JP2017019833A - Clostridium difficile proliferation inhibitor - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a method capable of suppressing proliferation of Clostridium difficile, and achieving alleviation of symptoms due to the suppression of proliferation, and furthermore suppression or prevention of infectious disease due to the suppression of proliferation simply and highly effectively.SOLUTION: There are provided a Clostridium difficile proliferation inhibitor having a propionic acid bacterium and/or a fermentation product thereof as active ingredients, and a prebiotic composition having Clostridium difficile proliferation inhibitory action. Further, there are also provided a Clostridium difficile proliferation inhibitor having 1,4-dihydroxy-2-naphthoic acid (DHNA) and/or organic acid as active ingredients, and a prebiotic composition having Clostridium difficile proliferation inhibitory action, and furthermore a food composition containing them.SELECTED DRAWING: Figure 1

Description

  The present invention relates to a Clostridium difficile growth inhibitor, a prebiotic composition having a Clostridial difficile growth inhibitory action, a food and beverage composition comprising the same, and a Clostridial difficile growth inhibition method.

  Clostridium difficile is a major causative agent of diarrhea or enteritis caused by antibacterial drugs, and is well known as a causative agent of pseudomembranous colitis. Symptoms of Clostridium difficile infection range from mild diarrhea symptoms to severe pathological conditions such as intestinal obstruction. In addition, this infection often recurs and is difficult to treat. It is also important as a causative bacterium for nosocomial infections, and in-hospital outbreaks have been frequently reported in hospitalized cases and nursing home residents. Since treatment with drugs is difficult in the clinical field in such cases, there is a strong demand for symptom relief, infection control and prevention with foods and beverages and prebiotics that can be used particularly easily.

  So far, countermeasures against various Clostridium difficile infections have been studied. For example, Patent Document 1 discloses a method of suppressing mammalian infection caused by Clostridium difficile by administering a therapeutically effective amount of an undigestible oligosaccharide comprising fructooligosaccharide. Patent Document 2 discloses that a hop extract or the like is added or administered to a food or drink or a beverage to suppress the growth of Clostridium difficile, and further to treat or prevent a disease induced thereby. ing. Further, Patent Document 3 discloses a bacteriocin called thuricin CD having a Clostridial difficile growth inhibitory effect as a novel antibacterial compound against these diseases with the increase of nosocomial infectious diseases caused by Clostridium difficile and the like. .

US Pat. No. 5,688,777 Japanese Patent Laid-Open No. 11-222104 Special table 2011-505126 gazette

  However, in the technology disclosed in the above-mentioned patent document, prebiotics and prebiotic-derived components that inhibit the growth of Clostridium difficile while promoting the growth of useful bifidobacteria are disclosed. None of the cases reported to date have been known.

  Therefore, the development of a technology that can suppress the growth of Clostridium difficile and that can relieve the symptoms, and further suppress and prevent the infections by using a simpler method and with higher effects, continues. It was desired.

  The present inventors have conducted extensive research in view of the above-mentioned conventional problems. As a result, culture supernatants such as Propionibacterium fredenreich, particularly ET-3 strain and ATCC 9614T strain, can proliferate Clostridium difficile. It was found that it can be suppressed. In addition, 1,4-dihydroxy-2-naphthoic acid (DHNA) and / or organic acids, which are highly produced by Propionibacterium Freuddenreich ET-3 strain, ATCC 9614T strain, etc., remarkably inhibit the growth of Clostridium difficile. I found out.

  Therefore, the present invention provides a prebiotic composition that significantly suppresses the growth of Clostridium difficile without affecting useful bacteria such as bifidobacteria and lactic acid bacteria, a food and beverage composition comprising the same, and Clostridium A method for inhibiting growth of Clostridium difficile is provided.

A preferred embodiment of the present invention is a Clostridium difficile growth inhibitor containing propionic acid bacteria and / or a fermented product thereof as an active ingredient.
Another embodiment of the present invention is a prebiotic composition having a Clostridium difficile growth inhibitory action comprising propionic acid bacteria and / or a fermented product thereof.

  The propionic acid bacteria in the growth inhibitor or prebiotic composition include Propionibacterium Freudrenreich ET-3 (Propionibacterium fredenreichichi ET-3) or Propionibacterium fredenreich ATCC9614T (Propionibacterium freidenTre 14). Preferably there is.

A preferred embodiment of the present invention is a Clostridium difficile growth inhibitor containing 1,4-dihydroxy-2-naphthoic acid (DHNA) and / or an organic acid as an active ingredient.
Another aspect of the present invention is a prebiotic composition having an inhibitory activity against Clostridium difficile growth comprising 1,4-dihydroxy-2-naphthoic acid (DHNA) and / or an organic acid. .

  The 1,4-dihydroxy-2-naphthoic acid (DHNA) in the above growth inhibitor or prebiotic composition may be Propionibacterium fredenreich ET-3 or Propionibacterium. -It is preferable that it is what is obtained from the fermented product of Freudenreich ATCC9614T (Propionibacterium fredenreichii ATCC9614T), or synthetic DHNA. Those skilled in the art can appropriately synthesize DHNA by a known method. Examples of the organic acid include acetic acid, formic acid, and propionic acid, and propionic acid is particularly preferable.

  Yet another embodiment of the present invention is a food or beverage composition having an inhibitory effect on the growth of Clostridium difficile comprising an effective amount of propionic acid bacteria and / or a fermented product thereof.

  The propionic acid bacterium in the above-mentioned food / beverage composition is preferably Propionibacterium fredenreich ET-3 (Propionibacterium fredenreichii ET-3) or Propionibacterium fredenreich ATCC 9614T (Propionibacterium fredenreich 96).

  Still another aspect of the present invention is a food or beverage composition having an inhibitory effect on the growth of Clostridium difficile comprising an effective amount of 1,4-dihydroxy-2-naphthoic acid (DHNA) and / or an organic acid. It is a thing.

  The 1,4-dihydroxy-2-naphthoic acid (DHNA) in the above food / beverage product composition may be Propionibacterium fredenreich ET-3 or Propionibacterium fredenreiich ATCC 9614T ( It is preferably obtained from a fermented product of Propionibacterium fredenreichii ATCC 9614T).

  Still another aspect of the present invention is a method for inhibiting the growth of Clostridium difficile, characterized by ingesting or administering a prebiotic composition comprising an effective amount of propionic acid bacteria and / or a fermented product thereof. It is.

  As the propionic acid bacteria in the above-mentioned method for inhibiting the growth of Clostridium difficile, Propionibacterium fludenreich ET-3 (Propionibacterium fredenreichichi ET-3) or Propionibacterium fredenreich ATCC9614Tc It is preferable that

  Yet another aspect of the present invention is characterized in that a prebiotic composition comprising an effective amount of 1,4-dihydroxy-2-naphthoic acid (DHNA) and / or an organic acid is ingested or administered. It is a method for inhibiting the growth of Clostridium difficile. Examples of administration subjects include humans and non-human mammals. In addition, the above-described growth inhibition method of the present invention includes, for example, in vivo, in vitro, and ex vivo methods.

  As the 1,4-dihydroxy-2-naphthoic acid (DHNA) in the Clostridium difficile growth inhibition method, Propionibacterium fredenreichii ET-3 or Propionibacterium It is preferably obtained from a fermented product of Umm Freudenreich ATCC 9614T (Propionibacterium fredenreichii ATCC 9614T).

  The Clostridium difficile growth inhibitor of the present invention has an effect of remarkably suppressing the growth of Clostridium difficile without affecting useful bacteria such as bifidobacteria and lactic acid bacteria, as described later. Therefore, examples of the utilization mode of the growth inhibitor of the present invention include a mode in which the growth inhibitor is used in combination with an antibiotic or an antibacterial agent having an effect on the growth of useful bacteria such as bifidobacteria and lactic acid bacteria. That is, the growth inhibitor of the present invention is useful as a concomitant agent with antibiotics and antibacterial agents.

  Examples of antibiotics used in combination with the growth inhibitor of the present invention include lincomycin antibiotics, β-lactam antibiotics, aminoglycoside antibiotics, macrolide antibiotics, tetracycline antibiotics, chloramphenicol. Antibiotics, peptide antibiotics, fosfomycin, and the like. Examples of the antibacterial agents of the present invention include sulfa drugs, antituberculosis drugs, quinolone antibacterial drugs, new quinolone antibacterial drugs, antiviral drugs, and antifungal drugs. Can be mentioned. Since these antibiotics and antibacterial agents may cause the growth of Clostridium difficile, they are used together for the purpose of suppressing the growth of Clostridium difficile. It is also possible to use the Clostridium difficile growth inhibitor of the present invention in combination with the effective action of an antibiotic effective against Clostridium difficile such as metronidazole, vancomycin, and fidaxomicin. It is included in the scope of the invention.

The present invention also relates to the use of propionic acid bacteria and / or fermented products thereof for the production of Clostridium difficile growth inhibitors.
The invention further relates to the use of 1,4-dihydroxy-2-naphthoic acid (DHNA) and / or organic acids for the production of Clostridium difficile growth inhibitors.

The present invention also relates to propionic acid bacteria and / or fermented products thereof for use in inhibiting the growth of Clostridium difficile.
The present invention further relates to 1,4-dihydroxy-2-naphthoic acid (DHNA) for use in inhibiting the growth of Clostridium difficile.

  The prebiotic composition of the present invention promotes the growth of bifidobacteria, which are useful bacteria in the intestine, and does not affect the growth of lactic acid bacteria. Therefore, the prebiotic composition of the present invention including Propionibacterium flouddenreich ET-3 and ATCC9614T and DHNA promotes the growth of bifidobacteria and the like, while Clostridium difficile ( The growth of Clostridium difficile can be suppressed. Therefore, the present invention can provide a very useful prebiotic composition having a Clostridial difficile growth inhibitory action, a food and beverage composition comprising the same, and a Clostridial difficile growth inhibitory method.

It is a graph which shows the growth inhibitory activity of Clostridium difficile of the prebiotic composition of this invention. It is a graph which shows the growth inhibitory activity of Clostridium difficile of the prebiotic composition of this invention. It is a graph which shows the influence which the prebiotic composition of this invention has on the proliferation of Bifidobacterium (Bifidobacterium longum OLB6001 strain). It is a graph which shows the influence which the prebiotic composition of this invention has on the growth of lactic acid bacteria (L. bulgaricus). It is a graph which shows the influence which the prebiotic composition of this invention has on the growth of lactic acid bacteria (S. thermophilus OLS3059 strain). It is a graph which shows the influence which the propionic acid of this invention has on the proliferation of Clostridium difficile. This is a copy of the deposit certificate with the deposit number FERM BP-8115 (Propionibacterium Freudenreich ET-3 strain). It is a copy of the certificate regarding the survival of the accession number FERM BP-8115 (Propionibacterium Freudenreich ET-3 strain).

  Hereinafter, the present invention will be described in detail, but the present invention is not limited to the individual forms described below.

Propionic acid bacteria have long been used as starters for Emmental cheese and the like, and their safety has been confirmed. Examples of propionic acid bacteria contained in the Clostridium difficile growth inhibitor or the prebiotic composition according to the present invention include Propionibacterium Freudenreich ET-3, Propionibacterium Freuden Reich ATCC 9614T (Propionibacterium fredenreichii ATCC 9614T), Propionibacterium fredenreich IFO 12424 (Propionibacterium fredenreichii piopro Ipi 12424), Propionibacterium ipro 12 Examples include microorganisms belonging to the genus Cuterium. Of these, the Propionibacterium freudenreich ET-3 strain and the Propionibacterium freudenreich ATCC 9614T strain are particularly preferable.
These bacterial cells may be used alone or in combination.

  The propionic acid bacterium and / or the fermented product thereof used in the Clostridium difficile growth inhibitor or the prebiotic composition according to the present invention can be obtained by culturing the aforementioned Propionibacterium microorganism according to a conventional method. The obtained fermented product itself and / or a processed product thereof can be used as a growth inhibitor or prebiotic composition according to the present invention. The treated product is, for example, a fermented product containing microbial cells, a filtrate obtained by filtering or sterilizing the fermented product, an extract extracted with a predetermined solvent, an evaporator or the like to obtain a fermented product, its filtrate, extract or supernatant. Concentrated concentrates and those obtained through processing steps such as dried products (freeze-dried, etc.) can be used.

  The preferred amount of propionic acid bacteria and / or fermented product thereof varies depending on the concentration of the fermented product. For example, in the case where a fermented propionic acid bacterium produced by the method described in Example 1 of International Publication Pamphlet No. WO2005 / 033323 is concentrated to a DHNA concentration of 100 μg / ml, the growth inhibitor of the present invention or It is 0.1 to 10 weight% with respect to a prebiotic composition, Preferably it is 0.3 to 5 weight%.

  As the 1,4-dihydroxy-2-naphthoic acid (DHNA) contained in the Clostridium difficile growth inhibitor or the prebiotic composition according to the present invention, either chemical synthesis or biosynthesis can be used, but preferably Is due to biosynthesis. In the present invention, it is particularly preferable to use DHNA which preferably produces Propionibacterium furedenreich ET-3 strain or Propionibacterium furedenreich ATCC9614T (Propionibacterium fredenreichiii ATCC9614T). For example, the fermentation product itself obtained by culturing a Propionibacterium microorganism according to a conventional method and / or a processed product thereof can be used as DHNA. The treated product is, for example, a fermented product containing bacterial cells, a filtrate obtained by filtering or sterilizing the fermented product, a crude product such as an extract extracted with a predetermined solvent, a purified product thereof, a fermented product by an evaporator, etc. Concentrates obtained by concentrating the filtrate / extract / supernatant liquid and those obtained through processing steps such as dried products (freeze-dried, etc.) can be used. The concentration of DHNA can be appropriately adjusted according to the use and the like, but is 1 to 250 μg / ml, preferably 1 to 5 μg / ml.

  This invention also provides the food-drinks composition containing a prebiotic composition.

  The food / beverage composition containing the prebiotic composition of the present invention is specifically a food / beverage composition, such as milk, soft drink, fermented milk (yogurt, yogurt drink), cheese, bread, biscuits, crackers. Even if the prebiotic composition of the present invention is added to a pizza crust, prepared milk powder, liquid food, food for the sick, food for infants, food such as milk powder for nursing women, nutritional food, etc. Good. In addition, the properties of the foods and drinks usually used, for example, solid (powder, granule, etc.), paste, liquid or suspension, liquid food, jelly, tablet, granule, capsule It can take any form of food or drink that can normally be distributed. Among these, it is preferable to provide the food / beverage composition of this invention with the form of a drink from the point which can be ingested easily, an absorptive point, etc.

  Other components are not particularly limited, but when the prebiotic composition of the present invention is used in a food or drink composition, water, protein, carbohydrates, lipids, vitamins, minerals, organic acids, Organic bases, fruit juices, flavors and the like can be used as main components. Examples of the protein include whole milk powder, skim milk powder, partially skimmed milk powder, casein, whey powder, whey protein, whey protein concentrate, whey protein isolate, α-casein, β-casein, κ-casein, β-lactoglobulin , Α-lactalbumin, lactoferrin, soy protein, egg protein, meat protein and other animal and vegetable proteins, hydrolysates thereof, butter, whey minerals, cream, whey, non-protein nitrogen, sialic acid, phospholipid, lactose, etc. And various milk-derived components. Examples of the saccharide include general saccharides, modified starch (in addition to dextrin, soluble starch, British starch, oxidized starch, starch ester, starch ether, etc.), dietary fiber, and the like. Examples of the lipid include animal oils such as lard, fish oil, etc., fractionated oils, hydrogenated oil, transesterified oil, etc .; palm oil, safflower oil, corn oil, rapeseed oil, coconut oil, fractionated oils thereof, Examples thereof include vegetable oils such as hydrogenated oils and transesterified oils. Examples of vitamins include vitamin A, carotene, vitamin B group, vitamin C, vitamin D group, vitamin E, vitamin K group, vitamin P, vitamin Q, niacin, nicotinic acid, pantothenic acid, biotin, inositol, choline. And minerals include, for example, calcium, phosphorus, potassium, magnesium, chlorine, sodium, copper, iron, manganese, zinc, selenium, chromium, molybdenum, vanadium, and whey mineral. Examples of the organic acid include malic acid, citric acid, lactic acid, tartaric acid, and the like. These components can be used singly or in combination of two or more, and a synthetic product and / or a food or drink containing a lot of these may be used.

  The said food-drinks composition can be manufactured in accordance with a conventional method by mixing the prebiotic composition of this invention and the appropriate quantity of the said required component.

The content of the prebiotic composition in the food / beverage product composition of the present invention can be arbitrarily determined according to its purpose, application and the like. Although this invention is not limited to this, As content, it is 5 weight% or more normally with respect to the whole quantity, More preferably, it is 10 weight% or more. By ingesting or administering this, it is possible to easily inhibit the growth of Clostridium difficile, and as a result, the symptoms can be alleviated and the infection can be suppressed or prevented.
It should be noted that all prior art documents cited in the present specification are incorporated herein by reference.

  EXAMPLES Hereinafter, although an Example is given and this invention is demonstrated further in detail, this invention is not limited by this. In the examples, “%” indicates weight% unless otherwise specified.

[Example 1]
-Preparation of culture supernatant of propionic acid bacteria that is the prebiotic composition of the present invention Propionibacterium freudenreich ET-3 strain (Incorporated administrative agency, National Institute of Advanced Industrial Science and Technology, as of August 9, 2001) The deposit center (deposited as FERM BP-8115 at 1st, 1st East, 1st Street, Tsukuba, Ibaraki, Japan) (postal code 305-8656)), Propionibacterium Freudrenreich ATCC 9614T (Propionibacterium) Um Freudenreich reference strain, purchased from American Type Culture Collection (ATCC)), Propionibacterium Freudenreich strain (No. 32-1) isolated from commercially available Emmental cheese for 48 hours under anaerobic conditions at 30 ° C And 3 more after pre-culture ℃ were cultured for 72 hours under anaerobic conditions. The culture is centrifuged (centrifugation conditions: 8000 rpm, 4 ° C., 10 minutes), the culture supernatant is recovered, filter sterilized, and the obtained culture supernatant is used as the prebiotic composition of the present invention at −80 ° C. Saved with.

Cultivation of Clostridium difficile The Clostridium difficile JCM1296T strain (Clostridial difficile reference strain) was purchased from the RIKEN BioResource Center and restored according to the attached manual.

Inhibition test 1% inoculation was carried out on BHI medium previously maintained in an anaerobic state with Clostridium difficile prepared as described above. To this, the prebiotic composition of the present invention or DHNA was added and cultured under anaerobic conditions at 37 ° C. for 24 hours. After thoroughly stirring the culture solution, OD ₆₅₀ was measured (Shimadzu spectrophotometer BioSpec 1600, manufactured by Shimadzu Corporation). The results are shown in FIG. 1 as a graph. In addition, about the obtained data, the multiple comparison test by Tukey-Kramer method was performed, and it was set as each group n = 3, * p <0.05, ** p <0.01. In addition, about Propionibacterium freudenreich ATCC9614T and Propionibacterium freudenreich ET-3, it was set as p <0.01 with respect to the control group.

[result]
In the prebiotic composition 5% the added system of the present invention, since the OD ₆₅₀ after 24 hours of culture was significantly reduced, the prebiotic composition growth inhibitory effectively Clostridium difficile of the invention I found out that

[Example 2]
-Preparation of DHNA contained in prebiotic composition of the present invention 100 mg of DHNA (manufactured by Sigma-Aldrich) was dissolved in 0.5 ml of DMSO to prepare a 1M DMSO solution. After 10-fold dilution with BHI medium to 100 mM, filter sterilization was performed. At the same time, a solution in which DMSO was diluted 10-fold with BHI medium was also prepared and sterilized by filter. All subsequent dilutions were performed with a 10-fold diluted DMSO solution so that 10% DMSO was included in the DHNA solution at all concentrations. Since 0.1% of these DHNA solutions were added to the bacterial culture as the prebiotic composition of the present invention, the final concentration of DMSO was 0.01%.

-Inhibition test The inhibition test similar to Example 1 was done about the growth inhibitory effect of the prebiotic composition of this invention with respect to Clostridium difficile used in Example 1. FIG. Specifically, the prebiotic composition of the present invention containing DHNA was added to each sample at each concentration (1 μM, 10 μM, 20 μM, 50 μM). The obtained results are shown in the graph of FIG. In addition, a multiple comparison test by the Tukey-Kramer method was performed, and each group was set to n = 3 and * p <0.05. In addition, about 10-50M, it was set as ** p <0.01 with respect to DMSO group.

  As shown in FIG. 2, it was shown that the growth of Clostridium difficile was markedly inhibited by the addition of 10 μM or more of DHNA.

[Example 3]
-Preparation of culture supernatant of bifidobacteria and lactic acid bacteria longum OLB6001 strain (FERM P-13610) and "Meiji Bulgaria Yogurt" (trade name: manufactured by Meiji Dairies). bulgaricus was cultured under anaerobic conditions at 37 ° C. for 18 hours. S. thermophilus OLS3059 strain (FERM P-15487) was cultured for 30 hours under anaerobic conditions at 30 ° C. The culture was centrifuged (centrifugation conditions: 8000 rpm, 4 ° C., 10 minutes), the culture supernatant was collected, filter sterilized, and the obtained culture supernatant was stored at −80 ° C. as a comparative composition ( Lactic acid bacteria are used in Example 4).

Effect of the prebiotic composition of the present invention on the growth of bifidobacteria Bifidobacterium longum OLB6001 strain added with DHNA or 0.01% DMSO used in Example 2 (control), and anaerobic conditions at 37 ° C. for 12 hours Cultured. After thoroughly stirring the culture solution, OD ₆₅₀ was measured (Shimadzu spectrophotometer BioSpec 1600, manufactured by Shimadzu Corporation). The results are shown in FIG. 3 as a graph. In addition, a multiple comparison test by the Tukey-Kramer method was performed, and each group was set to n = 3 and * p <0.05. In addition, about 10-50M, it was set as ** p <0.01 with respect to DMSO group.

[result]
It has been found that the addition of the prebiotic composition containing DHNA of the present invention significantly promotes the growth of OLB6001 strain. That is, it was shown that the prebiotic composition of the present invention does not affect useful bacteria but rather has a growth promoting effect. Therefore, it was found that when the growth of Clostridium difficile is inhibited, useful bacteria are not affected by the growth (rather, they have a growth promoting effect).

[Example 4]
Effect of the prebiotic composition of the present invention on the growth of lactic acid bacteria The prebiotic composition containing the DHNA of the present invention (DHNA concentration: 1 μM to 50 μM) used in Example 3 bulgaricus and S. In order to examine the influence on the thermophilus OLS3059 strain, the same test as in Example 3 was performed, and the results are shown in FIGS. As in Example 3, the addition of a prebiotic composition containing DHNA of the present invention resulted in L.P. bulgaricus and S. It was found that there is no effect on the growth of the thermophilus OLS3059 strain. Therefore, it was found that the useful bacteria have no influence on the growth when inhibiting the growth of Clostridium difficile.
[Example 5]
-Effect of propionic acid of the present invention on the growth of Clostridium difficile At the same time, 1% of a culture solution of Clostridium difficile (C. difficile JCM1296 strain) cultured for 24 hours and grown to OD ₆₅₀ of 1.2 was added to the culture Various organic acids were added to a final concentration of 100 μM, and the OD ₆₅₀ value after 24 hours was measured. The results are shown in FIG. 6 as a graph.
As shown in FIG. 6, it was shown that the growth of Clostridium difficile was inhibited by the addition of propionic acid.

  The Clostridium difficile growth inhibitor or prebiotic composition according to the present invention does not affect or is effective against the growth of bifidobacteria and lactic acid bacteria, which are useful bacteria in the intestine. It can inhibit the growth of certain Clostridium difficile. Therefore, based on the prebiotic composition of the present invention containing the Propionibacterium Freuddenreich ET-3 and ATCC9614T strains and DHNA, etc., a very useful prebiotic composition having a Clostridial difficile growth inhibitory action and It is possible to provide a food / beverage composition comprising the same and a method for inhibiting Clostridium difficile growth, and the industrial applicability is very high.

Claims (9)

  A Clostridium difficile growth inhibitor comprising a propionic acid bacterium and / or a fermented product thereof as an active ingredient.   The propionic acid bacterium is Propionibacterium fredenreich ET-3 (Propionibacterium fredenreichii ET-3) or Propionibacterium fredenreich ATCC 9614T (Drug of Propionibacterium fredenreichii ATCC9614T).   A Clostridium difficile growth inhibitor comprising 1,4-dihydroxy-2-naphthoic acid (DHNA) and / or an organic acid as an active ingredient.   The 1,4-dihydroxy-2-naphthoic acid (DHNA) is from Propionibacterium fredenreich ET-3 (Propionibacterium fredenreichii ET-3) or Propionibacterium fredenreich ATCC9614T (Propionibacterium fredenreich 96 fermentation) The medicine according to claim 3, which is obtained.   The combined use with an antibiotic or an antimicrobial agent which comprises the chemical | medical agent in any one of Claims 1-4.   A method for inhibiting the growth of Clostridium difficile, comprising ingesting or administering a prebiotic composition comprising an effective amount of propionic acid bacteria and / or a fermented product thereof.   The propionic acid bacterium is Propionibacterium fredenreich ET-3 (Propionibacterium fredenreichii ET-3) or Propionibacterium furedenreich ATCC 9614T (propionbacterium fredenreichii ATCC9614T) A method for inhibiting the growth of Clostridium difficile.   A method of inhibiting the growth of Clostridium difficile comprising ingesting or administering a prebiotic composition comprising an effective amount of 1,4-dihydroxy-2-naphthoic acid (DHNA) and / or an organic acid .   The 1,4-dihydroxy-2-naphthoic acid (DHNA) is from Propionibacterium fredenreich ET-3 (Propionibacterium fredenreichii ET-3) or Propionibacterium fredenreich ATCC9614T (Propionibacterium fredenreich 96 fermentation) The method for inhibiting the growth of Clostridium difficile according to claim 8, wherein the method is obtained.

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