Enterococcus hirae QT4713, microbial agent and use thereof in alleviating DSS-induced colon inflammation

Abstract

The present invention relates to the field of microbial technology. The present invention provides an Enterococcus hirae QT4713, whose taxonomic name is Enterococcus hirae, which was deposited in the China Center for Type Culture Collection on December 11, 2023, and the deposit address is Wuhan University, Wuhan, China, and the deposit number is CCTCC NO: M20232523. The present invention also provides a microbial agent comprising the Enterococcus hirae QT4713 strain and its application, which has a strong prebiotic function unique to plateau lactic acid bacteria, as well as strong anti-inflammatory ability, gastrointestinal fluid tolerance and safety, and can better alleviate DSS-induced colon inflammation in mice by regulating intestinal flora.

Description

Technical Field

[0001] The present invention relates to the technical field of microorganisms, and in particular to Enterococcus hirae QT4713, a microbial agent and application thereof in alleviating DSS-induced colon inflammation. Background Art

[0002] Inflammatory bowel disease (IBD) is a disease characterized by chronic recurrent inflammation of the small intestine and colon, mainly including ulcerative colitis (UC) and Crohn's disease (CD). Ulcerative colitis (UC) is one of the main forms of IBD and an important factor in inducing colon cancer. It is characterized by colon inflammation and destruction of colon epithelial structure, and its incidence is gradually increasing worldwide. UC is one of the main forms of ulcerative colitis and an important factor in inducing colon cancer. It is characterized by colon inflammation and destruction of colon epithelial structure. It is caused by a combination of factors including immune dysregulation, intestinal epithelial barrier destruction and genetic susceptibility, but the specific pathogenic mechanism is still unclear. Its clinical treatment methods include biological agents, salicylic acid and immunomodulators, but the efficacy is unstable and some have obvious side effects.

[0003] In recent years, many studies have shown that lactic acid bacteria can promote intestinal health through immune regulation, enhancing intestinal barrier function and regulating intestinal homeostasis, showing great potential in the treatment and relief of UC.

[0004] The unique geographical environment and rich natural resources of the Qinghai-Tibet Plateau contain a large number of characteristic bacterial resources. This special geographical environment gives them diversity in biological and genetic characteristics. At the same time, taking into account the particularity and preciousness of probiotic bacterial resources in high-altitude and cold areas, it is necessary to protect and develop and utilize these high-quality bacterial resources, establish a high-quality bacterial resource library, and screen out bacterial resources with obvious regional characteristics and unique biological activity, which is very necessary for studying the anti-inflammatory properties of plateau bacterial species. Summary of the invention

[0005] The purpose of the present invention is to provide an Enterococcus hirae QT4713, which has the powerful prebiotic function unique to plateau lactic acid bacteria, as well as strong anti-inflammatory ability, gastrointestinal fluid tolerance and safety, and can better alleviate DSS-induced colon inflammation in mice by regulating intestinal flora.

[0006] In order to achieve the above-mentioned object of the invention, the present invention provides the following technical solutions:

[0007] The present invention provides an Enterococcus hirae QT4713, whose taxonomic name is Enterococcus hirae, which was deposited in the China Center for Type Culture Collection on December 11, 2023, with the deposit address being Wuhan University, Wuhan, China, and the deposit number being CCTCC NO: M 20232523.

[0008] The present invention also provides a microbial agent, comprising the above-mentioned Enterococcus hirae QT4713.

[0009] Preferably, the effective concentration of Enterococcus hirae QT4713 in the microbial agent is (0.5-1.5)×10 8 cfu / mL.

[0010] The present invention also provides the use of the Enterococcus hirae QT4713 or the microbial agent in preparing a medicine for relieving colon inflammation.

[0011] Preferably, the colon inflammation is induced by DSS.

[0012] By adopting the above technical solution, the present invention has the following beneficial effects:

[0013] The Enterococcus hirae QT4713 described in the technical solution of the present invention is screened and obtained from the Qinghai-Tibet Plateau, and has a strong prebiotic function unique to plateau lactic acid bacteria, and has strong anti-inflammatory ability, gastrointestinal fluid tolerance and safety. The Enterococcus hirae QT4713 can promote intestinal health by immunomodulation, enhancing intestinal barrier function and regulating intestinal homeostasis; it can also regulate intestinal flora and maintain the homeostasis of intestinal microecology, thereby alleviating DSS-induced colon inflammation.

[0014] The embodiment of the present invention shows that E.hirae QT4713 can reduce the DAI score of colitis mice, reduce colon tissue inflammation and goblet cell loss and crypt structure destruction, reduce colon epithelial cell apoptosis, enhance the expression of MUC-2, occludin, and VEGF5 in colon epithelium, thereby reducing intestinal damage in colitis mice and protecting their intestinal barrier; it can also reduce systemic oxidative stress in colitis mice and reduce the expression of inflammatory factors in colon tissue. The abundance of potentially harmful bacteria (Escherichia and Alistipes) in the intestines of colitis mice can be reduced, the abundance of beneficial bacteria (Lactobacillus, Akkermansia, and Bifidobacterium) can be increased, the structure of intestinal microorganisms in colitis mice can be adjusted, and the content of its metabolite SCFAs can be increased to slow down the level of intestinal inflammation and promote intestinal barrier function. BRIEF DESCRIPTION OF THE DRAWINGS

[0015] Figure 1This is the phylogenetic tree of strain E. hirae QT4713;

[0016] Figure 2 is the strain morphology of strain E. hirae QT4713;

[0017] Figure 3 The DAI scores of mice in different groups;

[0018] Figure 4 The colon length of mice in different groups ( Figure 4 A in the figure is the colon of mice in each group. Figure 4 B in the figure is the statistical result of colon length);

[0019] Figure 5 HE and PAS-AB staining and histological scores of colon tissues of mice in different groups ( Figure 5 A in the figure shows HE and PAS-AB staining. Figure 5 B in the figure is histological score);

[0020] Figure 6 The contents of T-AOC, MDA, GSH-Px in serum and MPO in colon tissue of mice in different groups ( Figure 6 A is T-AOC content, B is MDA content, C is GSH-Px content, and D is MPO content);

[0021] Figure 7 The contents of IL-1β, IL-6, IL-8 and TNF-α in colon tissue of mice in different groups ( Figure 7 A is the IL-1β content, B is the IL-6 content, C is the IL-8 content, and D is the TNF-α content);

[0022] Figure 8 The results of TUNEL immunofluorescence sections and apoptosis rates of colon epithelial cells in different groups of mice ( Figure 8 A is the immunofluorescence section result, and B is the apoptosis rate);

[0023] Fig. 9 The immunofluorescence expression results of MUC-2, occludin, and VEGF5 in different groups of mice;

[0024] Fig.10 is the content of SCFAs in the colon contents of mice in different groups ( Fig.10A is the content of acetic acid, B is the content of propionic acid, C is the content of butyric acid, D is the content of valeric acid, E is the content of isobutyric acid, and F is the content of isovaleric acid).

[0025] Fig.11 The Alpha diversity of intestinal microorganisms in different groups of mice ( Fig.11 A is the Shannon index and B is the Simpson index);

[0026] Fig.12 Beta diversity of intestinal microorganisms and ANOSIM test results of mice in different groups ( Fig.12 A in the figure is the microbial Beta diversity result, and B is the ANOSIM test result);

[0027] Fig.13 is the relative abundance of intestinal microorganisms in different groups of mice ( Fig.13 A is the relative abundance of Akkermansi, B is the relative abundance of Bifidobacterium, C is the relative abundance of Lactobacillus, D is the relative abundance of Mucispirillum, E is the relative abundance of Enterococcus, F is the relative abundance of Escherichia, and G is the relative abundance of Alistipes).

[0028] Biological Deposit Description

[0029] The present invention relates to Enterococcus hirae QT4713, whose taxonomic name is Enterococcus hirae, which was deposited in the China Center for Type Culture Collection on December 11, 2023, with the deposit address being Wuhan University, Wuhan, China, and the deposit number being CCTCCNO: M 20232523. DETAILED DESCRIPTION

[0030] The present invention provides an Enterococcus hirae QT4713, whose taxonomic name is Enterococcus hirae, which was deposited in the China Center for Type Culture Collection on December 11, 2023, with a deposit address of Wuhan University, Wuhan, China, and a deposit number of CCTCC NO: M 20232523. The Enterococcus hirae QT4713 described in the present invention belongs to lactic acid bacteria.

[0031] The present invention also provides a microbial agent, comprising the Enterococcus hirae QT4713.

[0032] In the present invention, the effective concentration of Enterococcus hirae QT4713 in the microbial agent is preferably (0.5-1.5)×10 8 cfu / mL, more preferably (0.8-1.2)×l0 8 cfu / mL, and more preferably 1×10 8 cfu / mL.

[0033] The present invention also provides the use of the Enterococcus hirae QT4713 or the microbial agent in preparing a medicine for relieving colon inflammation.

[0034] In the present invention, the colon inflammation is induced by DSS.

[0035] The technical solutions provided by the present invention are described in detail below in conjunction with the embodiments, but they should not be construed as limiting the protection scope of the present invention.

[0036] Example 1

[0037] A microbial agent, the preparation process is:

[0038] The QT4713 bacterial suspension stored at -80°C was inoculated into the sterilized MRS agar medium and cultured at 37°C for 18 h. The third generation culture was collected after three consecutive subcultures. The third generation culture was then centrifuged at 13,000 rpm at 4°C for 10 min. The cells were collected and washed with PBS three times and resuspended in PBS to adjust the bacterial concentration to 1×10 8 CFU / mL is obtained.

[0039] Example 2 Isolation and Identification of Lactic Acid Bacteria QT4713

[0040] (I) Acquisition of Enterococcus hirae QT4713

[0041] Single colonies were selected from the strains (strain 133, strain 120, and strain QT4713) screened and activated in the Qinghai-Tibet Plateau, and single colonies were selected from the control strain Lactobacillus rhamnosus GG (LGG) (purchased from Ningbo Biotechnology Co., Ltd.), and placed together at 37°C for 18 hours. After three consecutive passages, the bacteria were collected by centrifugation at 13,000 rpm for 10 minutes. After washing and resuspending three times with PBS, the bacteria were collected again, and then resuspended with DMEM high-glucose medium, and the number of bacteria was adjusted according to the infection multiplicity (MOI) of 500:1. The adjusted bacterial suspension was then placed in a 56°C metal bath for 30 minutes to heat-inactivate the bacteria for use.

[0042] RAW264.7 cells (purchased from Wuhan Pronocell Biotechnology Co., Ltd.) were passed to the 3rd or 4th generation, and the cells were collected by gentle pipetting and the cell concentration was adjusted to 2.5×10 5 cell / mL, and evenly inoculated into 24-well plates, cultured in a 37°C, 5% CO2 incubator until fusion, discarded the supernatant, and washed 3 times with PBS. Then divided into 6 groups, the control group only added DMEM high-glucose medium, the LPS group added LPS with a final concentration of 1μg / mL, and the four strain groups added 500μL / well of the prepared corresponding bacterial suspension, and added LPS with a final concentration of 1μg / mL. The cells of each group were cultured in a 37°C, 5% CO2 incubator for 16h, the supernatant was collected, and the NO content in the supernatant was determined. The results are shown in Table 1.

[0043] The strain 133, strain 120 and strain QT4713 described in the present invention are strains screened by the inventors from the Qinghai-Tibet Plateau, and the corresponding strain names are named by the inventors themselves.

[0044] Table 1 NO content in the supernatant of different groups

[0045] Group NO content (μmol / L) Control Group 3.70±0.30 LPS group 14.23±0.55 Control strain LGG 7.72±0.58 strain 133 3.60±0.10 strain 120 2.89±0.24 Strain QT4713 2.81±0.19

[0046] NO is a small molecule produced by endothelial cells, inflammatory cells and epithelial cells, and has a variety of physiological and pathological effects. NO is an important signal factor in the mechanism of inflammation. The NO synthase family regulates its production and plays a regulatory role in the generation of pro-inflammatory mediators. NO itself can react with superoxide free radicals to produce peroxynitrite ions, which in turn leads to inflammatory reactions. In diseases related to inflammation, it can be observed that activated macrophages produce excessive NO. Therefore, inhibiting the production of NO is one of the important ways to treat inflammatory diseases. Therefore, by comparing the inhibitory effects of different strains on NO, the strain with the strongest inhibitory effect, strain QT4713, was screened out.

[0047] (II) Identification of Enterococcus hirae QT4713

[0048] The colony and bacterial morphology of the strain QT4713 obtained above were observed, and physiological and biochemical identification was performed. The results showed that the colony was milky white, round, convex, with regular edges, smooth and moist surface, easy to pick, Gram-positive bacteria, spherical under the microscope, without spores. Catalase negative, oxidase negative.

[0049] The DNA of strain QT4713 was extracted using a Gram-negative bacteria genomic DNA extraction kit (purchased from Beijing Solebow Technology Co., Ltd.) and sent to BioMed to clone the 16S rRNA sequence of QT4713 and sequence the 16S rRNA sequence. The sequencing results were compared with Genbank by BLAST to determine the phylogenetic evolutionary position of QT4713. The phylogenetic tree is shown in Figure 1 The strain QT4713 was further identified as Enterococcus hirae. The morphology of the strain E. hirae QT4713 is shown in Figure 2 shown.

[0050] Test Example 1

[0051] (I) Construction of mouse colitis model

[0052] A mouse colitis model was established to evaluate the effects of E. hirae QT4713 on colonic and systemic oxidative stress responses, colon tissue inflammation and intestinal microorganisms in mice.

[0053] The selected mice were female mice, of the C57BL / 6J strain, 8-10 weeks old, weighing 22±2g, purchased from the Medical Experiment Center of Lanzhou University. After 7 days of adaptive feeding, the mice were randomly divided into 4 groups (10 / group), namely, the Control group (blank control group, CON), the DSS group (model control group, DSS), the LAB group (healthy mouse LAB intervention group, LAB) and the LAB+DSS group (colitis mouse LAB intervention group, LAB+DSS).

[0054] The experimental period was 14 days. The Control group and the DSS group were given PBS gavage (200 μL / mouse) from 1 to 7 days. From the 8th day, the Control group was given PBS gavage (200 μL / mouse) + distilled water to drink freely, and the DSS group was given PBS gavage (200 μL / mouse) + 3% DSS (w / v) solution to drink freely. The LAB group and the LAB+DSS group were given E.hirae QT4713 gavage (200 μL / mouse) from 1 to 7 days. From the 8th day, the LAB group was given E.hirae QT4713 gavage (200 μL / mouse) + distilled water to drink freely, and the LAB+DSS group was given E.hirae QT4713 gavage (200 μL / mouse) + 3% DSS (w / v) solution to drink freely. On the 15th day, the mice in each group were treated for subsequent experiments.

[0055] (II) E. hirae QT4713 reduces the DAI score of colitis mice

[0056] The DAI scores were performed on the mice in the above groups.

[0057] The rating results are as follows Figure 3 As shown in the figure, from the second day of modeling to the sixth day, the DAI score of the DSS model group was the highest, which was significantly higher than that of the CON group, LAB group and LAB+DSS group. During the DSS modeling period, the mice in the DSS model group experienced weight loss, loose stools, and fecal occult blood. As the modeling time prolonged, the weight of the mice further decreased, the feces were unformed, diarrhea occurred, and there was visible blood in the stool. The DAI score reached the highest on the sixth day. However, by oral gavage intervention with E.hirae QT4713, the DAI score of mice in the LAB+DSS group was significantly reduced by the sixth day, indicating that compared with the DSS group, the intervention of E.hirae QT4713 significantly reduced the DAI score of colitis model mice, and alleviated the symptoms of weight loss, diarrhea, and blood in the stool caused by colitis.

[0058] (III) E. hirae QT4713 increases colon length in colitis mice

[0059] After treatment, the length of the entire colon from the end of the cecum to the anus was measured and recorded.

[0060] The changes of colon length in each group of mice are as follows Figure 4 As shown in the figure, there was no significant difference in the colon length between the CON group and the LAB group, indicating that oral administration of E. hirae QT4713 alone had no effect on the colon length of mice. However, compared with the CON group, the colon length of the mice in the DSS model group was significantly shortened, and there was obvious colon bleeding and blackening, which was caused by DSS-induced colon inflammation. Compared with the DSS model group, the colon length of the LAB+DSS group increased significantly, and the bleeding and blackening of the colon were also significantly improved.

[0061] (IV) E. hirae QT4713 alleviates DSS-induced colon pathological damage in mice

[0062] Colon pathological sections can effectively reflect the degree of inflammation and pathological changes in the mouse colon. By performing HE staining and AB-PAS staining on the mouse colon tissue, the inflammatory infiltration and goblet cell loss were observed.

[0063] The colon tissue of mice was taken and fixed in 4% paraformaldehyde solution for 24 hours. The colon tissue was dehydrated by soaking in 85% and 95% anhydrous ethanol and xylene in turn. Then it was embedded in paraffin and paraffin sections were made. The prepared paraffin sections were dewaxed and stained with hematoxylin-eosin (HE staining). Hematoxylin staining for 5 minutes, rinsed with running water, blueing for 10 seconds, eosin staining for 5 minutes, and finally dehydrated and sealed. The state of the colon was observed under an optical microscope and images were collected. The prepared paraffin sections were dewaxed to water and stained according to the instructions of the AB-PAS staining solution set. After staining, the sections were dehydrated and sealed. The loss of goblet cells in the colon tissue was observed under a microscope and images were collected.

[0064] like Figure 5 As shown in the figure, the intestinal walls of mice in the CON and LAB groups were intact, the glands were densely distributed and arranged in an orderly manner, the crypt structure was intact, no inflammatory cell infiltration was observed, and even more goblet cells were found in the LAB group. However, compared with the CON group, the colon tissue sections of mice in the DSS group showed obvious histopathological characteristics of ulcerative colitis, such as glandular atrophy, extensive erosion of the mucosal surface, ulceration, goblet cell loss, inflammatory cell infiltration, and crypt loss. However, the preventive intervention of E. hirae QT4713 alleviated the colonic lesions caused by DSS. In the LAB+DSS group, the epithelial structure was basically intact, the glands were densely distributed, and the loss of goblet cells and crypts and the infiltration of inflammatory cells were significantly alleviated.

[0065] (V) E. hirae QT4713 alleviates oxidative stress in colitis mice

[0066] GSH-Px, MDA, T-AOC and MPO content detection kits (all purchased from Beijing Solebow Technology Co., Ltd.) were used to measure the levels of GSH-Px, MDA, T-AOC in the serum of the above-mentioned groups of mice and MPO in the colon tissue, so as to analyze the relief of oxidative stress in colitis mice by E. hirae QT4713.

[0067] according to Figure 6 It can be seen that compared with the CON group, the levels of MDA in the serum and MPO in the colon tissue of the mice in the DSS group increased, and compared with the DSS group, the intervention of E.hirae QT4713 reduced the levels of MDA in the serum and MPO in the colon tissue of the mice in the LAB+DSS group; compared with the CON group, the activities of GSH-Px and T-AOC in the serum of the mice in the DSS group were significantly reduced, and compared with the DSS group, the intervention of E.hirae QT4713 significantly enhanced the activities of GSH-Px and T-AOC in the serum of the mice in the LAB+DSS group. This shows that the intervention of E.hirae QT4713 can reduce the oxidative stress of the whole body and colon of mice with colitis.

[0068] (VI) E. hirae QT4713 reduces the expression of inflammatory factors in the colon group of colitis mice

[0069] Colon tissue homogenates were prepared according to the instructions of ELISA kits (purchased from Shanghai ELISA Biotechnology Co., Ltd.), and the levels of IL-6, IL-8, IL-1β and TNF-α in colon tissues were detected. The normality and statistical significance of the data were tested by Welch's variance analysis using IBM SPSS Statistics.

[0070] The results are as follows Figure 7 As shown in the results, DSS induction increased the levels of IL-6, IL-8, IL-1β and TNF-α in the colon tissue of the mice in the model group, but the intervention of E. hirae QT4713 significantly decreased the levels of IL-6, IL-8, IL-1β and TNF-α in the LAB+DSS group, especially the content of IL-1β.

[0071] (VII) E. hirae QT4713 reduces apoptosis of colonic epithelial cells in mice with colitis

[0072] The in situ terminal transferase labeling (TUNEL) method was used to detect apoptotic cells in the colon in situ. The prepared paraffin sections were dewaxed to water, incubated with proteinase K at 37°C for 22 minutes, and incubated at room temperature for 5 minutes with a membrane permeabilization solution (0.1% Triton). After equilibration at room temperature, TUNEL staining was performed according to the instructions of the TUNEL assay kit (purchased from Wuhan Saiweier Biotechnology Co., Ltd.). After staining, 4',6-diamidino-2-phenylindole (DAPI) was added and incubated for 10 minutes to counterstain the cell nucleus, and the sections were sealed with an anti-fluorescence quencher. The sections were observed under a fluorescence microscope and images were collected.

[0073] The staining results are as follows Figure 8 As shown. Grayscale analysis of the fluorescence images was performed to obtain the apoptosis rate of colon cells in each group of mice. As can be seen from the figure, compared with the CON group, the apoptosis rate of colon cells in the DSS model group was significantly increased; however, compared with the DSS model group, the apoptosis rate in the LAB+DSS group was significantly decreased due to the protective intervention of E.hirae QT4713; compared with the CON group, the apoptosis rate of colon cells in the LAB group was also reduced.

[0074] (VIII) E. hirae QT4713 promotes the expression of MUC-2, occludin and VEGF5 in the colon tissue of colitis mice

[0075] The prepared paraffin sections were dewaxed to water, antigen repaired, circled with a brush, and then blocked with hydrogen peroxide. Incubated with BAS for 30 minutes, added with primary antibody and incubated overnight at 4°C, washed 3 times with PBS, added with secondary antibody of the corresponding species, and incubated at room temperature for 50 minutes. Washed, added with DAPI and incubated for 10 minutes to counterstain the cell nucleus, and sealed with anti-fluorescence quencher. The sections were observed under a fluorescence microscope and images were collected to determine the expression of tight junction protein mucin 2 (MUC-2), occludin, and VEGF5.

[0076] Immunofluorescence was used to determine the expression of MUC-2, Occludin, and VEGF5 in the colon epithelial tissues of each group of mice. Fig. 9 As shown, the expression of MUC-2 in the epithelial tissue of the DSS model group was significantly lower than that of the CON group, which may be related to the loss of goblet cells in the colon epithelium of the DSS group. However, compared with the DSS model group, the expression of MUC-2 in the LAB+DSS group was significantly increased, and the expression of MUC-2 in the LAB group was also significantly higher than that in the CON group. The changing trend of the expression of the tight junction protein occludin in various mouse epithelial tissues was consistent with that of MUC-2. Compared with the CON group, the treatment with DSS caused its expression to decrease. However, the protective intervention of E. hirae QT4713 significantly increased the expression of occludin in the colon epithelial tissue of mice in the LAB+DSS group compared with the DSS group. In addition, according to Fig. 9 It can also be seen that compared with the CON group, the expression level of VEGF5 in the colon epithelial tissue of the DSS model group was significantly decreased. However, compared with the DSS model group, the LAB+DSS group reversed the decrease in VEGF5 expression caused by DSS due to the intervention of E. hirae QT4713, making the expression level of VEGF5 in the colon epithelium of this group of mice significantly higher than that of the DSS model group.

[0077] (IX) E. hirae QT4713 increases the content of SCFAs in the intestinal contents of colitis mice

[0078] After thawing the collected mouse colon contents on ice, 20 mg was weighed and placed in a 2 mL centrifuge tube. A small steel ball was placed, and 1 mL of 0.5 M phosphoric acid solution (v / v) was added. The mixture was ground at 30 Hz for 60 s, vortexed at 2500 rpm / min for 10 min, sonicated at 4°C for 5 min, and then centrifuged at 12000 rpm / min for 10 min at 4°C. 100 μL of the supernatant was transferred to a 1.5 mL centrifuge tube, mixed with 500 μL of MTBE extractant containing an internal standard, vortexed at 2500 rpm / min for 3 min, sonicated at 4°C for 5 min, and then centrifuged at 12000 rpm / min for 10 min at 4°C. 200 μL of the supernatant was transferred to the inner liner of the injection bottle and analyzed by GC-MS / MS. The main conditions of GC-MS / MS include: chromatographic column DB-FFAP, carrier helium, column flow rate 1.2mL / min, front injection temperature 250℃, injection volume 1μL. The temperature program is set as follows: starting temperature 100℃ for 5min, heating at a rate of 10℃ / min to 150℃ and maintaining for 3min. Ion source temperature: 230℃, quadrupole temperature: 150℃.

[0079] SCFAs are the end products of dietary fiber fermentation by beneficial intestinal bacteria. They can protect the intestinal mucosa, maintain the integrity of the intestinal barrier, promote the absorption of nutrients, regulate immune responses, and inhibit intestinal inflammation. Changes in intestinal microflora can affect the level of SCFAs in the intestine. The levels of SCFAs in the intestinal contents of various mice were measured, and the results are shown in Fig.10 . As can be seen from the figure, the contents of acetic acid, propionic acid, butyric acid, valeric acid, isobutyric acid, and isovaleric acid in the DSS model group were all lower than those in the CON group. However, the preventive administration of E.hirae QT4713 significantly increased the contents of propionic acid and butyric acid in the intestinal contents of the mice in the LAB+DSS group compared with the DSS model group. The contents of acetic acid, valeric acid, isobutyric acid, and isovaleric acid also increased significantly compared with the DSS model group, but there was no statistical significance. Compared with the CON group mice that did not receive intervention, the normal mice significantly increased the content of valeric acid in their intestinal contents after receiving the intervention of E.hirae QT4713, and the content of butyric acid also increased significantly, but there was no statistical significance. This shows that the intervention of E.hirae QT4713 increased the content of SCFAs in the intestines of mice.

[0080] (X) E. hirae QT4713 regulates intestinal flora disturbance in colitis mice

[0081] The colon contents of each group of mice were sequenced by 16S rRNA, and the sequencing results were analyzed.

[0082] DSS induction can lead to colon congestion and edema, intestinal wall thickening and ulcer formation, which in turn leads to an imbalance of intestinal flora. Fig.11 It can be seen that compared with the DSS model control group, the Shannon index and Simpson index of the LAB+DSS group increased significantly, indicating that the Alpha diversity of microorganisms in this group increased significantly and the richness of intestinal microorganisms changed significantly.

[0083] Depend on Fig.12 It can be seen that the distribution of intestinal microbial communities in the DSS-induced model group mice and normal mice is significantly different. After the preventive intervention of E. hirae QT4713, the intestinal microbial communities of the colitis model mice are also significantly separated compared with the DSS model group. The ANOSIM test shows that the differences between the groups are greater than the differences within the groups, indicating that there are significant differences between different groups.

[0084] Depend on Fig.13 It can be seen that compared with the CON group, the abundance of Akkermansi in the intestinal microorganisms of the DSS model mice was significantly reduced; Mucispirillum, Escherichia, Alistipes and other genera were significantly enriched, indicating that the induction of DSS increased the abundance of conditional pathogens in the intestinal microorganisms of mice; compared with the CON group, the intervention of E.hirae QT4713 significantly enriched Bifidobacterium and Lactobacillus in the healthy mice of the LAB group (p<0.05), but the abundance of Akkermansi bacteria was reduced (p<0.05). However, compared with the DSS group, after the preventive intervention of E.hirae QT4713 in the colitis model mice, Akkermansi, Bifidobacterium, Lactobacillus, and Enterococcus were significantly enriched in the intestinal microorganisms of the mice, and the abundance of Escherichia, Alistipes and Mucispirillum in the intestinal microorganisms of the colitis mice was also reduced.

[0085] From the above embodiments and test examples, it can be seen that the lactic acid bacteria E.hirae QT4713 provided by the present invention can reduce the DAI score of colitis mice, reduce colon tissue inflammation and goblet cell loss and crypt structure destruction, reduce colon epithelial cell apoptosis, and enhance the expression of MUC-2, occludin, and VEGF5 in the colon epithelium, thereby reducing the intestinal damage of colitis mice and protecting their intestinal barrier; it can also reduce the systemic oxidative stress of colitis mice and reduce the expression of inflammatory factors in colon tissue. E.hirae QT4713 reduces the abundance of potentially harmful bacteria (Escherichia, Alistipe) in the intestines of colitis mice, increases the abundance of beneficial bacteria (Lactobacillus, Akkermansia, Bifidobacterium), regulates the structure of intestinal microorganisms in colitis mice, and increases the content of its metabolite SCFAs to slow down the level of intestinal inflammation and promote intestinal barrier function.

[0086] The above is only a preferred embodiment of the present invention. It should be pointed out that for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principle of the present invention. These improvements and modifications should also be regarded as the scope of protection of the present invention.

Claims

1. A strain of Enterococcus hirae Enterococcus hirae )QT4713, characterized in that The taxonomic name is Enterococcus hirae , deposited in the China Center for Type Culture Collection on December 11, 2023, with the deposit address being Wuhan University, Wuhan, China, and the deposit number being CCTCC NO: M 20232523.

2. A microbial agent, characterized in that: It comprises the Enterococcus hirae QT4713 described in claim 1.

3. The microbial agent according to claim 2, characterized in that: The effective concentration of lactic acid bacteria QT4713 in the microbial agent is (0.5-1.5)×10 8 cfu / mL.

4. Use of the Enterococcus hirae QT4713 according to claim 1 or the microbial agent according to claim 2 in the preparation of a medicament for alleviating colon inflammation, wherein the colon inflammation is induced by DSS.

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