La symbiose Homme-Microbiote dans la santé et la maladie

Joël Doré Micalis & MetaGenoPolis, INRA Jouy-en-Josas, France

Immune maturation

&

development of the microbiota

‘unique’ symbiosis : microbiota being recognized as a component of ‘self’

Maintained symbiosis : health and well-being

Disruption of tolerance : Risk of immune-mediated

disorders

Disruption of ecological balance :

Risk of infection

Homo sapiens ‘symbiosus’ ; a man-microbe mutualism that starts at birth

Diet, functional food, microbiota

Man-Microbe symbiosis

Re-evaluation using culture-independent

approaches

The dominant intestinal microbiota :

accessible using molecular approaches

21-32% Suau et al. 1999 21-37%

Hayashi et al. 2002

culturable

~ 30%

30% Tannock et al. 2000

Whole Genome Shotgun

sequencing

DNA

extraction

Assembly and annotation

Reference gene catalog

and gene counts Lessons from early human intestinal tract metagenomics: Reference gene catalogs, highlighting both core metagenome & rare genes

• 3.3 million genes Qin Nature 2010 to 10 million genes Li Nature Biotech 2014 Co-abundant gene clustering and metagenomic species Nielsen Nature Biotech 2014 Mouse gene catalog ; environment dependent Xiao Nat Biotech 2015

3 enterotypes / preferred ecological arrangements Arumugam Nature 2011 stability at SNPs level (strains) Schloissnig Nature 2012 nutrition and intestinal metabolome are linked Shoaie Cell Metabol 2015

=> metagenomic disbiosis ; diagnostic signatures ; predictive models.. • in T2D Qin Nature 2012, in Obesity Le Chatelier Nature 2013, in Liver cirrhosis Qin Nature 2014

=> low gene count is a key stratifier Cotillard Nature 2013

A technical revolution since the turn of the century

Metagenome: combined genomes of dominant microbes

Enterotypes may be regarded as preferred patterns in the ecological landscape of all possible arrangements

Data d

ensity (Fractio

n o

f data clo

se to a cen

tral po

int )

‘Density

plots’

For

~400

samples

Scheffer, Nature 2001

Prevotella

Bacteroides

Ruminococcus

Low to high

gene count

low high gene count

n=277

68 « species » significantly linked to gene count (richness/diversity)

nu

mb

er o

f in

div

idu

als

High richness Microbiota

Low richness Microbiota

Human microbiomes differ at the level of gene richness (diversity)

Impact of diet: Wu Science 2012

Low to high

gene count

Metagenomic view of the fecal microbiota

• Each dominant microbiota gathers on average over 500,000 genes representing 100’s of bacterial species (Qin et al Nature 2010) ; this is over 25 fold the size of the human genome for each individual.

• A reference catalog of some 10 million non-redundant genes.

• A small proportion of genes constitute a metagenomic core.

• Individuals are nonetheless different by genes, gene richness, metagenomic species and enterotypes (Arumugam et al. Nature 2011)

• The microbiota can be characterized by quantitative metagenomic profiling. (Cotillard et al. Nature 2013)

• Genomes of yet uncultured metagenomic species can be assembled (Nielsen et al. Nature Biotech 2014)

The human intestinal microbiota, a true organ…

100 trillion bacteria per individual microbiota

Interface between food and the epithelium

In contact with the 1st pool of immune cells and the 2nd pool of neural cells of the body

…with a major role

in health & disease !

yet ignored and neglected

barrier

Physio

defenses Microbiote intestinal

Nutrition

courtesy of Walter Wahly

1.3 MY

100’000 to 130’000 generations with fiber-rich diet (>60% of energy from fruits, veg, roots, nuts,..)

2 to 3 gen. with <10% fiber diet

‘Natural’ history of the genus Homo : we changed nutrition

with potentially major impact on man-microbe symbiosis

Altering mother-to-child vertical transfer of the microbiota,

Through several generations via: – Duration of gestation (preterm births)

– Mode of delivery (cesarean section >30% in Europe ; >80% in different places in the world)

– Hygiene of neonatal environment

– Exposure to antibiotics in mothers, neonates and infants

– Early life food and feeding mode (formula milk ; weaning diet)

– Maternal microbiomes

Extended hygiene hypothesis (Bach NEJM 2002)

“disappearing microbiota hypothesis” (Blaser EMBO-Report 2006)

& ... Missing Microbes (Blaser 2014)

‘Natural’ history of the genus Homo : we changed perinatal

management and environment

with potentially major impact on man-microbe symbiosis

HEALTH continuum DISEASE

Bach JF, N Eng J Med 2002

Chronic, immune-mediated diseases have

increased steadily for the past 60 years

Crohn’s disease Mangin 2004, Manichanh 2006, Sokol 2008, De Crutz 2012, Docktor 2012 Ulcerative colitis Lepage 2011 IBS Rajilic 2011, Carroll 2012, Durban 2012 Obesity Le Chatelier 2013; Cotillard 2013 Type-2 diabetes Burcelin 2011 Type-1 diabetes Giongo 2011 Coeliac disease Nadal et al., 2007; Collado et al., 2009 Allergy Abrahamsson 2012, Hanski 2012, Russel 2012 Autism Finegold et al., 2002; Paracho et al., 2005 AAD C.difficile Rea Mary 2012 Cystic fibrosis Han 2012, Del Campo 2014, Li 2014

Low species richness / low gene count

of the intestinal microbiota

is a feature of disbiosis associated with diseases

Low species richness and low

Faecalibacterium prausnitzii

on the microbiome side

Dysbiosis is an alteration of man-microbe symbiosis, with recurrent

features :

Altered gut-barrier permeability and

low grade inflammation

on the host side

( Indications from animal models, effects of antibiotics or probiotics, clinical studies; …

Microbiota Host Reduced microbiome richness Low grade inflammation - gene count - CRP, lymphocytes, calpro

Reduced levels of Firmicutes Increased gut permeability - butyrate producers - blood LPS, zonulin,..

- Faecalibacterium - fecal zonulin, tissue LBP, …

Increased levels of pathobionts - proteobacteria, Gram negative

Increased Bacteroides to

Ruminococcus ratio of

enterotypes

Features of dysbiosis

as altered man-microbe symbiosis:

Microbiome gene count as stratifier

Le Chatelier et al, Nature 2013

Low to High

gene count (French or Danes) Known species n=10

Unknown species n=48

signature species (n= 58)

ROC analysis

4 species

LGC HGC

Low gene count (low bacterial richness) individuals have less healthy metabolic & inflammatory traits:

Microbiota gene count / diversity is a health-associated stratifier

Increased adiposity, insulin resistance, dyslipidaemia, inflammation, that predispose to type 2 diabetes, cardio-vascular disease, cancer

Dietary habits

impact gene richness 3 dietary patterns are determined

based on food frequency records,

using k-means for 26 food categories.

Cluster 2

Cluster 3 Cluster 1

Karine Clément, head of ICAN, in partnership with Danone Research

‘Healthyness’

Canonical discriminant analysis Stratified Kruskal-Wallis test

among the 3 dietary clusters. Columns show the age-adusted

mean value of the parameters in cluster 1, cluster 2 and cluster 3,

respectively. # tendency p0.1

‘Healthier’ dietary habits are associated with a higher gene diversity

and a higher microbiota stability upon changes in diet

Kong et al, PLOS-One 2014

yoghurt

water

vegetables

fruits

soups

Potatoes including chips

sweets

Sweetened soft drinks

‘richness in fibers’

- +

Tap et al, EM 2015

Determined in early childhood, species richness differentiates human population-groups

Yatsunenko, ..Gordon, Nature 2012

Species richness

Spec

ies

rich

nes

s

Impact, over generations, of :

• Birth ‘management’ ? • Nutritional transition? • Repeated antibiotherapy? • …

… are we altering human biology?

Spp richness establishes in early childhood, ... by vertical transfer across generations?

intervention stabilization

intervention 1200-1500 Kcal

A low gene count microbiome predicts a lesser response; especially for obesity-associated adiposity, low grade inflammation and insulin resistance

LGC HGC

: high protein, low fat and low glycemic index carb with high/diverse fiber content

Cotillard et al, Nature 2013

Low gene-count in obesity:

predicts poor responders to a calory-restricted diet

+30%

Low gene count

High gene count

A high diverse fiber diet can correct low gene-count

in overweight and obesity

Low

High

Low

High

6 weeks intervention

6 weeks stabilization

Hypothesis: a large diversity in primary substrates (plant fibers) may promote diversification throughout the microbial food chain & improve gene richness: a new paradigm for personalized preventive nutrition / medicine?

intervention : high protein, low fat and low glycemic index carb with a highly diverse fiber content

+25%

Cotillard et al, Nature 2013

Sustained alteration

of the Gut Microbiota

Sustained low grade to

overt inflammation Dysbiosis

stressor

stressor

Homo sapiens ‘dysbioticus’

Critical transition* in chronic immune diseases (concept)

Eubiosis

of the gut microbiota

Physiological

Immune tone symbiosis

Reversible imbalance

of the gut microbiota crosstalk

genetic predisposition, infection, diet lifestyle & environmental triggers

Transient low grade

inflammation

Homo sapiens ‘modernicus’

Homo sapiens ‘symbiosus’

* Scheffer 2001, Kefi 2014

Microbiota Host

Today’s medicine addresses symptoms

Sustained alteration

of the Gut Microbiota

Sustained low grade to

overt inflammation Dysbiosis

The new paradigm : a holistic approach to intestinal health

Eubiosis

of the gut microbiota

Physiological

Immune tone symbiosis

Reversible imbalance

of the gut microbiota crosstalk

genetic predisposition, infection, diet lifestyle & environmental triggers

Transient low grade

inflammation

Homo sapiens ‘modernicus’

Homo sapiens ‘symbiosus’

* Scheffer 2001, Kefi 2014

Microbiota Host

Holistic approaches : Microbiome-host targeted

strategies

Holistic approaches : microbiome-host

targeted strategies

From Homo dysbioticus back to Homo symbioticus Restoring Symbiosis

FMT Autologous solutions:

• preventive

and

• curative microbiota-host

sustained dysbiosis Homo sapiens ‘dysbioticus’

microbiota-host symbiosis

microbiota-host altered crosstalk

Homo sapiens ‘modernicus’

Homo sapiens ‘symbioticus’

Same triggers and surgery, chemotherapy, radiotherapy,

antibiotics

Restoration by: • functional foods • nutrition, incl. fibers

Restoration by: • microbiotherapy / FMT

genetics, infections, diet lifestyle, environmental

triggers

Take home messages : • Humans share a core microbiome and yet they differ by genes, species, enterotypes (ecology) and gene count (microbiota diversity).

• microbiome gene count is a key stratifier in several immune disorders including major diseases of modern world, that have increased in incidence since 1950’s

• dysbiosis is an altered state of microbe-host symbiosis with auto-aggravating signals from both sides

• microbiota modulation should be considered as a target for personalized nutrition and as a reinforcement/adjuvant strategy in current therapy

• nutrition - fibers and live microbes - may be strategic bioactives for the maintenance, preservation or restoration of man-microbe symbiosis

INRA Jouy-en-Josas Christel Béra-Maillet Hervé Blottière Marion Leclerc Patricia Lepage Catherine Juste Nicolas Lapaque Tomas de Wouters Antonella Cultrone Malgorsata Nepelska Elsa Jacouton ChenHong Zhang Julien Tap Stanislas Mondot Omar Lakhdari

European Community & ANR-France

Philippe Seksik Harry Sokol Philippe Marteau

S Dusko Ehrlich, Jean Weissenbach (Genoscope, Evry), Wang Jun (BGI, Shenzhen), Peer Bork (EMBL Heidelberg), Francisco Guarner (Val d’Hebron Hospital Barcelona), Oluf Pedersen (SDC Copenhagen), Maria Rescigno (IEO Milan), Liping Zhao (Shanghai JiaoTong University), Jim Versalovic (Baylor College of Medicine, Houston), Baghi Singh (Western Ontario, London) and EU-MetaHIT and IHMS Consortia

Karine Clément (INSERM U972, CR des Cordeliers), Denis Le Paslier & Eric Pelletier, (CEA-Genoscope), Liping Zhao (Shanghai JiaoTong University) and ANR MicroObese consortium

Philippe Langella and col. Bruno Pot Corinne grangette and col.

Micro Obes

Merci

de votre attention

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