Antiviral drugs to prevent influenza from becoming severe

Abstract

The present invention relates to an anti-influenza virus agent containing Bifidobacterium bifidum as an active ingredient.

Description

[Technical Field]

[0001] The present invention relates to an anti-influenza virus agent for suppressing the aggravation of influenza. [Background technology]

[0002] Influenza is an infectious disease caused by infection with the influenza virus. The influenza virus is highly contagious, infecting approximately 10 million people in Japan every year. In addition, because influenza is epidemic, once an epidemic begins, it spreads to many people in a short period of time. Elderly people and those with chronic diseases are more likely to develop severe symptoms when they contract influenza, and are at higher risk of developing complications such as pneumonia and even death.

[0003] Anti-influenza drugs such as oseltamivir (trade name Tamiflu®) are commonly used to treat influenza. Oseltamivir must be administered early, within 48 hours of the onset of symptoms. However, if testing is performed too early, the virus may not be detected and a diagnosis of influenza may not be made, making it difficult to administer at the appropriate time. Therefore, there is a need for the development of a product that can be easily taken like a food product and that, when taken in advance, can be used to prevent the severity of influenza virus infection.

[0004] Patent Document 1 discloses that Bifidobacterium lactis strain BL-04 is effective in treating or preventing respiratory diseases such as rhinitis and bronchitis.

[0005] Patent Document 2 discloses a composition for treating or preventing infectious diseases and disorders related to the immune system, including infectious diseases, which comprises Bifidobacterium longum NCC2705.

[0006] Bifidobacterium bifidum OLB6378 strain, a type of bifidobacterium, has been confirmed to have the effect of enhancing intestinal mucosal immunity and to have the effect of preventing and treating allergies (Non-Patent Document 1 and Patent Document 3).

[0007] However, it has not been shown whether these microorganisms reduce the severity of influenza. [Prior art documents] [Patent documents]

[0008] [Patent Document 1] Special Publication No. 2014-517003 [Patent Document 2] Special Publication No. 2012-526752 [Patent Document 3] Japanese Patent Application Laid-Open No. 2006-273852 [Non-patent literature]

[0009] [Non-Patent Document 1] Tanaka K. et al., Nutrients, (2017) 9, 195; doi:10.3390 / nu9030195 Summary of the Invention [Problem to be solved by the invention]

[0010] An objective of the present invention is to provide an anti-influenza virus agent for suppressing the aggravation of influenza. [Means for solving the problem]

[0011] As a result of extensive research to solve the above problems, the present inventors discovered a Bifidobacterium bifidum strain that can suppress the aggravation of influenza, thereby completing the present invention.

[0012] That is, the present invention includes the following. [1] An anti-influenza virus agent containing Bifidobacterium bifidum as the active ingredient. [2] The anti-influenza virus agent according to [1] above, wherein Bifidobacterium bifidum has an inhibitory effect on the proliferation of influenza viruses in the respiratory tract. [3] The anti-influenza virus agent according to [1] or [2] above, wherein the Bifidobacterium bifidum is Bifidobacterium bifidum OLB6378 strain (accession number NITE BP-31). [4] The anti-influenza virus agent according to any one of the above [1] to [3], for suppressing the aggravation of influenza. [5] The anti-influenza virus agent according to any one of [1] to [4] above, wherein the Bifidobacterium bifidum is selected from the group consisting of a culture of Bifidobacterium bifidum cells, a heat-treated product, a dried product, a concentrate, and a combination of two or more of them. [6] A food for suppressing the aggravation of influenza, comprising the anti-influenza virus agent according to any one of [1] to [5] above. [7] A pharmaceutical for suppressing the aggravation of influenza, comprising the anti-influenza virus agent according to any one of [1] to [5] above. [Effects of the Invention]

[0013] According to the present invention, it is possible to prevent the aggravation of influenza virus infection. This specification includes the disclosure of Japanese Patent Application No. 2018-127101, from which the present application claims priority. [Brief explanation of the drawings]

[0014] [Figure 1]Figure 1 shows the effect of the OLB6378 strain on the survival rate of mice infected with influenza virus in the lower respiratory tract. *p=0.05 (compared to the water-administered group). n=9. In the figure, squares represent the survival rate of the oseltamivir-administered group (positive control group), black circles represent the survival rate of the OLB6378-administered group, and white circles represent the survival rate of the water-administered group. [Figure 2] Figure 2 shows the effect of the OLB6378 strain on the infectious virus titer in bronchoalveolar lavage fluid of mice infected with influenza virus in the lower respiratory tract. **p<0.01, ***p<0.001 (compared to the water-administered group). DETAILED DESCRIPTION OF THE INVENTION

[0015] The present invention will be described in detail below. The present invention relates to the use of Bifidobacterium bifidum, preferably Bifidobacterium bifidum having an inhibitory effect on the proliferation of influenza viruses in the respiratory tract, as an anti-influenza virus agent.

[0016] "Bifidobacterium bifidum" used in the present invention is a species of the genus Bifidobacteria. In the present invention, bacteria can be classified into Bifidobacterium bifidum according to standard taxonomic standards. The strain of Bifidobacterium bifidum used in the present invention is not particularly limited, but preferably has an inhibitory effect on influenza virus proliferation in the respiratory tract.

[0017] In the present invention, Bifidobacterium bifidum "has an inhibitory effect on influenza virus proliferation in the respiratory tract" means that it has the ability to inhibit the proliferation of influenza viruses in the respiratory tract, i.e., the upper and / or lower respiratory tract. The "upper respiratory tract" typically refers to the nasal cavity, pharynx, and larynx. The "lower respiratory tract" typically refers to the trachea, bronchi, and lungs. Bifidobacterium bifidum used in the present invention can inhibit the proliferation of influenza viruses in at least one (e.g., two or more) of the respiratory tract, such as the nasal cavity, pharynx, larynx, trachea, bronchi, and lungs.

[0018] In the present invention, "inhibition" of influenza virus proliferation in the respiratory tract refers to a reduction in the infectious virus titer of influenza virus recovered from the respiratory tract when a test substance (here, Bifidobacterium bifidum) is administered, compared to when water is administered instead of the test substance. Such a reduction in infectious virus titer is preferably a statistically significant reduction, and may be a reduction that is statistically significant, for example, by Dunnett's test. Such a reduction in infectious virus titer when a test substance is administered may also be a reduction of 30% or more, or 40% or more, for example, 50% or more, on average, compared to when water is administered instead of the test substance.

[0019] An example of Bifidobacterium bifidum that can be used in the present invention is the Bifidobacterium bifidum OLB6378 strain. The Bifidobacterium bifidum OLB6378 strain was deposited on October 26, 2004 (original deposit date) at the National Institute of Technology and Evaluation (NPMD) (Room 122, 2-5-8 Kazusa Kamatari, Kisarazu City, Chiba Prefecture, Japan, Postal Code 292-0818) under accession number NITE P-31. On January 18, 2006, the original deposit was transferred to a deposit under the Budapest Treaty, and the accession number was changed to NITE BP-31. The Bifidobacterium bifidum OLB6378 strain has the effect of inhibiting influenza virus proliferation in the respiratory tract. The current depositor of Bifidobacterium bifidum OLB6378 is Meiji Co., Ltd.

[0020] The present invention relates to an anti-influenza virus agent containing, as an active ingredient, Bifidobacterium bifidum, preferably Bifidobacterium bifidum having an inhibitory effect on the proliferation of influenza viruses in the respiratory tract. The anti-influenza virus agent according to the present invention may contain an effective amount of such Bifidobacterium bifidum.

[0021] The anti-influenza virus agent of the present invention can be used to inhibit the proliferation of influenza viruses, typically in the respiratory tract. The influenza viruses to which the agent is applied may be, but are not limited to, type A (such as the Soviet type, pdm09 type, or Hong Kong type), type B, type C, or a new strain. The influenza A virus may be, but is not limited to, a subtype such as H1N1, H1N2, H2N2, H2N3, H3N2, H5N1, H5N6, H7N7, H7N9, or H10N8.

[0022] In the present invention, Bifidobacterium bifidum can be used in any form in anti-influenza virus agents and foods and pharmaceuticals described below, as long as it retains its anti-influenza virus activity, preferably the ability to inhibit influenza virus proliferation in the respiratory tract and the ability to suppress the aggravation of influenza, as described below. In one embodiment, the Bifidobacterium bifidum used in the anti-influenza virus agents, foods, and pharmaceuticals may be bacterial cells prepared by any method and / or any processed bacterial cell product, such as a culture of Bifidobacterium bifidum bacterial cells, a heat-treated product, a dried product (e.g., a lyophilized product such as a freeze-dried powder), a concentrate, or a combination of two or more thereof. Bifidobacterium bifidum may be killed or live bacterial cells. In one embodiment, Bifidobacterium bifidum may be a heat-treated concentrated bacterial liquid or a lyophilized product thereof. In one embodiment, the heat treatment of Bifidobacterium bifidum can be carried out at 70 to 90°C (e.g., 75 to 85°C), for example, for 5 to 30 minutes, but is not limited to these conditions.

[0023] The above-mentioned Bifidobacterium bifidum according to the present invention and anti-influenza virus agents containing the same have the effect of suppressing the aggravation of influenza. In the present invention, "severe influenza" refers to the development of severe symptoms that may reduce the survival rate after influenza virus infection. Examples of such severe symptoms include complications such as bronchitis, pneumonia, influenza encephalopathy, encephalitis, otitis media, and febrile convulsions, as well as high fever and severe general fatigue. In the present invention, "suppressing the aggravation of influenza" encompasses reducing the probability of developing such severe symptoms after influenza virus infection, preventing the onset of severe symptoms or reducing their severity, and promoting early recovery from severe symptoms. Therefore, the anti-influenza virus agents according to the present invention can be used to suppress the aggravation of influenza. Suppression of the aggravation of influenza may be assessed, for example, by improving the survival rate after influenza virus infection. If the survival rate after influenza virus infection is increased when the test substance (here, Bifidobacterium bifidum) is administered compared to when water is administered instead of the test substance, it can be determined that the test substance suppresses the aggravation of influenza. Such an increase in survival rate is preferably a statistically significant increase, and may be an increase that shows a statistically significant difference, for example, by a log-rank test using the Kaplan-Meier method. The anti-influenza virus agent of the present invention (and the above-mentioned Bifidobacterium bifidum) can suppress the aggravation of influenza, thereby reducing the risk of influenza aggravation.

[0024] The present invention also provides a food or medicine comprising the anti-influenza virus agent of the present invention (or the above-mentioned Bifidobacterium bifidum). The food and medicine comprising the anti-influenza virus agent of the present invention (or the above-mentioned Bifidobacterium bifidum) can suppress the aggravation of influenza in a subject who ingests or administers it. The food and medicine of the present invention can suppress the proliferation of influenza viruses in the respiratory tract. The food and medicine of the present invention can be used to suppress the aggravation of influenza. The food and medicine of the present invention can also be used to reduce the risk of aggravation of influenza.

[0025] The anti-influenza agents, foods, and medicines according to the present invention may contain additives acceptable for oral ingestion or pharmaceutically acceptable. Examples of additives acceptable for oral ingestion or pharmaceutically acceptable include, but are not limited to, carriers (e.g., solid or liquid carriers), excipients, surfactants, binders, disintegrants, lubricants, solubilizers, suspending agents, coating agents, colorants, flavorings, preservatives, buffers, pH adjusters, diluents, stabilizers, propellants, antioxidants, thickeners, and sweeteners. These additives may be used alone or in combination of two or more, and may be used appropriately depending on the dosage form of the preparation and the type or shape of the food. Examples of carriers include water, organic solvents acceptable for oral administration, collagen, polyvinyl alcohol, polyvinylpyrrolidone, carboxyvinyl polymer, sodium alginate, water-soluble dextran, water-soluble dextrin, sodium carboxymethyl starch, pectin, xanthan gum, gum arabic, casein, gelatin, agar, glycerin, propylene glycol, polyethylene glycol, petrolatum, paraffin, stearyl alcohol, stearic acid, human serum albumin, mannitol, sorbitol, lactose, and surfactants acceptable for oral administration.

[0026] The term "food" as used herein is not particularly limited, and includes beverages, non-beverage foods, and functional foods. The term "food" as used herein may also be a food composition. The type of food product used herein is not particularly limited, and examples of beverages include fermented milk (e.g., yogurt drinks), lactic acid bacteria beverages, dairy beverages (e.g., coffee milk, fruit milk), tea beverages (e.g., green tea, black tea, oolong tea), fruit and vegetable beverages (e.g., beverages containing orange, apple, grape, or other fruit juice, or tomato, carrot, or other vegetable juice), alcoholic beverages (e.g., beer, happoshu, wine), oral rehydration solutions, carbonated beverages, soft drinks, and water. For information on the production methods of various beverages, existing reference books, such as "Latest Soft Drinks" (2003) (Korin Co., Ltd.), can be consulted. Examples of foods include fermented milk (e.g., set yogurt, soft yogurt, cheese), dairy products, modified milk powder, confectioneries (e.g., chocolate, gummies, chewing gum, biscuits, cookies, jellies, tablets, frozen desserts), and instant foods. Existing reference books can be used as references for the manufacturing methods of various foods.

[0027] The food of the present invention may be a functional food. The term "functional food" as used herein refers to a food that has a certain functionality for the living body, and includes, for example, health functional foods, including foods for specified health uses (including conditional FOSHUs [foods for specified health uses]) and foods with nutrient function claims in Japan, foods with functional claims, and foods for special dietary uses, as well as so-called health foods in general, such as nutritional supplements, health supplements, supplements (in various dosage forms, such as tablets, coated tablets, sugar-coated tablets, capsules, and liquids), and beauty foods (e.g., diet foods). Furthermore, the functional food of the present invention also includes health foods to which a health claim based on the Codex Alimentarius (Joint FAO / WHO Food Standards Commission) food standards is applicable.

[0028] The functional food of the present invention may be in the form of a solid preparation such as a tablet, granule, powder, pill, or capsule; a liquid preparation such as a liquid, suspension, or syrup; or a gel or paste; or may be in the form of a regular food or beverage (e.g., a beverage, yogurt, or confectionery).

[0029] The food product of the present invention may be intended for subjects who are susceptible to influenza or who are susceptible to developing severe influenza symptoms when infected. The food product of the present invention may be intended for consumption by newborns (particularly including newborns, healthy newborns, premature babies, premature babies, and low birth weight babies), infants, young children, sick people, frail healthy people, unwell healthy people, pregnant women, elderly people (typically 60 years of age or older), chronically ill people (e.g., patients with heart disease, respiratory disease, renal dysfunction, immune dysfunction, or metabolic diseases such as diabetes), and their family members and other close contacts (e.g., medical professionals, childcare and education professionals, and life support workers). The food product of the present invention may also be a special-purpose food, such as food for sick people, powdered milk for pregnant and nursing women, infant formula, infant formula, food for the elderly, or food for caregivers. The food product of the present invention may be powdered milk, liquid food, semi-liquid food, powdered liquid food, nursing care food, infant food, thickened food, energy paste, nutritional powder, nutritionally fortified food, liquid milk, etc.

[0030] The Bifidobacterium bifidum or anti-influenza virus agent of the present invention may be incorporated into a food product by any suitable method available to those skilled in the art. For example, the Bifidobacterium bifidum of the present invention may be prepared in a liquid, gel, solid, powder, or granular form and then incorporated into a food product. Alternatively, the Bifidobacterium bifidum or anti-influenza virus agent of the present invention may be directly mixed or dissolved in the ingredients of a food product. The Bifidobacterium bifidum or anti-influenza virus agent of the present invention may be applied to, coated on, impregnated into, or sprayed onto a food product. The Bifidobacterium bifidum or anti-influenza virus agent of the present invention may be uniformly dispersed or unevenly distributed throughout the food product. Capsules containing the Bifidobacterium bifidum or anti-influenza virus agent of the present invention may also be prepared. The Bifidobacterium bifidum or anti-influenza virus agent of the present invention may be encapsulated in an edible film, edible coating, or the like. Furthermore, Bifidobacterium bifidum or the anti-influenza virus agent according to the present invention may be added with an appropriate excipient, etc., and then molded into a shape such as a tablet. Foods and drinks containing Bifidobacterium bifidum or the anti-influenza virus agent according to the present invention may be further processed, and such processed products are also encompassed within the scope of the present invention. In one embodiment, the Bifidobacterium bifidum cell concentrate according to the present invention may be heat-treated and added to a food raw material (e.g., powdered milk preparation, liquid food raw material, etc.), followed by heat sterilization and concentration / drying.

[0031] The amount of Bifidobacterium bifidum or anti-influenza virus agent of the present invention to be added to a food product is not particularly limited and may vary depending on the case. The specific amount of Bifidobacterium bifidum or anti-influenza virus agent of the present invention to be added can be determined appropriately by those skilled in the art, taking into consideration the type of food and the desired taste and texture. Typically, the amount of Bifidobacterium bifidum to be added can be 0.0001 to 100% by weight (mass), for example 0.0005 to 100% by weight (mass), or 0.001 to 99% by weight (mass), in terms of the dry weight equivalent of Bifidobacterium bifidum cells (solid cell content) relative to the total weight of the food product. In one embodiment, Bifidobacterium bifidum may be incorporated into a solid food, such as a powdered food (formulated powdered milk, powdered seasoning, etc.), a granular food, a flaked food, a disintegrating solid food, etc., at a ratio of the dry weight equivalent of Bifidobacterium bifidum cells (solid bacterial content) relative to the total weight of the food of 0.0001 to 100% by weight (mass), for example 0.001 to 99% by weight (mass), 0.01 to 10% by weight (mass), or 0.01 to 0.1% by weight (mass). Such solid foods may be dissolved in a drinkable liquid medium such as water or a beverage, and prepared as a beverage (liquid milk (liquid milk), oral rehydration solution, etc.), liquid food (soup, curry, etc.), liquid seasoning, liquid food, semi-liquid food, thickened food, etc., for consumption. In another embodiment, Bifidobacterium bifidum may be blended into liquid or semi-liquid foods, such as soup, curry, fermented milk, beverages (liquid milk, oral rehydration solution, etc.), liquid seasonings, liquid foods, semi-liquid foods, thickened foods, etc., at a ratio of 0.0001 to 100% by weight (mass), for example 0.0005 to 10% by weight (mass), or 0.001 to 5% by weight (mass), based on the total weight of the food, equivalent to the dry weight of Bifidobacterium bifidum bacteria (solid bacterial content).The above-mentioned amount of Bifidobacterium bifidum expressed in terms of the dry weight equivalent of the bacterial cells does not necessarily mean that Bifidobacterium bifidum is incorporated into foods, etc. in the form of dried bacterial cells. The Bifidobacterium bifidum of the present invention can be incorporated into foods, etc. in any form as long as it retains its anti-influenza virus activity, particularly the activity of inhibiting the proliferation of influenza viruses in the respiratory tract and / or the activity of suppressing the aggravation of influenza.

[0032] The food product of the present invention may further contain any food ingredient. The food product of the present invention may contain water, protein, carbohydrates, lipids, vitamins, minerals, amino acids, organic acids, organic bases, fruit juice, flavors, etc. Examples of proteins include whole milk powder, skim milk powder, partially skim milk powder, casein, whey powder, whey protein, whey protein concentrate, whey protein isolate, α-casein, β-casein, κ-casein, β-lactoglobulin, α-lactalbumin, lactoferrin, soy protein, egg protein, meat protein, and other animal and plant proteins, their hydrolysates, butter, whey minerals, cream, whey, non-protein nitrogen, sialic acid, phospholipids, and various milk-derived components such as lactose. Examples of carbohydrates include common sugars, modified starches (dextrin, soluble starch, British starch, oxidized starch, starch esters, starch ethers, etc.), and dietary fiber. Examples of lipids include animal fats and oils such as lard, fish oil, and their fractionated, hydrogenated, and interesterified oils; and vegetable fats and oils such as palm oil, safflower oil, corn oil, rapeseed oil, coconut oil, and their fractionated, hydrogenated, and interesterified oils. Examples of vitamins include vitamin A, carotenes, B vitamins, vitamin C, D vitamins, vitamin E, K vitamins, vitamin P, vitamin Q, niacin, nicotinic acid, pantothenic acid, biotin, inositol, choline, and folic acid. Examples of minerals include calcium, potassium, magnesium, sodium, copper, iron, manganese, zinc, selenium, and whey minerals. Examples of organic acids include malic acid, citric acid, lactic acid, and tartaric acid. These components can be used alone or in combination.

[0033] The medicament according to the present invention may be in any dosage form, such as a solid preparation such as a tablet, granule, powder, pill, or capsule, or a liquid preparation such as a gel, solution, suspension, or syrup. The "medicament" according to the present invention may be a pharmaceutical composition.

[0034] The anti-influenza agent, food, and medicine according to the present invention may further contain other substances having anti-influenza activity.

[0035] The anti-influenza agent, food, and medicine according to the present invention may be for oral administration or for parenteral administration, but are preferably for oral administration.

[0036] The present invention also provides a method for treating or preventing influenza, which comprises administering the anti-influenza agent, food, or medicine of the present invention to a subject. The present invention also provides a method for suppressing the aggravation of influenza, which comprises administering the anti-influenza agent, food, or medicine of the present invention to a subject. In the present invention, "administration" includes "ingestion" as used with respect to foods, "administration" as used with respect to medicines, and in vivo "treatment" of cells or tissues. In the present invention, "oral administration" (also referred to as oral or enteral administration) includes not only oral administration or ingestion, but also administration by enteral feeding via a nasogastric tube, gastrostomy tube, enterostomy tube, or the like. The present invention also provides a method for reducing the risk of aggravation of influenza, which comprises administering the anti-influenza agent, food, or medicine of the present invention to a subject. The present invention further provides the use of the above-mentioned Bifidobacterium bifidum of the present invention, for example, the Bifidobacterium bifidum OLB6378 strain, in the manufacture of the anti-influenza agent, food, or medicine of the present invention.

[0037] The dosage of the anti-influenza agent, food, and medicine according to the present invention can be varied over a wide range at the discretion of a person skilled in the art, taking into consideration the age and weight of the subject to be administered, the route of administration, the number of administrations, etc. The dosage of Bifidobacterium bifidum according to the present invention is not particularly limited, but may be, for example, 1 x 10 per dose. 5 ~1×10 11 cells, 1 x 10 6 ~1×10 10 cells, or 1 × 10 7 ~1×10 10 It may be an amount sufficient to produce cells.

[0038] In one embodiment, the anti-influenza agents, foods, and medicines of the present invention may be administered to a subject once or more times a day, for example, twice or more times a day. The anti-influenza agents, foods, and medicines of the present invention may be administered to a subject continuously, for example, daily. The anti-influenza agents, foods, and medicines of the present invention may be administered to a subject for one week, two weeks, three weeks, four weeks, or more (for example, one year or more).

[0039] The anti-influenza agent, food, and medicine according to the present invention can be administered to a subject before or after infection with influenza virus, but is preferably administered before influenza virus infection.

[0040] The subjects to which the anti-influenza agents, foods, and medicaments of the present invention are administered may be any animal, preferably a mammal, such as a human, a gorilla, a chimpanzee, a dog, a cat, a mouse, a rat, a rabbit, a horse, a cow, a sheep, a goat, or the like, or cells thereof. The subjects to which the anti-influenza agents, foods, and medicaments of the present invention are administered may be subjects who are susceptible to influenza or who are susceptible to the development of severe influenza when infected. In a preferred embodiment, the human subjects may be, as described above, newborns, infants, young children, sick people, frail healthy individuals, unwell healthy individuals, pregnant women, elderly people, chronic disease patients, etc., as well as their family members and other close contacts. However, the subjects are not limited to these, and the anti-influenza agents, foods, and medicaments of the present invention can be used for a wide range of subjects at risk of influenza. [Example]

[0041] The present invention will be described in more detail below using examples, although the technical scope of the present invention is not limited to these examples.

[0042] [Example 1] Preparation of OLB6378 concentrated powder Bifidobacterium bifidum OLB6378 strain (accession number NITE BP-31) was anaerobically cultured in a whey protein hydrolysis medium (a medium containing enzymatically hydrolyzed whey protein as the main nitrogen source) under neutral conditions to obtain a concentrated bacterial suspension. The concentrated bacterial suspension was incubated at 80°C for 10 minutes or more and then freeze-dried. The resulting freeze-dried product contained 2.5 x 10 11 The freeze-dried product of the heat-treated OLB6378 strain thus prepared was designated OLB6378 concentrated powder and was tested for its anti-influenza virus activity as described below.

[0043] [Example 2] Evaluation of the anti-influenza virus activity of OLB6378 concentrated powder The anti-influenza virus activity of the OLB6378 concentrated powder prepared in Example 1 was evaluated in terms of "survival rate" and "infectious virus titer in the respiratory tract (virus multiplication level)" after influenza virus infection.

[0044] a) Survival rate BALB / c mice (female, 8 weeks old) (CLEA Japan) were orally administered water (water administration group) or a 20% suspension of OLB6378 concentrated powder (OLB6378 / Bifidobacteria administration group) at a dose of 0.2 ml / animal / day for 4 weeks. Three weeks after the start of oral administration, the mice were treated with the anesthetic somnopentyl (R) The animals were anesthetized by intraperitoneal administration of 5 mg / kg body weight of the active ingredient, sodium pentobarbital, and 20 μl (2 × LD ) of a suspension of mouse-adapted influenza virus strain A / PR / 8 / 34 (H1N1) was administered. 50Mice were inoculated intranasally into the right nostril with 0.1% BSA-PBS (equivalent to twice the LD50 dose) to induce lower respiratory tract infection with influenza virus. As a positive control, instead of water or a 20% suspension of OLB6378 concentrated powder, the anti-influenza drug oseltamivir (1 mg / kg body weight / day) was orally administered from 1 hour before to 6 days after influenza virus inoculation as described above. The survival rate of mice was recorded for 21 days after the start of virus inoculation. Survival was assessed using the Kaplan-Meier log-rank test.

[0045] The results are shown in Figure 1. In mice orally administered water (water-administered group), deaths began to be observed 8 days after virus inoculation (infection). Ten days after infection, survival rates dropped sharply to approximately 11%. Twenty days after infection, all mice had died (0% survival rate). In contrast, in mice orally administered a 20% suspension of OLB6378 concentrated powder (OLB6378-administered group), deaths began to be observed 9 days after virus inoculation (infection). However, survival rates remained approximately 56% at 10 days after infection and approximately 22% at 21 days after infection. The improvement in survival rate with administration of OLB6378 concentrated powder was statistically significant compared to the water-administered group (p=0.05, Figure 1).

[0046] In addition, no deaths were observed in the positive control group in which mice were orally administered the anti-influenza drug oseltamivir and then infected with influenza virus in their lower respiratory tracts (Figure 1).

[0047] b) Infectious virus titer (virus multiplication level) BALB / c mice (female, 8 weeks old) (CLEA Japan) were orally administered water (water administration group) or a 20% suspension of OLB6378 concentrated powder (OLB6378 administration group; test group) at a dose of 0.2 ml / animal / day for 25 days (from 21 days before to 3 days after virus inoculation). Three weeks after the start of oral administration, the mice were anesthetically administered with somnopentyl (R)The animals were anesthetized by intraperitoneal administration of 5 mg / kg body weight of the active ingredient, sodium pentobarbital, and 20 μl (2 × LD ) of a suspension of mouse-adapted influenza virus strain A / PR / 8 / 34 (H1N1) was administered. 50 Influenza virus infection was induced by intranasal inoculation of 0.1% BSA-PBS (equivalent to twice the LD50 concentration) into the right nasal cavity. As a positive control, instead of water or a 20% suspension of OLB6378 concentrated powder, the anti-influenza drug oseltamivir (1 mg / kg body weight / day) was orally administered from 1 hour before to 3 days after the influenza virus inoculation. Four days after the virus inoculation, bronchoalveolar lavage fluid (BALF) was prepared from the mice. The infectious virus titer in the BALF was measured by the plaque assay. Specifically, MDCK cells (a cell line derived from canine kidney tubular epithelial cells) were exposed to the BALF, and the number of plaques formed due to cell degeneration caused by virus infection was counted. The number of plaques formed per mL of BALF was used to evaluate the infectious virus titer. Statistical analysis was performed using analysis of variance (ANOVA) and Dunnett's test.

[0048] The results are shown in Figure 2. Four days after influenza virus infection, mice orally administered water (water-administered group) had high infectious virus titers in bronchoalveolar lavage fluid, indicating that lower respiratory tract infection had been established. On the other hand, mice orally administered a 20% suspension of OLB6378 concentrated powder (OLB6378-administered group) showed a statistically significant decrease in infectious virus titers in bronchoalveolar lavage fluid compared to the water-administered group (a decrease of more than 53% compared to the water-administered group) (p < 0.01, Figure 2).

[0049] Furthermore, in the positive control group, in which mice were orally administered the anti-influenza drug oseltamivir and then infected with influenza virus in their lower respiratory tracts, a significant decrease in infectious virus titer in bronchoalveolar lavage fluid was observed compared to the water-administered group (p < 0.001, Figure 2).

[0050] The above survival rate and infectious virus titer (virus growth level) test results showed that the OLB6378 strain inhibits virus growth in the bronchi and lungs after influenza virus infection and improves survival rate, indicating that the OLB6378 strain has anti-influenza virus activity, particularly in suppressing the aggravation of influenza. [Industrial Applicability]

[0051] The present invention can be used to provide an anti-influenza product that is effective in suppressing the aggravation of influenza. By taking the anti-influenza virus agent of the present invention daily in the form of a food or the like, it is possible to suppress the aggravation of symptoms during influenza virus infection. The present invention can provide an anti-influenza product that is useful for suppressing the aggravation of influenza symptoms in subjects who are prone to the aggravation of influenza, such as newborns (particularly newborns), infants, children, sick people, elderly people, and patients with chronic diseases.

[0052] All publications, patents, and patent applications cited herein are hereby incorporated by reference in their entirety.

Claims

1. An anti-influenza virus agent containing Bifidobacterium bifidum as an active ingredient, which improves the survival rate after influenza virus infection by suppressing the aggravation of influenza.

2. 2. The anti-influenza virus agent according to claim 1, wherein Bifidobacterium bifidum has an inhibitory effect on the proliferation of influenza viruses in the respiratory tract after infection with influenza viruses.

3. 3. The anti-influenza virus agent according to claim 1 or 2, wherein the Bifidobacterium bifidum is Bifidobacterium bifidum OLB6378 strain (accession number NITE BP-31).

4. The anti-influenza virus agent according to any one of claims 1 to 3, wherein the Bifidobacterium bifidum is selected from the group consisting of a culture of Bifidobacterium bifidum cells, a heat-treated product, a dried product, a concentrate, and a combination of two or more of them.

5. A food for improving survival rate after influenza virus infection by suppressing the aggravation of influenza, comprising the anti-influenza virus agent according to any one of claims 1 to 4.

6. A pharmaceutical for improving the survival rate after influenza virus infection by suppressing the aggravation of influenza, comprising the anti-influenza virus agent according to any one of claims 1 to 4.

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