Aggiornamenti in Gastroenterologia
Latina, 19 novembre 2014
RUOLO DEL MICROBIOTA
NELLE PATOLOGIE
DELL’APPARATO DIGERENTE
Antonio Gasbarrini
Medicina Interna e Gastroenterologia
Policlinico Universitario Gemelli
Universita’ Cattolica, Roma
At birth the human body is (almost) sterile
Vaginal microbiota (mother)
Fecal microbiota
(mother)
Diet
Skin microbiota
(mother/father/parents/
babysitter/animals..)
Ambient
Native CORE microbiota
(6-48 months of life)
GUT BARRIER
Microbiota
Mucosal
Barrier
Epithelial
barrier
Endocrine
system
Vascular and
lymphatic systems
Acquired
and
Innate
immunity
Neuroenteric system
Digestive enzymes
Not only Bacteria..
Bacteria
Virus
Bacteriophages
Protozoa Helminth
Yeast Parasite
Archea
Mucosal
Barrier
Epithelial
barrier
Endocrine
system
Vascular and
lymphatic systems
Acquired
and
Innate
immunity
Neuroenteric system
Digestive enzymes
Yeast: GUT MYCOME
• Commensal to the gut at
low concentrations
• In the healthy gut dominant
species are:
Wallemia, Trichocomaceae, Rhodotorula, Saccharomycetaceae,
Pleosporaceae, Agaricaceae, Metschnikowiaceae,
Cystofilobasidiaceae, Ascomycota, Amphisphaeriaceae...
• Yeast’s functions in the gut it is not clear
• Yeasts overgrowth/dysbiosis (related to host genetic
and physiology, lifestyle, antibiotic usage, immune
system disfunction) could be pathogenic fo the gut
Dollive S, et al. Genome Biol 2012
Cui et al. Genome Medicine 2013 Thewes S, Mol Microbiol 2007
GUT MYCOME
Liver Cirrhosis
Candida (33.78%), uncult. fungi
(12.53%), Aspergillus (7.99%),
Simplicillium (5.65%), Chaetomium
(2.46%), Galactomyces (2.33%),
Rhizopus (1.96%), Wallemia (1.10%),
Fusarium (1.1%), Iodophanus (0.1%),
Penicillium (0.49%), Saccharomyces
(25.18%), uncult. Pezizomycotina
(0.86%), uncult. Pucciniomycotina
(1.10%), uncult.Agaricomycotina
(0.74%), Aureobasidium (0.61%),
Hyphozyma and Asterotremella
(0.49%), Cryptococcus (0.49%),
Doratomyces (0.37%)
Ott SJ, et al. Scand J Gastroenterol 2008
Virus: GUT VIROME
Random pyrosequencing of virus-enriched metagenomes have
been isolated from bovine rumen and human gut
In the gut have been isolated up to 28.000 different viral
genotypes
The majority (∼78%) of sequences did not match any previously
described virus
Pro phages outnumbered lytic phages approximately 2:1
Metabolic profiling revealed an enrichment of sequences with
putative functional roles in DNA and protein metabolism, but a
low proportion of sequences assigned to carbohydrate and
amino acid metabolism
Berg Miller et al, Environ Microbiol 2011
Bacteria: GUT BACTERIOME
900-1200 gr, >5.000.000 genes
>9 phyla
95% genes identity
>1000 species
99% genes identity
>15000 strains
100% genes identity
Microbiome
Leser et al, Environ Microbiol 2011
Metabolome
Qin et al, Nature 2011
BACTERIAL TAXONOMY
Genes identity
DOMINIUM
REGNUM
95%
PHYLUM
CLASSE
ORDO
FAMILIA
GENUS
99%
SPECIES
100%
SUBSPECIES (STRAIN)
MOLECULAR BACTERIOLOGY:
most abundant PHYLA in the GUT (>70%)
BACTEROIDETES
FIRMICUTES
Eckburg et al, Science 2005
HUMAN GUT
ENTEROTYPE
INDIVIDUAL ENTEROTYPE
of pe
oplea
sle
a
n or obe
seca
n bemadesole
ly on theba
sisof the
ir gut
microbiotawith90%a
ccura
cy46,47, but the
ydonot se
pa
ra
teintodis
tinct
microbiota-base
d clus
te
rson commonly use
d principal coordina
te
s
plots, which a
reuse
d to ide
ntify s
ta
tis
tical diffe
re
nce
sbe
twe
e
n groups.
Thus, multiplesta
tis
tical te
chnique
sarenee
de
dtoshowfullythediffe
rnces in the microbiota between different physiological states (Fig. 2).
Some differences in the microbiota can contribute directly to disease
ta
te
s. Gnotobioticmicetha
t we
rera
ise
dge
rm-fre
ethe
n colonize
dwith
hemicrobiotafroma
n obe
semousega
ine
dfa
t morera
pidlytha
n those
7,45
olonized with themicrobiotaof alean mouse
. A phe
notypecan
e
me
rgefromdiffe
re
nt compos
itional ba
ckgrounds, which ma
yindica
te
ha
ts
pe
cificcompone
ntsof themicrobiotacan e
xe
rt la
rgee
ffe
ctsor tha
t
many different changes can lead to the same functional result.
Diffe
re
nce
sin fa
e
cal microbial communitydive
rs
ity, compos
ition a
nd
unction ha
vealsobe
e
n corre
la
te
dwith Crohn’sdise
ase9, ulce
ra
tivecoli10
48
s , irrita
blebowe
ls
yndrome(IBS
) , Clos
tridiumdifficile
- as
s
ocia
te
d
50
dis
ea
s
e(CDAD)49 and a
cutedia
rrhoe
a
.S
ome
time
s, thena
tureof the
microbiotade
via
tion fromhe
alth iscons
iste
nt acros
sindividualswith
hes
a
medis
ea
s
e
. For ins
ta
nce
, atwin s
tudy of IBD found ma
rke
da
nd
eproducible deviations in patients with ileal Crohn’s disease relative to
hecontrols
,a
ndmores
ubtle
, but cha
ra
cte
ris
tic, cha
nge
sin pa
tie
ntswith
51
olonicCrohn’sdis
e
a
se
,a
nd s
pe
cificfunctional diffe
re
nce
swe
realso
obse
rve
dfromme
ta
bolicprofilingof thesa
mesa
mple
s24. Othe
r dise
a
se
s
re associated with marked deviations from health that are inconsistent
cross individuals. For instance, individuals with recurrent CDAD had
aphylum-le
ve
l dive
rs
ity tha
t wa
sve
ry diffe
re
nt fromcontrolsbut not
mila
r toeachothe
r49. Manydiseasestudie
sareconfoundedbye
xte
ns
ive
seof tre
a
tme
nts, suchasa
ntibiotics, tha
t ma
yobscuretruedise
ase
-assoia
te
dcha
nge
s, highlightingtheurge
nt ne
e
dfor pros
pe
ctivelongitudinal
udies that establish cause and effect.
Parallels betw een host physiological states
S
tudiesof themicrobiotaoften target onespecific diseaseor state,
but comparisonsof themicrobiotaacrossmany diseasescan show
ommon changesin thegut environment. Disturbed mucouslayersthat linetheintestinal cell wall and concomitant inflammation
re seen i n i ndi vi duals wi th I BD, coeli ac di sease, H I V enteropathy,
52
acutediarrhoea, diverticulosis, carcinomaand IBS
. Given these
aral lels, an i ncrease or decrease i n abundance of si mi l ar mi crobes
acrossdifferent disturbancesmight beexpected53, but elucidation
Phylum level diversity can have
a marked variation even across
healthy adults in the same
population. Each individual has
many unique phylotypes not
found in the other.
R
E
VIE
W INSIG
HT
Ph ylu m
Fir m icu t es
Act in ob act er ia
Bact er oid et es
Pr ot eob act er ia
Fu sob act er ia
Ten er icu t es
Sp ir och aet es
Cyan ob act er ia
Ver r u com icr ob ia
TM 7
Fu n ct ion
Cen t r al car b oh yd r at e m et ab olism
Cofact or an d vit am in b iosyn t h esis
Oligosacch ar id e an d p olyol t r an sp or t syst em
Pu r in e m et ab olism
ATP syn t h esis
Ph osp h at e an d am in o acid -t r an sp or t syst em
Am in oacyl t r an sfer RNA
Pyr im id in e m et ab olism
Rib osom e
Ar om at ic am in o-acid m et ab olism
Figure 4 | Functional redundancy. The functi onal redundancy in microbi al
ecosystems may mirror that in macroecosystems. As shown in the HM P data
set 14, oral communiti es (top panels) and faecal communiti es (bottom panels)
analysed using 16SrRNA (coloured by microbial phyla, left panels) show
tremendous abundance diversity. The same samples analysed by shotgun
Lozupone et al. Nature 2012
U.S. adults sampled >5 times up to 296 weeks apart revealed that they harbored
195±48 bacterial strains, representing 101 ± 27 species
..in stable conditions, microbiota is stable, with 60% of strains
remaining over the course of 5 years: stable core of dominant
species
Animal-based diet had a greater impact on the
gut microbiota than the plant-based diet
Abundance of 22 microbial clusters
significantly changed while on the
animal-based diet (especially
Firmicutes, Bacteroidetes,
Proteobacteria, Tenericutes,
Verrucomicrobia)
Only 3 clusters showed significant
abundance changes while on the plantbased diet
Prevotella spp reduced in
vegetarians after animal-based diet
David – Nature 2013
The human gut microbiome can rapidly switch between
herbivorous and carnivorous functional profiles
It may reflect past selective pressures during human
evolution
Microbial communities that could quickly, and
appropriately, shift their functional repertoire in response to
diet change would have subsequently enhanced human
dietary flexibility
Examples of this flexibility may persist today in the form
of the wide diversity of modern human diets
David – Nature 2013
Microbiota composition is affected by life events
Ottmann N et al. Front Cell Infect Microb 2012
INTERV
NO-SMOKE
SMOKE
EFFECTS OF GUT MICROBIOTA ON
HOST HEALTH
Barrier effect
Immunocompetence/Tolerance
Synthesis
Metabolism
Drug metabolism
Behavior conditioning
…specific effects in each GI tract!
EFFECTS OF GUT MICROBIOTA ON
HOST HEALTH
Barrier effect
Immunocompetence/Tolerance
Synthesis
Metabolism
Drug metabolism
Behavior conditioning
Small Bowel
Lifelong immunostimulation by enteric
commensal and pathogenic bacteria
Maynard CL et al. Nature 2012
EFFECTS OF GUT MICROBIOTA ON
HOST HEALTH
Barrier effect
Immunocompetence/Tolerance
Synthesis
Metabolism
Drug metabolism
Behavior conditioning
Colon
CARBOHYDRATES
METABOLISM
Microbiota plays an
essential role in
catabolism of dietary
fibers into
metabolizable
monosaccharides and
disaccharides.
Ibrahim M et al,
Bioch Bioph Res Comm 2012
GUT microbiota of ruminant has a powerful
metabolic action: herbivores derive 70% of their
energy intake from microbial breakdown of
dietary plant polysaccharides
HJ Flint et al. Nature Review Microbiol 2008
‘‘NUTRIENT SENSOR PATHWAY’’
Tilg H, J Hepatology 2010
GUT BARRIER
Microbiota
Mucosal
Barrier
Epithelial
barrier
Endocrine
system
Vascular and
lymphatic systems
Acquired
and
Innate
immunity
Neuroenteric system
Digestive enzymes
MUCO-EPITHELIAL BARRIER
IMPORTANCE OF THE MUCUS LAYERS
• The inner mucus layer is
dense and does not allow
bacteria to penetrate, thus
keeping the epithelial cell
surface free from bacteria
• The inner mucus layer is
converted into the outer layer,
which is the habitat of the
commensal flora
Good
bacteria
Loosely
adherent mucus
layer
Firmly adherent
mucus layer
Epithelial cells
Johansson, M.E., Proc Natl Acad Sci U S A, 2010
GUT MICROBIOTA HAS TO BE
UNDER CONTROL
 Mucosal barrier integrity
 Secretion of :
 Gastric acid
 Biliary salts
 Antimicrobial substances
 Mucosal pH
 Local mucosal and systemic immunity
 Intestinal motility
 Interactions among different bacteria species
EUBIOSIS
Failure of MICROBIOTA control’s mechanisms
Quali-quantitative alterations of oral,
esophageal, gastric, small bowel and/or
colonic microbiota
DYSBIOSIS
Digestive and extradigestive diseases
DYSBIOSIS
LEAKY GUT