A probiotic composition for preventing and / or treating vaginal infections and diseases associated therewith and uses thereof

Abstract

This invention provides a probiotic composition for the prevention and / or treatment of vaginal infections and related diseases, and its use therein. The probiotic composition comprises *Lactobacillus plantarum* and *Lactobacillus fermentum*, wherein the gene sequence of the molecular marker of *Lactobacillus plantarum* has the nucleotide sequence shown in SEQ ID NO: 1, or the nucleotide sequence thereof is shown in SEQ ID NO: 1; and the gene sequence of the molecular marker of *Lactobacillus fermentum* has the nucleotide sequence shown in SEQ ID NO: 2, or the nucleotide sequence thereof is shown in SEQ ID NO: 2. The probiotic composition of this invention achieves better efficacy in treating vaginal infections, especially vulvovaginal candidiasis, compared to single strains therein and existing positive control drugs such as metronidazole suppositories.

Description

Technical Field

[0001] This invention belongs to the field of biomedicine. Specifically, this invention relates to a probiotic composition for the prevention and / or treatment of vaginal infections and related diseases, and its uses. Background Technology

[0002] With economic development and improved living standards, the incidence of female reproductive tract diseases is increasing, especially vaginitis, which seriously affects women's health and quality of life.

[0003] Among vaginal infections, vulvovaginal candidiasis (VVC) is relatively common. Studies have found that approximately 78% of women will experience VVC at least once in their lifetime, with 65% experiencing three or fewer annual episodes and 35% experiencing four or more annual episodes. VVC typically manifests as vulvar itching, burning, painful urination, and cottage cheese-like vaginal discharge, impacting quality of life.

[0004] VVC is typically treated with azole and polyene antifungal drugs, such as metronidazole suppositories. However, long-term use of these drugs has led to phenotypic changes and mutations in virulence factors in Candida albicans, resulting in increased infections with non-white Candida albicans and resistance to traditional treatments. Lactobacillus can be used as adjunctive therapy in combination with azole drugs, but there is currently a lack of evidence to support its effectiveness as a monotherapy for VVC. Summary of the Invention

[0005] To address the above problems, the present invention aims to provide a probiotic composition for the prevention and / or treatment of vaginal infections and related diseases, and its uses. The probiotic composition of the present invention achieves better efficacy in treating vaginal infections, especially VVC, compared to single strains therein and existing positive control drugs such as metronidazole suppositories.

[0006] The above-mentioned objective of the present invention is achieved by providing the following technical solution:

[0007] In a first aspect, the present invention provides a probiotic composition for the prevention and / or treatment of vaginal infections and related diseases, comprising *Lactobacillus plantarum* and *Lactobacillus fermentum*, wherein the gene of the molecular marker of said *Lactobacillus plantarum* has a nucleotide sequence selected from one of the following:

[0008] (1) The nucleotide sequence shown in SEQ ID NO: 1;

[0009] (2) A nucleotide sequence having at least 97%, 98%, 99% or higher homology to the nucleotide sequence shown in SEQ ID NO: 1;

[0010] (3) The nucleotide sequence shown in SEQ ID NO: 1 has one or more, for example, 1, 2, 3, 4, 5 or more nucleotide substitutions, deletions or insertions;

[0011] Preferably, the gene for the molecular marker of *Lactobacillus plantarum* has a nucleotide sequence as shown in SEQ ID NO: 1, or its nucleotide sequence is as shown in SEQ ID NO: 1;

[0012] The gene for the molecular marker of the fermenting *Lactobacillus mucinus* has a nucleotide sequence selected from one of the following:

[0013] (1) The nucleotide sequence shown in SEQ ID NO: 2;

[0014] (2) A nucleotide sequence having at least 97%, 98%, 99% or higher homology to the nucleotide sequence shown in SEQ ID NO: 2;

[0015] (3) The nucleotide sequence shown in SEQ ID NO: 2 has one or more, for example, 1, 2, 3, 4, 5 or more nucleotide substitutions, deletions or insertions;

[0016] Preferably, the gene sequence of the molecular marker of the fermenting Lactobacillus mucinus has the nucleotide sequence shown in SEQ ID NO: 2, or the nucleotide sequence shown in SEQ ID NO: 2.

[0017] According to some embodiments of the present invention, the probiotic composition further comprises Lactobacillus curvatureii, Lactobacillus janniae, or Lactobacillus gasseri.

[0018] According to some embodiments of the present invention, the probiotic composition further comprises any two of Lactobacillus curvaturei, Lactobacillus janniae, and Lactobacillus gasseri.

[0019] According to some embodiments of the present invention, the probiotic composition further comprises Lactobacillus curvaturei, Lactobacillus janniae, and Lactobacillus gasseri.

[0020] According to some embodiments of the present invention, the ratio of viable Lactobacillus plantarum to Lactobacillus fermentum in the probiotic composition is 1:(0.01-100), preferably 1:(1-100).

[0021] According to some embodiments of the present invention, the ratio of viable counts of *Lactobacillus curvaturei*, *Lactobacillus plantarum*, and *Lactobacillus fermentum* is (0.01-100):1:(0.01-100), preferably (0.01-100):1:(1-100);

[0022] According to some embodiments of the present invention, the ratio of viable counts of Lactobacillus janniae, Lactobacillus plantarum and Lactobacillus fermentum is (0.01-100):1:(0.01-100), preferably (0.01-100):1:(1-100).

[0023] According to some embodiments of the present invention, the ratio of viable counts of Lactobacillus gasseri, Lactobacillus plantarum and Lactobacillus fermentum is (0.01-100):1:(0.01-100), preferably (0.01-100):1:(1-100).

[0024] According to some embodiments of the present invention, the total number of live bacteria in the probiotic composition is not less than 2 × 10⁻⁶. 6 CFU / g, preferably 2×10 6 Up to 5×10 9 CFU / g.

[0025] Preferably, the viable count of each single strain in the probiotic composition is not less than 1 × 10⁻⁶. 6 CFU / g, preferably 1×10 6 Up to 9×10 8 CFU / g.

[0026] According to some embodiments of the present invention, the gene for the molecular marker of the *Lactobacillus curvature* has a nucleotide sequence selected from one of the following:

[0027] (1) The nucleotide sequence shown in SEQ ID NO: 3;

[0028] (2) A nucleotide sequence having at least 97%, 98%, 99% or higher homology to the nucleotide sequence shown in SEQ ID NO: 3;

[0029] (3) The nucleotide sequence shown in SEQ ID NO: 3 has one or more, for example, 1, 2, 3, 4, 5 or more nucleotide substitutions, deletions or insertions;

[0030] Preferably, the gene sequence of the molecular marker of the *Lactobacillus curvature* has the nucleotide sequence shown in SEQ ID NO: 3, or the nucleotide sequence thereof is shown in SEQ ID NO: 3;

[0031] Preferably, the strain preservation number of the *Lactobacillus curvatureensis* is CGMCC No. 26503.

[0032] According to some embodiments of the present invention, the gene for the molecular marker of the *Lactobacillus japonicus* has a nucleotide sequence selected from one of the following:

[0033] (1) The nucleotide sequence shown in SEQ ID NO: 4;

[0034] (2) A nucleotide sequence having at least 97%, 98%, 99% or higher homology to the nucleotide sequence shown in SEQ ID NO: 4;

[0035] (3) The nucleotide sequence shown in SEQ ID NO: 4 has one or more, for example, 1, 2, 3, 4, 5 or more nucleotide substitutions, deletions or insertions;

[0036] Preferably, the gene sequence of the molecular marker of the Lactobacillus janniae has the nucleotide sequence shown in SEQ ID NO: 4, or the nucleotide sequence thereof is shown in SEQ ID NO: 4;

[0037] Preferably, the strain preservation number of the Lactobacillus janniae is CGMCC No. 26505.

[0038] According to some embodiments of the present invention, the gene for the molecular marker of the *Lactobacillus gasseri* has a nucleotide sequence selected from one of the following:

[0039] (1) The nucleotide sequence shown in SEQ ID NO: 5;

[0040] (2) A nucleotide sequence having at least 97%, 98%, 99% or higher homology to the nucleotide sequence shown in SEQ ID NO: 5;

[0041] (3) The nucleotide sequence shown in SEQ ID NO: 5 has one or more, for example, 1, 2, 3, 4, 5 or more nucleotide substitutions, deletions or insertions;

[0042] Preferably, the gene sequence of the molecular marker of the Lactobacillus gasseri has the nucleotide sequence shown in SEQ ID NO: 5, or the nucleotide sequence thereof is shown in SEQ ID NO: 5;

[0043] Preferably, the strain preservation number of the Lactobacillus gasseri is CGMCC No. 26504.

[0044] Preferably, the strain preservation number of the *Lactobacillus plantarum* is CGMCC No. 26502.

[0045] Preferably, the strain preservation number of the fermenting *Lactobacillus mucinus* is CGMCC No. 26501.

[0046] In a second aspect, the present invention provides a microbial preparation for the prevention and / or treatment of vaginal infections and related diseases, comprising the probiotic composition according to the first aspect of the present invention.

[0047] According to some embodiments of the present invention, the microbial preparation further comprises pharmaceutically acceptable excipients.

[0048] According to some embodiments of the present invention, the dosage form of the microbial preparation is powder, suppository, tablet, capsule, granule, emulsion or powder inside a capsule.

[0049] Thirdly, the present invention provides a pharmaceutical composition for the prevention and / or treatment of vaginal infections and related diseases, the pharmaceutical composition comprising a probiotic composition according to the first aspect of the present invention or a microbial preparation according to the second aspect of the present invention, as well as other antimicrobial agents.

[0050] Fourthly, the present invention provides the use of the probiotic composition according to the first aspect of the invention or the microbial preparation according to the second aspect of the invention in the preparation of a medicament for the prevention and / or treatment of vaginal infections and related diseases.

[0051] Preferably, the vaginal infection and related diseases are vulvovaginal candidiasis.

[0052] The present invention has at least the following beneficial effects:

[0053] The dual probiotic composition provided by this invention has achieved better results in treating vaginal infections, especially VVC, compared to single strains and existing positive control drugs such as metronidazole suppositories.

[0054] This invention demonstrates through in vivo experiments on mice that the different proportions of the dual probiotic combinations of this invention have a significant effect on reducing the pathogenic bacterial load in VVC model mice, and the antibacterial performance is further enhanced with the addition of one or more of Lactobacillus curvaturei, Lactobacillus janniae, and Lactobacillus gasseri.

[0055] This invention demonstrates through co-culture antibacterial experiments that, based on the dual probiotic composition of this invention, the addition of one, two, or three of Lactobacillus curvaturei, Lactobacillus janniae, and Lactobacillus gasseri can all have a certain inhibitory effect on CA. Furthermore, the antibacterial performance is further enhanced with the addition of one or more of Lactobacillus curvaturei, Lactobacillus janniae, and Lactobacillus gasseri. Attached Figure Description

[0056] The embodiments of the present invention will now be described in detail with reference to the accompanying drawings, wherein:

[0057] Figure 1 This shows the CA (Candida albicans ATCC 10231) load in the vaginal irrigation fluid of VVC model mice in the animal efficacy experiment of single bacteria and composition in Example 1; where, compared with the model group, *P<0.05, **P<0.01, ***P<0.001.

[0058] Figure 2 This shows the CA load in the vaginal irrigation fluid of VVC model mice in the composition ratio screening experiment of Example 1; where, compared with the model group, *P<0.05, **P<0.01, ***P<0.001.

[0059] Figure 3 The results of the co-culture antibacterial experiment of the multi-probiotic composition in Example 1 are shown; among them, compared with the blank control group, ## ## P < 0.0001; compared with the model group, ****P < 0.0001; compared with the dual-combination group, Δ P < 0.05. Detailed Implementation

[0060] The present invention will be further described in detail below with reference to specific embodiments. The embodiments given are only for illustrating the present invention and are not intended to limit the scope of the present invention.

[0061] The *Lactobacillus curvatureii* HY1467 (CGMCC No. 26503), *Lactobacillus japonicus* HY1335 (CGMCC No. 26505), *Lactobacillus gasseri* HY1124 (CGMCC No. 26504), *Lactobacillus plantarum* HY2946 (CGMCC No. 26502), and *Lactobacillus fermentum* HY757 (CGMCC No. 26501) used in the following examples can all be purchased.

[0062] In addition, the following strain used in the following examples can also be obtained by purchase: Candida albicans ATCC 10231, abbreviated as CA.

[0063] Upon identification, the nucleotide shown in SEQ ID NO: 1 is a specific nucleotide sequence marker for *Lactobacillus plantarum* HY2946. After multiple sequencing and alignment tests, the homology range between the molecular marker of *Lactobacillus plantarum* HY2946 and the nucleotide sequence shown in SEQ ID NO: 1 was determined. Specifically, the nucleotide sequence of the molecular marker of *Lactobacillus plantarum* HY2946 has at least 97%, 98%, 99%, or higher homology with the nucleotide sequence shown in SEQ ID NO: 1 (for sequencing and identification methods, please refer to the examples in the applicant's previous patent application 202410369860.6).

[0064]

[0065]

[0066] Upon identification, the nucleotide shown in SEQ ID NO: 2 was confirmed to be a specific nucleotide sequence marker for *Lactobacillus fermentum* HY757. Multiple sequencing and alignment tests determined the homology range between the molecular marker of *Lactobacillus fermentum* HY757 and the nucleotide sequence shown in SEQ ID NO: 2. Specifically, the nucleotide sequence of the molecular marker of *Lactobacillus fermentum* HY757 exhibits at least 97%, 98%, 99%, or higher homology with the nucleotide sequence shown in SEQ ID NO: 2 (for sequencing and identification methods, please refer to the examples in the applicant's previous patent application 202410088916.0).

[0067]

[0068] Upon identification, the nucleotide shown in SEQ ID NO: 3 was confirmed to be a specific nucleotide sequence marker for *Lactobacillus curvature* HY1467. Multiple sequencing and alignment tests determined the homology range between the molecular marker of *Lactobacillus curvature* HY1467 and the nucleotide sequence shown in SEQ ID NO: 3. Specifically, the nucleotide sequence of the molecular marker of *Lactobacillus curvature* HY1467 exhibits at least 97%, 98%, 99%, or higher homology with the nucleotide sequence shown in SEQ ID NO: 3 (for sequencing and identification methods, please refer to the examples in the applicant's previous patent application 202410069354.5).

[0069]

[0070]

[0071] Upon identification, the nucleotide shown in SEQ ID NO:4 was confirmed to be a specific nucleotide sequence marker for *Lactobacillus japonicus* HY1335. Multiple sequencing and alignment tests determined the homology range between the molecular marker of *Lactobacillus japonicus* HY1335 and the nucleotide sequence shown in SEQ ID NO:4. Specifically, the nucleotide sequence of the molecular marker of *Lactobacillus japonicus* HY1335 exhibits at least 97%, 98%, 99%, or higher homology with the nucleotide sequence shown in SEQ ID NO:4 (for sequencing and identification methods, please refer to the examples in the applicant's previous patent application 202410071896.6).

[0072]

[0073] Upon identification, the nucleotide shown in SEQ ID NO: 5 was confirmed to be a specific nucleotide sequence marker for Lactobacillus gasseri HY1124. Multiple sequencing and alignment tests determined the homology range between the molecular marker of Lactobacillus gasseri HY1124 and the nucleotide sequence shown in SEQ ID NO: 5, indicating that the nucleotide sequence of the molecular marker of Lactobacillus gasseri HY1124 has at least 97%, 98%, 99%, or higher homology with the nucleotide sequence shown in SEQ ID NO: 5 (for sequencing and identification methods, please refer to the examples in the applicant's previous patent application 202410073451.1).

[0074]

[0075]

[0076] Example 1: Animal efficacy experiment of VVC

[0077] 1. Animal efficacy experiments of single bacteria and combinations

[0078] 1.1 Experimental Methods

[0079] 1.1.1 Laboratory Animals

[0080] Healthy SPF-grade Balb / c mice, female, 6-8 weeks old, were used.

[0081] 1.1.2 Experimental Grouping

[0082] After adaptive culture, animals were randomly divided into four main groups: blank control group (NC), model group (M), positive control group (double 5mg), and experimental group. The double 5mg group served as a positive control, using 5mg metronidazole suppositories (this dose was calculated from commonly used clinical dosages). The experimental group was further divided into single-strain treatment groups (D, E) and dual-strain combination treatment groups (DE). Specifically, D received single-strain treatment with *Lactobacillus plantarum* HY02946; E received single-strain treatment with *Lactobacillus fermentum* HY00757; and DE received dual-strain treatment (D and E represent *Lactobacillus plantarum* HY2946 and *Lactobacillus fermentum* HY757, respectively).

[0083] 1.1.3 Animal Experiments

[0084] Animals in the model group, positive control group, and experimental group were injected subcutaneously with estradiol benzoate injection daily from D-3 (3 days before modeling) to D-1. They were pretreated vaginally with cefuroxime sodium + levofloxacin hydrochloride antibiotic solution. From D0 to D2, they were continuously given CA (15 μL / animal) for modeling. From D3 to D7 after modeling, the model group (M) received no drug treatment; they were treated with 5 mg / animal metronidazole suppositories. The experimental groups D, E, and DE received 2 × 10 mg of metronidazole suppositories. 7 Single or dual-strain strains (CFU / animal) were administered for 5 consecutive days. Estradiol benzoate was subcutaneously injected once on days 2 and 5 in the model group, positive control group, and experimental group. On day 8, the vagina was irrigated, and the diluted irrigating solution was spread onto Candida albicans identification medium (chromogenic medium). After incubation at 37°C for 48 hours, the number of green colonies on the culture dishes was observed and counted. One-way ANOVA was performed using GraphPad Prism 5 software, with P < 0.05 considered statistically significant.

[0085] 1.2 Experimental Results

[0086] Experimental results are as follows Figure 1 As shown, the pathogenic load in the VVC animal pharmacodynamic model after treatment with the dual-strain combination DE was lower than that of a single strain and 5mg metronidazole suppositories, indicating that the dual-strain combination is more effective in treating VVC than either single strain or the positive control drug metronidazole suppositories.

[0087] 2. Screening of the ratio of dual-strain combinations

[0088] 2.1 Experimental Methods

[0089] 2.1.1 Laboratory Animals

[0090] The selection of laboratory animals is the same as in 1.1.1.

[0091] 2.1.2 Experimental Grouping

[0092] The animal experiments included a blank control group (NC), a model group (M), and nine experimental groups (see the table below for experimental group information).

[0093] Table 1. Strain composition and drug concentration in different experimental groups

[0094] experimental group Strains and dosage concentration T1 6D+6E T2 6D+7E T3 6D+8E T4 7D+6E T5 7D+8E T6 8D+6E T7 8D+8E T8 7D+7E

[0095] Note: D and E represent *Lactobacillus plantarum* HY02946 and *Lactobacillus fermentum* HY00757, respectively. 6, 7, and 8 represent the corresponding drug concentrations of 1.0 × 10⁻⁶. 6 CFU / each, 1.0×10 7 CFU / each, 1.0×10 8 CFU / each.

[0096] 2.1.3 Animal Experiments

[0097] The modeling and administration methods were the same as in 1.1.3. During the treatment, different strain combinations were administered according to the groupings and concentrations in Table 1, with continuous administration for 5 days. One-way ANOVA was performed using GraphPad Prism 5 software, with P < 0.05 considered statistically significant.

[0098] 2.2 Experimental Results

[0099] Depend on Figure 2 The experimental results show that different strain combinations significantly reduced the incidence of pathogenic CA in animals, indicating a therapeutic effect on VVC. The T2 and T5 strain combinations showed the best efficacy against VVC.

[0100] 3. In vitro efficacy test of multi-strain combination

[0101] 3.1 Experimental Methods

[0102] 3.1.1 Strains and Experimental Grouping

[0103] Table 2. Strain Information

[0104] strain number Identification Name HY2946 Lactiplantibacillus plantarum HY757 Fermenting Lactobacillus fermentum HY1467 Lactobacillus crispatus HY00744 Lactobacillus crispatus HY00714 Lactobacillus crispatus HY01628 Lactobacillus crispatus HY1335 Lactobacillus jensenii HY02405 Lactobacillus jensenii HY01361 Lactobacillus jensenii HY1124 Lactobacillus gasseri HY02772 Lactobacillus gasseri HY01880 Lactobacillus gasseri HY02172 Lactobacillus gasseri

[0105] Note: In the table above, HY00744, HY00714, and HY01628 were selected by the inventor from the samples collected by the inventor from the embodiments of the prior patent application with application number 202410069354.5; HY02405 and HY01361 were selected by the inventor from the samples collected by the inventor from the embodiments of the prior patent application with application number 202410071896.6; and HY02772, HY01880, and HY02172 were selected by the inventor from the samples collected by the inventor from the embodiments of the prior patent application with application number 202410073451.1.

[0106] Lactobacillus plantarum HY2946(D) and Lactobacillus fermentum HY757(E) were randomly mixed with Lactobacillus curvaturei, Lactobacillus janniae, and Lactobacillus gasseri from Table 2 to prepare solutions of dual, triple, quadruple, and pentavalent strains of Lactobacillus containing D and E. The total concentration of the bacterial solution was adjusted to 10. 5 -10 9 Within the CFU / mL range, high-throughput screening experiments were then performed.

[0107] 3.1.2 Experimental Methods

[0108] Co-culture of pathogenic bacteria and lactobacilli: Using blank BHIS medium (purchased from Oxoid) as a negative control, take CA glycerol tubes and inoculate 380 μL of CA into Erlenmeyer flasks containing 38 mL of BHIS medium at a 1% inoculation ratio. As a positive control, inoculate 380 μL of CA and 380 μL of the combined bacterial solution into Erlenmeyer flasks containing 38 mL of BHIS medium at a 1% inoculation ratio, gently shake the bacterial solution, and incubate at 37℃ in an anaerobic workstation.

[0109] Viable cell count in co-culture of CA and the composition: Viable cell counts of CA at 0 h were performed using the plate count method. At 5 h and 20 h of incubation, the positive control and co-culture solutions were gently mixed separately, and viable cell counts of CA were performed using Candida chromogenic medium (Chromagar, France).

[0110] 3.2 Experimental Results

[0111] The results of the co-culture antibacterial experiment showed that the combined dual, triple, quadrivalent, and pentavalent probiotic combinations all had good inhibitory effects on CA (causing bacterial infection). Furthermore, the antibacterial performance tended to increase with the addition of different probiotic species. The inhibition rate of the dual-strain combination was approximately 89.5%, while the inhibition rates of the multiple-strain combinations were all higher than 89.5%. The triple, quadrivalent, and pentavalent combinations containing D and E, and further containing HY1467, HY1335, and / or HY1124, showed the best antibacterial effects, with inhibition rates ranging from 95% to 100%. In addition, the experimental results also showed that the concentration of each bacterium below 10... 6 The antibacterial effect is poor when CFU / mL is high, and it is less effective when it is higher than 10. 8 The antibacterial effect was not significantly improved at CFU / mL.

[0112] 4. In vivo efficacy test of multi-strain combination

[0113] 4.1 Experimental Methods

[0114] 4.1.1 Laboratory Animals

[0115] The selection of laboratory animals is the same as in 1.1.1.

[0116] 4.1.2 Experimental Grouping

[0117] The treatment of the blank control group (NC) and the model group (M) is the same as in 1.1.3. The in vivo efficacy of the multi-combination was verified using the following experimental groups as examples. The composition and dosage of each experimental group are shown in the table below.

[0118] Table 3. Composition and drug dosage of different experimental groups

[0119] experimental group Composition and dosage Dual Combination 6D+7E Five-component combination 7A+7B+7C+6D+7E

[0120] Note: A, B, C, D, and E represent *Lactobacillus curvatureii* HY1467, *Lactobacillus japonicus* HY1335, *Lactobacillus gasseri* HY1124, *Lactobacillus plantarum* HY2946, and *Lactobacillus fermentum* HY757, respectively. 6, 7, and 8 represent viable bacterial counts of 1.0 × 10⁻⁶. 6 CFU / each, 1.0×10 7 CFU / each.

[0121] 4.1.3 Animal Experiments

[0122] The modeling and administration methods were the same as in 1.1.3. Different compositions were administered for 5 consecutive days during the treatment. One-way ANOVA was performed using GraphPad Prism 5 software, with P < 0.05 considered statistically significant.

[0123] 4.2 Experimental Results

[0124] Experimental results are as follows Figure 3 As shown, both the five-component combination and the two-component combination significantly reduced the pathogenic bacterial load in VVC model mice (P < 0.0001), and the five-component combination was significantly better than the two-component combination in reducing the pathogenic bacterial load.

[0125] The above descriptions are merely several exemplary embodiments of the present invention and are not intended to limit the present invention in any way. Although the present invention has been disclosed above with reference to preferred embodiments, it is not intended to limit the present invention. Any equivalent or related embodiments obtained by those skilled in the art through some modifications or variations made to the above-disclosed technical content without departing from the scope of the present invention are within the scope of the present invention.

Claims

1. A probiotic composition for the prevention and / or treatment of vaginal infections and related diseases, comprising *Lactobacillus plantarum* and *Lactobacillus fermentum*. in, The gene for the molecular marker of *Lactobacillus plantarum* has a nucleotide sequence selected from one of the following: (1) The nucleotide sequence shown in SEQ ID NO: 1; (2) A nucleotide sequence having at least 97%, 98%, 99% or higher homology to the nucleotide sequence shown in SEQ ID NO: 1; (3) The nucleotide sequence shown in SEQ ID NO: 1 has one or more, for example, 1, 2, 3, 4, 5 or more nucleotide substitutions, deletions or insertions; Preferably, the gene for the molecular marker of *Lactobacillus plantarum* has a nucleotide sequence as shown in SEQ ID NO: 1, or its nucleotide sequence is as shown in SEQ ID NO: 1; The gene for the molecular marker of the fermenting *Lactobacillus mucinus* has a nucleotide sequence selected from one of the following: (1) The nucleotide sequence shown in SEQ ID NO: 2; (2) A nucleotide sequence having at least 97%, 98%, 99% or higher homology to the nucleotide sequence shown in SEQ ID NO: 2; (3) The nucleotide sequence shown in SEQ ID NO: 2 has one or more, for example, 1, 2, 3, 4, 5 or more nucleotide substitutions, deletions or insertions; Preferably, the gene sequence of the molecular marker of the fermenting Lactobacillus mucinus has a nucleotide sequence as shown in SEQ ID NO: 2, or its nucleotide sequence is as shown in SEQ ID NO:

2.

2. The probiotic composition according to claim 1, wherein, The probiotic composition also contains Lactobacillus curvatureii, Lactobacillus janniae, or Lactobacillus gasseri.

3. The probiotic composition according to claim 1, wherein, The probiotic composition also includes any two of Lactobacillus curvaturei, Lactobacillus janniae, and Lactobacillus gasseri.

4. The probiotic composition according to claim 1, wherein, The probiotic composition also contains Lactobacillus curvaturei, Lactobacillus janniae, and Lactobacillus gasseri.

5. The probiotic composition according to any one of claims 1 to 4, wherein, The ratio of viable Lactobacillus plantarum to Lactobacillus fermentum in the probiotic composition is 1:(0.01-100), preferably 1:(1-100); Preferably, the ratio of viable counts of *Lactobacillus curvaturei*, *Lactobacillus plantarum*, and *Lactobacillus fermentum* is (0.01-100):1:(0.01-100), and more preferably (0.01-100):1:(1-100). Preferably, the ratio of viable counts of Lactobacillus janniae, Lactobacillus plantarum, and Lactobacillus fermentum is (0.01-100):1:(0.01-100), and more preferably (0.01-100):1:(1-100). Preferably, the ratio of viable counts of Lactobacillus gasseri, Lactobacillus plantarum, and Lactobacillus fermentum is (0.01-100):1:(0.01-100), and more preferably (0.01-100):1:(1-100).

6. The probiotic composition according to any one of claims 1 to 5, wherein, The total number of live bacteria in the probiotic composition is not less than 2 × 10⁻⁶. 6 CFU / g, preferably 2×10 6 Up to 5×10 10 CFU / g; Preferably, the viable count of each single strain in the probiotic composition is not less than 1 × 10⁻⁶. 6 CFU / g, preferably 1×10 6 Up to 9×10 9 CFU / g.

7. The probiotic composition according to any one of claims 1 to 6, wherein, The strain preservation number of the *Lactobacillus plantarum* is CGMCC No. 26502; Preferably, the strain preservation number of the fermenting *Lactobacillus mucinus* is CGMCC No. 26501.

8. A microbial preparation for the prevention and / or treatment of vaginal infections and related diseases, comprising a probiotic composition according to any one of claims 1 to 7.

9. The microbial preparation according to claim 8, wherein, The microbial preparation also contains pharmaceutically acceptable excipients.

10. The microbial preparation according to claim 8 or 9, wherein, The dosage form of the microbial preparation is powder, suppository, tablet, capsule, granule, emulsion, or powder inside a capsule.

11. A pharmaceutical composition for the prevention and / or treatment of vaginal infections and related diseases, said pharmaceutical composition comprising a probiotic composition according to any one of claims 1 to 7 or a microbial preparation according to any one of claims 8 to 10, and other antimicrobial agents.

12. Use of the probiotic composition according to any one of claims 1 to 7 or the microbial preparation according to any one of claims 8 to 10 in the preparation of a medicament for the prevention and / or treatment of vaginal infections and related diseases; Preferably, the vaginal infection and related diseases are vulvovaginal candidiasis.

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