Warrosiella spp. LC-95 and application thereof

Abstract

The invention belongs to the technical field of biological medicines, and provides a Roseburia zhanii LC-95, which is preserved in the China General Microbiological Culture Collection Center on January 29, 2024, the China General Microbiological Culture Collection Center is called as CGMCC for short, the preservation number is CGMCC NO.29797, and the address of the preservation unit is No.3, Yard 1, Beichen West Road, Chaoyang District, Beijing. The strain has the effects of relieving constipation and cognitive impairment of middle-aged and elderly people, improving exercise coordination, relieving anxiety and the like.

Description

Technical Field

[0001] The present invention belongs to the technical field of biomedicine. Background Art

[0002] As the global aging population continues to intensify, the health of middle-aged and elderly people has become a focus of social attention. Chronic symptoms such as constipation, cognitive decline, decreased motor coordination, and anxiety, leading to functional constipation, anxiety disorders, and degenerative diseases such as Alzheimer's disease (AD) and Parkinson's disease (PD), are relatively common among middle-aged and elderly people, posing challenges to their quality of life and physical health [1,2].

[0003] Constipation is one of the common gastrointestinal problems in middle-aged and elderly people. Its main characteristics include difficult bowel movements, hard stools, and reduced bowel movement frequency. Constipation not only causes abdominal distension, discomfort, and pain, but may also lead to more serious health problems such as anal fissures and hemorrhoids [3,4]. The causes of constipation are diverse, including lifestyle, diet, drug side effects, nerve or muscle problems, chronic diseases, etc.; different types of constipation require different treatments [4]. Currently, the treatment of constipation mainly includes changing eating habits, increasing dietary fiber intake, drinking enough water, regular physical exercise to promote intestinal motility, and relying on various drugs such as mild laxatives, moisturizers, and intestinal stimulants [1,4]. However, there are some problems with existing constipation treatments; some drugs may cause dependence or side effects, and not everyone responds well to these drugs [1,5]. Therefore, it is very important to seek a more effective, personalized, and side-effect-free treatment for constipation.

[0004] In addition, the cognitive function and memory ability of middle-aged and elderly people usually decline with age, which affects their ability to live independently[2]. The weakening of motor coordination also makes middle-aged and elderly people more vulnerable to injury. At the same time, middle-aged and elderly people often face mental health problems such as anxiety and psychological stress, which further reduces their quality of life[6]. Currently, there are no drugs or therapies that can cure degenerative diseases such as Alzheimer's disease (AD) and Parkinson's disease (PD), and this problem needs to be solved urgently.

[0005] In order to cope with these chronic symptoms, live bacterial drugs have attracted more and more attention as a microecological therapy with few side effects and high acceptance. Live bacterial drugs contain active probiotics, which help maintain the balance of the host's intestinal flora and may have a positive impact on constipation, cognitive function, motor coordination, anxiety and other problems through various mechanisms. Studies have shown that live bacterial drugs may improve constipation symptoms by increasing intestinal motility, improving intestinal health, promoting the production of neuroprotective factors, and adjusting the immune system [7]. In addition, live bacterial drugs may also improve cognitive and emotional problems by affecting the gut-brain axis [8]. It is of great significance to explore the functional probiotics derived from the intestines of healthy people for the above indications and develop microecological live bacterial drugs.

[0006] References:

[0007] 1.Vazquez Roque,M.and EPBouras,Epidemiology and management ofchronic constipation in elderly patients.Clin Interv Aging,2015.10:p.919-30.

[0008] 2. Scarmeas, N., CANastasiou, and M. Yannakoulia, Nutrition and prevention of cognitive impairment. Lancet Neurol, 2018.17(11):p.1006-1015.

[0009] 3. Rao, SS, K. ​​Rattanakovit, and T. Patcharatrakul, Diagnosis and management of chronic constipation in adults. Nat Rev Gastroenterol Hepatol, 2016.13(5):p.295-305.

[0010] 4. Black, CJ and AC Ford, Chronic idiopathic constipation in adults: epidemiology, pathophysiology, diagnosis and clinical management. Med J Aust, 2018.209(2):p.86-91.

[0011] 5. Luthra, P., et al., Efficacy of drugs in chronic idiopathic constipation: a systematic review and network meta-analysis. Lancet Gastroenterol Hepatol, 2019. 4(11): p. 831-844.

[0012] 6. Ngandu, T., et al., A 2-year multidomain intervention of diet, exercise, cognitive training, and vascular risk monitoring versus control to prevent cognitive decline in at-risk elderly people (FINGER): a randomised controlled trial. Lancet, 2015. 385(9984): p. 2255-63.

[0013] 7. Ma, T., et al., Effect of the probiotic strain, Lactiplantibacillus plantarum P9, on chronic constipation: A randomized, double-blind, placebo-controlled study. Pharmacol Res, 2023. 191: p. 106755.

[0014] 8. Kim, C.S., et al., Probiotic Supplementation Improves Cognitive Function and Mood with Changes in Gut Microbiota in Community-Dwelling Older Adults: A Randomized, Double-Blind, Placebo-Controlled, Multicenter Trial. J Gerontol A Biol Sci Med Sci, 2021. 76(1): p. 32-40. Summary of the Invention

[0015] In order to effectively relieve constipation in middle-aged and elderly people, relieve anxiety, improve cognitive memory ability in middle-aged and elderly people to slow down Alzheimer's disease (AD), and improve motor coordination to relieve chronic symptoms such as Parkinson's disease (PD), the first purpose of the present invention provides Roseburia zhanii, whose representative strain LC-95 has been deposited in the General Microbiology Center of the China Culture Collection Administration on January 29, 2024. The center is abbreviated as CGMCC, the deposit number is: CGMCCNO.29797, and the deposit unit address is: No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing.

[0016] A second object of the present invention is to provide a microbial agent, the active ingredients of which include but are not limited to live Roseburia zhanii.

[0017] The third purpose of the present invention is the use of a live Roseburia bacterial agent, which can relieve constipation.

[0018] The fourth purpose of the present invention is to use a live Roseburia inoculum, which can relieve anxiety, improve cognitive memory, and enhance motor coordination. BRIEF DESCRIPTION OF THE DRAWINGS

[0019] Figure 1 .Cell morphology and size under LC-95 transmission electron microscope.

[0020] Figure 2 Colony morphology of LC-95 live bacterial strain.

[0021] Figure 3 Flow chart of the experimental design for oral administration of LC-95 live bacteria. Specific implementation method:

[0022] Example

[0023] Modified GAM liquid medium (g / L): 10 g casein peptone, 3 g soy peptone, 15 g tryptone, 13.5 g digested serum, 5 g yeast extract, 2 g beef meal, 1.2 g bovine liver extract, 3 g glucose, 0.3 g soluble starch, 0.5 g L-cysteine ​​hydrochloride, 0.5 g L-arginine, 0.3 g L-tryptophan, 2 g sodium bicarbonate, 2.5 g potassium dihydrogen phosphate, 3 g NaCl, 0.15 g sodium thioglycolate, 2.46 g sodium acetate, 0.01 g hemin, 0.001 g resazurin, 10% (v / v) clarified rumen fluid, distilled water to 1 L, pH 7.2 ± 0.1, autoclaved at 115°C for 25 min. For solid medium, add an additional 15 g agar.

[0024] 1. Acquisition, isolation and identification of strains

[0025] The bacterial strain was obtained by the patent inventor on October 5, 2019, through anaerobically separation from fresh feces provided by the feces donor after he signed an informed consent form.

[0026] Colony morphology: In modified GAM broth medium (each liter of modified GAM broth medium contains: casein peptone 10 g, soy peptone 3 g, tryptone 15 g, digested serum 13.5 g, yeast extract 5 g, beef powder 2 g, bovine liver extract 1.2 g, glucose 3 g, soluble starch 0.3 g, L-cysteine ​​hydrochloride 0.5 g, L-arginine 0.5 g, L-tryptophan 0.3 g, sodium bicarbonate 2 g, potassium dihydrogen phosphate 2.5 g, NaCl 3 g, sodium thioglycolate 0.15 g, sodium acetate 2.46 g, hemin 0.01 g, resazurin 0.001 g, clarified rumen fluid 10% (v / v), distilled water to 1 L, pH 7.2±0.1, autoclaved at 115℃ for 25 minutes. 15g of agar should be added to the solid medium.) This bacterium is strictly anaerobic. Under transmission electron microscopy, the cells are rod-shaped with flagella around them, about 2.5-5.9μm long and 0.9-1.5μm wide. Figure 1 ), cultured under anaerobic conditions at 37°C for 3-7 days, the colony diameter is 1-5 mm, off-white, smooth and moist, with irregular edges and no water-soluble pigment ( Figure 2 ).

[0027] Molecular identification method: Full 16S rRNA sequence analysis identified and classified the strain as Roseburia zhanii, also known as Zhanluoshiye in Chinese. The strain was named LC-95. The strain was deposited with the General Microbiology Center of the China General Culture Collection Administration (CGMCC) on January 29, 2024, with the deposit number CGMCC No. 29797. The deposit address is No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing. The patented strain deposit number is CGMCC No. 29797. The 16S rRNA sequence homology with Roseburia zhanii is 72.35%, and the sequence is shown in SEQ ID NO. 1 in the sequence listing.

[0028] 2. Cultivation and preservation methods:

[0029] Roseburia zhanii LC-95 was activated once in modified GAM broth containing 10% clarified rumen fluid, and then inoculated into modified GAM liquid medium with the same composition at a 0.1%-10% inoculum size and cultured anaerobically at 37°C for 2-7 days.

[0030] Long-term storage method for Roseburia zhanii LC-95: After centrifugation at 10,000 rpm at room temperature for 2 minutes, collect the cells, wash once with PBS buffer solution, resuspend in a solution containing 15%-25% skim milk powder at a ratio of 5:1, freeze-dry and seal for storage, or freeze in liquid nitrogen in a preservation solution containing 15% glycerol and 85% serum and store at -80°C.

[0031] 3. Uses and effects of strains:

[0032] 3.1 Animal Experiment on Relieving Constipation with LC-95 Live Bacteria

[0033] 3.1.1 Materials and Methods

[0034] 1) Modified GAM medium (g / L):

[0035] Casein peptone 10 g, soy peptone 3 g, tryptone 15 g, digested serum 13.5 g, yeast extract 5 g, beef meal 2 g, bovine liver extract 1.2 g, glucose 3 g, soluble starch 0.3 g, L-cysteine ​​hydrochloride 0.5 g, L-arginine 0.5 g, L-tryptophan 0.3 g, sodium bicarbonate 2 g, potassium dihydrogen phosphate 2.5 g, NaCl 3 g, sodium thioglycolate 0.15 g, sodium acetate 2.46 g, hemin 0.01 g, resazurin 0.001 g, clarified rumen fluid 10% (v / v), distilled water to 1 L, pH 7.2 ± 0.1, sterilized at 115°C for 25 minutes by autoclaving before use. For solid media, add an additional 15 g of agar.

[0036] 2) LC-95 live bacteria culture method:

[0037] LC-95 is activated on modified GAM solid medium and incubated anaerobically at 37°C for 1-2 days. A single colony is then transferred to modified GAM liquid medium for seed culture at 37°C for 1-2 days. Subsequently, the seed culture is inoculated into modified GAM liquid medium with the same composition at a 1-5% inoculum level for expansion and incubated anaerobically at 37°C for 1-2 days.

[0038] 3) Animal Experiment Design:

[0039] The experiment was conducted using 8-week-old male C57BL / 6J mice. After acclimation for 1 week, each group of mice was fed a high-fat diet until 28 weeks, referred to as HFD (High Fat Diet) mice. Starting from the 29th week, the live bacteria group (6 mice) was gavaged with 200 μL of 1×10 cells per day. 9CFU of LC-95 live bacteria; the control group (6 mice) were injected with the same dose of PBS buffer solution every day. The experimental period was 32 weeks. The specific animal experimental design is as follows Figure 2 shown.

[0040] Endpoint sampling: A. Plasma samples (anticoagulant blood was centrifuged at 6000 rpm for 10 minutes, and the supernatant was collected); serum samples (non-anticoagulant blood was centrifuged at 3000 rpm for 15 minutes, and the supernatant was collected); B. Liver samples (livers were photographed and weighed, the middle lobe was divided into two halves and fixed with 4% paraformaldehyde, and then stained with H&E; the remaining liver was accurately weighed to 100 mg and quickly frozen in liquid nitrogen in three tubes for relevant indicator testing); C. Brain samples (the whole brain was dissected, the left hemisphere was fixed with 4% paraformaldehyde, and the right hemisphere was quickly frozen in liquid nitrogen); D. Gastrointestinal tissues and contents (the entire gastrointestinal tract was photographed and measured in length; stomach tissue was frozen; the small intestine was divided into three sections: duodenum, jejunum, and ileum, and frozen; the cecum and its contents were frozen separately; the colorectum and its contents were frozen separately); D. Organ samples (the apex of the heart was frozen, and the other sections were fixed; the spleen was frozen; the left kidney was fixed, and the right kidney was frozen; the left back skin block was fixed, and the right back skin block was frozen).

[0041] 4) Index detection:

[0042] Constipation index detection: fecal water content, black stool time of mice, small intestinal propulsion rate

[0043] a) Mouse fecal moisture content: 2-3 pellets of fresh feces were collected from each mouse and placed in a weighed EP tube. The wet weight was measured. The tube was then dried in a 65°C oven to a constant weight. The dry weight of the feces was then measured to calculate the fecal moisture content.

[0044] Fecal moisture content = (fecal wet weight - fecal dry weight) / fecal wet weight * 100%

[0045] b) Black stool excretion time in mice: After fasting for 20 hours but not water, mice were gavaged with 300 μL of semisolid nutrient paste. Time was counted after gavage, and the time of the first black stool excretion, the number of feces excreted by the mice, and the total weight of feces excreted by the mice within 6 hours were recorded. During the experiment, the mice had normal food and free access to water.

[0046] c) Mouse small intestinal propulsion rate test: Before sacrifice, mice were gavaged with 400 μL of semisolid nutrient paste. 15 minutes after sacrifice, the intestinal segments were photographed, and the propulsion distance of the semisolid nutrient paste in the small intestine and the total length of the small intestine were recorded to calculate the small intestinal propulsion rate.

[0047] Small intestine propulsion rate = distance from the pylorus to the front end of the nutrient paste propulsion / total length of the small intestine * 100%

[0048] 3.1.2 Experimental results:

[0049] 1) LC-95 live bacteria agent increases the water content and moistness of mouse feces:

[0050] The results in Table 1 show that the LC-95 live bacteria agent can significantly increase the water content of mouse feces and has a significant effect on increasing the wetness of mouse feces.

[0051] Table 1. Water content of mouse feces (%)

[0052]

[0053] 2) LC-95 live bacteria agent shortens the time it takes for mice to pass their first black stool and increases the number and weight of stools:

[0054] The data results in Table 2 show that LC-95 live bacteria agent can significantly shorten the time of first black stool excretion in mice, increase the number of stool particles and feces weight in mice, and promote intestinal peristalsis in mice.

[0055] Table 2. Time of first black stool excretion in mice (s)

[0056]

[0057] Table 3. Number of fecal pellets in mice in 6 hours

[0058]

[0059] Table 4. 6-hour stool weight of mice (mg)

[0060]

[0061] 3) LC-95 live bacteria agent promotes small intestinal propulsion rate in mice:

[0062] The results in Table 5 show that the LC-95 live bacteria agent can significantly promote the propulsion rate of the small intestine of mice and increase the intestinal peristalsis level of mice.

[0063] Table 5. Mouse small intestinal propulsion rate (%)

[0064]

[0065] 3.2LC-95 live bacteria agent relieves anxiety, improves cognitive memory, and enhances motor coordination.

[0066] 3.2.1 Materials and Methods

[0067] 1) Modified GAM medium (g / L):

[0068] Casein peptone 10 g, soy peptone 3 g, tryptone 15 g, digested serum 13.5 g, yeast extract 5 g, beef meal 2 g, bovine liver extract 1.2 g, glucose 3 g, soluble starch 0.3 g, L-cysteine ​​hydrochloride 0.5 g, L-arginine 0.5 g, L-tryptophan 0.3 g, sodium bicarbonate 2 g, potassium dihydrogen phosphate 2.5 g, NaCl 3 g, sodium thioglycolate 0.15 g, sodium acetate 2.46 g, hemin 0.01 g, resazurin 0.001 g, clarified rumen fluid 10% (v / v), distilled water to 1 L, pH 7.2 ± 0.1, autoclaved at 115°C for 25 min. For solid medium, add an additional 15 g agar.

[0069] 2) LC-95 live bacteria culture method:

[0070] LC-95 is activated on modified GAM solid medium and incubated anaerobically at 37°C for 1-2 days. A single colony is then transferred to modified GAM liquid medium for seed culture at 37°C for 1-2 days. Subsequently, the seed culture is inoculated into modified GAM liquid medium with the same composition at a 1-5% inoculum level for expansion and incubated anaerobically at 37°C for 1-2 days.

[0071] 3) Animal Experiment Design:

[0072] Animal Experiment Design:

[0073] The experiment was conducted using 8-week-old male C57BL / 6J mice. After acclimation for 1 week, each group of mice was fed a high-fat diet until 28 weeks, referred to as HFD (High Fat Diet) mice. Starting from the 29th week, the live bacteria group (6 mice) was gavaged with 200 μL of 1×10 cells per day. 9 CFU of LC-95 live bacteria; the control group (6 mice) were injected with the same dose of PBS buffer solution every day. The experimental period was 32 weeks. The specific animal experimental design is as follows Figure 2 shown.

[0074] Endpoint sampling: A. Plasma samples (anticoagulant blood was centrifuged at 6000 rpm for 10 minutes, and the supernatant was collected); serum samples (non-anticoagulant blood was centrifuged at 3000 rpm for 15 minutes, and the supernatant was collected); B. Liver samples (livers were photographed and weighed, the middle lobe was divided into two halves and fixed with 4% paraformaldehyde, and then stained with H&E; the remaining liver was accurately weighed to 100 mg and quickly frozen in liquid nitrogen in three tubes for relevant indicator testing); C. Brain samples (the whole brain was dissected, the left hemisphere was fixed with 4% paraformaldehyde, and the right hemisphere was quickly frozen in liquid nitrogen); D. Gastrointestinal tissues and contents (the entire gastrointestinal tract was photographed and measured in length; stomach tissue was frozen; the small intestine was divided into three sections: duodenum, jejunum, and ileum, and frozen; the cecum and its contents were frozen separately; the colorectum and its contents were frozen separately); D. Organ samples (the apex of the heart was frozen, and the other sections were fixed; the spleen was frozen; the left kidney was fixed, and the right kidney was frozen; the left back skin block was fixed, and the right back skin block was frozen).

[0075] 4) Behavioral testing indicators:

[0076] a) Elevated plus maze test;

[0077] Rodents tend to move in closed arms due to their dark preferences, but will also move in open arms out of curiosity and exploratory tendencies. When faced with novel stimuli, animals experience both an urge to explore and fear, leading to conflicting behaviors between exploration and avoidance, which can cause anxiety. The elevated plus maze is a classic experiment for behavioral research, particularly anxiety and depression.

[0078] During the test, mice were placed in the central area and allowed to move freely in an undisturbed environment for 5 minutes. The automated analysis system recorded the time spent exploring each area of ​​the open arms, closed arms, and central area, as well as the number of times they entered and exited the open and closed arms.

[0079] b) Y-maze spontaneous alternation test;

[0080] The spontaneous alternation experiment exploits rodents' curiosity about novelty—the tendency to explore areas they haven't visited before—to test their short-term working memory. Cognitively normal mice remember the arms they've visited and tend to enter arms they've visited less frequently. This short-term working memory requires the interaction of multiple different regions, including the hippocampus and prefrontal cortex.

[0081] During the test, mice were placed with their backs facing the central area into one arm of a Y-maze (defined as the starting arm) and allowed to freely explore each arm for 8 minutes. A successful entry into an arm was considered once if the mouse's torso entered that arm. A successful entry into three arms without duplication was considered accurate, and the accuracy rate was calculated. The total number of times a mouse entered each arm of the Y-maze within 8 minutes can be used to assess its activity level. A lower accuracy rate indicates more severe impairment of its short-term working memory.

[0082] Spontaneous alternation rate (%) = total number of alternations in a triplet containing all three arm numbers / possible number of alternations when entering the maze arm (total number of arm entries - 2) × 100%

[0083] c) Y-maze novel arm recognition test;

[0084] The novel arm recognition experiment consists of two phases: training and testing. A camera was placed 1.5 meters above the maze to record the entire process. During the training phase, a removable partition separated a random arm (the novel arm) from the other two arms of the Y-maze. The mouse was placed at the end of one arm (the starting arm) of the Y-maze with its back facing the central area. It was allowed to freely explore the two arms of the Y-maze for 10 minutes. The mouse was then returned to its cage for 2-4 hours (testing was allowed within 24 hours). During the testing phase, the partition was removed, and the mouse was again placed at the end of the starting arm with its back facing the central area. It was allowed to freely explore the three arms of the Y-maze for 5 minutes. A mouse was considered to have entered one arm once if both its trunk and all four limbs entered the arm. The entire process of the mouse's movement within the maze was recorded, with frequency = number of entries into the novel arm divided by the total number of entries into each arm. The shorter the frequency of novel arm entries and the time spent exploring the novel arm, the more severe the impairment of its memory ability.

[0085] d) Open mine identification test;

[0086] The open field test is a method to evaluate the autonomous behavior, exploratory behavior and stress level of experimental animals in a novel environment.

[0087] The animals were placed in the center of the chamber floor, which was equipped with a central area (30 cm x 30 cm) and an edge area. Video recording and timing were performed for 6 minutes. At the end of the experiment, each mouse's total distance crawled, number of entries into the central compartment, duration of stay, and percentage of distance covered were recorded. After each experiment, each mouse was wiped clean with 75% ethanol to minimize olfactory cue recognition.

[0088] e) novel object recognition test;

[0089] Novel object recognition is an experimental method to judge the memory and cognitive abilities of experimental mice. If the mice have a longer ability to explore new things than old things, it means that their cognitive abilities are normal.

[0090] The experiment was divided into an adaptation period, a familiarization period, and a test period. The mice were first acclimated to the cognitive box for 10 minutes, and then two identical objects were placed in the cognitive box. After 5 minutes of familiarization, the mice were taken out. One hour later, one of the objects was replaced with another object of different shape and color as a novel object. The number of times the mice explored the new and old objects, the time and distance they spent around the new and old objects were recorded, and the mice's cognitive status was tested. The recognition index (RI) was calculated as new object / (new object + old object) × 100%.

[0091] f) Fatigue rotarod test;

[0092] Assess the motor coordination and fatigue levels of mice. Each mouse was placed on a rotarod fatigue apparatus at a speed of 20 rpm / min for 3 consecutive days of rotarod fatigue training. For the formal rotarod experiment, mice were placed on the electric rotarod fatigue apparatus at speeds increasing from 1 rpm / min to 20 rpm / min. The rotarod dwell time was recorded from the start of the experiment until the mouse fell off the rotarod. This dwell time was used as an indicator of the mouse's condition.

[0093] 3.2.2 Experimental results:

[0094] 1) LC-95 live bacteria improves anxiety in mice:

[0095] The results in Tables 6-8 show that the LC-95 live bacteria agent can significantly increase the distance mice move in the open arms and the number and time they enter the open arms in the elevated plus maze test, thereby alleviating the anxiety state of mice.

[0096] Table 6. Elevated Cross-Region Length Percentage (%)

[0097]

[0098] Table 7. Elevated Cross-Area Time Percentage (%)

[0099]

[0100] Table 8. Percentage of Elevated Cross-Area Frequency (%)

[0101]

[0102] 2) LC-95 live bacteria agent alleviates short-term memory impairment in mice:

[0103] The data results in Table 9 show that the LC-95 live bacteria agent can significantly increase the spontaneous alternation rate of mice in the Y maze and alleviate the short-term memory impairment of mice.

[0104] Table 9. Y-maze - Spontaneous Alternation Rate Percentage (%)

[0105]

[0106] 3) LC-95 live bacteria agent improves the memory ability of mice:

[0107] The results in Table 10 show that the LC-95 live bacteria agent can significantly increase the frequency of mice entering the new arm in the new arm recognition experiment, improve the memory damage of mice, and alleviate the memory ability of mice.

[0108] Table 10. New Arm Identification - Frequency Percentage (%)

[0109]

[0110] 4) LC-95 live bacteria enhance the exploratory ability of mice:

[0111] The results in Tables 11-13 show that LC-95 live bacteria agent can increase the frequency of mice entering the central area in an open mine, the time they stay in the central area and the distance ratio, improve the exploration ability of mice, and relieve the stress level of mice.

[0112] Table 11. Mine-region distance percentage (%)

[0113]

[0114] Table 12 Mine-residence time (s)

[0115]

[0116] Table 13. Mine-area entry times (%)

[0117]

[0118] 5) LC-95 live bacteria enhances cognitive abilities in mice:

[0119] The results in Tables 14-15 show that LC-95 live bacteria agent can significantly increase the distance mice pass around new objects and the ratio of their stay time in the new object recognition experiment, thereby improving the cognitive ability of mice.

[0120] Table 14. New object recognition - area distance percentage (%)

[0121]

[0122] Table 15. New Object Recognition-Region Time Percentage (%)

[0123]

[0124] 6) LC-95 live bacteria agent improves the motor balance ability of mice:

[0125] The results in Tables 16 and 17 show that the LC-95 live bacteria agent can significantly prolong the distance and residence time of mice on the rotarod in the fatigue rotarod test, and improve the mice's exercise ability and balance ability.

[0126] Table 16. Fatigue Rotating Rod-Drop Time (s)

[0127]

[0128] Table 17. Fatigue Rotating Rod-Distance (m)

[0129]

[0130]

Claims

1. A Roseburia zhanii LC-95, whose deposit number is CGMCC NO.29797.

2. A microbial agent, the active ingredient of which is the Roseburia LC-95 described in claim 1.

3. A composition comprising the Roseburia LC-95 according to claim 1.

4. Use of the Roseburia LC-95 according to claim 1 in the preparation of a medicine or food for relieving constipation.

5. Use of the Roseburia LC-95 according to claim 1 in the preparation of medicines or foods for alleviating anxiety.

6. Use of the Roseburia LC-95 according to claim 1 in the preparation of a medicine or food for alleviating short-term memory impairment.

7. Use of the Roseburia LC-95 according to claim 1 in the preparation of medicines or foods for improving exploratory ability.

8. Use of the Roseburia LC-95 according to claim 1 in the preparation of medicines or foods for improving cognitive and memory functions.

9. Use of the Roseburia LC-95 according to claim 1 in the preparation of a medicine or food for improving athletic ability and balance ability.

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