Symbiotic bacteria in animals

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Symbiotic bacteria in animals

• Oct 3 2006
• Nancy Moran
• Professor, Ecology and Evolutionary Biology

Reading The gut flora as a forgotten organ by A.
OHara and F Shanahan EMBO Reports. 2006
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What is symbiosis?

• Term typically used for a chronic association of
  members of more than one genetic lineage, without
  overt pathogenesis
• Often for mutual benefit, which may be easy or
  difficult to observe
• Exchange of nutrients or other metabolic
  products, protection, transport, structural
  integrity

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Microbes in animal evolution

• Bacteria present by 3.9 bya, Archaea and
  Eukaryota by gt2 bya
• The Earth is populated by ecologically diverse
  microbes
• Animals appear about 1 bya
• Animals evolved in microbial soup
• Innate immune system probably universal among
  animal phyla pathogenic infection was a constant
  selection pressure
• But animals also evolved codependence on
  microbes, some of which are required for normal
  development and reproduction

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evolutionary innovations through symbiosis
examples

• Eukaryotic cell (mitochondria)
• Photosynthesis in eukaryotes (plastids)
• Colonization of land by plants (mycorrhizae)
• Nitrogen fixation by plants (rhizobia)
• Animal life at deep sea vents (chemoautotrophic
  life systems)
• Use of many nutrient-limited niches by animal
  lineages

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Why do hosts and symbionts cooperate so often?

• Persistent association allows both to increase
  their persistence and replication.
• Coinheritance
• Long-term infection
• Intimate metabolic exchange generating immediate
  beneficial feedback

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Symbiosis- main variables

• Route of infection (maternal, horizontal,
  mixture)
• Mechanisms of benefiting or exploiting hosts
• Location of symbionts in host body
• intracellular, between cells, in specialized
  organ or in other tissues, within gut lumen, etc.
  
• Molecular mechanisms of invading host tissues or
  cells similarities and differences between
  symbionts and pathogens

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Routes of transmission

• Vertical (parent to offspring)
• Horizontal
• May live in the environment (outside hosts), or
  not
• Mixture of vertical and horizontal
• Eg acquire from other individuals in the same
  family or colony (termites, humans )

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• Vertical transmission (parent to offspring)
• Infection of eggs, seeds, embryos, or babies
• Usually maternal only
• Has evolved in many invertebrate symbioses with
  bacteria, viruses and fungi
• Can be transovariolar (within the mothers body)
  or some other route (e.g. fecal-oral for gut
  inhabitants)

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Ways that vertically transmitted microbes can
increase in frequency

• Increase host survival reproduction (mutualism)
• Reproductive manipulation
• Turn presumptive male hosts into females
• Cause all-female progeny so that all offspring
  are carriers (son-killers)
• Cause hosts to be parthenogenetic (all female)
• Cytoplasmic incompatibility infected males
  sterilize uninfected females
• All of these are known to occur--caused by
  bacterial symbionts in insects Wolbachia and
  spiroplasmas

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Ways that vertically transmitted microbes can
increase in frequency

• Increase host survival reproduction (mutualism)
  
• Very common
• Why might vertical transmission be associated
  with mutualistic effects on hosts?
• Most famous cases are the lineages leading to
  organelles
• Mitochondria evolved from the alpha-Proteobacteria
  about 2 billion years ago
• Chloroplasts evolved from cyanobacteria about 1
  billion years ago

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Vertically transmitted symbiont can ultimately
fuse with the host to form a super-organism
--mutually obligate relationship --very unlike
pathogens
Eukaryotic genomes are littered with hundreds of
genes from mitochondria and plastids--now
apparent from plant and animal genome sequences.
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(Phylogenetic evidence for gene transfer from
organelles)
Cyanobacteria Cyanobacteria Eukaryote-
Plant Cyanobacteria Bacteria Bacteria Bacteria Ba
cteria Eukaryote-protozoan Eukaryote-protozoan Euk
aryote-animal Eukaryote-fungal
e.g. Arabidopsis genome has gt1000 genes from
cyanobacteria
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Vertically transmitted bacteria in animal
hosts--2 examples

1. Insect-nutritional mutualists (aphids and
   Buchnera)
2. Symbionts providing defense against natural
   enemies of hosts

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Beneficial microbes in animal hosts--examples

• Insect-nutritional mutualists (aphids Buchnera)
• Many invertebrates have specialized
  intracellular associations with bacteria that
  make nutrients
• Examples marine bivalves, leeches, many insects

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Tree of Life, N. Pace
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Aphids-Buchnera

• Intracellular bacteria in specialized host cells
• Vertically transmitted-mother to offspring
• Infection dates to gt100 million years
• Rather closely related to E. coli, but genome
  much reduced (only 600 of 4000 ancestral genes
  retained)
• Provides nutrients to host, allowing use of a
  diet that otherwise would be inadequate.

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maternal bacteriocytes containing symbionts
late embryos
early embryos with symbionts visible
1 mm
J. Sandström
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Buchnera aphidicola within pea aphid bacteriocyte
1mm
J. White
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Aphid eggs containing Buchnera from mother
0.5 mm
A. Mira
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-gtStrict vertical transmission since ancient
infection of ancestral host
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Schizaphis graminum on barley
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trp plasmid in Buchnera (Schizaphis graminum)
genomic adaptation to make more nutrients for
hosts
ori
chorismate
trpEG plasmid 14.3 kb
ori
anthranilate synthase
ori
ori
anthranilate
tryptophan
chromosome
trpD
trpA
trpB
trpC(F)
Lai, Baumann Baumann PNAS 1994
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The Buchnera gene set (570 genes) is a subset of
that of E. coli (4500 genes)
Shigenobu et al 2000 Nature
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Essential amino acid biosynthetic pathways
argA argB argC argD argE carAB
argF argG argH Glutamate---gt ---gt ---gt ---gt
---gt Ornithine ---gt ---gt ---gt ---gt ARG
ilvHI ilvC ilvD ilvE Pyruvate ---gt ---gt ---gt
---gt VAL ilvA
ilvHI ilvC ilvD ilvE Threonine ---gt
a-Ketobutyrate ---gt ---gt ---gt ---gt ILE
Pyruvate ilvHI ilvC ilvD leuA leuCD
leuB ilvE Pyruvate ---gt ---gt ---gt ---gt ---gt
---gt ---gt LEU aroH aroB
aroD aroE aroK aroA aroC PEPErythrose ---gt ---gt
---gt ---gt ---gt ---gt ---gt Chorismate 4-Phosph
ate pheA pheA hisC
Chorismate ---gt ---gt ---gt PHE
trpEG trpD trpC trpC trpAB
Chorismate ---gt ---gt ---gt ---gt ---gt TRP
thrA asd thrA thrB
thrC Aspartate ---gt ---gt ---gt Homoserine ---gt
---gt THR metB metC metE
Homoserine ---gt ---gt ---gt MET
thrA asd dapA dapB dapD dapC dapE dapF
lysA Aspartate ---gt ---gt ---gt ---gt ---gt ---gt
---gt ---gt ---gt LYS hisG
hisI hisA hisHF hisB hisC hisB hisD PRPP ATP
---gt ---gt ---gt ---gt ---gt ---gt ---gt ---gt HIS
Nonessential amino acid biosynthetic pathways
tyrA tyrA hisC Chorisimate ---gt ---gt
---gt TYR proB proA proC Glutamate
---gt ---gt ---gt PRO
serA serC serB 3-Phosphoglycerate ---gt ---gt
---gt SER glyA Serine ---gt GLY
cysE cysK Serine ---gt ---gt CYS
gtBD/gdhA 2-oxoglutarate ---gt GLU
glnA Glutamate ---gt GLN
aspCtyrB Oxaloacetate ---gt ASP
asnB/asnA Aspartate ---gt ASN
alaB/avtA Pyruvate ---gt ALA
GENE / product present in Buchnera GENE /
product absent in Buchnera
(based on Shigenobu et al 2000)
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But other symbionts appear not to have not left a
legacy of many genes transferred to host genomes,
at least not in animals so far sequenced (e.g.,
Drosophila)
Eukaryotic genomes contain many genes from
organelles, apparent from eukaryotic genome
sequences.
Why this difference?
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Heritable mutualistic bacteria (maternal
transmission)
Not much like pathogens-host has taken over
mechanisms of invading host cells and has
coevolved to maintain the association

• Mitochondria
• Chloroplasts
• Obligate nutritional symbionts (e.g. Buchnera
  in aphids)
• Facultative maternally transmitted symbionts

Much more like pathogens--have to invade naïve
hosts, overcome immune responses, but typically
benefit hosts
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Similarities between facultative symbionts and
pathogens at the molecular level

• Use of toxins that target eukaryotic cells and
  manipulate the cell cycle
• Use of secretion systems that deliver effector
  molecules to the host cytoplasm, sometimes enable
  host cell invasion
• Eg Type III Secretion Systems used by Salmonella
  and Yersinia pestis (mammalian pathogens) and by
  mutualistic symbionts of animals and plants
• Similar trends in genome evolution proliferation
  of insertion sequences (transposable elements)
  and inactivation of many ancestral genes

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Mutualistic effects of facultative symbionts on
aphids
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Hamiltonella defensa confers protection against
parasitoid wasps Kill developing parasite larva
within aphid body Increases aphid survival
reproduction
Oliver, et al. PNAS 2003 2005
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Other cases of vertically transmitted symbionts
providing defense Polyketides produced by
symbionts of beetles

• Many drug candidates from marine and terrestrial
  invertebrates are suspected metabolites of
  uncultured bacterial symbionts.
• Polyketides used as anti-tumor drugs

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Symbionts providing defense Polyketides
produced by symbionts of beetles and sponges
Biosynthesis is encoded in a 75kb acquired
chromosome fragment Used as anti-tumor drugs
J Piel 2002 PNAS 99 14002
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Why are vertically transmitted symbionts rare in
vertebrates?

• Other animal phyla studied have maternally
  transmitted symbionts, often originating hundreds
  of times (eg arthropods, molluscs)
• Acquired immunity system prohibits this type of
  symbiosis?
• Vertebrates typically have very large numbers of
  bacterial taxa associated with surfaces and gut

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Horizontally transmitted or environmentally
acquired symbionts

• Common and often clearly mutualistic
• Examples
• squid and Vibrio fischeri symbionts reacquired
  every day from seawater, special signalling
  system for recognizing the right bacteria
• Termite gut microbes
• Mammalian gut microbes
• Mouth-in habiting bacteria

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Commensal bacteria in mammalian guts- Case of
humans
In a person, bacterial cells outnumber somatic
and germ cells by gt10 fold Human intestinal
microbiota 500-1,000 different species,
aggregate biomass of 1.5 kg per person Number
of genes in the human microbiome may exceed
number of human genes by 100-fold
Xu Gordon, PNAS, 2003
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Recent research on the human gut
microbiota Summarized in A. OHara and F.
Shanahan, The gut flora as a forgotten organ
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Bacteria in mammalian gut

• Infected during birth
• Big change in community at weaning, from mostly
  aerobes to mostly anaerobes
• Differences between individuals that reinstate
  themselves following antibiotic treatment
• Some common bacterial types across individuals
• Some species with specialized communities

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Digestive tract of a cow
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Symbiotic bacteria in mammalian guts- Bacteroides
thetaiotaomicron in Mouse JI Gordon lab
(Washington University)

• Normally infection of the gut occurs at birth
• Gnotobiotic germ-free from birth
• Infection of gnotobiotic mice with single strain
  of B. thetaiotaomicron (LV Hooper et al 2001
  Science)
• Infection had major effects on expression of gt100
  mouse genes including genes modulating
  fundamental intestinal functions, some of these
  are affected similarly in zebra fish
• Major effects on development of intestine,
  vascularization

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Commensal bacteria in mammalian guts- Bacteroides
thetaiotaomicron

• DEVELOPMENT
• induction of capillary networks in intestine,
  etc.
• NUTRITION
• Absorption and processing of carbohydrates
  lipids germ-free mice require 30 more calories
  
• IMMUNITY AND DEFENSE
• Neutralization of dietary toxins
• Mucosal barrier protects against infectious
  microbes
• Bacterial surface molecules affect immune system
  functioning
• and development

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Intestinal vascularization of gut is dependent
on presence of bacteria
Germ-free conventional B.
thetaiotamicron only
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Commensal bacteria in mammalian guts- Bacteroides
thetaiotaomicron genome

• Gene content of the bacterium reflects its
  nutritional role esp in carbohydrate metabolism
• 172 glycosylhydrolases for breaking down
  carbohydratess into easily absorbed sugars, many
  of these are secreted from bacterial cells)
• Clear capacity for continued gene turnover and
  acquisition of new DNA and genes (phage, etc. ).

Symbionts, particularly consortia of commensal
bacteria, can be a means of acquiring novel
metabolic functions in eukaryotes
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Undigested carbohydrate polymers bind to surface
of Bt Much of Bt genome is devoted to making
binding proteins plus surface-localized
glycohydrolases that liberate simple sugars from
the carbohydrates. Sugars available to be used
by host, Bt, other bacteria
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B. thetaiotamicron upregulates a large set of its
genes upon colonization of the mouse intestine
64 enzymes for digesting polysaccharides in
dietary fiber Xylan, pectin, arabinose degrading
enzymes. Many of these are secreted by the
bacteria. Expression (transcription) is
affected by mouse diet. Shows adaptation to the
gut-bound lifestyle. Host mucous provides an
endogenous source of glycans used by Bt when
dietary supply is low. Bt embed in the mucosal
layer (next slide)
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Scanning electron microscope images showing
distribution of B. thetaiotaomicron within its
intestinal habitat. (A) Low-power view of the
distal small intestine of B. thetaiotaomicron
monoassociated gnotobiotic mice, showing a villus
(arrow) viewed from above. (B to D) Progressively
higher power views showing B. thetaiotaomicron
associated with luminal contents (food particles,
shed mucus) arrows in (B) and (C) and embedded
in the mucus layer overlying the epithelium
boxed region in (C), larger image in (D). Scale
bars, 50 µm (A), 5 µm (B) and (C), 0.5 µm (D).
Sonnenberg et al 2005 Science 3071955
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B. thetaiotamicron in mammalian guts

• Represents an extended phenotype--uses genes for
  host benefit and regulates them adaptively in
  response to host environment (diet)
• Retains capacity to acquire new genes, based on
  presence of integrases, phage different strains
  differ in gene content.

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Methanogens (Archaea) use hydrogen gas
(generated by carb digestion) to make methane,
thereby increasing efficiency of energy
conversion Manipulation of microbial gut
community could lower propensity for obesity?
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Consequences of interfering with gut community?

• Antibiotics-eradicate most bacteria in gut,
  followed by unusual progression back to original
  state
• Gut bacteria are environmentally acquired--Overly
  hygienic conditions-may not develop full
  diversity of gut community
• Association with Irritable Bowel Syndrome,
  Crohns disease
• May affect development of immune system
• Consequences for digestive efficiency,
  metabolism, tendency to fat deposition, obesity

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Methanobrevibacter smithii (Archaea) Methanogen De
termines efficiency of caloric uptake
"Changes in microbial ecology prompted by Western
diets, and/or differences in microbial ecology
between individuals living in these societies,
may function as an 'environmental' factor that
affects predisposition toward energy storage and
obesity. Backhad et al. Proc Natl Acad Sci USA
2004 101 15718-15723