Drosophila

Angela E Douglas

Drosophila: a superb model system

for microbiome science
Drosophila &
gut microbiota
axenic
Drosophila

bacteria
culturable

resynthesize
the association
(gnotobiotic flies)

1. Composition of microbiota
2. Axenic flies and gnotobiotic flies
3. Genetic & genomic resources
of Drosophila
4. Implications for animal welfare,
translatability, and challenges

Composition of the gut microbiota

Drosophila &
gut microbiota

Low diversity:
15-80 OTUs per fly
Acetobacteraceae
(Acetobacter, Gluconobacter)

axenic
Drosophila

bacteria
culturable

resynthesize
the association
(gnotobiotic flies)

Lactobacillales
(Lactobacillus, Vagococcus, Leuconostoc)

-proteobacteria
(Stenotrophomonas, Erwinia, Providencia)

The Inconstant Microbiota

of Drosophila
Graphical Taxonomy
Networks
3 wild populations
CantonS in lab-1
(2 timepoints)
CantonS in lab-2
(3 timepoints)

Wong et al. 2013

Mice (and people) have

an inconstant microbiota too .

Science (2016) 353: 741

http://www.sciencemag.org/news/2016/08/mouse-microbes-may-make-scientific-studies-harder-replicate

The Inconstant Microbiota

of Drosophila
Relative abundance (%)

79 lines of D. melanogaster
reared under uniform conditions

12-85 OTUs
Chaston et al. 2015

50
40
30
20
10

ct

iv
or
po ans
m
A. or
tro um
L. pic
al
pl
an is
ta
ru
m
L.
br
ev
is
O
th
er

fru

resynthesize
the association
(gnotobiotic flies)

% 16S reads

bacteria
culturable

L.

axenic
Drosophila

Ourstandardizedmicrobiota

A.

Drosophila &
gut microbiota

Drosophila &
gut microbiota

axenic
Drosophila

How fit is the axenic fly?

bacteria
culturable

resynthesize
the association
(gnotobiotic flies)

4,000 conventional
& 4,000 axenic flies

Wong et al. 2014

Impact of gut bacteria on

Drosophila on 16 diets
axenic
conventional

Wong et al. 2014

Nutritional rescue of axenic Drosophila

on low nutrient diet

0.8
0.6

1.0

proportion pupated

Proportion surviving
to pupa

25 g yeast + 200 g glucose liter-1

1.0

0.8
0.6
0.4
0.2

Gut microorganisms spare

0.0 dietary requirement
1.0
of
Drosophila
for
B vitamins,
0.2
0.8
especially riboflavin
0.6
0.0
none casein B-vitamins
diet supplement
bacteria
conventional
axenic

0.4

B vitamins
0.2

animal

0.0

no
ne
th a
ia ll
m
r
i
ni bo ine
co fl
a
pa tinic vin
nt a
ot ci
h d
py ena
rid te
ox
in
e
fo bio
lic tin
ac
id

0.4

single dietary omissions

of B vitamins

Wong, Dobson and Douglas, 2014

Nutrient allocation in Drosophila

on nutritionally-adequate diets
g TAG mg-1 weight
mean + s.e.

150

p<0.001

125
100
75

p<0.001

50
25

g glucose mg-1 weight

mean + s.e.

0
15

10

p<0.005

hyperglycemia and
hyperlipidemia
in axenic flies

p<0.001

0
male

female

conventional flies
axenic flies

Ridley et al. 2012

Identifying the Gut Microorganisms that Protect Drosophila

against Hyperglycemia and Hyperlipidemia

Drosophila line

Drosophila line

Correlation coefficient
Bacterial taxon

fly TAG content

Fly glucose content

Acetobacteraceae

-0.43 (p=0.002, n=49)

n.s.

Komagataeibacter

-0.38 (p=0.001, n=10)

-0.38 (p=0.001, n=14)

Lactobacillus OTU7

+0.44 (p=0.002, n=21)

n.s.

Chaston et al. 2015

50

10

er

1000
c
800

120

c
c

-1

-1

g triglyceride mg protein
(mean + s.e.)

bacterial species

O
th

L.
br
ev
is

uc
tiv
or
an
s
A.
po
m
or
um
A.
tro
pic
ali
s
L.
pla
nta
ru
m

Identifying the Gut Microorganisms that Protect Drosophila

against Hyperglycemia and Hyperlipidemia

600
b
400
200

a,b
a

co
nv
en
tio
5- na
s
l
A. pec
po ies
m
A. oru
tro m
pi
ca
l
L
. b is
L.
fru re
ct vis
i
L. vor
pl an
an s
ta
ru
m
ax
en
ic

Acetobacter protect against

hyperlipidemia by competing with
host for dietary sugar (Huang &
Douglas 2015)
Bacterial genetic determinants
identified (Chaston et al. 2014)

100
80

a
a

60
40
20
0
co
nv
en
tio
5- na
s
l
A. pec
po ies
m
A. oru
tro m
pi
ca
l
L
. b is
L.
fru re
ct vis
i
L. vor
pl an
an s
ta
ru
m
ax
en
ic

20

g glucose mg protein
(mean + s.e.)

30

L.
fr

% 16S reads

40

Lactobacillus and Acetobacter

reduce glucose levels
Some bacterial taxa and microbial
communities fail to protect against
hyperglycemia (unpub)
Newell & Douglas, 2014

Drosophila: a superb model system

for microbiome science
Drosophila &
gut microbiota
axenic
Drosophila

bacteria
culturable

resynthesize
the association
(gnotobiotic flies)

1. Composition of microbiota
2. Axenic flies and gnotobiotic flies
3. Genetic & genomic resources
of Drosophila
4. Translatability and challenges
of the Drosophila system

Harnessing the Genetic and Genomic Resources

of Drosophila for Microbiome Research
1. Comprehensivepanelsofmutantsmaintainedat
stockcenters
2. UASGAL4systemforstrictspatiotemporal
controlovergeneexpression
3. CRISPRforgeneediting
4. DrosophilaGeneticResourcePanel
200inbredlineswithsequencedgenomes:
(Mackayetal.2012;Huangetal.2014)
5.DrosophilaGlobalDiversityPanel:
84inbredlinesfrom5continentswith
sequencedgenomes(Grenieretal.2015)

Drosophila genetic resources and

regulation of the gut microbiota
Acidic region maintained by H+-V-type ATPase
Vha100-4-RNAi

ProximalpH7

pH4

Dissected gut of larva fed on m-cresol purple pH dye

Overend et al. 2016

Dominant bacterium Acetobacter

controlled by pH of acidic region
1.2e+5

Acetobacter

parent 1
RNAi
parent 2

1.0e+5
8.0e+4
6.0e+4
4.0e+4

gut region

distal

transition

neutral

0.0

acidic

2.0e+4
proximal

Acetobacter abundance
(mean + s.e.)

1.4e+5

Overend et al. 2016

325
300

g TAG
-1
mg dry wt

250
90
80
70
60
50
16

Response index

275

g glucose
-1
mg dry wt

g protein
-1
mg dry wt

Drosophila genomic resources and

microbiota-dependent nutritional traits
30
15
0
-15
-30
4
2
0
-2
8

14

12

10

-4

Drosophila line
AX GN
Averaged across all DGRP tested
AX= axenic; GN=gnotobiotic

1. Host genotype is a
major determinant of
elevated
in axenic microbiotadependent traits
flies
2. Apply GWAS to
identify genetic
determinants of
variation in
microbiotadependent traits
3. Validate candidate
genes by quantifying
phenotype of
mutants
Dobson et al., 2015

100

2. Effect of mutation evident in

axenic fly, i.e. microbiota
may mask phenotypic
consequences of host
genetic variation
Dobson et al., 2015

rugose

75

n.s.

50
25
0
50

bg

rg

bg

rg

g lipid per fly (mean + s.e.)

1. Many candidate genes

expressed in gut and/or
neurons; and with
homologs in mammals,
including humans

Drosophila genomic resources and

microbiota-dependent nutritional traits

Dscam3

n.s.

40
30
20
10
0

bg

dscam

gnotobiotic
flies

bg

dscam

axenic
flies

Impact of Gut Microbiome on

Transcriptional Networks

chico in gnotobiotic flies (TPM)

11.2

chico in axenic flies (TPM)

Transcriptome of axenic and gnotobiotic flies of 17 Drosophila lines

11.0

11.0
10.8
10.6
10.4

p=0.004
10.2
11.4

11.6

11.8

12.0

12.2

12.4

12.6

foxo in gnotobiotic flies (TPM)

10.8
10.6
10.4
10.2

p>0.05
10.0
11.6

11.7

11.8

11.9

12.0

12.1

foxo in axenic flies (TPM)

Dobson et al. 2016

12.2

Transcriptome-wide coexpression is significantly weaker

in axenic flies than gnotobiotic
flies (p = 2 x 10-16)
48% of expressed genes
assigned to 11 transcriptional
modules of co-expressed genes
7 modules with different coexpression between gnotobiotic
and axenic flies

Microbiome promotes co-expression

of specific transcriptional modules
*

median co-expression index

0.20

0.15

0.10

0.05

0.00

GN
AX
not differentially co-expressed

AX
GN
differentially co-expressed

transcriptional modules
Gain of function in microbiome-dependent transcriptional modules
Dobson et al. 2016

Microbiome promotes co-expression

of specific transcriptional modules
*

median co-expression index

0.20

0.15

0.10

0.05

0.00

GN
AX
not differentially co-expressed

AX
GN
differentially co-expressed

1. How general is this effect? (males of one species on one diet)

Gain of function in microbiome-dependent transcriptional modules
2. Is reduced coexpression associated with dysbiosis
(microbiomes that fail to support health)
Dobson et al. 2016

Drosophila:
a superb model system
for microbiome science
Association is amenable to manipulation
Genetic and genomic tools
Large and complex experimental designs
1. Inconstant microbiota, with community composition
shaped by a mix of deterministic and stochastic factors
2. Nutritional benefit of microbiota that both provides
nutrients and competes for nutrients
3. Drosophila genetic determinants of microbiotadependent effects on energy storage are conserved
across animal kingdom to humans
4. Penetrance of deleterious mutations can be
increased by perturbation of the microbiota
5. Microbiota-dependent co-expression may promote
healthy phenotype

Drosophila:
Implications for welfare
of laboratory animals

https://upload.wikimedia.org/wikipedia/
commons/f/f2/Lab_mouse_mg_3263.jpg

Association is amenable to manipulation

Genetic and genomic tools
Large and complex experimental designs

Appropriate use of non-vertebrate models

can substantially reduce animal
experiments
..without compromising scientific quality

Drosophila:
For enhanced quality
of microbiome science
Translatability
Drosophila to fast-track understanding of
fundamental principles of animal-microbial
associations
Mammalian models and human data to verify
relevance to humans
Challenges
Strengthen the framework and infrastructure for
integrating model systems with biomedical and
clinical science

Acknowledgements
Karen Adair
Alyssa Bost
Eduardo Bueno
John Chaston
Adam Dobson
Leanne Donahue
Sara Hermann
Jia Hsin Huang
Yuan Luo
John McMullen
Emma Ridley
Adam Wong

Cornell University
Andy Clark
Brian Lazzaro
Memorial Sloan Kettering
Cancer Center
Justin Cross
University of Glasgow, UK
Julian Dow
Gayle Overend