Streptococcus non-lactolyticus as well as application and microbial agent thereof

Abstract

The invention relates to the technical field of microorganisms, and discloses streptococcus alactolyticus, application of the streptococcus alactolyticus and a microbial agent, the strain name of the streptococcus alactolyticus is streptococcus alactolyticus 0625, the strain of the streptococcus alactolyticus is preserved in the China General Microbiological Culture Collection Center on December 01, 2025, and the preservation number is CGMCC No.36838. The streptococcus alactolyticus has the advantages that the streptococcus alactolyticus can be used for preparing the streptococcus alactolyticus; experiments prove that the streptococcus alactolyticus 0625 provided by the invention can be used for effectively improving the intestinal inflammation, and a solid foundation is laid for the development of related micro-ecological treatment products.

Description

Technical Field

[0001] This invention relates to the field of microbial technology, specifically to a non-lactolytic streptococcus, its application, and microbial agents. Background Technology

[0002] Gut health is crucial for maintaining overall physiological balance. Inflammatory bowel diseases, such as inflammatory bowel disease (IBD), are experiencing a continuous rise in incidence worldwide. These diseases are protracted, prone to relapse, and severely impact patients' quality of life. Current clinical treatment strategies primarily rely on anti-inflammatory drugs, immunosuppressants, and even biologics. While these methods can control symptoms to some extent, they generally suffer from significant side effects, the potential for drug resistance with long-term use, and high treatment costs. Therefore, developing safe, effective, and novel alternative or adjuvant therapies is of great clinical need and practical significance.

[0003] With the deepening of microbiology research, the function of the gut microbiota, as a "forgotten organ" of the human body, is receiving increasing attention. Utilizing probiotics to regulate the balance of the gut microbiota has been proven to be a potentially effective way to intervene in intestinal inflammation. However, current probiotic products on the market and in research mostly focus on a few traditional genera such as Lactobacillus and Bifidobacterium. These general-purpose strains have significant limitations: First, they are highly strain-specific, with vastly different efficacy among strains, making the screening of strains with stable and significant anti-inflammatory activity costly and time-consuming; second, many exogenous probiotics face difficulties in colonization and low survival rates in the host's complex internal environment, especially under inflammatory conditions, leading to a significant reduction in their actual efficacy; third, existing research on the anti-inflammatory mechanisms of probiotics is still not sufficiently in-depth and systematic, limiting their precise application.

[0004] Against this backdrop, isolating native bacterial strains from the gut of healthy hosts has become a more promising technical approach. These native strains, through a long process of co-evolution, have developed natural adaptability to the gut environment, making them easier to colonize and function in vivo. However, efficiently screening specific strains with both excellent gut adaptability and clear, potent anti-inflammatory functions from the vast array of gut microbiota, and validating their efficacy using reliable in vivo models, remains a major technical bottleneck for those skilled in the art.

[0005] Therefore, there is an urgent need in this field for a novel microbial resource that has a clear source, well-defined function, and has been rigorously scientifically verified to effectively improve intestinal inflammation, in order to fill the gaps in the specificity, efficacy, and mechanism of existing probiotic products. Summary of the Invention

[0006] The present invention aims to provide a microbial resource that can improve intestinal inflammation, thereby filling the gaps in the specificity, efficacy and mechanism of existing probiotic products.

[0007] To achieve the above objectives, the first aspect of the present invention provides a non-lactose-degrading Streptococcus (Lactococcus non-lactose-degrading strain). streptococcus alactolyticus ) strain, the non-lactolytic streptococcus ( streptococcus alactolyticus The strain name is streptococcus alactolyticus 0625, this strain was deposited at the China General Microbiological Culture Collection Center on December 1, 2025, with accession number CGMCC No. 36838.

[0008] A second aspect of the present invention provides a microbial inoculant comprising the non-lactolytic streptococcal strain described in the first aspect, and a pharmaceutically or feed-grade acceptable carrier.

[0009] The third aspect of the present invention provides the non-lactolytic streptococcus described in the first aspect ( streptococcus alactolyticus The use of the strain or the microbial agent described in the second aspect in the preparation of compositions for the prevention and / or treatment of intestinal inflammation.

[0010] A fourth aspect of the present invention provides a food, health product, or feed additive comprising the non-lactolytic streptococcus described in the first aspect (… streptococcus alactolyticus ) strains or the microbial agents described in the second aspect.

[0011] Compared with the prior art, the present invention has at least the following beneficial effects: (1) The non-lactolytic streptococcus provided by this invention has a clear function and significant therapeutic effect. This specific non-lactolytic streptococcus strain isolated from the intestines of healthy mammals has been found and verified to have excellent anti-intestinal inflammation function. Through a rigorous DSS-induced mouse enteritis model, it was confirmed that this strain can significantly improve the core symptoms of intestinal inflammation, effectively alleviate the weight loss of experimental animals and inhibit colonic shortening, providing a solid experimental basis for its development for the prevention or treatment of intestinal inflammatory diseases.

[0012] (2) The non-lactolytic streptococcus provided by this invention is an innovative source with excellent adaptability. The strain originates from the intestine of a healthy mammal. As a host native strain, it has a natural affinity and adaptability to the intestinal environment. Compared with traditional exogenous probiotics, it is expected to achieve better colonization and survival in the host intestine, thereby exerting its beneficial functions of regulating immunity and alleviating inflammation more persistently and stably, solving the problems of difficult colonization and unstable effects of existing probiotics.

[0013] (3) The non-lactolytic streptococcus provided by this invention has broad application prospects and high safety. This strain provides a novel core strain resource for developing new microecological preparations for maintaining intestinal health and intervening in intestinal inflammation. It can not only be used as an active ingredient in the preparation of pharmaceuticals, but also be widely used in functional foods, health products and special animal feed additives, etc., providing a safe and natural solution with few side effects to meet the urgent needs for intestinal health in different scenarios.

[0014] Information on the preservation of biological materials The strain information preserved in this invention is: a non-lactolytic streptococcus ( streptococcus alactolyticus ) strain, its name streptococcus alactolyticus 0625, categorized and named as follows: streptococcus alactolyticus The strain has the accession number CGMCC No. 36838 and is deposited at the China General Microbiological Culture Collection Center, located at No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing, on December 1, 2025. Attached Figure Description

[0015] Figure 1 This is a comparison chart of the body weight of mice on day 21 in Test Example 1 of this invention; Figure 2 These are comparative photographs of the colon length of mice on day 21 in Test Example 1 of this invention; Figure 3 This is a comparison image of the colon length of mice on day 21 in Test Example 1 of this invention. Detailed Implementation

[0016] The endpoints and any values ​​of the ranges disclosed herein are not limited to the precise ranges or values, and these ranges or values ​​should be understood to include values ​​close to these ranges or values. For numerical ranges, the endpoint values ​​of the various ranges, the endpoint values ​​of the various ranges and individual point values, and individual point values ​​can be combined with each other to obtain one or more new numerical ranges, which should be considered as specifically disclosed herein.

[0017] As previously stated, the first aspect of this invention provides a non-lactose-degrading Streptococcus (Lactococcus non-lactose-degrading strain). streptococcus alactolyticus ) strain, the non-lactolytic streptococcus ( streptococcus alactolyticus The strain name is streptococcus alactolyticus 0625, this strain was deposited at the China General Microbiological Culture Collection Center on December 1, 2025, with accession number CGMCC No. 36838.

[0018] In a preferred embodiment, the non-lactolytic streptococcus ( streptococcus alactolyticusThe strain was isolated from the feces of C57BL / 6 mice.

[0019] Preferably, the non-lactolytic streptococcus ( streptococcus alactolyticus This strain has the function of improving intestinal inflammation.

[0020] As previously stated, a second aspect of the present invention provides a microbial agent comprising the non-lactolytic streptococcal strain described in the first aspect, and a pharmaceutically or feed-grade acceptable carrier.

[0021] Preferably, the non-lactolytic streptococci in the bacterial agent ( streptococcus alactolyticus The viable count of the strain was 1×10⁻⁶. 8 CFU / mL to 1×10 10 CFU / mL.

[0022] As previously stated, the third aspect of this invention provides the non-lactolytic streptococcus described in the first aspect ( streptococcus alactolyticus The use of the strain or the microbial agent described in the second aspect in the preparation of compositions for the prevention and / or treatment of intestinal inflammation.

[0023] Preferably, the intestinal inflammation is intestinal inflammation induced by sodium dextran sulfate.

[0024] As previously stated, a fourth aspect of the present invention provides a food, health product, or feed additive comprising the non-lactose-degrading Streptococcus as described in the first aspect (… streptococcus alactolyticus ) strains or the microbial agents described in the second aspect.

[0025] The present invention will be described in detail below through examples. Unless otherwise specified, the raw materials used are all commercially available products.

[0026] Preparation of MRS solid medium: Using (CHINOOK) MRS solid medium, weigh 64.25g of it, add 1000ml of distilled water, heat to boiling, stir until completely dissolved, dispense into containers, sterilize at 121℃ for 15min, and then cool in a laminar flow hood onto plates for later use.

[0027] Example 1 This embodiment is used to illustrate the non-lactolytic streptococcus provided by the present invention. streptococcus alactolyticus The strain was isolated and identified according to the following steps: (1) Take 0.1g of feces from healthy C57BL / 6 mice, dissolve and mix with 5mL of sterile PBS (phosphate buffer solution), and serially dilute to 10. -7 100 μL gradient was drawn at 10 -2 -10 -7The diluted solution was spread onto MRS solid medium plates and incubated in a 5 vol% CO2, 37°C incubator for 48 h. Select plates with appropriate dilution gradients, pick single colonies of different morphologies and streak them onto new MRS solid medium plates, incubate at 37°C for 24 h, and purify the strains. Repeat the purification process 3-4 times until pure colonies are obtained, and then perform subsequent identification. (2) DNA of the strains obtained from the single colonies in step (1) was extracted and isolated. The bacterial 16S rDNA gene was sequenced using the SAnger sequencing method. The universal primers were 27F-AGAGTTTGATCCTGGCTCAG and 1492R-GGTTACCTTGTTACGACTT. Colony PCR was performed, and the amplified products were sent to the company for sequencing. Finally, it was determined that the four non-lactose-degrading streptococcal strains obtained in this screening were all the same strain after multiple alignment (forward sequence). The strains were classified and named as follows. streptococcus alactolyticus One of the strains was named streptococcus alactolyticus 0625, sent to the China General Microbiological Culture Collection Center for preservation.

[0028] streptococcus alactolyticus The 16S rDNA sequence of 0625 is (SEQ ID NO.1):

[0029] Test Example 1 The following animal experiments were used to verify the effect of this strain in improving intestinal inflammation. Non-lactolytic streptococci ( streptococcus alactolyticus Tests on the ability of strains to improve intestinal inflammation: S1. Thirty male c57 mice with similar weight (15±0.5g) and similar health status were selected and divided into three groups: control group (CON), DSS (sodium dextran sulfate) group (DSS), and sodium dextran sulfate and non-lactose streptococcus group (DSS+SA), with 10 mice in each group.

[0030] S2. At 9:00 a.m. every day, the CON group was given saline by gavage for 21 consecutive days, during which time they drank water normally. Every morning at 9:00, the DSS group was given saline by gavage for 21 consecutive days. They were allowed to drink water normally from day 1 to day 13, and from day 14 to day 21, they were fed with a 2.5 wt% sodium dextran sulfate solution instead of water.

[0031] Non-lactose-degrading Streptococcus (NLD) was administered to the DSS+SA group at 9:00 AM daily. streptococcus alactolyticus The bacterial culture of the strain was centrifuged, diluted with physiological saline, and then administered by gavage at a volume of 0.2 mL per animal, with a bacterial concentration of 1 × 10⁻⁶. 9 CFU / mL, administered by gavage for 21 consecutive days, with normal drinking water from day 1 to day 13, and 2.5wt% sodium dextran sulfate solution used instead of water for feeding from day 14 to day 21.

[0032] The rodent enclosure is managed using a 12-hour light and 12-hour dark cycle, with suitable temperature and humidity, and free access to water and food.

[0033] S3. Observe the health status of the mice daily and record their weight from day 14 to day 21.

[0034] Figure 1 The body weight of mice on day 21 is shown. It is evident that, compared to the control group, the body weight of mice in the DSS group treated with DSS was significantly lower (P < 0.01); while the body weight of mice in the DSS+SA group was significantly higher than that of the DSS group (P < 0.01). This indicates that the DSS model was successfully established and reduced mouse body weight, and that gavage with *Streptococcus non-Lactosomiasis* significantly improved the body weight loss in mice caused by DSS enteritis.

[0035] Figure 2 and Figure 3The figure shows a comparison of colon length in mice on day 21. Compared with the control group, the colon length in the DSS group was significantly shortened (P < 0.001); however, this phenomenon was significantly reversed in the DSS+SA group compared with the DSS group (P < 0.05). This indicates that the non-lactose-degrading streptococcus provided by this invention can significantly improve the colon shortening in mice caused by DSS enteritis.

[0036] The above results show that, through in vivo animal experiments, this invention has confirmed that the non-lactolytic streptococci isolated from the feces of C57BL / 6 mice (…) streptococcus alactolyticus This strain significantly improved DSS-induced intestinal inflammation, specifically by effectively alleviating weight loss and inhibiting colonic shortening in model animals. This strain provides a safe and effective core bacterial resource for developing novel probiotics for the prevention and / or treatment of intestinal inflammation.

[0037] The preferred embodiments of the present invention have been described in detail above; however, the present invention is not limited thereto. Within the scope of the inventive concept, various simple modifications can be made to the technical solutions of the present invention, including combinations of various technical features in any other suitable manner. These simple modifications and combinations should also be considered as the content disclosed in the present invention and are all within the protection scope of the present invention.

Claims

1. A non-lactolytic streptococcus ( streptococcus alactolyticus ) strain, characterized in that, The non-lactolytic streptococci ( streptococcus alactolyticus The strain name is streptococcus alactolyticus 0625, this strain was deposited at the China General Microbiological Culture Collection Center on December 1, 2025, with accession number CGMCC No. 36838.

2. The non-lactolytic streptococcal strain according to claim 1, characterized in that, The non-lactolytic streptococci ( streptococcus alactolyticus The strain was isolated from the feces of C57BL / 6 mice.

3. A microbial inoculant, characterized in that, Contains the non-lactolytic streptococci as described in claim 1 or 2 ( streptococcus alactolyticus ) strains, and pharmaceutically or feed-acceptable vectors.

4. The microbial agent as described in claim 3, characterized in that, The non-lactolytic streptococcus in the bacterial agent ( streptococcus alactolyticus The viable count of the strain was 1×10⁻⁶. 8 CFU / mL to 1×10 10 CFU / mL.

5. The non-lactolytic streptococcus as described in claim 1 or 2 ( streptococcus alactolyticus The use of the strain or the microbial agent of claim 3 in the preparation of a composition for the prevention and / or treatment of intestinal inflammation.

6. The application according to claim 5, characterized in that, The intestinal inflammation mentioned is intestinal inflammation induced by sodium dextran sulfate.

7. A food, health product, or feed additive, characterized in that, Contains the non-lactolytic streptococci as described in claim 1 or 2 ( streptococcus alactolyticus ( ) strain or the microbial agent as described in claim 3.

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