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Title:
COMPOSITIONS AND METHODS FOR PROMOTING GUT HEALTH
Document Type and Number:
WIPO Patent Application WO/2019/157566
Kind Code:
A1
Abstract:
The present disclosure relates to compositions and methods for the treatment of gastrointestinal disorders such as ulcerative colitis by suppressing and/or promoting the growth of certain bacteria or the activity of certain metabolic pathways. In some embodiments, the disclosure relates to methods and compositions for use in faecal microbiota transplantation (FMT).

Inventors:
KAMM MICHAEL A (AU)
Application Number:
PCT/AU2019/050126
Publication Date:
August 22, 2019
Filing Date:
February 15, 2019
Export Citation:
Click for automatic bibliography generation   Help
Assignee:
ST VINCENTS HOSPITAL MELBOURNE LTD (AU)
International Classes:
A61K35/74; A61P1/00; C12N1/20; C12Q1/689
Domestic Patent References:
WO2017210428A12017-12-07
WO2017116235A12017-07-06
Other References:
ROSSEN, N.G.M. ET AL.: "The microbiome and its therapeutic potential in inflammatory bowel diseases. Chapter 7, Fecal transplantation in ulcerative colitis: microbiota shifts and signatures that determine long-term success of therapy", PHD THESIS UNIVERSITY OF AMSTERDAM, 2016, pages 136 - 170, XP055631489, Retrieved from the Internet [retrieved on 20190517]
FUENTES S. ET AL.: "Microbial shifts and signatures of long-term remission in ulcerative colitis after faecal microbiota transplantation", THE ISME JOURNAL, vol. 11, no. 8, 2017, pages 1877 - 1889, XP055631500
PARAMSOTHY S. ET AL.: "OP019 In faecal microbiota transplantation (FMT) for ulcerative colitis, fusobacterium is associated with a lack of remission, while metabolic shifts to starch degradation and short-chain fatty acid production are associated with remission (FOCUS study", JOURNAL OF CROHN'S AND COLITIS, vol. 12, no. 1, 16 January 2018 (2018-01-16), pages S013 - S014
MOAYYEDI P. ET AL.: "Fecal Microbiota Transplantation Induces Remission in Patients With Active Colitis in a Randomized Controlled Trial", GASTROENTEROLOGY, vol. 149, no. 1, 2015, pages 102 - 109, XP055631556
VALCHEVA R. ET AL.: "Beta-Fructans Reduce Inflammation in Mild to Moderate Ulcerative Colitis Through Specific Microbiota Changes Associated With Improved Butyrate Formation and MUC2 Expression", GASTROENTEROLOGY, vol. 142, no. 5, 2012, pages S-196, XP055631568
KANAUCHI O. ET AL.: "Germinated Barley Foodstuff Feeding", DIGESTION, vol. 63, no. 1, 2001, pages 60 - 67
EL HAGE R. ET AL.: "Emerging Trends in ''Smart Probiotics'': Functional Consideration for the Development of Novel Health and Industrial Applications", FRONTIERS IN MICROBIOLOGY, 1889, 29 September 2017 (2017-09-29), pages 1 - 11, XP055631575
OHKUSA T. ET AL.: "Newly Developed Antibiotic Combination Therapy for Ulcerative Colitis: A Double-Blind Placebo-Controlled Multicenter Trial", AM. J. GASTROENTEROL., vol. 105, no. 8, 2010, pages 1820 - 1829
Attorney, Agent or Firm:
LOKAN, Nigel (AU)
Download PDF:
Claims:
Claims

1. A method of performing FMT therapy on a subject in need thereof, the method comprising the step of administering a faecal sample to the subject, wherein the faecal sample is enriched with one or more bacteria.

2. The method of claim 1 wherein the one or more bacteria are selected from the group consisting of Eubacterium, Ruminococcus, Lachnospiraceae, Roseburia, Faecalibacterium, Eubacterium, Clostridium XVIII, Dorea, Coprococcus and Bacteroides fragilis.

3. The method of claim 1 wherein the one or more bacteria is a member of the Firmicutes phylum.

4. The method of any one of claims 1 to 3 wherein the one or more bacteria is Eubacterium hallii.

5. The method of any one of claims 1 to 4 wherein the one or more bacteria promote a metabolic activity in the subject, wherein the metabolic activity is selected from the group consisting of benzoate degradation, glycerophospholipid metabolism, secondary bile acid biosynthesis, ppGpp biosynthesis, pyruvate fermentation to acetate and lactate, fatty acid biosynthesis, propanoate metabolism, ansamycin biosynthesis and starch degradation.

6. A method of performing FMT therapy on a subject in need thereof, the method comprising the step of administering a faecal sample to the subject, wherein the faecal sample is enriched with Eubacterium hallii.

7. The method of any one of claims 1 to 6 wherein the subject in need thereof suffers from a gastrointestinal disorder.

8. The method of claim 7 wherein the gastrointestinal disorder is ulcerative colitis.

9. A method of performing FMT therapy on a subject in need thereof, the method comprising the step of administering a faecal sample to the subject, wherein the faecal sample is depleted in one or more bacteria.

10. The method of claim 9 wherein the one or more bacteria are selected from the group consisting of Fusobacterium, Sutterella, Veillonella, Haemophilus, Streptococcus, Bacteroides, Akkermansia, Escherichia, Megamonas, Clostridium XI Va, Prevotella, Dialister, Veillonella, Bilophila, Parvimonas, Bacteroides uniformis and Bacteroides coprocola.

1 1 . The method of claim 9 or claim 10 wherein the one or more bacteria is Fusobacterium spp.

12. The method of any one of claims 9 to 1 1 wherein the one or more bacteria is Fusobacterium gonidiaformans.

13. The method of any one of claims 9 to 12 wherein the one or more bacteria suppress a metabolic activity in the subject, wherein the metabolic activity is selected from the group consisting of heme biosynthesis, lipopolysaccharide biosynthesis, ubiquinone biosynthesis, terpenoid quinine biosynthesis, terpenoid backbone biosynthesis, lysine biosynthesis and oxidative phosphorylation.

14. A method of performing FMT therapy on a subject in need thereof, the method comprising the step of administering a faecal sample to the subject, wherein the faecal sample is depleted in Fusobacterium spp.

15. The method of any one of claims 9 to 14 wherein the subject in need thereof suffers from a gastrointestinal disorder.

16. The method of claim 15 wherein the gastrointestinal disorder is ulcerative colitis.

17. A faecal sample for use in FMT therapy, wherein the faecal sample is enriched with one or more bacteria.

18. The faecal sample of claim 17 wherein the one or more bacteria are selected from the group consisting of Eubacterium, Ruminococcus, Lachnospiraceae, Roseburia, Faecalibacterium, Eubacterium, Clostridium XVIII, Dorea, Coprococcus and Bacteroides fragilis.

19. The faecal sample of claim 17 wherein the one or more bacteria is a member of the Firmicutes phylum.

20. The faecal sample of any one of claims 17 to 19 wherein the one or more bacteria is Eubacterium hallii.

21 . The faecal sample of any one of claims 17 to 20 wherein the one or more bacteria promote a metabolic activity in the subject, wherein the metabolic activity is selected from the group consisting of benzoate degradation, glycerophospholipid metabolism, secondary bile acid biosynthesis, ppGpp biosynthesis, pyruvate fermentation to acetate and lactate, fatty acid biosynthesis, propanoate metabolism, ansamycin biosynthesis and starch degradation.

22. A faecal sample for use in FMT therapy, wherein the faecal sample is depleted in one or more bacteria.

23. The faecal sample of claim 22 wherein the one or more bacteria are selected from the group consisting of Fusobacterium, Sutterella, Veillonella, Haemophilus, Streptococcus, Bacteroides, Akkermansia, Escherichia, Megamonas, Clostridium XlVa, Prevotella, Dialister, Veillonella, Bilophila, Parvimonas, Bacteroides uniformis and Bacteroides coprocola.

24. The faecal sample of claim 22 or claim 23 wherein the one or more bacteria is Fusobacterium spp.

25. The faecal sample of any one of claims 22 to 24 wherein the one or more bacteria is Fusobacterium gonidiaformans.

26. The faecal sample of any one of claims 22 to 25 wherein the one or more bacteria suppress a metabolic activity in the subject, wherein the metabolic activity is selected from the group consisting of heme biosynthesis, lipopolysaccharide biosynthesis, ubiquinone biosynthesis, terpenoid quinine biosynthesis, terpenoid backbone biosynthesis, lysine biosynthesis and oxidative phosphorylation.

27. A method of screening a faecal sample to determine its suitability for use in FMT therapy, the method comprising testing the faecal sample for the presence or absence of one or more bacteria,

wherein when the one or more bacteria are detected, the faecal sample is deemed suitable for use in FMT therapy,

or wherein when the one or more bacteria are not detected, the faecal sample is deemed unsuitable for use in FMT therapy.

28. The method of claim 27 wherein the one or more bacteria are selected from the group consisting of Eubacterium, Ruminococcus, Lachnospiraceae, Roseburia, Faecalibacterium, Eubacterium, Clostridium XVIII, Dorea, Coprococcus and Bacteroides fragilis.

29. The method of claim 27 wherein the one or more bacteria is a member of the Firmicutes phylum.

30. The method of any one of claims 27 to 29 wherein the one or more bacteria is Eubacterium hallii.

31 . The method of any one of claims 27 to 30 wherein the one or more bacteria promote a metabolic activity in the subject, wherein the metabolic activity is selected from the group consisting of benzoate degradation, glycerophospholipid metabolism, secondary bile acid biosynthesis, ppGpp biosynthesis, pyruvate fermentation to acetate and lactate, fatty acid biosynthesis, propanoate metabolism, ansamycin biosynthesis and starch degradation.

32. The method of any one of claims 27 to 31 wherein the testing includes shotgun DNA sequencing, whole genome sequencing, PCR, 16S genomic DNA sequencing, 16S complementary DNA sequencing, 16S RNA sequencing or culturing the one or more bacteria.

33. A method of screening a faecal sample to determine its suitability for use in FMT therapy, the method comprising testing the faecal sample for the presence or absence of one or more bacteria, wherein when the one or more bacteria are detected, the faecal sample is deemed unsuitable for use in FMT therapy,

or wherein when the one or more bacteria are not detected, the faecal sample is deemed suitable for use in FMT therapy.

34. The method of claim 33 wherein the one or more bacteria are selected from the group consisting of Fusobacterium, Sutterella, Veillonella, Haemophilus, Streptococcus, Bacteroides, Akkermansia, Escherichia, Megamonas, Clostridium XI Va, Prevotella, Dialister, Veillonella, Bilophila, Parvimonas, Bacteroides uniformis and Bacteroides coprocola.

35. The method of claim 33 or claim 34 wherein the one or more bacteria is Fusobacterium spp.

36. The method of any one of claims 33 to 35 wherein the one or more bacteria is Fusobacterium gonidiaformans.

37. The method of any one of claims 33 to 36 wherein the one or more bacteria suppress a metabolic activity in the subject, wherein the metabolic activity is selected from the group consisting of heme biosynthesis, lipopolysaccharide biosynthesis, ubiquinone biosynthesis, terpenoid quinine biosynthesis, terpenoid backbone biosynthesis, lysine biosynthesis and oxidative phosphorylation.

38. The method of any one of claims 33 to 37 wherein the testing includes shotgun DNA sequencing, whole genome sequencing, PCR, 16S genomic DNA sequencing, 16S complementary DNA sequencing, 16S RNA sequencing or culturing the one or more bacteria.

39. A method of performing FMT therapy on a subject in need thereof, the method comprising:

screening one or more faecal samples by the method of any one of claims 27 to 38 to identify a faecal sample suitable for use in FMT therapy; and

administering the faecal sample to the subject.

40. The method of claim 39 wherein the subject in need thereof suffers from a gastrointestinal disorder.

41 . The method of claim 40 wherein the gastrointestinal disorder is ulcerative colitis.

42. A method of diagnosing a subject as being potentially responsive to FMT therapy, the method comprising detecting the presence of one or more bacteria in a sample obtained from the subject, wherein when the one or more bacteria are detected, the subject is diagnosed as being potentially responsive to FMT therapy.

43. The method of claim 42 wherein the one or more bacteria are selected from the group consisting of Eubacterium, Ruminococcus, Lachnospiraceae, Roseburia, Faecalibacterium, Eubacterium, Clostridium XVIII, Dorea, Coprococcus and Bacteroides fragilis.

44. The method of claim 42 wherein the one or more bacteria is a member of the Firmicutes phylum.

45. The method of any one of claims 42 to 44 wherein the one or more bacteria is Eubacterium hallii.

46. The method of any one of claims 42 to 45 wherein the one or more bacteria promote a metabolic activity in the subject, wherein the metabolic activity is selected from the group consisting of benzoate degradation, glycerophospholipid metabolism, secondary bile acid biosynthesis, ppGpp biosynthesis, pyruvate fermentation to acetate and lactate, fatty acid biosynthesis, propanoate metabolism, ansamycin biosynthesis and starch degradation.

47. The method of any one of claims 42 to 46 wherein a responsive patient displays clinical and/or endoscopic, response or remission.

48. The method of any one of claims 42 to 47 wherein the one or more bacteria are detected by shotgun DNA sequencing, whole genome sequencing, PCR, 16S genomic DNA sequencing, 16S complementary DNA sequencing, 16S RNA sequencing or culturing the one or more bacteria.

49. The method of any one of claims 42 to 48 wherein the sample is a faecal sample or an intestinal biopsy.

50. A method of diagnosing a subject as being unresponsive to FMT therapy, the method comprising detecting the presence of one or more bacteria in a sample obtained from the subject, wherein when the one or more bacteria are detected, the subject is diagnosed as being unresponsive to FMT therapy.

51 . The method of claim 50 wherein the one or more bacteria are selected from the group consisting of Fusobacterium, Sutterella, Veillonella, Haemophilus, Streptococcus, Bacteroides, Akkermansia, Escherichia, Megamonas, Clostridium XI Va, Prevotella, Dialister, Veillonella, Bilophila, Parvimonas, Bacteroides uniformis and Bacteroides coprocola.

52. The method of claim 50 or claim 51 wherein the one or more bacteria is Fusobacterium spp.

53. The method of any one of claims 50 to 52 wherein the one or more bacteria is Fusobacterium gonidiaformans.

54. The method of any one of claims 50 to 53 wherein the one or more bacteria suppress a metabolic activity in the subject, wherein the metabolic activity is selected from the group consisting of heme biosynthesis, lipopolysaccharide biosynthesis, ubiquinone biosynthesis, terpenoid quinine biosynthesis, terpenoid backbone biosynthesis, lysine biosynthesis and oxidative phosphorylation.

55. The method of any one of claims 50 to 54 wherein the one or more bacteria are detected by shotgun DNA sequencing, whole genome sequencing, PCR, 16S genomic DNA sequencing, 16S complementary DNA sequencing, 16S RNA sequencing or culturing the one or more bacteria.

56. The method of any one of claims 50 to 54 wherein the sample is a faecal sample or an intestinal biopsy.

57. A method of overcoming a lack of response to FMT in a subject, the method comprising administering a composition to the subject wherein the composition is enriched with one or more bacteria.

58. The method of claim 57 wherein the composition comprises a faecal sample.

59. The method of claim 57 or claim 58 wherein the one or more bacteria are selected from the group consisting of Eubacterium, Ruminococcus, Lachnospiraceae, Roseburia, Faecalibacterium, Eubacterium, Clostridium XVIII, Dorea, Coprococcus and Bacteroides fragilis.

60. The method of claim 57 or claim 58 wherein the one or more bacteria is a member of the Firmicutes phylum.

61 . The method of any one of claims 57 to 60 wherein the one or more bacteria is Eubacterium hallii.

62. The method of any one of claims 57 to 61 wherein the one or more bacteria promote a metabolic activity in the subject, wherein the metabolic activity is selected from the group consisting of benzoate degradation, glycerophospholipid metabolism, secondary bile acid biosynthesis, ppGpp biosynthesis, pyruvate fermentation to acetate and lactate, fatty acid biosynthesis, propanoate metabolism, ansamycin biosynthesis and starch degradation.

63. A method of overcoming a lack of response to FMT in a subject, the method comprising administering a faecal sample to the subject wherein the faecal sample is depleted in one or more bacteria.

64. The method of claim 63 wherein the one or more bacteria are selected from the group consisting of Fusobacterium, Sutterella, Veillonella, Haemophilus, Streptococcus, Bacteroides, Akkermansia, Escherichia, Megamonas, Clostridium XI Va, Prevotella, Dialister, Veillonella, Bilophila, Parvimonas, Bacteroides uniformis and Bacteroides coprocola.

65. The method of claim 63 or claim 64 wherein the one or more bacteria is Fusobacterium spp.

66. The method of any one of claims 63 to 65 wherein the one or more bacteria is Fusobacterium gonidiaformans.

67. The method of any one of claims 63 to 66 wherein the one or more bacteria suppress a metabolic activity in the subject, wherein the metabolic activity is selected from the group consisting of heme biosynthesis, lipopolysaccharide biosynthesis, ubiquinone biosynthesis, terpenoid quinine biosynthesis, terpenoid backbone biosynthesis, lysine biosynthesis and oxidative phosphorylation.

68. A method of treating ulcerative colitis in a subject in need thereof, the method comprising administering to the subject a therapeutically effective amount of one or more bacteria.

69. The method of claim 68 wherein the one or more bacteria are selected from the group consisting of Eubacterium, Ruminococcus, Lachnospiraceae, Roseburia, Faecalibacterium, Eubacterium, Clostridium XVIII, Dorea, Coprococcus and Bacteroides fragilis.

70. The method of claim 68 wherein the one or more bacteria is a member of the Firmicutes phylum.

71 . The method of any one of claims 68 to 70 wherein the one or more bacteria is Eubacterium hallii.

72. The method of any one of claims 68 to 71 wherein the one or more bacteria promote a metabolic activity in the subject, wherein the metabolic activity is selected from the group consisting of benzoate degradation, glycerophospholipid metabolism, secondary bile acid biosynthesis, ppGpp biosynthesis, pyruvate fermentation to acetate and lactate, fatty acid biosynthesis, propanoate metabolism, ansamycin biosynthesis and starch degradation.

73. The method of any one of claims 68 to 72 wherein the therapeutically effective amount of one or more bacteria is administered to the subject by FMT.

74. A method of treating ulcerative colitis in a subject in need thereof, the method comprising enhancing the growth of one or more bacteria in the subject.

75. The method of claim 74 wherein the one or more bacteria are selected from the group consisting of Eubacterium, Ruminococcus, Lachnospiraceae, Roseburia, Faecalibacterium, Eubacterium, Clostridium XVIII, Dorea, Coprococcus and Bacteroides fragilis.

76. The method of claim 74 wherein the one or more bacteria is a member of the Firmicutes phylum.

77. The method of any one of claims 74 to 76 wherein the one or more bacteria is Eubacterium hallii.

78. The method of any one of claims 74 to 77 wherein the one or more bacteria promote a metabolic activity in the subject, wherein the metabolic activity is selected from the group consisting of benzoate degradation, glycerophospholipid metabolism, secondary bile acid biosynthesis, ppGpp biosynthesis, pyruvate fermentation to acetate and lactate, fatty acid biosynthesis, propanoate metabolism, ansamycin biosynthesis and starch degradation.

79. A method of preparing an FMT sample, the method comprising:

culturing one or more bacteria thereby to produce a culture; and

adding the culture or a part thereof to a faecal sample thereby to produce the FMT sample.

80. The method of claim 79 wherein the one or more bacteria are selected from the group consisting of Eubacterium, Ruminococcus, Lachnospiraceae, Roseburia, Faecalibacterium, Eubacterium, Clostridium XVIII, Dorea, Coprococcus and Bacteroides fragilis.

81 . The method of claim 79 wherein the one or more bacteria is a member of the Firmicutes phylum.

82. The method of any one of claims 79 to 81 wherein the one or more bacteria is Eubacterium hallii.

83. The method of any one of claims 79 to 82 wherein the one or more bacteria promote a metabolic activity in the subject, wherein the metabolic activity is selected from the group consisting of benzoate degradation, glycerophospholipid metabolism, secondary bile acid biosynthesis, ppGpp biosynthesis, pyruvate fermentation to acetate and lactate, fatty acid biosynthesis, propanoate metabolism, ansamycin biosynthesis and starch degradation.

84. An FMT sample when prepared by the method of any one of claims 79 to 83.

85. A probiotic composition comprising a therapeutically effective amount of one or more bacteria.

86. The probiotic composition of claim 85 wherein the one or more bacteria are selected from the group consisting of Eubacterium, Ruminococcus, Lachnospiraceae, Roseburia, Faecalibacterium, Eubacterium, Clostridium XVIII, Dorea, Coprococcus and Bacteroides fragilis.

87. The probiotic compostion of claim 85 wherein the one or more bacteria is a member of the Firmicutes phylum.

88. The probiotic composition of any one of claims 85 to 87 wherein the one or more bacteria is Eubacterium hallii.

89. The probiotic composition of any one of claims 85 to 88 wherein the one or more bacteria promote a metabolic activity in the subject, wherein the metabolic activity is selected from the group consisting of benzoate degradation, glycerophospholipid metabolism, secondary bile acid biosynthesis, ppGpp biosynthesis, pyruvate fermentation to acetate and lactate, fatty acid biosynthesis, propanoate metabolism, ansamycin biosynthesis and starch degradation.

90. A method of overcoming a lack of responsiveness to FMT therapy in a subject in need thereof, the method comprising reducing the level of one or more bacteria in the subject.

91 . The method of claim 90 wherein the subject in need thereof suffers from a gastrointestinal disorder.

92. The method of claim 91 wherein the gastrointestinal disorder is ulcerative colitis.

93. The method of any one of claims 90 to 92 wherein the one or more bacteria are selected from the group consisting of Fusobacterium, Sutterella, Veillonella, Haemophilus, Streptococcus, Bacteroides, Akkermansia, Escherichia, Megamonas, Clostridium XI Va, Prevotella, Dialister, Veillonella, Bilophila, Parvimonas, Bacteroides uniformis and Bacteroides coprocola.

94. The method of any one of claims 90 to 93 wherein the one or more bacteria is Fusobacterium spp.

95. The method of any one of claims 90 to 94 wherein the one or more bacteria is Fusobacterium gonidiaformans.

96. The method of any one of claims 90 to 95 wherein the one or more bacteria suppress a metabolic activity in the subject, wherein the metabolic activity is selected from the group consisting of heme biosynthesis, lipopolysaccharide biosynthesis, ubiquinone biosynthesis, terpenoid quinine biosynthesis, terpenoid backbone biosynthesis, lysine biosynthesis and oxidative phosphorylation

97. A method of identifying an ulcerative colitis patient at risk of not responding to FMT therapy, the method comprising testing for the presence or absence of one or more bacteria selected from the group consisting of Fusobacterium, Sutterella, Veillonella, Haemophilus, Streptococcus, Bacteroides, Akkermansia, Escherichia, Megamonas, Clostridium XI Va, Prevotella, Dialister, Veillonella, Bilophila, Parvimonas, Bacteroides uniformis and Bacteroides coprocola in a sample obtained from the patient, wherein the presence of the one or more bacteria is indicative of the patient having an increased risk of not responding to FMT therapy.

98. The method of claim 97 wherein the one or more bacteria is Fusobacterium spp.

99. The method of claim 97 or claim 98 wherein the one or more bacteria is Fusobacterium gonidiaformans.

100. A method of treating ulcerative colitis in a subject in need thereof, the method comprising suppressing the growth of one or more bacteria in the subject.

101. The method of claim 100 wherein the one or more bacteria are selected from the group consisting of Fusobacterium, Sutterella, Veillonella, Haemophilus, Streptococcus, Bacteroides, Akkermansia, Escherichia, Megamonas, Clostridium XI Va, Prevotella, Dialister, Veillonella, Bilophila, Parvimonas, Bacteroides uniformis and Bacteroides coprocola.

102. The method of claim 100 or claim 101 wherein the one or more bacteria is Fusobacterium spp.

103. The method of any one of claims 100 to 102 wherein the one or more bacteria is Fusobacterium gonidiaformans.

104. The method of any one of claims 100 to 103 wherein the one or more bacteria suppress a metabolic activity in the subject, wherein the metabolic activity is selected from the group consisting of heme biosynthesis, lipopolysaccharide biosynthesis, ubiquinone biosynthesis, terpenoid quinine biosynthesis, terpenoid backbone biosynthesis, lysine biosynthesis and oxidative phosphorylation.

Description:
COMPOSITIONS AND METHODS FOR PROMOTING GUT HEALTH Field of the invention

[0001 ] The present invention relates to compositions and methods for the treatment of gastrointestinal disorders such as ulcerative colitis by suppressing and/or promoting the growth of certain bacteria or the activity of certain metabolic pathways. In some embodiments, the invention relates to methods and compositions for use in faecal microbiota transplantation (FMT).

Background of the invention

[0002] Any discussion of the prior art throughout the specification should in no way be considered as an admission that such prior art is widely known or forms part of the common general knowledge in the field.

[0003] There is interest in the potential role of therapeutic microbial manipulation using faecal microbiota transplantation (FMT) in the treatment of a variety of gastrointestinal conditions, such as ulcerative colitis (UC) [Borody Current opinion in gastroenterology, Kelly Gastroenterology ]. Most currently available conventional treatments for UC target the immune system through a range of mediators of importance in the inflammatory cascade [Cohen, BMJ\. Flowever, many of those treatments do not provide adequate disease control or are associated with harmful side effects.

[0004] A number of studies including randomised controlled trials [Moayyedi Gastroenterology, Paramsothy Lancet, Costello ECCO / DDW\ and systematic reviews / meta-analyses [Paramsothy JCC, Costello APT, Allegretti IBD ] suggest that FMT may be useful in the treatment of active UC patients. One recent study suggested that multi donor FMT therapy for active UC may be superior to placebo therapy, with 27% of patients achieving the primary endpoint of clinical remission with endoscopic remission or response as opposed to 7.5% with placebo, with a clinical response rate of over 50% [Paramsothy Lancef\. Flowever, the underlying microbial basis, predictors of therapeutic outcome and the ultimate active bacteria and ingredient(s) of FMT mediating benefit in UC remain largely unknown [Moayeddi Gastroenterology, Rossen Gastroenterology, Fuentes ISME\. It would be useful to identify bacteria that play a role in the pathogenesis or alleviation of gastrointestinal disorders such as ulcerative colitis. It would also be useful to identify bacteria that play a role in causing or perpetuating ongoing inflammation, and bacteria that may ameliorate, diminish or cure inflammation. The identification of such bacteria may aid in modifying the constitution of faecal microbiota transplantation, or assist in the development of treatments utilising, or derived from, such bacteria.

[0005] In this context, there is a need for compositions and methods for use in FMT therapy to treat gastrointestinal disorders.

[0006] It is an object of the present invention to overcome or ameliorate at least one of the disadvantages of the prior art or to provide a useful alternative.

Summary of the invention

[0007] In a first aspect, the present invention provides a method of performing FMT therapy on a subject in need thereof, the method comprising the step of administering a faecal sample to the subject, wherein the faecal sample is enriched with one or more bacteria.

[0008] In one embodiment of the first aspect, the one or more bacteria are selected from the group consisting of Eubacterium, Ruminococcus, Lachnospiraceae, Roseburia, Faecalibacterium, Eubacterium, Clostridium XVIII, Dorea, Coprococcus and Bacteroides fragilis. The one or more bacteria may be a member of the Firmicutes phylum. Preferably, the one or more bacteria is Eubacterium hallii.

[0009] In one embodiment of the first aspect, the one or more bacteria promote a metabolic activity in the subject, wherein the metabolic activity is selected from the group consisting of benzoate degradation, glycerophospholipid metabolism, secondary bile acid biosynthesis, ppGpp biosynthesis, pyruvate fermentation to acetate and lactate, fatty acid biosynthesis, propanoate metabolism, ansamycin biosynthesis and starch degradation.

[0010] In an embodiment of the first aspect, there is provided a method of performing FMT therapy on a subject in need thereof, the method comprising the step of administering a faecal sample to the subject, wherein the faecal sample is enriched with Eubacterium hallii.

[001 1 ] In a further embodiment of the first aspect, the subject in need thereof suffers from a gastrointestinal disorder. The gastrointestinal disorder may be ulcerative colitis. [0012] In a second aspect, the present invention provides a method of performing FMT therapy on a subject in need thereof, the method comprising the step of administering a faecal sample to the subject, wherein the faecal sample is depleted in one or more bacteria.

[0013] In one embodiment of the second aspect, the one or more bacteria are selected from the group consisting of Fusobacterium, Sutterella, Veillonella, Haemophilus, Streptococcus, Bacteroides, Akkermansia, Escherichia, Megamonas, Clostridium XlVa, Prevotella, Dialister, Veillonella, Bilophila, Parvimonas, Bacteroides uniformis and Bacteroides coprocola. The one or more bacteria may be Fusobacterium spp. Preferably, the one or more bacteria is Fusobacterium gonidiaformans.

[0014] In one embodiment of the second aspect, the one or more bacteria suppress a metabolic activity in the subject, wherein the metabolic activity is selected from the group consisting of heme biosynthesis, lipopolysaccharide biosynthesis, ubiquinone biosynthesis, terpenoid quinine biosynthesis, terpenoid backbone biosynthesis, lysine biosynthesis and oxidative phosphorylation.

[0015] In an embodiment of the second aspect, there is provided a method of performing FMT therapy on a subject in need thereof, the method comprising the step of administering a faecal sample to the subject, wherein the faecal sample is depleted in Fusobacterium spp.

[0016] In a further embodiment of the second aspect, the subject in need thereof suffers from a gastrointestinal disorder. The gastrointestinal disorder may be ulcerative colitis

[0017] In a third aspect, the present invention provides a faecal sample for use in FMT therapy, wherein the faecal sample is enriched with one or more bacteria.

[0018] In one embodiment of the third aspect, the one or more bacteria are selected from the group consisting of Eubacterium, Ruminococcus, Lachnospiraceae, Roseburia, Faecalibacterium, Eubacterium, Clostridium XVIII, Dorea, Coprococcus and Bacteroides fragilis. The one or more bacteria may be a member of the Firmicutes phylum. Preferably, the one or more bacteria is Eubacterium hallii.

In one embodiment of the third aspect, the one or more bacteria promote a metabolic activity in the subject, wherein the metabolic activity is selected from the group consisting of benzoate degradation, glycerophospholipid metabolism, secondary bile acid biosynthesis, ppGpp biosynthesis, pyruvate fermentation to acetate and lactate, fatty acid biosynthesis, propanoate metabolism, ansamycin biosynthesis and starch degradation.

[0019] In a fourth aspect, the present invention provides a faecal sample for use in FMT therapy, wherein the faecal sample is depleted in one or more bacteria.

[0020] In one embodiment of the fourth aspect, the one or more bacteria are selected from the group consisting of Fusobacterium, Sutterella, Veillonella, Haemophilus, Streptococcus, Bacteroides, Akkermansia, Escherichia, Megamonas, Clostridium XlVa, Prevotella, Dialister, Veillonella, Bilophila, Parvimonas, Bacteroides uniformis and Bacteroides coprocola. The one or more bacteria may be Fusobacterium spp. Preferably, the one or more bacteria is Fusobacterium gonidiaformans.

[0021 ] In one embodiment of the fourth aspect, the one or more bacteria suppress a metabolic activity in the subject, wherein the metabolic activity is selected from the group consisting of heme biosynthesis, lipopolysaccharide biosynthesis, ubiquinone biosynthesis, terpenoid quinine biosynthesis, terpenoid backbone biosynthesis, lysine biosynthesis and oxidative phosphorylation.

[0022] In a fifth aspect, the present invention provides a method of screening a faecal sample to determine its suitability for use in FMT therapy, the method comprising testing the faecal sample for the presence or absence of one or more bacteria, wherein when the one or more bacteria are detected, the faecal sample is deemed suitable for use in FMT therapy, or wherein when the one or more bacteria are not detected, the faecal sample is deemed unsuitable for use in FMT therapy.

[0023] In one embodiment of the fifth aspect, the one or more bacteria are selected from the group consisting of Eubacterium, Ruminococcus, Lachnospiraceae, Roseburia, Faecalibacterium, Eubacterium, Clostridium XVIII, Dorea, Coprococcus and Bacteroides fragilis. The one or more bacteria may be a member of the Firmicutes phylum. Preferably, the one or more bacteria is Eubacterium hallii. [0024] In one embodiment of the fifth aspect, the one or more bacteria promote a metabolic activity in the subject, wherein the metabolic activity is selected from the group consisting of benzoate degradation, glycerophospholipid metabolism, secondary bile acid biosynthesis, ppGpp biosynthesis, pyruvate fermentation to acetate and lactate, fatty acid biosynthesis, propanoate metabolism, ansamycin biosynthesis and starch degradation.

[0025] In one embodiment of the fifth aspect, the testing includes shotgun DNA sequencing, whole genome sequencing, PCR, 16S genomic DNA sequencing, 16S complementary DNA sequencing, 16S RNA sequencing or culturing the one or more bacteria.

[0026] In a sixth aspect, the present invention provides a method of screening a faecal sample to determine its suitability for use in FMT therapy, the method comprising testing the faecal sample for the presence or absence of one or more bacteria, wherein when the one or more bacteria are detected, the faecal sample is deemed unsuitable for use in FMT therapy, or wherein when the one or more bacteria are not detected, the faecal sample is deemed suitable for use in FMT therapy.

[0027] In one embodiment of the sixth aspect, the one or more bacteria are selected from the group consisting of Fusobacterium, Sutterella, Veillonella, Haemophilus, Streptococcus, Bacteroides, Akkermansia, Escherichia, Megamonas, Clostridium XlVa, Prevotella, Dialister, Veillonella, Bilophila, Parvimonas, Bacteroides uniformis and Bacteroides coprocola. The one or more bacteria may be Fusobacterium spp. Preferably, the one or more bacteria is Fusobacterium gonidiaformans.

[0028] In one embodiment of the sixth aspect, the one or more bacteria suppress a metabolic activity in the subject, wherein the metabolic activity is selected from the group consisting of heme biosynthesis, lipopolysaccharide biosynthesis, ubiquinone biosynthesis, terpenoid quinine biosynthesis, terpenoid backbone biosynthesis, lysine biosynthesis and oxidative phosphorylation.

[0029] In one embodiment of the sixth aspect, the testing includes shotgun DNA sequencing, whole genome sequencing, PCR, 16S genomic DNA sequencing, 16S complementary DNA sequencing, 16S RNA sequencing or culturing the one or more bacteria.

[0030] In a seventh aspect, the present invention provides a method of performing FMT therapy on a subject in need thereof, the method comprising: screening one or more faecal samples by the method of the fifth aspect or the sixth aspect to identify a faecal sample suitable for use in FMT therapy; and administering the faecal sample to the subject.

[0031 ] In one embodiment of the seventh aspect, the subject in need thereof suffers from a gastrointestinal disorder. The gastrointestinal disorder may be ulcerative colitis.

[0032] In an eighth aspect, the present invention provides a method of diagnosing a subject as being potentially responsive to FMT therapy, the method comprising detecting the presence of one or more bacteria in a sample obtained from the subject, wherein when the one or more bacteria are detected, the subject is diagnosed as being potentially responsive to FMT therapy.

[0033] In one embodiment of the eighth aspect, the one or more bacteria are selected from the group consisting of Eubacterium, Ruminococcus, Lachnospiraceae, Roseburia, Faecalibacterium, Eubacterium, Clostridium XVIII, Dorea, Coprococcus and Bacteroides fragilis. The one or more bacteria may be a member of the Firmicutes phylum. Preferably, the one or more bacteria is Eubacterium hallii.

[0034] In one embodiment of the eighth aspect, the one or more bacteria promote a metabolic activity in the subject, wherein the metabolic activity is selected from the group consisting of benzoate degradation, glycerophospholipid metabolism, secondary bile acid biosynthesis, ppGpp biosynthesis, pyruvate fermentation to acetate and lactate, fatty acid biosynthesis, propanoate metabolism, ansamycin biosynthesis and starch degradation.

[0035] In one embodiment of the eighth aspect, a responsive patient may display clinical and/or endoscopic, response or remission.

[0036] In one embodiment of the eighth aspect, the one or more bacteria are detected by shotgun DNA sequencing, whole genome sequencing, PCR, 16S genomic DNA sequencing, 16S complementary DNA sequencing, 16S RNA sequencing or culturing the one or more bacteria.

[0037] In one embodiment of the eighth aspect, the sample is a faecal sample or an intestinal biopsy.

[0038] In a ninth aspect, the present invention provides a method of diagnosing a subject as being unresponsive to FMT therapy, the method comprising detecting the presence of one or more bacteria in a sample obtained from the subject, wherein when the one or more bacteria are detected, the subject is diagnosed as being unresponsive to FMT therapy.

[0039] In one embodiment of the ninth aspect, the one or more bacteria are selected from the group consisting of Fusobacterium, Sutterella, Veillonella, Haemophilus, Streptococcus, Bacteroides, Akkermansia, Escherichia, Megamonas, Clostridium XlVa, Prevotella, Dialister, Veillonella, Bilophila, Parvimonas, Bacteroides uniformis and Bacteroides coprocola. The one or more bacteria may be Fusobacterium spp. Preferably, the one or more bacteria is Fusobacterium gonidiaformans.

[0040] In one embodiment of the ninth aspect, the one or more bacteria suppress a metabolic activity in the subject, wherein the metabolic activity is selected from the group consisting of heme biosynthesis, lipopolysaccharide biosynthesis, ubiquinone biosynthesis, terpenoid quinine biosynthesis, terpenoid backbone biosynthesis, lysine biosynthesis and oxidative phosphorylation.

[0041 ] In one embodiment of the ninth aspect, the one or more bacteria are detected by shotgun DNA sequencing, whole genome sequencing, PCR, 16S genomic DNA sequencing, 16S complementary DNA sequencing, 16S RNA sequencing or culturing the one or more bacteria.

[0042] In one embodiment of the ninth aspect, the sample is a faecal sample or an intestinal biopsy.

[0043] In a tenth aspect, the present invention provides a method of overcoming a lack of response to FMT in a subject, the method comprising administering a composition to the subject wherein the composition is enriched with one or more bacteria. [0044] In one embodiment of the tenth aspect, the composition comprises a faecal sample.

[0045] In one embodiment of the tenth aspect, the one or more bacteria are selected from the group consisting of Eubacterium, Ruminococcus, Lachnospiraceae, Roseburia, Faecalibacterium, Eubacterium, Clostridium XVIII, Dorea, Coprococcus and Bacteroides fragilis. The one or more bacteria may be a member of the Firmicutes phylum. Preferably, the one or more bacteria is Eubacterium hallii.

[0046] In one embodiment of the tenth aspect, the one or more bacteria promote a metabolic activity in the subject, wherein the metabolic activity is selected from the group consisting of benzoate degradation, glycerophospholipid metabolism, secondary bile acid biosynthesis, ppGpp biosynthesis, pyruvate fermentation to acetate and lactate, fatty acid biosynthesis, propanoate metabolism, ansamycin biosynthesis and starch degradation.

[0047] In an eleventh aspect, the present invention provides a method of overcoming a lack of response to FMT in a subject, the method comprising administering a faecal sample to the subject wherein the faecal sample is depleted in one or more bacteria.

[0048] In one embodiment of the eleventh aspect, the one or more bacteria are selected from the group consisting of Fusobacterium, Sutterella, Veillonella, Haemophilus, Streptococcus, Bacteroides, Akkermansia, Escherichia, Megamonas, Clostridium XlVa, Prevotella, Dialister, Veillonella, Bilophila, Parvimonas, Bacteroides uniformis and Bacteroides coprocola. The one or more bacteria may be Fusobacterium spp. Preferably, the one or more bacteria is Fusobacterium gonidiaformans.

[0049] In one embodiment of the eleventh aspect, the one or more bacteria suppress a metabolic activity in the subject, wherein the metabolic activity is selected from the group consisting of heme biosynthesis, lipopolysaccharide biosynthesis, ubiquinone biosynthesis, terpenoid quinine biosynthesis, terpenoid backbone biosynthesis, lysine biosynthesis and oxidative phosphorylation.

[0050] In a twelfth aspect, the present invention provides a method of treating ulcerative colitis in a subject in need thereof, the method comprising administering to the subject a therapeutically effective amount of one or more bacteria or bacterial derivatives. [0051 ] In one embodiment of twelfth aspect, the one or more bacteria are selected from the group consisting of Eubacterium, Ruminococcus, Lachnospiraceae, Roseburia, Faecalibacterium, Eubacterium, Clostridium XVIII, Dorea, Coprococcus and Bacteroides fragilis. The one or more bacteria may be a member of the Firmicutes phylum. Preferably, the one or more bacteria is Eubacterium hallii.

[0052] In one embodiment of twelfth aspect, the one or more bacteria promote a metabolic activity in the subject, wherein the metabolic activity is selected from the group consisting of benzoate degradation, glycerophospholipid metabolism, secondary bile acid biosynthesis, ppGpp biosynthesis, pyruvate fermentation to acetate and lactate, fatty acid biosynthesis, propanoate metabolism, ansamycin biosynthesis and starch degradation.

[0053] In one embodiment of twelfth aspect, the therapeutically effective amount of one or more bacteria is administered to the subject by FMT.

[0054] In a thirteenth aspect, the present invention provides a method of treating ulcerative colitis in a subject in need thereof, the method comprising enhancing the growth of one or more bacteria in the subject.

[0055] In one embodiment of the thirteenth aspect, the one or more bacteria are selected from the group consisting of Eubacterium, Ruminococcus, Lachnospiraceae, Roseburia, Faecalibacterium, Eubacterium, Clostridium XVIII, Dorea, Coprococcus and Bacteroides fragilis. The one or more bacteria may be a member of the Firmicutes phylum. Preferably, the one or more bacteria is Eubacterium hallii.

[0056] In one embodiment of the thirteenth aspect, the one or more bacteria promote a metabolic activity in the subject, wherein the metabolic activity is selected from the group consisting of benzoate degradation, glycerophospholipid metabolism, secondary bile acid biosynthesis, ppGpp biosynthesis, pyruvate fermentation to acetate and lactate, fatty acid biosynthesis, propanoate metabolism, ansamycin biosynthesis and starch degradation.

[0057] In a fourteenth aspect, the present invention provides a method of preparing an FMT sample, the method comprising: culturing one or more bacteria thereby to produce a culture; and adding the culture or a part thereof to a faecal sample thereby to produce the FMT sample.

[0058] In one embodiment of the fourteenth aspect, the one or more bacteria are selected from the group consisting of Eubacterium, Ruminococcus, Lachnospiraceae, Roseburia, Faecalibacterium, Eubacterium, Clostridium XVIII, Dorea, Coprococcus and Bacteroides fragilis. The one or more bacteria may be a member of the Firmicutes phylum. Preferably, the one or more bacteria is Eubacterium hallii.

[0059] In one embodiment of the fourteenth aspect, the one or more bacteria promote a metabolic activity in the subject, wherein the metabolic activity is selected from the group consisting of benzoate degradation, glycerophospholipid metabolism, secondary bile acid biosynthesis, ppGpp biosynthesis, pyruvate fermentation to acetate and lactate, fatty acid biosynthesis, propanoate metabolism, ansamycin biosynthesis and starch degradation.

[0060] In a fifteenth aspect, the present invention provides an FMT sample when prepared by the method of the fourteenth aspect.

[0061 ] In a sixteenth aspect, the present invention provides a probiotic composition comprising a therapeutically effective amount of one or more bacteria.

[0062] In one embodiment of the sixteenth aspect, the one or more bacteria are selected from the group consisting of Eubacterium, Ruminococcus, Lachnospiraceae, Roseburia, Faecalibacterium, Eubacterium, Clostridium XVIII, Dorea, Coprococcus and Bacteroides fragilis. The one or more bacteria may be a member of the Firmicutes phylum. Preferably, the one or more bacteria is Eubacterium hallii.

[0063] In one embodiment of the sixteenth aspect, the one or more bacteria promote a metabolic activity in the subject, wherein the metabolic activity is selected from the group consisting of benzoate degradation, glycerophospholipid metabolism, secondary bile acid biosynthesis, ppGpp biosynthesis, pyruvate fermentation to acetate and lactate, fatty acid biosynthesis, propanoate metabolism, ansamycin biosynthesis and starch degradation. [0064] In a seventeenth aspect, the present invention provides a method of overcoming a lack of responsiveness to FMT therapy in a subject in need thereof, the method comprising reducing the level of one or more bacteria in the subject.

[0065] In one embodiment of the seventeenth aspect, the subject in need thereof suffers from a gastrointestinal disorder. The gastrointestinal disorder may be ulcerative colitis.

[0066] In one embodiment of the seventeenth aspect, the one or more bacteria are selected from the group consisting of Fusobacterium, Sutterella, Veillonella, Haemophilus, Streptococcus, Bacteroides, Akkermansia, Escherichia, Megamonas, Clostridium XI Va, Prevotella, Dialister, Veillonella, Bilophila, Parvimonas, Bacteroides uniformis and Bacteroides coprocola. The one or more bacteria may be Fusobacterium spp. Preferably, the one or more bacteria is Fusobacterium gonidiaformans.

[0067] In one embodiment of the seventeenth aspect, the one or more bacteria suppress a metabolic activity in the subject, wherein the metabolic activity is selected from the group consisting of heme biosynthesis, lipopolysaccharide biosynthesis, ubiquinone biosynthesis, terpenoid quinine biosynthesis, terpenoid backbone biosynthesis, lysine biosynthesis and oxidative phosphorylation.

[0068] In a eighteenth aspect, the present invention provides a method of identifying an ulcerative colitis patient at risk of not responding to FMT therapy, the method comprising testing for the presence or absence of one or more bacteria selected from the group consisting of Fusobacterium, Sutterella, Veillonella, Haemophilus, Streptococcus, Bacteroides, Akkermansia, Escherichia, Megamonas, Clostridium XI Va, Prevotella, Dialister, Veillonella, Bilophila, Parvimonas, Bacteroides uniformis and Bacteroides coprocola in a sample obtained from the patient, wherein the presence of the one or more bacteria is indicative of the patient having an increased risk of not responding to FMT therapy.

[0069] The one or more bacteria of the eighteenth may be Fusobacterium spp. Preferably, the one or more bacteria is Fusobacterium gonidiaformans.

[0070] In a nineteenth aspect, the present invention provides a method of treating ulcerative colitis in a subject in need thereof, the method comprising suppressing the growth of one or more bacteria in the subject. [0071 ] In one embodiment of the nineteenth aspect, the one or more bacteria are selected from the group consisting of Fusobacterium, Sutterella, Veillonella, Haemophilus, Streptococcus, Bacteroides, Akkermansia, Escherichia, Megamonas, Clostridium XI Va, Prevotella, Dialister, Veillonella, Bilophila, Parvimonas, Bacteroides uniformis and Bacteroides coprocola. The one or more bacteria may be Fusobacterium spp. Preferably, the one or more bacteria is Fusobacterium gonidiaformans.

[0072] In one embodiment of the nineteenth aspect, the one or more bacteria suppress a metabolic activity in the subject, wherein the metabolic activity is selected from the group consisting of heme biosynthesis, lipopolysaccharide biosynthesis, ubiquinone biosynthesis, terpenoid quinine biosynthesis, terpenoid backbone biosynthesis, lysine biosynthesis and oxidative phosphorylation.

Detailed description of the invention

[0073] Bacterial flora of the gut and bowel is increasingly being viewed as an important virtual organ. Faecal microbiota transplantation (also known as faecal bacteriotherapy or stool transplant) is one means by which bacterial floral of the gut and bowel can be modified, for example, to treat or prevent a particular disease or disorder that is regulated by this virtual organ. FMT typically involves implantation or administration of gut microbiota such as colonic bacteria into the gut or bowel of a subject. FMT therapy may be performed, for example, by way of infusions through a colonoscope, encapsulated formulation, oral administration, enema, nasogastric tube, sigmoidoscope, orogastric tube or nasojejunal tube.

[0074] In a first aspect of the present invention, there is provided a method of performing FMT therapy on a subject in need thereof, the method comprising the step of administering a faecal sample to the subject, wherein the faecal sample is enriched with one or more bacteria.

[0075] In one embodiment of the first aspect, the one or more bacteria are selected from the group consisting of Eubacterium, Ruminococcus, Lachnospiraceae, Roseburia, Faecalibacterium, Eubacterium, Clostridium XVIII, Dorea, Coprococcus and Bacteroides fragilis. The one or more bacteria may be a member of the Firmicutes phylum. Preferably, the one or more bacteria is Eubacterium hallii. [0076] In one embodiment of the first aspect, the one or more bacteria promote a metabolic activity in the subject, wherein the metabolic activity is selected from the group consisting of benzoate degradation, glycerophospholipid metabolism, secondary bile acid biosynthesis, ppGpp biosynthesis, pyruvate fermentation to acetate and lactate, fatty acid biosynthesis, propanoate metabolism, ansamycin biosynthesis and starch degradation.

[0077] In an embodiment of the first aspect, there is provided a method of performing FMT therapy on a subject in need thereof, the method comprising the step of administering a faecal sample to the subject, wherein the faecal sample is enriched with Eubacterium hallii.

[0078] In a further embodiment of the first aspect, the subject in need thereof suffers from a gastrointestinal disorder. The gastrointestinal disorder may be ulcerative colitis. However, those skilled in the art will be aware of other gastrointestinal disorders that may be treated by the methods and compositions of the present invention, for example, spastic colon, mucous colitis, collagenous colitis, Crohn's disease, Johne' s disease (paratuberculosis), bloating, gastrointestinal dysbiosis, diarrhea, microscopic colitis, idiopathic inflammatory bowel disease, antibiotic-associated colitis, idiopathic or simple constipation, diverticular disease, Acquired Immune Deficiency Syndrome (AIDS) enteropathy and autistic spectrum disorder (ASD). The methods and compositions of the present invention may also be used to increase bacterial diversity in a subject's gastrointestinal tract and/or to maintain a healthy gut and reduce the likelihood of developing a gastrointestinal disorder.

[0079] In a second aspect, the present invention provides a method of performing FMT therapy on a subject in need thereof, the method comprising the step of administering a faecal sample to the subject, wherein the faecal sample is depleted in one or more bacteria.

[0080] In one embodiment of the second aspect, the one or more bacteria are selected from the group consisting of Fusobacterium, Sutterella, Veillonella, Haemophilus, Streptococcus, Bacteroides, Akkermansia, Escherichia, Megamonas, Clostridium XlVa, Prevotella, Dialister, Veillonella, Bilophila, Parvimonas, Bacteroides uniformis and Bacteroides coprocola. The one or more bacteria may be Fusobacterium spp. Preferably, the one or more bacteria is Fusobacterium gonidiaformans. [0081 ] In one embodiment of the second aspect, the one or more bacteria suppress a metabolic activity in the subject, wherein the metabolic activity is selected from the group consisting of heme biosynthesis, lipopolysaccharide biosynthesis, ubiquinone biosynthesis, terpenoid quinine biosynthesis, terpenoid backbone biosynthesis, lysine biosynthesis and oxidative phosphorylation.

[0082] In an embodiment of the second aspect, there is provided a method of performing FMT therapy on a subject in need thereof, the method comprising the step of administering a faecal sample to the subject, wherein the faecal sample is depleted in Fusobacterium spp.

[0083] In a further embodiment of the second aspect, the subject in need thereof suffers from a gastrointestinal disorder. The gastrointestinal disorder may be ulcerative colitis

[0084] In a third aspect, the present invention provides a faecal sample for use in FMT therapy, wherein the faecal sample is enriched with one or more bacteria.

[0085] A sample may be considered "enriched" with one or more bacteria when positive steps have been taken to increase the concentration of the one or more bacteria in the sample. The positive steps may include, for example, culturing the one or more bacteria and using the culture to enrich a composition such as a faecal sample. The positive steps may also include testing multiple samples, such as multiple faecal samples, and selecting (and optionally combining) those samples which have the highest concentration of the one or more bacteria.

[0086] In one embodiment of the third aspect, the one or more bacteria are selected from the group consisting of Eubacterium, Ruminococcus, Lachnospiraceae, Roseburia, Faecalibacterium, Eubacterium, Clostridium XVIII, Dorea, Coprococcus and Bacteroides fragilis. The one or more bacteria may be a member of the Firmicutes phylum. Preferably, the one or more bacteria is Eubacterium hallii.

In one embodiment of the third aspect, the one or more bacteria promote a metabolic activity in the subject, wherein the metabolic activity is selected from the group consisting of benzoate degradation, glycerophospholipid metabolism, secondary bile acid biosynthesis, ppGpp biosynthesis, pyruvate fermentation to acetate and lactate, fatty acid biosynthesis, propanoate metabolism, ansamycin biosynthesis and starch degradation. [0087] In a fourth aspect, the present invention provides a faecal sample for use in FMT therapy, wherein the faecal sample is depleted in one or more bacteria.

[0088] A sample may be considered "depleted" in one or more bacteria when positive steps have been taken to reduce the concentration of the one or more bacteria in the sample. The positive steps may include, for example, selectively reducing the concentration or growth of the one or more bacteria in a composition such as a faecal sample. The positive steps may also include testing multiple samples, such as multiple faecal samples, and selecting (and optionally combining) those samples which have the lowest concentration of the one or more bacteria.

[0089] Those skilled in the art will be aware of several techniques that may be used to reduce the level of a specific bacterium or one or more bacteria in a sample or a subject. For example, the skilled person can readily determine suitable antimicrobial or antibiotic compositions active against certain types of bacteria. Alternatively, an antibody or antigen-binding protein may target and optionally neutralise one or more target bacteria. As a further example, one or more bacteria may be targeted using a virus or using genetic techniques such as gene silencing (or RNA silencing or RNA interference) or other nuclease-based techniques (eg, CRISPR).

[0090] In one embodiment of the fourth aspect, the one or more bacteria are selected from the group consisting of Fusobacterium, Sutterella, Veillonella, Haemophilus, Streptococcus, Bacteroides, Akkermansia, Escherichia, Megamonas, Clostridium XlVa, Prevotella, Dialister, Veillonella, Bilophila, Parvimonas, Bacteroides uniformis and Bacteroides coprocola. The one or more bacteria may be Fusobacterium spp. Preferably, the one or more bacteria is Fusobacterium gonidiaformans.

[0091 ] In one embodiment of the fourth aspect, the one or more bacteria suppress a metabolic activity in the subject, wherein the metabolic activity is selected from the group consisting of heme biosynthesis, lipopolysaccharide biosynthesis, ubiquinone biosynthesis, terpenoid quinine biosynthesis, terpenoid backbone biosynthesis, lysine biosynthesis and oxidative phosphorylation.

[0092] In a fifth aspect, the present invention provides a method of screening a faecal sample to determine its suitability for use in FMT therapy, the method comprising testing the faecal sample for the presence or absence of one or more bacteria, wherein when the one or more bacteria are detected, the faecal sample is deemed suitable for use in FMT therapy, or wherein when the one or more bacteria are not detected, the faecal sample is deemed unsuitable for use in FMT therapy.

[0093] Those skilled in the art will understand that one or more bacteria may be identified using nucleic acid sequencing methods known in the art, as well as other suitable techniques such as, for example, diagnostic PCR, antibody-based techniques, nucleic acid hybridisation-based techniques, biochemical assays and selective culturing.

[0094] In one embodiment of the fifth aspect, the one or more bacteria are selected from the group consisting of Eubacterium, Ruminococcus, Lachnospiraceae, Roseburia, Faecalibacterium, Eubacterium, Clostridium XV III, Dorea, Coprococcus and Bacteroides fragilis. The one or more bacteria may be a member of the Firmicutes phylum. Preferably, the one or more bacteria is Eubacterium hallii.

[0095] In one embodiment of the fifth aspect, the one or more bacteria promote a metabolic activity in the subject, wherein the metabolic activity is selected from the group consisting of benzoate degradation, glycerophospholipid metabolism, secondary bile acid biosynthesis, ppGpp biosynthesis, pyruvate fermentation to acetate and lactate, fatty acid biosynthesis, propanoate metabolism, ansamycin biosynthesis and starch degradation.

[0096] In one embodiment of the fifth aspect, the testing includes shotgun DNA sequencing, whole genome sequencing, PCR, 16S genomic DNA sequencing, 16S complementary DNA sequencing, 16S RNA sequencing or culturing the one or more bacteria.

[0097] In a sixth aspect, the present invention provides a method of screening a faecal sample to determine its suitability for use in FMT therapy, the method comprising testing the faecal sample for the presence or absence of one or more bacteria, wherein when the one or more bacteria are detected, the faecal sample is deemed unsuitable for use in FMT therapy, or wherein when the one or more bacteria are not detected, the faecal sample is deemed suitable for use in FMT therapy. [0098] In one embodiment of the sixth aspect, the one or more bacteria are selected from the group consisting of Fusobacterium, Sutterella, Veillonella, Haemophilus, Streptococcus, Bacteroides, Akkermansia, Escherichia, Megamonas, Clostridium XlVa, Prevotella, Dialister, Veillonella, Bilophila, Parvimonas, Bacteroides uniformis and Bacteroides coprocola. The one or more bacteria may be Fusobacterium spp. Preferably, the one or more bacteria is Fusobacterium gonidiaformans.

[0099] In one embodiment of the sixth aspect, the one or more bacteria suppress a metabolic activity in the subject, wherein the metabolic activity is selected from the group consisting of heme biosynthesis, lipopolysaccharide biosynthesis, ubiquinone biosynthesis, terpenoid quinine biosynthesis, terpenoid backbone biosynthesis, lysine biosynthesis and oxidative phosphorylation.

[00100] In one embodiment of the sixth aspect, the testing includes shotgun DNA sequencing, whole genome sequencing, PCR, 16S genomic DNA sequencing, 16S complementary DNA sequencing, 16S RNA sequencing or culturing the one or more bacteria.

[00101] In a seventh aspect, the present invention provides a method of performing FMT therapy on a subject in need thereof, the method comprising: screening one or more faecal samples by the method of the fifth aspect or the sixth aspect to identify a faecal sample suitable for use in FMT therapy; and administering the faecal sample to the subject.

[00102] In one embodiment of the seventh aspect, the subject in need thereof suffers from a gastrointestinal disorder. The gastrointestinal disorder may be ulcerative colitis.

[00103] In an eighth aspect, the present invention provides a method of diagnosing a subject as being potentially responsive to FMT therapy, the method comprising detecting the presence of one or more bacteria in a sample obtained from the subject, wherein when the one or more bacteria are detected, the subject is diagnosed as being potentially responsive to FMT therapy.

[00104] In one embodiment of the eighth aspect, the one or more bacteria are selected from the group consisting of Eubacterium, Ruminococcus, Lachnospiraceae, Roseburia, Faecalibacterium, Eubacterium, Clostridium XVIII, Dorea, Coprococcus and Bacteroides fragilis. The one or more bacteria may be a member of the Firmicutes phylum. Preferably, the one or more bacteria is Eubacterium hallii.

[00105] In one embodiment of the eighth aspect, the one or more bacteria promote a metabolic activity in the subject, wherein the metabolic activity is selected from the group consisting of benzoate degradation, glycerophospholipid metabolism, secondary bile acid biosynthesis, ppGpp biosynthesis, pyruvate fermentation to acetate and lactate, fatty acid biosynthesis, propanoate metabolism, ansamycin biosynthesis and starch degradation.

[00106] In one embodiment of the eighth aspect, a responsive patient may display clinical and/or endoscopic, response or remission.

[00107] In one embodiment of the eighth aspect, the one or more bacteria are detected by shotgun DNA sequencing, whole genome sequencing, PCR, 16S genomic DNA sequencing, 16S complementary DNA sequencing, 16S RNA sequencing or culturing the one or more bacteria.

[00108] In one embodiment of the eighth aspect, the sample is a faecal sample or an intestinal biopsy.

[00109] In a ninth aspect, the present invention provides a method of diagnosing a subject as being unresponsive to FMT therapy, the method comprising detecting the presence of one or more bacteria in a sample obtained from the subject, wherein when the one or more bacteria are detected, the subject is diagnosed as being unresponsive to FMT therapy.

[001 10] In one embodiment of the ninth aspect, the one or more bacteria are selected from the group consisting of Fusobacterium, Sutterella, Veillonella, Haemophilus, Streptococcus, Bacteroides, Akkermansia, Escherichia, Megamonas, Clostridium XI Va, Prevotella, Dialister, Veillonella, Bilophila, Parvimonas, Bacteroides uniformis and Bacteroides coprocola. The one or more bacteria may be Fusobacterium spp. Preferably, the one or more bacteria is Fusobacterium gonidiaformans.

[001 1 1] In one embodiment of the ninth aspect, the one or more bacteria suppress a metabolic activity in the subject, wherein the metabolic activity is selected from the group consisting of heme biosynthesis, lipopolysaccharide biosynthesis, ubiquinone biosynthesis, terpenoid quinine biosynthesis, terpenoid backbone biosynthesis, lysine biosynthesis and oxidative phosphorylation.

[001 12] In one embodiment of the ninth aspect, the one or more bacteria are detected by shotgun DNA sequencing, whole genome sequencing, PCR, 16S genomic DNA sequencing, 16S complementary DNA sequencing, 16S RNA sequencing or culturing the one or more bacteria.

[001 13] In one embodiment of the ninth aspect, the sample is a faecal sample or an intestinal biopsy.

[001 14] In a tenth aspect, the present invention provides a method of overcoming a lack of response to FMT in a subject, the method comprising administering a composition to the subject wherein the composition is enriched with one or more bacteria.

[001 15] In one embodiment of the tenth aspect, the composition comprises a faecal sample.

[001 16] In one embodiment of the tenth aspect, the one or more bacteria are selected from the group consisting of Eubacterium, Ruminococcus, Lachnospiraceae, Roseburia, Faecalibacterium, Eubacterium, Clostridium XVIII, Dorea, Coprococcus and Bacteroides fragilis. The one or more bacteria may be a member of the Firmicutes phylum. Preferably, the one or more bacteria is Eubacterium hallii.

[001 17] In one embodiment of the tenth aspect, the one or more bacteria promote a metabolic activity in the subject, wherein the metabolic activity is selected from the group consisting of benzoate degradation, glycerophospholipid metabolism, secondary bile acid biosynthesis, ppGpp biosynthesis, pyruvate fermentation to acetate and lactate, fatty acid biosynthesis, propanoate metabolism, ansamycin biosynthesis and starch degradation.

[001 18] In an eleventh aspect, the present invention provides a method of overcoming a lack of response to FMT in a subject, the method comprising administering a faecal sample to the subject wherein the faecal sample is depleted in one or more bacteria. [001 19] In one embodiment of the eleventh aspect, the one or more bacteria are selected from the group consisting of Fusobacterium, Sutterella, Veillonella, Haemophilus, Streptococcus, Bacteroides, Akkermansia, Escherichia, Megamonas, Clostridium XI Va, Prevotella, Dialister, Veillonella, Bilophila, Parvimonas, Bacteroides uniformis and Bacteroides coprocola. The one or more bacteria may be Fusobacterium spp. Preferably, the one or more bacteria is Fusobacterium gonidiaformans.

[00120] In one embodiment of the eleventh aspect, the one or more bacteria suppress a metabolic activity in the subject, wherein the metabolic activity is selected from the group consisting of heme biosynthesis, lipopolysaccharide biosynthesis, ubiquinone biosynthesis, terpenoid quinine biosynthesis, terpenoid backbone biosynthesis, lysine biosynthesis and oxidative phosphorylation.

[00121 ] In a twelfth aspect, the present invention provides a method of treating ulcerative colitis in a subject in need thereof, the method comprising administering to the subject a therapeutically effective amount of one or more bacteria or bacterial derivatives.

[00122] A therapeutically effective amount of a composition is the amount, when administered to an individual in need thereof, either in a single dose or as part of a series, that is effective for treating the relevant condition. The effective amount will vary depending on the health and physical condition of the individual to be treated, the formulation of the composition, the assessment of the medical situation, and other relevant factors. It is expected that the amount will fall in a relatively broad range that can be determined through routine trials.

[00123] In certain embodiments, a therapeutically effective amount of a composition comprises at least about 10 5 , 10 6 , 10 7 , 10 8 , 10 9 , 10 1 °, 10 11 , 10 12 or 10 13 cfu. A therapeutically effective amount may be from about 10 8 cfu to about 10 14 cfu, from about 10 9 cfu to about 10 13 cfu, from about 10 1 ° cfu to about 10 12 cfu, from about 10 9 cfu to about 10 14 cfu, from about 10 9 cfu to about 10 12 cfu, from about 10 9 cfu to about 10 11 cfu, from about 10 9 cfu to about 10 1 ° cfu, from about 10 1 ° cfu to about 10 14 cfu, from about 10 1 ° cfu to about 10 13 cfu, from about 10 11 cfu to about 10 14 cfu, from about 10 11 cfu to about 10 13 cfu, from about 10 12 cfu to about 10 14 cfu, or from about 10 13 cfu to 10 14 about cfu. In certain embodiments, a therapeutically effective amount of a composition comprises at least about 10 5 , 10 6 , 10 7 , 10 8 , 10 9 , 10 1 °, 10 1 1 , 10 12 , or 10 13 cells or spores. A therapeutically effective amount may be from about 10 4 to about 10 5 , from about 10 5 to about 10 6 , from about 10 6 to about 10 7 , from about 10 7 to about 10 8 , from about 10 5 to about 10 8 , from about 10 6 to about 10 8 , from about 10 5 to about 10 9 , from about 10 5 to about 10 1 °, from about 10 8 to about 10 14 , from about 10 9 to about 10 13 , from about 10 1 ° to about 10 12 , from about 10 9 to about 10 14 , from about 10 9 to about 10 12 , from about 10 9 to about 10 11 , from about 10 9 to about 10 1 °, from about 10 1 ° to about 10 14 , from about 10 1 ° to about 10 13 , from about 10 11 to about 10 14 , from about 10 11 to about 10 13 , from about 10 12 to about 10 14 , or from about 10 13 to about 10 14 cells or spores.

[00124] In one embodiment of the twelfth aspect, the one or more bacteria are selected from the group consisting of Eubacterium, Ruminococcus, Lachnospiraceae, Roseburia, Faecalibacterium, Eubacterium, Clostridium XVIII, Dorea, Coprococcus and Bacteroides fragilis. The one or more bacteria may be a member of the Firmicutes phylum. Preferably, the one or more bacteria is Eubacterium hallii.

[00125] In one embodiment of the twelfth aspect, the one or more bacteria promote a metabolic activity in the subject, wherein the metabolic activity is selected from the group consisting of benzoate degradation, glycerophospholipid metabolism, secondary bile acid biosynthesis, ppGpp biosynthesis, pyruvate fermentation to acetate and lactate, fatty acid biosynthesis, propanoate metabolism, ansamycin biosynthesis and starch degradation.

[00126] In one embodiment of the twelfth aspect, the therapeutically effective amount of one or more bacteria is administered to the subject by FMT.

[00127] In a thirteenth aspect, the present invention provides a method of treating ulcerative colitis in a subject in need thereof, the method comprising enhancing the growth of one or more bacteria in the subject.

[00128] In one embodiment of the thirteenth aspect, the one or more bacteria are selected from the group consisting of Eubacterium, Ruminococcus, Lachnospiraceae, Roseburia, Faecalibacterium, Eubacterium, Clostridium XVIII, Dorea, Coprococcus and Bacteroides fragilis. The one or more bacteria may be a member of the Firmicutes phylum. Preferably, the one or more bacteria is Eubacterium hallii.

[00129] In one embodiment of the thirteenth aspect, the one or more bacteria promote a metabolic activity in the subject, wherein the metabolic activity is selected from the group consisting of benzoate degradation, glycerophospholipid metabolism, secondary bile acid biosynthesis, ppGpp biosynthesis, pyruvate fermentation to acetate and lactate, fatty acid biosynthesis, propanoate metabolism, ansamycin biosynthesis and starch degradation.

[00130] In a fourteenth aspect, the present invention provides a method of preparing an FMT sample, the method comprising: culturing one or more bacteria thereby to produce a culture; and adding the culture or a part thereof to a faecal sample thereby to produce the FMT sample.

[00131] Those skilled in the art will be aware that a faecal sample may be enriched in one or more bacterial by means other than by the addition of a culture. For example, the one or more bacteria may be isolated or purified from a source such as a natural source and then added to a faecal sample.

[00132] In one embodiment of the fourteenth aspect, the one or more bacteria are selected from the group consisting of Eubacterium, Ruminococcus, Lachnospiraceae, Roseburia, Faecalibacterium, Eubacterium, Clostridium XVIII, Dorea, Coprococcus and Bacteroides fragilis. The one or more bacteria may be a member of the Firmicutes phylum. Preferably, the one or more bacteria is Eubacterium hallii.

[00133] In one embodiment of the fourteenth aspect, the one or more bacteria promote a metabolic activity in the subject, wherein the metabolic activity is selected from the group consisting of benzoate degradation, glycerophospholipid metabolism, secondary bile acid biosynthesis, ppGpp biosynthesis, pyruvate fermentation to acetate and lactate, fatty acid biosynthesis, propanoate metabolism, ansamycin biosynthesis and starch degradation.

[00134] In a fifteenth aspect, the present invention provides an FMT sample when prepared by the method of the fourteenth aspect.

[00135] In a sixteenth aspect, the present invention provides a probiotic composition comprising a therapeutically effective amount of one or more bacteria. [00136] In one embodiment of the sixteenth aspect, the one or more bacteria are selected from the group consisting of Eubacterium, Ruminococcus, Lachnospiraceae, Roseburia, Faecalibacterium, Eubacterium, Clostridium XVIII, Dorea, Coprococcus and Bacteroides fragilis. The one or more bacteria may be a member of the Firmicutes phylum. Preferably, the one or more bacteria is Eubacterium hallii.

[00137] In one embodiment of the sixteenth aspect, the one or more bacteria promote a metabolic activity in the subject, wherein the metabolic activity is selected from the group consisting of benzoate degradation, glycerophospholipid metabolism, secondary bile acid biosynthesis, ppGpp biosynthesis, pyruvate fermentation to acetate and lactate, fatty acid biosynthesis, propanoate metabolism, ansamycin biosynthesis and starch degradation.

[00138] In a seventeenth aspect, the present invention provides a method of overcoming a lack of responsiveness to FMT therapy in a subject in need thereof, the method comprising reducing the level of one or more bacteria in the subject.

[00139] In one embodiment of the seventeenth aspect, the subject in need thereof suffers from a gastrointestinal disorder. The gastrointestinal disorder may be ulcerative colitis.

[00140] In one embodiment of the seventeenth aspect, the one or more bacteria are selected from the group consisting of Fusobacterium, Sutterella, Veillonella, Haemophilus, Streptococcus, Bacteroides, Akkermansia, Escherichia, Megamonas, Clostridium XI Va, Prevotella, Dialister, Veillonella, Bilophila, Parvimonas, Bacteroides uniformis and Bacteroides coprocola. The one or more bacteria may be Fusobacterium spp. Preferably, the one or more bacteria is Fusobacterium gonidiaformans.

[00141] In one embodiment of the seventeenth aspect, the one or more bacteria suppress a metabolic activity in the subject, wherein the metabolic activity is selected from the group consisting of heme biosynthesis, lipopolysaccharide biosynthesis, ubiquinone biosynthesis, terpenoid quinine biosynthesis, terpenoid backbone biosynthesis, lysine biosynthesis and oxidative phosphorylation.

[00142] In a eighteenth aspect, the present invention provides a method of identifying an ulcerative colitis patient at risk of not responding to FMT therapy, the method comprising testing for the presence or absence of one or more bacteria selected from the group consisting of Fusobacterium, Sutterella, Veillonella, Haemophilus, Streptococcus, Bacteroides, Akkermansia, Escherichia, Megamonas, Clostridium XlVa, Prevotella, Dialister, Veillonella, Bilophila, Parvimonas, Bacteroides uniformis and Bacteroides coprocola in a sample obtained from the patient, wherein the presence of the one or more bacteria is indicative of the patient having an increased risk of not responding to FMT therapy.

[00143] The one or more bacteria of the eighteenth aspect may be Fusobacterium spp. Preferably, the one or more bacteria is Fusobacterium gonidiaformans.

[00144] In a nineteenth aspect, the present invention provides a method of treating ulcerative colitis in a subject in need thereof, the method comprising suppressing the growth of one or more bacteria in the subject. Those skilled in the art will understand that suppressing the growth of one or more bacteria may include slowing the growth of the bacteria or eliminating the bacteria. The present invention also contemplates methods of selecting to avoid one or more bacteria.

[00145] In one embodiment of the nineteenth aspect, the one or more bacteria are selected from the group consisting of Fusobacterium, Sutterella, Veillonella, Haemophilus, Streptococcus, Bacteroides, Akkermansia, Escherichia, Megamonas, Clostridium XI Va, Prevotella, Dialister, Veillonella, Bilophila, Parvimonas, Bacteroides uniformis and Bacteroides coprocola. The one or more bacteria may be Fusobacterium spp. Preferably, the one or more bacteria is Fusobacterium gonidiaformans.

[00146] In one embodiment of the nineteenth aspect, the one or more bacteria suppress a metabolic activity in the subject, wherein the metabolic activity is selected from the group consisting of heme biosynthesis, lipopolysaccharide biosynthesis, ubiquinone biosynthesis, terpenoid quinine biosynthesis, terpenoid backbone biosynthesis, lysine biosynthesis and oxidative phosphorylation.

[00147] In the methods described herein, the patient may also be treated with a further therapeutic agent such as an anti-inflammatory drug or an immunosuppressant. In certain embodiments, the method of the present invention comprises pretreating a subject with an antibiotic composition prior to administering a therapeutic composition. Suitable antibiotics include, for example, rifabutin, clarithromycin, clofazimine, vancomycin, rifampicin, nitroimidazole, chloramphenicol, rifaximin, rifamycin derivative, rifabutin, rifapentine, rifalazil, bicozamycin, aminoglycoside, gentamycin, neomycin, streptomycin, paromomycin, verdamicin, mutamicin, sisomicin, netilmicin, retymicin, kanamycin, aztreonam, aztreonam macrolide, clarithromycin, dirithromycin, roxithromycin, telithromycin, azithromycin, bismuth subsalicylate, fidaxomicin, amikacin, arbekacin, netilmicin, paromomycin, rhodostreptomycin, tobramycin, apramycin or a combination thereof.

[00148] The composition of the present invention may take many different forms. For example, the composition may be formulated as an enteric coated capsule or an enteric coated microcapsule, or formulated as part of or administered together with a food, a food additive, a dairy-based product, a soy-based product or a derivative thereof, a jelly, or a yogurt. The composition may be provided as a powder optionally in combination with a food or drink. A food or drink may be a dairy-based product or a soy- based product. In certain embodiments, a food or food supplement contains enteric- coated microcapsules containing a composition of the present invention.

[00149] The composition of the present invention may comprise a liquid culture. In certain embodiments, the composition is lyophilized, pulverized and powdered. It may then be infused, dissolved such as in saline, as an enema. Alternatively the powder may be encapsulated as enteric-coated capsules for oral administration. These capsules may take the form of enteric-coated microcapsules. A powder may be provided in a palatable form for reconstitution for drinking or for reconstitution as a food additive. In one aspect, a powder may be reconstituted to be infused via naso-duodenal infusion. In further embodiments, the composition of the present invention may be administered in a liquid, frozen, freeze-dried, spray-dried, lyophilized or powder form. The composition may be formulated as a delayed or gradual enteric release form. In certain embodiments, the composition comprises an excipient, a saline, a buffer, a buffering agent, or a fluid- glucose-cellobiose agar (RGCA) media. The composition may also comprise a cryoprotectant. A cryoprotectant may include polyethylene glycol, skim milk, erythritol, arabitol, sorbitol, glucose, fructose, alanine, glycine, proline, sucrose, lactose, ribose, trehalose, dimethyl sulfoxide (DMSO), glycerol, or a combination thereof. The composition of the present invention may comprise an acid suppressant, an antacid, an H 2 antagonist, a proton pump inhibitor or a combination thereof. In certain embodiments, the composition is substantially free of non-living matter such as acellular material like residual fiber, DNA, viral coat material, and non-viable material. [00150] In certain embodiments, the composition of the present invention comprises faecal microbiota. The preparation of faecal microbiota may involve, for example, ethanol treatment, detergent treatment, heat treatment, irradiation or sonication. In one embodiment, the preparation of a faecal microbiota involves a separation step such as by way of density gradients, filtration (e.g., sieves, nylon mesh) or chromatography. In certain embodiments, a faecal microbiota used herein comprises a donor's entire faecal microbiota. In certain embodiments, the composition comprises a faecal microbiota substantially free of eukaryotic cells from the donor. The faecal sample or composition may also be supplemented with bacteria grown in a culture.

[00151] Those skilled in the art will understand that prebiotics may also be used in accordance with the present invention to promote the growth of certain bacteria. A prebiotic nutrient can be any ingredient that stimulates the stability, growth and/or activity of the faecal flora, in particular, bacteria. By way of example, polyols, fructooligosaccharides (FOSs), oligofructoses, inulins, galactooligosaccharides (GOSs), xylooligosaccharides (XOSs), polydextroses, monosaccharides such as tagatose, and/or mannooligosaccharides may be used as prebiotics to practice the present invention. In one embodiment, the prebiotics are added to prevent "shock" to the faecal flora subsequent to their isolation or purification, freezing, freeze-drying, spray-drying, reconstitution in solution and the like.

[00152] The terms "comprise", "comprises", "comprised" or "comprising", "including" or "having" and the like in the present specification and claims are used in an inclusive sense, ie, to specify the presence of the stated features but not preclude the presence of additional or further features.

[00153] The term "about" is understood to refer to a range of +/- 10%, preferably +/- 5% or +/- 1% or, more preferably, +/- 0.1 %.

Examples

[00154] In a randomised controlled FOCUS Study, multi-donor faecal microbiota transplant (FMT) was effective in the treatment of active ulcerative colitis (UC). Flere the inventors characterise the bacterial taxonomic and functional changes associated with FMT in UC and those predictive of therapeutic outcomes. Although the examples refer primarily to UC, those skilled in the art will understand that the methods and compositions described herein may be suitable for treating other gastrointestinal disorders.

Study design

[00155] Active UC patients were randomised in a double blind controlled trial to intensive multi-donor FMT or placebo enemas 5 days per week for 8 weeks. Patients randomised to placebo were eligible for open label FMT after the double-blind study period. FMT infusions were constituted from the blended homogenised stool of 3 to 7 unrelated donors to increase microbial heterogeneity. Each patient received all their FMT infusions from the same donor batch to ensure consistency and reproducibility of the infused faecal microbiota.

[00156] 314 faecal and 160 colonic biopsy samples were collected at fixed intervals from 70 patients. 1 13 faecal samples were collected from the 14 individual donors and 21 donor batches. DNA and RNA were extracted, RNA converted to cDNA, then 16S rRNA gene sequencing performed using 2x300 bp lllumina MiSeq chemistry. Raw sequences were analysed using mothur. Shotgun metagenomics was performed on 285 faecal samples using 2x250 bp HiSeq 2500 chemistry and analysed using MetaPhlAn2 and HUMANn2.

Trial samples

[00157] Samples were collected from individual faecal donors, multi-donor FMT batches and study patients for molecular microbiological analyses and gastrointestinal microbial community profiling. Faecal samples (1 13 in total) were collected from all of the 14 individual donors (55 samples) and the 21 multi-donor FMT batches (50 samples) utilised in the trial, as well as 4 placebo batches (8 samples) to serve as a control. Samples (blinded and open label) for molecular microbiological analyses were collected from all 70 study patients recruited at the two Sydney-based trial sites. Study patient faecal samples (314 in total) were collected at screening, then every 4 weeks during study therapy (blinded, and open label if applicable) and at final review 8 weeks after finishing blinded or open-label FMT therapy. Study patient colonic biopsies (160 in total) were collected at the study endoscopic evaluation at the start of treatment, end of treatment (week 8), and end of open label treatment if applicable.

Sample handing and sequencing [00158] All samples were immediately stored at -80°C upon collection until nucleic acid extraction. Faecal samples were homogenised and both DNA and RNA were extracted using the MOBIO PowerViral RNA/DNA Isolation kit. Faecal RNA was then isolated from DNA using the MOBIO On-Spin Column DNase kit and Bioline Isolate II RNA micro clean-up kit. Colonic biopsy samples were homogenised and bacterial DNA and RNA extracted using the Macherey-Nagel RNA Isolation Kit. Colonic RNA was then isolated from DNA using the MOBIO On-Spin Column DNase kit and Macherey-Nagel RNA clean-up kit. Faecal and colonic RNA was then converted to cDNA using the SensiFAST cDNA Synthesis Kit (Bioline).

[00159] The 16S rRNA gene fragment of the extracted DNA and RNA converted to cDNA was amplified using the Immolase DNA polymerase (95°C for 10 min, 35 cycles of 94°C for 30 s, 55°C for 10 s, 72°C for 45 s, followed by a final step of 72°C for 10 min) and the primers F27-519R. Indices and lllumina sequencing adapters were attached using the Nextera XT Index Kit according to the manufacturer’s instructions. Amplicon sequencing was performed with lllumina MiSeq 2x300 bp chemistry at the Ramaciotti Centre for Genomics. Shotgun metagenomics was performed on DNA extracted from 285 donor and patient faecal samples using Nextera XT DNA library prep kit and 2x250 bp HiSeq 2500 chemistry. This resulted in five datasets including faecal 16S DNA, faecal 16S cDNA, biopsy 16S DNA, biopsy 16S cDNA, and faecal shotgun DNA.

Raw read analyses

[00160] Quality filtering of 16S rRNA sequences was conducted using mothur [Schloss Applied and Environmental Microbiology] and followed the mothur MiSeq SOP [Kozich Applied and Environmental Microbiology ]. Paired-end sequences were merged into contigs, and poor quality contigs removed based on alignment quality and ambigious base calls. A multiple sequence alignment was constructed using the SILVA SEED 16S rRNA reference alignment, and poorly aligned sequenced removed. To eliminate artefacts of sequencing at high frequency, rare sequences with high similarity to abundant sequences were clustered together. Chimeric sequences were removed. Sequences were taxonomically classified and those without classification at the kingdom level (unknown), or classified as mitochondrial or chloroplast were removed. Quality filtered sequences were then clustered into operational taxonomic units at 97% similarity using the opti-clust average neighbour algorithm, and consensus taxonomies of the OTUs obtained using the classifications of sequences within each OTU. The resulting OTU count by sample data matrix was used for data analysis.

[00161] Shotgun metagenomic reads were first analysed with DeconSeq [Schmieder PLoS One] for identification and filtering of human DNA sequences. Sequencing reads were assessed for quality using FastQC (version 0.1 1.2). SolexaQA was then applied to calculate sequence quality statistics and perform quality filtering of the lllumina reads. Paired-end raw reads were trimmed with the BWA trimming mode at a threshold of Q13 (P=0.05) using the read trimmer module DynamicTrim. Filtered reads that were less than 25 bp in length were then discarded using LengthSort. The average microbial read counts per sample were 4590171 ± 1 19145 reads. MetaPhlAn2 [Truong Nat Methods] was employed to generate taxonomic profiles from the shotgun reads, while HUMAnN2 (HMP Unified Metabolic Analysis Network) [Flail Genome medicine] was used to determine the metabolic contributions within the samples. The HUMAnN2 pipeline involved mapping of the metagenomic reads against Uniref orthologous gene family, MetCyc UniPathway, and KEGG.

Experimental design

[00162] During the initial double-blind FMT trial, patients were allocated into treatment or placebo groups (FMT : two levels - Placebo or FMT, factor type - fixed) and each patient was sampled at three time points over an eight-week period (Time: three levels - 0, 4 and 8 weeks, factor type - fixed). Each patient was included in the experimental design as a random factor. After 8 weeks, the placebo group received FMT and were sampled at weeks 4 and 8 under an open-label (non-blind) FMT trial. All patients receiving FMT were also sampled at 8 weeks post FMT completion (follow-up), either blinded or open-label.

[00163] To examine which microbial taxa differed between patients showing remission, the inventors first combined the data from the blinded and open-label trials, and then created groups based on remission (Remission, two levels - Yes or No, factor type - fixed) and treatment group (FMT : three levels - Placebo, FMTblind or FMTopen). The effect of remission was then examined across a number of contrasts, including those comparing remission within the blinded trial (Remission among Placebo and FMTblind), and those comparing remission regardless of trial (Remission among Placebo, FMTblind and FMTopen). Patient remission was measured using four different remission classifications - primary endpoint, clinical remission, endoscopic response, and endoscopic remission (steroid free endoscopic Mayo score of 0) [Paramsothy Lancef\.

[00164] The effect of donor batch on remission was examined by allocating donor batches into two groups based on the number of patient remissions observed for each batch (DonorRemission, two levels - Yes or No, factor type - fixed). If more than 50% of the patients receiving a particular donor batch showed remission, the donor batch was allocated to the DonorRemission = Yes group, while all other donors were allocated to the DonorRemission = No group.

Microbial communities

[00165] Microbial communities were examined with respect to the above analyses in terms of alpha-diversity and beta-diversity, as well as comparing each taxon individually. Prior to diversity comparisons, the OTU counts were rarefied to account for uneven sequencing depths among samples. A number of OTUs and Shannon diversity index were used as alpha diversity measures, and used linear mixed models (LMMs) to examine the effects of the various predictors mentioned above. Means and 95% confidence intervals for figures were derived from the LMMs. Models were created using the R packages LME4 [Bates Journal of Statistical Software] and ImerTest [Kuznetsova Journal of Statistical Software ].

[00166] Bray-Curtis (BC) dissimilarity coefficient for beta-diversity comparisons was employed, and prior to calculation of BC dissimilarities, OTU counts were transformed into square-root relative abundances. The BC distance matrix was visualised using non-metric multidimensional scaling (nMDS). PERMANOVA was used to examine the effects of the various predictors mentioned above. Dissimilarities, figures and models were created using the R package‘vegan’ [Oksanen, Vegan: Community Ecology Package ]. For per taxon comparisons, un-rarefied OTU counts in negative binomial generalised linear models (GLMs) were used, and employed contrasts to examine the comparisons of interest within each analysis. Models were created using the R package DESeq2 [Love Genome Biology ]. Linear Discriminant Analysis Effect Size (LEfSe) analyses were also performed [Segata Genome Biology ].

Summary of results [00167] a-diversity consistently increased following FMT across all datasets (faecal 16S DNA & cDNA, colonic 16S DNA & cDNA, shotgun metagenomics) a- diversity saturated by week 4 with trends towards greater a-diversity with remission b- diversity across all datasets demonstrated FMT significantly shifted global microbial composition, with a transference from Bacteroides to Prevotella dominance. Fusobacterium consistently associated with lack of remission, as did, to a lesser extent, Sutterella, Veillonella and Flaemophilus. There was little consistency in taxa associated with remission (most commonly members of Firmicutes e.g. Clostridium XVIII, Ruminococcus, Lachnospiraceae). Presence of a Streptococcus OTU in donors was generally associated with ineffective batches. Shotgun metagenomics of faecal samples demonstrated shifts in overall bacterial metabolic functions with FMT, with specific pathways associated with remission (starch degradation, short chain fatty acid production) or lack of remission (heme biosynthesis). Taxa contributing to beneficial pathways included Eubacterium, Ruminococcus, Lachnospiraceae and Roseburia.

FMT induces changes in microbial diversity and composition independent of outcome

[00168] The effect of FMT on the gut microbiome of UC patients was comprehensively assessed independent of therapeutic outcome. An increase in a- diversity, as measured by number of OTUs and Shannon’s diversity, was observed in the resident faecal microbiota (16S rRNA gene) of patients undergoing blinded FMT therapy (Figure 1 ). To determine the strength of these changes, the data was fitted into a model of diversity explained by time and treatment with patient ID as a random factor (Figure 2). The trends in number of OTUs across time were significantly different between patients on FMT and those on placebo (ChiSq=46.6, df=2, P<0.001 ; Figure 3). This was also observed for Shannon’s diversity (ChiSq=6.5, df=2, P=0.039; Figure 3). Pairwise comparisons across time and treatment found significant differences in a- diversity between baseline and post-FMT samples but not between baseline and post placebo samples (Figure 4). Differences were also identified between post-FMT and post-placebo at week 4 and week 8 but not at baseline (Figure 5). The above findings were replicated in the patients that underwent open label FMT therapy (Figures 6 and 7). Importantly, changes in a-diversity appeared to saturate at week 4 of FMT (Figure 3), and changes were maintained at final follow-up 8 weeks post-FMT therapy (Figures 6 and 7). [00169] A significant shift in b-diversity (composition) was also observed in the faecal microbiota (16S rRNA gene) post-FMT but not post-placebo, and these changes were sustained at follow-up (Figures 8 and 9). Fleat map analysis showed a range of OTUs were either transplanted or depleted post-FMT but not during placebo, including those belonging to Bacteroides, Prevotella, Clostridium XIVa, Parabacteroides, Blautia, and Parasutterella (Figure 10).

[00170] To validate these changes in diversity and composition in the resident faecal microbiota using more robust techniques, a subset of faecal DNA samples (n=285) were shotgun sequenced, and taxonomic information was extracted. An increase in the number of OTUs in post-FMT samples but not post-placebo samples when compared to baseline samples was confirmed (Figure 1 1 ), and this difference was found to be significant when data was fitted into a model similar to the above (P=0.027; Figures 12 and 13). However, increases in Shannon’s diversity were not replicated in the shotgun sequencing data (Figures 12 and 13). Changes in b-diversity following FMT therapy were also identified in the taxonomic information arising from the shotgun sequencing (Figure 14), and these were remarkably similar to changes observed using 16S rRNA gene sequencing (Figure 15). The shift in b-diversity following FMT therapy could be more clearly seen when a constraint on the factor‘patient’ was applied to eliminate inter-patient variability (Figure 16). In addition to changes in composition, FMT therapy increased homogeneity (reduced dispersion) in the taxonomic profiles across patient samples to a level observed in the individual donors (Figure 17).

[00171] To determine if FMT therapy had similar effects on the active/viable faecal microbiota, 16S rRNA transcript sequencing was performed on faecal cDNA. While a higher number of OTUs was observed in the active faecal microbiota when compared to the faecal microbiota derived from DNA sequencing (Figure 18), similar trends in both a- diversity and b-diversity were observed post-FMT therapy or post-placebo (Figures 18 to 20). Heatmap analysis also showed that the same OTUs were either transplanted or depleted in the active faecal microbiota post-FMT (Figure 21 ).

[00172] The resident and active gut mucosal microbiota derived from colonic biopsy samples were also assessed by 16S rRNA gene and transcript sequencing. Similar to faecal samples, a higher number of OTUs was observed in the active mucosal microbiota when compared to the resident mucosal microbiota (Figure 22). FMT therapy but not placebo led to increases in a-diversity and shifts in b-diversity in the mucosal microbiota as was observed for the faecal microbiota (Figures 22-25).

Specific bacterial taxa are consistently associated with therapeutic outcome

[00173] FMT-treated patients who achieved the primary outcome (steroid free clinical remission with endoscopic remission or response) tended to have higher baseline and post-therapy species richness than those who did not achieve the primary outcome [Paramsothy Lancef\. This higher baseline and post-FMT species richness was also observed in the mucosal microbiome (Figure 26); however, while these results showed a trend, they did not reach statistical significance.

[00174] Changes in global microbial composition were also associated with primary outcome. Analysis of shotgun sequencing data identified differences at week 8 of FMT and at 8 weeks follow-up between patients who went into remission and those that did not (Week 8: t=1 .26, P=0.077, Permutations=999, df=51 ; Follow-up: t=1.41 , P=0.018, Permutations=999, df=50). Cluster analysis of shotgun taxonomic results highlighted the differences between baseline and post-placebo samples as compared to post-FMT samples (Figure 27). Patients who achieved the primary outcome also appeared to cluster tightly together unlike those that did not, the latter forming two separate groups that highlight inter-patient variability in lack of remission (Figure 27). Further, despite intense FMT therapy, five patients did not appear to have a dramatic change in overall microbial structure, their baseline samples clustering tightly with their post-FMT samples (Figure 27). Surprisingly, one of these patients achieved the primary outcome, and on further analysis, the overall structure of their microbiota was similar at baseline and post-FMT except for replacement of key species Bacteroides clarus (1 1.5% to 0.06%) and Akkermansia muciniphila (1 1.1 % to 0%) with Faecalibacterium prausnitzii (4.9% to 1 1.1 %), Eubacterium rectale (0.19% to 9.9%), and Eubacterium siraeum (0.96% to 14.2%).

[00175] To identify specific microbial taxa significantly associated with primary outcome across all patients, DESeq2 analyses were performed on all datasets (faecal and mucosal 16S rRNA gene and transcript, as well as faecal shotgun taxonomic data), with only the top taxa being presented as potential biomarkers. A range of microbial taxa were identified to be associated with lack of remission (primary outcome) including Fusobacterium, Sutterella, Haemophilus, Escherichia, Megamonas, Clostridium XI Va, Prevotella, Dialister, Veillonella and Bilophila (Figures 28 to 32), and these associations were in some datasets clearer when blinded and open label patients were stratified (Figures 33 to 36). The most consistent association with lack of achieving primary outcome was with Fusobacterium gonidiaformans, with this taxon identified in faecal 16S rRNA gene (Figure 33), mucosal 16S rFtNA gene and transcript (Figures 30 and 31 ), and shotgun sequencing data (Figure 32). Of interest, Prevotella OTU2 ( Prevotella copri in shotgun data) appeared to flourish in a range of patients post-FMT (Figures 15 and 21 ); however, this OTU was associated with lack of remission (Figures 28, 30 and 31 ) and patients who achieved remission tended to be those who resisted dominance by Prevotella, having lower levels relative to the patients who did not achieve remission (Figure 37). There was less consistency in taxa associated with remission across the differing datasets - these most commonly involved members of Firmicutes e.g. Clostridium XVIII, Ruminococcus, Lachnospiraceae, Roseburia inulinivorans, and Eubacterium hallii. The associations among a range of these microbial taxa and primary outcome were confirmed using LEfSe (Figure 38). Notably, one patient (1003) who had a strikingly positive response to FMT [Paramsothy Lancet] had high levels of Eubacterium hallii post-FMT.

[00176] To further examine the consistency of these associations, DESeq2 analyses were replicated against three other therapeutic outcomes including the stricter endpoint of complete endoscopic remission (steroid free endoscopic Mayo 0), endoscopic response, and clinical remission. Fusobacterium gonidiaformans, Sutterella wadsworthensis, Haemophilus, Escherichia, Megamonas, Clostridium XlVa, Prevotella, Dialister, Veillonella and Bilophila were all found to be consistently associated with lack of endoscopic remission (Figures 39 to 43), and this was also observed when blinded and open label patients were stratified (Figures 44 to 47). Interestingly, another member of the oral microbiota, Parvimonas was also found to be associated with lack of endoscopic remission in the active mucosal microbiota (Figure 42). Results from the analysis of endoscopic response and clinical remission were less consistent, likely due to the less strict nature of these endpoints; however, a range of the above taxa (e.g. Fusobacterium, Haemophilus, Escherichia, Dialister and Veillonella) were still identified to be associated with negative outcomes (Figures 48 to 57).

FMT results in functional changes associated with therapeutic outcome [00177] Microbial functional outputs were defined using HUMANn2, and changes across FMT therapy and therapeutic outcome were characterised, with analysis focusing on outputs from KEGG and MetaCyc pathways. FMT, and not placebo, resulted in significant changes in microbial KEGG pathways (Figures 58 and 59), and this was replicated in the MetaCyc pathways (Figure 60). Despite intense FMT, patient microbial functional profiles remained significantly different to that of the donors (Figure 59). Similar to the taxonomic profiles, FMT increased homogeneity (reduced dispersion) in the functional profiles across patient samples, but these levels did not reach those of the individual donors or batch donor samples (Figure 61 ). Due to the significant patient variability that was observed in the data (Figures 58 and 60), a constraint on the factor ‘patient’ was applied, which showed a clearer delineation between FMT and placebo (Figure 62).

[00178] Differences in global microbial functional profiles were associated with primary outcome, but this was only observed when blinded and open label patients were stratified (Figure 63). Specific pathways associated with primary outcome were then identified using DESeq2. Pathways such as benzoate degradation, glycerophospholipid metabolism, secondary bile acid biosynthesis, ppGpp biosynthesis, pyruvate fermentation to acetate and lactate (short chain fatty acid biosynthesis), biosynthesis of ansamycins, and starch degradation were all associated with positive primary outcome (Figures 64 and 65). Importantly, taxa contributing to beneficial pathways included Eubacterium, Ruminococcus, Lachnospiraceae, Roseburia, Dorea and Coprococcus, consistent with the taxonomic analysis associating these species with positive therapeutic outcome. Furthermore, the relationship between short chain fatty acid biosynthesis and positive primary outcome was confirmed in the predicted metagenome (PICRUSt) of the mucosal microbiome (Figure 66). In contrast, heme biosynthesis, lipopolysaccharide biosynthesis, ubiquinone and other terpenoid quinine biosynthesis, lysine biosynthesis and oxidative phosphorylation pathways were all associated with negative primary outcome (Figures 64 and 65). The relationships between a range of these pathways and primary outcome were confirmed using LEfSe (Figures 67 and 68).

[00179] Similar to the taxonomic analysis, DESeq2 analyses were replicated against the three other therapeutic outcomes of complete endoscopic remission, endoscopic response, and clinical remission. The results from these endpoints showed consistent associations as those observed for the primary study outcome (Figures 69 to 74). Donor taxonomic and functional profiles are associated with therapeutic outcome

[00180] Despite intense FMT therapy over 8 weeks, patient taxonomic and functional profiles remained different to those of the individual donors and batch donor samples. Thus, specific factors associated with donor suitability were evaluated by analysis of the donor faecal samples (16S rRNA gene and transcript, as well as shotgun metagenomic datasets) relative to the four different therapeutic outcomes.

[00181] Donor batch samples were categorised based on the total number of samples and number of patients that achieved a positive primary outcome (Figure 75), with batch samples leading to >50% remission classified as effective batch samples and the rest as ineffective batches a-diversity and b-diversity within effective and ineffective batches were compared in all datasets (Figures 76 to 80), with no clear patterns emerging between the two groups. While some differences in global b-diversity were observed, this was likely due to the high inter-donor variability (Figure 78).

[00182] Specific taxonomic differences between effective and ineffective batches were then analysed by DESeq2 (Figures 81 to 83). Bacteroides OTU187 was found to be higher in abundance in effective batches (Figure 81 ), and consistently Bacteroides fragilis was identified as the strongest taxonomic marker in these batches (Figure 83). In contrast, Bacteroides uniformis and Bacteroides coprocola were associated with ineffective batches (Figure 83). Other donor microbial taxa associated with ineffective batches included Clostridium XlVa (OTU173), a taxon that was associated with negative primary outcomes, and Streptococcus (OTU56), which was found in both the 16S rRNA gene and transcript datasets (Figures 81 and 82). Surprisingly, low abundance OTUs belonging to beneficial taxa such as Faecal i bacterium (OTU271 ) and Ruminococcus (OTU259) were found in higher abundance in ineffective batches (Figures 81 and 82). Flowever, these results were not replicated in the shotgun sequencing data (Figure 83). Similar to the taxonomic data, no clear patterns in overall pathway compositions were found between effective and ineffective batches (Figures 84 and 86). Flowever, a range of pathways were identified to be in higher abundance in effective and ineffective batches (Figures 85 and 87). Specifically, pathways such as fatty acid biosynthesis and propanoate metabolism were higher in effective batches while terpenoid backbone biosynthesis and bacterial chemotaxis were higher in ineffective batches. [00183] Similar analyses were conducted for the three other therapeutic endpoints. The strict endpoint of endoscopic remission (Figures 88 to 100) and the less strict endpoint of endoscopic response (Figures 101 to 1 13) showed similar outcomes to the primary study endpoint. One notable difference was the clustering of effective batches at the higher end of a-diversity when shotgun taxonomic data was classified by endoscopic response (Figure 107). This higher level of a-diversity was also identified for effective batches classified by the clinical remission endpoint (Figures 1 14, 1 15 and 120). In fact, classification of donor batch effectiveness based on the clinical remission endpoint (Figures 1 14 to 126) showed the strongest signs of consistency with the results of the patient analysis. For example, Sutterella wadsworthensis, previously associated with lack of remission in patients, was associated with ineffective batches in clinical remission (Figure 122). Further, pathways such as secondary bile acid biosynthesis, glycerophospholipid metabolism and biosynthesis of ansamycins were all associated with positive patient outcomes, and were associated with effective batches (Figure 124). Moreover, heme biosynthesis was a strong marker for negative primary outcome in patients, and was also found to be higher in ineffective batches (Figure 126).

Discussion

[00184] There was a surprisingly high level of concordance not only between the trends observed in the 16S rRNA gene (resident) and transcript (active) datasets, but also between those observed in the faecal (luminal) and colonic biopsy (mucosal) datasets.

[00185] FMT was shown to induce a distinct increase in the a-diversity as well as significant shifts in b-diversity (composition) of the microbiota of UC patients. Flowever, at the same time FMT resulted in the recipient microbiota of UC patients becoming more homogeneous (decreased dispersion). These do not appear to be transient effects because (1 ) the changes were observed in both the resident and active (16S rRNA gene and transcript) faecal and mucosal microbiomes, (2) they are replicated using shotgun metagenomics and (3) they are durable at 8 weeks post FMT.

[00186] Alpha-diversity increased with remission but this difference did not reach significance, though it must be noted that the study was not sufficiently powered to assess for this distinction. While some associations between remission and global b- diversity were observed there was substantial inter-patient variability, as demonstrated by the cluster data of the post-FMT samples. Nevertheless, DESeq2 analyses did identify bacterial taxonomic changes in UC patients associated with therapeutic outcome, although identification of changes that consistently associated with lack of remission was more successful than bacterial taxonomic changes associated with remission. The most prominent taxa associating with lack of remission in our cohort included Fusobacterium (specifically Fusobacterium gonidiaformans), Sutterella (specifically Sutterella wadsworthensis), Haemophilus, and Escherichia.

[00187] Little is known about the non-oral species Fusobacterium gonidiaformans, but it has been previously isolated from the intestinal and urogenital tract of humans [Citron Clinical Infectious Diseases ].

[00188] More definitive data regarding the potential metabolic pathways of importance in FMT outcomes for UC were provided by the shotgun metagenomic functional outputs in UC patients post FMT, which unlike the taxonomic analyses revealed more consistent associations with positive therapeutic outcomes. Important pathways linked with remission included benzoate degradation, glycerophospholipid metabolism, secondary bile acid biosynthesis, ppGpp biosynthesis and biosynthesis of ansamycins. The positive relationship between patient outcome and alarmone (ppGpp) biosynthesis, also referred to as the stringent response [Jain J Microbiol ], suggests that it is beneficial for FMT to act as a stressor to the UC dysbiotic microbiome. Further, ansamycins are a family of bacterial secondary metabolites that act as antimicrobial compounds and have been shown to target Gram-positive and Gram-negative bacteria, as well as bacteriophages and some poxviruses [Wehrli Medicinal Chemistry ]. It can be speculated that production of these compounds would likely have a substantial effect on the composition of the gut microbiome.

[00189] Importantly, metabolic shifts to starch degradation and short chain fatty acid (SCFA) biosynthesis (such as pyruvate fermentation to acetate and lactate) were associated with remission. This functional finding was consistent with the taxonomic analyses that revealed many of the organisms that associated with remission were Firmicutes species and known producers of SCFAs. The organism Eubacterium hallii may be of particular interest in this regard, as an increase in E. hallii in UC patients post FMT was associated with remission across a number of the datasets, and consistent with these taxonomic findings is the fact that analyses of the microbial functional outputs data showed that Eubacterium contributed to the identified beneficial metabolic pathways.

[00190] Pathways that were found to associate with negative therapeutic outcomes or lack of remission included heme biosynthesis, lipopolysaccharide biosynthesis, ubiquinone and other terpenoid quinine biosynthesis and oxidative phosphorylation. Heme biosynthesis and lipopolysaccharide biosynthesis have been shown to contribute to bacterial pathogenesis and increased inflammation, respectively [Raetz Ann Rev Biochem, Choby J Mol Biol]. The two pathways ubiquinone and other terpenoid quinine biosynthesis and oxidative phosphorylation are connected through the compound ubiquinone-n, and their association with lack of remission may be a reflection of the beneficial nature of anaerobic fermenters identified in our patients, as well as the harmful effects of resultant reactive oxygen species. Lysine biosynthesis was also associated with lack of remission, which could putatively be secondary to its contribution to bacterial peptidoglycan biosynthesis, or alternatively, a preference towards pyruvate fermentation to SCFAs rather than pyruvate metabolism to lysine.

[00191] Analyses of donor specific factors associated with therapeutic outcomes were performed in an attempt to more directly inform on profiles that may be of importance in the development of future microbial therapeutic in UC, though it should be noted that such analyses were restricted due to the limited sample size. Perhaps the most notable finding was the association of Bacteroides fragilis with effective donor batches, which may possibly relate to it serving to counteract the dominance of Prevotella in UC patients post FMT. Streptococcus, a bacterium commonly associated with the oral cavity, was found in both the 16S rRNA gene and transcript datasets to be associated with lack of remission. Notably, other oral microbial taxa such as Dialister, Veillonella and Parvimonas were associated with negative patient outcomes, and intestinal colonisation by oral bacteria has been previously associated with inflammatory bowel diseases [Atarashi Science]. Consistent taxonomic and functional profile associations across both faecal donor clinical remission data and UC patient data include the negative impact of Sutterella wadsworthensis and heme biosynthesis pathways on patient outcomes and the positive impact of pathways involved in the biosynthesis of ansamycins.

[00192] FMT treatment consistently increased a-diversity and shifted microbial composition on b-diversity measures. Fusobacterium consistently associated with lack of remission while there was less consistency in taxa associated with remission. Shotgun metagenomics demonstrated specific pathways associated with therapeutic outcomes with FMT treatment, with particular species contributing to beneficial pathways. These findings may be important in both understanding the pathophysiology of the microbiota in UC and shaping future bacterial therapy.

[00193] Although the invention has been described with reference to specific embodiments, it will be appreciated by those skilled in the art that the invention may be embodied in many other forms..

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