(19) TZZ Z _T

(11) EP 2 750 682 B1

(12) EUROPEAN PATENT SPECIFICATION

(45) Date of publication and mention (51) Int Cl.: of the grant of the patent: A61K 35/74 (2015.01) A61P 1/12 (2006.01) 18.05.2016 Bulletin 2016/20 A61P 1/00 (2006.01) A61P 17/10 (2006.01) A61P 19/02 (2006.01) A61P 35/00 (2006.01) (21) Application number: 12772317.9 (86) International application number: (22) Date of filing: 11.10.2012 PCT/EP2012/070177

(87) International publication number: WO 2013/053836 (18.04.2013 Gazette 2013/16)

(54) COMPOSITION COMPRISING ANAEROBICALLY CULTIVATED HUMAN INTESTINAL MICROBIOTA ZUSAMMENSETZUNG, ENTHALTEND ANAEROBISCH KULITIVIERTE MENSCHLICHE DARMFLORA COMPOSITION COMPRENANT DE LA FLORE INTESTINALE CULTIVE DE FAÇON ANAEROBIQUE

(84) Designated Contracting States: • BAKKEN JOHAN S: "Fecal bacteriotherapy for AL AT BE BG CH CY CZ DE DK EE ES FI FR GB recurrent Clostridium difficile infection", GR HR HU IE IS IT LI LT LU LV MC MK MT NL NO ANAEROBE,, vol. 15, no. 6, 1 January 2009 PL PT RO RS SE SI SK SM TR (2009-01-01), pages285-289, XP009161367, ISSN: 1095-8274 (30) Priority: 11.10.2011 SE 1150940 • JOHANNES AAS ET AL: "Recurrent Clostridium 11.01.2012 SE 1250011 difficile Colitis: Case Series Involving 18 Patients Treated with Donor Stool Administered via a (43) Date of publication of application: Nasogastric Tube", CLINICAL INFECTIOUS 09.07.2014 Bulletin 2014/28 DISEASES, vol. 36, no. 5, 1 March 2003 (2003-03-01), pages 580-585, XP55043539, ISSN: (73) Proprietor: Achim Biotherapeutics AB 1058-4838, DOI: 10.1086/367657 162 44 Vällingby (SE) • BORODY T J ET AL: "Treatment of ulcerative colitis using fecal bacteriotherapy", JOURNAL (72) Inventors: OF CLINICAL GASTROENTEROLOGY, RAVEN • BERSTAD, Arnold PRESS LTD., NEW YORK, NY, US, vol. 37, no. 1, N-0783 Oslo (NO) 1 January 2003 (2003-01-01), pages 42-47, • MIDTVEDT, Tore XP008093566, ISSN: 0192-0790, DOI: N-1346 Gjettum (NO) 10.1097/00004836-200307000-00012 • NORIN, Elisabeth • Sebastian Amaral Anders: "Mit Fäkalien heilen", S-162 44 Vällingby (SE) FrankfurterRundschau ,2 May 2011 (2011-05-02), • BENNO, Peter 2 May 2011 (2011-05-02), XP002686975, Retrieved S-181 33 Lidingö (SE) from the Internet: • DAHLGREN, Atti-La URL:http://www.fr-online.de/wissenschaft/b CH-2000 Neuchatel (CH) akterientherapie-mit-faekalien-heilen,1472 788,8400888.html [retrieved on 2012-11-08] (74) Representative: Fenix Legal KB Stureplan 4C, 4 tr 114 35 Stockholm (SE)

(56) References cited: WO-A1-02/07741 WO-A1-2011/033310

Note: Within nine months of the publication of the mention of the grant of the European patent in the European Patent Bulletin, any person may give notice to the European Patent Office of opposition to that patent, in accordance with the Implementing Regulations. Notice of opposition shall not be deemed to have been filed until the opposition fee has been paid. (Art. 99(1) European Patent Convention). EP 2 750 682 B1

Printed by Jouve, 75001 PARIS (FR) (Cont. next page) EP 2 750 682 B1

• SERINO M ET AL: "Intestinal microflora and metabolic diseases", DIABETES & METABOLISM, PARIS, AMSTERDAM, NL, vol. 35, no. 4, 1 September 2009 (2009-09-01), pages 262-272, XP026642909, ISSN: 1262-3636, DOI: 10.1016/J.DIABET.2009.03.003 [retrieved on 2009-05-05]

2 1 EP 2 750 682 B1 2

Description for the treatment of gastrointestinal disorders. WO 2011/033310 describes encapsulated dosage form FIELD OF THE INVENTION comprisingflora extracted fromfaeces for use in the treat- ment of gastrointestinal disorders. [0001] The present invention relates to an anaerobic 5 [0008] Faecal transplantation is a known method, but micro-ecological system comprising anaerobically culti- not without risk. Until now faecal material from donors of vated human intestinal microbiota. The present invention close relatives has been used. A main problem with this also relates to a composition of anaerobically cultivated procedure is the risk of transmission of genes resistant microbiota that constitutes a functional seeding culture to certain antibiotics. The transfer of such genes may for re-establishing normality of a disturbed human micro- 10 have serious consequences for the patient when antibi- biome and gastrointestinal functions. otic treatment is needed in the future. Another main prob- [0002] The present invention further relates to the com- lem with this therapy is the risk of transmission of poten- positionfor useas a pharmaceutical andfor useas proph- tially contagious agents present in the donor’s faeces, ylaxis of diseases and a method for preparing the com- which argue for careful screening of the faeces to be position. 15 given. This screening is time-consuming and expensive. Further this type of treatment is perceived by many as TECHNICAL BACKGROUND highly unesthetical. [0009] Previous publications mentioning faecal trans- [0003] The gastrointestinal microbiota consists of sev- plantation: eral complex micro-ecological systems, where interac- 20 tions in-between the individual microbial strains and be- - Johan S. Bakken Fecal bacteriotherapy for recurrent tween the microbiota and the host takes place. The in- Clostridium difficile infection Anaerobe 15 (2009) teractions with the host occur through direct contact with 285-289. the mucosal wall and indirectly e.g. through metabolites - C Jorup-Rönström, A Håkansson, A-K Person, T and signalling substances. 25 Midtvedt, E Norin. Feceskultur framgångsrik terapi [0004] This intestinal ecological balance can be dis- vid Clostridium difficile-diarré. Läkartidningen nr 46 turbed by external and internal physical, chemical and 2006 volym 103, 3603-3605. biological factors or agents, the consequence of which - A. Gustafsson, A. Berstad, S. Lund-Tønnesen, T. can be local e.g. disturbed metabolism, gastrointestinal Midtvedt, E. Norin.The Effect of Faecal Enema on motility alteration, altered functions/crosstalks, and/or 30 Five Microflora-Associated Characteristicsin Pa- lead to systemic effects involving other organ systems. tients with Antibiotic-Associated Diarrhoea. Scand J [0005] Clostridium difficile associated diarrhoea Gastroenterol 1999;34:580-586. CDAD was first described around 1/3 of a century ago. - A. Gustavsson, S. Lund-Tonnesen, A. Berstad, T. However, in spite of thousands of publications in Pub Midtvedt & E. NorinFaecal Short-Chain Fatty Acids Med of the pathogenesis, epidemiology, clinical diagno- 35 in Patients with Antibiotic-AssociatedDiarrhoea, be- sis and various therapeutic approaches CDAD continues fore and after Faecal Enema Treatment. Scand J to persist as a costly, leading cause of infectious health- Gastroenterol 1998;33:721-727. care-associated gastrointestinalillness and the problems are assumed to increase worldwide. It was recently stat- SUMMARY OF THE INVENTION ed that today we experienced a tenfold increase of CDAD 40 in Europe, USA and Canada. Increasing therapeutic fail- [0010] The present invention relates to a composition ures of metronidazole and vancomycin treatments are comprising strictly anaerobically cultivated human intes- reported and new antibiotics, as fidaxomicin, oritavancin, tinal microbiota selected from at least three of the fow nitazoxanide, REP3123, and NVB303 are on their way phyla Bacterioidete, , Proteobacteria and Ac- to the market. However, it has to be recognized that even 45 tinobacteria for use in the prevention and treatment of before these drugs have been used, a certain number of diseases. The present invention further relates to a meth- Clostridium difficile strains are expressing a reduced sen- od to prepare such a composition. Also described is the sitivity to these new drugs. Additionally, whatever their administration of a therapeutic or prophylactic amount of clinical efficacy may turn out to be, they will certainly be the invention to a human. expensive in use. 50 [0011] Also described is a composition comprising [0006] It has been stated that some patients continue anaerobically cultivated intestinal microbiota that consti- to manifest relapsing diarrhoea after completed treat- tutes a functional seeding culture for re-establishing nor- ment with novel drugs, and therefore other strategies mality of a disturbed human microbiota and gastrointes- have been focused on e.g. faecal transplantation from a tinal functions and the use of such composition for the close relative or a synthetic composition comprising a 55 prevention and treatment of disease. mixture of different . [0012] This invention also relates to a composition ac- [0007] WO 02/07741 describes a synthetic composi- cording to the claims for use in the pharmaceutical treat- tion comprising a preparation of a predetermined flora ment of gastrointestinal and/or systemic/metabolic dis-

3 3 EP 2 750 682 B1 4 orders that has its origin in a disturbed or dysfunctional [0024] The functional ecologic system according to the gastrointestinal ecosystem. present invention has the ability to re-establish and/or [0013] The invention also relates to the use of such an stabilise intestinal microbiota and/or normal intestinal anaerobically cultivated human microbiota for pharma- functions. ceutical treatment or prevention of disease. The compo- 5 [0025] Another aspect of the present invention is a sition may also contain other microbial factors (e.g. bac- composition comprising a micro-ecological system com- teriophages) or signal substances active against diseas- prising anaerobically cultivated human intestinal micro- es. biota for use in the treatment or prophylaxis of diseases. [0026] Preferably, the bacterial content of the compo- DETAILED DESCRIPTION OF THE INVENTION 10 sition is more than 106 per ml medium, more preferably more than 109 per ml medium. [0014] The present invention is defined by the claims. [0027] In one embodiment of the present invention the [0015] A first aspect of the present invention is a com- composition comprises an anaerobic micro-ecological position comprising anaerobically cultivated human in- system obtained from faeces from a single donor. Culti- testinal microbiota as defined in the claims. 15 vation of faeces from a healthy single donor is necessary [0016] In the present application the term "microbiota" for obtaining a functional micro-ecological system that is is used for the microbial flora, i.e. the microorganisms stable over time. that typically inhabit the gastrointestinal system. [0028] Preferably, the donor is a healthy individual with [0017] The composition according to the present in- a well-functional gastrointestinal system. vention constitutes a functional cultivate for re-establish- 20 [0029] Preferably, the faeces is obtained from a single ing normality of a disturbed human microbiota and gas- stool sample. Also preferably, the faeces were obtained trointestinal functions. This composition has surprisingly prior to 1995. shown to be useful as in the treatment or prophylaxis of [0030] In a preferred embodiment of the present inven- disease. tion, the composition is kept under strict anaerobic con- [0018] The composition according to the present in- 25 ditions until the final administration. Also, preferably the vention interacts in a synergistic way with the damaged composition is kept under anaerobic conditions during intestinal microbiota and re-establish its physiological the final administration. and metabolic functions. The invention may also be used [0031] Another aspect of the present invention is a as a prophylactic to avoid the disturbance of the intestinal method for preparing a composition comprising an anaer- microbiota in the first place. 30 obic micro-ecological system comprising anaerobically [0019] The composition contains anaerobic bacteria of cultivated human intestinal microbiota according to the at least three of the following four Phyla: Bacterioidetes, claims. Firmicutes, Proteobacteria and Actinobacteria. [0032] The composition according to the present in- [0020] According to another embodiment the compo- vention has been cultivated under strict anaerobic con- sition isfree from thefollowing microbes: hepatitis viruses 35 ditions. By anaerobic cultivation the proliferation of the A, B and C, cytomegalo virus, Epstein-Barr virus, human anaerobic bacteria is promoted, and further proliferation immunodeficiency virus (HIV), Calici- and Rotavirus, Sal- of aerobic bacterial is suppressed. monella, Shigella, Campylobacter, Yersinia, and proto- [0033] In a preferred embodiment of the present inven- zoan cysts. Preferably it is also free from material select- tion, the composition is prepared by cultivating a stool ed from Extended spectrum- and Metallo-beta-lactama- 40 sample obtained from a healthy individual. Preferably, ses, metabolites of pharmaceutical substances, and xe- the healthy individual has normal functional values of co- nobiotics. prostanol, urobilins, mucin, fecal tryptic activity, short [0021] Preferably the microbiota in the composition ac- chain fatty acids and beta-aspartylglycine. Also prefera- cording to the present invention have not been exposed bly, the sample should be obtained prior to 1995. More to antibiotics taken into clinical use after 1995. 45 preferably the stool sample is investigated to be free from [0022] This invention comprises an anaerobically cul- hepatitis viruses A, B and C, cytomegalovirus, Epstein- tivated human intestinal microbiota for the pharmaceuti- Barr virus, human immunodeficiency virus (HIV), Calici- cal treatment of gastrointestinal and/or systemic/meta- and Rotavirus. Furthermore, the stool sample is bolic disorders that has its origin in a dysfuntional gas- screened to be free from Salmonella, Shigella, Campy- trointestinal ecosystem. 50 lobacter, Yersinia, Clostridium difficile and the presence [0023] An advantage with the composition according of protozoan cysts. to the present invention is that it consists of functional [0034] The cultivating medium used in the method ac- micro-ecological system that is stable and homogenous cording to the invention promotes the proliferation of the during 15 years of cultivation. No other bacterial strains anaerobic bacteria of at least three of the following four enter the system during cultivation. The microbial strains 55 Phyla: Bacterioidetes, Firmicutes, Proteobacteria and in the micro-ecological system interact and communicate Actinobacteria. with each other. A micro-ecological system is homoge- [0035] In one embodiment the cultivation medium is a nous. yeast based medium, preferably a bacteriological pep-

4 5 EP 2 750 682 B1 6 tone yeast based medium. The cultivation medium com- Bacteria;"Bacteroidetes";"Bacteroidia";"Bacteroi- prises added cholesterol, preferably cholesterol of ani- dales";"Prevotellaceae";Prevotella, Bacteria;"’Bacter- mal origin. Even more preferably the medium comprises oidetes";"Bacteroidia"-,"Bacteroidales";"Rikenellace- added freeze dried hen yolk, preferably in an amount of ae";Alistipes, from 0.5 to 5 % more preferably 1.25 % w/v (weight per 5 Bacteria;"Bacteroidetes";"Bacteroidia";"Bacteroidales"; volume). Bacteroidaceae;Bacteroides , [0036] Preferably, the cultivation contains an anaero- Bacteria;"Firmicutes";"Bacilli";"Lactobacillales";"Entero- bic indicator, preferably resazurin. coccaceae’;Enterococcus, Bacteria;"Firmi- [0037] The cultivating medium used in the method ac- cutes";"";Clostridiales;"Eubacteriaceae";Eu- cording to the invention promotes the proliferation of the 10 bacterium Bacteria.;"Firmicutes";"Clostridia";Clostridi- anaerobic bacteria of at least three of the following four ales;"Eubacteriaceae";unclassified_"Euba cteriaceae", Phyla: Bacterioidetes, Firmicutes, Proteobacteria and Bacteria;"Firmicutes";"Clostridia";Clostridiales;"Lachno- Actinobacteria in one and the same sample. Further, the spiraceae";Anaerostipes, Bacteria;"Firmi- preferred cultivating conditions suppresses the prolifer- cutes";"Clostridia";Clostridiales;"Lachnospiraceae";Do- ation of aerobic bacteria. 15 rea, Bacteria;"Firmicutes";"Clostridia";Clostridi- [0038] In a preferred embodiment the composition is ales;,"Lachnospiraceae",Roseburia, Bacteria;"Firmi- re-cultivatedunder strict anaerobic conditions,preferably cutes";"Clostridia";Clostridiales;"Lachnospirace- under nitrogen flow every second week. ae";unclassified_"Lac hnospiraceae", [0039] Preferably, the faecal sample is anaerobically Bacteria;"Firmicutes";"Clostridia";Clostridiales;"Pepto- cultivated for at least 15 years. 20 streptococcaceae";Sporacetig enium, [0040] Cultivating a faecal sample over time according Bacteria;"Firmicutes";"CIostridia";Clostridiales;"Peptost to the method of the present invention promotes the pro- reptococcaceae" ;unclassifie d_"Peptostreptococcace- liferation of the desired bacteria, while aerobic bacteria ae", Bacteria;"Firmicutes’;"Clostridia";Clostridiales;"Ru- are suppressed. minococcaceae";, Bacteria;"Firmi- [0041] An advantage with the method according to the 25 cutes";"Clostridia";Clostridiales;"Ruminococcace- present invention is that it enables proliferation of all de- ae";, Bacteria;"Firmicutes";"Clostrid- sired bacteria in one and the same sample. Thus, no ia";Clostridiales;"";Oscillibacter, Bac- mixing of different cultures is necessary. This has the teria;"Firmicutes";"Clostridia";Clostridiales;"Ruminococ- advantage that the obtained composition comprises a caceae";unclassified_"R uminococcaceae", micro-ecological system that has a stabilizing effect on 30 Bacteria;"Firmicutes";"Clostridia";Clostridiales;Clostrid- the intestinal microbiota and is able to re-establish the iaceae;"Clostridiaceae1";Clo stridium, communication between the host and the intestine. Fur- Bacteria;"Firmicutes";"Clostridia,,;Clostridiales;Clostrid- ther, by anaerobically cultivating microbiota in one sam- iaceae;"Clostridiaceae1";Sar cina, ple, according to the claimed invention, the same micro- Bacteria;"Firmicutes";"Clostridia";Clostridiales;Clostrid- ecologicalsystem isreproduced in each compositionwith 35 iaceae;"Clostridiaceae1";uncl retained physiological and/or pharmaceutical effect over assified_"Clostridiaceae1", a long time, such as over at least 15 years. Racteria;"Firmicutes";"Clostridia";Clostridiales;lncertae- [0042] Further, the micro-ecological system according SedisXI;Anaerococcus, Bacteria;"Firmicutes";"Clostrid- to the present invention is cultivated under strict anaer- ia";Clostridiales;IncertaeSedisXI;Finegoldia, Bacte- obic conditions. 40 ria;"Firmicutes";"Clostridia";Clostridiates;IncertaeSedis- [0043] The human intestinal flora consists of several XI;Peptoniphilus, Bacteria;"Firmicutes";"Clostrid- hundreds of different microbes, of which the vast majority ia";Clostridiales;IncertaeSedisXI;unclassified_Incerta are strictly anaerobic. eSedisXI, In one embodiment of the invention the following bacterial Bacteria;"Firmicutes";"Clostridia";Clostridiales;Incertae- together comprise more than 1% preferably45 SedisXIII;Anaerovorax, Bacteria;"Firmicutes";"Clostrid- more than 20%, and more preferably more than 50% of ia";Clostridiales;IncertaeSedisXIII;Mogibacterium, Bac- the total bacterial content of the composition; teria;"Firmicutes";"Clostridia";Clostridiales;IncertaeSe- Bacteria;"Actinobacteria";Actinobacteria;Coriobacteri- disXIV;Blautia, Bacteria;"Firmicutes";"Clostrid- dae;Coriobacteriales;"Corioba cterineae";Coriobacte- ia";Clostridiales;unclassifed_Clostridiales, Bacteria;"Fir- riaceae;Collinsella, 50 micutes";"Clostridia";Clostridiales;Veillonellaceae;Veil- Bacteria;"Actinobacteria";Actinobacteria;Coriobacteri- lonella, Bacteria;"Firmicutes";"Erysipelotrichi";"Erysip- dae;Coriobacteriales;"Corioba cterineae";Coriobacte- elotrichales";Erysipelotrichaceae;Hold emania, riaceae;Eggerthella, Bacteria;"Proteobacteria";Betaproteobacteria;Burkhol- Bacteria;"Actinobacteria";Actinobacteria;Coriobacteri- deriales;Alcaligenaceae;Para sutterella, dae;Coriobacteriales;"Corioba cterineae";Coriobacte- 55 [0044] Diseases that can be treated or prevented by riaceae;Olsenella, the invention are for example diarrhoeas following anti- Bacteria;"Bacteroidetes";"Bacteroidia";"Bacteroi- biotic treatment, especially diarrhoeas caused by dales";"Porphyromonadaceae";Par abacteroides, Clostridium difficile which can be very long lasting and

5 7 EP 2 750 682 B1 8 difficult to treat by conventional methods and can in rare Means of administration cases lead to death. Other therapeutic areas are Irritable Bowel Syndrome (IBS), Celiac Disease and Inflammato- [0050] The product can preferably be administered ac- ry Bowel Diseases for example Ulcerative Colitis, cording to the following non limiting examples: in the form Crohn’s Disease, Microscopic Colitis, and Pauchitis. An- 5 of a solution through e.g. a naso-duodenal tube, gastro- other treatment area is iatrogenic disturbance/dysbiosis scope, colo/sigmoideoscope, enema or in freeze dried of the intestinal microbiome, e.g., following radiation ther- form in a suitable galenic preparation, e.g. a gastric acid apy, chemotherapy and in connection with transplanta- resistant capsule, nano-encapsulated or a suppository. tions. Examples of other diseases that can be treated or prevented with the invention are: neurological diseases 10 Other ingredients (vehicles) for example Parkinson’s Disease, Alzheimer’s Disease, Lou Gehrig’s Disease ALS-Amyotrophic Lateral Sclero- [0051] Other substances and components which facil- sis, Multiple Sclerosis; behavioural/psychiatric disorders itates the administration or supports the effect of the in- e.g. Autism, Asperger’s Syndrome , Attention Deficit Hy- vention may be added to the composition. peractivity Disorder (ADHD) and Depression; Rheuma- 15 The treatment with the invention may or may not be pre- tologic diseases e.g. Rheumatoid arthritis; and system- ceded by an antibiotic treatment and/or motility reducing ic/metabolic disorders for example: Hypertension, Obes- substances. itas and type-2 Diabetes, and neoplastic/tumour diseas- [0052] The invention is described by the following non es of the gastrointestinal tract. The invention can also be limiting examples. used in the treatment of Chronic Fatigue Syndrome. The 20 invention can also be used in the treatment of dermato- Example1 logical diseases such as acne vulgaris. [0045] Also described is the use of such a anaerobi- [0053] Microbiota according to the present invention cally cultivated human microbiota for the production of a were obtained from a stool sample from a healthy single pharmaceutical treatment or prevention of disease. 25 donor prior to 1995. Both the donor and the stool were [0046] The composition according to the present in- thoroughly examined. The functionalstatus of the donor’s vention maybe administrated to a human by a device intestinal microbiota were found to have normal function- selected from a naso-duodenal tube, gastroscope, co- al values of coprostanol, urobilins, mucin, fecal tryptic lo/sigmoideoscope, and enema, or in freeze dried form activity, short chain fatty acids and beta-aspartylglycine. in a suitable galenic preparation, e.g. gastric acid resist- 30 The tests for hepatitis viruses A, B and C, cytomegalo- ant capsule, nano-encapsulation or suppository. virus, Epstein-Barr virus, human immunodeficiency virus [0047] In one embodiment the composition may also (HIV), Calici- and Rotavirus were all negative. Further- contain other microbial factors (e.g. bacteriophages) or more, the faeces was screened for presence of Salmo- signalling substances active against diseases. nella, Shigella, Campylobacter, Yersinia, Clostridium dif- 35 ficile and protozoan cysts, and all these investigations Forms of preparations were negative. [0054] A sample of faeces from the interior of the stool The composition may be provided in the following forms sample (Bristol scale type 2) was taken from the above- of preparations: mentioned healthy donor and immediately suspended in 40 30 ml of a pre-reduced sterilized bacteriological peptone [0048] yeast based medium (Difco, USA), with addition of 1.25 % (w/v) freeze dried hen yolk, Fresenius-Kabi Sweden 1. Freshly thawn from frozen solution with resazurin as an anaerobic indicator. This composi- 2. Freeze-dried cultivated product tion was re-cultivated under nitrogen flow every second 3. Fresh from the culture 45 week. The bacterial content of the cultivated product 4. Live micro-encapsulated culture were found to be > 109 per ml medium. [0055] The product contained the following species: Dosage Bacteria;"Actinobacteria";Actinobacteria;Coriobac- [0049] The invention is applied to the human gastro- 50 teridae;Coriobacteriales;"Corioba cterineae";Corio- intestinal tract at least one time per day during the course bacteriaceae;Collinsella, of a number of days, e.g. 1-30 days, depending on the Bacteria;"Actinobacteria";Actinobacteria;Coriobac- nature, cause and severity of the problem. Per adminis- teridae;Coriobacteriales;"Corioba cterineae";Corio- tration 1-100 ml of the product is given, preferably 20-40 bacteriaceae;Eggerthella, ml, or the equivalent amount in freeze-dried form. 55 Bacteria;"Actinobacteria";Actinobacteria;Coriobac- teridae;Coriobacteriales;"Corioba cterineae";Corio- bacteriaceae;Olsenella, Bactecia;"Bacteroidetes";"Bacteroidia";"Bacteroi-

6 9 EP 2 750 682 B1 10 dales";"Porphyromonadaceae";Par abacteroides, certaeSedisXIV;Blautia, Bacteria;"Firmicutes";"Clostridia";Clostridi- Bacteria;"Bacteroidetes";"Bacteroidia" ;"Bacteroida ales;unclassified_Clostridiales, les";"Prevotellaceae";Prevotella, Bacteria;"Firmicutes";"Clostridia";Clostridiafes;Veil- Bacteria;"Bacteroidetes";"Bacteroidia";"Bacteroi- 5 lonellaceae;Veillonella, dales";"Rikenellaceae";Alistipes, Bacteria;"Firmicutes";"Erysipelotrichi";"Erysipelot- Bacteria;"Bacteroidetes";"Bacteroidia";"Bacteroi- richales";Erysipelotrichaceae;Hold emania, and dales";Bacteroidaceae;Bacteroides Bacteria;"Proteobacteria";Betaproteobacteria;Bur- Bacteria;"Firmicutes";"Bacilli";"Lactobacilla- kholderiales;Alcaligenaceae;Para sutterella, les";"Enterococcaceae";Enterococcus, 10 Bacteria;"Firmicutes";"Clostridia";Clostridiales;"Eu- [0056] After addition of 10% glycerol the composition bacteriaceae";Eubacterium can be stored at -70°C. Bacteria;"Firmicutes";"Clostridia’;Clostridiales;"Eu- bacteriaceae";unclassified_"Euba cteriaceae", Example 2 Bacteria;"Firmicutes";"Clostridia";Clostridi- 15 ales;"Lachnospifaceae";Anaerostipes, [0057] The fresh or frozen stool sample from an indi- Bacteria;"Firmicutes";"Clostridia";Clostridi- vidual who has not been exposed to antibiotics or anti- ales;"Lachnospiraceae";Dorea, biotic resistance genes e.g. through a bacterial infection, Bacteria;"Firmicutes";"Clostridia";Clostridi- is inoculated under anaerobic conditions on to 30 ml pep- ates;"Lachnospiraceae";Roseburia, 20 tone-yeast medium (Difco,USA), containing cholesterol Bacteria;"Firmicutes";"Clostridia";Clostridi- of animal origin ( e.g. 1.25% freeze-dried hen yolk, Fre- ales;"Lachnospiraceae";unclassified_"Lac hnos- senius-Kabi,Sweden), with resazurin as an anaerobic in- piraceae", dicator. This composition is re-cultivated every second Bacteria;"Firmicutes";"Clostridia";Clostridi- week. The bacterial content of the cultivated product ales;"Peptostreptococcaceae";Sporacetig enium, 25 should be > 106 preferably > 109 per ml medium. After Bacteria;"Firmicutes";"Clostridia";Clostridi- addition of 10% glycerol the composition can be stored ales;"Peptostreptococcaceae";unclassifie d_"Pep- at -70°C. tostreptococcaceae", Bacteria;"Firmicutes";"Clostridia";Clostridiales;"Ru- Example 3 minococcac,eae";Anaerofilum, 30 Bacteria;"Ficmicutes";"Clostridia";Clostridi- [0058] A number of 32 patients, aged 27-94 years (20 ales;"Ruminococcaceae";Anaerotruncus, females, median age 78 years and 12 males, median Bacteria;"Firmicutes";"Clostridia";Clostridiales;"Ru- age 75 years) were treated with a composition according minococcaceae";Oscillibacter, to the invention. Out of these patients, 22 had underlying Bacteria;"Firmicutes";"Clostridia";Clostridiales;"Ru- 35 chronic diseases as cancer (breast, colon, prostate) and minococcaceae";unclassified-"R uminococcaceae", cardiovascular or pulmonary diseases. In 11 patients ce- phalosporins were given, in 9 patients clindamycin were Bacteria;"Firmicutes";"Clostridia";Clostridiales;Clos given, in 4 patients ciprofloxacin were given, in 2 patients tridiaceae;"Clostridiaceae1 ";Clo stridium, isoxazolylpenicillin were given, another 3 patients were 40 given some other antibiotics, and finally, 1 patient was Bacteria;"Firmicutes";"Clostridia";Clostridiales;Clos given azitromycin when recurrent Clostridium Difficile in- tridiaceae;"Clostridiaceae1 ";Sar cina, fection (RCDI) first was diagnosed. 2 patients did not Bacteria;"Firmicutes";"Clostridia";Clostridi- obtain specific antibiotic pretreatment before receiving ales;Clostridiaceae;"Closstridiaceae1";uncl the composition according to the invention. assified_"Clostridiaceae1", 45 [0059] Routinely, a volume of 30 ml of the invention Bacteria;"Firmicutes";"Clostridia";Clostridi- according to Example 1 was given rectally by a rectally ales;lncertaeSedisXI;Anaerococcus, introduced catheter in Sigmoideum or by colonoscopy. Bacteria;"Firmicutes";"Clostridia";Clostridiales;In- Analysis of clostridium toxin was performed according to certaeSedisXI;Finegoldia, general practice at Karolinska Hospital, Huddinge, Swe- Bacteria;"Firmicutes";"Clostridia";Clostridiales;In- 50 den. Out of the 32 patients receiving the invention, 22 certaeSedisXI;Peptoniphilus, patients were cured. These 22 patients experienced a Bacteria;"Firmicutes";"Clostridia";Clostridiales;In- dramatic improvement in quality of life from living with certaeSedisXI;unclassified_Incerta eSedisXI, diarrhoeaduring several months to a life with normal bow- Bacteria;"Firmicutes";"Clostridia";Clostridiales;In- el habits within some few days. Fifteen of these patients certaeSedisXIII;Anaerovorax, 55 received the invention only once, the remaining received Bacteria;"Firmicutes";"Clostridia";Clostridi- the invention one to two more times. Four more patients ales;lncertaeSedisXIII;Mogibacterium, experienced a considerable improvement, and in the re- Bactecia;"Firmicutes";"Clostridia";Clostridiales;In- maining 4 patients repeated antibiotic therapies had to

7 11 EP 2 750 682 B1 12 be given. Another 2 patients with unknown cause of the Bacteria;"Bacteroidetes";"Bacteroidia";"Bacter- diarrhoea were also cured. The follow-up time was been oidales";"Rikenellaceae";Alistipes, 5-68 months, median 26 months. No adverse effects by Bacteria;"Bacteroidetes";"Bacteroidia";"Bacter- the procedure were observed. oidales";Bacteroidaceae; Bacteroides, 5 Bacteria;"Firmicutes";"Bacilli";"Lactobacilla- Example 4 les";"Enterococcaceae"; Enterococcus, Bacteria;"Firmicutes";"Clostridia";Clostridi- [0060] Ten patients with post-infectious gastroenteritis ales;"Eubacteriaceae";Eubacterium, related to Giardia lamblia, were suffering from IBS like Bacteria;"Firmicutes";"Clostridia";Clostridi- problems. Conventional IBS treatment was not success- 10 ales;"Eubacteriaceae";unclassified_ "Eubacte- ful. It was decided that these patients should receive the riaceae", composition according to the invention. One dose of 30 Bacteria;"Firmicutes";"Clostridia";Clostridi- ml was administrated over three consecutive days, using ales;"Lachnospiraceae";Anaerostipes, a gastro-jejunal tube. Surprisingly, all patients recovered Bacteria;"Firmicutes";"Clostridia";Clostridi- shortly after the administration of the invention, and re- 15 ales;"Lachnospiraceae";Dorea, mained without symptoms for a long period of time. Bacteria;"Firmicutes";"Clostridia";Clostridi- ales;"Lachnospiraceae";Roseburia, Bacteria;"Firmicutes";"Clostridia";Clostridi- Claims ales;"Lachnospiraceae";unclassified _"Lachno- 20 spiraceae", 1. A composition comprising strictly anaerobically cul- Bacteria;"Firmicutes";"Clostridia";Clostridi- tivated human intestinal microbiota selected from at ales;"Peptostreptococcaceae";Spor aceti- least three of the four Phyla,Bacterioidetes, Firmi- genium, cutes, Proteobacteria and Actinobacteria for use in Bacteria;"Firmicutes";"Clostridia";Clostridi- the prevention and treatment of diseases. 25 ales;"Peptostreptococcaceae";uncla ssified_"Peptostreptococcaceae", 2. The composition according to claim 1,character- Bacteria;"Firmicutes";"Clostridia";Clostridi- ised in that it is free from extended spectrum-and ales;"Ruminococcaceae";Anaerofilum, metallo-beta-lactamases, metabolites of pharma- Bacteria;"Firmicutes";"Clostridia";Clostridi- ceutical substances, or xenobiotics. 30 ales;"Ruminococcaceae"; Anaerotruncus, Bacteria;"Firmicutes";"Clostridia";Clostridi- 3. The composition according to any one of claims 1-2, ales;"Ruminococcaceae";Oscillibacter, characterised in that it is free from the following Bacteria;"Firmicutes";"Clostridia";Clostridi- microbes: hepatitis viruses A, B and C, cytomegalo ales;"Ruminococcaceae";unclassified_"Ru mi- virus, Epstein-Barr virus, human immunodeficiency 35 nococcaceae", virus (HIV), Calici- and Rotavirus, Salmonella, Shig- ella, Campylobacter, Yersinia, and protozoan cysts. Bacteria;"Firmicutes";"Clostridia";Clostridiales; Clostridiaceae;"Clostridiaceae1 ";Clostridium, 4. The composition according to any one of claims 1-3, characterised in that thefollowing bacterial species 40 Bacteria;"Firmicutes";"Clostridia";Clostridiales; together comprise morethan 1 %of thetotal bacterial Clostridiaceae;"Clostridiaceae1 ";Sarcina, content of the invention: Bacteria;"Firmicutes";"Clostridia";Clostridiales; Bacteria;"Actinobacteria";Actinobacteria;Corio- Clostridiaceae;"Clostridiaceae1 ";unclassified_ bacteridae;Coriobacteriales;"Co riobacterine- 45 "Clostridiaceae1", ae";Coriobacteriaceae;Collinsella, Bacteria;"Firmicutes";"Clostridia";Clostridi- Bacteria;"Actinobacteria";Actinobacteria;Corio- ales;IncertaeSedisXI;Anaerococcus, bacteridae;Coriobacteriales;"Co riobacterine- Bacteria;"Firmicutes";"Clostridia";Clostridi- ae";Coriobacteriaceae; Eggerthella, ales;IncertaeSedisXI;Finegoldia, Bacteria;"Actinobacteria";Actinobacteria;Corio- 50 Bacteria;"Firmicutes";"Clostridia";Clostridi- bacteridae;Coriobacteriales;"Co riobacterine- ales;IncertaeSedisXI;Peptoniphilus, ae";Coriobacteriaceae;Olsenella, Bacteria;"Firmicutes";"Clostridia";Clostridi- ales;IncertaeSedisXI;unclassified_In certaeSe- Bacteria;"Bacteroidetes";"Bacteroidia";"Bacter disXI, oidales";"Porphyromonadaceae ";Parabacteroi 55 Bacteria;"Firmicutes";"Clostridia";Clostridi- des, ales;IncertaeSedisXIII;Anaerovorax, Bacteria;"Bacteroidetes";"Bacteroidia";"Bacter- Bacteria;"Firmicutes";"Clostridia";Clostridi- oidales";"Prevotellaceae"; Prevotella, ales;IncertaeSedisXIII;Mogibacterium,

8 13 EP 2 750 682 B1 14

Bacteria;"Firmicutes";"Clostridia";Clostridi- trum- und Metallo-beta-Laktamasen, Metaboliten ales;IncertaeSedisXIV;Blautia, pharmazeutischer Substanzen oder Xenobiotika ist. Bacteria;"Firmicutes";"Clostridia";Clostridi- ales;unclassified_Clostridiales, 3. Zusammensetzung gemäß irgendeinem der An- Bacteria;"Firmicutes";"Clostridia";Clostridi- 5 sprüche 1 bis 2, dadurch gekennzeichnet, dass ales;Veillonellaceae;Veillonella, sie frei von folgenden Mikroben ist: Bacteria;"Firm icutes";"Erysipelotrichi";"Erysipelotrichales";Er Hepatitis-Viren A, B und C, Zytomegalie-Virus, ysipelotrichaceae ;Holdemania, and Epstein-Barr-Virus, humanes Immundefizienz- Bacteria;"Proteobacteria";Betaproteobacte- 10 Virus (HIV), Calici- und Rotavirus, Salmonella, ria;Burkholderiales;Alcaligenaceae; Parasut- Shigella, Campylobacter, Yersinia und Protozo- terella. en-Zysten.

5. The composition according to any one of claims 1-3, 4. Zusammensetzung gemäß irgendeinem der An- characterised in that the bacterial content of the 15 sprüche 1 bis 3, dadurch gekennzeichnet, dass composition comprises one or more of the bacterial die folgenden bakteriellen Spezies zusammen mehr groups mentioned in claim 4. als 1 % des gesamten Bakteriengehalts der Erfin- dung ausmachen: 6. The composition according to any one of claims1-5 for use in the treatment of diarrhoea, such as Irritable 20 Bacteria; "Actinobacteria"; Actinobacteria; Cori- Bowel Syndrome (IBS), or antibiotic associated di- obacteridae; Coriobacteriales; "Coriobacterine- arrhea(AAD), especially related to Clostridium ae"; Coriobacteriaceae; Collinsella, difficile , Clostridium difficile associated diarrhoea Bacteria; "Actinobacteria"; Actinobacteria; Cori- ( CDAD) and recurrent Clostridium difficile infections obacteridae; Coriobacteriales; "Coriobacterine- (RCDI). 25 ae"; Coriobacteriaceae; Eggerthella, Bacteria; "Actinobacteria"; Actinobacteria; Cori- 7. The composition according to any one of claims 1-6 obacteridae; Coriobacteriales; "Coriobacterine- for use in administration to a human by a device se- ae"; Coriobacteriaceae; Olsenella, lected from a naso-duodenal tube, gastroscope, co- Bacteria; "Bacteroidetes"; "Bacteroidia"; "Bac- lo/sigmoideoscope, and enema, or in freeze dried 30 teroidales"; "Porphyromonadaceae"; Parabac- form in a suitable galenic preparation, e.g. gastric teroides, acid resistant capsule, nano-encapsulated or sup- Bacteria; "Bacteroidetes"; "Bacteroidia"; "Bac- pository. teroidales"; "Prevotellaceae"; Prevotella, Bacteria; "Bacteroidetes"; "Bacteroidia";"Bacte- 8. A method for preparing a composition comprising 35 roidales"; "Rikenellaceae"; Alistipes, strictly anaerobically cultivated human intestinal Bacteria; "Bacteroidetes"; "Bacteroidia"; "Bac- microbiota selected from at least three of the four teroidales"; Bacteroidaceae; Phyla, Bacterioidetes, Firmicutes, Proteobacteria Bacteroides, and Actinobacteria, the method characterised by Bacteria; "Firmicutes"; "Bacilli"; "Lactobacilla- anaerobic cultivation of a stool sample in a cultivation 40 les"; "Enterococcaceae"; medium that promotes the proliferation of the anaer- Enterococcus, obic bacteria of at least three of the following four Bacteria; "Firmicutes"; "Clostridia"; Clostridia- Phyla: Bacterioidetes, Firmicutes, Proteobacteria les; "Eubacteriaceae"; Eubacterium, and Actinobacteria, and characterised in that aer- Bacteria; "Firmicutes"; "Clostridia"; Clostridia- obic bacteria are suppressed during cultivation. 45 les; "Eubacteriaceae"; nicht klassifizierte "Eu- bacteriaceae", Bacteria; "Firmicutes"; "Clostridia"; Clostridia- Patentansprüche les; "Lachnospiraceae"; Anaerostipes, Bacteria; "Firmicutes"; "Clostridia"; Clostridia- 1. Zusammensetzung, welche streng anaerob kulti- 50 les; "Lachnospiraceae"; Dorea, vierte humane intestinale Mikrobiota, ausgewählt Bacteria; "Firmicutes"; "Clostridia"; Clostridia- aus wenigstens drei der vier Stämme (Phyla) Bac- les; "Lachnospiraceae"; Roseburia, terioidetes, Firmicutes, Proteobacteria und Actino- Bacteria; "Firmicutes"; "Clostridia"; Clostridia- bacteria, umfasst, zur Verwendung bei der Vorbeu- les; "Lachnospiraceae"; nicht klassifizierte gung und Behandlung von Krankheiten. 55 "Lachnospiraceae", Bacteria; "Firmicutes"; "Clostridia"; Clostridia- 2. Zusammensetzung gemäß Anspruch 1,dadurch les; "Peptostreptococcaceae"; Sporacetigeni- gekennzeichnet, dass sie frei von Extended-Spec- um,

9 15 EP 2 750 682 B1 16

Bacteria; "Firmicutes"; "Clostridia"; Clostridia- sammenhang mit Clostridium difficile, Clostridium- les; "Peptostreptococcaceae"; nicht klassifizier- difficile-assoziierte Diarrhö (CDAD) und rezidivie- te "Peptostreptococcaceae", rende Clostridium-difficile-Infektionen (RCDI). Bacteria; "Firmicutes"; "Clostridia"; Clostridia- les; "Ruminococcaceae"; Anaerofilum, 5 7. Zusammensetzung gemäß irgendeinem der An- Bacteria; "Firmicutes"; "Clostridia"; Clostridia- sprüche 1 bis 6 zur Verwendung bei der Verabrei- les; "Ruminococcaceae"; chung an einen Menschen durch eine Vorrichtung, Anaerotruncus, ausgewählt aus nasoduodenaler Sonde, Gastros- Bacteria; "Firmicutes"; "Clostridia"; Clostridia- kop, Koloskop/Sigmoidoskop und Klistier, oder in les; "Ruminococcaceae"; Oscillibacter, 10 gefriergetrockneter Form in einer geeigneten gale- Bacteria; "Firmicutes"; "Clostridia"; Clostridia- nischen Präparation, z.B. Magensäure-resistenten les; "Ruminococcaceae"; nicht klassifizierte Kapsel, Nano-Verkapselung oder Suppositorium. "Ruminococcaceae", Bacteria; "Firmicutes"; "Clostridia"; Clostridia- 8. Verfahren zur Herstellung einer Zusammensetzung, les; Clostridiaceae; "Clostridiaceae 1 "; Clostri- 15 welche streng anaerob kultivierte humane intestina- dium, le Mikrobiota, ausgewählt aus wenigstens drei der Bacteria; "Firmicutes"; "Clostridia"; Clostridia- vier Stämme (Phyla) Bacterioidetes, Firmicutes, les; Clostridiaceae; "Clostridiaceae 1"; Sarcina, Proteobacteria und Actinobacteria, umfasst, wobei Bacteria; "Firmicutes"; "Clostridia"; Clostridia- das Verfahren gekennzeichnet ist durch die ana- les; Clostridiaceae; "Clostridiaceae 1"; nicht20 eorobe Kultivierung einer Stuhlprobe in einem Kul- klassifizierte "Clostridiaceae 1", turmedium, welches die Proliferation der anaeoro- Bacteria; "Firmicutes"; "Clostridia"; Clostridia- ben Bakterien von wenigstens drei der folgenden les; Incertae Sedis XI; Anaerococcus, vier Stämme (Phyla) fördert: Bacterioidetes, Firmi- Bacteria; "Firmicutes"; "Clostridia"; Clostridia- cutes, Proteobacteria und Actinobacteria, undda- les; Incertae Sedis XI; Finegoldia, 25 durch gekennzeichnet ist, dass aerobe Bakterien Bacteria; "Firmicutes"; "Clostridia"; Clostridia- während der Kultivierung unterdrückt werden. les; Incertae Sedis XI; Peptoniphilus, Bacteria; "Firmicutes"; "Clostridia"; Clostridia- les; Incertae Sedis XI; nicht klassifizierte Incer- Revendications tae Sedis XI, 30 Bacteria; "Firmicutes"; "Clostridia"; Clostridia- 1. Composition comprenant un microbiote intestinal les; Incertae Sedis XIII; Anaerovorax, humain cultivé strictement dans des conditions ana- Bacteria; "Firmicutes"; "Clostridia"; Clostridia- érobies sélectionné parmi au moins trois des quatre les; Incertae Sedis XIII; Mogibacterium, phylums Bacteroidetes, Firmicutes, Proteobactéria Bacteria; "Firmicutes"; "Clostridia"; Clostridia- 35 et Actinobacteria pour une utilisation dans la préven- les; Incertae Sedis XIV; Blautia, tion et le traitement de maladies. Bacteria; "Firmicutes"; "Clostridia"; Clostridia- les; nicht klassifizierte Clostridiales, 2. Composition selon la revendication 1, caractérisée Bacteria; "Firmicutes"; "Clostridia"; Clostridia- en ce qu’ elle est exempte de métallo-bêta-lactama- les; Veillonellaceae; Veillonella, 40 ses et de bêtalactamases à spectre élargi, de méta- Bacteria; "Firmicutes"; "Erysipelotrichi"; "Erysi- bolites de substances pharmaceutiques ou de xé- pelotrichales"; Erysipelotrichaceae; Holdema- nobiotiques. nia, und Bacteria; "Proteobacteria"; Betaproteobacteria; 3. Composition selon l’une quelconque des revendica- Burkholderiales; Alcaligenaceae; Parasutterel- 45 tions 1 à 2, caractérisée en ce qu’elle est exempte la. des microbes suivants : virus de l’hépatite A, B et C, cytomégalovirus, virus d’Epstein-Barr, virus de l’im- 5. Zusammensetzung gemäß irgendeinem der An- munodéficience humaine (VIH), Calici- et Rotavirus, sprüche 1 bis 3, dadurch gekennzeichnet, dass Salmonella, Shigella, Campylobacter, Yersinia, et der bakterielle Gehalt der Zusammensetzung eine 50 kystes de protozoaires. oder mehrere der in Anspruch 4 genannten Bakte- riengruppen einschließt. 4. Composition selon l’une quelconque des revendica- tions 1 à 3, caractérisée en ce que les espèces 6. Zusammensetzung gemäß irgendeinem der An- bactériennessuivantes comprennent ensemble plus sprüche 1 bis 5 zur Verwendung bei der Behandlung 55 de 1 % de la teneur en bactéries totale de l’invention : von Diarrhö, wie beispielsweise Reizdarmsyndrom ("Irritable Bowel Syndrome", IBS) oder Antibioti- Bacteria;"Actinobacteria";Actinobacteria;Corio- kaassoziierte Diarrhö (AAD), insbesondere im Zu- bacteridae;Co riobacteriales;"Co riobacteri-

10 17 EP 2 750 682 B1 18 neae";Coriobacteriaceae;Collinsella, les;Clostridia ceae;"Clostridiaceae1 "; non Bacteria;"Actinobacteria";Actinobacteria;Corio- classifiés_"Clostridiaceae1", bacteridae;Co riobacteriales;"Co riobacteri- Bacteria;"Firmicutes";"Clostridia";Clostridia- neae";Coriobacteriaceae; Eggerthella, les;IncertaeSe disXI;Anaerococcus, Bacteria;"Actinobacteria";Actinobacteria;Corio- 5 Bacteria;"Firmicutes";"Clostridia";Clostridia- bacteridae;Co riobacteriales;"Co riobacteri- les;IncertaeSe disXI;Finegoldia, neae";Coriobacteriaceae;Olsenella, Bacteria;"Firmicutes";"Clostridia";Clostridia- Bacteria;"Bacteroidetes";"Bacteroidia";"Bacte- les;IncertaeSe disXI;Peptoniphilus, roidales";"Por phyromonadaceae Bacteria;"Firmicutes";"Clostridia";Clostridia- ";Parabacteroides, 10 les;IncertaeSe disXI; non classifiés_In Bacteria;"Bacteroidetes";"Bacteroidia";"Bacte- certaeSedisXI, roidales";"Pre votellaceae"; Bacteria;"Firmicutes";"Clostridia";Clostridia- Prevotella, les;IncertaeSe disXIII;Anaerovorax, Bacteria;"Bacteroidetes";"Bacteroidia";"Bacte- Bacteria;"Firmicutes";"Clostridia";Clostridia- roidales";"Rik enellaceae";Alistipes, 15 les;IncertaeSe disXIII;Mogibacterium, Bacteria;"Bacteroidetes";"Bacteroidia";"Bacte- Bacteria;"Firmicutes";"Clostridia";Clostridia- roidales";Bact eroidaceae; les;IncertaeSe disXIV;Blautia, Bacteroides, Bacteria;"Firmicutes";"Clostridia";Clostridiales; Bacteria;"Firmicutes";"Bacilli";"Lactobacilla- non classifiés Clostridiales, les";"Enteroco ccaceae"; 20 Bacteria;"Firmicutes";"Clostridia";Clostridia- Enterococcus, les;Veillonell aceae;Veillonella, Bacteria;"Firmicutes";"Clostridia";Clostridia- les;"Eubacteri aceae";Eubacterium, Bacteria;"Firmicutes";"Erysipelotrichi";"Erysipe Bacteria;"Firmicutes";"Clostridia";Clostridia- lotrichales ";Erysipelotrichaceae les;"Eubacteri aceae"; non classifiés _ 25 ;Holdemania, et "Eubacteriaceae", Bacteria;"Proteobacteria";Betaproteobacte- Bacteria;"Firmicutes";"Clostridia";Clostridia- ria;Burkholderiale s;Alcaligenaceae; les;"Lachnospi raceae";Anaerostipes, Bacteria;"Firmicutes";"Clostridia";Clostridia- Parasutterella. les;"Lachnospi raceae";Dorea, 30 Bacteria;"Firmicutes";"Clostridia";Clostridia- 5. Composition selon l’une quelconque des revendica- les;"Lachnospi raceae";Roseburia, tions 1 à 3, caractérisée en ce que le contenu bac- Bacteria;"Firmicutes";"Clostridia";Clostridia- térien de la composition comprend un ou plusieurs les;"Lachnospi raceae"; non classifiés des groupes de bactéries mentionnés dans la reven- _"Lachnospiraceae", 35 dication 4. Bacteria;"Firmicutes";"Clostridia";Clostridia- les;"Peptostre ptococcaceae";Spor 6. Composition selon l’une quelconque des revendica- acetigenium, tions 1 à 5, pour une utilisation dans le traitement Bacteria;"Firmicutes";"Clostridia";Clostridia- d’une diarrhée, telle que le syndrome du côlon irri- les;"Peptostre ptococcaceae";non 40 table (SCI), ou une diarrhée associée aux antibioti- classifiés_"Peptostreptococcaceae", ques (DAA), en particulier associée à Clostridium Bacteria;"Firmicutes";"Clostridia";Clostridia- difficile, une diarrhée associée à Clostridium difficile les;"Ruminococ caceae";Anaerofilum, (DACD) et les infections récurrentes à Clostridium Bacteria;"Firmicutes";"Clostridia";Clostridia- difficile (IRCD). les;"Ruminococ caceae"; 45 Anaerotruncus, 7. Composition selon l’une quelconque des revendica- Bacteria;"Firmicutes";"Clostridia";Clostridia- tions 1 à 6, pour une utilisation dans l’administration les;"Ruminococ caceae";Oscillibacter, à un être humain par un dispositif choisi parmi un Bacteria;"Firmicutes";"Clostridia";Clostridia- tube nasoduodénal, ungastroscope, un colo/sigmoï- les;"Ruminococ caceae";non classifiés_"Rumi- 50 déoscope, et un lavement ou sous une forme lyo- nococcaceae", philisée dans une préparation galénique appropriée, Bacteria;"Firmicutes";"Clostridia";Clostridia- par exemple une capsule résistant à l’acide gastri- les;Clostridia que, une nano-capsule ou un suppositoire. ceae;"Clostridiaceae1 ";Clostridium, Bacteria;"Firmicutes";"Clostridia";Clostridia- 55 8. Procédé de préparation d’une composition compre- les;Clostridia ceae;"Clostridiaceae1 nant un microbiote intestinal humain cultivé stricte- ";Sarcina, ment dans des conditions anaérobies sélectionné Bacteria;"Firmicutes";"Clostridia";Clostridia- parmi au moins trois des quatre phylums Bacteroi-

11 19 EP 2 750 682 B1 20 detes, Firmicutes, Proteobactéria et Actinobacteria, le procédé étant caractérisé par une culture anaé- robie d’un échantillon de selles dans un milieu de culture qui favorise la prolifération de bactéries ana- érobies d’au moins trois des quatre phylums5 suivants : Bacteroidetes, Firmicutes, Proteobactéria et Actinobacteria, et caractérisé en ce que les bac- téries anaérobies sont supprimées au cours de la culture. 10

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REFERENCES CITED IN THE DESCRIPTION

This list of references cited by the applicant is for the reader’s convenience only. It does not form part of the European patent document. Even though great care has been taken in compiling the references, errors or omissions cannot be excluded and the EPO disclaims all liability in this regard.

Patent documents cited in the description

• WO 0207741 A [0007] • WO 2011033310 A [0007]

Non-patent literature cited in the description

• JOHAN S. Bakken Fecal bacteriotherapy for recur- • A. GUSTAFSSON ; A. BERSTAD ; S. rent Clostridium difficile infection Anaerobe, 2009, LUND-TØNNESEN ; T. MIDTVEDT ; E. NORIN. The vol. 15, 285-289 [0009] Effect of Faecal Enema on Five Microflora-Associat- • C JORUP-RÖNSTRÖM ; A HÅKANSSON ; A-K ed Characteristics in Patients with Antibiotic-Associ- PERSON ; T MIDTVEDT ; E NORIN. Feceskultur ated Diarrhoea. Scand J Gastroenterol, 1999, vol. 34, framgångsrik terapi vid Clostridium difficile-diarré. 580-586 [0009] Läkartidningen, 2006, vol. 103 (46), 3603-3605 • A. GUSTAVSSON ; S. LUND-TONNESEN ; A. [0009] BERSTAD ; T. MIDTVEDT ; E. NORIN. Faecal Short-Chain Fatty Acids in Patients with Antibiot- ic-Associated Diarrhoea, before and after Faecal En- ema Treatment. Scand J Gastroenterol, 1998, vol. 33, 721-727 [0009]

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