Role of Microbiota in Health and Disease

Dr Ailsa Hart Director IBD Unit, St Mark’s Hospital, London

Honorary Senior Clinical Lecturer, Imperial College, London

Gut microbiota – structure and function

Role of gut microbiota in IBD

Modulation of gut microbiota as a therapy

- antibiotics, probiotics, prebiotics

- faecal transplant

Role of microbiota in other diseases states

Introduction

The gut microbiota

1014 gut bacteria and 1013 cells in body

Most densely populated ecosystem on Earth

4 major phyla (Bacteroidetes, Firmicutes, Actinobacteria, Proteobacteria)

Provide traits we have not had to evolve on our own

“Virtual” organ

Genes in gut flora 100 times our own genome

Function of gut microbiota Germ-free animals

Exist and survive

But abnormalities of:

Immune function (oral tolerance)

Metabolic function (altered enzymes)

Physiological function (altered motility)

Trophic function (altered cell turnover)

The Meta HIT project

The human microbiome project

• 5 year project launched by NIH published in Nature • 242 healthy men/women - samples from different body sites • 10,000 different types of organism found • Is there a core set of microbes that all humans share?

• Diversity of microbes across human beings

• Diet, host genetics, early microbial exposure • Unique communities of microbes at different body sites • At specific body sites, many microbes had similar genes/functions

The human microbiome project

Challenges thinking of one-microbe model of disease More likely “function” of group of microbes changes

Gut microbiota “pathology”

e.g. inflammatory bowel diseases functional bowel disease liver diseases non-GI diseases - obesity, type I diabetes, atopy/allergy

Genetics of IBD

1980

’s

2001

20

07

2008

20

11

2012

>160 independent IBD susceptibility loci

- Information about pathways involved in disease process - Over 2/3 of genes are shared between UC &CD - 30 CD-specific and 23 UC-specific - Overlap between ankylosing spondylitis and psoriasis - Overlap between susceptibility loci for IBD & mycobacterial infection

1Jostins et al Nature 491 (7422):119-124; 2Lees et al Gut 2011;60:1739-53 (diagram)

> 75,000 cases & controls

>160 independent IBD susceptibility loci

Genetics of IBD

Experimental models of colitis

Germ-Free No Colitis

Animal models of colitis

Experimental models of colitis

Bacterial Colonisation

Germ-Free

Colitis

No Colitis

Animal models of colitis

In humans…

Faecal stream diversion alleviates Crohn’s

Reanastomosis triggers recurrence

Infusion of luminal contents into excluded normal

bowel induces inflammation

What part of the gut microbiota drives inflammation?

Single organism?

Expansion or relative contraction?

“Functional” changes?

Changes in mycobiome or virome?

Reduced diversity of faecal microbiota in CD

Reduced Firmicutes in Crohn’s disease

- Reduction of a major member of Firmicutes, F.

prausnitzii, associated with higher risk of

postoperative recurrence of ileal CD.

Experimental replacement of F. prausnitzii had anti-inflammatory effects

Reduced diversity of faecal microbiota in UC

Reduced diversity of faecal microbiota in pouchitis

McLaughlin et al. (St Mark’s) Ann Surg 2010 July;252(1):90-8

UC FAP Non-pouchitis pouchitis

What are the specific changes in Crohn’s disease?

Gevers et al. 2014 Cell Host & Microbe 15, 383-93

What are the specific changes in Crohn’s disease?

Fusobacteriaceae • Biomarker • Progression of colorectal cancer

Gevers et al. 2014 Cell Host & Microbe 15, 383-93

What are the specific changes in Crohn’s disease?

Pastueurellacaea, Veillonellaceae, pathogenic E. coli • link with ulcer formation

Gevers et al. 2014 Cell Host & Microbe 15, 383-93

Machiels K, et al. Gut 2014;63:1275–1283

Decrease in Faecalibacterium prausnitzii and Roseburia hominus in UC (decrease in butyrate production)

What are the specific changes in UC?

Major shifts in oxidative

stress pathways Decreased carbohydrate

metabolism Decreased amino acid

synthesis In ileal Crohn’s, increases

in virulence and secretion pathways

But changes in “function” of microbiota….

Morgan et al. Genome Biology Sept 2012

Microbial function more consistently altered than microbial composition

Mucosa/stool samples from 231 IBD patients & controls 16S gene pyrosequencing/shotgun metagenomics

Fungal microbiota in IBD

Inflamm Bowel Dis 2014;0:1–10

Virome in IBD

• In depth analysis of stool • Expansion of Caudovirales

bacteriophages • Validated in distinct cohorts • Household and non-household

controls • ? Bacteriophages key in altering

bacterial microbiota

Norman et al. Cell 2015 160(3):447-60

Challenges Clinical • phenotype • confounders

• age, gender, smoking • ethnicity, diet, surgery • medications

• “healthy” controls

Sampling • faeces v mucosa

• axial and longitudinal variation

• replication • multiple samples from same region • longitudinal sampling

Technical • 16S sequence • metagenomics • metatranscriptomics • metabonomics • … economics

Communication • clinicians • microbial ecologists • bioinformatics • statisticians

How can the microbiota be modified?

In inflammatory bowel diseases…

• Antibiotics – pouchitis; post-operative CD

• Probiotics – pouchitis, mild to moderate UC

• Prebiotics – no benefit

Algorithm management of pouchitis

Consider budesonide, immunomodulators, anti-TNF, alicaforsen

36% relapse

E. coli Nissle

34% relapse

Mesalazine 1.5 g

327 patients

E. Coli Nissle in remission maintenance of UC (Kruis et al. Gut 2004 53(11):1617-23)

Equivalence in maintaining remission

VSL#3 in mild/moderate UC (Sood et al. Clin Gastroenterol Hepatol 2009 Nov;7(11):1202-9)

Multicentre, double-blind, placebo-controlled trial

147 patients with mild to moderate UC

Given VSL#3 (3.6 x 1012 CFU/8 sachets/ twice a day) 12 weeks

Primary endpoint: decrease in UCDAI >50% at 6 wks

33% VSL#3; 10% placebo (p=0.001)

1 sachet contains 450 billion bacteria

Prebiotics and Crohn’s disease - 103 patients with active CD (CDAI>220) - randomised to FOS or placebo for 4 weeks

FOS is ineffective in active Crohn’s disease Is there a role in maintaining remission / preventing onset

in those with high risk phenotype?

Benjamin et al. Gut 2011 Jul; 60(7):923-9

What is faecal microbial transplantation (FMT)?

“Administration of faecal material containing gut microbiota from a healthy person (donor) to a patient with a disease or condition

related to dysbiosis or an alteration in their normal gut microbiota”

Faecal transplantation - history

Landy et al. (St Mark’s ) Aliment Pharmacol Ther 2011; 34: 409-15

4th century China - human “faecal suspension” for dysentery

16th century China - human “faecal suspension” known as “yellow soup”

17th century “Transfaunation” for animals unable to ruminate

“Coprophagia” – camel dung ingested by Bedouin

1958; first therapy in humans for pseudomembranous colitis

Publications on faecal transplantation

Faecal microbial transplantation

• Patients’ acceptability of FMT

• Trial data

• Clostridium difficile

• Inflammatory bowel diseases

• Key (unresolved) issues relating to FMT

• Microbiome in health/ disease state

• Method of FMT (how to give; donor; fresh v frozen stool; safety)

How acceptable is faecal transplantation for patients

• Refractory pouchitis – 2/3 of patients willing to consider FMT (St Mark’s)

• Refractory C. difficile

• 97% of patients who had FT willing to have further FT; 53% choose FT as first-line therapy1

• Survey with efficacy data given: 85% chose FT, 15% chose antibiotics2

• When aware of details of the therapies, only 4% changed choice from FT to antibiotics2

• Women and young people find FMT less appealing than men and older people

• Enema and colonoscopy more acceptable than NG route

• Hospital setting preferable to patients’ home

• 77% willing to pay for FMT

1Brandt et al. Am J Gastro 2012;107:1079-87 and 2Zipursky et al. Clin Infect Dis 2012:55:1652-8

Faecal transplantation

• Patients’ acceptability of FT

• Trial data

• Clostridium difficile

• Inflammatory bowel diseases (including pouchitis)

• Key (unresolved) issues relating to FT

• Microbiome in health/ disease state

• Method of FT (how to give; donor; fresh v frozen stool; safety)

Faecal transplantation in C. difficile

Recurrent CDI – ↓ phylogenetic richness and ↓Bacteroidetes/ Firmicutes1

Case series; meta-analyses; systematic reviews, RCT

Overall success rates of >91%2

1Chang et al. J Infect Dis 2008;197:435-8 and 2 Sofi et al. Scand J Gastro 2013;48 (3) 266-73

Faecal transplantation in C. difficile

Multi-centre long-term follow-up colonoscopic FMT for recurrent CDI1

cure rate of 98%

symptoms for ~ 1 year before FMT and 74% better in 3 days

Stool resembles donor stool in 2 weeks2,3

Persistence for over 30 days post FMT2,3

1 Brandt et al. Am J Gastro 2012;107:1079-87, Grehan et al. J Clin Gastroenterol 2010;44:551-61, Khoruts

et al. J Clin Gastroenterol 2010; 44:354-60

First randomised controlled trial

Aimed for 120 patients – stopped early

13/16 (81%) in FMT group – resolution of diarrhoea

4/13 (31%) in vancomycin group

After FT, ↑diversity (similar to healthy donors)

Van Nood et al. NEJM 2013;368:407-15

Faecal transplantation for C. difficile

Faecal transplantation in IBD

Damman et al. Am J Gastro. 2012; 107: 1451-9; Anderson et al. Aliment Pharm Ther 2012;36:503-16

41 patients with IBD (27 UC; 12 CD; 2 indeterminate) Majority (19/25) had reduction in symptoms & 15/24 remission

Resolution of C. diff infection in 15/15

Meta-analysis of 9 cohort studies, 8 case studies, 1 RCT

Overall 45% (54/119) achieved clinical remission

In cohort studies, 36% achieved clinical remission

Sub-group analysis, young patients 64% achieved clinical remission

Colman and Rubin J Crohn’s Colitis 2014 December 1;8(12):1569-81

Faecal transplantation in UC

RCT of 75 UC patients

38 patients faecal enemas; 37 patients

water enemas (50ml)

Weekly enemas for 6 weeks

24% (9/38) FMT achieved remission

versus 5% (2/37) placebo (p=0.03)

7/9 FMT patients histological healing

Donor dependence; immunosuppressant

use; early disease

1 year – 8/9 in remission (nil therapy)

1Moayyedi et al. Gastroenterology 2015; 149 (1): 102-9, 2 Noortje et al. Gastroenterology 2015; 149 (1): 110-18

RCT of 48 UC patients

Naso-duodenal FMT (healthy

donor) versus FMT (own stool)

Week 0 and 3

30.4% (7/23) FMT achieved

remission and endoscopic response

versus 20% (5/25) placebo (p=0.51)

Faecal transplantation in UC

Faecal transplantation

• Patients’ acceptability

• Trial data

• Clostridium difficile

• Inflammatory bowel diseases (including pouchitis)

• Key (unresolved) issues relating to FT

• Microbiome in health/ disease state

• Method of FT (how to give; donor; fresh v frozen stool; safety)

Methods of faecal transplantation donor screening

History Donor stool Donor serology Antibiotic use (3 months) C. Diff toxin Hep A IgM

Tattooes/ blood products (3 months) Ova, cysts, parasites Hep BsAg

High risk sexual behaviour (3 months) Giardia stool Ag Hep B core IgG/M

IBD H. Pylori stool Ag Ab Hep B sAg

IBS/ constipation Cryptosporidium Ag HIV 1&2

CRC/ polyps Isospora Syphilis

Immunocompromised Rotavirus Hep C Ab

Morbid obesity

Metabolic syndrome

Chronic fatigue

Atopy

Bakken et al. Clin Gastroenterol Hepatol 2011;9:1044-9

AABB Donor History Questionnaire Documents, available at http://www.fda.gov/BiologicsBloodVaccines/BloodBloodProducts/ApprovedProducts/LicensedProductsBLAs/BloodDonorScreening/ucm164185.htm

Methods of faecal transplantation related versus unrelated donors

related unrelated

Methods of faecal transplantation fresh versus frozen stool

Fresh used within 6-8 hours

Frozen stored -80oC (1-8 weeks), thawed 2-4 hours on ice bath

Hamilton et al. Am J Gastroent 2012;107:761-7

43 patients with recurrent C. diff

92% cure rate 90% cure rate

Methods of faecal transplantation mode of delivery

75% 25%

Better patient acceptance Entire colon infused Possibly more effective

Less acceptable for patients Entire GI tract exposed to FT Obviates need for endoscopy

Methods of faecal transplantation safety

>3 month follow up in multi-centre study of 77 patients with RCDI

Transient GI symptoms after FMT common

Autoimmune disease (e.g. RA, Sjogren’s, ITP)

FMT has been performed in patients on steroids, thiopurines, anti-TNFs and

immunocompromised patients (CLL, lymphoma, renal transplant)

2 recent studies in UC – patients developed fever and ↑ CRP

Unresolved Issues

Can microbiota be altered in IBD with FMT?

If so, does genetic pressure / indigenous bacteria affect ability to change?

Active “ingredient(s)”?

Donor - mixture of phylogenetically diverse bacteria – which?

Issues of mode of delivery, fresh versus frozen donor stool, frequency, dose

“Designer capsule” of selected micro-organisms

Selecting the “right” stage of disease to treat

Obesity

Obesity

Calorie extraction from food varies

depending on gut microbiota

“Obese microbiota” more effective at

extracting energy from food

Could manipulation of gut microbiota influence obesity?

Malnutrition

Calorie extraction from food varies

depending on gut microbiota

“Obese microbiota” more effective at

extracting energy from food

Could manipulation of gut microbiota influence malnutrition?

Obesity associated metabolic disorders and gut microbiota

• “Inflammation-associated” microbiome

• Lower potential for butyrate production

• Reduced bacterial diversity and/or gene richness

• FMT from lean donors to individuals with metabolic

syndrome increased their insulin sensitivity1

1. Vrieze et al. Gastroenterol. 2012;143:913-6

Liver disease and gut microbiota

• Liver receives 70% of blood supply from intestine

• Alterations of gut microbiota in NAFLD, alcoholic liver disease

and autoimmune liver diseases

• FMT could transfer NAFLD phenotype from mice with liver

steatosis to germ-free mice1

• Meta-analysis: “probiotics can reduce aminotransfrerases,

total cholesterol, and improve insulin resistance in NAFLD”2

1. Le Roy et al. GUT. 2013;62:1787-94; 2. Ma et al. World J Gastroenterol 2013;19:6911-18

Summary

Role of microbiota in health and disease

Modulation of gut microbiota as a therapeutic intervention

Mechanistic approach

Far reaching potential in GI (and non-GI) disease