A strain of Bifidobacterium animalis subspecies lactis ZYhyy-003 and its postbiotic preparation and application

Abstract

The present invention relates to the field of microbial technology, and provides a strain of Bifidobacterium animalis subspecies lactis ZYhyy‑003 and a postbiotic preparation and application thereof. Bifidobacterium animalis subsp.lactis )ZYhyy‑003 was deposited in the General Microbiology Center of China National Committee for the Preservation of Microbiological Cultures on August 5, 2024, with the deposit address at No. 3, Yard No. 1, Beichen West Road, Chaoyang District, Beijing, and the deposit number is CGMCC No. 31553. The animal Bifidobacterium lactis subsp. ZYhyy‑003 postbiotic preparation described in this application can increase the food intake of pets, maintain the healthy fecal state of pets, enhance the intestinal barrier function of pets and maintain the balance of intestinal flora, and has the characteristics of high safety and stability, easy storage and production, long shelf life and wide range of application scenarios, and can be used as the preferred additive for pet gastrointestinal conditioning related products.

Description

Technical Field

[0001] The invention belongs to the technical field of microorganisms, and particularly relates to a strain of animal Bifidobacterium lactis subspecies ZYhyy-003 and a postbiotic preparation and application thereof. Background Art

[0002] With the booming development of the pet industry, pet health issues have gradually received widespread attention. Among them, gastrointestinal health, as a key factor affecting the overall health and quality of life of pets, has received much attention from pet owners and industry experts. However, gastrointestinal dysfunction is becoming increasingly common in pets, manifested in symptoms such as loss of appetite, indigestion, diarrhea, and constipation, which not only seriously affects the normal growth and reproduction of pets, but also brings significant economic and psychological burdens to pet owners.

[0003] The main means of regulating pet gastrointestinal function on the current market include drug therapy and probiotic supplementation. Although drug therapy can quickly relieve symptoms, its side effects and the imbalance of intestinal microbiota and drug resistance that may result from long-term use limit its application. Probiotics are widely regarded as a more natural means of regulation because of their function of regulating the balance of intestinal flora. However, the efficacy of existing probiotic products is usually affected by multiple factors such as strain type, dosage, and delivery method, and many strains are difficult to survive effectively and exert long-term effects after entering the intestine. Therefore, the actual effect of existing probiotic products in improving pet gastrointestinal health is still insufficient.

[0004] Bifidobacterium animalis ( Bifidobacterium animalis ) is a widely studied probiotic with significant application potential in improving animal intestinal health, especially showing positive effects in balancing intestinal flora and enhancing digestive ability. However, the currently used animal Bifidobacterium strains still have the following significant problems:

[0005] Existing strains are easily inactivated under high-temperature processing conditions, which limits their widespread application in pet food; they are not effective enough in enhancing intestinal immune function and improving gastrointestinal health in the long term; and strict requirements on production environment, storage conditions and delivery methods limit their flexible application.

[0006] Therefore, there is an urgent need for a more efficient and stable solution to optimize pet gastrointestinal health management. This approach should be able to significantly improve pets' digestive function while reducing dependence on medications, providing more lasting health benefits, and thus comprehensively improving the overall health of pets. Summary of the invention

[0007] In order to overcome the problems of the prior art, the purpose of the present invention is to screen and obtain probiotics that can improve the gastrointestinal function of pets, and propose a strain of animal Bifidobacterium lactis subspecies ZYhyy-003 and its postbiotic preparation and application, which provide a new technical approach for pet gastrointestinal health management and provide a valuable reference for the development of functional nutritional preparations in the pet food industry.

[0008] The object of the present invention is achieved in that:

[0009] The first aspect of the present invention provides a strain of Bifidobacterium lactis subsp. ZYhyy-003 for improving the gastrointestinal function of pets, Bifidobacterium lactis subsp. Bifidobacterium animalis subsp. lactis ) ZYhyy-003 was deposited in the General Microbiology Center of China Microbiological Culture Collection Administration on August 5, 2024. The deposit address is No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, and the deposit number is CGMCC No. 31553.

[0010] The second aspect of the present invention provides a postbiotic preparation of animal Bifidobacterium lactis subspecies ZYhyy-003 for improving the gastrointestinal function of pets, which is prepared by uniformly mixing the postbiotic powder of animal Bifidobacterium lactis subspecies ZYhyy-003 described in the first aspect and glucose, and the postbiotic powder is prepared by inactivating, concentrating and freeze-drying the bacterial liquid of animal Bifidobacterium lactis subspecies ZYhyy-003.

[0011] Furthermore, the preparation of the animal Bifidobacterium lactis subspecies ZYhyy-003 bacterial liquid comprises the following steps:

[0012] (1) Take out the frozen Bifidobacterium animalis subsp. lactis ZYhyy-003 from the -80°C refrigerator, quickly thaw it in a 37°C water bath, inoculate the strain into 5 mL of liquid MRS medium, and culture it anaerobically at 37°C for 24 to 48 hours until the bacterial solution becomes turbid;

[0013] (2) The turbid bacterial solution obtained in step (1) is inoculated on an MRS solid culture medium for activation, the activated bacteria are inoculated in a primary culture medium, and a seed solution is obtained through primary culture; the seed solution is inoculated in a secondary culture medium, and a bacterial solution is obtained through secondary culture;

[0014] (3) Centrifuge the bacterial solution at 4°C, 3000-5000 rpm for 15 min, collect the cells, wash with sterile saline, and resuspend to obtain a suspension. Adjust the concentration of the suspension to greater than 1×10 9 CFU / mL, that is, the bacterial liquid of animal Bifidobacterium lactis subspecies ZYhyy-003 was obtained.

[0015] Furthermore, in step (1), the strain inoculation amount is 0.02-0.04%.

[0016] Furthermore, in step (2), the primary culture medium and the secondary culture medium are both MRS liquid culture medium. The preparation method of MRS liquid culture medium is as follows: 20.00 g of glucose, 10.00 g of peptone, 5.00 g of yeast extract, 10.00 g of beef extract, 1.00 g of Tween 80, 5.00 g of sodium acetate, 0.58 g of magnesium sulfate, 0.25 g of manganese sulfate and 2.00 g of potassium dihydrogen phosphate are dissolved in 1 L of deionized water and sterilized at 121°C for 15 min.

[0017] Furthermore, in step (2), the inoculation amount of the seed liquid inoculated into the secondary culture medium is 1% to 2%.

[0018] Furthermore, in step (2), the primary culture condition is culturing at 36-38°C for 24-48 h; the secondary culture condition is culturing at 36-38°C for 24-48 h.

[0019] Furthermore, the sterilization conditions are as follows: sterilization at 121°C for more than 30 min, applying the inactivated bacterial solution on an MRS plate, and placing it at 37°C for anaerobic culture for more than 48 h to ensure that no bacterial colonies grow.

[0020] The third party of the present invention provides the use of the animal Bifidobacterium lactis subspecies ZYhyy-003 described in the first aspect and the postbiotic preparation described in the second aspect in the preparation of products for improving the gastrointestinal function of pets.

[0021] Furthermore, the pet includes a cat, a dog, a rabbit, a piglet, a mink, a golden bear, a chinchilla, a guinea pig or a meerkat.

[0022] Furthermore, the product is one or more of the following:

[0023] (1) The product can increase the pet's food intake;

[0024] (2) The product can maintain the healthy fecal state of pets;

[0025] (3) The product can enhance the intestinal barrier function of pets;

[0026] (4) The product can maintain the balance of pet’s intestinal flora.

[0027] Strain deposit information:

[0028] Bifidobacterium animalis subsp. lactis ZYhyy-003, depository institution: General Microbiology Center of China Microbiological Culture Collection Administration (CGMCC); address: No. 3, Yard No. 1, Beichen West Road, Chaoyang District, Beijing, Postal Code: 100101; deposit date: August 5, 2024; deposit number CGMCC No. 31553; classification name: Bifidobacterium animalis subsp. lactis Bifidobacterium animalis subsp. lactis .

[0029] The advantages and beneficial effects of the present invention are:

[0030] 1. The present invention screened out a probiotic bacteria that can improve the gastrointestinal function of pets. The strain was preserved and identified as Bifidobacterium animalis subsp. lactis, with a preservation number of CGMCC No. 31553; classification name: Bifidobacterium animalis subsp. lactis Bifidobacterium animalis subsp. lactis .

[0031] 2. Experiments have confirmed that the effect of the animal Bifidobacterium lactis subspecies ZYhyy-003 postbiotic preparation provided by the present invention is better than the postbiotic solid preparation prepared by Lactobacillus plantarum that has been developed and marketed and has the function of regulating the gastrointestinal function of pets. Specifically, the animal Bifidobacterium lactis subspecies ZYhyy-003 postbiotic preparation described in this application can increase the food intake of pets, maintain the healthy fecal state of pets, enhance the intestinal barrier function of pets and maintain the balance of intestinal flora, and has the characteristics of high safety and stability, easy storage and production, long shelf life and wide range of application scenarios, and can be used as a preferred additive for regulating pet gastrointestinal related products. BRIEF DESCRIPTION OF THE DRAWINGS

[0032] The present invention will be further described below in conjunction with the accompanying drawings and embodiments.

[0033] Figure 1 Growth of postbiotic solids prepared from Bifidobacterium animalis subspecies lactis ZYhyy-003 described in Example 2 on MRS plates;

[0034] Figure 2 is the food intake of each group of cats in Example 3;

[0035] Figure 3 is the intestinal barrier function index of each group of cats in Example 3;

[0036] Figure 4 is the alpha diversity of cat feces flora in each group of Example 3;

[0037] Figure 5 The figure shows the difference in flora in cat feces of each group in Example 3. DETAILED DESCRIPTION

[0038] The examples are provided to better illustrate the present invention, but the present invention is not limited to the examples. Therefore, those skilled in the art may make non-essential improvements and adjustments to the implementation scheme according to the above invention content, which still fall within the protection scope of the present invention.

[0039] The endpoints and any values ​​of the ranges disclosed in this article 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 each range, the endpoint values ​​of each range and the individual point values, and the individual point values ​​can be combined with each other to obtain one or more new numerical ranges, which should be considered as specifically disclosed in this article.

[0040] The present invention will be described in detail below by way of examples. It should be understood that the following examples are only used to further explain and illustrate the content of the present invention in detail, and are not used to limit the present invention.

[0041] Example 1: Isolation and identification of bacterial species

[0042] 1. Source of the strain: feces of healthy pet cats in catteries and animal stray rescue stations in Yancheng District, Luohe City, Henan Province. The strain was screened and isolated from feces of healthy pet cats collected in large quantities during the period from July 1, 2023 to August 1, 2023.

[0043] 2. Screening and purification of strains

[0044] (1) Sampling: Use sterile tweezers to collect 10 g of fecal sample on a clean bench and set aside.

[0045] (2) Sample preparation:

[0046] ① Take 90 mL of sterilized saline (0.85%) and place it in a sterile conical flask, then add the sample from step (1), shake and set aside;

[0047] ② Perform gradient dilution of the dilution solution in step ①, diluting 10 -1 , 10 -2 , 10 -3 , 10 -4 and 10 -5 , numbered 1, 2, 3, 4 and 5, for use.

[0048] (3) Prepare MRS solid medium: dissolve 20.00 g of glucose, 10.00 g of peptone, 5.00 g of yeast extract, 10.00 g of beef extract, 1.00 g of Tween 80, 5.00 g of sodium acetate, 0.58 g of magnesium sulfate, 0.25 g of manganese sulfate, and 2.00 g of potassium dihydrogen phosphate in 1 L of deionized water. Add 17 g of agar and sterilize at 121°C for 15 min.

[0049] (4) Cultivation: 10 -1 , 10 -2 , 10 -3 , 10 -4 and 10 -5 The solution was spread on MRS solid medium plates using a spreading rod and cultured at 37°C for 48 h.

[0050] (5) Purification of strains: Select milky white raised colonies of different shapes and sizes for isolation and purification, and select a single colony to streak three zones on an MRS solid culture medium plate. After streaking, culture the plate at 37°C under anaerobic conditions for 48 h. Repeat the above operation for a total of three streakings to purify the pure strain.

[0051] 3. Identification of strains

[0052] The strain was cultured anaerobically at 37°C on an MRS solid medium plate for 48 h, and a single colony was picked for PCR. The primers were 16S rRNA universal primers 27F and 1492R, and the PCR reaction conditions were: 95°C pre-denaturation for 3 min; 95°C denaturation for 15 s, 60°C annealing for 15 s, 72°C extension for 90 s, 35 cycles; 72°C final extension for 5 min. The obtained 16S rRNA was subjected to BLAST analysis with the sequences of known strains on the GenBank website, and the identification result was: animal Bifidobacterium lactis subsp. Bifidobacterium animalis subsp. lactis ).

[0053] The identified strain was named Bifidobacterium animalis subsp. lactis ZYhyy-003, and the strain was sent to the General Microbiological Center of China Microbiological Culture Collection Administration for preservation. The preservation information is as follows;

[0054] Bifidobacterium animalis subsp. lactis ZYhyy-003, depository institution: General Microbiology Center of China Microbiological Culture Collection Administration (CGMCC); address: No. 3, Yard No. 1, Beichen West Road, Chaoyang District, Beijing; deposit date: August 5, 2024; deposit number CGMCC No. 31553; classification name: Bifidobacterium animalis subsp. lactis Bifidobacterium animalis subsp. lactis .

[0055] Example 2: Preparation of animal Bifidobacterium lactis subspecies ZYhyy-003 postbiotic solid preparation

[0056] In this example, animal Bifidobacterium lactis subspecies ZYhyy-003 was used to prepare a postbiotic solid preparation, and the specific method was as follows:

[0057] (1) Prepare MRS liquid medium: dissolve 20.00 g of glucose, 10.00 g of peptone, 5.00 g of yeast extract, 10.00 g of beef extract, 1.00 g of Tween 80, 5.00 g of sodium acetate, 0.58 g of magnesium sulfate, 0.25 g of manganese sulfate, and 2.00 g of potassium dihydrogen phosphate in 1 L of deionized water and sterilize at 121°C for 15 min.

[0058] (2) Animal Bifidobacterium lactis subsp. ZYhyy-003 was activated on MRS solid medium, and the activated bacteria were inoculated into MRS liquid medium, and cultured at 37°C for 48 h to obtain seed solution; the seed solution was inoculated into MRS liquid medium at an inoculum rate of 1%, and cultured at 37°C for 48 h to obtain bacterial solution; after centrifuging the bacterial solution, the bacteria were collected, washed with sterile saline, and resuspended to obtain a suspension; the concentration of the suspension was adjusted to obtain 1.0×10 10 CFU / mL of bacterial suspension.

[0059] (3) The bacterial suspension was sterilized at 121°C for 30 min, and the inactivated bacterial suspension was spread on an MRS plate and incubated at 37°C for more than 48 hours. The sterilized bacterial suspension was freeze-dried to obtain postbiotic powder; according to the number of live bacteria in the bacterial suspension, the postbiotic powder was mixed with glucose (purchased from Shanghai MacLean Biochemical Technology Co., Ltd.) to prepare a plate containing 2.0×10-100 cells of animal Bifidobacterium lactis subsp. ZYhyy-003. 10 CFU / g of postbiotic solid preparation.

[0060] The postbiotic solid preparation was taken and coated on an MRS plate in the same manner as above, and then placed at 37°C for anaerobically culture for more than 48 hours.

[0061] The results are as follows Figure 1 As shown, there are a large number of live bacteria in the non-sterile bacterial suspension after coating, while no live bacteria are detected in the sterilized bacterial suspension and postbiotic solid preparation.

[0062] Example 3: Effect of animal Bifidobacterium lactis subsp. ZYhyy-003 postbiotic solid preparation on gastrointestinal health of pet cats

[0063] In order to explore the effect of the animal Bifidobacterium lactis subsp. ZYhyy-003 postbiotic solid preparation described in Example 2 on the gastrointestinal health of pets, this example takes pet cats as an example and conducts the following animal experiments:

[0064] (1) Experimental groups

[0065] In this experiment, 16 pet cats from Zhongyuan Food Laboratory were randomly divided into 4 groups, 4 cats in each group, namely positive control group, negative control group, experimental group 1 and experimental group 2. The specific treatments are as follows:

[0066] Positive control group: A commercially available solid preparation of postbiotics for regulating pet gastrointestinal function (Lactobacillus plantarum ( Lactobacillus plantarum ) Postbiotics) were dissolved in 10 mL of sterile saline and administered orally once a day.

[0067] Negative control group: 10 mL of sterile saline was administered once a day.

[0068] Experimental group 1: The animal Bifidobacterium lactis subsp. ZYhyy-003 postbiotic solid preparation prepared in Example 2 was dissolved in 10 mL of sterile saline and then orally administered at a dose of 250 mg / kg body weight, once a day.

[0069] Experimental group 2: The animal Bifidobacterium lactis subsp. ZYhyy-003 postbiotic solid preparation prepared in Example 2 was dissolved in 10 mL of sterile saline and then orally administered at a dose of 500 mg / kg body weight, once a day.

[0070] All pet cats had completed vaccination and deworming before the experiment. The experimental environment was a 12-hour day and night cycle, the temperature was controlled at 18-26°C, and the relative humidity was 30%-60%. The cattery cleaned feces twice a day and disinfected. Each cat was fed twice a day, 40 grams of basic feed each time, and the feed was a baked staple food that met the nutritional standards of the Association of American Feed Control Officials (AAFCO, 2023) and the National Research Council (NRC, 2006). Free drinking water was provided. The week before the experiment was the adaptation period, and the following 28 days were the formal experimental period. Before the formal experiment, the feces of each cat were collected and stored in a -80°C refrigerator. During the experiment, the appetite changes, digestion speed and gastrointestinal discomfort symptoms of each cat were recorded every day. After the experiment, the blood and feces of the cats were collected respectively. The blood was centrifuged at 3000 rpm for 15 minutes to obtain serum to determine the intestinal barrier function. The feces were analyzed together with the feces collected before the experiment by 16S rRNA gene sequencing technology to analyze the changes in the pet's intestinal microbiome.

[0071] All data were statistically analyzed using GraphPad Prism 10.1.2. The significance level is: ns indicates no significant difference, * indicates p < 0.05, ** indicates p < 0.01, *** indicates p< 0.001, **** indicates p < 0.0001.

[0072] This study strictly complies with international animal use guidelines and the Regulations on the Administration of Laboratory Animals in China, and has been approved by the Animal Experiment Ethics Committee of China Agricultural University (approval number: AW11104202-5-2). All experiments were conducted under the premise of minimizing animal suffering.

[0073] (2) Effect of animal Bifidobacterium lactis subsp. ZYhyy-003 postbiotic solid preparation on cat food intake

[0074] Before the experiment began, a baseline test of the average daily food intake of all pet cats was performed. The food intake of each cat was recorded daily and the average was calculated every 7 days for a total of 4 weeks.

[0075] The average daily feed intake of the positive control group, negative control group, experimental group 1, and experimental group 2 was 60, 61, 59, and 60 g / day, respectively. There was no significant difference in the average daily feed intake of pet cats during the one-week adaptation period, indicating that the appetite of the four groups of pets was at a similar level. Figure 2 As shown in the figure, starting from the third week of feeding, the average feed intake of cats in experimental groups 1 and 2 was higher than that in the positive control group and the negative control group, indicating that the animal Bifidobacterium lactis subsp. ZYhyy-003 postbiotic solid preparation has a positive effect on the feeding behavior of cats. This may be related to the postbiotic solid preparation regulating intestinal health, improving metabolism and enhancing appetite.

[0076] (3) Evaluation of the effect of animal Bifidobacterium lactis subsp. ZYhyy-003 postbiotic solid preparation on cat fecal status

[0077] The stool scores were scored using the Bristol feces classification method (see Table 1), and were recorded by professionals after each week of the experiment. The experiment lasted for 4 weeks.

[0078]

[0079] The effect of the animal Bifidobacterium lactis subsp. ZYhyy-003 postbiotic solid preparation on the fecal status of cats is shown in Table 2. Whether in experimental group 1 or experimental group 2, the fecal quality has been in the healthy range, and no severe signs of constipation or diarrhea were shown. In contrast, the negative control group showed signs of mild diarrhea in the third and fourth weeks (scores of 5.75 and 6.0). The pet cats in the experimental group maintained a high intestinal health score throughout the experimental period, indicating that the postbiotic preparation has a positive effect on the intestinal health of pet cats. The intestinal health of the negative control group decreased significantly in the third and fourth weeks, which may reflect gastrointestinal problems caused by the lack of effective intervention measures.

[0080]

[0081] (4) Effect of animal Bifidobacterium lactis subsp. ZYhyy-003 postbiotic solid preparation on intestinal barrier function in cat serum

[0082] like Figure 3 As shown in Figure 2, there was no significant difference in D-lactate levels between the positive control group and the negative control group, but the D-lactate level in the negative control group was slightly higher, indicating that intestinal permeability may have increased. The D-lactate levels in both experimental groups 1 and 2 were significantly lower than those in the negative control group ( p < 0.05), especially in experimental group 2 (high dose), the reduction effect was more significant. This shows that the animal Bifidobacterium lactis subspecies ZYhyy-003 postbiotic solid preparation described in Example 2 can effectively reduce the concentration of metabolites related to intestinal permeability and protect the intestinal barrier function. The excellent effect of experimental group 2 further proves that the effect of the postbiotic solid preparation is dose-dependent.

[0083] The diamine oxidase (DAO) activity in the negative control group was significantly higher than that in the positive control group ( p < 0.01), indicating that the intestinal barrier was damaged and mucosal cells released DAO. The DAO activity of experimental groups 1 and 2 was significantly lower than that of the negative control group ( p < 0.05 and p < 0.001), the DAO activity of experimental group 2 was close to the level of the positive control group, but lower than the positive control group, and significantly better than experimental group 1. This shows that the animal Bifidobacterium lactis subspecies ZYhyy-003 postbiotic solid preparation described in Example 2 can effectively repair the intestinal mucosal barrier and reduce intestinal damage. In addition, the better effect of the high-dose postbiotic solid preparation shows that it has a stronger effect in improving intestinal barrier function.

[0084] The level of lipopolysaccharide (LPS) in the negative control group was significantly higher than that in the positive control group ( p < 0.05), indicating that increased intestinal permeability may have triggered endotoxin penetration and systemic inflammatory response. The LPS levels in experimental groups 1 and 2 were significantly lower than those in the negative control group ( p < 0.05 and p < 0.01), among which the LPS level of experimental group 2 was close to that of the positive control group and significantly better than that of experimental group 1. This indicates that the postbiotic solid preparation can significantly reduce the inflammatory response caused by intestinal barrier damage, which may be related to its regulation of intestinal flora and enhancement of intestinal mucosal barrier function.

[0085] (5) Effect of animal Bifidobacterium lactis subsp. ZYhyy-003 postbiotic solid preparation on cat fecal microorganisms

[0086] The α diversity of the bacterial community changes asFigure 4 As shown in the figure, Chao index is mainly used to evaluate the richness of the community. The higher the value, the richer the species of the community. The Chao index of the negative control group was significantly lower than that of the positive control group ( p < 0.01), indicating that intestinal barrier damage may lead to a decrease in the diversity and richness of the microbiota. The Chao index of experimental group 1 (low dose) and experimental group 2 (high dose) were significantly higher than those of the negative control group, and the Chao index of experimental group 2 was significantly close to that of the positive control group ( p <0.001). This result shows that the animal Bifidobacterium lactis subsp. ZYhyy-003 postbiotic solid preparation described in Example 2 can effectively improve the richness of intestinal flora and improve the flora disorder caused by barrier damage, and the effect of the high-dose group is better than that of the low-dose group, showing a dose-dependency.

[0087] The Shannon index is used to evaluate the diversity and uniformity of the flora. The higher the value, the more uniform and diverse the structure of the flora. There was no significant difference in the Shannon index between the positive control group and the negative control group, indicating that the effect of barrier damage on the uniformity of flora diversity may be weak or not obvious in the short term. There was no significant difference in the Shannon index between experimental group 1 and experimental group 2 compared with the negative control group, indicating that the animal Bifidobacterium lactis subspecies ZYhyy-003 postbiotic solid preparation described in Example 2 did not significantly improve the uniformity of the flora. This may be because the postbiotic solid preparation mainly acts to enrich specific probiotic species, but has little effect on overall diversity and uniformity.

[0088] The changes in the differential bacterial flora Figure 5 As shown in Figure 2, the animal Bifidobacterium lactis subsp. lactis ZYhyy-003 postbiotic solid preparation can significantly regulate the structure of cat intestinal flora and reduce the potential harmful bacteria at 28 days. Blautia The relative abundance of probiotics was significantly increased Bifidobacterium The negative control group showed an imbalance of microbiota caused by damaged intestinal barrier. Blautia The abundance increased and Bifidobacterium The abundance of Blautia Abundance and enhancement Bifidobacterium This indicates that the solid preparation of postbiotics plays an important role in improving the imbalance of intestinal flora, promoting the growth of probiotics and maintaining intestinal health, and is dose-dependent.

[0089] Finally, it should be noted that the above is only used to illustrate the technical solution of the present invention and is not a limitation. Although the present invention is described in detail with reference to the preferred arrangement scheme, a person skilled in the art should understand that the technical solution of the present invention can be modified or replaced by equivalents without departing from the spirit and scope of the technical solution of the present invention.

Claims

1. A postbiotic preparation of Bifidobacterium lactis subspecies ZYhyy-003 for improving gastrointestinal function of pets, characterized in that: The postbiotic powder is prepared by uniformly mixing the postbiotic powder of animal Bifidobacterium lactis subspecies ZYhyy-003 and glucose, wherein the postbiotic powder is prepared by inactivating, concentrating and freeze-drying the bacterial liquid of animal Bifidobacterium lactis subspecies ZYhyy-003; Bifidobacterium animalis subsp. lactis ( Bifidobacterium animalis subsp.lactis ) ZYhyy-003 was deposited in the General Microbiology Center of China Microbiological Culture Collection Administration on August 5, 2024. The deposit address is No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, and the deposit number is CGMCC No. 31553.

2. The postbiotic preparation according to claim 1, characterized in that The preparation of the animal Bifidobacterium lactis subspecies ZYhyy-003 bacterial liquid comprises the following steps: (1) Take out the frozen Bifidobacterium animalis subsp. lactis ZYhyy-003 from the -80°C refrigerator, quickly thaw it in a 37°C water bath, inoculate the strain into 5 mL of liquid MRS medium, and culture it anaerobically at 37°C for 24 to 48 hours until the bacterial solution becomes turbid; (2) The turbid bacterial solution obtained in step (1) is inoculated on an MRS solid culture medium for activation, the activated bacteria are inoculated in a primary culture medium, and a seed solution is obtained through primary culture; the seed solution is inoculated in a secondary culture medium, and a bacterial solution is obtained through secondary culture; (3) Centrifuge the bacterial solution at 4°C, 3000-5000 rpm for 15 min, collect the cells, wash with sterile saline, and resuspend to obtain a suspension. Adjust the concentration of the suspension to greater than 1×10 9 CFU / mL, that is, the bacterial liquid of animal Bifidobacterium lactis subspecies ZYhyy-003 was obtained.

3. The postbiotic preparation according to claim 2, characterized in that In step (1), the strain inoculation amount is 0.02~0.04%.

4. The postbiotic preparation according to claim 2, characterized in that In step (2), the primary culture medium and the secondary culture medium are both MRS liquid culture medium.

5. The postbiotic preparation according to claim 2 or 4, characterized in that In step (2), the inoculation amount of the seed liquid inoculated into the secondary culture medium is 1% to 2%.

6. The postbiotic preparation according to claim 2, characterized in that In step (2), the primary culture condition is culturing at 36-38°C for 24-48 h; the secondary culture condition is culturing at 36-38°C for 24-48 h.

7. The postbiotic preparation according to claim 1, characterized in that The inactivation conditions are as follows: sterilization at 121°C for more than 30 min, applying the inactivated bacterial solution to an MRS plate, and placing it at 37°C for anaerobic culture for more than 48 h to ensure that no bacterial colonies grow.

8. Use of the animal Bifidobacterium lactis subsp. ZYhyy-003 postbiotic preparation according to any one of claims 1 to 7 in the preparation of a product for improving gastrointestinal function of pets; The product is one or more of the following: (1) The product can increase the pet's food intake; (2) The product can maintain the healthy fecal state of pets; (3) The product can enhance the intestinal barrier function of pets; (4) The product can maintain the balance of pet’s intestinal flora.

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