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00 mois 0000 1 © Confidential | 05 Feb 2014| Préparation Rdv DR-IP| S Thompson GUT MICROBIOTA AND PROBIOTICS IN HUMAN MURIEL DERRIEN, PHD

M derrien yakult vf

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© Confidential | 05 Feb 2014| Préparation Rdv DR-IP| S Thompson

GUT MICROBIOTA AND PROBIOTICS IN HUMAN

MURIEL DERRIEN, PHD

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GUT MICROBIOTAGENERAL CONCEPTS01

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© Confidential | 00 mois 2012 | Nom de la présentation | Auteur2

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GUT MICROBIOTA – A RICH ECOSYSTEM

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3 © Yakult 2015 | M.DERRIEN

http://www.nature.com/nrmicro/journal/v7/n2/box/nrmicro2052_BX2.html

Bacteria Archaea

Virus-prophages Eukarya

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BACTERIA IN NUMBERS

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http://protomag.com

Human is a reservoir of bacteria !

90% of our body is microbial origin (Savage, 1977)

Genome of microbiota (= microbiome) is 100> human genome

© Yakult 2015 | M.DERRIEN

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BACTERIA ALONG THE GI TRACT

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Kovatcheva-Datchary et al, Prokaryotes

© Yakult 2015 | M.DERRIEN

Microbiota diversity and number

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BALANCED VERSUS UNBALANCED MICROBIOTA

© Yakult 2015 | M.DERRIEN

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HOW TO MODULATE MICROBIOTA FOR HEALTH?

2018

DiseaseDisorder

Crohn’s disease

Colorectal cancerDiabetes

Obesity

Allergies

Ulcerative colitisIrritable bowel

syndrome

C. difficile infection

Health

© Yakult 2015 | M.DERRIEN

Nutrition Pharmaceutics

Healthy diet

Probiotics

Commensals

Fecal TransplantSynthetic communitiesPrebiotics F. prausnitzii

« No specificsymptom »

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PROBIOTICS AND GUT MICROBIOTA0200 mois 0000

© Confidential | 00 mois 2012 | Nom de la présentation | Auteur8

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PROBIOTIC “TRAVEL” WITHIN THE INTESTINAL MICROBIOTA

http://www.mynaturall.com/intestinal-tract

© Yakult 2015 | M.DERRIEN

Matrix

Dose (1010CFU/day)

Oral

Stomach

pH–gastric tolerance

102-3 CFU / low diversity

pH 2-3 – fast transit –O2

Colon

SubdominantpH advantage (proximal)

High number (1012) / diversity

pH 5 -7 / indigestible substrates/ low transit –Low O2

Small intestine

Mid number 106-8 CFU/ diversity

Dominant – Bile tolerant

pH 6-7. Major site immune system – Simple sugars-

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© Confidential | 05 Feb 2014| Préparation Rdv DR-IP| S Thompson10

POTENTIAL MODES OF ACTION OF PROBIOTICS ON GUTMICROBIOTA

Derrien and van Hylckama Vlieg, Trends Microbiol, In press

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TOOLS TO ANALYSE REPONSE OF MICROBIOTA TO PROBIOTICS

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Metagenomics (genetic potential of microbiota)

© Yakult 2015 | M.DERRIEN

Cultivation approaches~ 20% bacteria cultivable« great plate count anomaly »

Metatranscriptomics/metaproteomics/metabolomics active bacteria

Years

Exhaustivity

/ activity

16S rRNA (microbiota composition, diversity)

Overall structure

Dominant / targeted

1980 2000

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Tested strain Cohort intake Observed results

L. rhamnosus GG

Healthy adults (Mb,Fc)

Test: n= 9

Control: n= 13

3 weeks

MPe

1010 CFU

No significant impact of gut microbiota

composition and stability

L. paracasei Zhang

Healthy subjects (M,F)

Test: n=24

28 days

chewable tablets

10.6 log10 CFU

Difference in the composition and diversity of

intestinal microbiota compared to baseline

L. paracasei DG

Healthy subjects (M, F)

Group A: n=14

Group B: n=16

4 weeks

Capsule

2.4 x 1010 CFU

↑ Proteobacteria, Coprococcus

↓ Blautia

6 commercially available

probiotics

18 healthy adults (M,F)

6 groups (3 ind/group)

8 weeks

MP

Between 108 and 1010

No significant changes in the overall structure

of gut microbiota

Changes in some OTUs

VSL#3f

IBS subjects (M,F)

Test: n=10

4 weeks

Lyophilized

1.8 x1012 bacteria

↓ Bacteroides

B. longum Bar33 + L.

helveticus Bar13

32 subjects (M,F)

Test: n= 16

Placebo: n= 16

1 month

In biscuit ( lyophilized)

109 CFU of each

No ↑ Clostridium cluster XI, Clostridium

difficile, Clostridium perfringens, Enterococcus

faecium, Campylobacter

LGG, L. rhamnosus, P.

freudenreichii B.animalis

ssp. lactis Bb12

IBS subjects (M,F)

Test :n =12

Control : n= 8

5 months

MP

1.2 ×109 CFU ⁄ d (each)

Stabilization of the microbiota

WHAT DO WE KNOW SO FAR FROM CLINICAL STUDIES IN ADULTS ?

© Yakult 2015 | M.DERRIEN

-> No major perturbation after probiotics when all individuals taken into account-> Some genera modulated-> Small cohorts

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QUESTIONS

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Most probiotics used were not originally selected for their ability of targeted modulation of the microbiome -> How do they impact gut microbiota

Is there a need for “personalized” probiotics taking into account the variability of the microbiome across the human population?

What level of microbiota modulation is expected and necessary to impact host health?

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DANONE PROBIOTICS AND GUTMICROBIOTA03

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© Confidential | 00 mois 2012 | Nom de la présentation | Auteur14

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IMPACT OF PROBIOTICS IN HEALTHY INDIVIDUALS

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1. Healthy women (16S rRNA analysis)

2. Healthy twin women (metatransciptomics + metabolomics)

3. Next: stragegy of stratification of gut microbiota

© Yakult 2015 | M.DERRIEN

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IMPACT OF A FERMENTED MILK PRODUCT (FMP) ON HEALTHY INDIVIDUALS

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Fermented milk product

© Yakult 2015 | M.DERRIEN

B. animalis subsp lactis CNCM I-2494

Lc. lactis subsp lactis CNCM I-1631

L. delbrueckii subsp bulgaricus (2)

S. thermophilus CNCM I-1630

µ

Stool sample

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4 WEEKS CONSUMPTION OF FMP IN HEALTHY WOMEN

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Tillisch et al, 2013, Gastroenterology

FMP

No product

Control (acidified milk)

V3 V5

45 healthy women

4 weeksStool collection

© Yakult 2015 | M.DERRIEN

Stool DNA 16S rRNASequencing

Analysis

Multivariate analysis Abundance of taxa Diversity of samples

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FECAL MICROBIOTA ANALYSIS (16S SEQUENCING)

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Post hoc analysis Tillisch et al 2011 Gastroenterology

No major perturbation of gut mcirobiota Inter-individuality microbiota higher than probiotic impact Only trend detected

No significant impact of FMP on dominant fecal microbiota (454 16S)

© Yakult 2015 | M.DERRIEN

Trend to increase Faecalibacteriumin FMP Group

Lo

g 2

ra

tio

F.p

ra

us

nit

zii

Acti

ve

Co

ntr

ol

No

P

-6

-4

-2

0

2

4

6

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EVOLUTION OF TECHNIQUES FOR GUT MICROBIOTAANALYSIS

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Metagenomics (genetic potential of microbiota)

© Yakult 2015 | M.DERRIEN

Cultivation approaches~ 20% bacteria cultivable« great plate count anomaly »

Metatranscriptomics/metaproteomics/metabolomics active bacteria

16S rRNA (microbiota composition, diversity)

Years

Exhaustivity

/ activity

Overall structure

Dominant / targeted

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7 pairs of healthywomen twins

Stool collection

FMP Consumption

7 weeks (twice a day)

Preconsumption

4 weeks

Postconsumption

4 weeks

16S rRNA (454, 2000 reads / sample)

Microbiome (Shotgun, 50 000 reads/ sample)

RNA seq (8-10 M reads / sample)

McNulty et al, 2011, Science Translational Medicine

TARGETED CLINICAL TRIAL FOR MICROBIOTA ANALYSIS

© Yakult 2015 | M.DERRIEN

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Adapted from McNulty et al, 2011, Science Translational Medicine

TRANSIENT COLONIZATION OF INGESTED STRAINS

❶ Transient colonization and elimination few days after cessation of

consumption in healthy adults

❷ Some individuals retain it longer than others

Re

lati

ve

ab

un

da

nc

e (

%)

Pre 1 2 4 7 Post0.0

0.1

0.2

0.3

B. animalis subsp lactis CNCM I- 2494 L. lactis CNCM I-1631

© Yakult 2015 | M.DERRIEN

Pre Post

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McNulty et al, 2011, Science Translational Medicine

FMP IMPACTS ACTIVITY OF CARBOHYDRATE METABOLISM BY THE COMMENSAL MICROBIOTA

Cellobiophosphoryla

se

Pectinesterase Levanase

pre FMP FMP post pre FMP FMP post pre FMP FMP post

Transient colonization

correlates to transcriptional

changes in the fecal gut

microbiota”

Human fecal metatranscriptome exhibited significant changes, including those

related to plant polysaccharide metabolism and SCFA production

-> expansion of the carbohydrate metabolizing capability of gut microbiota

(Bacteroides , Eub. rectale, Rum. bromii)

Starch and sucrose metabolism

© Yakult 2015 | M.DERRIEN

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FMP INCREASES SCFA IN VITRO

© Yakult 2015 | M.DERRIEN

Dialysis

Dialysis

Ascending Transverse Descending

Veiga et al, 2014 Sci reports

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NEXT : CAN WE IDENTIFY RESPONDERS OF PROBIOTICMODULATION?

Rationale: Microbiota degree of response to diet has been shown to depend on microbiota structure at baseline (Salonen et al, ISME J 2014)

© Yakult 2015 | M.DERRIEN

Variability of extent of response of microbiota to FMP

Microbiota response to probiotics

Stable microbiota = Non responders

Non responders have higher microbiota diversity at baseline

Microbiota response to diet

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CONCLUSIONS AND FUTURE DIRECTIONS

Impact of probiotics in human

❶ No major change in the composition of dominant fecal microbiota

-> Strategy of stratification of baseline microbiota (enterotypes, richness, …),

larger cohorts

© Yakult 2015 | M.DERRIEN

❷ When use of dedicated models -> Impact on activity of resident microbiota

(butyrate producers…)

❸ Stool analysis probably understimates our peception of activity of probiotics

on GI microbiota (-> ileum, proximal colon)

❹ Need to specifically screen probiotics that modulate gut microbiota

❺ End game :need to understand which level of microbiota modulation is

beneficial for host

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THANKS FOR YOUR ATTENTION !

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MODE OF ACTION OF PROBIOTICS ON GUT MICROBIOTA

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27 © IPC 2014 | M.DERRIEN Derrien and van Hylckama Vlieg, Trends Microbiol, In press