Breast milk-derived bifidobacterium breve for relieving inflammatory bowel disease and application of breast milk-derived bifidobacterium breve

Abstract

The invention discloses breast milk-derived bifidobacterium breve for relieving inflammatory bowel diseases and application of the breast milk-derived bifidobacterium breve. The bifidobacterium breve is preserved in the China General Microbiological Culture Collection Center (CGMCC), the preservation date is November 13, 2024, and the preservation number is CGMCC No. 32608. The bifidobacterium breve has the function of regulating the immunity of the intestinal tract, can effectively enhance the barrier of the intestinal tract, and plays a positive role in relieving the symptoms of colitis; compared with a traditional medicine for treating colitis, a medicine or a probiotic preparation prepared from the bifidobacterium breve has certain advantages and has a wide application prospect.

Description

Technical Field

[0001] The present application relates to the fields of microbial technology and pharmaceutical technology, and in particular to a strain of breast milk-derived Bifidobacterium breve for alleviating inflammatory bowel disease and a use thereof. Background Art

[0002] Breastfeeding has always been considered the most beneficial feeding method for infant health and development. More and more studies have confirmed that breastfeeding can improve the survival rate and health level of newborns and reduce the risk of infection and various developmental disorders. Breast milk is the most important source of infant intestinal flora. The rich microbial community in breast milk can be vertically transmitted to infants through lactation, among which bifidobacteria and lactic acid bacteria are the main probiotics. These probiotics have a positive effect on the growth and development, immune system and nervous system of infants, and can reduce the occurrence of allergic diseases, especially in building intestinal barriers and regulating immune responses.

[0003] Inflammatory bowel disease (IBD), including ulcerative colitis (UC) and Crohn's disease (CD), is a common digestive system disease. The exact pathogenesis of IBD is still unclear, and it is currently believed that several factors are involved, including dysregulated immune response, changes in intestinal microorganisms, genetic susceptibility, and environmental factors. The most common symptoms of IBD patients are abdominal pain, diarrhea, rectal bleeding, and tenesmus. Currently, aminosalicylic acid preparations can be used to treat mild to moderate IBD patients in clinical practice, but 30% of them have poor treatment effects. Although glucocorticoids can relieve symptoms in patients with moderate to severe IBD, they are not a long-term solution due to side effects. In the case of unsatisfactory treatment results, approximately 15% of IBD patients (within 20 years of diagnosis) must undergo colectomy. Therefore, people have been looking for new options for the effective treatment of IBD.

[0004] Given that one of the characteristics of IBD is the reduction in the diversity of the intestinal microbiota, many scientists have shifted their attention to probiotic therapy to restore the homeostasis of the intestinal microbiota. Probiotic therapy is a natural, safe and effective intervention for the treatment of gastrointestinal diseases. Among the many probiotics, bifidobacteria and lactic acid bacteria have particularly prominent probiotic functions. Previous studies have confirmed that Bifidobacterium breve has functions such as anti-infection, anti-depression, regulation of the host immune system, and promotion of host nutrient absorption. However, at this stage, there is no patent for the use of breast milk-derived Bifidobacterium breve to relieve colitis. Therefore, it is necessary to study the effect of a single breast milk-derived Bifidobacterium breve on relieving colitis. This not only provides a new perspective on Bifidobacterium breve in reducing intestinal inflammation, but also provides new ideas for the future treatment of IBD. Summary of the invention

[0005] Based on this, it is necessary to provide at least one strain of breast milk-derived Bifidobacterium breve for alleviating inflammatory bowel disease and its use.

[0006] In the first aspect of the present application, a strain of Bifidobacterium breve SHMB8001 is provided, which is deposited in the General Microbiological Center of China Microorganism Culture Collection Administration, with a deposit date of November 13, 2024 and a deposit number of CGMCC No.32608.

[0007] In some embodiments, the Bifidobacterium breve SHMB 8001 is derived from breast milk, and the 16S rDNA sequence of the Bifidobacterium breve SHMB 8001 comprises a nucleotide sequence as shown in SEQ ID NO: 1.

[0008] In a second aspect of the present application, a derivative of Bifidobacterium breve SHMB 8001 as described in the first aspect is provided, wherein the derivative is an inactivated cell, extract, culture, culture supernatant, fermentation product, fermentation supernatant or a combination thereof of Bifidobacterium breve SHMB 8001.

[0009] In a third aspect of the present application, a composition is provided, comprising a first component, wherein the first component comprises one or more of the Bifidobacterium breve SHMB 8001 as described in the first aspect and the derivative as described in the second aspect.

[0010] In some embodiments, the composition is a pharmaceutical composition, a food composition, or a feed composition.

[0011] In some embodiments, the number of viable bacteria of Bifidobacterium breve SHMB 8001 in the composition is not less than 1×10 9 CFU / g or 1×10 9 CFU / mL.

[0012] In some embodiments, the composition further comprises a second component; the second component comprises a probiotic, a postbiotic, a prebiotic, an antimicrobial agent, an immunomodulator, an anticancer agent, an inflammatory therapeutic agent, or a combination thereof.

[0013] In some embodiments, the composition further comprises a pharmaceutically acceptable carrier and / or excipient.

[0014] In a fourth aspect of the present application, there is provided use of the Bifidobacterium breve SHMB 8001 as described in the first aspect, the derivative as described in the second aspect, or the composition as described in the third aspect in the preparation of a medicament for treating inflammatory diseases.

[0015] In some embodiments, the inflammatory disease satisfies one or more of the following conditions (1)-(2):

[0016] (1) Inflammatory diseases associated with overexpression of one or more of the pro-inflammatory cytokines TNF-α, IL-1β and IL-6; and,

[0017] (2) selected from the group consisting of inflammatory bowel disease, irritable bowel syndrome, celiac disease, gastroenteritis, duodenitis, jejunitis, ileitis, peptic ulcer, pouchitis, Collin's ulcer, appendicitis, colitis, diverticular disease, adenocarcinoma, proctitis, mucositis, chemotherapy-related enteritis, radiation-related enteritis, short bowel disease, chronic diarrhea, familial adenomatous polyposis and perianal fistula; optionally, the inflammatory bowel disease is selected from the group consisting of Crohn's disease and ulcerative colitis.

[0018] In a fifth aspect of the present application, a method for culturing Bifidobacterium breve SHMB 8001 as described in the first aspect is provided, the method comprising:

[0019] The Bifidobacterium breve SHMB 8001 was cultured in a culture medium.

[0020] In some embodiments, the culturing of the Bifidobacterium breve SHMB 8001 satisfies one or more of the following conditions 1) to 3):

[0021] 1) The culture medium is an MRS liquid culture medium, and the MRS liquid culture medium optionally contains 0.5 g / L cysteine;

[0022] 2) cultured under anaerobic conditions; and,

[0023] 3) Incubate at 37°C.

[0024] The Bifidobacterium breve SHMB 8001 in the present application has at least the following advantages:

[0025] It has the function of regulating intestinal immunity, can effectively enhance the intestinal barrier, and plays a positive role in alleviating the symptoms of colitis. Animal experiments have shown that Bifidobacterium breve SHMB 8001 showed a protective effect on DSS-induced colitis in mice, which was manifested by significantly alleviating the degree of weight loss, disease activity index score, shortening of colon length and histological damage induced by DSS in mice. The content of tight junction proteins in colon tissue increased significantly, the levels of proinflammatory cytokines TNF-α, IL-1β and IL-6 decreased significantly, the diversity of intestinal flora increased significantly, and the disorder of flora was alleviated. The drugs or probiotic preparations prepared using Bifidobacterium breve SHMB 8001 provided in this application have certain advantages over traditional drugs for the treatment of colitis and have broad market prospects. BRIEF DESCRIPTION OF THE DRAWINGS

[0026] In order to more clearly illustrate the technical solutions in the implementation methods and examples of the present application and to more completely understand the present application and its beneficial effects, the following is a brief introduction to the drawings required for use in the description of the implementation methods or examples. Obviously, the drawings described below are only some embodiments of the present application. For those skilled in the art, other drawings can be obtained based on these drawings without creative work. It should also be noted that the drawings are all drawn in a simplified form and are only used to conveniently and clearly assist in explaining the present application.

[0027] Figure 1 The colony morphology of Bifidobacterium breve SHMB 8001 on MRS plate solid culture medium in one embodiment of the present application is shown.

[0028] Figure 2 The morphology of Bifidobacterium breve SHMB 8001 under 100× oil microscope after Gram staining in one embodiment of the present application.

[0029] Figure 3 FIG. 1 is a schematic diagram of the homologous evolutionary tree of Bifidobacterium breve SHMB 8001 in one embodiment of the present application.

[0030] Figure 4 This is the growth curve of Bifidobacterium breve SHMB 8001 in one embodiment of the present application.

[0031] Figure 5 This is the hemolytic detection of Bifidobacterium breve SHMB 8001 in one embodiment of the present application.

[0032] Figure 6 The figure shows the changes in body weight of mice in different groups during DSS modeling in one embodiment of the present application.

[0033] Figure 7 This is the change in the disease activity index DAI index of different groups of mice in one embodiment of the present application.

[0034] Figure 8 The colon lengths of mice in different groups in one embodiment of the present application.

[0035] Fig. 9 This is H&E staining of colon tissues of different groups of mice in one embodiment of the present application.

[0036] Fig.10 This is the histopathological score of mice in different groups in one embodiment of the present application.

[0037] Fig.11 The content of Claudin-1 (CLDN1) in the colon tissue of different groups of mice in one embodiment of the present application.

[0038] Fig.12 The content of occludin in the colon tissue of different groups of mice in one embodiment of the present application.

[0039] Fig.13 It is the content of TNF-α in the colon tissue of different groups of mice in one embodiment of the present application.

[0040] Fig.14 It is the content of IL-1β in the colon tissue of different groups of mice in one embodiment of the present application.

[0041] Fig.15 The content of IL-6 in the colon tissue of different groups of mice in one embodiment of the present application.

[0042] Fig.16 It is the Chao diversity index of the intestinal flora of different groups of mice in one embodiment of the present application.

[0043] Fig.17 It is the ACE diversity index of the intestinal flora of different groups of mice in one embodiment of the present application.

[0044] Fig.18 It is the Shannon diversity index of the intestinal flora of different groups of mice in one embodiment of the present application.

[0045] Fig.19 This is the principal coordinate analysis of the β-diversity index of the intestinal flora of different groups of mice in one embodiment of the present application.

[0046] Figure 1-Figure 19 In the table, “*”, “**” and “***” all indicate significant differences compared with the DSS group. The more * there are, the greater the significant difference is. The error is presented in the form of Mean±SEM (mean±standard error).

[0047] A strain of Bifidobacterium breve SHMB 8001, taxonomically named Bifidobacterium breve, was deposited in the General Microbiology Center of China Culture Collection Administration on November 13, 2024, with the deposit number CGMCC No.32608, and the deposit address is the Institute of Microbiology, Chinese Academy of Sciences, No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing. DETAILED DESCRIPTION

[0048] In order to facilitate the understanding of the present application, the present application will be described more fully below with reference to the relevant drawings. The preferred embodiments of the present application are given in the drawings. However, the present application can be implemented in many different forms and is not limited to the embodiments described herein. On the contrary, the purpose of providing these embodiments is to make the understanding of the disclosure of the present application more thorough and comprehensive.

[0049] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as those commonly understood by those skilled in the art to which this application belongs. The terms used herein in the specification of this application are only for the purpose of describing specific embodiments and are not intended to limit this application. The term "and / or" used herein includes any and all combinations of one or more of the related listed items.

[0050] In this application, unless otherwise specified, "one or more" means any one of the listed items or any combination of the listed items. Similarly, "one or more" and the like, when used in other ways to mean "one or more", are also understood in the same way unless otherwise specified.

[0051] The terms "combination thereof", "any combination thereof", "any combination thereof" and the like used in this application include all suitable combinations of any two or more of the listed items.

[0052] In this application, the word "suitable" in "suitable combination", "suitable method", "any suitable method", etc. is based on the ability to implement the technical solution of this application, solve the technical problems of this application, and achieve the expected technical effects of this application.

[0053] In this application, "further", "further", "particularly", "for example", "such as", "example", "for example", etc. are used for descriptive purposes, indicating that the previous and subsequent different technical solutions are related in terms of the content covered, but should not be understood as a limitation on the previous technical solution, nor can it be understood as a limitation on the protection scope of this application. In this application, unless otherwise specified, A (such as B) means that B is a non-limiting example of A, and it can be understood that A is not limited to B.

[0054] In this application, "optionally", "optional", and "optional" mean optional, that is, any one of the two parallel solutions of "yes" or "no". If there are multiple "optional" in a technical solution, unless otherwise specified and there is no contradiction or mutual restriction, each "optional" is independent. Unless otherwise specified, the descriptions of "optionally include", "optionally contain", etc. in this application, taking "optionally include" as an example, mean "may include or not include".

[0055] The terms "contain", "include" and "include" used in this application are synonymous, which are inclusive or open-ended and do not exclude additional, uncited members or features. Members or features include materials or components, structures, elements, instruments, etc.; non-limiting examples of members or features include actions, conditions for the occurrence of actions, timing, states, etc.

[0056] In this application, the technical features or technical solutions described in an open language include closed technical features or technical solutions composed of the listed contents, and also include open technical features or technical solutions containing the listed contents.

[0057] In the present application, exemplary descriptions such as "in some implementation modes (or examples)" and "in one implementation mode (or example)" may include but are not limited to the following meanings: these solutions may be combined with other solutions in a suitable manner to form new technical solutions.

[0058] In the present application, in "the first aspect", "the second aspect", "the third aspect", "the fourth aspect", etc., the terms "first", "second", "third", "fourth", etc. are used only for descriptive purposes and cannot be understood as indicating or implying relative importance or quantity, nor can they be understood as implicitly indicating the importance or quantity of the indicated technical features. Moreover, "first", "second", "third", "fourth", etc. only serve the purpose of non-exhaustive enumeration and description, and it should be understood that they do not constitute a closed limitation on quantity.

[0059] In the present application, when it comes to a numerical interval (i.e., a numerical range), unless otherwise specified, the distribution of the optional numerical values ​​in the numerical interval is considered to be continuous, and includes the two numerical endpoints (i.e., the minimum and maximum values) of the numerical interval, and each numerical value between the two numerical endpoints. Unless otherwise specified, when the numerical interval only refers to an integer in the numerical interval, it includes the two endpoint integers of the numerical range, and each integer between the two endpoints, which is equivalent to directly listing each integer. When multiple numerical ranges are provided to describe features or characteristics, these numerical ranges can be merged. In other words, unless otherwise specified, the numerical range disclosed herein should be understood to include any and all sub-ranges included therein. The "numerical value" in the numerical interval can be any quantitative value, such as a number, a percentage, a ratio, etc. "Numerical interval" allows broadly including numerical interval types such as percentage intervals, ratio intervals, and ratio intervals.

[0060] In this application, if there are multiple steps involved in the method flow, unless there is a clear different description in this document, there is no strict order restriction for the execution of these steps, and they can be executed in other orders than described. Moreover, any step can include multiple sub-steps or multiple stages, and these sub-steps or stages are not necessarily executed at the same time, but can be executed at different times, and their execution order is not necessarily sequential, but can be executed in turn, alternating or simultaneously with other steps or parts of sub-steps or stages of other steps.

[0061] The Bifidobacterium breve SHMB 8001 provided in this application is isolated from breast milk samples of healthy mothers in the breast milk bank of Shanghai Children's Hospital. The strain has no toxic side effects such as hemolysis on humans. Experiments have found that the Bifidobacterium breve can significantly alleviate the weight loss, disease activity index score, and shortening of colon length induced by DSS in mice; intervention with Bifidobacterium breve SHMB 8001 can significantly reduce the damage to colon tissue; intervention with Bifidobacterium breve SHMB 8001 can significantly inhibit the reduction of tight junction proteins Claudin-1 and Occludin; intervention with Bifidobacterium breve SHMB8001 can significantly reduce the concentrations of proinflammatory cytokines TNF-α, IL-1β, and IL-6; it can significantly reduce the concentrations of proinflammatory cytokines TNF-α, IL-1β, and IL-6, can significantly improve the diversity of intestinal flora, and alleviate the disorder of intestinal flora. Therefore, the drug prepared using the Bifidobacterium breve SHMB 8001 shows certain advantages over conventional drugs for treating colitis, and the strain can be used to prepare probiotic preparations, etc., and has broad application prospects.

[0062] The present application provides a strain of Bifidobacterium breve SHMB 8001, which is deposited in the General Microbiology Center of China Microorganism Culture Collection Administration, with a deposit date of November 13, 2024 and a deposit number of CGMCC No.32608.

[0063] In some embodiments, the Bifidobacterium breve SHMB 8001 is derived from breast milk.

[0064] In some embodiments, the 16S rDNA sequence of Bifidobacterium breve SHMB 8001 comprises the nucleotide sequence shown in SEQ ID NO: 1.

[0065] In some embodiments, the Bifidobacterium breve SHMB 8001 has at least one or more of the following characteristics:

[0066] (1) The colonies on MRS solid medium are milky white, round, convex, smooth-edged, and sticky;

[0067] (2) After Gram staining, it appears purple under the microscope and may appear as a short rod, Y-shaped rod, or V-shaped rod;

[0068] (3) Helps prevent and / or treat colitis.

[0069] The present application also provides derivatives of Bifidobacterium breve SHMB 8001 as described above.

[0070] In some embodiments, the derivative is an inactivated cell, an extract, a culture, a culture supernatant, a fermentation product, a fermentation supernatant or a combination thereof of the Bifidobacterium breve SHMB 8001.

[0071] The present application also provides a composition comprising a first component, wherein the first component comprises one or more of the above-mentioned Bifidobacterium breve SHMB 8001 and the derivatives.

[0072] The composition in the present application may further include a second component; the second component includes probiotics, postbiotics, prebiotics, antimicrobial agents, immunomodulators, anticancer agents, inflammatory therapeutic agents or a combination thereof.

[0073] The composition described in the present application may be a composition in a common form in the art, for example, a pharmaceutical composition, a food composition, a feed composition, etc.

[0074] In some embodiments, the pharmaceutical composition further comprises other drugs that can be used to alleviate inflammatory bowel disease. Exemplarily, the pharmaceutical composition further comprises mesalazine.

[0075] In some embodiments, the pharmaceutical composition is a drug.

[0076] In some embodiments, the food composition is a food composition for special medical purposes.

[0077] In some embodiments, the food composition may be a fermented food or a fermented beverage.

[0078] In some embodiments, the composition is a solid composition. In some embodiments, the amount of Bifidobacterium breve SHMB 8001 in the solid composition is not less than 1×10 9 CFU / g.

[0079] In some embodiments, the composition is a liquid composition. In some embodiments, the amount of Bifidobacterium breve SHMB 8001 in the liquid composition is not less than 1×10 9 CFU / mL.

[0080] In some embodiments, the composition further comprises a pharmaceutically acceptable carrier and / or excipient.

[0081] The term "pharmaceutically acceptable" means that the specified carrier, vehicle, diluent, excipient and / or salt is generally chemically and / or physically compatible with the other ingredients comprising the formulation, and physiologically compatible with the recipient thereof.

[0082] As used in this application, the term "pharmaceutically acceptable carrier and / or excipient" refers to a carrier and / or excipient that is pharmacologically and / or physiologically compatible with a subject and an active ingredient, which is well known in the art (see, e.g., Remington's Pharmaceutical Sciences. Edited by Gennaro AR, 19th ed. Pennsylvania: Mack Publishing Company, 1995), and includes, but is not limited to, pH regulators, surfactants, ionic strength enhancers, agents that maintain osmotic pressure, agents that delay absorption, diluents, preservatives, stabilizers, and the like. For example, pH regulators include, but are not limited to, phosphate buffers. Surfactants include, but are not limited to, cationic, anionic, or nonionic surfactants, such as Tween-80. Ionic strength enhancers include, but are not limited to, sodium chloride. Agents that maintain osmotic pressure include, but are not limited to, sugars, NaCl, and the like. Agents that delay absorption include, but are not limited to, monostearate and gelatin. Diluents include, but are not limited to, water, aqueous buffers (such as buffered saline), alcohols, and polyols (such as glycerol), and the like. Preservatives include, but are not limited to, various antibacterial and antifungal agents, such as thimerosal, 2-phenoxyethanol, parabens, chlorobutanol, phenol, sorbic acid, etc. Stabilizers have the meanings generally understood by those skilled in the art, and are capable of stabilizing the desired activity of the active ingredient in the drug, including, but not limited to, sodium glutamate, gelatin, SPGA, sugars (such as sorbitol, mannitol, starch, sucrose, lactose, dextran, or glucose), amino acids (such as glutamic acid, glycine), proteins (such as dried whey, albumin or casein) or their degradation products (such as lactalbumin hydrolysate), etc.

[0083] The name or form of the composition may be determined according to the application scenario and specific purpose of the composition, etc. For example, the composition may be a liquid or solid microbial preparation, which can be used to balance intestinal flora.

[0084] In some embodiments, the composition is a microbial preparation. Exemplarily, the preparation method of the microorganism comprises:

[0085] The seed liquid of the Bifidobacterium breve SHMB 8001 is inoculated into an MRS liquid culture medium (containing 0.5 g / L cysteine) at a certain inoculation amount of 0.5% (v / v)-5% (v / v) (for example, 2% (v / v)), and anaerobically cultured at 37° C. for a period of time (for example, 24 hours) to obtain a culture solution; the culture solution is centrifuged to obtain bacterial cells, and the bacterial cells are washed 3 times with physiological saline and then resuspended to obtain a microbial preparation.

[0086] The present application also provides a method for preparing a microbial preparation, which comprises, for example: inoculating the seed liquid of the Bifidobacterium breve SHMB 8001 into an MRS liquid culture medium containing cysteine ​​(e.g., cysteine ​​hydrochloride) at a certain inoculation amount (e.g., 2% (v / v)), and culturing anaerobically at 37° C. to obtain a culture solution; centrifuging the culture solution to obtain bacterial cells, washing the bacterial cells with physiological saline and then resuspending them to obtain a microbial preparation.

[0087] The concentration of cysteine ​​in the MRS liquid culture medium may be 0.5 g / L. The culture time may be determined according to the proliferation law of Bifidobacterium breve SHMB 8001 and the concentration requirement of the microbial preparation. For example, the culture may be performed for more than 24 hours. In addition, the number of times of washing with physiological saline may be, for example, more than 3 times.

[0088] In some embodiments, the microbial preparation can be further used to prepare fermented food or fermented beverage.

[0089] The present application also provides use of the Bifidobacterium breve SHMB 8001, derivatives or compositions described above in the preparation of a medicament for alleviating inflammatory diseases.

[0090] In this application, "alleviate" and "prevent and treat" have the same meaning. It should be understood that "prevent and treat" includes aspects such as prevention, treatment, and adjuvant therapy. As used herein, "prevent and treat" refers to alleviating, delaying progression, attenuating, preventing, or maintaining an existing disease or condition. "Prevent and treat" also includes curing one or more symptoms of a disease or condition, preventing its development, or alleviating to a certain extent. In this application, "prevent and treat" and "prevent and / or treat", "prevent, treat, or prevent and treat" have the same meaning and can be used interchangeably.

[0091] In some embodiments, the relief of inflammatory bowel disease (e.g., colitis) includes at least one of the following functions:

[0092] (1) Alleviate the degree of weight loss;

[0093] (2) Reduce colon tissue damage;

[0094] (3) Increase the content of colon tight junction proteins Claudin-1 and Occludin;

[0095] (4) reduce the expression of at least one of the proinflammatory cytokines TNF-α, IL-1β, and IL-6;

[0096] (5) Improve intestinal flora diversity and / or improve intestinal flora disorders.

[0097] In some embodiments, the inflammatory disease is an inflammatory disease associated with overexpression of one or more of the proinflammatory cytokines TNF-α, IL-1β, and IL-6.

[0098] In some embodiments, the inflammatory disease is selected from the group consisting of inflammatory bowel disease, irritable bowel syndrome, celiac disease, gastroenteritis, duodenitis, jejunitis, ileitis, peptic ulcer, pouchitis, Collin's ulcer, appendicitis, colitis, diverticular disease, adenocarcinoma, proctitis, mucositis, chemotherapy-related enteritis, radiation-related enteritis, short bowel disease, chronic diarrhea, familial adenomatous polyposis, and perianal fistula.

[0099] In some embodiments, the inflammatory bowel disease is selected from the group consisting of Crohn's disease and ulcerative colitis.

[0100] In this application, "drug" includes any agent, compound, composition or mixture that provides physiological and / or pharmacological effects in vivo or in vitro, and often provides beneficial effects. The scope of the physiological and / or pharmacological effects of "drug" in vivo is not particularly limited, and it can be a systemic effect or only a local effect. The activity of the "drug" is not particularly limited, and it can be an active substance that can interact with other substances, or it can be an inert substance that does not interact.

[0101] The present application also provides a method for alleviating inflammatory diseases, which comprises administering an effective dose of the composition as described above to a subject.

[0102] "Administering," "giving," and "treating" when applied to animals, humans, experimental subjects, cells, tissues, organs, or biological fluids, refers to the contact of an exogenous drug, therapeutic agent, diagnostic agent, immunogenic agent, or composition with an animal, human, subject, cell, tissue, organ, or biological fluid. "Administering," "giving," and "treating" can refer to, for example, treatment, pharmacokinetics, diagnosis, research, and experimental methods. Treatment of cells includes contact of an agent with a cell, and contact of an agent with a fluid, wherein the fluid is in contact with the cell. "Administering," "giving," and "treating" also mean in vitro and ex vivo treatment of, for example, a cell by an agent, a diagnosis, a combination composition, or by another cell. "Treatment," when applied to humans, veterinary medicine, or research subjects, refers to therapeutic treatment, prophylactic or preventive measures, research and diagnostic applications. Some examples are provided below.

[0103] The present application also provides a method for culturing the Bifidobacterium breve SHMB 8001, the method comprising:

[0104] The Bifidobacterium breve SHMB 8001 was cultured in a culture medium.

[0105] The culture medium used may be a culture medium conventionally used in the art for culturing Bifidobacterium breve.

[0106] In some embodiments, the culture medium is MRS liquid medium.

[0107] Exemplarily, the MRS liquid culture medium contains: 10 g / L tryptone, 10 g / L beef extract, 5 g / L yeast powder, 20 g / L glucose, 2 g / L anhydrous sodium acetate, 0.25 g / L manganese sulfate monohydrate, 2 g / L diammonium hydrogen citrate, 2.6 g / L dipotassium hydrogen phosphate trihydrate, 0.5 g / L magnesium sulfate heptahydrate, 1 mL / L Tween80, and 0.5 g / L cysteine ​​hydrochloride.

[0108] In some embodiments, the culture medium is a MRS liquid culture medium containing cysteine. In some embodiments, the concentration of cysteine ​​in the MRS liquid culture medium is 0.5 g / L.

[0109] In some embodiments, the Bifidobacterium breve SHMB 8001 is cultured under anaerobic conditions.

[0110] In some embodiments, the culture temperature is set at about 37°C.

[0111] Some examples are provided below.

[0112] The embodiments of the present application will be described in detail below in conjunction with examples. It should be understood that these examples are only used to illustrate the present application and are not intended to limit the scope of the present application. The experimental methods for which the conditions are not specified in the following examples are preferably referred to the guidance given in the present application, and can also be based on the experimental manual or normal conditions in the art, or can also be based on the conditions recommended by the manufacturer, or with reference to experimental methods known in the art.

[0113] The culture medium involved in the following examples is as follows:

[0114] MRS solid medium: tryptone 10 g / L, beef extract 10 g / L, yeast powder 5 g / L, glucose 20 g / L, anhydrous sodium acetate 2 g / L, manganese sulfate monohydrate 0.25 g / L, diammonium hydrogen citrate 2 g / L, dipotassium hydrogen phosphate trihydrate 2.6 g / L, magnesium sulfate heptahydrate 0.5 g / L, Tween80 1 mL / L, cysteine ​​hydrochloride 0.5 g / L, and agar 20 g / L.

[0115] MRS liquid medium: tryptone 10 g / L, beef extract 10 g / L, yeast powder 5 g / L, glucose 20 g / L, anhydrous sodium acetate 2 g / L, manganese sulfate monohydrate 0.25 g / L, diammonium hydrogen citrate 2 g / L, dipotassium hydrogen phosphate trihydrate 2.6 g / L, magnesium sulfate heptahydrate 0.5 g / L, Tween80 1 mL / L, and cysteine ​​hydrochloride 0.5 g / L.

[0116] The mice involved in the following examples are 6-week-old to 8-week-old female SPF (Specific pathogen free) grade C57BL / 6J mice purchased from Jicui Pharmaceutical Company.

[0117] Example 1: Isolation, identification and preservation of breast milk-derived Bifidobacterium breve SHMB 8001

[0118] The details include:

[0119] (1) Collection of breast milk samples: First, the mother who donates milk needs to clean the nipple and the surrounding area with sterile water. Then wear sterile gloves to collect breast milk and store it in a sterile milk storage bottle. After collection, the breast milk sample should be immediately placed in a refrigerator at 4°C and cultured within 24 hours (h).

[0120] (2) Isolation of breast milk-derived Bifidobacterium breve SHMB 8001: Take 50 μL-100 μL of breast milk sample from healthy mothers from the breast milk bank of Shanghai Children's Hospital, spread it on MRS solid medium (containing 0.5 g / L cysteine), and culture it anaerobically at 37°C for 72 h. Observe and record the colony morphology. Figure 1 As shown, a milky white, smooth, round and protruding colony was streaked on MRS solid medium (containing 0.5 g / L cysteine) and purified under anaerobic conditions at 37°C. This operation was repeated 3 times to obtain a purified single colony. The purified colony was inoculated into MRS liquid medium (containing 0.5 g / L cysteine) and cultured anaerobically at 37°C for 36 h-48 h to obtain strain SHMB 8001.

[0121] (3) Identification of human milk-derived Bifidobacterium breve SHMB 8001: Figure 2 As shown, it appears purple under the microscope after Gram staining, and the morphology may be short rods, Y-shaped rods or V-shaped rods. The strains identified by Gram staining were extracted with a bacterial DNA extraction kit (Tian Gen, catalog number: #DP302-02) and amplified and sequenced. The nucleotide sequence of the amplified 16S rDNA is shown in SEQ ID NO: 1:

[0122] GGGCGGGTGCTACCATGCAGTCGAACGGGATCCATCGGGCTTTGCCTGGTGGTGAGAGTGGCGAACGGGTGAGTAATGCGTGACCGACCTGCCCCATGCACCGGAATAGCTCCTGGAAACGGGTGGTAATGCCGGATGCTCCATCACACCGCATGGTGTGTTGGGAAAGCCTTTGCGGCATGGGATGGGGTCGCGTCCTATCAGCTTGATGGCGGGGTAACGGCCCACCATGGCTTCGACGGGTAGCCGGCCTGAGAGGGCGACCGGCCACATTGGGACTGAGATACGGCCCAGACTCCTACGGGAGGCAGCAGTGGGGAATATTGCACAATGGGCGCAAGCCTGATGCAGCGACGCCGCGTGAGGGATGGAGGCCTTCGGGTTGTAAACCTCTTTTGTTAGGGAGCAAGGCACTTTGTGTTGAGTGTACCTTTCGAATAAGCACCGGCTAACTACGTGCCAGCAGCCGCGGTAATACGTAGGGTGCAAGCGTTATCCGGAATTATTGGGCGTAAAGGGCTCGTAGGCGGTTCGTCGCGTCCGGTGTGAAAGTCCATCGCTTAACGGTGGATCCGCGCCGGGTACGGGCGGGCTTGAGTGCGGTAGGGGAGACTGGAATTCCCGGTGTAACGGTGGAATGTGTAGATATCGGGAAGAACACCAATGGCGAAGGCAGGTCTCTGGGCCGTTACTGACGCTGAGGAGCGAAAGCGTGGGGAGCGAACAGGATTAGATACCCTGGTAGTCCACGCCGTAAACGGTGGATGCTGGATGTGGGGCCGTTCCACGGGTTCCGTGTCGGAGCTAACGCGTTAGCATCCCGCCTGGGGGAGTACGGCCGCAAGGCTAAAACTCAAAGAAATTGGCGGGGGCCCGCCAAACGGGGGAGGATGCGGGTTAATTCGAGGGAACGCGAGAAACTTTCCTGGGGTTGAATGTTT。The obtained sequences were compared with NCBI-BLAST, and the strain was assigned to Bifidobacterium breve and named Bifidobacterium breve SHMB 8001. The evolutionary tree was drawn using MEGA 11 software, such as. Figure 3 As shown in the figure, strain SHMB 8001 and Bifidobacterium breve DSM 20213 = JCM 1192 clustered together, so it can be confirmed that strain SHMB 8001 and the type strain are the same species, Bifidobacterium breve.

[0123] (4) Storage of breast milk-derived Bifidobacterium breve SHMB 8001: Pick a single colony of Bifidobacterium breve SHMB 8001 and inoculate it into MRS liquid culture medium (containing 0.5 g / L cysteine), and culture it anaerobically at 37 °C for 36 h to obtain a bacterial solution. Place the bacterial solution in a cryopreservation tube under sterile conditions at 6000 r / min and centrifuge for 3 min to collect the bacteria. Wash with sterile PBS buffer solution, centrifuge at 6000 r / min for 3 min to obtain the washed bacteria, and repeat this operation 3 times. Resuspend the bacteria with sterile 30% (v / v) glycerol and store the cryopreservation tube in a -80 °C refrigerator.

[0124] Example 2: Preparation of Bifidobacterium breve SHMB 8001 bacterial suspension

[0125] (1) Pipette 10 μL of the bacterial solution of Bifidobacterium breve SHMB 8001 from the cryopreservation tube and streak it on MRS solid medium (containing 0.5 g / L cysteine), and culture it anaerobically at 37 ℃ for 48 h to obtain a single colony. Pick a single colony and inoculate it into MRS liquid medium (containing 0.5 g / L cysteine), and culture it anaerobically at 37 ℃ for 36 h to obtain an activated solution. Inoculate the activated solution into MRS liquid medium (containing 0.5 g / L cysteine) at an inoculum volume of 2% (v / v), and culture it anaerobically at 37 ℃ for 24 h. Repeat the activation culture three times to obtain an activated bacterial solution.

[0126] (2) The activated bacterial solution was inoculated into MRS liquid medium (containing 0.5 g / L cysteine) at an inoculum volume of 2% (v / v) and cultured at 37 °C for 24 h to obtain the culture medium. The culture medium was centrifuged to obtain the bacterial cells, which were washed three times with physiological saline and resuspended, and the viable cell count was adjusted to 1×10 9 CFU / mL to obtain bacterial suspension.

[0127] Example 3: In vitro probiotic activity test of Bifidobacterium breve SHMB 8001

[0128] This example tests the in vitro beneficial activity of Bifidobacterium breve SHMB 8001, including growth curve, antimicrobial sensitivity and hemolytic activity tests, specifically including the following:

[0129] (1) Growth curve detection: The activated bacterial solution in Example 2 was inoculated into 200 mL of MRS liquid culture medium (containing 0.5 g / L cysteine) at an inoculum volume of 2% (v / v). After mixing, 10 mL of the culture was dispensed into test tubes and anaerobically cultured at 37 °C for 0 h, 3 h, 5 h, 8 h, 10 h, 15 h, 20 h, 24 h, 36 h, 48 h and 72 h. The absorbance OD of the culture at 600 nm was measured at each time point. 600 (and deduct the OD of the culture medium of the control group 600 MRS liquid culture medium (containing 0.5 g / L cysteine) was used as blank control, and the culture time was used as the horizontal axis, and OD 600 As the vertical axis, the growth curve of Bifidobacterium breve SHMB 8001 was drawn

[0130] The growth curve drawn is as follows Figure 4 As shown in the figure, there is no obvious change in the OD value of the bacterial solution from 0 h to 8 h, and the strain is in the growth lag phase to adapt to the new environment. The OD value of the bacterial solution increases logarithmically from 8 h to 24 h, which is the logarithmic growth phase of the strain. During this period, the strain has vigorous metabolism, the strongest ability to resist adverse environments, and the morphology and physiological characteristics of the cell population are most consistent. The OD value of the bacterial solution is basically stable from 24 h to 72 h, entering the plateau phase of strain growth, and the cell metabolic activity gradually decreases.

[0131] (2) Detection of Antimicrobial Sensitivity: Determining the sensitivity of probiotics to antimicrobial drugs is an important indicator for measuring their safety. This example adopts the method recommended by the International Dairy Federation standard document (ISO 10932 / IDF 233 standard, 2010). Before the experiment, antibiotic microdilution plates were prepared using 96-well plates, with 50 μL of antibiotics of different concentration gradients placed in each well, including aminoglycoside gentamicin 0.5 μg / mL-256 μg / mL, macrolide erythromycin 0.016 μg / mL-8 μg / mL, lincosamide clindamycin 0.032 μg / mL-16 μg / mL, β-lactam ampicillin 0.032 μg / mL-16 μg / mL, glycopeptide vancomycin 0.25 μg / mL-128 μg / mL, quinolone levofloxacin 0.25 μg / mL-128 μg / mL, and anti-tuberculosis drug rifampicin 0.125 μg / mL-64 μg / mL. After preparation, the plates were stored at -20 ℃. Pick a single colony cultured for 48 h from the MRS solid medium and transfer it to a transparent glass tube containing 2 mL of saline solution. Adjust the concentration of the bacterial solution until the transmittance reaches the McFarland standard of 1.0. Dilute the above-mentioned saline solution containing bacteria 500 times into LSM liquid medium (90% IST liquid medium (Shandong Top, Product No. M8120) + 10% MRS liquid medium). Thaw the antibiotic microdilution plate prepared in advance at room temperature, inoculate 50 μL of the above-mentioned LSM liquid medium containing bacteria in each well, and determine the minimum inhibitory concentration (MIC) and epidemiological cut-off value (ECOFF) after anaerobic culture at 37°C for 48 h.

[0132] Table 1: Test results of antimicrobial susceptibility of Bifidobacterium breve SHMB 8001

[0133]

[0134] As shown in Table 1 , Bifidobacterium breve SHMB 8001 was sensitive to antimicrobial drugs including gentamicin, erythromycin, clindamycin, ampicillin, vancomycin, levofloxacin and rifampicin.

[0135] (3) Hemolytic detection: 10 μL of the bacterial solution of Bifidobacterium breve SHMB 8001 was taken and streaked onto Columbia blood agar medium (Bickman Biotech) containing 5% (v / v) defibrinated sheep blood. The culture was then anaerobically cultured at 37°C for 48 h. The hemolytic characteristics of the strain were determined based on the following changes: if it is α-hemolytic, a 1 mm-2 mm grass-green, translucent hemolytic ring can be observed around the colonies, indicating an incomplete hemolytic phenomenon; if it is β-hemolytic, a 2 mm-4 mm clearly defined, completely transparent hemolytic ring can be observed around the colonies, indicating a complete hemolytic phenomenon; if it is γ-hemolytic, there is no hemolytic phenomenon around the colonies, indicating incomplete hemolysis.

[0136] like Figure 5 As shown, after Bifidobacterium breve SHMB 8001 was cultured anaerobically at 37°C for 48 hours on Columbia blood agar medium, no hemolysis was observed around the colonies, indicating γ-hemolysis, that is, Bifidobacterium breve SHMB 8001 was not hemolytic.

[0137] Example 4: Effect of Bifidobacterium breve SHMB 8001 on the symptoms of DSS-induced colitis in mice

[0138] The DSS-induced colitis modeling method in mice and the intervention steps of Bifidobacterium breve SHMB 8001 are as follows:

[0139] (1) Prepare 3.0% dextran sulfate sodium (DSS) solution: Prepare 3.0% dextran sulfate sodium (DSS) solution with sterile water.

[0140] (2) Animal Experiment Design: Twenty healthy female C57BL / 6J mice aged 6 to 8 weeks were randomly divided into 4 groups, 5 mice in each group: blank control group, DSS group, Bifidobacterium breve SHMB 8001 intervention group (SHMB 8001+DSS) and mesalazine drug intervention group. The Bifidobacterium breve SHMB 8001 bacterial suspension was prepared according to the method of Example 2. The experimental scheme and the treatment methods of each group of mice are shown in Table 2.

[0141] Table 2: Experimental plan and treatment of mice in each group

[0142]

[0143] Note: "Day-X" means "Day X", for example, "Day-0" means Day 0. "mg / kg / d" means the number of milligrams given per kg of body weight per day.

[0144] (3) Analysis method of test results:

[0145] Disease activity index (DAI): The score was based on the weight change score of the mice (0% decrease: 0 points; 1%-5% decrease: 1 point; 6%-10% decrease: 2 points; 11%-18% decrease: 3 points; decrease >18%: 4 points), stool characteristics score (normal: 0 points; soft but formed: 1 point; loose: 2 points; very loose and moist: 3 points; watery stool: 4 points) and blood in stool score (no occult blood: 0 points; occult blood positive: 2 points; mild gross blood in stool: 3 points; severe gross blood in stool: 4 points). DAI = weight change score + stool characteristics score + blood in stool score, with a total score of 12 points. The higher the score, the more severe the colitis.

[0146] Histological assessment: Scoring was based on the mouse colon tissue damage score (no damage: 0 points; isolated focal epithelial damage: 1 point; mucosal erosion and ulceration: 2 points; extensive damage across the intestinal wall: 3 points) and the lamina propria inflammatory cell infiltration score (rare: 0 points; some neutrophil infiltration: 1 point; submucosal inflammatory cell clusters: 2 points; transmural cell infiltration: 3 points). Histopathological score = tissue damage score + lamina propria inflammatory cell infiltration score. The higher the score, the more severe the colitis. Refer to the relevant literature (Wirtz S, Popp V, Kindermann M, et al. Chemically induced mouse models of acute and chronic intestinal inflammation. Nat Protoc. 2017; 12(7): 1295-1309. doi: 10.1038 / nprot.2017.044).

[0147] (4) During the modeling period, the body weight of each group of mice was measured every day (the results are shown in Figure 6 ) and the disease activity index DAI (results as Figure 7 After the model was established, the mice were killed, and the entire colon (from the end of the cecum to the anus) was taken to measure the length of the colon (the results are shown in Figure 8 as shown).

[0148] Depend on Figure 6 It can be seen that on the 7th day of modeling, the average body weight of mice in the DSS group decreased by 22.18%, while the average body weight of mice in the Bifidobacterium breve SHMB8001 intervention group and the mesalazine drug intervention group decreased by 9.04% and 8.02%, respectively, indicating that both Bifidobacterium breve SHMB 8001 and mesalazine can alleviate the weight loss of mice caused by DSS modeling, and the degree of relief in the two groups is similar.

[0149] Depend on Figure 7It can be seen that on the 7th day of modeling, the DAI index of mice in the DSS group reached a full score of 12.0, while the DAI index of mice in the Bifidobacterium breve SHMB8001 intervention group and the mesalazine drug intervention group was significantly lower than that in the DSS group, at only 6.4 and 6.2.

[0150] Depend on Figure 8 It can be seen that the average colon length of mice in the blank control group was 6.1 cm, and the average colon length of mice in the DSS group was only 4.1 cm. However, the intervention of Bifidobacterium breve SHMB 8001 and mesalazine significantly alleviated the shortening of colon length caused by DSS, and the average colon length reached 4.8 cm and 4.7 cm, respectively.

[0151] (5) After the mice were killed, 0.5 cm of the distal colon tissue was taken for fixation, dehydration, embedding, and HE staining. The HE staining of the colon tissue of mice in different groups was observed. Fig. 9 As shown in the figure, the colon tissue of mice in the DSS group showed obvious damage, including severe inflammatory cell infiltration, destruction of crypt and glandular structures, disappearance of goblet cells, and edema of the submucosal layer. The colon tissue structure of the Bifidobacterium breve SHMB 8001 intervention group was similar to that of the mesalazine intervention group, close to the blank control group, with relatively complete glands and crypts, more goblet cells, and only a small amount of inflammatory cell infiltration and submucosal edema.

[0152] Histopathological scoring Fig.10 As shown, the pathological score of the DSS group reached 4.6 points, while the scores of the Bifidobacterium breve SHMB8001 intervention group and the mesalazine intervention group were significantly lower than that of the DSS group, at 3.0 points and 2.6 points, respectively.

[0153] The above results show that the Bifidobacterium breve SHMB 8001 of the present application has a good alleviating effect on the symptoms of DSS-induced colitis in mice, and its alleviating effect is close to that of the traditional clinical drug mesalazine.

[0154] Example 5: Effect of Bifidobacterium breve SHMB 8001 on the content of intestinal tight junction proteins in colitis mice

[0155] The content of intestinal tight junction protein was determined by Western blot analysis: 300 μL of RIPA lysis buffer, 3 μL of phenylmethylsulfonylfuoride (PMSF) and 12 μL of 25× cocktail were mixed at a ratio of 100:1:4 to prepare protein lysis buffer. About 30 mg of tissue was weighed and placed in the prepared protein lysis buffer. The sample was homogenized for 30 s using a homogenizer. The homogenized protein was placed on ice for 30 min, then centrifuged at 4°C and 15000 rpm for 15 min, and the supernatant was taken. The protein concentration was detected using a BCA protein concentration quantification kit. The protein solution was mixed with 5× Loading buffer at a ratio of 4:1 and heated at 100°C for 15 min. The protein was separated by 10% SDS-polyacrylamide gel electrophoresis (SDS-PAGE), and the protein loading amount per well was set at 30 μg. The membrane was transferred using NC membrane at a constant current of 300 mA for 60 min, and blocked with 5% skim milk prepared in 1×PBST (phosphate buffer) for 1 h-2 h. The membrane was washed three times with 1×PBST, each time for 10 min, and then incubated with primary antibody at 4°C overnight. The membrane was washed three times with 1×PBST and then incubated with secondary antibody at 4°C for 1 h. β-actin was used as an internal reference, and horseradish peroxidase-conjugated β-actin antibody (HRP-Conjugated anti-β-actin, Qihe Biotechnology, Cat. No.: 103030004) was used to incubate at 4°C for 2 h. After washing the membrane three times with 1×PBST, the protein bands were detected using a fluorescent chemiluminescence imaging system (BIO-RAD, ChemiDoc Imaging Syst).

[0156] After the modeling in Example 4 was completed, the mice were killed, and the colon tissues were taken to detect the tight junction protein content in the supernatant of the colon tissues according to the Western blot analysis method, including the tight junction proteins Claudin-1 and Occludin.

[0157] like Fig.11 As shown, compared with the DSS group, both Bifidobacterium breve SHMB 8001 and mesalazine intervention were able to significantly increase the concentration of tight junction protein Claudin-1, among which the mesalazine group was closer to the blank control group.

[0158] like Fig.12As shown, compared with the DSS group, both Bifidobacterium breve SHMB 8001 and mesalazine intervention can significantly increase the concentration of tight junction protein Occludin, and the Bifidobacterium breve SHMB 8001 group and the mesalazine group are similar. It is proved that the Bifidobacterium breve SHMB 8001 intervention of the present application can strengthen the mucosal epithelial barrier and improve the colonic mechanical barrier of colitis mice.

[0159] Example 6: Effects of Bifidobacterium breve SHMB 8001 on cytokine levels in the colon of colitis mice

[0160] After the modeling in Example 4 was completed, the mice were killed, and the concentrations of colon tissue cytokines TNF-α, IL-1β and IL-6 were measured using a commercially available ELISA kit (Beijing Sizhengbai Biotechnology Co., Ltd.) according to the manufacturer's instructions.

[0161] like Figure 13-Figure 15 As shown in the results, DSS treatment significantly increased the concentrations of proinflammatory cytokines TNF-α, IL-1β, and IL-6 in colonic tissues, while both Bifidobacterium breve SHMB 8001 and mesalazine interventions significantly reduced the concentrations of TNF-α, IL-1β, and IL-6, and the concentrations of Bifidobacterium breve SHMB 8001 group and mesalazine group were similar.

[0162] Example 7: Effect of Bifidobacterium breve SHMB 8001 on the diversity of intestinal flora in colitis mice

[0163] The mice after modeling in Example 4 were killed, and the feces of the mice after modeling were collected. The genomic DNA in the feces was extracted and 16S rDNA sequencing was performed. The analysis results of the α diversity and β diversity of the bacterial flora are as follows: Figure 16-Figure 19 shown.

[0164] Previous studies have shown that DSS modeling can cause disturbances in the intestinal flora of mice, so the effect of intervention with Bifidobacterium breve SHMB 8001 on the intestinal flora structure of colitis mice was further evaluated.

[0165] like Figure 16-18 As shown, the analysis of alpha diversity showed that the richness and diversity (calculated by Chao, Ace and Shannon diversity index) in the DSS group, the B. breve SHMB 8001 group and the mesalazine group were lower than those in the control group. Compared with the DSS group, an increasing trend of alpha diversity was observed in the B. breve SHMB 8001 group and the mesalazine group, achieving differences, but no significant differences were found.

[0166] like Fig.19As shown, the intestinal flora of mice in the DSS group was different from that of mice in the blank control group. After intervention with Bifidobacterium breve SHMB 8001 and mesalazine, the intestinal flora of mice moved toward the blank control group, and the Bifidobacterium breve SHMB 8001 group and the mesalazine group were very similar.

[0167] The above results indicate that Bifidobacterium breve SHMB 8001 can effectively increase the diversity of intestinal flora in colitis mice and improve the intestinal flora disorder caused by DSS modeling.

[0168] The technical features of the above-described embodiments may be arbitrarily combined. To make the description concise, not all possible combinations of the technical features in the above-described embodiments are described. However, as long as there is no contradiction in the combination of these technical features, they should be considered to be within the scope of this specification.

[0169] The above-mentioned embodiments only express several implementation methods of the present application, and the descriptions thereof are relatively specific and detailed, but they cannot be understood as limiting the scope of the invention patent. It should be pointed out that, for a person of ordinary skill in the art, several variations and improvements can be made without departing from the concept of the present application, and these all belong to the protection scope of the present application. Therefore, the protection scope of the patent of the present application shall be subject to the attached claims, and the description and drawings may be used to interpret the contents of the claims.

Claims

1. Bifidobacterium breve SHMB 8001, characterized in that It is deposited in the General Microbiology Center of China Microorganism Culture Collection Administration, the deposit date is November 13, 2024, and the deposit number is CGMCC No.32608.

2. The Bifidobacterium breve SHMB 8001 according to claim 1, characterized in that The Bifidobacterium breve SHMB 8001 is derived from breast milk, and the 16S rDNA sequence of the Bifidobacterium breve SHMB 8001 comprises a nucleotide sequence as shown in SEQ ID NO:

1.

3. The derivative of Bifidobacterium breve SHMB 8001 according to claim 1 or claim 2, characterized in that The derivative is the inactivated bacteria, extract, culture, culture supernatant, fermentation product, fermentation supernatant or a combination thereof of the Bifidobacterium breve SHMB 8001.

4. A composition, characterized in that It comprises a first component, which comprises one or more of Bifidobacterium breve SHMB 8001 as claimed in claim 1 or claim 2 and a derivative as claimed in claim 3.

5. The composition according to claim 4, characterized in that The composition is a pharmaceutical composition, a food composition or a feed composition; Optionally, the number of Bifidobacterium breve SHMB 8001 in the composition is not less than 1×10 9 CFU / g or 1×10 9 CFU / mL.

6. The composition according to claim 4 or claim 5, characterized in that The composition further comprises a second component; the second component comprises one or more of a probiotic, a postbiotic, a prebiotic, an antimicrobial agent, an immunomodulator, an anticancer agent, an inflammatory therapeutic agent, and any combination of the foregoing; Optionally, the composition further comprises a pharmaceutically acceptable carrier and / or excipient.

7. Use of Bifidobacterium breve SHMB 8001 according to claim 1 or claim 2, the derivative according to claim 3, or the composition according to any one of claims 4 to 6 in the preparation of a medicament for alleviating inflammatory bowel disease.

8. The use according to claim 7, characterized in that The inflammatory bowel disease is colitis.

9. A method for culturing Bifidobacterium breve SHMB 8001 according to claim 1 or claim 2, characterized in that: The method comprises: The Bifidobacterium breve SHMB 8001 was cultured in a culture medium.

10. The method according to claim 9, characterized in that The cultivation of the Bifidobacterium breve SHMB 8001 satisfies one or more of the following conditions 1) to 3): 1) The culture medium is an MRS liquid culture medium, and the MRS liquid culture medium optionally contains 0.5 g / L cysteine; 2) cultured under anaerobic conditions; and, 3) Incubate at 37°C.

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