage forms, injections, and suppositories.

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