HIGH THROUGHPUT DISCOVERY OF

MICROBIAL HOST REGULATORY

CIRCUITS USING C. ELEGANS

BUCK S. SAMUEL
Assistant Professor
Alkek Center for Metagenomics and Microbiome Research
Baylor College of Medicine
bucks@bcm.edu | @microbeMinded

You have evolved from worm

to man, but much within you is
still worm. -- Nietzsche

[Scanning Electron Micrograph of Feces] National Geographic

Microbial changes correlate with many

diseases cause or effect?
Healthy

Diseased

Morrison & Baxter 2012

Selected contributions to science of C. elegans

Genetic control of behavior
First cloning and sequencing of a myosin gene
First germline stem cell niche identified
First complete metazoan cell lineage
Discovery of apoptosis (cell death) genes
Identification of heterochronic genes (developmental timing)
First complete wiring diagram of a nervous system
Discovery of the first axon guidance genes
Genetic control of lifespan by insulin signaling
First microRNA (lin-4) and its mRNA target (lin-14) described
Identification of nonsense-mediated decay genes
Introduction of GFP as a biological marker
First demonstration of specific olfactory receptor/ligand pair
First metazoan genome sequenced
Discovery of RNA interference (RNAi)
Conservation and ubiquity of miRNAs
First full genome-wide profiling of gene function via RNAi
Transgenerational inheritance and its mediation by piRNA
Identification of surveillance immunity

By leveraging the power of C. elegans we can rapidly identify

new pathways that control host microbe interactions

Outline
C. elegans as a high-throughput gnotobiotic system

Influence of natural microbiome on C. elegans

Potential for application to human systems

The C. elegans gnotobiotics tool box

Facile gnotobiotics
[every experiment starts germ-free]

Robust high-throughput assays

[metabolism, lifespan, stress, immunity, etc.]

Defined natural microbiome

C. elegans, a host primed for gnotobiotics

*** NO ANIMAL PROTOCOL REQUIRED ***

Short lifespan (~15days; most assays <3days)

Small (1mm) size 384-well plate screening

Defined cell lineage and plan (959 cells)

Transparent live imaging of host-microbe

interactions in real time

20,000 microbiomes ~ 3-6 months work

Host-microbe interactions in C. elegans within the

intestine

Used to explore responses to

human pathogens for decades

Conserved innate epithelial immune

pathways [e.g., p38 MAPK, TGF-]

Endocrine regulation of metabolism

[insulin, neuropeptides, etc.]

Similar intestinal niches and

pressures for colonization [low O2, 35 pH, mucins, antimicrobials, etc.]

Intestinal functions comparable

Adapted from Balla et al. 2013

Addressing the challenges ahead: new

tools for gnotobiotics in C. elegans

Intestinal lumen (and its microbiome) are forgotten organs

Functional assays of digestive and intestinal function

Live-imaging of microbiome function

Platform for systems biology of both host and microbe

Kaleta et al., 2016

Szigeti et al., 2014

Yilmaz et al., 2016

Outline
C. elegans as a high-throughput gnotobiotic system

Influence of natural microbiome on C. elegans

Potential for application to human systems

Not a soil nematode

more like a fruit
worm

PROLIFERATING
adult

L4

apple

L1

petasites

apple

orange

optunia

grapes

apple

bryony

slug

snail

NON-PROLIFERATING PROLIFERATING

L2d

apple

dauers
snail

L2
L3
Apple Orchard, Orsay, France

various invertebrates

[invertebrate dispersal]

embryos
dauer

hawthorn

C. elegans retains a microbiome in the wild

Marie-Anne Felix

Simple (5-15 strains)

~10,000 CFUs / animal

Highest colonization in
early adulthood

Gut microbiome composition differs from its habitats

Meta analysis of Samuel et al., PNAS (2016); Dirksen et al., BMC Biol (2016); Berg et al., AEM (2016)

Composition of the C. elegans core microbiome

Meta analysis of Samuel et al., PNAS (2016); Dirksen et al., BMC Biol (2016); Berg et al., AEM (2016)

Platform for understanding strain-level variation on

host physiology
~550
MICROBES

CLASSIFY BASED ON:

IMPACT ON
GROWTH/METABOLI
SM
Growth rates
Brood delivery
Fat storage
Lifespan

INDUCTION OF
PATHOGEN
REPORTERS

INDUCTION OF STRESS
REPORTERS

hsp-6p::GFP (mitochondria)
hsp-4p::GFP (ER)
dhs-9p::GFP (proteosome)

BACTERIAL

irg-1p::GFP (bZip)
F35E2.5p::GFP (p38 MAPK)
clec-60p::GFP (Wnt)

PATHOGEN or
COMMENSAL?

VIRAL
RNAi effectiveness
sur-5p::GFP (Tg silencing)
scm::GFP (hairpin silencing)

Cultivated isolates of nearly 80% of

core OTUs [>75% by abundance]

Strain-level variation in responses

Definite spectrum of impact on host

physiology

Samuel et al., PNAS 2016

Prospectus much within the worm is still human

Simple to make and maintain germ-free

Defined natural microbiome
Conserved host responses and niches

GOAL: Complete genetic understanding of microbial

impact on host health

By understanding genetic pathways

we can reprogram and restore health

Healthy

Diseased

Morrison & Baxter 2012

Many thanks!!
BCM/TMC, labs of:
Joseph Petrosino
Rob Britton
Gretchen Diehl
Joseph Hyser
Lynn Zechiedrich
Mary Estes

*** POSITIONS AVAILABLE ***

Gary Ruvkun

SAMUEL LAB:
Christopher Ayoub
Fan Zhang, Ph.D.
Ali Strtak
Jessica Weckhorst

Collaborators:
Marie-Anne Felix (INRS)
Hinrich Schulenburg (U Kiel)
Michael Shapira (UC Berkeley)
Katherine Gregory (Brigham and Womens Hospital)
Allan Walker (MGH)