Pediococcus acidilactici, composition and application thereof

Abstract

The invention relates to the field of microorganisms, in particular to pediococcus acidilactici, a composition and application of the pediococcus acidilactici. The pediococcus acidilactici provided by the invention is preserved in the China General Microbiological Culture Collection Center on June 19, 2024, and the preservation number is CGMCC (China General Microbiological Culture Collection Center) NO.31001. The pediococcus acidilactici provided by the invention is obtained by screening from excrement of healthy kittens, has relatively strong acid resistance and bile salt resistance, is extremely strong in intestinal adhesion capacity, has a good inhibition effect on common pathogenic bacteria, is sensitive to various antibiotics, has good probiotic potential, and can be used for preparing a feed additive for the kittens. The invention provides a new candidate strain for pet food, health care and treatment products or probiotic preparations, and has wide application prospects in the aspects of preparation of pet food, health care products, medicines and the like.

Description

Technical Field

[0001] The present invention relates to the technical field of microbiology, and particularly relates to a Pediococcus acidilactici, a composition and their applications. Background Art

[0002] According to statistics, in 2022, the scale of China's pet consumption market exceeded 300 billion yuan, and the consumption of pet foods with scientific proportioning, comprehensive nutrition and health care functions has gradually increased. Probiotics are defined as "living microorganisms that have beneficial effects on the health of the host when consumed in sufficient quantities". In pet foods, lactic acid bacteria are often added as probiotics. With the continuous increase in the number of pet cats in families, cat-specific probiotics have attracted more and more attention, and there is a broad market prospect for the development of cat-specific probiotics. Summary of the Invention

[0003] The purpose of the present invention is to provide a Pediococcus acidilactici, a composition and their applications. The Pediococcus acidilactici has excellent probiotic properties and provides new candidate strains for cat milk replacers, pet foods and health care treatment products.

[0004] To achieve the above invention purposes, the present invention provides the following technical solutions:

[0005] The present invention provides a strain of Pediococcus acidilactici, and the preservation number of the Pediococcus acidilactici is CGMCC NO.31001, which is preserved in the China General Microbiological Culture Collection Center, and the preservation date is June 19, 2024.

[0006] Among them, Pediococcus acidilactici is the Latin name of Pediococcus acidilactici. The Pediococcus acidilactici of the present invention is preserved in the China General Microbiological Culture Collection Center, located at the Institute of Microbiology, Chinese Academy of Sciences, No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, China. In the specification of the present invention, the Pediococcus acidilactici claimed by the present invention is numbered P4, and in some embodiments, P4 is equivalent to the Pediococcus acidilactici claimed by the present invention.

[0007] In some embodiments, the Pediococcus acidilactici is isolated from the feces of kittens.

[0008] Extracting probiotic groups from the feces of kittens has many significant advantages. The strains themselves are derived from the pet intestine, and can better adapt to the host intestinal environment compared with strains from other sources, and can precisely regulate the balance of the intestinal flora of kittens. Such probiotics are tolerant to the gastrointestinal environment, have a close symbiotic relationship with kittens, are stably colonized, and have high safety, no toxic side effects, and rarely cause allergies, which is an excellent choice to promote the healthy growth of puppies.

[0009] In some embodiments, the 16s rRNA gene sequence of the Pediococcus acidilactici is as shown in SEQ ID NO: 01.

[0010] In some embodiments, the Pediococcus acidilactici has an inhibitory effect on bacteria, including Gram-positive bacteria and Gram-negative bacteria, and the bacteria are selected from at least one of Escherichia coli, Staphylococcus aureus, Salmonella, Methicillin-resistant Staphylococcus aureus, Klebsiella pneumoniae; and / or

[0011] The Pediococcus acidilactici is non-hemolytic.

[0012] The Pediococcus acidilactici of the present invention has an inhibitory effect on Gram-positive bacteria and Gram-negative bacteria, is non-hemolytic, has good high safety, and can be used as a probiotic to prepare pet food.

[0013] Furthermore, the inhibitory zone diameter of the Pediococcus acidilactici P4 derived from kittens of the present invention against Klebsiella pneumoniae is ≥ 13 mm; the inhibitory zone diameter of the Pediococcus acidilactici P4 derived from kittens of the present invention against Methicillin-resistant Staphylococcus aureus is ≥ 15 mm; the inhibitory zone diameter of the Pediococcus acidilactici P4 derived from kittens of the present invention against Salmonella is ≥ 20 mm; the inhibitory zone diameter of the Pediococcus acidilactici P4 derived from kittens of the present invention against Escherichia coli is ≥ 24 mm; the inhibitory zone diameter of the Pediococcus acidilactici P4 derived from kittens of the present invention against Staphylococcus aureus is ≥ 26 mm.

[0014] In some embodiments, the Pediococcus acidilactici is sensitive to ampicillin, penicillin, chloramphenicol, cefazolin.

[0015] The present invention is sensitive to a variety of antibiotics, can avoid the problem of antibiotic resistance to a certain extent, prevent the imbalance of intestinal flora, and has a certain safety.

[0016] In some embodiments, the Pediococcus acidilactici has acid tolerance; and / or

[0017] The Pediococcus acidilactici has bile salt tolerance; and / or

[0018] The Pediococcus acidilactici has intestinal adhesion ability.

[0019] The Pediococcus acidilactici of the present invention is acid-resistant, bile salt-resistant and has intestinal adhesion ability, can stably survive and colonize in the host body, can play a probiotic role in the intestine, and regulate gastrointestinal dysfunction caused by the disorder of the intestinal flora structure. Further, the survival rate of P4 is ≥43.62% after surviving for 3 h under the condition of pH 2, ≥79.78% after surviving for 3 h under the condition of pH 2.5, and ≥100.00% after surviving for 3 h under the condition of pH 3. The survival rate of P4 is ≥86.18% after surviving for 3 h under the condition of 0.3% bile salt, ≥98.39% after surviving for 3 h under the condition of 0.1% bile salt, and ≥92.47% after surviving for 3 h under the condition of 0.2% bile salt. P4 has extremely strong cell adhesion ability, and the adhesion rate reaches 65.51%, with good probiotic potential. Adhesion and colonization in the intestine are important prerequisites for lactic acid bacteria to exert their ecological effects and physiological functions, and are one of the important bases for evaluating their probiotic potential. The adhesion of lactic acid bacteria to epithelial cells is the main basis for their probiotic functions. The adhered lactic acid bacteria can exist in the intestine and resist the invasion of pathogenic bacteria, produce metabolites such as antibacterial peptides to help effectively kill pathogenic bacteria, achieve functions such as preventing the intestine from being infected by pathogenic bacteria, and enhance the immune capacity of the body.

[0020] In some embodiments, the Pediococcus acidilactici has antioxidant properties.

[0021] The Pediococcus acidilactici of the present invention has certain antioxidant properties, among which the DPPH free radical scavenging rate is greater than 37%, and the ABTS free radical scavenging rate is greater than 44%.

[0022] In summary, the Pediococcus acidilactici provided by the present invention has acid production ability and adhesion ability, regulates the gastrointestinal flora, and maintains the balance of the intestinal microecology. It has an antagonistic effect on pathogenic microorganisms in animals, can competitively inhibit pathogenic microorganisms, enhance the immune function of the animal body, produce beneficial metabolites, activate the activity of acid protease, participate in the metabolism of the body, and prevent the generation of harmful substances. At the same time, the Pediococcus acidilactici of the present invention is derived from the feces of puppies, has various physiological functions such as regulating the gastrointestinal flora, enhancing the immune function, and anti-inflammatory effect. Isolating and screening excellent Pediococcus acidilactici from kittens has important scientific significance and practical application value, can meet the market demand, promote the development of the probiotic industry, promote scientific research, improve the health level of animals, and promote the development of related application fields.

[0023] On the basis of the above technical solutions, the present invention also provides a composition, and the composition contains the above-mentioned Pediococcus acidilactici or its metabolites.

[0024] In some embodiments, the composition is a probiotic preparation, a functional food, a health product or a medicine.

[0025] Among them, pet foods include but are not limited to: yogurt, milk powder, cheese, snacks and / or freeze-dried staple foods or pet dry foods, etc.

[0026] In addition, the present invention also provides the use of the aforementioned Pediococcus acidilactici or the above composition in the preparation of a drug or food for treating, preventing or alleviating gastrointestinal dysfunction.

[0027] Compared with the prior art, the beneficial effects of the present invention are as follows:

[0028] (1) The present invention provides a new strain of Pediococcus acidilactici P4 derived from young cats, which has strong adhesion ability and good probiotic ability.

[0029] (2) The Pediococcus acidilactici P4 derived from young cats provided by the present invention has no hemolytic property and high biosafety.

[0030] (3) The Pediococcus acidilactici P4 derived from young cats provided by the present invention has good acid tolerance and bile salt tolerance.

[0031] (4) The Pediococcus acidilactici P4 derived from young cats provided by the present invention is a strain with excellent probiotic properties selected after isolation, screening and identification from cat feces. It has a good inhibitory effect on common pathogenic bacteria and is sensitive to a variety of antibiotics, providing a new candidate strain for pet foods, health care treatment products or probiotic preparations, and having broad application prospects in the preparation of pet foods, health products, drugs, etc. Description of the Drawings

[0032] In order to more clearly illustrate the specific embodiments of the present invention or the technical solutions in the prior art, the following will briefly introduce the drawings required to be used in the description of the specific embodiments or the prior art. Obviously, the following drawings are some embodiments of the present invention. For those of ordinary skill in the art, other drawings can be obtained based on these drawings without creative efforts.

[0033] Figure 1 It is the acid tolerance test result at pH = 2 in Example 1.

[0034] Figure 2 It is the bile salt tolerance test result at a bile salt concentration of 0.3% in Example 1.

[0035] Figure 3 It is the antibacterial ability result of Pediococcus acidilactici P4 derived from young cats in Example 1.

[0036] Figure 4 It is the BLAST database annotation result in Example 2.

[0037] Figure 5It is the colony morphology of Pediococcus acidilactici P4 from young kittens in Example 2.

[0038] Figure 6 It is the growth curve of Pediococcus acidilactici P4 from young kittens in Example 3.

[0039] Figure 7 It is the result of the in vitro acid tolerance ability of Pediococcus acidilactici P4 from young kittens in Example 3.

[0040] Figure 8 It is the result of the in vitro bile salt tolerance ability of Pediococcus acidilactici P4 from young kittens in Example 3.

[0041] Figure 9 It is the result of the hemolytic ability of Pediococcus acidilactici P4 from young kittens in Example 3.

[0042] Figure 10 It is the result of the adhesion experiment of Pediococcus acidilactici P4 from young kittens in Example 3.

[0043] Figure 11 It is the result of the DPPH free radical scavenging test of Pediococcus acidilactici P4 from young kittens in Example 3.

[0044] Figure 12 It is the result of the ABTS free radical scavenging test of Pediococcus acidilactici P4 from young kittens in Example 3. Detailed implementation mode

[0045] The following examples facilitate a better understanding of the present invention, but do not limit the present invention. The experimental methods in the following examples are all conventional methods unless otherwise specified. The test materials used in the following examples are all purchased from conventional biochemical reagent companies unless otherwise specified. In the following examples, all quantitative tests are set with three repeated experiments, and the results are averaged.

[0046] Example 1: Screening and identification of Pediococcus acidilactici P4 from young kittens

[0047] 1. Cultivation and isolation of strains

[0048] Select fresh young kitten fecal samples to screen strains. After gradient dilution with sterilized PBS buffer (purchased from Beijing Saiweier Biotechnology Co., Ltd.), coat them on MRS solid medium containing 1% CaCO3, and culture them in a 37°C anaerobic incubator for 48 hours. Pick white or milky white single colonies with obvious transparent circles for Gram staining and microscopic examination. Select strains that are rod-shaped and Gram-positive bacteria, and select the relatively well-growing Pediococcus acidilactici strains P3, P4, P7, and P8 from numerous strains for enrichment culture in a 37°C anaerobic incubator; mix the bacterial cells with sterilized 50% glycerol and store them in a -80°C refrigerator for later use.

[0049] 2. Screening of strains

[0050] Under aseptic conditions, inoculate the strains of P3, P4, P7, and P8 after glycerol preservation into MRS broth medium, and culture them anaerobically at 37°C for 24 h, and activate them continuously for three generations; after centrifuging the bacterial solution, wash the medium with sterilized PBS 2-3 times;

[0051] Conduct acid resistance, bile salt resistance, and antibacterial tests on the strains of P3, P4, P7, and P8.

[0052] The test steps for acid resistance, bile salt resistance, and adhesion are as follows:

[0053] (1) Acid resistance test

[0054] Resuspend the washed medium in MRS broth medium with a pH of 2, culture it anaerobically at 37°C, take samples for gradient dilution at 0 h and 3 h respectively, coat them on the MRS solid medium plate, and count after anaerobic culture at 37°C for 48 h.

[0055] Survival rate of the strain (%) = lg(N1 / N0) * 100%;

[0056] Where: N0 is the viable count at 0 h (cfu / mL); N1 is the viable count of the strain after 3 h of acid resistance (cfu / mL).

[0057] The experimental results are as Figure 1 shown. The survival rate of strain P4 is the highest, up to 43.62%.

[0058] (2) Bile salt resistance test

[0059] Resuspend the washed medium in MRS broth medium containing 0.3% bile salt and culture it, culture it anaerobically at 37°C, take samples for gradient dilution at 0 h and 3 h respectively, coat them on the MRS solid medium plate, and count after anaerobic culture at 37°C for 48 h.

[0060] Survival rate of the strain (%) = lg(N1 / N0) * 100%;

[0061] Where: N0 is the viable count at 0 h (cfu / mL); N1 is the viable count of the strain after 3 h of bile salt resistance (cfu / mL).

[0062] The experimental results are as Figure 2 shown. The survival rate of strain P4 is 86.18%.

[0063] (3) Determination of the antibacterial ability of Pediococcus acidilactici P4 from kittens

[0064] Pet food contains a large amount of nutrients. In daily life, food is accompanied by spoilage and deterioration, which not only endangers the health of pets but also causes economic losses. The contamination, growth, and reproduction of microorganisms are the main reasons for food spoilage and deterioration. Therefore, the antibacterial performance of P4 is one of the safety indicators for its application in food.

[0065] Using five common foodborne pathogenic bacteria, namely Escherichia coli, Staphylococcus aureus, Salmonella, Methicillin-resistant Staphylococcus aureus, and Klebsiella pneumoniae, as indicator bacteria. After activating the indicator bacteria for three generations, the Oxford cup method was used for the preliminary determination of antibacterial activity.

[0066] Using the Oxford cup method, the specific method is as follows: Take the activated strain and inoculate it into MRS broth medium at an inoculation amount of 2% (V / V), and culture it anaerobically at 37°C for 24 h. Perform 10-fold serial dilutions on the activated Escherichia coli, Staphylococcus aureus, and Salmonella suspensions, and select the bacterial suspension with a dilution of 10 -2 for use. Add 5 mL of LB agar medium to the culture dish for bottom sealing. After cooling and solidifying, place Oxford cups evenly. Take 100 μL of each gradient indicator bacteria suspension and add it to 15 mL of LB agar at 50 - 60°C, mix well, and slowly pour it into the plate with Oxford cups. After condensation, use forceps to pull out the Oxford cups to form round holes with a size of 8×6×10. Add 200 μL of the strain fermentation supernatant to the round holes, diffuse it in a 4°C refrigerator for 12 h, and then culture it anaerobically at 37°C for 9 - 12 h. Observe whether there is an antibacterial zone, and measure the diameter of the antibacterial zone with a vernier caliper. Each group of experiments is repeated three times in parallel, and the results are expressed as x±s.

[0067] The experimental results are as Figure 3 shown. Bacterium P4 has strong inhibitory effects on both Gram-positive bacteria (Staphylococcus aureus, Methicillin-resistant Staphylococcus aureus) and Gram-negative bacteria (Escherichia coli, Salmonella, Klebsiella pneumoniae), and the diameter of the antibacterial zone is above 13 mm. Among them, the inhibitory effect on Staphylococcus aureus is the strongest, and the diameter of the antibacterial zone of the supernatant is 26 mm.

[0068] From Figure 1 , Figure 2 and Figure 3 the results, it can be seen that Pediococcus acidilactici P4 from kittens has extremely strong antibacterial ability, up to 26 mm, which is much higher than that of other Pediococcus acidilactici from kittens; and after surviving for 3 h under the acid-resistant condition of pH = 2 and the bile salt-resistant condition of 0.3% bile salt concentration, the survival rates reach 43.62% and 86.18% respectively, indicating good acid resistance and bile salt resistance.

[0069] Example 2: Identification of Pediococcus acidilactici P4 from kittens

[0070]

[0071] The sequencing results were used for homology search in the gene bank by applying BLAST on the NCBI website. The results showed that the highest similarity was 99.93% with Pediococcus acidilactici. The results are as Figure 4 shown, and 99.93% of the genes corresponded to the genus Pediococcus acidilactici.

[0072] The strain P4 was diluted and spread on the plate to observe the colony state. As Figure 5 shown, combining with the morphological characteristics of the P4 colonies, the strain P4 was identified as Pediococcus acidilactici. In addition, P3, P7, and P8 belong to the same species of Pediococcus acidilactici.

[0073] Example 3: Determination of the probiotic properties of Pediococcus acidilactici P4 from kittens

[0074] 1. Determination of the growth curve of Pediococcus acidilactici P4 from kittens

[0075] The cultured P4 bacterial solution was activated for three generations and inoculated into MRS broth medium, placed in an incubator at 37°C, and anaerobically cultured. The viable counts of each bacterium were measured at 0, 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, and 24 h respectively;

[0076] The growth curve of Pediococcus acidilactici P4 from kittens was plotted. The results are shown in Figure 6 the figure.

[0077] 2. Determination of the acid and bile salt tolerance of Pediococcus acidilactici P4 from kittens in vitro

[0078] The cultured P4 bacterial solution was activated for three generations and inoculated into MRS broth medium with pH values of 2.5, 3 and bile salt concentrations of 0.1%, 0.2% respectively. It was placed in an incubator at 37°C and anaerobically cultured. Samples were taken for plate counting at 0 h and 3 h respectively;

[0079] The experimental results are shown in Figure 7 and Figure 8 respectively; As can be seen from Figure 7 , when the bacteria were cultured in an environment with pH 2.5 and pH 3 for 0 - 3 h, the growth of the bacteria was stable, and the viable counts were all above 10 8 cfu / mL, indicating strong acid tolerance.

[0080] As can be seen from Figure 8 , the bacteria grew relatively stably in the medium with a bile salt concentration of 0.1%; after culturing in the medium with a bile salt content of 0.2% for 3 h, the survival rate decreased by 6%, indicating strong bile salt tolerance.

[0081] 3. Determination of the antibiotic sensitivity of Pediococcus acidilactici P4 from kittens

[0082] The excessive use of antibiotics in the pharmaceutical industry can lead to an enhanced drug resistance of pathogenic strains, and the flora in the human intestine is prone to disorder and imbalance. Therefore, obtaining lactic acid bacteria sensitive to antibiotics has become one of the important indicators for their safety in food applications.

[0083] Adjust the concentrations of the fermentation broths of each strain cultured for 24 h to 10 8 CFU / mL respectively. Add 100 μL of the bacterial liquid to the MRS solid plate, spread it evenly, and let it dry. Place 5 - 6 drug sensitivity discs in each plate on average. After diffusing in a 4°C refrigerator for 12 h, culture anaerobically at 37°C for 9 - 12 h. The sensitivity of the strain to the drug is judged with reference to the instructions for using the drug sensitivity paper disc.

[0084] The results are shown in Table 1. As can be seen from Table 1, Pediococcus acidilactici P4 is extremely sensitive to ampicillin, penicillin, chloramphenicol, and cefazolin (cephalosporins), and is tolerant to gentamicin, ciprofloxacin, and norfloxacin.

[0085] Table 1 Antibiotic sensitivity test

[0086] Antibiotic Name Gentamicin Ampicillin Penicillin Ciprofloxacin Chloramphenicol Cefazolin Norfloxacin Sensitivity R S S R S S R

[0087] S: susceptible, sensitive; I: intermediate, moderately sensitive; R: resistance, tolerant

[0088] 4. Determination of the hemolytic ability of Pediococcus acidilactici P4 from young cats

[0089] Dip a inoculation loop into a small amount of bacterial liquid and streak it on a Columbia blood agar plate. Culture anaerobically at 37°C for 24 - 36 h, and observe whether there is a hemolytic zone on the plate. According to the hemolytic phenomenon, it can be divided into α-hemolysis (greenish hemolysis), β-hemolysis (completely transparent hemolytic ring), and γ-hemolysis (non-hemolysis).

[0090] The hemolysis experiment can detect the safety of the strain and determine whether it has hemolytic activity. The results are as Figure 9 shown. This strain did not show any hemolytic phenomenon on the blood plate, showing γ-hemolysis. Staphylococcus aureus was selected as the positive control, and a completely transparent hemolytic ring appeared around Staphylococcus aureus, showing β-hemolysis. The above results indicate that this strain of Pediococcus acidilactici has no hemolytic activity and is a safe and non-toxic strain.

[0091] 5. Adhesion test

[0092] After resuscitating Caco-2 cells (human colon cancer cells), inoculate them into a cell culture flask and add fresh DMEM complete medium (DMEM high-glucose medium containing double antibodies purchased from Beijing Solarbio Science & Technology Co., Ltd., 20% fetal bovine serum). Culture them in an incubator at 37°C and 5% CO2, change the cell culture medium every two days. When the cell adherent growth fusion state reaches about 70%-80%, use 0.25% trypsin-EDTA (purchased from Beijing Solarbio Science & Technology Co., Ltd.) mixed digestive solution for passage. The last time, use DMEM incomplete medium.

[0093] Preparation of FITC solution: FITC (fluorescein isothiocyanate) labeling solution (purchased from Beijing Solarbio Science & Technology Co., Ltd.): Before preparation, place the FITC stored at 4°C at room temperature for 30 min. Accurately weigh 0.5 g and dissolve it in 1 mL of dimethyl sulfoxide (DMSO) to prepare a solution of 500 mg / mL. Dilute it 1000 times (take 10 μL and add it to a 9.99 mL DMSO brown 15 mL EP tube) to obtain a labeling solution with a working concentration of 500 μg / mL, and store it at 4°C.

[0094] Fluorescent labeling: Take 1 mL of freshly cultured probiotic bacterial suspension and centrifuge it in a brown 1.5 mL EP tube (4°C, 4000 rpm, 5 min). Wash it continuously 3 times with sterile PBS. After the last wash, resuspend the bacteria with 1 mL of FITC solution that has been placed at room temperature for 30 min in advance, and incubate it with shaking in the dark at 37°C for 2 h. After the dark incubation, centrifuge it (4°C, 4000 rpm, 5 min) and wash the strain 3 times with sterile PBS to wash away the unbound FITC labeling solution, and resuspend it with 1 mL of DMEM cell culture medium. Take 200 μL and add it to a black 96-well plate to measure the relative fluorescence intensity value (RFU) of the strain under the conditions of wavelength 485 nm (absorption wavelength) and wavelength 530 nm (emission wavelength), which is recorded as the relative fluorescence intensity value R0 of the strain before adhesion.

[0095] Adhesion test: Passage Caco-2 cells for more than 3 generations continuously and prepare for the experiment.

[0096] Before the experiment, adjust the cell concentration to 5×10 5cells / mL. 1 mL of cell suspension was evenly spread in each well of a 24-well cell culture plate and cultured further. After observing that the cells in the 24-well plate grew to a monolayer (about 24 h of culture), the cell culture medium was discarded. After washing three times with sterile PBS, 0.6 mL of the completed-labeled probiotic suspension was added. Three replicate wells were set for each strain, and they were incubated at 37 °C and 5% CO2 in the dark for 2 h. After taking them out, the culture medium was discarded, and the strains that did not adhere to the cells were removed by washing 3 times with sterile PBS; 0.3 mL of trypsin was added to each well to digest the cells for 5 min, and the digestion was terminated with DMEM cell culture medium. The relative fluorescence intensity value (RFU) of the cell suspension was measured under the same wavelength conditions and recorded as the relative fluorescence intensity value R of the adhered strain.

[0097] Adhesion rate (%) calculation formula: R / R0×100%

[0098] CFU (colony-forming unit): The total number of bacterial communities in a unit volume; when counting viable bacteria in culture, a colony formed by the growth and reproduction of a single bacterial cell or a group of aggregated bacterial cells on a solid medium is called a colony-forming unit, and it is used to express the number of viable bacteria.

[0099] The experimental results are shown in Figure 10 as shown, the adhesion rate of strain P4 was as high as 65.51%.

[0100] 6. DPPH free radical scavenging assay

[0101] The bacterial cells were collected and the bacterial suspension concentration was adjusted to 10 -8 , and a DPPH free radical scavenging ability kit (purchased from Beijing Solarbio Science & Technology Co., Ltd.) was used. The specific operation was carried out according to the steps in the instruction manual.

[0102]

[0103] DPPH free radical scavenging rate of the sample (%) = (1 - (A measurement - A control) / A blank)×100%

[0104] The experimental results are shown in Figure 11 , and the DPPH free radical scavenging rate of the bacterial suspension of strain P4 was 37.67%, showing certain antioxidant properties.

[0105] 7. ABTS free radical scavenging assay

[0106] The bacterial cells were collected and the bacterial suspension concentration was adjusted to 10 -8 , and an ABTS free radical scavenging ability kit (Beijing Solarbio Science & Technology Co., Ltd.) was used. The specific operation was carried out according to the steps in the instruction manual.

[0107]

[0108]

[0109] Sample ABTS radical scavenging rate (%) = (1 - (A measured - A control) / A blank) × 100%

[0110] The experimental results are shown in Figure 12 , the ABTS radical scavenging rate of the supernatant of strain P4 was 86.89%, indicating certain antioxidant activity.

[0111] Finally, it should be noted that the above are only the preferred embodiments of the present invention and are not used to limit the present invention. Although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art can still modify the technical solutions described in the foregoing embodiments or perform equivalent replacements for some of the technical features. Any modifications, equivalent replacements, improvements, etc. made within the spirit and principle of the present invention shall be included within the protection scope of the present invention.

Claims

1. A strain of Pediococcus acidilactici, characterized in that, The preservation number of the Pediococcus acidilactici is CGMCC NO. 31001, which is preserved in the China General Microbiological Culture Collection Center, and the preservation date is June 19, 2024.

2. The Pediococcus acidilactici according to claim 1, characterized in that, The Pediococcus acidilactici is isolated from the feces of kittens.

3. The Pediococcus acidilactici according to claim 1, characterized in that, The 16s rRNA gene sequence of the Pediococcus acidilactici is as shown in SEQ ID NO:

01.

4. The Pediococcus acidilactici according to claim 1, characterized in that, The Pediococcus acidilactici has an inhibitory effect on bacteria, and the bacteria include Gram-positive bacteria and Gram-negative bacteria, and the bacteria are selected from at least one of Escherichia coli, Staphylococcus aureus, Salmonella, Methicillin-resistant Staphylococcus aureus, Klebsiella pneumoniae; and / or The Pediococcus acidilactici has no hemolytic property.

5. The Pediococcus acidilactici according to claim 1, characterized in that, The Pediococcus acidilactici is sensitive to ampicillin, penicillin, chloramphenicol, and cefazolin.

6. The Pediococcus acidilactici according to claim 1, characterized in that, The Pediococcus acidilactici has acid tolerance ability; and / or The Pediococcus acidilactici has bile salt tolerance ability; and / or The Pediococcus acidilactici has intestinal adhesion ability.

7. The Pediococcus acidilactici according to claim 1, characterized in that, The Pediococcus acidilactici has antioxidant property.

8. A composition, characterized in that, The composition comprises the Pediococcus acidilactici or its metabolite according to any one of claims 1 to 7.

9. The composition according to claim 8, wherein The composition is a probiotic preparation, a functional food, a health product or a medicine.

10. Use of the Pediococcus acidilactici according to any one of claims 1 to 7 or the composition according to claim 8 or 9 in the preparation of a drug or food for treating, preventing or alleviating gastrointestinal dysfunction.

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