Lactobacillus plantarum D27 for antagonizing vibrio cholerae and application of lactobacillus plantarum D27

Abstract

The invention discloses lactobacillus plantarum D27 for antagonizing vibrio cholerae and application thereof, and belongs to the technical field of microorganisms, the lactobacillus plantarum D27 is preserved in China Center for Type Culture Collection (CCTCC) on September 4, 2024, the preservation address is No. 299 on eight road, Wuchang District, Wuhan City, Hubei Province, and the preservation number is CCTCC M 20241910. The lactobacillus plantarum D27 disclosed by the invention can effectively antagonize vibrio cholerae, inhibit the formation of a biofilm of the vibrio cholerae and destroy a cell structure of the vibrio cholerae, also has the characteristics of high yield of lactic acid and exopolysaccharide and good self-aggregation and co-aggregation, and can be used for preparing foods or medicines for antagonizing vibrio cholerae pollution; comprising lactobacillus plantarum liquid, lactobacillus plantarum powder, lactobacillus plantarum capsules, lactobacillus plantarum tablets and the like.

Description

Technical Field

[0001] The invention relates to the technical field of microorganisms, and in particular to a lactobacillus plantarum D27 capable of antagonizing Vibrio cholerae and an application thereof. Background Art

[0002] Vibrio cholerae is the pathogen that causes the infectious disease "cholera" of type A. Studies have shown that the bacteria usually contaminate water or food, spread from environmental hosts to human hosts, and cause "cholera" diseases that endanger human life. Clinical symptoms mainly include gastric ulcers, acute diarrhea, severe vomiting, severe dehydration, and even death. According to a report by the World Health Organization, "cholera" causes approximately 1.3 to 4 million cases of infection and 21,000 to 143,000 deaths each year. Since 2021, the "cholera" epidemic has surged, and the mortality rate has hit the highest record in nearly a decade.

[0003] At present, antibiotics are mainly used for treatment, which can effectively shorten the duration of diarrhea and the severity of infection to control the spread of "cholera". Doxycycline, azithromycin, ciprofloxacin and other antibiotics are used clinically for treatment. However, long-term and excessive use of antibiotics can lead to the production of drug-resistant Vibrio cholerae. Studies have shown that most epidemic strains of Vibrio cholerae are multidrug-resistant, leading to higher mortality rates, more secondary infections, and higher medical costs. In addition, the long-term and excessive use of antibiotics in aquaculture also creates food safety risks and environmental pollution problems.

[0004] Lactic acid bacteria are food-grade probiotics and are widely used in the food industry. Several literature reports have been published on the research on lactic acid bacteria producing metabolites such as organic acids, lactic acid and bacteriocins to inhibit the growth of Vibrio cholerae. Compared with the reported antagonistic bacteria Lactobacillus brevis, Lactobacillus rhamnosus and Lactobacillus casei, Lactobacillus plantarum has a better tolerance to high acid and high bile salts in the digestive tract. In addition, Lactobacillus plantarum also has multiple probiotic functions such as regulating the host gastrointestinal flora, inhibiting pathogens, and enhancing immunity. Summary of the invention

[0005] The main purpose of the present invention is to provide a Lactobacillus plantarum D27 antagonistic to Vibrio cholerae, which can effectively antagonize Vibrio cholerae, inhibit its biofilm formation, destroy its cell structure, has high production of lactic acid and extracellular polysaccharides, and good self-aggregation and co-aggregation properties.

[0006] Another object of the present invention is to provide the application of Lactobacillus plantarum D27 for antagonizing Vibrio cholerae.

[0007] To achieve the above object, the present invention adopts the following technical solution:

[0008] The invention provides a Lactobacillus plantarum D27 for antagonizing Vibrio cholerae. The Lactobacillus plantarum D27 is deposited in China Center for Type Culture Collection (CCTCC), the deposit address is No. 299, Bayi Road, Wuchang District, Wuhan City, Hubei Province (on the campus of Wuhan University), the deposit date is September 4, 2024, the deposit number is CCTCC M 20241910, and the classification name is Lactobacillus plantarum.

[0009]

[0010] Preferably, the Lactobacillus plantarum D27 can effectively antagonize Vibrio cholerae, inhibit its biofilm formation and destroy its cell structure.

[0011] Preferably, the Lactobacillus plantarum D27 has high production of lactic acid and extracellular polysaccharides, as well as self-aggregation and co-aggregation properties.

[0012] The present invention also provides a Lactobacillus plantarum agent for antagonizing Vibrio cholerae, which comprises the Lactobacillus plantarum D27 for antagonizing Vibrio cholerae.

[0013] Preferably, the Lactobacillus plantarum agent is selected from one or more of Lactobacillus plantarum liquid, Lactobacillus plantarum powder, Lactobacillus plantarum capsules or Lactobacillus plantarum tablets.

[0014] The present invention also provides use of any of the above-mentioned Lactobacillus plantarum D27 antagonizing Vibrio cholerae, or the above-mentioned Lactobacillus plantarum bacterial agent antagonizing Vibrio cholerae in the preparation of food or medicine antagonizing Vibrio cholerae contamination.

[0015] Preferably, the types of food include ready-to-eat, fresh, refrigerated and frozen.

[0016] Preferably, the dosage form of the drug includes liquid, powder, tablet and capsule.

[0017] Compared with the prior art, the present invention has the following beneficial effects:

[0018] (1) The plant lactobacillus D27 of the present invention can effectively antagonize Vibrio cholerae, inhibit its biofilm formation, destroy its cell structure, and has high production of lactic acid and extracellular polysaccharides as well as good self-aggregation and co-aggregation properties.

[0019] (2) The plant lactobacillus D27 of the present invention can be used for the preparation of food or medicine, and the prepared food or medicine also has efficacy. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] Figure 1 The experimental results of LpD27-CFS inhibiting the biofilm formation of Vibrio cholerae in the examples; AD: Vibrio cholerae B5-82, b10-49, 7-6-19 and GIM1.449 strains, *p<0.05, **p<0.01, ***p<0.001.

[0021] Figure 2 These are the experimental results of LpD27-CFS in the example that reduces CSH and increases CMF of Vibrio cholerae; A: CSH; B: CMF; *p<0.05, **p<0.01, ***p<0.001.

[0022] Figure 3 The experimental results of LpD27-CFS in the example show that it increases the IMP of Vibrio cholerae; AD: Vibrio cholerae B5-82, b10-49, 7-6-19, and GIM 1.449 strains, respectively.

[0023] Figure 4 This is the scanning electron microscopy result of LpD27-CFS destroying the cell surface structure of Vibrio cholerae in the example. DETAILED DESCRIPTION

[0024] The technical scheme of the present invention is further described below by way of examples. Obviously, the described embodiments are only a part of the embodiments of the present invention, rather than all of the embodiments. Based on the embodiments of the present invention, all other implementation methods obtained by those skilled in the art without creative work are within the scope of protection of the present invention.

[0025] The present invention proposes a Lactobacillus plantarum D27 for antagonizing Vibrio cholerae. The Lactobacillus plantarum D27 is a lactic acid bacterium that can effectively antagonize Vibrio cholerae, inhibit its biofilm formation, and destroy its cell structure. The Lactobacillus plantarum D27 has high lactic acid and extracellular polysaccharide production, as well as good self-aggregation and co-aggregation properties. The 16S rRNA sequence is shown in SEQ ID NO: 1. The Lactobacillus plantarum D27 has been preserved by the China Center for Type Culture Collection (CCTCC) on September 4, 2024, with a preservation address of No. 299, Bayi Road, Wuchang District, Wuhan City, Hubei Province (on the campus of Wuhan University), a preservation number of CCTCC M 20241910, a classification name of Lactobacillus plantarum, and a survival state of survival.

[0026] Embodiment 1

[0027] Experiment on Antagonism of Lactobacillus plantarum D27 against Vibrio cholerae

[0028] (I) Antibacterial activity test of Lactobacillus plantarum D27

[0029] Preparation of cell-free supernatant (LpD27-CFS) of Lactobacillus plantarum D27: Lactobacillus plantarum D27 was inoculated into sterilized De Man Rogosa and Sharp (MRS) liquid medium, activated by passage twice at a volume fraction of 1%, cultured at 37° C. for 48 h, centrifuged at 4° C. and 4000 r / min for 10 min, and the supernatant was collected and filtered through a 0.22 μm sterile microporous filter membrane for standby use.

[0030] The tested Vibrio cholerae GIM1.449 standard strain was purchased from Guangdong Institute of Microbiology, and the Vibrio cholerae b10-49, 7-6-19 and B5-82 strains were self-preserved strains. The Vibrio cholerae strains were inoculated into sterilized Tryptic Soy Broth (TSB) liquid medium, subcultured twice at a volume fraction of 1%, and then cultured at 37°C to the mid-logarithmic growth phase (mid-LGP). Then, the OD 600 The absorbance value is 0.08-0.13, set aside.

[0031] Take the above Vibrio cholerae suspension and evenly spread it on the sterilized Mueller-Hinton Agar (MHA) agar plate (200 μL / plate). After the bacterial solution is absorbed, place a sterilized Oxford cup (7.8×6.0×10.0 mm) evenly on it. Take 150 μL LpD27-CFS and add it to each Oxford cup. Use MRS liquid culture medium as the negative control group and antibiotic streptomycin solution (100 μg / mL) as the positive control group. Place the MHA agar plate horizontally in a 37°C constant temperature incubator and culture for 16 to 18 hours. Observe the size of the inhibition zone and measure the diameter of the inhibition zone (Diameter of Inhibition Zone, DIZ). Repeat each group of experiments three times.

[0032] As shown in Table 1, the DIZ value of LpD27-CFS for inhibiting Vibrio cholerae 7-6-19 was 20.69±0.29; the DIZ value for inhibiting Vibrio cholerae GIM1.449 was 18.87±0.29; the DIZ value for inhibiting Vibrio cholerae B5-82 was 18.76±0.58; the DIZ value for inhibiting Vibrio cholerae b10-49 was 16.70±1.15. There was no inhibition zone in the negative control, and the DIZ value of the positive control was 23.03±0.17. The results showed that LpD27-CFS could inhibit Vibrio cholerae and had a similar effect as the positive control antibiotic.

[0033] Table 1: Experimental results of LpD27-CFS inhibiting Vibrio cholerae

[0034] Tested strains DIZ(mm) Ki(%) Vibrio cholerae GIM1.449 18.87±0.29 97.17±1.95 Vibrio cholerae B5-82 18.76±0.58 80.22±1.37 Vibrio cholerae b10-49 16.70±1.15 94.84±0.17 Vibrio cholerae 7-6-19 20.69±0.29 97.57±9.19

[0035] (II) Bactericidal index test of Lactobacillus plantarum D27

[0036] The Vibrio cholerae strain cultured to mid-LGP was collected by centrifugation, washed with sterile 1× PBS buffer (pH 7.2-7.4) and resuspended. A Vibrio cholerae suspension (80 μL / well, 1×10 6CFU / mL), then add LpD27-CFS (20 μL / well), mix thoroughly, and incubate at 37°C for 3 h. Then, add TSB liquid medium (100 μL / well) and continue to culture at 37°C for 21 h. Set up a positive control group and mix 1×PBS buffer solution (20 μL / well) with Vibrio cholerae suspension (80 μL / well, 1×10 6 CFU / mL). Two blank control groups were set up: the blank control group of LpD27-CFS, LpD27-CFS (20 μL / well) was mixed with TSB liquid culture medium (80 μL / well); the blank control group of 1×PBS, 1×PBS buffer (20 μL / well) was mixed with TSB liquid culture medium (80 μL / well). The OD of each well was measured using a multifunctional microplate reader. 600 The absorbance value was used to calculate the killing index (Ki) according to the following formula: Ki% = [1-corrLpD27 / corrAB] × 100%; where CorrLpD27 = OD 600 (LpD27-CFS + Vibrio cholerae suspension) - LpD27-CFS blank control value; corrAB = OD 600 (1×PBS buffer + Vibrio cholerae suspension) - 1×PBS blank control value, each group of experiments was repeated three times.

[0037] As can be seen from Table 1, the Ki value of LpD27-CFS against Vibrio cholerae 7-6-19 is 97.57±9.19%; the Ki value against Vibrio cholerae GIM1.449 is 97.17±1.95%; the Ki value against Vibrio cholerae b10-49 is 94.84±0.17%; the Ki value against Vibrio cholerae B5-82 is 80.22±1.37%, indicating that LpD27-CFS can effectively kill Vibrio cholerae strains.

[0038] (III) Experiment on inhibition of biofilm formation of Vibrio cholerae by Lactobacillus plantarum D27

[0039] Referring to the above method, different concentrations of LpD27-CFS (0%, 5%, 10%, 20%) were used to treat Vibrio cholerae strains at 37°C for 3 h. Then, the OD of the bacterial solution was adjusted with TSB liquid medium. 600 The absorbance value was 0.4, and it was inoculated into a 24-well sterilized bacterial culture plate (1 mL / well) and cultured at 37°C for 12 h, 24 h, 36 h, 48 h, and 60 h. The floating bacteria in each well were removed by suction, washed three times with 1× PBS buffer (1 mL / well), stained with 0.25% crystal violet at room temperature for 30 min, and then washed three times with 1× PBS buffer. After drying at room temperature for 10 min, 95% ethanol (2 mL / well) was added, and the OD of each well was measured using a multifunctional microplate reader.600 Absorbance values, each test was repeated three times.

[0040] like Figure 1 As shown in the control group with 0% LpD27-CFS concentration, the biofilm of Vibrio cholerae B5-82 formed slowly from 0 to 24 h, increased rapidly from 24 to 48 h, and reached the maximum value (OD 600 =1.09±0.01); decreased sharply at 48-60h (OD 600 =0.39±0.03). However, after being treated with LpD27-CFS (5-20%) for 3h, the biofilm of Vibrio cholerae B5-82 was significantly reduced in the formation, maturation and diffusion stages (p<0.05), and showed a reduction pattern that was dependent on the concentration of LpD27-CFS. Similarly, compared with the control group with a LpD27-CFS concentration of 0%, the maximum biomass (48h) of biofilm formation of Vibrio cholerae b10-49, 7-6-19 and GIM 1.449 strains was significantly reduced (0.10-0.88 times, p<0.05) after being treated with 5-20% LpD27-CFS for 3h. The results showed that Lactobacillus plantarum D27 can effectively reduce the formation of Vibrio cholerae biofilm.

[0041] (IV) Experiment on increasing the cell surface hydrophobicity (CSH) of Vibrio cholerae by Lactobacillus plantarum D27

[0042] The suspension of Vibrio cholerae (1 mL, mid-LGP, OD 600 =0.55-0.60), 1 mL of 98% hexadecane was added respectively, the mixture was shaken for 1 min, and the mixture was allowed to stand at room temperature for 30 min. The OD 600 Absorbance values, each test was repeated three times.

[0043] like Figure 2 A shows that, compared with the control group with a LpD27-CFS concentration of 0%, the CSH of the tested Vibrio cholerae strains treated with 5-20% LpD27-CFS for 3 hours significantly decreased (p<0.05), and showed a decrease in a LpD27-CFS concentration-dependent manner. Among them, the CSH of Vibrio cholerae b10-49 and 7-6-19 strains decreased sharply after being treated with 5% LpD27-CFS for 3 hours (p<0.01).

[0044] (VI) Experiment on increasing cell membrane fluidity (CMF) of Vibrio cholerae by Lactobacillus plantarum D27

[0045] 10 mM 1,6-diphenyl-1,3,5-hexatrine (DPH) solution (2.0 μL / well) was added to the mid-LGP Vibrio cholerae suspension (200 μL / well) treated with LpD27-CFS at different concentrations (0%, 5%, 10%, 20%), and the horizontal polarization (Ivh) and vertical polarization (Ivv) fluorescence of each well were detected by a multifunctional microplate reader, 360 / 40 excitation filter, 460 / 40 detection filter and 400 nm dichroic mirror, and calculated according to the following formula: rDPH = (Ivv-G×Ivh) / (Ivv+2×G×Ivh), where G = 0.85. Each group of experiments was repeated three times.

[0046] like Figure 2 B shows that compared with the control group with a LpD27-CFS concentration of 0%, the CMF of the tested Vibrio cholerae strains treated with 5-20% LpD27-CFS for 3 hours increased significantly (p<0.05), and showed an increase in a LpD27-CFS concentration-dependent manner. After 5% LpD27-CFS treatment for 3 hours, CMF increased by 0.41-0.53 times; after 10% LpD27-CFS treatment for 3 hours, CMF increased by 0.56-0.63 times; after 20% LpD27-CFS treatment for 3 hours, CMF increased by 0.66-0.69 times.

[0047] (IV) Lactobacillus plantarum D27 increases the inner membrane permeability (IMP) of Vibrio cholerae cells

[0048] The mid-LGP Vibrio cholerae suspension treated with LpD27-CFS at different concentrations (0%, 5%, 10%, 20%) was added to a 96-well sterilized bacterial culture plate (200 μL / well), and then 10 mM o-nitrophenyl-β-D-galactopyranoside (ONPG, 2.5 μL / well) was added and incubated at 37°C. The OD value of each well was measured every 30 minutes using a multifunctional microplate reader. 415 Absorbance value. Each group of experiments was repeated three times.

[0049] like Figure 3It can be seen that compared with the control group with a LpD27-CFS concentration of 0%, the IMP of Vibrio cholerae GIM 1.449 strain increased by 0.08 times after 5% LpD27-CFS treatment for 4 hours, while the IMP of the other three strains had no significant difference (p>0.05). However, the IMP of the four tested Vibrio cholerae strains showed a significant increase trend after 10% and 20% LpD27-CFS treatment for 4 hours (p<0.05).

[0050] The CSH, CMF and IMP of cells are important parameters for bacteria to maintain the integrity of cell membrane structure and are also the main targets of antimicrobial drugs. The above experimental results show that Lactobacillus plantarum D27 can effectively reduce the CSH of Vibrio cholerae, increase its CMF and IMP, and show an LpD27-CFS concentration-dependent effect.

[0051] 7. Scanning electron microscopic observation of the destruction of Vibrio cholerae cell surface structure by Lactobacillus plantarum D27

[0052] The mid-LGP Vibrio cholerae were treated with different concentrations (0%, 5%, 10%, 20%) of LpD27-CFS, incubated at 37°C for 3h, centrifuged at 4°C, 4000rpm for 10min, the supernatant was discarded, and the cells were washed twice with 1×PBS buffer. Then, the cells were fixed with 2.5% glutaraldehyde at 4°C for 12h, and then dehydrated with gradient ethanol (30%, 50%, 80%, 90%, 100%) for 15min. Finally, 20μL of the sample was dropped on a coverslip and dried at room temperature. The cell morphology of Vibrio cholerae was observed using a scanning electron microscope (5.0kV, 30,000×), and each group of experiments was repeated three times.

[0053] like Figure 4 It can be seen that in the control group with a LpD27-CFS concentration of 0%, the cells of the tested Vibrio cholerae B5-82, b10-49, 7-6-19, and GIM1.449 strains were full in shape, flat on the surface, and complete in structure. In contrast, after being treated with 5% LpD27-CFS for 3 hours, the cell surface of Vibrio cholerae B5-82 was slightly wrinkled; after being treated with 10% LpD27-CFS for 3 hours, obvious cracks and depressions appeared on the cell surface; after being treated with 20% LpD27-CFS for 3 hours, the cell surface was severely wrinkled and depressed, intracellular substances leaked, and the cell structure was broken. Similarly, after treatment with 5% to 20% LpD27-CFS for 3 h, the cell structure of Vibrio cholerae B10-49, 7-6-19, and GIM1.449 strains also showed a LpD27-CFS concentration-dependent damage pattern, which is consistent with the above-mentioned experimental results that the growth of Vibrio cholerae was inhibited and important parameters of the cell membrane structure were changed.

[0054] Embodiment 2

[0055] Determination of pH value, lactic acid, polysaccharide and protein content in CSF of Lactobacillus plantarum D27

[0056] The pH value of LpD27-CFS was measured using a pH meter, and the experiment was repeated three times.

[0057] The lactic acid content of LpD27-CFS was determined by the p-hydroxybiphenyl colorimetric method: 0.5 mL of 20% copper sulfate solution was added to 5 mL of LpD27-CFS, and after thorough mixing, 0.5 mL of the mixture was slowly added to 6 mL of pre-cooled concentrated sulfuric acid, and after thorough mixing, the mixture was placed in a boiling water bath for 5 min, and then quickly removed and placed in an ice water bath for cooling. Then 125 μL of 1.5% p-hydroxybiphenyl solution was added, and the mixture was quickly mixed, placed in a 30°C water bath for 30 min, and then placed in a boiling water bath for 90 s, and then quickly removed and placed in an ice water bath for cooling, and the OD of the reaction solution was determined using a multifunctional microplate reader. 565 The standard solution of lithium lactate (0.5 mg / mL) was prepared, and the standard curve was drawn. The lactic acid content of LpD27-CFS was calculated according to the standard curve. Each group of experiments was repeated three times.

[0058] The polysaccharide content of LpD27-CFS was determined by the phenol-sulfuric acid method: 1 mL of 5% phenol solution and 5 mL of concentrated sulfuric acid solution were added to 2 mL of appropriately diluted LpD27-CFS, mixed thoroughly and allowed to stand for 5 min, then reacted in a boiling water bath for 15 min, quickly removed and cooled in an ice water bath, and the OD of the reaction solution was determined using a multifunctional microplate reader. 490 Prepare glucose (100 μL / mL) standard solution, draw a standard curve, and calculate the polysaccharide content of LpD27-CFS according to the standard curve. Each group of experiments was repeated three times.

[0059] The protein content of LpD27-CFS was determined by Bradford method: LpD27-CFS was appropriately diluted, and the protein concentration determination kit was used to perform the operation according to the experimental steps in the instruction manual. The OD value of the reaction solution was determined using a multifunctional microplate reader. 595 The absorbance value of the sample was 0.0447 W / mL. A bovine serum albumin (BSA, 5 mg / mL) standard solution was prepared, and a standard curve was drawn. The protein content of LpD27-CFS was calculated based on the standard curve. Each group of experiments was repeated three times.

[0060] As shown in Table 2, the pH value of LpD27-CFS is 3.67±0.00; the lactic acid content is 11.08±0.09 mg / mL; the polysaccharide content is 1.30±0.28 mg / mL; and the protein content is 88.21±0.40 μg / mL, indicating that Lactobacillus plantarum D27 has a strong ability to produce lactic acid and polysaccharides.

[0061] Table 2: pH value of CSF of Lactobacillus plantarum D27 and the results of determination of lactic acid, polysaccharide and protein content

[0062] Test items Standard curve <![CDATA[R 2 ]]> Test results pH / / 3.67±0.00 Lactic acid (mg / mL) y=0.0026x+0.0889 0.9921 11.08±0.09 Polysaccharide (mg / mL) y=0.0092x+0.0749 0.9955 1.30±0.28 Protein (μg / mL) y=0.0014x-0.0130 0.9947 88.21±0.40

[0063] Embodiment 3

[0064] CSH, Self-Aggregation Capacity (SAC), and Co-Aggregation Capacity (CAC) of Lactobacillus plantarum D27 with Vibrio cholerae

[0065] CSH determination of Lactobacillus plantarum D27: Referring to the above-mentioned determination method of Vibrio cholerae CSH, each group of experiments was repeated three times.

[0066] SAC determination of Lactobacillus plantarum D27: Lactobacillus plantarum D27 was inoculated into sterilized MRS liquid medium at a volume fraction of 1%, cultured at 37°C until mid-LGP, centrifuged at 4°C, 4000 rpm for 15 min, the supernatant was discarded, the cells were washed twice with 1× PBS buffer, and then resuspended to a concentration of OD 600 The absorbance value was 0.5±0.05. Take 4 mL of bacterial suspension and vortex mix for 10 seconds, incubate at 37°C for 2 hours, take the upper liquid sample, and use a multifunctional microplate reader to measure its OD 600 Absorbance value. SAC was calculated according to the following formula: SAC (%) = [1-(A2 / A0)] × 100%, where A2 represents the absorbance value at 2 h of incubation, and A0 represents the absorbance value at 0 h of incubation. Each group of experiments was repeated three times.

[0067] Determination of CAC of Lactobacillus plantarum D27 and Vibrio cholerae: Prepare the suspension of Lactobacillus plantarum D27 according to the above method; prepare the suspension of Vibrio cholerae b10-49 strain similarly, taking 2mL of Lactobacillus plantarum D27 and 2mL of Vibrio cholerae b10-49 suspension as the representative strain. Vortex mix for 10s. Incubate at 37℃ for 2h, take the upper liquid sample, and use a multifunctional microplate reader to measure its OD 600 Absorbance value. CAC was calculated according to the following formula: CAC (%) = [1-2A mix / (A Lp +A Vc )]×100%, where A mix Indicates the absorbance value of the mixed bacterial suspension at 2h, A Lp and A Vc It represents the absorbance value of Lactobacillus plantarum D27 and Vibrio cholerae b10-49 before mixing. Each group of experiments was repeated three times.

[0068] CSH, SAC, and CAC are important evaluation parameters for probiotics to adhere to and colonize host intestinal epithelial cells, remove intestinal pathogens, and play a beneficial role. As shown in Table 3, the CSH of Lactobacillus plantarum D27 is 46.97±0.76%; SAC is 45.02%; and the CAC with Vibrio cholerae b10-49 strain is 36.07%. The results show that Lactobacillus plantarum D27 has good CSH, SAC, and co-aggregation characteristics with Vibrio cholerae.

[0069] Table 3: Experimental results of CSF, SAC, and CAC of Lactobacillus plantarum D27

[0070] Tested strain Detection indicators Test results (%) Lactobacillus plantarum D27 CSH 46.97±0.76 Lactobacillus plantarum D27 SAC 45.02±1.82 Lactobacillus plantarum D27 and Vibrio cholerae b10-49 CAC 36.07±1.22

[0071] In summary, the present invention provides a Lactobacillus plantarum D27 for antagonizing Vibrio cholerae. The Lactobacillus plantarum D27 can effectively antagonize Vibrio cholerae, inhibit its biofilm formation, and destroy its cell structure. It has high lactic acid and extracellular polysaccharide production and good self-aggregation and co-aggregation properties. It can be used for the preparation of food or medicine. The prepared food or medicine also has the effect of antagonizing Vibrio cholerae contamination and has broad application prospects.

[0072] Finally, it should be noted that the above embodiments are only used to illustrate the technical solution of the present invention rather than to limit it. Although the present invention has been described in detail with reference to the preferred embodiments, those skilled in the art should understand that they can still modify or replace the technical solution of the present invention with equivalents, and these modifications or equivalent replacements cannot cause the modified technical solution to deviate from the scope of the technical solution of the present invention.

Claims

1. A Lactobacillus plantarum D27 antagonistic to Vibrio cholerae, characterized in that: The Lactobacillus plantarum D27 is deposited in the China Center for Type Culture Collection (CCTCC), the storage address is No. 299, Bayi Road, Wuchang District, Wuhan City, Hubei Province, the storage date is September 4, 2024, the storage number is CCTCC M 20241910, and the classification name is Lactobacillus plantarum.

2. Plant Lactobacillus D27 according to claim 1, characterized in that, The Lactobacillus plantarum D27 can effectively antagonize Vibrio cholerae, inhibit its biofilm formation and destroy its cell structure.

3. Plant Lactobacillus D27 according to claim 1, characterized in that, The plant lactobacillus D27 has high lactic acid and extracellular polysaccharide production, as well as self-aggregation and co-aggregation properties.

4. A plant lactobacillus agent for antagonizing Vibrio cholerae, characterized in that: The invention comprises the Lactobacillus plantarum D27 antagonistic to Vibrio cholerae according to any one of claims 1 to 3.

5. The plant lactobacillus agent according to claim 4, characterized in that The Lactobacillus plantarum agent is selected from one or more of Lactobacillus plantarum liquid, Lactobacillus plantarum powder, Lactobacillus plantarum capsules or Lactobacillus plantarum tablets.

6. Use of the Lactobacillus plantarum D27 according to any one of claims 1 to 3 or the Lactobacillus plantarum agent according to claim 4 or 5 in the preparation of food or medicine for antagonizing Vibrio cholerae contamination.

7. The use according to claim 6, characterized in that: The types of food include ready-to-eat, fresh, refrigerated and frozen.

8. The use according to claim 6, characterized in that: The dosage forms of the drug include liquid, powder, tablet and capsule.

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