Application of Akkermansia muciniphila in preparation of product for resisting klebsiella pneumoniae infection

Abstract

The invention discloses an application of Akkermansia muciniphila in the preparation of a product for resisting klebsiella pneumoniae infection, and particularly discloses an application of Akkermansia muciniphila in the preparation of a product for resisting klebsiella pneumoniae infection. The invention provides an application of A. muciniphila bacteria or a culture solution thereof or any one of the following applications: 1) preparing a product for resisting host infection with klebsiella pneumoniae; 2) preparing a product for reducing the pathogenicity of klebsiella pneumoniae to a host; (3) preparing a product for improving the Klebsiella pneumoniae infection resistance of a host; and 4) application in preparation of a product for activating the AhR signal channel of the host intestinal tract. Compared with the traditional antibiotic therapy, the A. muciniphila, as the emerging second-generation probiotics, can effectively resist klebsiella pneumoniae infection, can reduce the dosage of antibiotics, and provides a new method for improving sharp increase of drug-resistant bacteria and difficulty in treatment caused by antibiotic abuse.

Description

Technical Field

[0001] The invention belongs to the field of probiotics resisting pathogenic bacteria infection, and particularly relates to application of Akkermansia muciniphila in preparing a product resisting Klebsiella pneumoniae infection. Background Art

[0002] Klebsiella pneumoniae (KP) belongs to the Enterobacteriaceae family and can cause a variety of diseases, including pneumonia, urinary tract infections (UTIs), bloodstream infections, and sepsis. It is one of the common opportunistic pathogens in clinical practice (Bengoechea and Sa Pessoa, 2019). Hypervirulent Klebsiella pneumoniae (hvKP) is an evolving variant that is more virulent than classical K. pneumoniae (cKP). It was first discovered in Southeast Asia and usually spreads among healthy individuals in the community and causes infection. It is the main cause of suppurative liver abscesses (Russo and Marr, 2019; Wyres et al., 2020). The latest epidemiological studies have shown that hvKP has begun to spread widely internationally, posing a serious threat to global human health (Lee et al., 2017). Antibiotic therapy is currently an important means of treating hvKP infection in clinical practice (Choby et al., 2020), but recent studies have found that hvKP is evolving towards multidrug resistance and has evolved into variant resistant strains that can produce beta-lactamase and carbapenemase (Liu et al., 2014). Among them, the proportion of carbapenemase-producing hvKP strains in my country has rapidly increased from 7.4% to 15% between 2013 and 2015 (Lee et al., 2017; Paczosa and Mecsas, 2016). This "superbug" produced by the fusion evolution of high virulence and multidrug resistance has greatly increased the difficulty of clinical antibiotic treatment, resulting in higher mortality and posing a new threat to the global public health network. Therefore, it is urgent to find new alternatives and / or new means of auxiliary antibiotic treatment.

[0003] Beneficial intestinal commensal bacteria have the function of regulating the host's immune function. They can directly block the invasion of exogenous pathogens by producing metabolites or competing for nutrients, and can also indirectly eliminate pathogens by activating the host's intestinal immune function (Pickard et al., 2017; Seekatz et al., 2018). At present, studies have shown that some beneficial bacteria in the intestine can promote host resistance to cKP, but no reports have been found on intestinal commensal bacteria resisting hvKP infection by regulating intestinal immune function. Studies have confirmed in mouse models that some lactobacilli and Escherichia coli Nissle1917 may resist sepsis caused by excessive growth of pathogenic KP by reducing the oxygen content in the intestine (Singer et al., 2019). In addition, a research group found that there are intestinal commensal bacteria in the human intestine that can resist KP colonization. For example, intestinal Bacteroides can enhance intestinal immunity through the IL-36 signaling pathway and macrophages, inhibiting the colonization of KP in the intestine (Sequeira et al., 2020), and acid-producing Klebsiella can directly compete with KP for specific carbon sources (including various β-glucosides) under the mediation of casA, which metabolizes β-glucosides, to achieve the effect of resisting KP (Osbelt et al., 2021). In addition, another study predicted through hierarchical logistic regression analysis that bacteria in the Desulfovibrio and Ruminococcaceae in the intestine may have the effect of resisting CRKP (Seekatz et al., 2018). It can be seen that the exploration of intestinal commensal bacteria that can resist hvKP colonization and infection may serve as a new target for the prevention and treatment of hvKP, which has great application potential. Summary of the invention

[0004] The present invention aims at the existing clinical problems and finds that the intestinal symbiotic bacteria Akkermansia muciniphila (A. muciniphila) has a new function of resisting Klebsiella pneumoniae infection. Exogenous supplementation of A. muciniphila can significantly reduce the infection rate and mortality rate of Klebsiella pneumoniae to the host.

[0005] In a first aspect, the present invention provides the use of A. muciniphila or its culture solution or its bacterial solution in any of the following:

[0006] 1) Preparation of products for preventing diseases caused by Klebsiella pneumoniae;

[0007] 2) preparing products for resisting Klebsiella pneumoniae infection in the body;

[0008] 3) preparing products that reduce the pathogenicity of Klebsiella pneumoniae to the body;

[0009] 4) preparing products that improve the body's ability to resist Klebsiella pneumoniae infection;

[0010] 5) Application in the preparation of products for increasing the expression of intestinal AhR signals, IL-22 and / or intestinal tight junction proteins.

[0011] In the above text, the A. muciniphila bacteria are live bacteria.

[0012] The above-mentioned resistance to Klebsiella pneumoniae infection in the body is specifically reflected in reducing the mortality rate of Klebsiella pneumoniae to the body.

[0013] In the above applications, the organism is a human or an animal.

[0014] In the above application, the animal is a mammal;

[0015] Furthermore, the mammal is specifically a rodent.

[0016] In a second aspect, the present invention provides a product, the active ingredient of which is A. muciniphila bacteria or its culture solution or its bacterial solution;

[0017] The product has any of the following functions:

[0018] 1) Prevent diseases caused by Klebsiella pneumoniae;

[0019] 2) Fight against Klebsiella pneumoniae infection;

[0020] 3) Reduce the pathogenicity of Klebsiella pneumoniae to the body;

[0021] 4) Improve the body's ability to resist Klebsiella pneumoniae infection;

[0022] 5) Increase the expression of intestinal AhR signaling, IL-22 and / or intestinal tight junction proteins.

[0023] In the above, in the embodiments of the present invention, the intestinal tight junction protein is ZO-1, Occludin and / or Caludin.

[0024] In the above, in the embodiments of the present invention, A. muciniphila is exemplified by A. muciniphila DSM22959. Klebsiella pneumoniae is exemplified by Klebsiella pneumoniae ATCC43816.

[0025] The present invention is achieved through the following technical solutions:

[0026] The inventors previously compared the changes in the intestinal flora of mice before and after Klebsiella pneumoniae infection, and targeted the beneficial symbiotic bacteria A.muciniphila that participates in regulating the host's resistance to Klebsiella pneumoniae invasion. First, it was determined whether exogenous supplementation of A.muciniphila could resist Klebsiella pneumoniae infection in mice. The results showed that exogenous intervention with A.muciniphila significantly reduced the mortality rate of Klebsiella pneumoniae in mice from 76.5% in the control group to 38.9%. Then, the inventors studied the mechanism of action of A.muciniphila in reducing the infection rate of Klebsiella pneumoniae from in vitro and in vivo perspectives, and co-cultured A.muciniphila DSM22959 live bacteria, culture supernatant and inactivated bacteria with the standard strain of Klebsiella pneumoniae ATCC43816, respectively, to detect the growth of Klebsiella pneumoniae. The results showed that the live bacteria, culture supernatant and inactivated bacteria of A. muciniphila DSM22959 had no significant effect on the growth of Klebsiella pneumoniae. However, when the immunosuppressant dexamethasone was used to inhibit the intestinal immune function of mice, the resistance efficiency of A. muciniphila to Klebsiella pneumoniae infection was significantly reduced, indicating that A. muciniphila may resist Klebsiella pneumoniae infection by regulating intestinal immune function.

[0027] The inventors hypothesized through literature research that A. muciniphila may achieve the effect of resisting Klebsiella pneumoniae infection by activating the intestinal AhR signaling pathway and promoting intestinal immune function. To confirm this hypothesis, the applicant conducted preliminary tests on the effects of exogenous A. muciniphila intervention on mouse intestinal AhR and immune function. The intervention effect was evaluated by detecting the expression levels of genes such as AhR, RORγt, IL-22, IL-17A and IFN-γ in the ileum of mice after exogenous A. muciniphila intervention by Q-PCR. The results showed that exogenous A. muciniphila intervention can significantly activate the intestinal AhR signaling pathway, promote the expression of intestinal IL-22, and significantly increase the expression of intestinal tight junction proteins (ZO-1, Occludin and Caludin) genes. The above results indicate that A. muciniphila intervention may enhance intestinal barrier function through the AhR signaling pathway. Then, AhR antagonists were used to inhibit the binding ability of mouse intestinal AhR to aromatic hydrocarbon ligands. The results showed that AhR antagonists significantly reduced the effect of A. muciniphila intervention in reducing the mortality rate of Klebsiella pneumoniae infection, confirming that A. muciniphila resists Klebsiella pneumoniae infection through the AhR signaling pathway.

[0028] Compared with the prior art, the present invention has beneficial effects: compared with traditional antibiotic therapy, A. muciniphila, as an emerging second-generation probiotic, can reduce the dosage of antibiotics while effectively resisting Klebsiella pneumoniae infection, providing a new method to improve the surge in drug-resistant bacteria and treatment difficulties caused by the abuse of antibiotics. BRIEF DESCRIPTION OF THE DRAWINGS

[0029] Figure 1 The effect of Klebsiella pneumoniae on the relative abundance of Akkermansia muciniphila in the intestinal flora of mice.

[0030] Figure 2 Effects of Klebsiella pneumoniae on mouse body weight (A) and abundance of A. muciniphila in the intestine (B). * Indicates P<0.05.

[0031] Figure 3 Klebsiella pneumoniae damages the host intestinal barrier function.

[0032] Figure 4 The effect of A. muciniphila intervention on the lethality of Klebsiella pneumoniae in mice; (A) Experimental design and treatment, (B) Mouse survival curve. * indicates P < 0.05.

[0033] Figure 5 Co-culture of A. muciniphila and Klebsiella pneumoniae.

[0034] Figure 6 The effect of exogenous A. muciniphila intervention on gene expression in mouse ileum; * indicates P < 0.05, ** indicates P < 0.01, and each point represents the mean of the results of a separate animal experiment.

[0035] Figure 7 AhR antagonists inhibit the effect of A.muciniphila against Klebsiella pneumoniae. DETAILED DESCRIPTION

[0036] Unless otherwise specified, the experimental methods used in the following examples are conventional methods.

[0037] Unless otherwise specified, the materials and reagents used in the following examples can be obtained from commercial sources.

[0038] The PBS used in the following examples is a PBS solution with a concentration of 0.1 M and a pH value of 7.2-7.4.

[0039] A.muciniphila DSM22959, strain name: Akkermansia muciniphila; selling company: BNCC; product catalog number: 341917;

[0040] Klebsiella pneumoniae ATCC43816 (hereinafter referred to as 43816), strain name: Klebsiella pneumoniae; selling company: American ATCC; product catalog number: 70028205.

[0041] Table 1 shows the primer sequences

[0042]

[0043] Example 1. Discovery of the resistance bacteria A. muciniphila screened by Klebsiella pneumoniae changing the intestinal flora of the host I. The resistance bacteria A. muciniphila screened by Klebsiella pneumoniae changing the intestinal flora of the host

[0044] In order to study and screen intestinal commensal bacteria that have the ability to resist Klebsiella pneumoniae infection, a Klebsiella pneumoniae asymptomatic colonization mouse model was first constructed, and the intestinal flora structure of mice after Klebsiella pneumoniae treatment was analyzed by 16s rRNA sequencing.

[0045] Method for constructing a mouse model without infection symptoms:

[0046] 1) Klebsiella pneumoniae bacterial culture: prepare 2 Columbia blood plates, take out ATCC43816 bacterial solution from the -80℃ bacterial library, use an inoculation loop to take frozen bacterial solution and draw a line on the blood plate in a biosafety cabinet, then place it in a 37℃ bacterial incubator and culture it for 12 hours;

[0047] 2) Use an inoculation loop to pick a single colony from the lined plate and inoculate it into LB medium for overnight culture;

[0048] 3) Carry out activation culture at a ratio of 1:50 for 1-2 hours to make the bacteria in the logarithmic growth phase, centrifuge at 10000 rpm / min for 5 minutes, collect the bacterial precipitate, and resuspend it with an appropriate amount of PBS to adjust the bacterial solution concentration to 5×10 8 cfu / mL, and 43816 bacterial liquid was obtained;

[0049] 4) Six 5-week-old SPF-grade C57BL / 6 mice were taken, and fecal samples were collected from the mice (recorded as 1 day before treatment), and then 200 μL 43816 bacterial solution was gavaged once;

[0050] 5) On the 5th day after infection (represented as 5 days after treatment in the figure), mouse feces were collected and 16s rRNA sequencing was performed to analyze the structure and differences of the mouse intestinal flora before and after 43816 treatment, and to screen for intestinal commensal bacteria that may have the ability to resist 43816 infection.

[0051] According to the above method, A. muciniphila was successfully screened, which may play an important role in promoting mice to resist the invasion of Klebsiella pneumoniae ( Figure 1 ).

[0052] The relative abundance of A. muciniphila in the intestine was significantly different between Klebsiella pneumoniae-infected and asymptomatically colonized mice

[0053] To further explore whether A. muciniphila plays a key role in host resistance to Klebsiella pneumoniae infection, this study established a symptomatic mouse model of Klebsiella pneumoniae infection.

[0054] Construction of mouse model of Klebsiella pneumoniae infection:

[0055] 1) Cultivate and prepare 43816 bacterial solution according to the method for constructing a mouse model without infection symptoms;

[0056] 2) Twelve 5-week-old SPF C57BL / 6 mice were selected, and fecal samples were collected. Then, 200 μL of 43816 bacterial solution was gavaged once a day for 2 consecutive days. PBS was used as the control.

[0057] 3) Record the weight and vitality of mice to distinguish infected mice (4 mice) from uninfected mice (8 mice), and collect fecal samples of mice (symptoms of severe infection of mice: 1. The animal is dying or unable to move; 2. Difficulty breathing, salivation or cyanosis; 3. Diarrhea or urinary incontinence; 4. Weight loss of 20% of the body weight before the experiment; 5. Inability to eat or drink).

[0058] 4) Using Q-PCR and total bacteria as the internal reference, we further detected and compared the difference in the relative abundance of A. muciniphila in the intestines of infected and uninfected mice (primers were A. muciniphila-F and A. muciniphila-R, and the internal reference primers were total bacteria-F and total bacteria-R).

[0059] The mouse body weight results were as follows Figure 2 As shown in A, D0 represents the day after intragastric administration, D5 represents the fifth day after intragastric administration, PBS represents the intragastric administration PBS group, and 43816 represents the intragastric administration 43816 bacterial solution. It can be seen that the body weight of infected mice was significantly reduced.

[0060] The relative abundance of A. muciniphila was detected as follows: Figure 2As shown in Figure B, it can be seen that the relative abundance of A. muciniphila in the feces of uninfected mice (marked as no infection symptoms in the figure) showed a significant upward trend (P=0.0573), and the relative abundance of A. muciniphila in the feces of infected mice (marked as severe infection in the figure) was significantly reduced (P<0.01), indicating that the relative abundance of A. muciniphila is directly correlated with Klebsiella pneumoniae infection.

[0061] 3. Klebsiella pneumoniae infection seriously damages the intestinal barrier

[0062] To investigate the effect of Klebsiella pneumoniae infection on the intestinal barrier function of mice, this study detected the intestinal barrier structure of mice after Klebsiella pneumoniae infection.

[0063] 1) Cultivate and prepare 43816 bacterial solution according to the method for constructing a mouse model without infection symptoms described above;

[0064] 2) 5-week-old SPF-grade C57BL / 6 mice were gavaged once with 200 μL of 43816 bacterial solution; the control group was gavaged once with 200 μL of PBS.

[0065] 3) Observe the status of mice, distinguish between severe infection and no infection symptoms based on the weight and vitality of mice, collect ileum samples from severely infected mice 5 days after gavage, and collect ileum samples from mice with no infection symptoms and control group mice at the same time, fix the intestinal samples with tissue fixative, and then perform HE section staining.

[0066] The results are as follows Figure 3 As shown, AC are the ileum morphology of the control group, mice without infection symptoms and mice with severe infection, respectively. It can be seen that Klebsiella pneumoniae severely damages the intestinal barrier function and causes infection.

[0067] Therefore, intestinal mucosal immunity is the main barrier against the invasion of Klebsiella pneumoniae.

[0068] Example 2: A. muciniphila intervention reduces the mortality rate of Klebsiella pneumoniae in mice

[0069] To further investigate whether A. muciniphila is involved in the infection efficiency of Klebsiella pneumoniae, an A. muciniphila intervention experiment was conducted to study the effect of A. muciniphila intervention on promoting resistance of mice to Klebsiella pneumoniae infection ( Figure 4 A).

[0070] 1) Experimental animals and grouping: 40 5-week-old C57BL6 male mice were selected and randomly divided into 4 groups of 10 mice each, 2 mice per cage, and fed and ate freely.

[0071] 2) Pick a single colony of A. muciniphila DSM22959 in BHI medium and culture it anaerobically for 72 h. Centrifuge at 6000 rpm / min for 5 min, collect the bacterial precipitate, and resuspend it in an appropriate amount of PBS to adjust the bacterial solution concentration to 5×10 8 cfu / mL, recorded as DSM22959 bacterial solution;

[0072] After ATCC43816 was cultured overnight in LB liquid medium, it was activated and cultured for 1-2 hours at a ratio of 1:50 to keep the bacteria in the logarithmic growth phase. The bacterial precipitate was collected by centrifugation at 6000 rpm / min for 5 minutes and resuspended in an appropriate amount of PBS to adjust the bacterial solution concentration to 5×10 8 cfu / mL, the bacterial solution was used for oral administration of mice and recorded as 43816 bacterial solution.

[0073] 3) Experimental treatment:

[0074] PBS group: 200 μL PBS solution was gavaged every day for 5 consecutive days, followed by 200 μL 43816 bacterial solution gavaged every day for two consecutive days;

[0075] AKK-1d group: 200 μL PBS solution was gavaged daily from day 1 to day 4, 200 μL DSM22959 bacterial solution was gavaged daily on day 5, and then 200 μL 43816 bacterial solution was gavaged daily for two consecutive days;

[0076] AKK-2d group: 200 μL PBS solution was gavaged daily on days 1-3, 200 μL DSM22959 bacterial solution was gavaged daily on days 4-5, and then 200 μL 43816 bacterial solution was gavaged daily for two consecutive days;

[0077] AKK-5d group: mice were gavaged with 200 μL DSM22959 bacterial solution for 5 consecutive days, and then gavaged with 200 μL 43816 bacterial solution for 2 consecutive days.

[0078] 4) Observe and record the survival rate of mice within 14 days after infection and draw a survival curve.

[0079] The results are as follows Figure 4 As shown in B, it can be seen that exogenous supplementation of A. muciniphila can effectively reduce the mortality rate of kvKP in mice.

[0080] Example 3: Study on the mechanism of A. muciniphila interfering with the lethality of Klebsiella pneumoniae in mice

[0081] 1. A. muciniphila does not affect the growth of Klebsiella pneumoniae when co-cultured with Klebsiella pneumoniae in vitro

[0082] In order to study the reasons why A. muciniphila reduces the mortality rate of Klebsiella pneumoniae, the applicant studied whether A. muciniphila has an antagonistic effect on Klebsiella pneumoniae through an in vitro co-culture experiment.

[0083] 1. Preparation of live A. muciniphila DSM22959 bacteria, culture supernatant and inactivated bacteria, and 43816 bacterial solution

[0084] A single colony of A. muciniphila DSM22959 was picked up and placed in BHI medium for anaerobically culture for 72 h. The supernatant and bacterial precipitate were collected by centrifugation at 6000 rpm / min for 5 min. The supernatant was filtered through a 0.2 μm filter to remove the bacteria and the filtrate was collected as the culture supernatant. The precipitate was washed twice with an appropriate amount of PBS and resuspended to adjust the bacterial solution concentration to 5 × 10 4 cfu / mL, recorded as DSM22959 live bacterial solution; 1 mL of DSM22959 live bacterial solution was taken and inactivated by pasteurization (70°C water bath for 30 minutes followed by rapid ice water bath for 10 minutes) to prepare DSM22959 inactivated bacterial solution;

[0085] After ATCC43816 was cultured overnight in LB liquid medium, it was activated and cultured for 1-2 hours at a ratio of 1:50 to keep the bacteria in the logarithmic growth phase. The bacterial precipitate was collected by centrifugation at 6000 rpm / min for 5 minutes, and then washed twice with an appropriate amount of PBS and resuspended to adjust the bacterial solution concentration to 5×10 4 cfu / mL, the bacterial solution was used for oral administration of mice and recorded as 43816 live bacterial solution.

[0086] 2. Co-cultivation

[0087] Take a 96-well plate (with lid), and divide 6 consecutive wells into 6 groups:

[0088] a. Control group (LB): 200 μL of LB medium was added to each well as a blank control;

[0089] b. Akkermansia live bacteria group (Akkermansia): add 200 μL of DSM22959 live bacteria solution to each well;

[0090] c.43816 group (43816): add 200 μL of 43816 live bacterial solution to each well;

[0091] d. 43816+akk group (43816+akk): 200 μL of 1:1 (volume ratio) mixed DSM22959:43816 live bacterial solution was added to each well;

[0092] e. 43816+akk supernate group: 200 μL of a 1:1 (volume ratio) mixture of DSM22959 supernatant and 43816 live bacterial solution was added to each well;

[0093] f. 43816+inactived akk group: 200 μL of 1:1 (volume ratio) mixed inactivated DSM22959:43816 live bacteria solution was added to each well;

[0094] The 96-well plate (with lid) was placed in a microplate reader to measure the growth curve. The culture temperature was set to 37° C. and the measurement was continued for 48 hours. The OD600 absorbance was measured every 10 minutes.

[0095] The results are as follows Figure 5 As shown, it can be seen that the live bacteria, culture supernatant and inactivated bacteria of A. muciniphila DSM22959 had no significant effect on the growth of 43816, indicating that A. muciniphila has no direct antagonistic effect on Klebsiella pneumoniae.

[0096] 2. Exogenous supplementation of A. muciniphila activates intestinal AhR signaling pathway

[0097] To clarify whether A. muciniphila activates the intestinal AhR signaling pathway and promotes intestinal immune function, a preliminary test on the effects of exogenous A. muciniphila intervention on intestinal AhR and immune function in mice was conducted.

[0098] The ileum of mice after 5 days of exogenous A. muciniphila intervention in Example 2 (i.e., the AKK-5d group was gavaged with 200 μL DSM22959 bacterial solution for 5 consecutive days) was used as the intervention group sample, and the genomic DNA of the intervention group samples was extracted. The relative expression levels of genes such as AhR, RORγt (denoted as ROR in the figure), IL-22, IL-17A and IFN-γ in the ileum of mice after exogenous A. muciniphila intervention (intervention group / control group) were detected by Q-PCR to evaluate the intervention effect.

[0099] The ileum samples that were gavaged with 200 μL PBS solution for 5 consecutive days were used as the control group samples.

[0100] The results are as follows Figure 6 As shown, it can be seen that exogenous A. muciniphila intervention can significantly activate intestinal AhR signaling, promote the expression of intestinal IL-22, and significantly increase the expression of intestinal tight junction protein (ZO-1, Occludin and Caludin) genes.

[0101] These results suggest that A. muciniphila intervention may enhance intestinal barrier function through AhR signaling.

[0102] The above preliminary experimental results show that A. muciniphila has no obvious antagonistic effect on Klebsiella pneumoniae in vitro, but exogenous supplementation of A. muciniphila significantly reduces the mortality rate of Klebsiella pneumoniae in mice, and exogenous supplementation of A. muciniphila significantly increases the expression levels of ileal AhR, RORγ and IL-22 genes.

[0103] This provides an important basis for the subsequent search for the hypothesis that A. muciniphila resists Klebsiella pneumoniae infection by regulating the host intestinal barrier function.

[0104] 3. AhR antagonists inhibit the resistance of A. muciniphila to Klebsiella pneumoniae

[0105] To further verify that A. muciniphila is the result of gene expression of the AhR signaling pathway in the living intestine, AhR antagonists were used to inhibit the binding ability of mouse intestinal AhR to aromatic hydrocarbon ligands.

[0106] 1) Prepare DSM22959 bacterial solution (referred to as AKK bacterial solution) and 43816 bacterial solution according to Example 2;

[0107] 2) Preparation of AhR inhibitor solution: AhR inhibitor was purchased from MCE (catalog number HY-141609), and the stock solution was prepared according to the instructions, diluted to 1 mg / mL, and used immediately after preparation;

[0108] 3) Experimental animals and grouping: 90 5-week-old C57BL6 male mice were selected and randomly divided into 3 groups: control group (28 mice); AKK intervention group (34 mice); AhR inhibitor group (28 mice). Two mice were placed in each cage and allowed to eat and drink freely.

[0109] 4) Drug administration

[0110] Control group: After continuous intragastric administration of PBS solution for 7 days (200 μL per day), 43816 bacterial solution was continuously intragastrically administered for 2 days (200 μL per day);

[0111] AKK treatment group: After continuous gavage with AKK bacterial solution for 5 days (200 μL per day), PBS solution was gavage for 2 days (200 μL per day), and then 43816 bacterial solution was gavage for 2 days (200 μL per day);

[0112] AhR inhibitor group: After continuous gavage with AKK bacterial solution for 5 days (200 μL per day), the mice were gavaged with AhR inhibitor solution for 2 days (200 μL per day), and then with 43816 bacterial solution for 2 consecutive days (200 μL per day).

[0113] 5) Observe and record the mortality rate of mice.

[0114] The results are as follows Figure 7 As shown, it can be seen that AhR antagonists significantly reduced the effect of A. muciniphila intervention in reducing the mortality rate of Klebsiella pneumoniae infection, indicating that A. muciniphila may resist Klebsiella pneumoniae infection through the AhR signaling pathway.

Claims

1. Use of A. muciniphila or its culture solution or its bacterial solution in any of the following: 1) Preparation of products for preventing diseases caused by Klebsiella pneumoniae; 2) preparing products for resisting Klebsiella pneumoniae infection in the body; 3) preparing products that reduce the pathogenicity of Klebsiella pneumoniae to the body; 4) preparing products that improve the body's ability to resist Klebsiella pneumoniae infection; 5) Preparing products that increase the expression of intestinal AhR signals, IL-22 and / or intestinal tight junction proteins.

2. The use according to claim 1, characterized in that: The organism is a human or an animal.

3. The use according to claim 2, characterized in that: The animal is a mammal.

4. The use according to claim 3, characterized in that: The mammals are specifically rodents.

5. A product, the active ingredient of which is A. muciniphila or its culture solution or its bacterial solution; The product has any of the following functions: 1) Prevent diseases caused by Klebsiella pneumoniae; 2) Fight against Klebsiella pneumoniae infection; 3) Reduce the pathogenicity of Klebsiella pneumoniae to the body; 4) Improve the body's ability to resist Klebsiella pneumoniae infection; 5) Increase the expression of intestinal AhR signaling, IL-22 and / or intestinal tight junction proteins.

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