Pediococcus pentosaceus ZRS217 and application thereof in high-quality daily diet for yaks

Abstract

The application discloses a pentosus ZRS217 and application thereof in high-energy daily rations for yaks, and belongs to the technical field of microorganism and ruminant nutrition regulation. The strain is preserved in the China General Microbiological Culture Collection Center (CGMCC) and has a preservation number of CGMCC No. 38958. The pentosus ZRS217 disclosed by the application can significantly improve the growth performance of yaks, effectively relieve oxidative stress and inflammatory response caused by high-energy daily rations, significantly increase the antioxidant enzyme activity in serum, significantly reduce the level of proinflammatory factors, protect the intestinal barrier function, reduce mucosal injury, improve liver health, reduce endotoxin level and reduce lipid deposition, and provides an efficient and safe microecological regulation scheme for the breeding of yaks on the plateau.

Description

Technical Field

[0001] This invention belongs to the field of microbial technology and ruminant nutrition regulation technology, specifically involving a type of Pediococcus pentosaceus ZRS217 and its application in high-precision diets for yaks. Background Technology

[0002] With the development of the livestock industry, high-concentration diets (i.e., diets with a significantly higher proportion of concentrate than roughage, such as a roughage-to-concentrate ratio of 30:70) have been widely used in the breeding practices of beef cattle, dairy cattle, and yaks in order to improve the growth performance and production efficiency of ruminants. However, long-term or excessive feeding of high-concentration diets can easily lead to rumen environment imbalance and trigger a series of metabolic and immune-related problems, collectively referred to as "high-concentration diet-induced injury".

[0003] Studies have shown that highly concentrated diets can significantly reduce rumen pH, promote the release of endotoxins (such as lipopolysaccharide, LPS), and allow them to enter the bloodstream via the gut-hepatic axis, inducing systemic oxidative stress and chronic inflammation. Specifically, this manifests as decreased serum total antioxidant capacity (T-AOC), superoxide dismutase (SOD), and catalase (CAT) activity, and increased malondialdehyde (MDA) levels. Pro-inflammatory factors such as interleukin-1β (IL-1β), IL-6, tumor necrosis factor-α (TNF-α), and interferon-γ (IFN-γ) levels are significantly elevated, while the expression of the anti-inflammatory factor IL-10 is suppressed. Furthermore, highly concentrated diets can damage the intestinal mucosal structure, leading to villus breakage, impaired epithelial barrier function, and promoting lipid deposition and LPS accumulation in the liver, ultimately affecting animal growth performance and health. This problem is particularly prominent in yak farming. As a unique livestock species of the plateau, yaks are highly adapted to low-energy, high-fiber natural pastures in terms of digestive physiology, and their tolerance to high-concentration diets is far lower than that of ordinary cattle.

[0004] Probiotics, as green and safe feed additives, have shown great potential in regulating the intestinal microecology and enhancing the host's antioxidant and immune functions. Among them, lactic acid bacteria (such as Lactobacillus and Pediococcus) are widely used due to their acid-producing, antibacterial, adhesive, and immunomodulatory properties. Pediococcus pentosaceus is a Gram-positive, facultative anaerobic lactic acid bacterium that has been used in food fermentation and animal feed. However, existing commercial Pediococcus pentosaceus strains generally suffer from poor gastrointestinal tolerance, weak antioxidant capacity, and limited effectiveness in alleviating stress from high-precision diets, making it difficult to meet the health needs of special ruminants such as yaks under high-precision feeding models.

[0005] Therefore, it is particularly important to develop a probiotic strain that can significantly improve the multidimensional damage to the oxidative-inflammatory-gut-liver induced by high-precision diets in yaks. Summary of the Invention

[0006] In view of the shortcomings of the prior art, the present invention provides a Pediococcus pentosaceus ZRS217 and its application in high-precision diets for yaks, which solves the problems of poor gastrointestinal tolerance, weak antioxidant capacity and limited effect on alleviating multi-system damage induced by high-precision diets in existing commercial probiotic strains, and provides an efficient and safe microecological regulation solution for the breeding of plateau yaks.

[0007] To achieve the above objectives, the present invention provides the following technical solution: This invention provides a strain of Pediococcus pentosaceus ZRS217 ( Pediococcus pentosaceus ZRS217, with accession number CGMCC No. 38958, was deposited on May 29, 2026, at the China General Microbiological Culture Collection Center (CGMCC), located at No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing. This strain was isolated from naturally fermented yogurt from farmers in Nagqu City, Tibet Autonomous Region.

[0008] Furthermore, the *Pediococcus pentosaceus* ZRS217 has the following characteristics: (a) Tolerance to artificial gastric fluid: The survival rate was 62.02% after treatment at 37°C for 3 hours in artificial gastric fluid at pH 1.5; Tolerance to artificial intestinal fluid: The survival rate was 55.22% after treatment at 37°C for 3 hours in artificial intestinal fluid at pH 6.8. (b) Bile salt tolerance: The survival rate was 72.92% after treatment at 37°C for 3 h in a medium containing 0.1% bile salts, 55.53% in a medium containing 0.2% bile salts, and 46.98% in a medium containing 0.3% bile salts. (c) It has antioxidant activity, with a DPPH free radical scavenging rate of 53.43% and a hydroxyl free radical scavenging rate of 36.55%.

[0009] The present invention also provides the application of the above-mentioned Pediococcus pentosaceus ZRS217 in the preparation of feed additives for improving damage induced by high-precision diets in ruminants.

[0010] Furthermore, the ruminant is a yak.

[0011] Furthermore, the damage induced by the high-precision diet includes at least one of the following: decreased growth performance, oxidative stress, inflammatory response, intestinal barrier damage, or liver metabolic disorder.

[0012] Furthermore, the amount of *Pediococcus pentosaceus* ZRS217 added is 1×10⁻⁶. 8 CFU / mL high-precision diet.

[0013] The present invention also provides a feed additive comprising the aforementioned Pediococcus pentosaceus ZRS217, with a viable bacterial concentration of not less than 1×10⁻⁶. 8CFU / mL.

[0014] The present invention also provides a high-precision diet for yaks, which comprises the feed additives described above.

[0015] Compared with the prior art, the present invention has the following beneficial effects: (1) The survival rate of *Pediococcus pentosus* ZRS217 of this invention reached 62.02% after treatment in artificial gastric fluid (pH 1.5) for 3 h and 55.22% after treatment in artificial intestinal fluid (pH 6.8) for 3 h, indicating that this strain has excellent tolerance to gastric acid and intestinal environment, and can effectively cross the gastrointestinal barrier and colonize and exert its function in the intestine. In terms of bile salt tolerance, the survival rates of the strain after treatment in culture media containing 0.1%, 0.2%, and 0.3% ox bile salts for 3 h were 72.92%, 55.53%, and 46.98%, respectively. In particular, it can still maintain a survival rate of nearly 50% even at a high concentration of 0.3% bile salts, which is significantly better than most commercial lactic acid bacteria strains, ensuring that it can still maintain its activity in the complex intestinal environment.

[0016] (2) The *Pediococcus pentosaceus* ZRS217 strain of this invention can effectively alleviate oxidative stress, significantly increasing serum total antioxidant capacity (T-AOC) to 6.17 μmol Trolox / mL, superoxide dismutase (SOD) activity to 20.48 U / mL, catalase (CAT) activity to 39.83 U / mL, and malondialdehyde (MDA) content to 0.46 nmol / mL (P<0.001). Regarding inflammation regulation, this strain can significantly reduce the levels of pro-inflammatory factors interleukin-1β (165.13 pg / mL), interleukin-6 (57.25 pg / mL), tumor necrosis factor-α (27.12 pg / mL), and interferon-γ (242.40 pg / mL), while significantly increasing the level of the anti-inflammatory factor interleukin-10 (41.20 pg / mL) (P<0.001). Histological observation of the intestines showed that this strain can effectively protect the intestinal barrier function and reduce mucosal damage such as villus breakage and shedding caused by high-refined diets. Regarding liver health, this strain can significantly reduce hepatic lipopolysaccharide (LPS) levels, alleviate hepatic lipid deposition, and improve metabolic disorders of total cholesterol, triglycerides, and low-density lipoprotein. Its overall efficacy is significantly superior to commercial Pediococcus pentosaceus. Attached Figure Description

[0017] Figure 1 This is a morphological diagram of Pediococcus pentosaceus ZRS217, as described in this invention. Figure 2 Gram-stained optical micrograph of Pediococcus pentosaceus ZRS217, as described in this invention; Figure 3The image shown is a scanning electron microscope (SEM) image of Pediococcus pentosaceus ZRS217 of this invention. Figure 4 This is the phylogenetic tree of Pediococcus pentosaceus ZRS217 of the present invention; Figure 5 The survival rate of Pediococcus pentosaceus ZRS217 of this invention in artificial gastric fluid (pH 1.5) and artificial intestinal fluid (pH 6.8); Figure 6 The survival rate of the *Pediococcus pentosaceus* ZRS217 strain of this invention under different bile salt concentrations (37℃, 3 h). Figure 7 The scavenging rate of DPPH and hydroxyl radicals by Pediococcus pentosaceus ZRS217 of this invention; Figure 8 HE staining results of jejunal tissue from yaks in each group (CON: basal diet group; T1: high-concentration diet group; T2: high-concentration diet + ZRS217 group; T3: high-concentration diet + commercial Pediococcus pentosaccharide group). Figure 9 Comparison of LPS content in yak livers among different groups (CON: basal diet group; T1: high-concentrate diet group; T2: high-concentrate diet + ZRS217 group; T3: high-concentrate diet + commercial Pediococcus pentosaccharide group). Figure 10 Oil Red O staining of liver (left) and serum TC, TG, LDL, and HDL levels (right); CON: basal diet, T1: high-concentration diet, T2: high-concentration diet + ZRS217, T3: high-concentration diet + commercial bacteria. Detailed Implementation

[0018] The technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0019] Unless otherwise specified, the experimental methods used in the following examples are conventional methods.

[0020] Unless otherwise specified, all materials and reagents used in the following examples are commercially available.

[0021] The technical solution of the present invention will be further described below with reference to the accompanying drawings and specific embodiments: Example 1 This example describes the screening, isolation, purification, identification, and preservation of Pediococcus pentosaceus ZRS217.

[0022] 1.1 Isolation and Purification of Strains The Pediococcus pentosaceus ZRS217 described in this invention was isolated from naturally fermented yak yogurt by farmers in Nagqu City, Tibet Autonomous Region.

[0023] Take 1.0 g of naturally fermented yak yogurt sample and add it to 9.0 mL of sterile physiological saline. After thorough shaking and mixing, prepare a 10⁻¹ dilution and then serially dilute it 10-fold. Spread 100 μL of the appropriate dilution onto MRS solid medium plates and incubate at 37℃ for 24–48 h. Select single colonies with regular morphology, neat edges, smooth surface, milky white or grayish white color, and suspected to be lactic acid bacteria. Repeat streak purification 3–5 times to obtain pure culture, named ZRS217.

[0024] 1.2 Observation of morphological characteristics Strain ZRS217 forms round or nearly round colonies on MRS solid medium. The colony surface is smooth and moist, with neat edges and opacity (e.g., Figure 1 (As shown). Gram staining revealed that this strain is Gram-positive, appearing spherical or nearly spherical under a microscope, mostly arranged singly, in pairs, or in short chains, and does not form spores (as shown). Figure 2 As shown). Scanning electron microscopy observation (e.g.) Figure 3 As shown in the figure, strain ZRS217 has spherical or ellipsoidal cells with relatively uniform cell morphology and a smooth surface, which is consistent with the typical morphological characteristics of Pediococcus strains.

[0025] 1.3 Molecular biological identification and preservation Genomic DNA was extracted from strain ZRS217, and its 16S rRNA gene sequence was amplified using the bacterial universal primers 27F 5'-AGAGTTTGATCCTGGCTCAG-3' (SEQ ID NO.1) and 1492R 5'-GGTTACCTTGTTACGACTT-3' (SEQ ID NO.2), resulting in a 1427bp 16S rRNA gene sequence (SEQ ID NO.3).

[0026] The sequence was submitted to the NCBI database for BLAST comparison, and the results showed that strain ZRS217 was similar to... Pediococcus pentosaceus The 16S rRNA gene sequence showed the highest similarity (99.72%). A phylogenetic tree was constructed based on the 16S rRNA gene sequence using MEGA software (e.g., Figure 4 As shown), the results indicated that strain ZRS217 was similar to Pediococcus pentosaceus (…). Pediococcus pentosaceusRelated strains clustered into the same branch. Based on colony morphology, microscopic morphology, physiological and biochemical characteristics, and 16S rRNA gene sequence analysis, strain ZRS217 was identified as *Pediococcus pentosaceus*, and named *Pediococcus pentosaceus*. Pediococcus pentosaceus ZRS217.

[0027] This strain was deposited on May 29, 2026, at the China General Microbiological Culture Collection Center (CGMCC), accession number: CGMCC No. 38958, address: No. 3, No. 1 Beichen West Road, Chaoyang District, Beijing.

[0028] Example 2 This example describes the determination of the biological characteristics of Pediococcus pentosaceus ZRS217.

[0029] 2.1 Artificial gastric juice tolerance test Pediococcus pentosaceus ZRS217 was inoculated into MRS liquid medium and incubated statically at 37°C for 24 h. The cells were collected by centrifugation, washed twice with sterile physiological saline, and resuspended to a concentration of approximately 1 × 10⁸ CFU / mL. 10 mL of the bacterial suspension was added to 90 mL of simulated gastric and intestinal fluid, respectively, and incubated at 37°C with shaking for 0 h and 3 h, respectively. Samples were taken at 0 h and 3 h, and the viable cell count (CFU / mL) was determined using the plate count method. The survival rate was calculated using the value at 0 h as a blank control.

[0030] Survival rate calculation formula: Survival rate (%) = ×100%.

[0031] Preparation of artificial gastric fluid: Take an appropriate amount of 1 mol / L HCl solution, dilute with water and adjust the pH to 1.5; add 1.0 g of pepsin to every 100 mL of solution, mix well and filter through a 0.2 μm sterile filter for later use.

[0032] The results are as follows Figure 5 As shown, the survival rate of strain ZRS217 after treatment in artificial gastric fluid at pH 1.5 for 3 hours was 62.02%, indicating that it has good gastric acid tolerance.

[0033] 2.2 Artificial Intestinal Fluid Tolerance Test The procedure is the same as the gastric juice test, except that the artificial gastric juice is replaced with artificial intestinal juice, and the treatment conditions remain the same (37℃, 3h).

[0034] Preparation of artificial intestinal fluid: Dissolve 6.8g of KH2PO4 in 500ml of water, and adjust the pH to 6.8 with 0.4% (w / w) NaOH. Add 1g of trypsin to every 100ml of liquid, mix well, filter through a 0.2μm sterile filter, and set aside.

[0035] The results are as follows Figure 5 As shown, strain ZRS217 had a survival rate of 55.22% after being treated in artificial intestinal fluid (pH 6.8) for 3 hours, indicating that it has good adaptability to the intestinal environment.

[0036] 2.3 Bile salt tolerance test Adjust the Pediococcus pentosacchari ZRS217 bacterial suspension to 1×10⁻⁶. 8 CFU / mL, at a 2% inoculum, was inoculated into MRS liquid medium containing 0.1%, 0.2%, and 0.3% ox bile salts, respectively (100 mL of MRS medium was fully dissolved, sterilized, and cooled to room temperature before adding 0.3 g of ox bile salts). Samples were taken after incubation at 37°C for 3 h, and the OD600 value of each bacterial solution was measured; the acid and bile salt tolerance of the strain was evaluated.

[0037] Survival rate (%) = ×100%; Where: OD1 is the OD value after bile salt treatment, and OD0 is the OD value without bile salt treatment.

[0038] The results are as follows Figure 6 As shown, the survival rates of strain ZRS217 after 3 hours of treatment in media containing 0.1%, 0.2%, and 0.3% bile salts were 72.92%, 55.53%, and 46.98%, respectively, indicating that it has good tolerance to different concentrations of bile salts. In particular, it can still maintain a survival rate of nearly 50% even at a high concentration of 0.3% bile salts, which is significantly better than most commercial lactic acid bacteria strains.

[0039] 2.4 Antioxidant Activity Assay (1) Determination of DPPH free radical scavenging ability Pediococcus pentosaceus ZRS217 cells were washed three times with phosphate-buffered saline (PBS, pH 7.4), resuspended, and the concentration was adjusted to 1.0 × 10⁻⁶. 9 CFU / mL; Add 1.0 mL of the bacterial suspension to 1.0 mL of 0.02 mol / L DPPH ethanol solution, mix thoroughly, and react at room temperature in the dark for 30 min; Simultaneously set up a control group (1.0 mL PBS + 1.0 mL DPPH solution) and a blank group (1.0 mL bacterial suspension + 1.0 mL anhydrous ethanol). After the reaction, centrifuge at 5000 rpm for 10 min, and measure the absorbance (OD value) of the supernatant at a wavelength of 517 nm. The calculation formula is as follows: DPPH free radical scavenging rate (%) = [1- ] × 100%.

[0040] The results are as follows Figure 7As shown, Pediococcus pentosaceus ZRS217 achieved a scavenging rate of 53.43% for DPPH free radicals, indicating that Pediococcus pentosaceus ZRS217 has a strong hydrogen atom donor capability and can effectively neutralize and stabilize the free radical DPPH•.

[0041] (2) Determination of hydroxyl radical scavenging ability Using o-phenanthroline-Fe 2+ The -H2O2 system generates •OH, and Fe is detected. 2+ The absorbance change of the o-phenanthroline complex at 536 nm indirectly reflects the •OH scavenging efficiency. The specific procedure is as follows: Take 0.5 mL of Pediococcus pentosaceus ZRS217 bacterial suspension (1.0 × 10⁻⁶). 9 Add 0.5 mL of o-phenanthroline (2.5 mmol / L), 1.0 mL of PBS (20 mmol / L), 0.5 mL of FeSO4 (2.5 mmol / L), and 0.5 mL of H2O2 (2.5 mmol / L) sequentially to the control group (CFU / mL). Mix well and incubate at 37°C for 60 min. The control group used an equal volume of double-distilled water instead of the bacterial suspension, while the blank group used double-distilled water instead of both the bacterial suspension and H2O2. After the reaction, centrifuge at 5000 rpm for 5 min, and measure the absorbance (OD value) of the supernatant at 536 nm using the following formula: Hydroxyl radical scavenging rate (%) = ×100%.

[0042] The results are as follows Figure 7 As shown, Pediococcus pentosaceus ZRS217 scavenged hydroxyl radicals at a rate of 36.55%.

[0043] The above results indicate that Pediococcus pentosus ZRS217 not only has excellent gastrointestinal tolerance but also exhibits significant in vitro antioxidant activity, providing a theoretical basis for its application in improving oxidative stress induced by high-precipitate diets.

[0044] Example 3 This example demonstrates the application effect of Pediococcus pentosaceus ZRS217 in high-precision yak diets.

[0045] 3.1 Group Design: Thirty-two healthy male yaks of similar health, weight, and age (around 3.5 years) were selected and randomly divided into four groups of eight yaks each: CON group (control group): fed with a basal diet with a roughage-to-concentrate ratio of 70:30; Group T1 (High-concentration diet model group): fed with a high-concentration diet, with a roughage-to-concentrate ratio of 30:70; Group T2 (experimental group): The *Pediococcus pentosaceus* ZRS217 strain of this invention was added to a high-concentration diet at a dosage of 1×10⁻⁶. 8CFU / mL; Group T3 (positive control group): *Pediococcus pentosaceus* was added to the high-concentration diet. Pediococcus pentosaceus CGMCC 1.12961 (purchased from China General Microbiological Culture Collection Center), with an addition dose of 1×10⁻⁶. 8 CFU / mL.

[0046] The basal diet and the high-concentration diet were fed at the same ratio. All yaks were housed individually in standard pens equipped with separate feed troughs and water buckets. During the trial, yaks were fed twice daily at 08:00 and 17:00, with free access to water and natural lighting. The trial included a 7-day pre-trial period (for acclimatization to the experimental environment and diet) and a 60-day formal trial period.

[0047] 3.2 Preparation of bacterial suspension: Pediococcus pentosaceus ZRS217 bacterial suspension: Pediococcus pentosaceus ZRS217 was inoculated into MRS liquid medium and incubated statically at 37°C for 24 h. The bacterial cells were collected by centrifugation, washed twice with sterile physiological saline, and resuspended. The bacterial concentration was adjusted to 1×10⁻⁶. 8 CFU / mL; Pediococcus pentosaceus bacterial suspension: Pediococcus pentosaceus (CGMCC 1.12961) was prepared using the same method to a viable count of 1×10⁻⁶. 8 CFU / mL bacterial suspension.

[0048] 3.3 Feeding method: Before daily feeding, prepare the corresponding bacterial suspension at a ratio of 1×10 8 The CFU / mL dose was added to the diets of groups T2 and T3, and fed immediately after thorough mixing. An equal volume of sterile saline was added to groups CON and T1 to ensure consistent treatment conditions for each group.

[0049] The diet used in this experiment was formulated from roughage and concentrate in different proportions, including: Roughage: Composed of oat hay and alfalfa hay; Concentrated feed: Composed of corn, soybean meal, wheat bran, extruded soybeans, NaCl, calcium source or buffer, and premix; by mass percentage, corn 50.00%, soybean meal 25.00%, wheat bran 12.50%, extruded soybeans 5.00%, NaCl 1.25%, calcium source or buffer 1.25%, and premix 5.00%.

[0050] Basic diet (CON group): 70% roughage + 30% concentrate (roughage to concentrate ratio 70:30); High-concentration diet (T1, T2, T3 groups): 30% roughage + 70% concentrate (roughage to concentrate ratio 30:70).

[0051] 3.4 Sample Collection and Index Measurement (1) Growth performance indicators Record initial weight, final weight, and average daily weight gain (ADG). (2) Serum biochemical indicators On day 60 of the experiment, 10 mL of blood was collected from the jugular vein before morning feeding. Serum was separated by centrifugation at 3000 rpm for 15 min and stored at -80℃ for later analysis. Antioxidant indicators: total antioxidant capacity (T-AOC), glutathione peroxidase (GSH-Px), superoxide dismutase (SOD), catalase (CAT), malondialdehyde (MDA); inflammatory factors: interleukin-1β (IL-1β), interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α), interferon-γ (IFN-γ), interleukin-10 (IL-10).

[0052] (3) Histological observation of the intestines After the experiment, four yaks were randomly selected from each group for slaughter, and jejunal tissue was collected, fixed with 4% paraformaldehyde, embedded in paraffin, sectioned, stained with hematoxylin and eosin (HE), and observed under a microscope for changes in the intestinal mucosal structure.

[0053] (4) Liver marker measurement Liver histology: Liver tissue was collected, fixed, embedded, sectioned, and stained with Oil Red O to observe the lipid droplet deposition in hepatocytes; Liver endotoxins: The content of lipopolysaccharide (LPS) in liver tissue was determined by ELISA. Serum lipid metabolism indicators: Measure the levels of serum total cholesterol (TC), triglycerides (TG), high-density lipoprotein (HDL), and low-density lipoprotein (LDL).

[0054] 3.5 Analysis of Experimental Results (1) The effects of the present invention Pediococcus pentosaceus ZRS217 on the growth performance of yaks are shown in Table 1.

[0055] Table 1

[0056] As shown in Table 1, there was no significant difference in the initial weight of yaks among the experimental groups before grouping (P = 0.971), indicating that the experimental groups were comparable. At the end of the experiment, there were significant differences in the final weight among the groups (P = 0.029), with group T2 having the highest final weight at 256.81 kg, significantly higher than group CON (P<0.05); groups T1 and T3 were at an intermediate level. Regarding average daily weight gain, the differences among the groups were extremely significant (P<0.001), with group T2 having the highest average daily weight gain at 0.72 kg / d, significantly higher than the other groups (P<0.05); groups T1 and T3 were next, and group CON had the lowest. This indicates that adding Pediococcus pentosaccharide ZRS217 of this invention to a high-concentration diet can improve the final weight and average daily weight gain of yaks, improve growth performance, and the effect is superior to commercial Pediococcus pentosaccharide.

[0057] (2) The effects of Pediococcus pentosaceus ZRS217 of the present invention on antioxidant indicators of yak serum are shown in Table 2.

[0058] Table 2

[0059] As shown in Table 2, the high-concentration diet significantly affected the antioxidant capacity of yak serum. Compared with the CON group, the total antioxidant capacity (T-AOC), superoxide dismutase (SOD), and catalase (CAT) levels in the T1 group were significantly reduced (P<0.05), while the malondialdehyde (MDA) content was significantly increased (P<0.05), indicating that the high-concentration diet induced significant oxidative stress. There was no significant difference in glutathione peroxidase (GSH-Px) among the groups (P = 0.617).

[0060] After adding Pediococcus pentosus ZRS217 of this invention, the serum antioxidant indicators of group T2 were significantly improved; among them, T-AOC of group T2 was 6.17 μmol Trolox / mL, SOD was 20.48 U / mL, and CAT was 39.83 U / mL, all of which were significantly higher than other treatment groups (P<0.05); at the same time, the MDA content of group T2 decreased to 0.46 nmol / mL, which was significantly lower than that of group T3 (P<0.05). Compared with the commercial Pediococcus pentosus group (group T3), the antioxidant indicators of group T2 were more significantly improved.

[0061] This indicates that the Pentosacchariformis ZRS217 of the present invention can increase the activity of antioxidant enzymes in the serum of yaks fed a high-precision diet and reduce the level of lipid peroxidation product MDA, thereby alleviating oxidative stress induced by a high-precision diet.

[0062] (3) The effects of the present invention Pediococcus pentosaceus ZRS217 on inflammatory factors in yak serum are shown in Table 3.

[0063] Table 3

[0064] As shown in Table 3, the high-concentration diet significantly induced serum inflammatory response in yaks. Compared with the CON group, the levels of pro-inflammatory factors interleukin-1β (IL-1β), interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α), and interferon-γ (IFN-γ) were significantly increased in the T1 group (P<0.05), while the level of anti-inflammatory factor interleukin-10 (IL-10) was significantly decreased (P<0.05), indicating that the high-concentration diet led to an increase in the body's inflammation level.

[0065] After adding Pediococcus pentosaceus ZRS217 of this invention, the serum inflammatory factor levels in group T2 were significantly improved. Specifically, the levels of IL-1β, IL-6, TNF-α, and IFN-γ in group T2 decreased to 165.13 pg / mL, 57.25 pg / mL, 27.12 pg / mL, and 242.40 pg / mL, respectively, all significantly lower than those in group T1 (P<0.05); at the same time, IL-10 in group T2 increased to 41.20 pg / mL, significantly higher than that in group T1 (P<0.05); compared with the commercial Pediococcus pentosaceus group (group T3), group T2 had lower overall levels of pro-inflammatory factors and higher levels of IL-10.

[0066] This indicates that the Pediococcus pentosaccharide ZRS217 of the present invention can inhibit the increase of serum pro-inflammatory factors induced by high-precipitate diets and promote the secretion of anti-inflammatory factor IL-10, thereby alleviating the inflammatory response in yaks fed high-precipitate diets. Its improvement effect is superior to that of commercial Pediococcus pentosaccharide.

[0067] (4) Histological observation of the intestine, the results of hematoxylin-eosin (HE) staining of the intestine are as follows: Figure 8 As shown.

[0068] from Figure 8 It can be seen that: CON group: the intestinal tissue structure was intact, the mucosal layer was continuous, the villi were arranged relatively neatly, and no obvious tissue damage was observed; T1 group: after treatment with high-concentration diet, the intestinal mucosal structure was significantly damaged, manifested as villi breakage and shedding, and the mucosal layer structure was disordered, indicating that high-concentration diet caused significant damage to the yak intestinal tissue; T2 group: after adding Pediococcus pentosaccharide ZRS217 of this invention, the intestinal tissue structure was significantly improved, the villi morphology was relatively intact, and the degree of mucosal damage was significantly reduced, indicating that Pediococcus pentosaccharide ZRS217 can effectively alleviate the intestinal tissue damage induced by high-concentration diet; T3 group: after adding commercial Pediococcus pentosaccharide, it also showed some improvement, but its intestinal structure recovery was not as good as that of T2 group.

[0069] (5) Liver lipopolysaccharide (LPS) test results are as follows Figure 9 As shown.

[0070] As shown in the figure, the liver LPS level in group T1 was significantly higher than that in group CON (P<0.05), indicating that high-concentration diets can lead to increased endotoxin levels in yaks and may induce liver inflammation or metabolic stress. Compared with group T1, the liver LPS level in group T2 was significantly reduced after the addition of Pediococcus pentosaccharide ZRS217 of this invention, and there was no significant difference compared with group CON (P>0.05), indicating that Pediococcus pentosaccharide ZRS217 can effectively alleviate the abnormal increase in liver endotoxins caused by high-concentration diets. The liver LPS level in group T3 was also reduced after treatment with commercial Pediococcus pentosaccharide, but the overall effect was not as good as that in group T2.

[0071] (6) Results of liver lipid metabolism and liver oil red staining are as follows: Figure 10 As shown.

[0072] Oil Red O staining results for the liver showed that: in the CON group, there were fewer lipid droplets in the liver tissue and the hepatocyte structure was relatively clear; in the T1 group, there were significantly more red lipid droplets in the liver tissue, suggesting that a high-precipitate diet can lead to increased lipid deposition in the liver of yaks; compared with the T1 group, the addition of Pediococcus pentosaceus ZRS217 to the T2 group significantly reduced the number of lipid droplets in the liver tissue and alleviated the degree of lipid deposition; the T3 group also showed some improvement after treatment with commercial Pediococcus pentosaceus, but the overall improvement effect was weaker than that of the T2 group.

[0073] Serum lipid metabolism parameters showed that, compared with the CON group, the levels of total cholesterol (TC), triglycerides (TG), and low-density lipoprotein (LDL) in the T1 group were significantly increased (P<0.05), while high-density lipoprotein (HDL) showed no significant change (P>0.05), indicating that a high-concentration diet can cause lipid metabolism disorders in yaks. After adding Pediococcus pentosaccharide ZRS217 of this invention, the levels of TC, TG, and LDL in the T2 group were significantly lower than those in the T1 group (P<0.05) and approached the levels of the CON group; at the same time, the HDL level in the T2 group increased; compared with the commercial Pediococcus pentosaccharide group (T3 group), the T2 group showed a more significant effect in improving lipid metabolism.

[0074] In summary, the *Pediococcus pentosaceus* ZRS217 strain of this invention exhibits significant multi-target protective effects in improving multi-system damage induced by high-precipitate diets in yaks. This strain not only significantly increases the final weight and average daily weight gain of yaks, improving growth performance, but its effects are significantly superior to commercial *Pediococcus pentosaceus* strains. Simultaneously, it effectively alleviates oxidative stress by enhancing the body's antioxidant capacity by increasing serum antioxidant enzyme activity and reducing lipid peroxidation product levels. Regarding inflammation regulation, *Pediococcus pentosaceus* ZRS217 significantly reduces pro-inflammatory factor levels and increases anti-inflammatory factor levels, effectively inhibiting the inflammatory response induced by high-precipitate diets. Furthermore, this strain also protects the intestinal barrier function, mitigating intestinal mucosal structural damage caused by high-precipitate diets. In terms of liver health, *Pediococcus pentosaceus* ZRS217 reduces liver endotoxin levels, alleviates lipid deposition, and improves lipid metabolism disorders. Its overall efficacy is significantly superior to commercial *Pediococcus pentosaceus* strains.

[0075] The foregoing has provided a detailed description of Pediococcus pentosaceus ZRS217 and its application in high-precision yak diets. Specific examples have been used to illustrate the principles and implementation methods of the invention. The descriptions of these embodiments are merely illustrative of the methods and core ideas of the invention. It should be noted that those skilled in the art can make various improvements and modifications to the invention without departing from its principles, and these improvements and modifications also fall within the scope of protection of the claims.

Claims

1. A type of Pediococcus pentosaceus ( Pediococcus pentosaceus ZRS217, characterized in that, The Pediococcus pentosaceus ZRS217 has the accession number CGMCC No.38958 and was deposited at the China General Microbiological Culture Collection Center (CGMCC) on May 29, 2026. The Pediococcus pentosaceus ZRS217 was isolated from naturally fermented yak yogurt from farmers in Nagqu City, Tibet Autonomous Region.

2. The Pediococcus pentosaceus ZRS217 according to claim 1, characterized in that, The Pediococcus pentosaceus ZRS217 has the following characteristics: (a) Tolerance to artificial gastric fluid: The survival rate was 62.02% after treatment at 37°C for 3 hours in artificial gastric fluid at pH 1.5; Tolerance to artificial intestinal fluid: The survival rate was 55.22% after treatment at 37°C for 3 hours in artificial intestinal fluid at pH 6.

8. (b) Bile salt tolerance: The survival rate was 72.92% after treatment at 37°C for 3 h in a medium containing 0.1% bile salts, 55.53% in a medium containing 0.2% bile salts, and 46.98% in a medium containing 0.3% bile salts. (c) It has antioxidant activity, with a DPPH free radical scavenging rate of 53.43% and a hydroxyl free radical scavenging rate of 36.55%.

3. The use of Pediococcus pentosaceus ZRS217 as described in claim 1 in the preparation of a feed additive for improving damage induced by high-precision diets in ruminants.

4. The application according to claim 3, characterized in that, The ruminant animal is the yak.

5. The application according to claim 3, characterized in that, The damage induced by the high-precision diet includes at least one of the following: decreased growth performance, oxidative stress, inflammatory response, intestinal barrier damage, or liver metabolic disorder.

6. The application according to claim 3, characterized in that, The amount of *Pediococcus pentosaceus* ZRS217 added was 1×10⁻⁶. 8 CFU / mL high-precision diet.

7. A feed additive comprising Pediococcus pentosaceus ZRS217 as described in claim 1, with a viable bacterial concentration of not less than 1 × 10⁻⁶. 8 CFU / mL.

8. A high-precision diet for yaks, comprising the feed additive of claim 7.

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