The human microbiome response to pre/probiotics. Discussion group 6

Organisers: Karen Scott and Glenn Gibson

Rapporteur: Paul Sheridan

Workshop timetable

DISCUSSION TOPICS 'effect of probiotics on indigenous microbiota' led by Maria Marco addressing the question what constitutes a probiotic effect? 'effect of prebiotics on indigenous microbiota' addressing the question what constitutes a prebiotic effect? led by Lindsay Hall How do we measure ‘the effect’? led by Colin Hill How relevant are samples - faeces vs gut. What biomarkers could we use? (or symptoms in patients) led by Paul O’Toole How have culture independent methods helped? What is the relative importance of contributions from different omics techniques? What is preferred? led by Julian Marchesi and Anne Salonen Role of metabolic studies led by Rodney Dietert Which populations should be targeted with pro/prebiotics? led by Omry Koren, Yuan-Kun Lee Summary discussion – finalise answers to questions

Probiotic effects on the microbiota depends

on:

Gastrointestinal

pathogens • Antimicrobials

• Competition for

nutrients and

binding sites

Age:

Infancy

Adulthood

Elderly

Diet

Geography

Inflammatory

bowel disease

Antibiotic-

associated diarrhea

Obesity

• Nutrient utilization

• Vitamin & EPS production

• SCFAs (butyrate)

• Mucin production

• Host antimicrobials

NEC

• Do probiotics affect microbiota composition? – YES and NO!

Depends on analysis method

composition vs activity

strains used

dose

diet

target population

Target site

BUT probiotic can have an effect on health independent of the microbiota

Prebiotics and the microbiota

• Increase in microbiota growth and fermentation – Increased bifidobacteria and lactobacillus – Increased SCFAs and impact on

• immune responses e.g. cytokines

• Obesity – Reduction in chronic low-grade inflammation link – Reduction in hypercholesterolemia – Impact on appetite?

• C. difficile infection – Reduced recurrence rate

• Allergy – Atopic dermatitis – Eczema

• Do prebiotics affect microbiota composition? – YES

Depends on analysis method

composition vs activity

types used

dose

diet

target population

target site

BUT prebiotic effect on health can be bigger than any detectable effect on the microbiota

How well do faecal samples reflect the in situ gut microbiota?

• They don’t!

• But can be used as a surrogate marker

accessible

analysable

comparative data available

Different microbiota of mucosae and stool

Bacteroidetes Firmicutes

Eckburg et al, 2005. Science 308: 1635-8

Lb. murinus Lb. murinus Lb. salivarius (Bac+)

Lb. pentosus Pd. pentosaceus

Control Probiotic

Clinical scores

Salmonella/g faeces (day 15) 107

106

105

104

103

***

Casey et al. 2007 Appl. Environ. Microbiol. 73:1858-1863 Walsh et al. 2008 FEMS Microbiol. Ecol. 64:317-327

Ileal microbiota (N=9; day 28)

Faecal microbiota (N=9; day 28)

15

20

25 ***

Weight gain/kg (day

15)

Control Probiotic

10

20

30

40

*** 25

50

75

100

***

Control Probiotic

Control Probiotic

% Diarrhoea (day 4)

Five-live probiotic challenge in pigs

How best do we measure intestinal health or disease?

• Focus on microbiota changes

OR

Focus on human study outcomes

“Best” case scenario:

• Microbiota alterations cause or modulate risk or severity of various diseases…

• Some diseases have been associated with altered microbiota

De Vos & De Vos, 2012. Nutrition Reviews 70 (S. ):S45–S56

Worst case follow-on….a thought exercise

• Microbiota modulation by pro- or prebiotics cannot be detected in stool

• Need surrogate/replacement biomarkers (predictor of later disease development) • Could be simply improvement of clinical markers… • Would this justify a pro/prebiotic-based health

claim? Condition Microbiota change Potential probiotic Potential surrogate Comment

Cdiff Diversity restoration Mixture? BAs FMT >90% effective

IBD Reduced diversity, Fprau Fprau, Clostridia Faecal calprotectin Most analysis of dysbiosis is on fecal

samples anyway Can inflammation, once triggered, be

stopped?

IBS Increased Firmicutes Bacteroidetes Bacteriocin producer

Rome III Conflicting signatures of dysbiosis

Obesity Increased Firmicutes Akkermansia Glucose metab BMI

Conflicting signatures of dysbiosis

T2D Clostridium clostridioforme ↑ Roseburia↓

Not clear Glucose metab Conflicting signatures of dysbiosis

How will omics techniques improved our understanding of the microbiome and what will be the contribution of the

individual omics techniques?

Which microbes are stimulated

and which repressed?

What functions are affected?

What is the effect on genetic potential?

Community-wide microbiome response to pre/probiotics

Zoetendal EG, M Rajilić-Stojanović & WM de Vos (2008) High throughput diversity and functionality

analysis of the gastrointestinal tract microbiota. Gut 57 : 1605-15

16S rRNA gene

Before vs. After supplementation

O-PLS-DA cross-validation scores plots and coefficient plots of urinary and faecal nuclear magnetic resonance spectral data obtained from sham control (blue) and Roux-en-Y gastric bypass (RYGB)-operated rats (red) at week 8,

Marrying together “omic” datasets

X Y

Correlation matrix

ob

serv

atio

ns

ob

serv

atio

ns

NMR/MS OTUs

Pearson’s correlation

OTUs

NM

R/M

S

Metabonomics Metataxonomics

Matches change in composition

with change in activity of the microbiota

Which populations (human) should be treated with pro/prebiotics?

Which populations (bacterial) should be treated with pro/prebiotics?

Target populations

Changes in the gut microbiome through life

Neuman and Koren, In press

• Akkermansia muciniphila (IBD versus CRC)

• Faecalibacterium praustnizii (Low in CD adults)

• Ruminococcus bromii – resistant starch degrader

• Roseburia/Eubacterium rectale group – butyrate producers

• Oxalobacter formigenes - oxalate degrader

Summary discussion

• Probiotics and prebiotics can affect the commensal microbiota – with caveats as discussed

• Faecal samples are an acceptable surrogate for changes in the large intestine

• Microbiota changes are only one marker

need to consider alternatives

• Alternative ‘omics tie together activity and composition

• Different populations require different approaches – infants, teenage, pregnant disease, elderly

Participants

Participants Karen Scott and Glenn Gibson

• Paul Sheridan

• Julian Marchesi

• Lindsay Hall

• Omry Koren

• Anne Salonen

• Yuan-Kun Lee

• Paul O’Toole

• Maria Marco

• Colin Hill

• Rodney Dietert

• Gun-Britt Fransson

• JoMay Chow

• Valerie Benoit

• Lori Lathrop Stern

• Marie-Emmanuelle Le Guern

• Sylvie Binda

• Koji Nomoto

• Benedicte Flambard

• Juliet Ansell