WO2018046485A1 - Utilisation de butyricimonas et/ou d'oxalobacter pour traiter le syndrome de l'intestin irritable - Google Patents

Utilisation de butyricimonas et/ou d'oxalobacter pour traiter le syndrome de l'intestin irritable Download PDF

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Publication number
WO2018046485A1
WO2018046485A1 PCT/EP2017/072207 EP2017072207W WO2018046485A1 WO 2018046485 A1 WO2018046485 A1 WO 2018046485A1 EP 2017072207 W EP2017072207 W EP 2017072207W WO 2018046485 A1 WO2018046485 A1 WO 2018046485A1
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composition
butyricimonas
oxalobacter
bacteria
composition according
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PCT/EP2017/072207
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English (en)
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Magnus SIMRÉN
Agnes Wold
Ingegerd Adlerberth
Fei Sjöberg
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Swecure Ab
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K35/00—Medicinal preparations containing materials or reaction products thereof with undetermined constitution
    • A61K35/66—Microorganisms or materials therefrom
    • A61K35/74—Bacteria
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P1/00—Drugs for disorders of the alimentary tract or the digestive system

Definitions

  • This invention pertains in general to the field of probiotic treatment. More particularly the invention relates to use of microorganisms for prevention or treatment of irritable bowel syndrome (IBS) in a human patient.
  • IBS irritable bowel syndrome
  • IBS Irritable bowel syndrome
  • IBS intracranial pressure
  • post-infectious IBS A small but significant relive in symptoms due to antibiotic treatment have been shown, and probiotics treatment may alleviate the symptoms according to a meta-analysis.
  • IBS is a heterogeneous disorder and can be subdivided by the Rome II criteria, which is based on stool frequency, stool form and absence or presence of defecation straining or urgency. According to this, IBS can be grouped into three symptom subtypes: diarrhea predominant (IBS-D), constipation predominant (IBS-C) and mixed subtype (IBS-A) with alternating episodes of both diarrhea and constipation, retrospective. IBS a disorder distinct from inflammatory bowel disease (e.g. ulcerous colitis and Crohn's disease).
  • WO 2012/142605 there are a large number of gastrointestinal diseases that can be, at least in part, attributable to an imbalance, disruption, or abnormal distribution of microflora in the gastrointestinal tract of a mammalian host.
  • present treatment of IBS includes dietary adjustments, medication, and psychological interventions.
  • Treatment methods aim to supply a large microbiota of bacteria to the gastrointestronal tract of a patient, to treat inflammatory disease and IBS.
  • freeze dried feces is used, comprising a wide range of microorganisms in a single formulation thus not taking the differences between various intestinal disorders and their cause into consideration.
  • US 2014/0328803 discloses a therapeutic composition containing nonpathogenic, germination-competent bacterial spores, for the prevention, control, and treatment of gastrointestinal diseases, disorders and conditions and for general nutritional health.
  • a population of germinatable bacterial spores is administered to populate a gastrointestinal tract in the subject.
  • the treatment aims to augmenting growth of at least one bacteria type not present in the gastrointestinal tract.
  • the present invention preferably seeks to mitigate, alleviate or eliminate one or more of the above identified deficiencies in the art and disadvantages singly or in any combination and solves at least the above mentioned problems by providing a composition comprising a therapeutically effective amount of any species of viable Butyricimonas and/or Oxalobacter bacteria and a carrier or excipient for use in the prevention or treatment of irritable bowel syndrome (IBS) in a subject.
  • IBS irritable bowel syndrome
  • the application further provides a method of treating or reducing a severity of at least one symptom of the gastrointestinal disease irritable bowel syndrome (IBS), comprising administering to a human subject a composition comprising bacterial strains of genus Butyricimonas and/or Oxalobacter in an amount effective to populate a gastrointestinal tract in the subject, thereby providing an amount effective to treat the gastrointestinal disease or reduce a severity of at least one symptom of the gastrointestinal disease in the subject.
  • IBS gastrointestinal disease irritable bowel syndrome
  • Fig. 1 depicts a graph which shows, for analyzed samples from IBS patients and control subjects, the number of DNA sequence reads per sample; and Fig. 2 depicts graphs which show microbiota comparisons between controls and IBS patients for A) bacterial genera Butyricimonas and B) bacterial genera
  • 16S ribosomal RNA (rRNA) sequencing was used to identify and compare bacteria present within the faeces samples.
  • the rRNA gene is the most conserved DNA in all cells.
  • the small ribosomal subunit contains the 16S rRNA, thus the comparison of 16s rDNA sequence can show evolutionary relatedness among microorganisms.
  • 16S rRNA gene sequencing is a powerful culture-free method for analysis of the microbial community within a complex sample. This enables the detection of strains that may not be found using other methods.
  • Butyricimonas are gram-negative obligate anaerobic bacteria that produce butyrate (main end products are butyric and isobutyric acids, but smaller amounts of acetic propionic and succinic acids are also produced), which serves as nourishment for colonocytes (colonic epithelial cells) and can have positive effects on the epithelium and mucosa. As known in the art, lack of butyrate can cause an increased permeability of the colon epithelium. Mucosal barrier defects have indeed been shown in IBS patients suffering from diarrhea (Dunlop, S. P. et al. Abnormal intestinal permeability in subgroups of diarrhea-predominant irritable bowel syndromes. Am. J. Gastroenterol.
  • Butyricimonas may thus serve to counter-act the increased permeability of the colon epithelium, and mucosal barrier defects often seen in IBS. Furthermore, intestinal colonization with Butyricimonas have also been shown to be decreased in other diseases. Patients with multiple sclerosis had alterations in the composition of the microbiota, with lower colonization rates with Butyricimonas compared to healthy controls (Jangi, S. et al. Alterations of the human gut microbiome in multiple sclerosis. Nat. Commun. 7: 12015 DOI: 10.1038/ncomms l2015 (2016)). The same study showed that Butyricimonas also had negative correlations with inflammatory markers, suggesting that reduction in Butyricimonas is associated with increased inflammation.
  • Oxalobacter is a genus of bacteria in the Oxalobacteraceae family. They are found in the human gastrointestinal microbiota, where Oxalobacter formigenes is a known oxalate-degrading anaerobic bacterium that colonizes the large intestine.
  • Oxalobacter uses oxalate as their sole carbon source, supporting their important role in removing oxalate from the human body. Elevated oxalate levels may cause oxalate stone formation, and lack of Oxalobacter formigenes colonization in the gastrointestinal tract may lead to recurrent kidney stone disease (Kaufmann et al in J Am Soc Nephrol 19: 1 197-1203, 2008; DOI: 10.1681/ASN.2007101058). Any role of elevated levels of oxalate in IBS has not been described in the art, but a link between IBS and kidney stone formation has been observed.
  • the relative risk of developing IBS was significantly higher in patients with kidney stone formation compared to patients devoid oxalate stone disease (Erdum et al in Digestive Diseases and Sciences, Vol. 50, No. 3 (March 2005), pp. 605-608; DOI: 10.1007/s 10620-005-2482-3 and Lei et al, in PLoS ONE 11(6): e0157701. DOI: 10.1371/journal.pone.0157701).
  • oxalate has been shown to promote production of the inflammatory mediators IL-6 and MCP- 1 from epithelial cell lines (Huang et al in Kidney International, Vol. 68 (2005), pp. 497-503; and Umekawa et al in Kidney International, Vol. 61 (2002), pp. 105-1 12), factors that potentially can recruit and activate inflammatory cells in the gut.
  • Butyricimonas and Oxalobacter can be used to prevent and treat IBS.
  • An embodiment thus relates to a composition
  • a composition comprising a therapeutically effective amount of any species of viable Butyricimonas and/or Oxalobacter bacteria and a carrier or excipient for use in the prevention or treatment of irritable bowel syndrome (IBS) in a subject.
  • IBS irritable bowel syndrome
  • another embodiment relates to a method of preventing, e.g. reducing the incidence of, treating or reducing a severity of at least one symptom of the gastrointestinal disease irritable bowel syndrome (IBS) in a human, the method comprising administration of viable Butyricimonas and/or Oxalobacter bacteria.
  • a composition comprising a therapeutically effective amount of viable Butyricimonas and/or Oxalobacter bacteria and a carrier or excipient, is administered, in a therapeutically effective dose, to a subject in need thereof.
  • an amount effective to treat the gastrointestinal disease or reduce a severity of at least one symptom of the gastrointestinal disease in the subject is provided.
  • Another embodiment relates to the use of viable Butyricimonas and Oxalobacter bacteria for the manufacture of a composition comprising a therapeutically effective amount of Butyricimonas and/or Oxalobacter bacteria and a carrier or excipient, for the use in prevention or treatment of irritable bowel syndrome (IBS).
  • IBS irritable bowel syndrome
  • composition may be a pharmaceutical or nutraceutical composition.
  • nutraceutical composition refers to a composition that, except for its basic nutritional value, also provides extra health benefits, such as alleviating IBS.
  • the bacteria to be used are purified and isolated.
  • the viable Butyricimonas and/or Oxalobacter bacteria are isolated bacteria. Once a strain has been isolated, it is often possible to culture said strain to provide a pure and homogeneous population of bacteria. Furthermore, this is often suitable for larger scale processes, and offers a high level of control of bacterial purity and level of contaminants.
  • the viable Butyricimonas and/or Oxalobacter bacteria are cultured bacteria.
  • composition should comprise a therapeutically effective amount of viable Butyricimonas and/or
  • Oxalobacter bacteria The actual content will depend on various factors; cf. the type of mammal, the manner of administration, and the nature and severity of the disorder. According to an embodiment, the content of Butyricimonas and/or Oxalobacter bacteria, i.e. the total amount of such viable bacteria, in the pharmaceutical composition is 10 5 cfu/g to 10 12 cfu/g, such as 10 7 cfu/g to 10 10 cfu/g. Further, not only the amount of viable bacteria in the formulation, but also the dose administered may be of importance for the treatment to be effective. A therapeutically effective dose is a dose that is sufficient to produce the desired effects in relation to the condition for which it is administered.
  • the exact dose may be dependent on the manner of administration, nature and severity of the disorder and/or disease and the general conditions, such as age and body weight of the subject. Further, different types of mammals may, or may not, need different doses.
  • the total amount of viable Butyricimonas and/or Oxalobacter bacteria administered per day is within the range of 10 4 to 10 11 cfu/ kg body weight, such as within the range of 10 6 to 10 9 cfu/ kg body weight.
  • the amount of each bacteria administered is within this range.
  • the composition may comprise other microorganisms than Butyricimonas and/or Oxalobacter bacteria. According to an embodiment, at least 10%, such as at least 25%, 35%, 50%, 60%, 75%, or 90% of the total number of colony forming
  • microorganisms in the composition are Butyricimonas and/or Oxalobacter bacteria.
  • the composition comprises Butyricimonas bacteria as well as other bacteria, at least 5%, such as at least 10%, 15%, 20%, 25%, 30%, 40%, 50%, 60%, or 70% of the total number of colony forming microorganisms in the composition are Butyricimonas bacteria.
  • the composition comprises Oxalobacter bacteria as well as other bacteria, at least 5%, such as at least 10%, 15%, 20%, 25%, 30%, 40%, 50%, 60%, or 70% of the total number of colony forming microorganisms in the composition may be Oxalobacter bacteria.
  • the composition may comprise Butyricimonas or Oxalobacter bacteria, it is preferred if both bacteria are present in the composition to be administered. Without being bound to theory, it is conceivable that the deficiency of Butyricimonas and Oxalobacter bacteria in the intestinal flora may lead to a dissonant synergy effect for the intestinal tract and that administration of both these bacteria thus may be additional beneficial.
  • Butyricimonas are symbiotic bacteria of old for the mammalian intestine, so long that the lining of the large intestine uses butyrate as its primary source of energy. It may play roles both in intestinal permeability (which in turn may turn out to be linked to autoimmune disorders of the gut) and gut inflammation. Oxalobacter deficiency in turn leads to problems with the metabolism of dietary oxalates. This could be a source for pain in people with damaged bowel lining (i.e. popularly called a "leaky gut”), a problem possibly caused or amplified by the Butyricimonas deficiency.
  • composition may be supplemented with other bacteria known to be beneficial, e.g. have probiotic potential, such as one or several of Streptococcus, Lactococcus, Leuconostoc, Pediococcus, Finegoldia, Ruminococcus, Roseburia,
  • probiotic potential such as one or several of Streptococcus, Lactococcus, Leuconostoc, Pediococcus, Finegoldia, Ruminococcus, Roseburia,
  • the composition is supplemented with at least one additional bacteria, such 1, 2, 3 or 4 types of bacteria, selected from the group consisting of Streptococci, Lactococcus, Leuconostoc,
  • Pediococcus Roseburia and Eubacterium, such as from Streptococcus, Lactococcus, Leuconostoc, Pediococcus, Odoribacter, Collinsella, Lactobacillus, Clostridium,.
  • said additional microorganism(s) are selected from the group consisting of Clostridium and Lactobacillus.
  • Butyricimonas and/or Oxalobacter may constitute at least 50%, such as at least 60%, 70%, 80%, 90%, or 95% of the total number of colony forming microorganisms in the pharmaceutical or nutraceutical composition.
  • the Butyricimonas and/or Oxalobacter to be included in pharmaceutically composition, as well as any additional bacterium/bacteria, may be prepared by fermentation carried out under various conditions. If more than one type of bacteria is to be present, they may either be cultured individually or co-cultured. After the fermentation, the bacteria are collected. In one embodiment, the bacteria in the composition is cultured individually or co-cultured. The collection may be performed by centrifugation, or using alternative cell harvesting methods such as ultra-filtration, direct spray drying, precipitation or sedimentation, as known in the art. The resulting wet pellets can then be dried by any method that preserves the activity of the bacteria. As known to the skilled person, such methods include freeze drying, spray drying, heat drying and sporulation. Further, in order to maintain a high viability and protect the bacteria during the drying step, a protective agent may be added to the bacteria before the drying step.
  • Commonly employed protective agents comprise polyhydroxy compounds, e.g. a mono-, di-, or polysaccharide, polyalcohols, e.g. sorbitol, and inorganic agents, such as calcium chloride.
  • the bacteria may be mixed with a pharmaceutically acceptable carrier.
  • Carriers and excipients including pharmaceutically acceptable carriers and excipients, suitable for the administration of microorganism are well-known to the skilled artisan. Such carrier should be compatible with the bacteria and not deleterious to the subject to be treated. Suitable carriers include water, microcrystalline cellulose, mannitol, glucose, defatted milk powder, polyvinylpyrrolidone, and starch, or a combination thereof.
  • Suitable excipients include alginate, carrageenan, chitosan, cellulose acetate phthalate and locust bean gum, or a combination thereof.
  • the composition may comprise a fiber, preferably a non-digestible fiber.
  • the fiber content in the composition may be 5 to 95 wt% on dry weight basis.
  • the composition may comprise at least 5 wt%, such as at least 10, 15, 20, 25, 30, 40, or 50 wt% fibers on dry weight basis.
  • the composition may comprise less than 95 wt%, such as less than 90, 80, 70, 60 or 50 wt% fibers on dry weight basis.
  • suitable fibers include oat fibers, rye fibers, inulin, rice fibers, Psylium seed husks, soybean oligosaccharides, pectin, resistant starch,
  • the composition comprises at least one fiber, wherein the fiber is selected from a group consisting of oat fibers, rye fibers, inulin, rice fibers, Psylium seed husks, soybean oligosaccharides, pectin, resistant starch, galactooligosaccharide, fructooligosaccharide (FOS), lactulose, isomaltooligosaccharides, lactitol, lactosucrose, lactulose, pyrodextrin, transgalactooligosaccharides, and xylooligosaccharides , such as fructooligosaccharide or inulin.
  • the fiber is selected from a group consisting of oat fibers, rye fibers, inulin, rice fibers, Psylium seed husks, soybean oligosaccharides, pectin, resistant starch, galactooligosaccharide, fructooligosaccharide
  • composition will typically be formulated for oral administration.
  • suitable oral formulations of the composition include among others solid
  • compositions such as tablets, capsules and powders, and liquid dispersions. Further, the composition may be formulated for the rectal administration.
  • the composition can be introduced via direct access to the compromised gut (tube, enema, suppository or similar mechanism). The composition may thus be administered rectally, e.g. by using a suppository.
  • the composition is for oral or rectal administration, where the composition for oral administration may be formulated in the form of a tablet, a capsule, a powder, or a liquid dispersion.
  • the carrier(s) or excipient(s) may, without limitation, be selected from the group consisting of: corn starch, gelatin, lactose, acacia, sucrose, microcrystalline cellulose, kaolin, mannitol, dicalcium phosphate, calcium carbonate, sodium chloride, alginic acid.
  • tablet binders such as acacia, methylcellulose, sodium carboxymethylcellulose, polyvinylpyrrolidone (Povidone3), hydroxypropyl methylcellulose, sucrose, starch, and ethylcellulose may be used.
  • a lubricant may be used in formulating a solid composition.
  • lubricants examples include magnesium stearates, stearic acid, silicone fluid, talc, waxes, oils, and colloidal silica.
  • disintegrators such as microglycolate, and alginic acid, may be present.
  • Solid compositions may further be formulated in manner such that the bacteria are released in the intestines, whereby avoiding exposure to the acidic conditions in the stomach.
  • examples of such formulations e.g. tablets with an enteric coating, are known in the art.
  • liquid compositions e.g. dispersions, comprising Butyricimonas and Oxalobacter
  • liquid food products e.g. nutraceutical compositions.
  • the composition is a food product.
  • food products include yogurt, milk, or other drinks and beverages, and a food supplement.
  • a nutraceutical composition may also be a composition in powder form.
  • a composition in powder form may be dispersed in liquid, such as a beverage, before being consumed. Further, a composition in powder form may be spread over an ordinary meal. When to be used in powder form, the composition may be a freeze dried composition.
  • the composition may be formulated as a suppository.
  • the suppository may be hard or a soft suppository.
  • anhydrous hydrophobic carriers may be used.
  • suitable anhydrous hydrophobic carriers include vegetable fat, e.g. a long-chain triglyceride, vegetable oil, e.g. a medium-chain triglyceride, and mixtures thereof.
  • the water content in the formulation should be as low as possible.
  • the water activity at 25°C of a soft suppository should preferably be less than 0.30, such as less than 0.25 or less than 0.20.
  • the water content in such a soft suppository may be less the 0.5 wt%, such as less than 0.1 wt%.
  • medium-chain triglyceride refers to a triglyceride comprising residues of carboxylic acids comprising 6 to 12 carbon atoms.
  • the term is intended to include separate triglycerides as well as mixtures of triglycerides.
  • medium-chain triglycerides are Ce-n fatty acid esters of glycerol.
  • the fatty acid may be the same or different and they may be saturated, monounsaturated or polyunsaturated.
  • a preferred example of a medium-chain triglyceride is an oil having vegetable origin.
  • long-chain triglyceride refers to a triglyceride comprising residues of carboxylic acids comprising more than 12 carbon atoms.
  • the term is intended to include separate triglycerides as well as mixtures of triglycerides.
  • long-chain triglycerides are C13-30, such as C16-22, fatty acid esters of glycerol.
  • the fatty acid may be the same or different and they may be saturated, monounsaturated or polyunsaturated.
  • a preferred example of a long-chain triglyceride is a fat having vegetable origin.
  • prevent/preventing should not be construed to mean that a condition and/or a disease never might occur again after use of the pharmaceutical composition to achieve prevention. Further, the term should neither be construed to mean that a condition not might occur, at least to some extent, after such use to prevent said condition. Rather, “prevent/preventing” is intended to mean that the condition to be prevented, if occurring despite such use, will be less severe than without such use. Thus, prevent may be interpreted as reducing the coincidence of a given condition. Further, the term may be interpreted as prophylactic treatment.
  • HAD Anxiety and Depression
  • Gastrointestinal Symptom Rating Scale-IBS (as known in the art); Bristol Stool Form Scale (BSF) - one week diary to assess stool form (as known in the art); and the Short Form-36 (SF-36) (as known in the art).
  • the subjects also underwent a rectal sensitivity test with a rectal barostat (as known in the art), and colonic transit time measurement using radiopaque markers (as known in the art).
  • Freshly voided feces were taken by the 23 of the IBS subjects and all 14 controls.
  • the samples were collected at home and immediately placed in sterile tubes which were placed in plastic bags in which an anaerobic atmosphere was created (AnaeroGen Compact, Oxoid Ltd, Basingstoke, UK).
  • the samples were kept refrigerated until transported to the laboratory. Portions of all samples were frozen at -80°C for pyrosequencing analysis.
  • Bacterial DNA was extracted from 200 mg of fecal sample according to the manufacturer's protocol (QIAamp DNA Stool Mini Kit, Qiagen, Hilden, Germany). To increase DNA yield, four glass beads (3.0 mm diameter) and 0.5 g zirconia beads (0.1 mm) were added to the ASL buffer and the suspension was homogenized at 5 m/s, 2x30 s (Thermo Savant FastPrep Cell disrupter, Holbrook, NY, USA), incubated at 95°C for 5 min, and then shaken at 1,200 rpm (Vibrax Shaker; IKA, Staufen, Germany) for 30 min at 5°C. The DNA was quantified spectrophotometrically (NanoDrop Technologies, Wilmington, DE, USA).
  • the VI -V3 region of the bacterial 16S rRNA genes was amplified using forward primer 27F, 5 '-AGAGTTTGATCCTGGCTCAG-3 ' (SEQ ID No. 1); reverse primer 534R, 5 '-ATTACCGCGGCTGCTGG -3' (SEQ ID No. 2) and the following PCR protocol: 98°C for 30 s, followed by 25 cycles of 98°C for 10 s, 56°C for 30 s, 72°C for 10 s, and a final extension at 72°C for 1 min.
  • the library was constructed according to the protocol of GATC Biotech (Konstanz, Germany) and sequenced on a Genome Sequencer FLX System (Roche).
  • OTUs Operational Taxonomic Units
  • IBS-A insulin-predominant IBS
  • IBS-C constipation-predominant IBS
  • IBS-A alternating type of IBS
  • IBS-D diarrhea-predominant IBS
  • IBS-C constipation-predominant IBS
  • the fecal microbiota of patients with IBS was compared to healthy controls using pyrosequencing.

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Abstract

L'invention concerne une composition comprenant une quantité thérapeutiquement efficace de bactéries viables d'espèces quelconques des genres Butyricimonas et/ou Oxalobacter ainsi qu'un support ou excipient, destinée à être utilisée chez le patient dans la prévention ou le traitement du syndrome de l'intestin irritable (SII). L'invention concerne en outre une méthode permettant de traiter au moins un symptôme d'une maladie gastro-intestinale, le syndrome de l'intestin irritable (SII), ou de diminuer la gravité de ce symptôme, la méthode comprenant l'administration à un patient humain d'une composition comprenant des souches bactériennes des genres Butyricimonas et/ou Oxalobacter en une quantité suffisante pour coloniser le tractus gastro-intestinal du patient, apportant ainsi une quantité efficace pour traiter la maladie gastro-intestinale ou diminuer la gravité d'au moins un symptôme de la maladie gastro-intestinale chez le patient.
PCT/EP2017/072207 2016-09-09 2017-09-05 Utilisation de butyricimonas et/ou d'oxalobacter pour traiter le syndrome de l'intestin irritable WO2018046485A1 (fr)

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Cited By (3)

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WO2021138571A1 (fr) * 2019-12-31 2021-07-08 Assembly Biosciences, Inc. Compositions comprenant des espèces bactériennes et procédés associés
WO2021137494A1 (fr) * 2020-01-02 2021-07-08 삼육대학교산학협력단 Composition pour la prévention ou le traitement du diabète comprenant une souche de butyricimonas sp. en tant que principe actif
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Cited By (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2021138571A1 (fr) * 2019-12-31 2021-07-08 Assembly Biosciences, Inc. Compositions comprenant des espèces bactériennes et procédés associés
WO2021137494A1 (fr) * 2020-01-02 2021-07-08 삼육대학교산학협력단 Composition pour la prévention ou le traitement du diabète comprenant une souche de butyricimonas sp. en tant que principe actif
WO2021173895A1 (fr) * 2020-02-28 2021-09-02 The Cleveland Clinic Foundation Analyse du microbiome pour le diagnostic et le traitement de maladie des calculs urinaires

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