Faecalibacterium prausnitzii strain CNCM I-4573 for the treatment and prevention of Clostridioides difficile infections

Abstract

The present invention relates to a bacterial strain of the species Faecalibacterium prausnitzii deposited at the CNCM under accession number CNCM I-4573 and / or its culture supernatant for use in the treatment and / or prevention of Clostridioides difficile infection in an individual.

Description

[Technical Field]

[0001] FIELD OF THE INVENTION The present invention relates to a Faecalibacterium prausnitzii strain for use in the treatment and prevention of Clostridioides difficile infection in an individual.

[0002] prior art Clostridioides difficile (also known as C. difficile and formerly known as Clostridium difficile) is an anaerobic, spore-forming, toxin-producing, Gram-positive bacterium transmitted between humans via the fecal-oral route. Elderly, immunocompromised individuals whose gut microbiota has been disrupted by antibiotic therapy are at greatest risk for this highly contagious, life-threatening, and potentially fatal diarrheal disease. After hypervirulent strains began to emerge in the early 2000s, C. difficile emerged as a major, globally distributed enteric pathogen.

[0003] Generally, C. difficile is considered a hospital-acquired nosocomial pathogen, although C. difficile infection can occur outside the hospital setting.

[0004] However, the disease is now emerging more frequently in individuals once considered low-risk. These cases tend to occur in younger individuals. Recent antibiotic exposure is an important risk factor in this population, but use of gastric acid-suppressing medications and comorbid conditions such as inflammatory bowel disease, chronic kidney disease, immunodeficiency, malignancy, and organ transplantation are also associated with C. difficile (CDI). In some rare cases, infection can even emerge spontaneously in otherwise healthy individuals in the absence of antibiotic exposure.

[0005] C. difficile is responsible for 15-25% of cases of antibiotic-associated bacterial diarrhea and is considered the leading cause of healthcare-associated diarrhea in adults. After an initial episode, the risk of first recurrence is high and problematic (approximately 20%), and this increases further after the first recurrence, reaching up to 60%. Symptoms associated with this infection can range from simple diarrhea to pseudomembranous colitis.

[0006] Accordingly, C. difficile has been classified by the Centers for Disease Control and Prevention (CDC) as one of the top three urgent threats of antibiotic resistance, and CDI has become the most common cause of antibiotic-associated diarrhea and gastroenteritis-related death in developed countries.

[0007] Currently, the standard treatment for CDI is antibiotics, with vancomycin or fidaxomicin being the most commonly prescribed compounds. Metronidazole is used only when vancomycin or fidaxomicin are unavailable. Fecal microbial transplantation (FMT) is used only as a last resort after multiple recurrences. Probiotics have also been used as a preventative treatment, but with mixed results, and a vaccine is not yet available. Within healthcare facilities, preventative measures include the judicious use of antibiotics and the implementation of infection control measures. The use of monoclonal antibodies has also been investigated with the aim of improving host resistance to CDI, but this approach has produced mixed results.

[0008] There is a need for new substances, particularly probiotics, or compositions containing same, for the treatment and / or prevention of C. difficile infection.

[0009] In particular, there is a need for novel substances, particularly probiotics, or compositions containing the same, that can reduce the risk of C. difficile infection after perturbation of the intestinal flora with antibiotics and / or proton pump inhibitors (PPIs).

[0010] In particular, there is a need for novel substances, particularly probiotics, or compositions comprising same, that are capable of reducing the risk of recurrence of C. difficile infection after a given episode.

[0011] There is a need for new substances, particularly probiotics, or compositions containing same, that are capable of delaying intestinal colonisation by C. difficile.

[0012] There is a need for new substances, in particular probiotics, or compositions comprising same, which allow the reduction or even absence of symptoms after infection with C. difficile.

[0013] There is a need for new substances, particularly probiotics, or compositions containing same, that allow for bacterial colonization of the intestinal tract, more rapid clearance of C. difficile from the intestinal tract, and also advantageously reduce or even eliminate symptoms following infection.

[0014] Summary of the Invention It is an object of the present invention to provide novel substances, in particular probiotics, and compositions for the treatment and / or prevention of Clostridioides difficile infection in individuals.

[0015] In the context of the present invention, the terms "prevent", "prevention" and "preventing" refer to a reduction in the risk or probability of a given phenomenon, i.e., in the present invention, a C. difficile infection, in particular the occurrence of symptoms after infection with C. difficile. In particular, the terms "prevent", "prevention" and "preventing" refer to a reduction in the risk or probability of a first episode of C. difficile infection (CDI) in an individual and / or a reduction in the risk or probability of a recurrence of C. difficile infection. For the prevention of a first episode, the target population may correspond in particular to any patient prescribed a treatment including antibiotic treatment and / or administration of a proton pump inhibitor (PPI), in particular any patient prescribed antibiotic treatment.

[0016] Recurrent CDI was defined as the recurrence of diarrhea that tested positive for C. difficile toxin within 8 weeks of completing effective anti- C. difficile therapy.

[0017] As used herein, the terms "treating," "treat," or "treatment" include not only the reduction and / or elimination of symptoms associated with a particular disorder or condition, but also the complete elimination of the disorder or condition under consideration.

[0018] The present invention is based on the inventors' discovery of the ability of the Faecalibacterium prausnitzii strain deposited at the CNCM under accession number CNCM I-4573 to prevent and / or treat Clostridioides difficile infection in individuals.

[0019] The results of the inventors' experiments have shown that a particular strain of Faecalibacterium prausnitzii (F. prausnitzii), deposited at the CNCM by the French National Institute for Agricultural Research (INRA) on January 21, 2012 under accession number CNCM I-4573, has the unexpected ability to treat and prevent infection of an individual with C. difficile in vivo, in particular the unexpected ability to delay colonization of the intestinal tract of the individual by C. difficile, to rapidly clear C. difficile from the intestinal tract, and to reduce symptoms following infection with C. difficile, or even to prevent the appearance of any symptoms following infection with C. difficile.

[0020] According to a first subject, the present invention relates to a bacterial strain of the species Faecalibacterium prausnitzii deposited at the CNCM under accession number CNCM I-4573 and / or its culture supernatant for use in the treatment and / or prevention of Clostridioides difficile infection in individuals.

[0021] Therefore, this strain identified by the inventors is a probiotic strain that can be used for the applications presented herein.

[0022] The individual according to the present invention may be selected from the group consisting of humans; Suidae, particularly pigs, more particularly piglets; Bovinae, particularly cattle, more particularly bulls, more particularly steers; Canidae, particularly dogs; Equidae, particularly horses; and Phasianidae, particularly poultry, more particularly chickens; preferably mammals, more particularly humans (J. Scott Weese; Journal of Veterinary Diagnostic Investigation 2020, Vol. 32(2) 213-221).

[0023] The bacterial strain of the species Faecalibacterium prausnitzii deposited at the CNCM under accession number CNCM I-4573 may be administered to the individual in live, inactive and / or dead form, preferably in live or inactive form, more particularly in live form.

[0024] Viable bacterial forms refer to culturable and / or metabolically active bacterial strains, in particular culturable and metabolically active bacterial strains.

[0025] The bacterial strain in inactive form according to the invention may be, for example, a lyophilisate prepared using techniques well known to those skilled in the art.

[0026] In some embodiments, Clostridioides difficile infection may occur after an individual has received at least one antibiotic drug therapy and / or at least one proton pump inhibitor (PPI) drug therapy. The bacterial strain of Faecalibacterium prausnitzii deposited in the CNCM under accession number CNCM I-4573 and / or its culture supernatant may be administered simultaneously and / or separately, particularly simultaneously, with at least one antibiotic drug therapy and / or at least one PPI drug therapy. The bacterial strain of Faecalibacterium prausnitzii deposited in the CNCM under accession number CNCM I-4573 and / or its culture supernatant may be administered simultaneously and / or after at least one antibiotic drug therapy and / or at least one PPI drug therapy.

[0027] The bacterial strain and / or its culture supernatant for its use according to the invention may be comprised in a composition comprising a physiologically acceptable medium, preferably in an oral or rectal composition, more particularly preferably in a pharmaceutical product.

[0028] The term "physiologically acceptable medium" is intended to mean a medium that is compatible with the body of the individual to which the composition must be administered. This is, for example, a non-toxic solvent such as water. In particular, the solvent is compatible with oral administration.

[0029] The compositions of the present invention are preferably for oral or rectal administration, especially for oral administration.

[0030] The composition for oral administration of the present invention may be selected from the group consisting of a food, a beverage, a pharmaceutical, a nutraceutical, a food additive, a supplement and a dairy product, and is particularly in the form of a pharmaceutical.

[0031] The composition for use according to the invention comprises, as a pharmaceutical, 10% or more of the F. prausnitzii strain of the invention. 4 ~10 15 Total cell count (TCC), more preferably 10 8 ~l0 1310 of the bacterial strain according to the present invention in a daily dose equivalent to TCC. 13 It may be suitable for administration as a daily dose equivalent to TCC. [Brief explanation of the drawings]

[0032] [Figure 1] Figure 1 shows the evaluation of weight loss in mice treated or not with F. prausnitzii I-4573 according to the invention. Mice were weighed and monitored for the occurrence of clinical signs after infection with C. difficile. Two groups of mice are examined: mice treated with PBS-glycerol 16% (vehicle); and mice treated with the F. prausnitzii I-4573 strain. Abscissa: day (D0-D20, D0 being the day when C. difficile R20291 was administered to mice previously treated with antibiotics). Ordinate: mouse weight (% compared to weight at D0). [Figure 2] Figure 1 shows the 7-day survival rate of mice treated with PBS-glycerol 16% (vehicle) or with F. prausnitzii strain I-4573. Abscissa: days (D0-D7). Ordinate: probability of survival (0-1). [Figure 3] Colonization of mice (10 per group) with C. difficile R20291 over time (D0-D20, where D0 is the day antibiotic-pretreated mice were given C. difficile R20291). Abscissas (arrows) are, from left to right: D0, D2, D3, D4, D5, D6, D8, D10, D14, and D20. Ordinate: % of mice with detectable C. difficile in the feces.

[0033] Detailed Description of the Invention The present inventors have conducted extensive research to identify the ability of specific strains of Faecalibacterium prausnitzii to treat and / or prevent Clostridioides difficile (C. difficile) infection in individuals.

[0034] Indeed, the inventors have unexpectedly demonstrated that the F. prausnitzii I-4573 strain not only delays intestinal colonization by C. difficile in individuals, but also has the ability to rapidly clear C. difficile from the intestinal tract and significantly reduce symptoms after infection with C. difficile, and in particular prevent the appearance of any symptoms after infection with C. difficile.

[0035] The Faecalis bacterium prausnitzii strain of the present invention F. prausnitzii is a major member of the phylum Firmicutes and is some of the most abundant commensal bacteria in the healthy human large intestinal microbiota.

[0036] F. prausnitzii is an extremely oxygen-sensitive (EOS) bacterium, making it difficult to cultivate, even under anaerobic conditions (Duncan et al. 2002, Int. J. Syst. Evol. Microbiol. 52(Pt 6): 2141-6 and Lopez-Siles et al. Appl. Environ Microbiol. January 2012; 78 (2):420-8). F. prausnitzii is particularly known to be one of the most abundant butyrate-producing bacteria in the human gastrointestinal tract, and butyrate, a short-chain fatty acid, is crucial for intestinal physiology, systemic function, and beneficial effects on human health (Macfarlane and Macfarlane (2011), J. Clin. Gastroenterol. 45 Suppl: S120-7).

[0037] A suitable daily dose of the bacterial strain according to the present invention is 10 mg / day when the F. prausnitzii strain of the present invention is used as a medicine. 7 ~10 14 It may be colony forming units (cfu).

[0038] The daily dose is also 10 mg / day when the F. prausnitzii strain of the present invention is used as a medicine. 4 ~10 15 Total cell count (TCC), preferably 10 8 ~l013 TCC range, e.g. 10 13 It may be the equivalent daily dose of TCC.

[0039] The F. prausnitzii strain specifically embodied in the present invention is a bacterial strain of the species Faecalibacterium prausnitzii deposited at the CNCM under accession number CNCM I-4573.

[0040] As previously mentioned, the Faecalibacterium prausnitzii strain and / or its culture supernatant of the present invention are for use in the treatment and / or prevention of Clostridioides difficile infection in an individual.

[0041] The Faecalibacterium prausnitzii bacterial strain deposited at the CNCM under accession number I-4573 for use according to the present invention may be in an isolated form. By "isolated form" according to the present invention, we mean a form isolated from any fecal material or intestinal sampling. Therefore, it is preferred that the bacterial strain of the present invention is not administered to patients in the form of fecal microbial transplant (FMT). Methods for isolating bacteria from the intestinal microbiota, such as those exemplified by Foditsch et al. (PLoS One. 2014; 9(12): e116465), are well known to those skilled in the art.

[0042] The individual according to the present invention may be selected from the group consisting of humans; Suidae, particularly pigs, more particularly piglets; Bovidae, particularly cattle, more particularly bulls, more particularly steers; Canidae, particularly dogs; Equidae, particularly horses; and Phasianidae, particularly poultry, more particularly chickens. In particular, the individual according to the present invention is a mammal, in particular a human.

[0043] The strains of the present invention are probiotics, whose activity is located in the intestinal tract. Probiotic bacteria in the present invention refer to bacteria that can exert a beneficial effect on the health of an individual when ingested live in sufficient amounts.

[0044] As a result, the bacterial strain of the species Faecalibacterium prausnitzii deposited at the CNCM under accession number CNCM I-4573 may be administered to an individual in need thereof in live, inactivated and / or killed form, preferably administered intraintestinally in live form.

[0045] The bacterial strains of the present invention may be administered to the gastrointestinal tract of the individual to be treated in various ways, namely orally or rectally. The bacteria according to the present invention are preferably administered orally.

[0046] According to a preferred embodiment, the bacterial strain of the invention is contained in a composition comprising a physiologically acceptable medium, such a composition being preferably for oral or rectal administration, in particular for oral administration, and in particular in pharmaceutical form.

[0047] composition The present invention also relates to a composition for use according to the invention, which comprises at least the Faecalibacterium prausnitzii bacterial strain I-4573 of the invention in a physiologically acceptable medium.

[0048] The compositions for use according to the present invention are intended for the digestive tract, particularly the intestinal tract.

[0049] Consequently, the compositions for use according to the invention may be selected from oral or rectal compositions. The compositions of the invention are preferably oral or rectal compositions, more preferably oral compositions.

[0050] Compositions for use in the present invention that are oral compositions are intended for oral administration to an individual.

[0051] Compositions for use in the present invention that are rectal compositions are intended for rectal administration to an individual.

[0052] Oral compositions may be in the form of suspensions, tablets, pills, capsules, granules or powders.

[0053] The composition for use according to the invention for oral administration may be selected from the group consisting of a food, a beverage, a pharmaceutical, a nutraceutical, a food additive, a supplement or a dairy product, in particular a pharmaceutical.

[0054] According to one preferred embodiment, the composition for use according to the invention is a pharmaceutical product.

[0055] Pharmaceuticals for oral administration may be present in capsules, gel capsules, soft capsules, tablets, dragees, pills, pastes, lozenges, gums, oral liquids or emulsions, syrups, or gels.

[0056] For example, the medicinal products according to the invention for oral administration may be in capsules, in particular in the form of a lyophilisate, preferably together with pharmaceutically acceptable excipients.

[0057] Advantageously, compositions for use according to the invention intended for oral administration may be provided with a gastric acid resistant coating to ensure that the bacterial strains of the invention contained therein are able to pass through the damaged stomach, so that release of the bacterial strains may occur initially in the upper gastrointestinal tract.

[0058] Medicaments for oral use according to the invention may also contain sweeteners, stabilizers, antioxidants, additives, flavorings and / or coloring agents.

[0059] The formulation is carried out by a conventional method for producing sugar-coated tablets, gel capsules, gels, sustained-release hydrogels, emulsions, tablets or capsules.

[0060] The compositions for use containing the bacterial strains of the present invention may be administered rectally.

[0061] Rectal administration can especially be carried out in the form of suppositories, enemas or foams, especially enemas.

[0062] The composition for use according to the invention may also be in the form of a nutritional composition.

[0063] Nutritional compositions for use according to the present invention may be in the form of yogurt, cereal bars, breakfast cereals, desserts, frozen meals, soups, pet food, suspensions, powders, tablets, gums or candies.

[0064] In particular, the composition for use according to the invention, when used as a medicine, contains 10% or more of the strain of the invention. 7 ~10 14 A daily dose equivalent to colony forming units (cfu), preferably 10 11 It may be appropriate to administer a daily dose equivalent to cfu.

[0065] For example, a composition for use according to the present invention may be administered to an individual in need thereof by administering 10 doses of the I-4573 bacterial strain of the present invention. 7 ~10 14 cfu, preferably 10 11 It can be administered in a single daily dose of 1 g containing an amount equivalent to a dose of cfu.

[0066] In another example, the composition for use according to the present invention may comprise administering to an individual in need thereof 10 doses of the I-4573 bacterial strain of the present invention. 7 ~10 14 cfu, preferably 10 11 It can be administered in a single daily dose of 0.2 g containing an amount equivalent to the amount of cfu.

[0067] As previously mentioned, the daily dosage of the F. prausnitzii strain of the present invention when used as a medicine is also 10 total cell counts (TCC). 4 ~10 15 and more preferably in the range of 10 8 ~10 13 The daily dose may be in the range of TCC, for example, 10 13 An amount equivalent to TCC is included.

[0068] The composition for use according to the invention may be administered to an individual in need thereof 1 to 10 times daily, each dose containing, for example, 5.10 mg of the I-4573 bacterial strain of the invention. 8 ~5.1011 By including the amount of TCC, the total daily dose of the I-4573 bacterial strain of the present invention administered to an individual is as described above.

[0069] In particular, the compositions for use according to the present invention may be administered to an individual for 1 day, 2 days, 3 days, 4 days, 5 days, 6 days, 1 week, 10 days, 14 days, 15 days, 3 weeks, 1 month, or more than 1 month. When the compositions for use according to the present invention are administered for more than 1 day, the same daily dose of the F. prausnitzii strain of the present invention may be administered every day, or the daily dose may vary over the course of treatment. In particular, the daily dose of the F. prausnitzii strain of the present invention may be higher at the beginning of treatment and then gradually decreased.

[0070] Compositions according to the present invention may also include at least one of antioxidants, fish oil, DHA, EPA, vitamins, minerals, phytonutrients, proteins, lipids, probiotics, and combinations thereof.

[0071] Thus, an individual according to the invention: - Individuals who will be starting or have already started antibiotic treatment; and / or - Individuals completing or who have just completed antibiotic treatment; and / or - Individuals who are initiating or have already started proton pump inhibitor (PPI) therapy; and / or - Individuals completing or who have just completed proton pump inhibitor (PPI) therapy; and / or - an individual who has been treated with, e.g., vancomycin and / or fidaxomicin, for an episode of C. difficile infection, e.g., a first episode of C. difficile infection; and / or - Individuals with a first episode of C. difficile infection or recurrent C. difficile infection It could be.

[0072] The Clostridioides difficile infection to be treated and / or prevented according to the present invention may in particular occur after the use of at least one antibiotic and / or PPI medication by the individual considered. Thus, the individual according to the present invention may in particular be an individual who is starting or has already started antibiotic and / or PPI treatment or an individual with dysbiosis.

[0073] Thus, the bacterial strain, its culture supernatant or a composition comprising same for use according to the invention may be administered simultaneously and / or separately with at least one antibiotic drug therapy (antibiotic) and / or at least one PPI drug therapy of the antibiotic treatment, in particular simultaneously, e.g. separately on the same day, or even with at least one or even more days between the administration of the bacterial strain and the administration of the antibiotic / PPI drug therapy.

[0074] In a particular embodiment, the bacterial strain for use according to the invention or its culture supernatant and the at least one antibiotic and / or PPI are administered to the individual in the same composition.

[0075] In a particular embodiment, the bacterial strain for use according to the invention or its culture supernatant and the at least one antibiotic and / or PPI are administered as separate compositions.

[0076] When the bacterial strain or its culture supernatant for use according to the invention and the at least one antibiotic and / or PPI are administered as separate compositions, the compositions can be administered to an individual simultaneously or separately via the same route or different routes.

[0077] At the same time, it is understood that the bacterial strain or its culture supernatant for use according to the present invention, or a composition comprising same, can be administered at the same time as the antibiotic and / or PPI, or at most on the same day or 2-3 days thereafter.

[0078] By separately it is understood that the bacterial strain or its culture supernatant for use according to the invention, or the composition comprising same, can be administered at least several days apart, for example at least 3 days apart.

[0079] The bacterial strain for use according to the invention or its culture supernatant or a composition comprising same may be administered simultaneously and separately with at least one antibiotic drug therapy (antibiotic) and / or at least one PPI drug therapy of an antibiotic treatment, for example, the bacterial strain for use according to the invention, its culture supernatant or a composition comprising same according to the invention may be administered simultaneously with the administration of the antibiotic / PPI drug therapy and also after the antibiotic and / or PPI treatment has ended.

[0080] The dosage and frequency of administration of the bacterial strain or its culture supernatant for use according to the invention may be adapted depending on the host response.

[0081] By way of example only, the frequency of administration of bacterial strains, culture supernatants or compositions comprising same for use according to the invention can be daily administration for 5 to 15 consecutive days, or administration can be every 2, 3, 4, 5, 6 or 7 days for up to several months.

[0082] In the case of individuals who are starting or have already started the above-mentioned treatment, in particular antibiotic and / or PPI treatment, the frequency of administration of the bacterial strain, culture supernatant or composition comprising same for use according to the invention can be daily administration throughout the entire duration of the individual's treatment, in particular throughout the entire duration of antibiotic and / or PPI treatment, as well as administration of the bacterial strain, culture supernatant or composition comprising same for use according to the invention: - may be initiated one or several days before the start of the above treatments, in particular antibiotic and / or PPI treatment; and / or - May be terminated one or several days after the end of the above treatments, especially antibiotic and / or PPI treatment.

[0083] As is known in the art, adjustments may be necessary for proteolysis, systemic versus local delivery, as well as age, weight, general health, sex, diet, time of administration, drug interactions, and severity of the condition, and these are readily determined by one of ordinary skill in the art with routine experimentation.

[0084] The antibiotic medication, or antibiotic and / or PPI medication, relevant according to the present invention may be any medication that alters the growth of, inhibits or destroys microorganisms of the gut microbiota.

[0085] Without limitation, such antibiotics may be selected from the group consisting of penicillins, tetracyclines, cephalosporins, quinolones, lincomycin, macrolides, sulfonamides, glycopeptides, aminoglycosides, carbapenems, and mixtures thereof, in particular amoxicillin, doxycycline, cephalexin, ciprofloxacin, clindamycin, metronidazole, azithromycin, sulfamethoxazole, and trimethoprim, clavulanate, ofloxacin, levofloxacin, and mixtures thereof.

[0086] The antibiotic may be selected from the group consisting of, for example, amoxicillin, doxycycline, cephalexin, ciprofloxacin, clindamycin, metronidazole, azithromycin, sulfamethoxazole and trimethoprim, clavulanate, levofloxacin, vancomycin, fidaxomicin, metronidazole, kanamycin, gentamicin, colistin, clindamycin, and mixtures thereof.

[0087] In a non-limiting manner, such PPIs can be selected from the group consisting of omeprazole, lansoprazole, pantoprazole, rabeprazole, esomeprazole and dexlansoprazole.

[0088] The present invention also provides: for treating and / or preventing Clostridioides difficile infection in an individual - the bacterial strain of the species Faecalibacterium prausnitzii deposited at the CNCM under accession number CNCM I-4573; and / or - the culture supernatant thereof; and / or - a composition comprising a bacterial strain or its culture supernatant in a physiologically acceptable medium Regarding the use of.

[0089] The present invention also provides a method for treating and / or preventing Clostridioides difficile infection in an individual, comprising administering to said individual: - the bacterial strain of the species Faecalibacterium prausnitzii deposited at the CNCM under accession number CNCM I-4573; and / or - the culture supernatant thereof; and / or - a composition comprising a bacterial strain or its culture supernatant in a physiologically acceptable medium The present invention relates to a method comprising administering

[0090] The present invention also provides: For the manufacture of a medicament for treating and / or preventing Clostridioides difficile infection in an individual. - the bacterial strain of the species Faecalibacterium prausnitzii deposited at the CNCM under accession number CNCM I-4573; and / or - the culture supernatant thereof; and / or - a composition comprising a bacterial strain or its culture supernatant in a physiologically acceptable medium Regarding the use of.

[0091] The invention will be explained in more detail below by means of the following examples, which are given by way of illustration only.

[0092] All references to percentages are by weight unless otherwise stated.

[0093] Example The F. prausnitzii strain according to the invention, deposited at the CNCM under accession number CNCM I-4573, was tested in vitro and in vivo for its ability to modulate the intestinal colonization of C. difficile, in particular in mice exhibiting induced dysbiosis.

[0094] 26-week-old conventionally raised C57 Black 6 (C57BL / 6) female mice were identified and housed in isolators with sterile food and water to protect them from the external environment. Mice were divided into two groups: - 10 treated with PBS-glycerol 16% (vehicle); - 10 animals are treated with F. prausnitzii strain CNCM I-4573.

[0095] To produce intestinal dysbiosis, 6 days before infection (D-6), all mice were given an antibiotic cocktail consisting of kanamycin 0.4 mg / mL, gentamicin 0.035 mg / mL, colistin 0.0567 mg / mL, metronidazole 0.215 mg / mL, and vancomycin 0.045 mg / mL in drinking water for 3 days (D-6 to D-3). On D-1, all mice were given a single 200 μL intraperitoneal (IP) dose of clindamycin 50 mg / kg.

[0096] From D-6 to D+14, each group received 200 μL of the strain corresponding to that group once a day. On D0, all mice received 2 × 10 5 Infection was performed by gavage with 500 μL of CFU / mL C. difficile strain R20291 (DSM27147). For this, a preculture was performed with C. difficile strain R20291 (DSM27147) the day before, and the purity of the suspension was verified by Gram staining the following morning. The culture was diluted with 27 mL of BHI along with 3 mL of preculture. After approximately 7 hours of anaerobic incubation, the suspension was counted and 2 × 10 cells were collected in 50 mL of reconstituted 1 × PBS. 5 After infection with C. difficile, mice are weighed daily for 8 days after infection, and then on D10, D14, and D20.

[0097] To monitor intestinal colonization by C. difficile, mouse feces were collected on D1, D2, D3, D5, D8, D12, D14, and D20 and resuspended in PBS to a final concentration of 10 mg / mL. -1 , 10 -2 and 10 -3 The suspension is diluted to 100 ml, spread onto BHI agar + 3% blood + C. difficile selective supplement (Biomerieux®) and incubated under anaerobic conditions at 37° C. for 48 hours. The undiluted suspension is also spread under the same conditions.

[0098] result All mice were given an antibiotic cocktail to induce dysbiosis, allowing for better colonization of the intestinal tract with C. difficile R20291 (DSM27147). On each day of the experiment (D-6 to D14), mice were fed F. prausnitzii CNCM I-4573 or PBS-glycerol 16% (vehicle). After infection with the toxigenic C. difficile strain R20291 (DSM27147), clinical scores (loss of body weight and activity, diarrhea, and mortality) were assessed, and colonization was monitored by fecal bacterial counts.

[0099] 1) Clinical evaluation Regarding the evaluation of clinical scores (FIG. 1), no weight loss, decreased activity, or diarrhea was observed in mice treated with the F. prausnitzii strain of the present invention.

[0100] On the other hand, mice in the corresponding control group (PBS-vehicle) showed weight loss from D2 onwards, followed by their recovery to normal weight on D6 post-infection, reduced activity and diarrhea.

[0101] The survival of the animals after challenge was monitored. As shown in Figure 2, after infection with C. difficile, two mice died in the PBS-glycerol 16% group (control-vehicle), and one mouse treated with F. prausnitzii CNCM I-4573 died 5 days after infection.

[0102] 2) Monitoring of establishment Regarding monitoring the colonization of mice with C. difficile (Fig. 3 ), all mice were colonized with C. difficile at D4.

[0103] Although mice in the inventive F. prausnitzii strain group did not appear to develop symptoms, they were fully colonized. There was a delay in colonization and more rapid clearance of C. difficile compared to the control group (vehicle group) (from D6, compared to D14 for the control group). At D10, C. difficile was still colonized in 4 of 4 mice treated with PBS-glycerol 16% (vehicle control group) and 1 of 6 mice treated with the inventive F. prausnitzii strain.

[0104] It was demonstrated that treatment based on administration of the F. prausnitzii strain of the present invention unexpectedly provided protection from C. difficile infection. Despite all mice being colonized, none developed clinical signs (stable weight, normal feces and activity). Colonization was delayed, and surprisingly, more rapid clearance of C. difficile was observed.

[0105] [Table 1]

Claims

1. A bacterial strain of the species Faecalibacterium prausnitzii deposited at the CNCM under accession number CNCM I-4573 and / or a culture supernatant thereof for use in the treatment and / or prevention of Clostridioides difficile infection in an individual.

2. 2. The bacterial strain and / or culture supernatant thereof for use according to claim 1, characterized in that the individual is selected from the group consisting of humans; Suidae, in particular pigs, more particularly piglets; Bovinae, in particular cattle, more particularly bulls, more particularly steers; Canidae, in particular dogs; Equidae, in particular horses; and Phasianidae, in particular poultry, more particularly chickens.

3. A bacterial strain and / or its culture supernatant for use thereof according to claim 1 or 2, wherein said individual is a mammal, more particularly a human.

4. A bacterial strain and / or its culture supernatant for use thereof according to any one of claims 1 to 3, characterized in that the bacterial strain of the species Faecalibacterium prausnitzii deposited at the CNCM under accession number CNCM I-4573 is administered to said individual in live, inactive and / or dead form, preferably in live or inactive form, more particularly in live form.

5. 5. The bacterial strain and / or culture supernatant thereof for use thereof according to any one of claims 1 to 4, characterized in that the Clostridioides difficile infection occurs after the use of at least one antibiotic drug therapy and / or at least one proton pump inhibitor (PPI) drug therapy by said individual.

6. A bacterial strain and / or its culture supernatant for use according to any one of claims 1 to 5, characterized in that the bacterial strain and / or its culture supernatant of the species Faecalibacterium prausnitzii deposited in the CNCM under accession number CNCM I-4573 is administered simultaneously with and / or after at least one antibiotic drug therapy and / or at least one PPI drug therapy.

7. A bacterial strain and / or its culture supernatant for use as described in any one of claims 1 to 6, wherein the bacterial strain of the species Faecalibacterium prausnitzii deposited at the CNCM under accession number CNCM I-4573 is contained in a composition comprising a physiologically acceptable medium.

8. A bacterial strain and / or its culture supernatant for use as claimed in claim 7, characterized in that the bacterial strain of the species Faecalibacterium prausnitzii deposited at the CNCM under accession number CNCM I-4573 is contained in an oral or rectal composition, in particular in an oral composition.

9. A bacterial strain and / or its culture supernatant for its use according to claim 7 or 8, characterized in that said bacterial strain is contained in a medicinal product.

10. The composition is a pharmaceutical formulation of 10 strains of bacteria. ４ ~10 １５ A daily dose equivalent to the total cell count (TCC) of 10 ８ ~l0 １３ Corresponding to TCC, more specifically 10 １３ A bacterial strain and / or its culture supernatant for its use according to any one of claims 7 to 9, characterized in that it is suitable for administration in the form of a daily dose corresponding to TCC.

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