RU2791561C2 - Lactobacillus crispatus kbl693 strain and its application - Google Patents

Abstract

FIELD: biotechnology.

SUBSTANCE: present invention relates to the field of biotechnology, namely to the Lactobacillus crispatus KBL693 strain and to its application. Lactobacillus crispatus KBL693 strain (identification number KCTC 13519BP) in accordance with the present invention weakens allergic cellular reactions, significantly improves the symptoms of atopic dermatitis and has anti-inflammatory and antifungal effects. Thus, this single strain by itself can achieve all the goals of mitigating atopic dermatitis and other allergic diseases and improving the condition with inflammatory diseases and autoimmune diseases and thereby successfully find use as a probiotic substance.

EFFECT: this strain, based on its antifungal activity, can be successfully used as a means for external skin treatment against various skin diseases caused by fungi, as well as in a cosmetic composition and functional strip to soften skin sensitivity.

25 cl, 16 dwg, 4 tbl, 7 ex

Description

Technical field

[1] The present invention relates to a strain of Lactobacillus crispatus KBL693 and its use. More specifically, the present invention relates to a functional health food composition having at least one effect selected from the group consisting of alleviation of allergy symptoms, alleviation of inflammation symptoms and immunoregulation; antifungal composition; and a pharmaceutical composition for the treatment of at least one disease selected from the group consisting of allergic diseases, including atopic dermatitis, and/or inflammatory or autoimmune diseases, containing at least one component selected from the group consisting of a new probiotic strain of Lactobacillus crispatus KBL693, cultures of said strain, lysates of said strain, and extracts of said strain.

[2]

State of the art

[3] Probiotics refers to microorganisms and products derived from them that have antimicrobial activity and enzymatic activity that help maintain the balance of intestinal microorganisms. In addition, probiotics, by definition, are live bacteria in the form of one or more strains that improve the intestinal microflora when administered to humans or animals in the form of dry cells or fermentation products. Probiotics must inhabit the human gut, be non-pathogenic and non-toxic, and survive for a sufficiently long time before entering the gut. In addition, probiotics must remain viable and active prior to dietary intake, be sensitive to antibiotics used to prevent infections, and not contain antibiotic resistance plasmids. In addition, probiotics must be resistant to acids, enzymes and bile in the environment present in the intestines.

[4] Such probiotics may include, for example, Bacillus sp. , having a remarkable ability to produce hydrolyzing enzymes, such as amylase, protease, lipase, cellulase and phosphatase, Lactobacillus sp. lactic acid-producing bacteria and photosynthetic bacteria that prevent malodor by using malodorous substances (such as ammonia, hydrogen sulfide, and amines) present in livestock faeces in metabolic processes. According to a recent report, probiotics have a variety of bodily function-enhancing effects, including gut health, and are therefore actively considered as mainstream therapeutic agents that can replace existing compound-based therapeutic agents.

[5]

[6] Allergy is a biochemical phenomenon characterized by a unique distorted response to a foreign substance (antigen, allergen). The foreign substance that causes symptoms is called an allergen, and the diseases in which these symptoms develop are called allergic diseases. Allergy is a pathological process in a living organism, the cause of which is the antigen-antibody reaction. In general, there are four types of allergy, depending on the time period during which the reaction occurs and the involvement of the complement. Type 1 allergy is an anaphylactic (immediate) type reaction in which the target organs are mainly the digestive system, skin and lungs, and common symptoms include gastrointestinal allergies, urticaria, atrophic dermatitis, allergic rhinitis, bronchial asthma, etc. d. It is known that the pathological mechanisms of type 1 allergy develop as follows: upon contact of antigens with IgE antibodies attached to the surface of mast cells and basophils, target cells are activated and secrete chemical mediators, for example, histamine, leukotrienes and PAF, followed by contraction of blood vessels and smooth muscles. This mechanism can often be seen in combination with type 4 (delayed) allergy. In other words, such an anaphylactic and allergic reaction may occur due to various changes in mast cells, etc. The activation of mast cells leading to their degranulation is due to the binding of antigen, anti-IgE antibody, lectin, etc. with Fc receptors, stimulation by anaphylotoxin, etc. or other drugs, such as calcium ionophore, compound 48/80, codeine and synthetic adrenocorticotropin.

[7] It is known that mast cells and blood basophils are the main cells of the body that cause various allergic diseases, such as allergic rhinitis, allergic dermatitis, asthma, food allergies and anaphylactic shock. These cells contain on their surface receptors (FcRI) for IgE, which are antibodies that cause allergies; these cells are stimulated by allergy-causing substances (antigens, allergens) and secrete their own allergy-causing substances (Kim K et al., Eur J Pharmacol, 581:191-203, 2008).

[8] Among allergic diseases, atopic dermatitis is a well-known chronic relapsing skin disease that occurs in newborns or children and may persist into adulthood. Like asthma or allergic rhinitis, atopic dermatitis is an inflammatory skin disease associated with local infiltration of T-lymphocytes producing IL-4 and IL-5. IL-4 is well known to control the development of T helper 2 (Th2) phenotype, which leads to excessive production of immunoglobulins (Ig) and eosinophilia, as well as an increase in serum IgE levels. Atopic dermatitis is found in 80-90% of subjects with positive skin test results for food and inhalant allergens.

[9] There are various agents for the treatment or prevention of allergic diseases and atopic dermatitis, but an effective method of treatment has not yet been found. Several drug therapies are known, however, even short-term use of a drug can lead to the development of resistance, and long-term use can cause serious side effects, and therefore these drug therapies for allergic diseases and atopic dermatitis have recently been avoided. In these circumstances, in the absence of treatment options that provide an absolute demonstrable effect, attempts to improve the symptoms of irritation, such as itching and hyperemia in allergies, often do not work.

[10]

[11] Under these circumstances, the present inventors have focused on research on probiotics to find a way to replace drug-based therapeutics for the treatment of allergic diseases (including atopic dermatitis) for which there are no satisfactory treatments. Thus, the present invention is accomplished by confirming that a new strain of Lactobacillus crispatus showed an exceptional therapeutic effect in relation to allergic diseases, for example, atopic dermatitis, and by confirming that said strain also showed an excellent effect in relation to anti-inflammatory, immunoregulatory and antifungal activities. .

[12]

[13] Brief description of the invention

[14] The purpose of the present invention is to provide a new strain showing exceptional action in alleviating the symptoms of allergies, for example, atopic dermatitis, alleviating the symptoms of inflammation and immunoregulation, as well as a number of applications.

[15] To achieve this goal, the present invention proposes a strain of Lactobacillus crispatus KBL693 (identification number KCTC 13519BP).

[16] In addition, the present invention provides a food composition or dietary supplement composition comprising at least one component selected from the group consisting of said strain, cultures of said strain, lysates of said strain, and extracts of said strain.

[17] In addition, the present invention provides a feed composition or a feed additive composition comprising at least one component selected from the group consisting of said strain, cultures of said strain, lysates of said strain, and extracts of said strain.

[18] In addition, the present invention provides an antifungal composition, for example, a composition for treating dandruff, comprising at least one component selected from the group consisting of said strain, cultures of said strain, lysates of said strain, and extracts of said strain.

[19] In addition, the present invention provides a pharmaceutical composition for the treatment of allergic diseases, for example, atopic dermatitis, and/or inflammatory or autoimmune diseases, containing at least one component selected from the group consisting of the specified strain, cultures of the specified strain, lysates of said strain; and extracts of said strain.

[20] In addition, the present invention provides a method for treating allergic diseases, including atopic dermatitis, and/or inflammatory or autoimmune diseases, comprising the introduction of at least one component selected from the group consisting of the specified strain, cultures of the specified strain, lysates of said strain and extracts of said strain to a subject in need thereof.

[21] In addition, the present invention provides a composition containing at least one component selected from the group consisting of the specified strain, cultures of the specified strain, lysates of the specified strain and extracts of the specified strain, for use in the prevention or treatment of allergic diseases, in including atopic dermatitis, and/or inflammatory or autoimmune diseases.

[22] In addition, the present invention provides the use of a composition for the preparation of a medicament for the prevention or treatment of allergic diseases, including atopic dermatitis, and/or inflammatory or autoimmune diseases, comprising at least one component selected from the group consisting of said strain, cultures of said strain, lysates of said strain, and extracts of said strain.

[23] Further, the present invention provides a cosmetic composition comprising at least one component selected from the group consisting of said strain, cultures of said strain, lysates of said strain, and extracts of said strain.

[24] In addition, the present invention provides a cosmetic patch or medical patch containing at least one component selected from the group consisting of said strain, cultures of said strain, lysates of said strain, and extracts of said strain.

[25]

Brief description of the drawings

[26] FIG. 1 shows the result of comparing the effect of inhibition of histamine secretion by treatment with various strains of Lactobacillus , including Lactobacillus crispatus KBL693, and the antihistamine agent ketotifen after induction of histamine production by an antigen-antibody reaction in RBL 2H3 cell lines.

[27] FIG. 2 shows a result confirming the effect of inhibition of IL-4 expression by various strains of Lactobacillus including Lactobacillus crispatus strain KBL693 after induction of an allergic reaction in EL4 cell lines.

[28] FIG. 3 shows a result confirming the effect of inhibition of IL-5 expression by various strains of Lactobacillus including Lactobacillus crispatus strain KBL693 after induction of an allergic reaction in EL4 cell lines.

[29] FIG. 4 shows a result confirming the effect of inhibition of IgE secretion by various strains of Lactobacillus including Lactobacillus crispatus strain KBL693 after induction of an allergic reaction in U266B1 cell lines.

[30] FIG. 5 shows the result of observing a significant quantitative increase in secretion of the anti-inflammatory cytokine IL-10 by treatment with strain KBL693 when treated with various strains of Lactobacillus after induction of an inflammatory response in RAW 264.7 cell lines.

[31] FIG. 6 shows a result confirming the significant immunoregulatory and anti-inflammatory effect of treatment with strain KBL693, taking into account the value of IL-10/TNF-α when treated with various strains of Lactobacillus after induction of an inflammatory response in RAW 264.7 cell lines.

[32] FIG. 7 shows a result confirming the significant immunoregulatory and anti-inflammatory effect of treatment with strain KBL693, taking into account the value of IL-10/IL-6 when treated with various strains of Lactobacillus after induction of an inflammatory response in RAW 264.7 cell lines.

[33] FIG. 8 shows the result of the drop test confirming the antifungal activity of Lactobacillus crispatus strain KBL693.

[34] FIG. 9 shows a result confirming the effect of lowering the dermatitis score by oral administration of the Lactobacillus crispatus strain KBL693 mouse model of induced atopic dermatitis.

[35] FIG. 10 shows a result confirming the itching effect of oral administration of Lactobacillus crispatus strain KBL693 to a mouse model of induced atopic dermatitis.

[36] FIG. 11 shows a result confirming the effect of reducing ear thickness by oral administration of Lactobacillus crispatus strain KBL693 to a mouse model of induced atopic dermatitis.

[37] FIG. 12 shows a result confirming the effect of skin thickness reduction by oral administration of Lactobacillus crispatus strain KBL693 to a mouse model of induced atopic dermatitis.

[38] FIG. 13(A) depicts an experimental design using an ovalbumin-induced asthma (OVA) mouse model, and FIG. 13(B) shows the observation of lung tissues collected in each group from the conventional control group (control-PBS), the induced asthma control group (OVA-PBS) and the experimental group (OVA-LC206) through an optical microscope after staining with H&E.

[39] FIG. 14 shows the result of determining the level of expression of total IL-5 and total IL-13 in lung tissues collected in each group from the conventional control group (control-PBS), the induced asthma control group (OVA-PBS) and the experimental group (OVA-KBL693). ).

[40] FIG. 15 shows a result confirming the effect of reducing airway hypersensitivity (AHR) by Lactobacillus crispatus strain KBL693 administered to house dust mite (HDM) asthma mouse models.

[41] FIG. 16 shows a result confirming the effect of reducing the number of eosinophils, IL-5 ⁺ CD4 T cells and IL-13 ⁺ CD4 T cells by administering Lactobacillus crispatus strain KBL693.

[42]

[43] Best Mode for Carrying Out the Invention

[44] Unless otherwise indicated, all technical and scientific terms used in this application have the same meaning as generally accepted among specialists in this field of technology. In general, the nomenclature used in this application is well known and widely used in the art.

[45]

[46] The present invention has made it possible to detect the anti-allergic effect of microorganisms derived from the human body, and the selected strain of Lactobacillus crispatus KBL693 (identification number KCTC 13519BP) has an exceptional allergy-suppressing effect. Analysis of the 16S rDNA of said strain demonstrates that said strain is a new strain that has never been known to the public.

[47]

[48] According to one embodiment of the present invention, the present invention relates to a novel probiotic strain of Lactobacillus crispatus KBL693 (identification number KCTC 13519BP), and said strain is characterized by the presence of a 16S rDNA sequence of SEQ ID NO: 1.

[49]

[50] <SEQ ID NO: 1> 16S rDNA sequence of Lactobacillus crispatus strain KBL693 (KCTC ID 13519BP)

[51]

CGCGGGGTGGCGCGAGCTATAATGCAGTCGAGCGAGCGGAACTAACAGATTTACTTCGGTAATGACGTTAGGAAAGCGAGCGGCGGATGGGTGAGTAACACGTGGGGAACCTGCCCCATAGTCTGGGATACCACTTGGAAACAGGTGCTAATACCGGATAAGAAAGCAGATCGCATGATCAGCTTTTAAAAGGCGGCGTAAGCTGTCGCTATGGGATGGCCCCGCGGTGCATTAGCTAGTTGGTAAGGTAAAGGCTTACCAAGGCGATGATGCATAGCCGAGTTGAGAGACTGATCGGCCACATTGGGACTGAGACACGGCCCAAACTCCTACGGGAGGCAGCAGTAGGGAATCTTCCACAATGGACGCAAGTCTGATGGAGCAACGCCGCGTGAGTGAAGAAGGTTTTCGGATCGTAAAGCTCTGTTGTTGGTGAAGAAGGATAGAGGTAGTAACTGGCCTTTATTTGACGGTAATCAACCAGAAAGTCACGGCTAACTACGTGCCAGCAGCCGCGGTAATACGTAGGTGGCAAGCGTTGTCCGGATTTATTGGGCGTAAAGCGAGCGCAGGCGGAAGAATAAGTCTGATGTGAAAGCCCTCGGCTTAACCGAGGAACTGCATCGGAAACTGTTTTTCTTGAGTGCAGAAGAGGAGAGTGGAACTCCATGTGTAGCGGTGGAATGCGTAGATATATGGAAGAACACCAGTGGCGAAGGCGGCTCTCTGGTCTGCAACTGACGCTGAGGCTCGAAAGCATGGGTAGCGAACAGGATTAGATACCCTGGTAGTCCATGCCGTAAACGATGAGTGCTAAGTGTTGGGAGGTTTCCGCCTCTCAGTGCTGCAGCTAACGCATTAAGCACTCCGCCTGGGGAGTACGACCGCAAGGTTGAAACTCAAAGGAATTGACGGGGGCCCGCACAAGCGGTGGAGCATGTGGTTTAATTCGAAGCAACGCGAAGAACCTTACCAGGTCTTGACATCTAGTGCCATTTGT AGAGATACAAAGTTCCCTTCGGGGACGCTAAGACAGGTGGTGCATGGCTGTCGTCAGCTCGTGTCGTGAGATGTTGGGTTAAGTCCCGCAACGAGCGCAACCCTTGTTATTAGTTGCCAGCATTAAGTTGGGCACTCTAATGAGACTGCCGGTGACAAACCGGAGGAAGGTGGGGATGACGTCAAGTCATCATGCCCCTTATGACCTGGGCTACACACGTGCTACAATGGGCAGTACAACGAGAAGCGAGCCTGCGAAGGCAAGCGAATCTCTGAAAGCTGTTCTCAGTTCGGACTGCAGTCTGCAACTCGACTGCACGAAGCTGGAATCGCTAGTAATCGCGGATCAGCACGCCGCGGTGAATACGTTCCCGGGCCTTGTACACACCGCCCGTCACACCATGGGAGTCTGCAATGCCCAAAGCCGGTGGCCTAACCTTCGGAAGGAGCCGTCTAAGTAGACAATGTGCA

[52] Thus, in the development of the present invention, experiments were carried out regarding the effectiveness of said strain, confirming that said strain has an exceptional effect in suppressing allergic disorders, for example, atopic dermatitis, immunoregulatory properties, attenuates the inflammatory response, and has antifungal activity. In addition, the present inventors have confirmed that these effects can be obtained not only under conditions suitable for living bacteria, but also under conditions of low temperature sterilization or high temperature sterilization.

[53]

[54] Accordingly, in yet another embodiment of the present invention, the present invention relates to a food composition or a food supplement composition comprising at least one component selected from the group consisting of a strain of Lactobacillus crispatus KBL693 (identification number KCTC 13519BP), cellular components of said strain, cultures of said strain, lysates of said strain, and extracts of said strain.

[55] The composition may be characterized in that it is a functional health food composition having at least one effect selected from the group consisting of alleviation of symptoms of allergies, for example, atopic dermatitis, alleviation of symptoms of inflammation, and immunoregulation.

[56] Said food composition or dietary supplement composition can be used as a food product effective in alleviating allergy symptoms such as atopic dermatitis, alleviating inflammation symptoms and/or promoting immunoregulation, as well as preventing the above phenomena, for example, as main or secondary ingredients. foodstuffs, nutritional supplements, functional health food composition or functional drinks, but not limited to.

[57] The term "food composition" refers to a natural or artificial product containing at least one nutrient, and, more preferably, refers to a product that has become edible as a result of certain processing, and usually covers all foods, nutritional supplements, functional health food composition; and functional drinks.

[58] A food product that may contain the specified food composition according to the present invention as an additive may include, for example, various types of foods, drinks, chewing gum, tea, vitamin complex or functional foods. In addition, foods of the present invention include specialty fortified foods (e.g., modified milk, baby food), processed meats, fish products, tofu, flour, noodles (e.g., ramen noodles, Asian noodles), baked goods, organically active food additives, seasonings (e.g. soy sauce, soy paste, red pepper paste, mixed paste), sauces, confectionery (e.g. snack foods), candy, chocolate, chewing gums, ice cream, dairy products (e.g. fermented milk , cheese), other processed foods, kimchi, salty foods (such as various types of kimchi, pickled foods), beverages (such as fruit juice, vegetable juice, soy milk, fermented drinks), and natural condiments (such as broth ramen noodle powder), but not limited to. Said foodstuffs, beverages or nutritional supplements can be obtained by conventional methods.

[59] The term "functional health food composition" is a group of food products, the effectiveness of which is enhanced by the ability to display and express their function for a predetermined purpose using physical, biochemical or bioengineering techniques, or processed foods designed to that the regulatory functions of the respective food composition in vivo , such as the function of adjusting the defecation rhythm in disease prevention and recovery, are sufficiently exhibited. Such a functional food product may contain nutritionally acceptable food additives and may additionally contain suitable carriers, excipients and diluents commonly used in their manufacture.

[60] In the present invention, the term "functional drinks" collectively refers to drinks intended to quench thirst or to enjoy the taste. There are no specific restrictions on this term, except that said drinks must contain a composition for alleviating symptoms of allergies, for example, atopic dermatitis, alleviating symptoms of inflammation and / or immunoregulation, and also for the prevention of these phenomena as the main ingredients in the indicated ratio, and also , like regular drinks, may contain various flavors or natural carbohydrates as additional ingredients.

[61] In addition to the above, the food product containing the food composition or dietary supplement composition of the present invention may contain various nutrients, vitamins, minerals (electrolytes), flavors such as synthetic and natural flavors, colors, and fillers ( cheese, chocolate, etc.), pectic acid and its salts, alginic acid and its salts, organic acids, colloidal thickeners, pH control agents, stabilizing agents, preservatives, glycerin, alcohol, carbonation agents used in carbonated beverages , etc., and each of the above ingredients can be used alone or in combination with each other.

[62] The food product containing the food composition according to the present invention may contain the composition according to the present invention in an amount of from 0.001 wt.% 100 ml wt.%, preferably from 1 wt.% to 99 wt.% in terms of the total weight of the food ; in the case of beverages, it can be from 0.001 g to 10 g, preferably from 0.01 g to 1 g per 100 ml. However, with long-term use for recreational or hygienic purposes or in order to maintain a state of health, the amount of the product may be lower than the above range; and since the effective ingredients do not have problems in terms of the safety profile, they can be used in an amount exceeding the specified range, and their amount is not limited to the above range.

[63] The nutritional composition of the present invention may comprise the Lactobacillus crispatus KBL693 strain alone or in combination with a suitable carrier, or may be formulated into a composition suitable for human or animal consumption. Thus, said composition can be added to a food product that does not contain probiotic bacteria or a pair of probiotic bacteria. For example, microorganisms that can be used in combination with the strain of the present invention in the preparation of the food of the present invention should be suitable for human or animal consumption and have probiotic activity in suppressing pathogenic, harmful bacteria or improving the balance of microflora in the intestinal tract of mammals when taken inside, but not limited to. Such probiotic microorganisms may include, for example, yeasts, such as Saccharomyces , Candida , Pichia , or Torulopsis , fungi, such as Aspergillus , Rhizopus , Mucor , or Penicillium , and bacteria of the genera Lactobacillus , Bifidobacterium , Leuconostoc , Lactococcus , Bacillus, Streptococcus, Propionibacterium, Enterococcus or Pediococcus . Подходящие конкретные пробиотические микроорганизмы могут включать, например, Saccharomyces cerevisiae, Bacillus coagulans, Bacillus licheniformis, Bacillus subtilis, Bifidobacterium bifidum, Bifidobacterium infantis, Bifidobacterium longum, Enterococcus faecium, Enterococcus faecalis, Lactobacillus acidophilus, Lactobacillus alimentarius, Lactobacillus casei, Lactobacillus curvatus, Lactobacillus delbruckii , Lactobacillus johnsonii, Lactobacillus farciminus, Lactobacillus gasseri, Lactobacillus helveticus, Lactobacillus rhamnosus, Lactobacillus reuteri, Lactobacillus sakei, Lactococcus lactis, or Pediococcus acidilactici . The nutritional composition according to the present invention may preferably further contain a mixture of probiotic microorganisms having a pronounced probiotic activity and excellent anti-allergic, anti-inflammatory and/or immunoregulatory activity, further enhancing their effects. Carriers that may be included in the food composition of the present invention may include, for example, bulking agents, high fiber supplements, encapsulating agents, and lipids well known in the art. The strain of Lactobacillus crispatus KBL693 according to the present invention may be in lyophilized or encapsulated form, or in the form of suspensions or dry culture powders.

[64]

[65] The composition according to the present invention can also be presented in the form of a feed additive or feed containing the specified strain.

[66] The feed additive according to the present invention may be in the form of a dry or liquid composition and, in addition, contain other non-pathogenic microorganisms in addition to the specified strain of Lactobacillus crispatus KBL693. Microorganisms that can be added to the feed additive may include, for example, Bacillus subtilis , which can produce protease, lipase, and carbohydrate converting enzymes, a strain of Lactobacillus , which has physiological activity and the ability to degrade organic compounds under anaerobic conditions, such as in the stomach of a cow , filamentous fungi such as Aspergillus oryzae showing effects on animal weight gain, milk production and feed digestibility (Slyter, LLJ Animal Sci. 1976, 43. 910-926), and yeasts such as Saccharomyces cerevisiae (Johnson, D. E et al., J. Anim. Sci. 1983, 56, 735-739; Williams, PEV et al., 1990, 211).

[67] The feed additive according to the present invention may further contain at least one enzymatic agent in addition to the specified strain of Lactobacillus crispatus KBL693. Additional enzymatic agents may be in dry or liquid form and may include, for example, steatolytic enzymes, for example, lipase, phytase, which forms phosphate and inositol phosphate upon decomposition of phytic acid, amylase, i.e. an enzyme for the hydrolysis of the α-1,4-glycosidic bond in the composition of, for example, starch and glycogen, phosphatase, i.e. an enzyme for the hydrolysis of an organic ester of phosphoric acid, a carboxymethyl cellulase for the decomposition of cellulose, a xylase for the decomposition of xylose, a maltase for the hydrolysis of maltose into two molecules of glucose, and carbohydrate-producing enzymes, such as invertase, which form a mixture of glucose and fructose upon hydrolysis of sucrose.

[68] When using the Lactobacillus crispatus strain KBL693 according to the present invention, feed raw materials such as peanuts, peas, beets, bagasse, grain products, animal gut powder and fish meal, as well as various grains and soy protein can be used as feed additives. They may be processed or unprocessed and can be used without restriction. Processing may include a process in which the feedstock is loaded and forced under pressure through a predetermined opening, and for proteins, extrusion may preferably be used, in which the proteins are degenerated to increase their availability. Extrusion denatures proteins by heat treatment to destroy anti-enzyme factors, which is advantageous. In addition, the digestibility of soy proteins can be improved by extrusion to inactivate feed depleting substances, such as trypsin inhibitor, one of the protease inhibitors present in soy. In addition, extrusion can improve protease hydrolyzability, which increases the nutritional value of soy proteins.

[69]

[70] According to another embodiment of the present invention, the present invention relates to an antifungal composition comprising at least one component selected from the group consisting of said strain, cultures of said strain, lysates of said strain, and extracts of said strain.

[71] The composition may be characterized by the manifestation of antifungal activity against one of the organisms selected from the group consisting of Malassezia furfur, Malassezia globosa and Malassezia restricta , but not limited to them.

[72] The composition may be a composition for the prevention, alleviation or treatment of seborrheic dermatitis or dandruff, and said seborrheic dermatitis may be located on the skin of the calvaria.

[73] Further, said composition may be a composition for the prevention, alleviation or treatment of urticaria, rash, dermatophytosis of the trunk, groin dermatophytosis or tinea pedis caused by a fungal infection.

[74] The specified strain and cultures of the specified strain, lysates of the specified strain and extracts of the specified strain may be contained in the composition in an amount of 0.1 wt. % to 50 wt.% of the total weight of the composition.

[75] Said antifungal composition may be a pharmaceutical composition, a cosmetic composition, or a health food composition, and may also be an external skin treatment agent.

[76] Said cosmetic composition can be presented in all dosage forms suitable for topical application, for example, liquid, oil-in-water emulsion, water-in-oil emulsion, suspension, solid, gel, powder, paste, foam or spray. These compositions of the above dosage forms can be obtained by traditional methods used in the art.

[77] In addition to the above ingredients, this composition may contain other ingredients in an amount that does not interfere with the manifestation of the main effect, preferably in an amount that provides a synergistic effect with respect to the main effect. The composition according to the present invention may contain a substance selected from the group consisting of vitamins, polypeptides, polysaccharides and sphingolipid. In addition, the cosmetic composition of the present invention may contain a humectant, an emollient, a surfactant, an ultraviolet absorber, a preservative, a sterilizer, an antioxidant, a pH adjusting agent, organic or inorganic pigments, a flavoring agent, a cooling agent, or an antiperspirant agent. Those skilled in the art can choose the combined percentage of these ingredients in a range that does not interfere with the achievement of the goal and manifestation of the effect according to the present invention, and it can be in the range from 0.01 wt.% to 5 wt.%, more specifically, from 0, 01 wt.% to 3 wt.% of the total weight of the composition.

[78] According to the above embodiment, the antifungal composition of the present invention may be an external skin treatment such as a cream, ointment, shampoo, or drug.

[79]

[80] According to another embodiment of the present invention, the present invention relates to a pharmaceutical composition for the treatment or prevention of allergic diseases, for example, atopic dermatitis, and/or inflammatory or autoimmune diseases, containing at least one component selected from the group consisting of cell components of Lactobacillus crispatus KBL693 strain, cultures of said strain, lysates of said strain and extracts of said strain.

[81] The pharmaceutical composition of the present invention may be in the form of cell components of live bacteria, a dry strain, cultures of said strain, lysates of said strain, or a composition in combination with a pharmaceutically acceptable carrier or medium. Carriers or media that can be used in the present invention may include a solvent, dispersant, coating, enhancer, controlled release formulation (i.e., sustained release formulation), or at least one inert excipient including starch, polyol, granules , fine cellulose, microcrystalline cellulose, eg Celphere, Celphere granules, diluent, lubricant, binder, disintegrant. The tablet of the above composition can optionally be coated using standard aqueous or non-aqueous techniques. Examples of a pharmaceutically acceptable carrier and excipient for use as a pharmaceutically acceptable inert carrier and these additional ingredients may include, for example, a binder, a filler, a disintegrant, a lubricant, an antimicrobial agent, and a coating agent.

[82] In addition, the pharmaceutical composition according to the present invention can be used as an external agent, containing a dosage form selected from the group consisting of ointments, creams, pastes, liquids and solutions for application to the skin, glycerogelatins, rubbing ointments, skin powders, aerosols and patches.

[83] In the present invention, these allergic diseases refer to conditions associated with the expression of IL-4 or IL-5, and may include, for example, eczema, allergic asthma, atopic dermatitis, allergic rhinitis, allergic conjunctivitis, urticaria, or an anaphylactic reaction. The present invention may preferably be characterized in that said diseases are selected from, but are not limited to, childhood eczema, allergic asthma, allergic rhinitis, atopic dermatitis, allergic conjunctivitis, and food allergies.

[84] In the present invention, In said inflammatory or autoimmune diseases or symptoms are diseases or symptoms that can be treated, alleviated or prevented by stimulating the expression or secretion of IL-10, which is an anti-inflammatory immunoregulatory cytokine, and may include, for example, rheumatoid arthritis , rheumatic fever, lupus, systemic scleroderma, atopic dermatitis, psoriasis, psoriatic arthritis, asthma, Guillain-Barré syndrome, myasthenia gravis, dermatomyositis, polymyositis, multiple sclerosis, autoimmune encephalomyelitis, periarteritis nodosa, Hashimoto's thyroiditis, temporal arteritis, juvenile diabetes, focal alopecia, pemphigus, aphthous stomatitis, autoimmune hemolytic anemia, Wegener's granulomatosis, Sjögren's syndrome, Addison's disease, Crohn's disease, Behçet's disease, edema, conjunctivitis, periodontitis, rhinitis, otitis media, chronic sinusitis, pharyngolaryngitis, tonsillitis, bronchitis, pneumonia, ulcer stomach, ha street, colitis, gout, eczema, acne, contact dermatitis, seborrheic dermatitis, ankylosing spondylitis, fibromyalgia, osteoarthritis, periarthritis of the shoulder joint, tendinitis, tenosynovitis, myositis, hepatitis, cystitis, nephritis, sepsis, vasculitis, bursitis, etc., but not limited to them.

[85]

[86] The term "treatment" refers to the relief or mitigation of diseases or conditions used in conjunction with the term, or one or more of their symptoms, the suppression of their progression or their prevention, unless it is used in another sense. The term "therapy" in the present invention refers to the act of "treating" as defined above. Accordingly, treatment or therapy regimens for a disease in a mammal may include one or more of the following:

[87] (1) Suppression of disease progression, i. e. suppression of its development

[88] (2) Preventing the spread of disease

[89] (3) Disease mitigation

[90] (4) Prevention of disease recurrence and

[91] (5) Temporary relief of the symptom of the disease

[92]

[93] The composition according to the present invention for the prevention or treatment of allergic diseases, for example, atopic dermatitis, and/or inflammatory or autoimmune diseases may contain a pharmaceutically effective amount of Lactobacillus crispatus strain KBL693 alone or in combination with at least one of the pharmaceutically acceptable carriers , excipients or diluents.

[94]

[95] In the present invention, the term "effective amount" means an amount high enough to cause the desired effect, but low enough to prevent serious side effects, as judged by physicians. The number of microorganisms introduced into the body with the composition according to the present invention can be adjusted according to the route of administration and the purpose of administration.

[96]

[97] The composition according to the present invention can be administered to the subject one or more times a day. Dosage form means a physically discrete unit suitable for standard administration to human subjects and other mammals, each unit containing a suitable pharmaceutical carrier and a predetermined amount of Lactobacillus crispatus KBL693 strain of the present invention to provide a therapeutic effect. The dosage form for oral administration to an adult patient preferably contains 0.001 g or more of the microorganism according to the present invention, and the dosage of the composition according to the present invention for oral use is about 0.001 g to 10 g, preferably from 0.01 g to 5 g per dose. A pharmaceutically effective amount of the microorganism according to the present invention is from 0.01 g to 10 g/day. At the same time, the dosage depends on the severity of the disease of the patient, the microorganism and the jointly used auxiliary effective ingredients. In addition, the total daily dosage can be divided into several times and continuously administered as needed. Accordingly, the above dosage ranges do not limit the scope of the present invention in any way.

[98]

[99] In addition, the term "pharmaceutically acceptable" as used above refers to a composition that is physiologically acceptable and does not cause an allergic reaction, such as gastrointestinal upset, dizziness, or the like, when administered to a human.

[100] the Composition according to the present invention can be formulated using methods known in the art, providing rapid, delayed or delayed release of the active ingredients after administration to a mammal. Dosage forms may be powders, granules, tablets, emulsions, syrups, aerosols, soft or hard gelatin capsules, sterile injectable solutions or sterile powders. In addition, the composition according to the present invention for the prevention or treatment of allergic diseases, for example, atopic dermatitis, and/or inflammatory or autoimmune diseases can be administered in various ways, including oral, transdermal, subcutaneous, intravenous or intramuscular administration. The dosage of active ingredients can be selected depending on various factors, for example, the route of administration, age, sex, body weight of the patient and the severity of his condition. The composition according to the present invention for the treatment of allergic diseases, for example atopic dermatitis, and/or inflammatory or autoimmune diseases, can be administered in combination with a known compound capable of preventing, alleviating or treating the respective symptoms.

[101]

[102] The pharmaceutical composition according to the present invention can be presented, in particular, in the form of a dosage form for oral administration in the composition with an enteric coating. As used herein, the term "enteric coating" includes any pharmaceutically acceptable coating that can persist in the stomach without being degraded by acidic gastric juice and can be sufficiently broken down in the intestine to release the active ingredients there. "Enteric coating" according to the present invention refers to a coating that can be preserved for 2 hours or more when in contact with artificial gastric juice, for example, HCl solution with pH 1 at 36-38°C, and can be further destroyed, preferably in artificial intestinal juice, for example, KH ₂ PO ₄ buffer solution with pH 6.8 for 30 minutes.

[103] The enteric coating of the present invention is applied to one core in an amount of about 16 mg to 30 mg, preferably 16 mg to 20 mg, or 25 mg or less. With a thickness of the enteric coating according to the present invention of 5 µm to 100 µm, preferably 20 µm to 80 µm, satisfactory results can be obtained. The enteric coating material can be selected from known polymeric materials. Suitable polymeric materials are listed in a number of known documents [L. Lachman et al., The Theory and Practice of Industrial Pharmacy, ^3rd ed., 1986, pp. 365~373; H. Sucker et al., Pharmazeutische Technologie, Thieme, 1991, pp. 355-359; Hagers Handbuchder pharmazeutischen Praxis, ^4th ed., Vol. 7, pp. 739~742, and 766~778, (SpringerVerlag, 1971); and Remington's Pharmaceutical Sciences, ^13th ed., pp. 1689~1691 (Mack Publ., Co., 1970)] and may include cellulose ester derivatives, cellulose ethers, an acrylic resin-methyl acrylate copolymer, and a copolymer of maleic acid and phthalic acid derivatives.

[104]

[105] The enteric coating of the present invention can be prepared using a conventional enteric coating method in which an enteric coating solution is sprayed onto a core. Suitable solvents used in the enteric coating process are alcohols, eg ethanol, ketones, eg acetone, halogenated hydrocarbon solvents, eg dichloromethane (CH ₂ Cl ₂ ), and mixtures of these solvents. A plasticizer, such as di-n-butyl phthalate or triacetylglycerol, is added to the coating solution in a ratio of about 1 : 0.05 to about 1 : 0.3 (coating material : plasticizer). considering the conditions of coating application. The atomization pressure can be adjusted in various ways, and in general, satisfactory results can be obtained at a spray pressure of from about 1 bar to about 1.5 bar.

[106]

[107] In yet another embodiment of the present invention, the present invention provides the use of said strain or said composition for the prevention or treatment of allergic diseases, e.g., atopic dermatitis, and/or inflammatory or autoimmune diseases, and the use of said strain or said composition to obtain a therapeutic agent for the treatment of the aforementioned diseases.

[108] Specifically, the present invention relates to a composition containing at least one component selected from the group consisting of the specified strain, cultures of the specified strain, lysates of the specified strain and extracts of the specified strain, for use in the prevention or treatment of allergic diseases, including including atopic dermatitis, and/or inflammatory or autoimmune diseases.

[109] In addition, the present invention relates to the use of a composition for the preparation of a medicinal product for the prevention or treatment of allergic diseases, including atopic dermatitis, and/or inflammatory or autoimmune diseases, containing at least one component selected from the group consisting of said strain, cultures of said strain, lysates of said strain, and extracts of said strain.

[110] As used herein, the term "prevention" refers to the prevention, delay, inhibition, or slowing down of diseases in order to reduce their impact.

[111] As used herein, the term "treatment" refers to caring for a subject suffering from a disease in order to alleviate, cure, or lessen the symptoms of the disease, or lessen or stop the progression of the disease.

[112]

[113] In yet another embodiment of the present invention, the present invention provides a method for the prevention or treatment of allergic diseases, for example, atopic dermatitis, and/or inflammatory or autoimmune diseases, comprising administering a pharmaceutically effective amount of said strain or said composition to a subject in need of prevention or in the treatment of said diseases or in the mitigation of said diseases.

[114] In addition, the present invention provides a method for the treatment of allergic diseases, for example, atopic dermatitis, and/or inflammatory or autoimmune diseases, comprising the introduction of a therapeutically effective amount of at least one component selected from the group consisting of the specified strain, cultures of the specified strain, lysates of said strain, and extracts of said strain to a subject in need thereof.

[115] Since the pharmaceutical composition used in the method for the prevention or treatment of these diseases, as well as the method of administration thereof, are described above, the overlapping elements of the composition and the method will not be included in this document in order to avoid unnecessary complexity of the present description.

[116] the Specified subject, which you can enter the composition for the prevention or treatment of these diseases, includes all animals, including humans. For example, the subject may be an animal, such as a dog, cat, or mouse.

[117]

[118] In yet another embodiment of the present invention, the present invention relates to a cosmetic composition containing a pharmaceutically effective amount of at least one component selected from the group consisting of cellular components of the Lactobacillus crispatus KBL693 strain, cultures of said strain, lysates of said strain, and extracts of said strain.

[119] The cosmetic composition can be characterized by the function of alleviating at least one condition of sensitive skin, selected from the group consisting of skin allergies, urticaria, atopic dermatitis, psoriasis, fungal infection and eczema.

[120] As used herein, the term "cosmetic composition" refers to a composition containing at least one component selected from the group consisting of cellular components of Lactobacillus crispatus KBL693 strain, cultures of said strain, lysates of said strain, and extracts of said strain, which may be presented in the form of a dosage form of any type. For example, cosmetics made using said cosmetic composition may include creams, tampons, lotions, essences, tonics, cream bases and cosmetic bases, and may be marketed in any of the above dosage forms to achieve the purpose of the present invention, but not limited to them. Ingredients contained in the cosmetic composition of the present invention, in addition to the above ingredients, include ingredients commonly used in cosmetic compositions, such as conventional adjuvants such as antioxidants, stabilizers, solubilizers, vitamins, pigments and fragrances, and carriers.

[121]

[122] In another embodiment of the present invention, the present invention relates to a functional patch containing at least one component selected from the group consisting of cellular components of the strain Lactobacillus crispatus KBL693, cultures of the specified strain, lysates of the specified strain and extracts of the specified strain, and said functional patch can be used for a cosmetic or medical purpose.

[123] The patch can be characterized by the function of alleviating at least one condition of sensitive skin, selected from the group consisting of skin allergies, urticaria, atopic dermatitis, psoriasis, fungal infection and eczema.

[124] In the present invention, the patch is usually a small sticky patch containing delivered substances, and the specified patch can be presented in various forms. The simplest form is a sticky one-piece product containing a reservoir containing the delivered substances, placed on a substrate. The reservoir is usually formed by a cosmetically or pharmaceutically acceptable self-adhesive material, however, in some cases, it can also be formed by non-adhesive materials presented in a thin adhesive layer suitable for a skin contact surface. The rate of introduction of substances delivered from the patch into the body of the subject wearing the patch can be changed, since the permeability of the skin to the delivered substances usually depends on individuals and the place on the skin, which can be chosen by specialists in this field of technology.

[125]

[126] The present invention will be described in detail hereinafter by way of examples. These examples are provided for the sole purpose of illustrating the present invention, and those skilled in the art will appreciate that the spirit of the present invention should not be considered to be limited to these examples.

[127]

[128] Example 1 Alleviating and curing effects of KBL693 on allergic reactions due to inhibition of histamine secretion in basophils.

[129] It has been reported that when an allergic reaction occurs, histamine is produced in the tissues causing an inflammatory reaction, and the suppression of histamine secretion results in alleviation of allergy symptoms by blocking the reaction caused by histamine. allergic reactions due to inhibition of histamine secretion. To this end, the ability of a total of eight human vaginal-derived Lactobacillus strains to inhibit histamine secretion was evaluated. The ability to suppress histamine secretion was confirmed by inducing degranulation of the RBL-2H3 cell line after cultivation, followed by measuring the activity of β-hexosaminidase co-secreted with histamine by colorimetric reaction with the substrate.

[130]

[131] 1-1. Incubation of RBL-2H3 Cell Lines

[132] RBL-2H3 cells (ATCC NO. CRL-2256) were cultured in DMEM supplemented with 10% FBS (fetal calf serum), penicillin (100 μg/ml), and streptomycin (100 μg/ml) at 37°C in an atmosphere of 5% CO2, and then reseeded every three days. RBL-2H3 cells were seeded on a 6-well plate at a concentration of 1×10 ⁶ cells/well, then cultured for 3 hours, and then treated with IgE (0.5 μg/ml) and incubated for 16 - 20 hours.

[133]

[134] 1-2. Incubation of strains and preparation of culture solution

[135] The Lactobacillus crispatus strains used were cultured in MRS medium supplemented with 0.5% cysteine, activated for a total of two passages 18 hours apart, and then used in the experiments. The resulting culture solution was centrifuged at 15,000× g for 3 minutes, and then the supernatant was collected.

[136]

[137] 1-3. Induction of degranulation after treatment with strains

[138] After removing the medium for RBL-2H3 cells previously seeded in a 6-well plate, the cells were washed twice with 1 ml of Siraganyan's buffer (pH 7.2). The cells were then treated with 120 μl of the previously obtained bacterial culture solution or, in the positive control group, with ketotifen (20 μg/ml) in each well, incubated for 20 minutes in an incubator with an atmosphere of 5% CO ₂ at 37°C, and then treated with 60 µl of antigen (DNP-HAS, 1 µg/ml) to induce degranulation by antigen-antibody reaction. As a negative control, degranulation was induced after treatment with 120 μl Siraganyan's buffer. The reaction was carried out in a 5% CO ₂ incubator at 37° C. for 20 minutes, and then the supernatant was collected.

[139]

[140] 1-4. Color Reaction Detection

[141] To detect β-hexosaminidase activity, 25 μl of the supernatant collected in Example 1-3 was transferred to each well of a 96-well plate, and then 25 μl of each substrate ( p -nitrophenyl-N-acetyl-D- glucosaminidase) followed by reaction at 37°C in an incubator with an atmosphere of 5% CO ₂ for 90 minutes. Then 200 µl of stop solution (Na2CO3/NaHCO3) was added to stop the reaction and the absorbance at 405 nm was measured. The absorbance after treatment with Lactobacillus crispatus of each type was compared with the negative control group and converted to a percentage to show the level of inhibition of degranulation.

[142]

[143] As a result, as can be seen in FIG. 1 and Table 1, ten types of Lactobacillus crispatus strains, including strain KBL693, showed significantly lower quantitative levels of β-hexosaminidase secretion compared to the negative control group, and nine of them showed even lower quantitative levels of β-hexosaminidase secretion by compared with levels using the commercially available antihistamine agent ketotifen, a positive control. As a result, strains of Lactobacillus crispatus , including strain KBL693, were found to exhibit remarkable degranulation preventing activity to effectively alleviate allergy symptoms caused by excessive histamine secretion.

[144]

[145]

Table 1

View The strain or agent used for the treatment β-hexosaminidase (fold change) - (Negative control) Siraganyan Buffer 1.0 - (Positive control) Ketotifen 0.2569 Lactobacillus crispatus KBL693 0.2140 Lactobacillus crispatus KBL694 0.2063 Lactobacillus crispatus KBL392 0.1904 Lactobacillus crispatus KBL702 0.2065 Lactobacillus crispatus KBL706 0.1943 Lactobacillus crispatus KBL707 0.2008 Lactobacillus crispatus KBL709 0.2128 Lactobacillus crispatus KBL696 0.2331 Lactobacillus crispatus KBL708 0.2464 Lactobacillus crispatus SNUV272-1 0.3632 Lactobacillus fermentum SNUV417 0.9284 Lactobacillus jensenii SNUV221 0.7088 Lactobacillus vaginalis SNUV351 1.0336

[146]

[147] Example 2 The mitigating and curing effects of KBL693 on allergic reactions due to inhibition of Th2-type cytokines in T cells.

[148] T-helper type 2 (Th2) related cytokines, such as IL-4 and IL-5, have been reported to contribute to chronic allergic reactions by enhancing Th2-related immune responses and increasing production IgE (Passante E, Inflamm. Res . 2009). The present invention attempts to further compare the effects of suppressing chronic allergic reactions among strains of Lactobacillus crispatus , showing a remarkable effect of inhibiting histamine secretion in Example 1. a line of mouse T cells.

[149]

[150] 2-1. Incubation of EL-4 Cell Lines

[151] EL4 cells (ATCC NO. TIB-39) were cultured in DMEM supplemented with 10% FBS, penicillin (100 μg/mL), and streptomycin (100 μg/mL) at 37°C in 5% CO2, and then reseeded every three days. EL4 cells were seeded on a 24-well plate at a concentration of 4×10 ⁵ cells/well, then cultured for 16-20 hours to induce allergic reactions and treated with strains.

[152]

[153] 2-2. Strain incubation and recovery

[154] A total of nine strains of Lactobacillus crispatus , including the used KBL693 (KBL693, KBL709, KBL702, KBL696, KBL708, KBL707, KBL692, KBL694 and KBL706) were cultured in MRS medium supplemented with 0.5% cysteine, activated for a total of two including subcultures at 18-hour intervals and then used in experiments. The resulting culture solution was centrifuged at 15,000× g for 3 minutes, the pellet was washed with PBS buffer. The strains were stained for 15 minutes with SYTO9 and PI using the LIVE/DEAD™ BacLight™ bacterial count and viability kit for flow cytometry (Thermo Fisher Scientific, USA). The beads contained in the kit were then added and the number of stained live bacteria was calculated using flow cytometric analysis.

[155]

[156] 2-3. Treatment of strains and quantitative measurement of secretion of IL-4 and IL-5 after the induction of allergic reactions

[157] To induce allergic reactions in EL4 cells previously seeded in a 24-well plate, each well was treated with 100 μl of PMA (20 ng/ml) and ionomycin (1 μg/ml). Then, 300 μl of previously obtained strains were processed in a cell:strain ratio of 1:10. After incubation for 24 hours in a 5% CO ₂ incubator at 37° C., the supernatant was collected and the Mouse IL-4 (Cat. No. 555232, BD OptEIA™) and Mouse IL-5 (Cat. No. 555236, BD OptEIA™) to measure the amount of secreted IL-4 and IL-5 according to the manufacturer's instructions.

[158]

[159] As a result, as shown in FIG. 2 and 3, these strains of Lactobacillus crispatus as a whole showed the effects of inhibiting the secretion of IL-4 and IL-5; in particular, it was found that when using strain KBL693, the amount of secreted IL-4 was the lowest and the amount of secreted IL-5 was the second highest in ascending order. Accordingly, it was found that KBL693 could have a therapeutic and prophylactic effect in relation to allergies by inhibiting the secretion of Th2-type cytokines mediating allergic reactions.

[160]

[161] Example 3 Effect of KBL693 on Alleviation and Treatment of IgE Inhibition of Allergic Reactions

[162] Immunoglobulin E (IgE) is a type of immunoglobulin; it is known to be involved in the development of allergic diseases. In general, the measurement of the total amount of IgE in serum is one of the highest priority methods for diagnosing allergic diseases. Thus, the present invention attempts to test the effect of inhibition of IgE secretion by strain KBL693 using the U266B1 cell line, which is a human B cell line.

[163]

[164] 3-1. Incubation of cell lines U266B1

[165] U266B1 cells (ATCC NO. TIB-196) were cultured in RPMI-1640 medium supplemented with 10% FBS, penicillin (100 µg/mL), and streptomycin (100 µg/mL) at 37°C under 5% CO2 and then reseeded every three days. U266B1 cells were seeded on a 24-well plate at a concentration of 1×10 ⁵ cells/well, then cultured for 16-20 hours to induce inflammatory reactions, and then treated with strains.

[166]

[167] 3-2. Strain incubation and recovery

[168] The strains used in the present experiment were prepared in the same manner as in Example 2-2.

[169]

[170] 3-3. Treatment of strains and quantitative measurement of IgE secretion after induction of allergic reactions

[171] To induce allergic reactions in U266B1 cells previously seeded in a 24-well plate, each well was treated with 100 μl of LPS (10 ng/ml) and IL-4 (5 ng/ml). Then, 300 μl of previously obtained strains were processed in a cell:strain ratio of 1:10. After incubation for 48 hours in a 5% CO ₂ incubator at 37° C., the supernatant was collected and human IgE ELISA kits (Catalog # E88-108, Bethyl Laboratories) were used for measurement according to the manufacturer's instructions.

[172]

[173] As a result, as shown in FIG. 4 confirmed that the groups treated with seven strains of Lactobacillus crispatus , including KBL693, showed effects of inhibiting IgE secretion compared to the negative control group. The present experiment showed that KBL693 could have a therapeutic and prophylactic effect in relation to allergic diseases due to the inhibition of IgE, the main factor involved in allergic reactions.

[174]

[175] Example 4 Analysis of Immunoregulatory and Inflammatory Inhibition Effects of KBL693

[176] In addition to the anti-allergic efficacy of KBL693, its immunoregulatory and inflammation-inhibiting effects were also tested. For this purpose, the ratio between IL-10, a typical cytokine with immunoregulatory function, and TNF-α and IL-6, which are cytokines - the main indicators of the inflammatory response (IL-10/TNF-α, IL-10/IL-6 ), using macrophages that play an important role in the inflammatory response.

[177]

[178] 4-1. Incubation of RAW264.7 Cell Lines

[179] RAW264.7 cells (ATCC NO. TIB-71) were cultured in DMEM supplemented with 10% FBS, penicillin (100 µg/mL), and streptomycin (100 µg/mL) at 37°C under 5% CO2 and then reseeded every three days. Incubated RAW264.7 cells were seeded on a 24-well plate at a concentration of 1×10 ⁵ cells/well, then cultured for 16-20 hours, and then used for example 4-3.

[180]

[181] 4-2. Incubation of strains and obtaining samples

[182] Culture solutions were corrected for the same amount of live bacteria and then obtained as three samples: live bacteria, pasteurized bacteria, and heat-killed bacteria. Samples of live bacteria, bacteria after pasteurization and bacteria killed by heat were obtained in accordance with the method used in example 2-2.

[183] Then, the pasteurization sample was subjected to a temperature of 70°C for 30 minutes, and a sample was obtained using the same centrifugation process. A sample of heat-killed bacteria was sterilized at 121°C for 15 minutes and obtained using the same process. MRS medium supplemented with 0.5% cysteine was used as a negative control group.

[184]

[185] 4-3. Treatment of strains and measurement of secreted inflammatory cytokines after induction of inflammatory responses

[186] To induce inflammatory responses in RAW264.7 cells previously seeded in a 24-well plate, each well was treated with 500 μl LPS (20 ng/ml). Then, 300 μl of the previously obtained strains were treated in a cell:strain ratio of 1:10 and incubated for 24 hours in an incubator with an atmosphere of 5% CO ₂ at 37°C. The supernatant of the cultured cell-strain mixture was harvested and a mouse TNF solid phase ELISA kit (Mouse TNF ELISA Set II, cat. no. 558534, BD OptEIA™), a mouse IL-6 solid phase ELISA kit (Mouse IL-6 ELISA Set, no. 555240, BD OptEIA™) and Mouse IL-10 ELISA Set, Cat No. 555252, BD OptEIA™) to measure the amount of each cytokine according to the manufacturer's instructions.

[187]

[188] As a result, as shown in FIG. 5 confirmed that, among strains of Lactobacillus crispatus, KBL693 was particularly effective in inducing IL-10 secretion, and thus had a remarkable effect in terms of immunoregulation and suppression of the inflammatory response. As shown in FIG. 6 and 7, based on the ability to secrete IL-10 adjusted for the pro-inflammatory cytokines TNF-α and IL-6, it was confirmed that the KBL693-treated group showed a significantly enhanced inflammation-suppressing effect of the immunoregulatory ability compared to the control group. Accordingly, KBL693 was found to have immunoregulatory and anti-inflammatory activity by secreting IL-10. In addition, these effects were similarly confirmed not only in live bacteria, but also in bacteria after pasteurization or bacteria killed by heat (see Table 2 and Table 3), indicating the possibility of using KBL693 to regulate immunity and suppress inflammatory responses in various forms, for example, in the form of dead bacteria.

[189]

[190] Table 2

IL-10/TNF-α live Killed by heat pasteurized Control -1.7574022 (N/O) -1.6268471 (N/O) -1.9978721 (N/O) KBL693 4.22079378 3.82025311 4.97474621

N/A: not determined.

[191] Table 3

IL-10/IL-6 live Killed by heat pasteurized Control -3.5789041 (N/O) -4.2268549 (N/O) -4.712413 (N/O) KBL693 4.95082596 4.9303055 6.46882207

N/A: not determined.

[192]

[193] Example 5 KBL693 Antifungal Assay

[194] The antifungal activity of KBL693 was confirmed by spot analysis. Approximately 1% KBL693 was inoculated into liquid MRS medium and then incubated in an incubator at 37° C. for approximately 24 hours under anaerobic conditions for stationary culture. Drops of the culture solution in which the cells were incubated were applied to the solid MRS medium prepared under anaerobic conditions at a rate of 10 μl at a time, and then incubated at 37° C. under anaerobic conditions for about 24 hours. Malassezia furfur KCTC 7545, a microscopic fungus, was obtained by inoculation in mYPG liquid medium prepared under aerobic conditions at 1% followed by incubation in an incubator at 37° C. for approximately 24-48 hours. Soft agar mYPG for use in evaluating antifungal efficacy was prepared with the addition of the components shown in Table 4, and 2.5 ml of the obtained culture medium was inoculated with a culture solution in which 500 μl of M. furfur were incubated. 2.5 ml mYPG soft agar inoculated with M. furfur was poured onto KBL693 dropped MRS medium and dried for 1 hour. The dried medium was incubated under aerobic conditions in an incubator at 37°C for 24-48 hours. When a clean zone was detected in the culture medium, antifungal activity was determined by measuring the distance from the outer boundary of the applied drop with lactic acid bacteria to the clean zone. The clean zone is an area with inhibited growth of fungi, and the antifungal activity of lactic acid bacteria was determined based on the distance to the clean zone.

[195] As a result of three repeated experiments, clear zones with an interval of approximately 4.33 mm were revealed in the area of the deposited droplets of KBL693 (Fig. 8), indicating the ability of KBL693 to effectively suppress the growth of microscopic fungi.

[196]

[197] Table 4

Components Share (g/l) Malt extract 5.0 Peptone 10.0 Glucose 20.0 Twin-40 1.0 Twin-80 1.0 agar 4.5

[198]

[199] Example 6. Effect of KBL693 on the mitigation of atopic conditions

[200] Based on a comprehensive review of in vitro screening results, animal tests were performed, selecting Lactobacillus crispatus KBL692, KBL693 and KBL702, which had the most pronounced immunoregulatory effect. The NC/Nga mice, an animal model of atopic skin disease, were used to test the mitigating effect on atopic conditions in the series of allergy-improving effects of KBL693.

[201] After dividing the NC/Nga mice into groups of five mice, hair was removed on the back of each mouse from the bottom of the ear to the top of the tail and the mice were left for 24 hours. Then, 200 µl of a 1% solution of DNCB (dinitrochlorobenzene) (acetone: olive oil=3:1) was applied to the dehaired area twice a week to induce atopic dermatitis. From the third week of induction of dermatitis, mice in the control group received 200 μl of PBS orally once a day; the cultured test strain was centrifuged, washed by PBS dilution and reconstitution, and then samples were obtained containing 2×10 ⁹ CFU per 200 μl of PBS, which were orally administered to mice in the experimental group in an amount of 200 μl/day. At the same time, mice in the positive control group were injected with 200 μl of dexamethasone (60 μg/ml). Then, during three weeks of bacteria administration, the dermatitis score of mice in the control and experimental groups was measured weekly, and at 3 weeks after the bacteria administration, the mice's scratching time and skin thickness were measured.

[202]

[203] 6-1. Performing dermatitis scoring

[204] To evaluate skin lesions caused by DNCB, the dermatitis score was measured as follows. The condition of the skin was monitored by taking pictures for 3 weeks at weekly intervals, starting from 3 weeks after the introduction of the strain. Checked four indicators - dryness, edema, erythema/bleeding and erosion/separation of the skin. The unaffected condition was scored as 0 points, the mild condition as 1 point, the moderate condition as 2 points, and the severe condition as 3 points, and an overall score was evaluated.

[205] As a result, as shown in FIG. 9, the DNCB dermatitis score was significantly reduced in the KBL693 strain group compared to the control group (negative control) where atopic dermatitis was induced and groups treated with other strains of Lactobacillus crispatus . As a result, the effect of treating atopic dermatitis by administering strain KBL693 was tested.

[206]

[207] 6-2. Effect on itch relief

[208] In order to test the mitigating effect on itching when the KBL693 strain was administered to mouse models suffering from DCNB-induced atopic dermatitis, scratching time was measured by videotaping the mouse models for 10 minutes 3 weeks after strain administration.

[209] As a result, as can be seen in FIG. 10, it appeared that the scratching time was significantly reduced in the experimental group treated with KBL693 compared with the control group (negative control) where atopic dermatitis was induced and groups treated with other strains of Lactobacillus crispatus , which confirmed a marked improvement in the symptoms of itching in atopic dermatitis for by introducing strain KBL693.

[210]

[211] 6-3. Reduction in skin thickness

[212] In order to test the effect of skin thickness reduction after administration of KBL693 to mouse models suffering from DCNB-induced atopic dermatitis, mouse ear thickness and dorsal skin thickness were measured with a caliper three weeks after strain administration, and a decrease in symptoms of edema due to atopic dermatitis was observed.

[213] As a result, as can be seen in FIG. 11 and 12, a significant decrease in the thickness of the ear and skin on the back was observed in the experimental group treated with KBL693 and the positive control group. In FIG. 11 and 12 693-7, 693-8 and 693-9, respectively, mean experimental groups in which KBL693 samples containing 2×10 ⁷ , 2×10 ⁸ and 2×10 ⁹ cfu per 200 μl PBS were used, which were injected mice in the experimental group at 200 µl/day.

[214]

[215] Example 7 Effect of KBL693 on Treatment and Prevention of Asthma in Ovalbumin Asthma (OVA) Mouse Models

[216] In order to test the effect of KBL693 (LC206) on allergic asthma, mouse models with OVA-induced asthma were prepared as follows, histopathological examination was performed, and the level of expression of IL-5 and IL-13 was evaluated.

[217]

[218] 7-1. Generation of mouse models with OVA-induced asthma

[219] Five week old Balb/c mice with an average body weight of approximately 15 g were subjected to acclimatization for one week, and then mice were selected that did not show pathologies on routine physical examination. As shown in FIG. 13(A), the mice used were assigned to the normal control group (control-PBS; no OVA administration or inhalation; n=8), the asthma-induced control group (OVA-PBS; OVA administration or inhalation; n=9), and the experimental group (OVA-LC206; n=8). Prior to sacrificing the mice at 31 days from the start of the experiment, the normal control group and the asthma-induced control group were orally administered 200 μl of PBS once a day, and the experimental group was orally administered 200 μl of KBL693 diluted with PBS to a concentration of at least 2×10 ⁹ once a day cfu/0.2 ml.

[220] Mice were sensitized by intraperitoneal injection of 200 μl of phosphate buffer (pH 7.4) in which 2 mg of aluminum hydroxide (Imject™ Alum Adjuvant, Thermofisher) and 20 μg of OVA (ovalbumin, Sigma-aldrich) were suspended, one week later (D7) and three weeks (D21) after acclimatization for one week, respectively. From day 28 to day 30 from the start of the experiment, 1% OVA was inhaled into the lungs via intranasal administration (three stimulations). 24 hours after the last stimulation (D31), pentobarbital was administered and then a bronchus incision was performed to collect lung tissue samples.

[221]

[222] 7-2. Histopathological examination

[223] Antigen-treated bronchi were infiltrated with inflammatory cells composed of eosinophils, neutrophils, and macrophages. Thus, in order to test the effect of KBL693 on asthma, a histopathological examination of lung tissue samples was performed in each group obtained in Example 7-1. The lung tissue samples obtained in Example 7-1 were prepared as 4 μm thick tissue sections using the conventional formalin-fixed paraffin-embedded (FFPE) tissue sectioning procedure and then stained with H&E hematoxylin and eosin Y (eosin Y; ThermoShandon , Pittsburgh, Pennsylvania, USA). After staining with H&E, tissue sections in each group were examined using an optical microscope (Fig. 13(B)).

[224] As a result, as shown in FIG. 13(B), in the control group with induced asthma (OVA-PBS), found infiltration of a large number of inflammatory cells, including eosinophils, around the bronchioles, as well as hyperproliferation of epithelial cells and thickening of the smooth muscles of the bronchi. On the other hand, in the experimental group treated with KBL693 (OVA-LC206), a significant reduction in inflammatory cell infiltration, including eosinophils, bronchial tissue thickness, and severe epithelial cell damage was observed. As a result, the therapeutic effect of KBL693 on OVA-induced allergic asthma was confirmed.

[225]

[226] 7-3. Analysis of the expression level of total IL-5 and total IL-13

[227] A flow cytometer (LSRFortessa X-20, BD) was used to evaluate the immunocytes in the lung tissue samples collected in each group. Antibodies to several markers were used to stain IL-5 ⁺ CD4 ⁺ T cells and IL-13 ⁺ CD4 ⁺ T cells (anti-mouse CD45, BioLegend; anti-mouse Cd3ε, BD; anti-mouse/human IL-5, BioLegend; anti-mouse IL-13 antibody, Invitrogen). IL-5 ⁺ - and IL-13 ⁺ cells were determined by counting cells producing IL-5 or IL-13 among lymphocytes with markers CD45 and CDε (Fig. 14).

[228]

[229] As a result, as shown in FIG. 14, it was found that the levels of IL-5 and IL-13 in mice in the control group with induced asthma (OVA-PBS) were significantly increased compared to the usual control group (PBS), while the levels of total IL-5 and total IL-13 in mice in the experimental group treated with KBL693 (OVA-KBL693) were significantly reduced compared to the control group with induced asthma. Accordingly, it was found that KBL693 according to the present invention could have a therapeutic and prophylactic effect in relation to allergic asthma by inhibiting the secretion of IL-5 and IL-13, which are Th2-type cytokines that mediate allergic reactions.

[230]

[231] Example 8 Effect of KBL693 on the Treatment and Prevention of Asthma in House Dust Mite Models of Asthma (HDM)

[232] House dust mites are an allergen that is the leading cause of exogenous asthma. In order to test the effect of KBL693 on allergic asthma, mouse models with HDM-induced asthma were obtained as described below, airway hypersensitivity was assessed and the proportion of eosinophils among CD45 ⁺ cells, the proportion of IL-5 ⁺ CD4 T cells among CD4 ⁺ T- cells and the proportion of IL-13 ⁺ CD4 T cells among CD4 ⁺ T cells.

[233]

[234] 8-1. Generation of Mouse Models with HDM-Induced Asthma

[235] Five-week-old Balb/c mice with an average body weight of approximately 15 g were acclimatized for one week, and then mice were selected that did not show pathologies on routine physical examination. The mice used were assigned to a normal control group (PBS control; no HDM injection), an asthma-induced control group (HDM-PBS; HDM administration), and an experimental HDM-KBL693 group, five mice from each group were used to evaluate airway hyperresponsiveness. (AHR), and eight mice from each group were used to evaluate immunocytes in the lungs.

[236] Mice were sensitized by intranasal administration of 50 μl of phosphate buffer (pH 7.4) in which 10 μg of HDM (Greer) was suspended one week after acclimatization for one week. One week after sensitization, 50 μl of phosphate buffer in which 10 μg of HDM had been suspended was inhaled into the lungs by intranasal administration for five days (D14~D18) (five stimulations). 24 hours after the last stimulation (D19), pentobarbital was administered, followed by an assessment of airway hyperresponsiveness (AHR, Example 8-2) and bronchus dissection to collect lung tissue samples.

[237]

[238] 8-2. AHR Score

[239] Pentobarbital anesthetized mice were connected to an AHR meter (FinePointe Resistance and Compliance, DSI-Buxco) and injected with various concentrations of methacholine dissolved in PBS (0, 5, 10, 20 and 40 mg/mL). The AHR value was then calculated by measuring the volume of air passed through the respiratory tract (FIG. 15).

[240] As a result, as shown in FIG. 15 confirmed that with increasing methacholine concentration, AHR (R _L ) slowly increased in the normal control group (CTRL) and rapidly increased in the induced asthma control (HDM) group. On the other hand, in the experimental group treated with KBL693 (HDM+KBL693), AHR was significantly reduced compared to the group with induced asthma (HDM) over the entire range of methacholine concentrations. This difference was more evident in the administration of high concentrations of methacholine compared to the area of its low concentrations. Accordingly, it was found that KBL693 could effectively suppress asthma-causing AHR and thus could be effectively used for the treatment and prevention of allergic asthma.

[241]

[242] 8-3. Determination of the proportion of eosinophils among CD45 ⁺ -cells, shares of IL-5 ⁺ CD4 T cells among CD4 ⁺ T cells and proportion of IL-13 ⁺ CD4 T cells among CD4 ⁺ T cells.

[243] A flow cytometer (LSRFortessa X-20, BD) was used to evaluate lung immunocytes. Antibodies to several markers were used to stain eosinophils, IL-5 ⁺ CD4 ⁺ T cells, and IL-13 ⁺ CD4 ⁺ T cells (anti-mouse CD45, BioLegend; rat anti-mouse Siglec-F, BD; anti-mouse Cd11b). , BD; anti-mouse Cd3ε antibody, BD; anti-mouse TCRβ antibody, BioLegend; anti-mouse CD4 antibody, BioLegend; anti-mouse/human IL-5 antibody, BioLegend; anti-mouse IL-13 antibody, Invitrogen). Eosinophils were determined by counting Siglec-f ⁺ Cd11b+ cells ⁺ cells among cells expressing CD45, a common leukocyte marker, and IL-5 ⁺ CD4 ⁺ T cells and IL-13 ⁺ CD4 ⁺ T cells were determined by counting cells producing IL-5 or IL-13 among CD4 ⁺ T cells with CDε, TCRβ and CD4 markers (FIG. 16).

[244] As a result, as shown in FIG. 16, it was confirmed that in the case of the control group with induced asthma (HDM), the proportion of immunocytes, i. eosinophils, IL-5 ⁺ CD4 T cells and IL-13 ⁺ CD4 T cells were significantly increased compared to the normal control group (PBS), while in the case of the experimental group treated with KBL693 (HDM+KBL693), the proportions of eosinophils , IL-5 ⁺ CD4 T cells and IL-13 ⁺ CD4 T cells were significantly reduced compared to the control group with induced asthma. Accordingly, it was found that KBL693 could have a therapeutic and prophylactic effect in relation to allergic asthma by inhibiting inflammatory cells, i.e. eosinophils, IL-5 ⁺ CD4 T cells and IL-13 ⁺ CD4 T cells.

[245]

[246] Specific aspects of the present invention are described in detail above, and for those skilled in the art it will be obvious that these specific aspects are only specific implementation options that do not limit the scope of the present invention. Thus, the scope of the present invention is largely defined by the following claims with the equivalents of the claims.

[247]

[248] Depositor name: Korea Research Institute of Bioscience & Biotechnology

[249] ID number: KCTC13519BP

[250] Access date: 20180427

[251]

Possibility of industrial application

[252] The strain of Lactobacillus crispatus KBL693 (ID number KCTC 13519BP) according to the present invention attenuates allergic cellular reactions, significantly improves the symptoms of atopic dermatitis, and has anti-inflammatory, immunoregulatory and antifungal effects. Thus, this single strain alone can achieve all the goals of mitigating allergic diseases and improving inflammatory diseases and autoimmune diseases, and thus successfully find use as a probiotic substance. In addition, this strain can be successfully used as an external skin treatment against various skin diseases caused by fungi, as well as in a cosmetic composition and a functional patch for alleviating skin sensitivity, based on its antifungal activity.

Sequence list text

[253] Electronic file attached

[254]

--->

<110> Ko BioLabs, Inc.

<120> Lactobacillus crispatus strain KBL 693 and its use

<130> P18-B095

<160> 1

<170> KoPatentIn 3.0

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<212> DNA

<213> Lactobacillus crispatus

<400> 1

cgcggggtgg cgcgagctat aatgcagtcg agcgagcgga actaacagat ttacttcggt 60

aatgacgtta ggaaagcgag cggcggatgg gtgagtaaca cgtggggaac ctgccccata 120

gtctgggata ccacttggaa acaggtgcta ataccggata agaaagcaga tcgcatgatc 180

agcttttaaa aggcggcgta agctgtcgct atgggatggc cccgcggtgc attagctagt 240

tggtaaggta aaggcttacc aaggcgatga tgcatagccg agttgagaga ctgatcggcc 300

acattgggac tgagacacgg cccaaactcc tacgggaggc agcagtaggg aatcttccac 360

aatggacgca agtctgatgg agcaacgccg cgtgagtgaa gaaggttttc ggatcgtaaa 420

gctctgttgt tggtgaagaa ggatagaggt agtaactggc ctttatttga cggtaatcaa 480

ccagaaagtc acggctaact acgtgccagc agccgcggta atacgtaggt ggcaagcgtt 540

gtccggattt attgggcgta aagcgagcgc aggcggaaga ataagtctga tgtgaaagcc 600

ctcggcttaa ccgaggaact gcatcggaaa ctgtttttct tgagtgcaga aggagagt 660

ggaactccat gtgtagcggt ggaatgcgta gatatatgga agaacaccag tggcgaaggc 720

ggctctctgg tctgcaactg acgctgaggc tcgaaagcat gggtagcgaa caggattaga 780

taccctggta gtccatgccg taaacgatga gtgctaagtg ttgggaggtt tccgcctctc 840

agtgctgcag ctaacgcatt aagcactccg cctggggagt acgaccgcaa ggttgaaact 900

caaaggaatt gacgggggcc cgcacaagcg gtggagcatg tggtttaatt cgaagcaacg 960

cgaagaacct taccaggtct tgacatctag tgccatttgt agagatacaa agttcccttc 1020

ggggacgcta agacaggtgg tgcatggctg tcgtcagctc gtgtcgtgag atgttgggtt 1080

aagtcccgca acgagcgcaa cccttgttat tagttgccag cattaagttg ggcactctaa 1140

tgagactgcc ggtgacaaac cggaggaagg tggggatgac gtcaagtcat catgcccctt 1200

atgacctggg ctacacacgt gctacaatgg gcagtacaac gagaagcgag cctgcgaagg 1260

caagcgaatc tctgaaagct gttctcagtt cggactgcag tctgcaactc gactgcacga 1320

agctggaatc gctagtaatc gcggatcagc acgccgcggt gaatacgttc ccgggccttg 1380

tacacaccgc ccgtcacacc atgggagtct gcaatgccca aagccggtgg cctaaccttc 1440

ggaaggagcc gtctaagtag acaatgtgca 1470

<---

Claims (25)

1. Strain of Lactobacillus crispatus KBL693 with identification number KCTC 13519BP for obtaining a probiotic agent. 2. The strain according to claim 1, characterized in that said strain contains the 16S rDNA sequence SEQ ID NO: 1. 3. A food composition containing an effective amount of at least one component selected from the group consisting of a strain according to claim 1, cultures of said strain, lysates of said strain, and extracts of said strain, and an acceptable carrier. 4. A food composition according to claim 3, characterized in that said composition has at least one action selected from the group consisting of alleviation of allergy symptoms, alleviation of inflammation symptoms, and immunoregulation. 5. A food composition according to claim 4, characterized in that said allergy symptoms are selected from the group consisting of eczema, allergic asthma, allergic rhinitis, atopic dermatitis, allergic conjunctivitis, and food allergy. 6. A feed composition containing an effective amount of at least one component selected from the group consisting of a strain according to claim 1, cultures of said strain, lysates of said strain and extracts of said strain, and an acceptable carrier. 7. An antifungal composition comprising an effective amount of at least one component selected from the group consisting of a strain according to claim 1, cultures of said strain, lysates of said strain, and extracts of said strain, and an acceptable carrier. 8. An antifungal composition according to claim 7, characterized in that said composition has antifungal activity against Malassezia furfur. 9. An external skin treatment comprising an effective amount of the antifungal composition of claim 7 and an acceptable carrier. 10. A medical patch for improving allergic diseases, containing an effective amount of at least one component selected from the group consisting of a strain according to claim 1, cultures of said strain, lysates of said strain and extracts of said strain, and an acceptable carrier. 11. A pharmaceutical composition for the treatment of allergic diseases, containing an effective amount of at least one component selected from the group consisting of a strain according to claim 1, cultures of said strain, lysates of said strain and extracts of said strain, and an acceptable carrier. 12. Pharmaceutical composition according to claim 11, characterized in that said allergic diseases are selected from the group consisting of eczema, allergic asthma, allergic rhinitis, atopic dermatitis, allergic conjunctivitis and food allergy. 13. A pharmaceutical composition for the treatment of inflammatory diseases, comprising an effective amount of at least one component selected from the group consisting of a strain according to claim 1, cultures of said strain, lysates of said strain and extracts of said strain, and an acceptable carrier. 14. The pharmaceutical composition according to claim 13, characterized in that these diseases are selected from the group consisting of rheumatoid arthritis, rheumatic fever, lupus, systemic scleroderma, atopic dermatitis, psoriasis, psoriatic arthritis, asthma, Guillain-Barré syndrome, myasthenia gravis, dermatomyositis, polymyositis, multiple sclerosis, autoimmune encephalomyelitis, periarteritis nodosa, Hashimoto's thyroiditis, temporal arteritis, juvenile diabetes, alopecia areata, pemphigus, aphthous stomatitis, autoimmune hemolytic anemia, Wegener's granulomatosis, Sjogren's syndrome, Addison's disease, Beckochett's disease, Crohn's disease , conjunctivitis, periodontitis, rhinitis, otitis media, chronic sinusitis, pharyngolaryngitis, tonsillitis, bronchitis, pneumonia, stomach ulcers, gastritis, colitis, gout, eczema, acne, contact dermatitis, seborrheic dermatitis, ankylosing spondylitis, fibromyalgia, osteoarthritis, shoulder periarthritis joints, tendonitis, tendosynovitis, myositis, hepatitis, cystitis, nephritis, sepsis, vasculitis and bursitis. 15. Pharmaceutical composition for the treatment of autoimmune diseases, containing an effective amount of at least one component selected from the group consisting of a strain according to claim 1, cultures of the specified strain, lysates of the specified strain and extracts of the specified strain, and an acceptable carrier. 16. Pharmaceutical composition according to claim 15, characterized in that said diseases are selected from the group consisting of rheumatoid arthritis, rheumatic fever, lupus, systemic scleroderma, atopic dermatitis, psoriasis, psoriatic arthritis, asthma, Guillain-Barré syndrome, myasthenia gravis, dermatomyositis, polymyositis, multiple sclerosis, autoimmune encephalomyelitis, periarteritis nodosa, Hashimoto's thyroiditis, temporal arteritis, juvenile diabetes, alopecia areata, pemphigus, aphthous stomatitis, autoimmune hemolytic anemia, Wegener's granulomatosis, Sjogren's syndrome, Addison's disease, Beckochett's disease, Crohn's disease , conjunctivitis, periodontitis, rhinitis, otitis media, chronic sinusitis, pharyngolaryngitis, tonsillitis, bronchitis, pneumonia, stomach ulcers, gastritis, colitis, gout, eczema, acne, contact dermatitis, seborrheic dermatitis, ankylosing spondylitis, fibromyalgia, osteoarthritis, shoulder periarthritis joints, tendonitis, tendosynovitis, myositis, hepatitis, cystitis, nephritis, sepsis, vasculitis and bursitis. 17. A method for treating allergic diseases, comprising administering at least one component selected from the group consisting of a strain according to claim 1, cultures of said strain, lysates of said strain, and extracts of said strain, to a subject in need thereof. 18. A method for treating inflammatory diseases, comprising administering at least one component selected from the group consisting of a strain according to claim 1, cultures of said strain, lysates of said strain, and extracts of said strain, to a subject in need thereof. 19. A method for treating autoimmune diseases, comprising administering at least one component selected from the group consisting of a strain according to claim 1, cultures of said strain, lysates of said strain, and extracts of said strain, to a subject in need thereof. 20. Use of a composition containing at least one component selected from the group consisting of a strain according to claim 1, cultures of said strain, lysates of said strain, and extracts of said strain, for the preparation of a prophylactic or therapeutic drug against allergic diseases. 21. Use of a composition containing at least one component selected from the group consisting of a strain according to claim 1, cultures of said strain, lysates of said strain, and extracts of said strain, for the preparation of a prophylactic or therapeutic drug against inflammatory diseases. 22. The use of a composition containing at least one component selected from the group consisting of a strain according to claim 1, cultures of the specified strain, lysates of the specified strain and extracts of the specified strain, for the production of a prophylactic or therapeutic drug against autoimmune diseases. 23. A cosmetic composition containing an effective amount of at least one component selected from the group consisting of a strain according to claim 1, cultures of the specified strain, lysates of the specified strain and extracts of the specified strain, and an acceptable carrier. 24. Cosmetic composition according to claim 23, characterized in that said cosmetic composition improves skin allergies, urticaria, atopic dermatitis, psoriasis, fungal infection or eczema. 25. Cosmetic composition according to claim 24, characterized in that said cosmetic composition is in the form of a cosmetic patch.

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