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© Confidential | 05 Feb 2014| Préparation Rdv DR-IP| S Thompson
GUT MICROBIOTA AND PROBIOTICS IN HUMAN
MURIEL DERRIEN, PHD
GUT MICROBIOTAGENERAL CONCEPTS01
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© Confidential | 00 mois 2012 | Nom de la présentation | Auteur2
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
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
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 »
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
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
QUESTIONS
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13 © Yakult 2015 | M.DERRIEN
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?
DANONE PROBIOTICS AND GUTMICROBIOTA03
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© Confidential | 00 mois 2012 | Nom de la présentation | Auteur14
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
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
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
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
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
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
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 !
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