Evaluating human gut microbiota and microbe-host phenotype relationships

Jeremiah FaithIcahn School of Medicine at Mount Sinai

Immunology Institute

Institute for Genomics and Multiscale Biology

2014 Advances in Inflammatory Bowel Diseases

Orlando, FL

Dec 6, 2014

Associations between our microbes and intestinal disease

• Crohn’s disease • Ulcerative colitis• Pouchitis • Necrotizing enterocolitis • Coeliac disease • Diarrhea (C. diff)• Colorectal cancer

What must be done1. Verify causation2. Identify microbial effector strains3. Modify effector strains to improve health

Identifying human microbial effector strains

in mice

Germ-free and gnotobiotic mice

• Germ-free: devoid of microbial life• Gnotobiotic: “known life”, colonized with defined collections of microbes• Humanized microbiota: gnotobiotic animals harboring a human community

Effector strain identification successes

• B. vulgatus accelerates colitis in gnotobiotic guinea pigs [A. Onderdonk et.al., 1980s]

• IL10-/- mice; HLA-B27 rats [B. Sartor 1990s-now]• B. fragilis matures immune system [Mazmanian, et.al., Cell 2005]• SFB increase Th17 in small intestine [Ivanov, et.al., Cell 2009]• K. pneumoniae and P. mirabilis elicit colitis in T-bet-/- x Rag2-/- [Garrett et. al.,

Cell Host Microbe 2010]• Clostridia increase Treg in colonic lamina propria [Atarashi, et.al., Science 2011]• E. coli NC101 modulates colorectal cancer (AOM/IL10) [Arthur, et.al., Science

2012]

We are getting better at this. But can we be more exhaustive

A systematic pipeline for identifying human effector strains in mice

Screening “humanized” mice

Which bacteria are causing this?!?

Faith, et.al., Sci Transl Med 2014

Diverse donors

Gnotobiotic miceHumanized with the

microbiota of a unique donor

%Fo

xP3+

am

ong

CD4+

Tce

lls

Donor1

Donor2

Donor3

Donor4

Donor5

Germ-fr

ee

High-throughput anaerobic bacterial isolation

Anaerobic isolation Protein extraction Identification(MALDI-TOF)

Each well contains a unique bacterial strain

Donor Goodman, et.al., PNAS 2011Faith, et.al., Science 2013

Large scale bacterial isolation

Each well contains a unique bacterial strain

(17 total microbes) Donor

Fractionate community

Gnotobiotic mice

Phenotype screen

Phenotype response

Community size

% T

regs

Serin

e

Adip

osity

Identifying effector strains%

FoxP

3+ a

mon

g CD

4+ T

cel

ls

Which bacteria are causing this?!?

%Fo

xP3+

am

ong

CD4+

Tce

lls

Donor1

Donor2

Donor3

Donor4

Donor5

Germ-fr

ee

Faith, et.al, Sci Transl Med 2014

Next steps for understanding host/microbe interactions in IBD

Diverse donors

Gnotobiotic miceHumanized with the

microbiota of a unique donor

UC, Crohn’s, Pouchitis, PSC

DSS, IL10, T-cell transfer

Next steps• Contributions of host/genotype?• Diversity of effector strains• Effector strains consistent across mouse colitis models?

Identifying human microbial effector strains

in humans!

Identifying effector strains

Hypothesis: members of human gut microbiota modulate complex disease risk

Microbial inheritance in complex disease1. The “strain” is the key unit of measure

a. acquired during the first three years of life.b. stable inhabitants for decades.c. decades to manifest clinical symptoms

Microbial inheritance in complex disease

Microbiome’s Future: optimal colonization at birth

H. pylori

cagA- cagA+

1. The “strain” is the key unit of measurea. acquired during the first three years of life.b. stable inhabitants for decades.c. decades to manifest clinical symptoms

Microbial inheritance in complex disease

Disease Agent Time to manifest clinical symptoms

Leprosy Mycobacterium leprae 10-30 years; (13% of household contacts are carriers; Araujo et.al., mem Inst Oswaldo Cruz 2012)

Whipple’s disease Tropheryma whipplei ~10 years (many asymptomatic carriers)

Peptic ulcer Helicobacter pylori ~10 years (few infected progress)

Gastric cancer Helicobacter pylori >50 years (few infected progress)

1. The “strain” is the key unit of measurea. acquired during the first three years of life.b. stable inhabitants for decades.c. decades to manifest clinical symptoms

Evidence of microbial role:MZ < 40% concordantGWAS loci involved in immune regulation, microbial recognition/defenseAntibiotics somewhat effectiveGerm-free models have no or significantly less pathology than colonized

Hypothesized:Crohn’s diseaseRheumatoid arthritisMultiple sclerosis

Treatment: slow the progressive pathology (like Pre-1982 peptic ulcer)

Microbial inheritance in complex disease: a simple model

• P(transmissioni) is the probability of transmission of strain i• P(accessi) is the probability of access to strain i• P(resistancei) is the resistance of the host to the colonization by strain i

Microbial inheritance in complex disease: consequences

• Family members share microbes!

High

Microbial diseases can “look” genetic

Majority of family members share strains

Faith, et.al., Science 2013

Microbial inheritance in complex disease: consequences

• Family members share microbes!• Sibling microbiotas should be more similar than

parents

Turnbaugh et al, Nature 2009

*p<10-5; **p<10-14; ***p<10-41

Schloss et al, Microbiome 2014

LowHigh

Microbial inheritance in complex disease: consequences

• Family members share microbes!• Sibling microbiotas should be more similar than

parents• Increase in disease risk in siblings, especially

sequential

1. Gastric Cancer risk 2x higher in sibships of 7 individuals compared to 1-3 [Blaser et.al., PLoS Med 2007] 2. Crohn’s disease risk higher in consecutive births relative to non-consecutive births [Hugot et.al., Eur J

Hum Genet 2003]

LowHigh

Microbial inheritance in complex disease: consequences

• Family members share microbes!• Sibling microbiotas should be more similar than

parents• Increase in disease risk in siblings, especially

sequential• Very hard for cohabiting adults to pass microbes

• Disease takes years to manifest• P(resistance) is HIGH

• Unrelated individuals do not!

Low

Microbial inheritance in complex disease1. The “strain” is the key unit of measure

a. acquired during the first three years of life.b. stable inhabitants for decades.c. decades to manifest clinical symptoms

We can use methods similar to human genetics!

gene

Microbial inheritance in complex disease:using the rules to identify effector strains

We can use methods similar to human genetics!

Case-control association: difficulty unrelated individuals do not share microbial strains

Crohn’s No Crohn’s

Microbial inheritance in complex disease:using the rules to identify effector strains

We can use methods similar to human genetics!

Familial association: difficulty powering study

Microbial inheritance in complex disease:moving forwards

Need methods to microbe-type at the strain-level

Thanks to

Faith laboratory

Eduardo ContijochSean LlewellynIlaria Mogno Cinkia FerminZhihua LiRuby NgEddie Vazquez

Immunology InstituteInstitute for Genomics and Multiscale BiologyIcahn School of Medicine atMount Sinai

Jeffrey Gordon, Philip Ahern, Andrew Goodman, Jean-Frederic Colombel

IBD Collaborators

Jose ClementeSergio LiraMiriam MeradJudy ChoInga PeterAri Grinspan

Funding

NIH GM108505SUCCESS