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Sarkis Mazmanian: Okay, thank you to the organizers, and to
Annette and Mike for the introduction. And so I'll talk to you about some of our work,
looking not solely at the immune system, but at colonization factors. And I think that
many of the introductory slides that I normally use are taken out of this presentation because
they have already been given. But I wanted to remind you of the data that we've already
seen, mostly yesterday, and then Maria's work as well, thinking about the succession of
organisms that colonize the infant, and how we assemble our microbiomes, and what are
the molecular mechanisms by which we assemble our microbiomes. And that's really what our
lab is interested in, is trying to understand, on an experimental level, on a molecular level,
how microbes colonize us, and subsequently, how they confer benefits in terms of immunologic
and neurologic health.
And so one introductory slide only just to remind you about this notion of dysbiosis
or alterations, and so this is just another way of looking at the data that we have already
seen. This is a healthy gut in people, looking across many regions of the gut, and highlighting
the importance or the prominence of bacteroidetes and Firmicutes in a healthy individual. But
then we see that the landscape is quite different in patients with IDD, lowering of the bacteroidetes
and Firmicutes, and an increase in the, mainly, the proteobacteria, which are minor species
in humans. And just by way of setting the context for the types of experiments that
we do, I think that this implies -- there's no cause or effect relationships here -- but
that this implies that there are bad players and good players, and perhaps health and disease
is in the balance of the proportions of these organisms.
And so, once again, we don't really do a lot of metagenomics, and so we don't look at community
profiles, but the approach that we have taken is to use model organisms, to use organisms
that we can genetically manipulate, associate into mice, and then look at outcomes and then
really, once again, get this molecular relationship between the bug and the host to try to understand
how the microbiome impacts us. And so the organisms -- one of the organisms that we
work with, is bacteroides fragilis, and so this is an organism that has been studied
for many, many years, initially by people like Sidney Feingold, who studied many of
these anaerobes, many of these bacteroides species, and bacteroidetes, as you know, is
the most common genus in most humans in the gastrointestinal tract.
It's a very unique organism in many ways, and so people like Dennis Kasper and Laurie
Comstock have shown that bacteroides expresses multiple capsular polysaccharides within each
genome, and so this is one thing that is very rare for organisms, because most bacteria
will only express on capsular polysaccharide per genome. There are obviously stereotypes,
but here we have multiple capsular polysaccharides, two of which have a very unique structure
in that they are positively charged and negatively charged within each subunit. So there is one
subunit of these very large molecules, and the molecule that we focused on is polysaccharide
A, PSA, because it was previously shown by Dennis and others that PSA induces CD4-positive
T-cell proliferation, and so we became interested in how that impacts the host during colonization
and symbiosis.
And so many years ago, we sort of took a leap of faith at the time, and wondered whether
or not this organism and this particular molecule would be protective in models of inflammatory
bowel disease. And we chose IBD because of its proximity to the microbiota. This is a
TNBS model of colitis. We've tested several other models as well, where you see an acute
weight loss in mice after induction of disease. But if the animals were fed a PSA orally,
you can see that they are ameliorated from the weight loss. This is the colitis that
ensues in TNBS treatment, but oral feeding of these animals with PSA shows pretty much
an unremarkable intestine. And we can do the treatment both prophylactically and therapeutically,
and get essentially the same result. And the idea is that someday PSA hopefully will be
a therapeutic for inflammatory bowel disease. And so that's the disease, and so we've looked
at the immune response, and here, once again, you can see that there is an increase in TH17
cells, these IL-17-producing CD4-positive T-cells that we've already heard about in
the TNBS in the colitic animals, but then highly reduced in animals that were treated
with the polysaccharide.
And so we wondered what the mechanism was, and there are multiple ways that one can imagine
TH17 cells, which are pro-inflammatory, could be reduced in number or inactivated, but it
appears that PSA doesn't cripple parts of the immune system or the pro-inflammatory
system. It actually activates other arms of the immune system, specifically regulatory
T-cells, and we've already, once again, heard a little bit about these as well. So regulatory
T-cells are marked by a transcription factor Foxp3, and here we have CD25 as another marker,
and, as you can see, both healthy animals and animals with colitis have the same relative
proportions in the intestine of these regulatory T-cells. But if we treat animals, even though
they were treated with the colitic agent, and if we administer PSA orally, you can see
the proportional increase in these Foxp3-positive cells. There's also a numerical increase,
and PSA also increases the transcription of Foxp3. And so what this implies is that, on
a cell-intrinsic basis, there's an increased regulatory activity. In fact, we measured
this in vitro -- I won't show this data in the sake of time -- but each regulatory T-cell
is more suppressive if it came from an animal with PSA.
And so following our work, several other groups have shown very similar results, in fact extended
some of the results in other systems, and these include single organisms like faecalibacterium
prausnitzii. There is a wonderful story, that this organism, and I think many of you already
know this, was isolated from patients, or was depleted in patients with inflammatory
bowel disease, signifying that the absence of the organism may be a risk factor for disease,
and then subsequently shown to have anti-inflammatory effects. Something very similar recently from
Herusticata's [spelled phonetically] lab showed that an organism called bifidobacterium breve
induces Interleukin-10, an anti-inflammatory cytokine, whereas another probiotic doesn't.
And then this was recently published just last year.
And then, once again, a very famous story from Kenya Honda, both in mice and humans,
that a consortium of clostridia-induced regulatory T-cells in the colon and expand their proportions,
and also protected against inflammatory bowel disease. And another story from Andrew MacPherson
showing that altered shaydysphora [spelled phonetically] can induce ITregs, or inducible
Tregs, here marked by Helios expression. So many examples, once again both individual
organisms as well as consortia, that induce regulatory T-cells, and later today we'll
hear about a wonderful story from Wendy Garrett, looking at the molecular mechanisms by which
some of these organisms may be inducing regulatory T-cells. So this notion of regulatory T-cell
induction appears to be quite broad in several organisms.
So, going back to PSAs, so then we characterize what the signaling pathways were, and I'm
just going to show you a preliminary -- or just some data from that, and so we've worked
out the entire signal transaction cascade, but here, TLR2 is clearly very important,
because this is the data I showed you. So this is colitis scores in wild-type mice untreated,
and then treated with PSA. If we do the same thing in TLR2 knockout animals, we see no
protection. In wild-type animals, PSA induces Interleukin-10, this anti-inflammatory cytokine
that I told you about, suppresses IL-17, and we see no such phenotype in TLR2 knockout
animals. And when we look at regulatory T-cell proportions, those are not increased in TLR2
knockouts. So TLR2 is clearly important in PSA signaling, and then work I'm not going
to talk about, we know that TLR2, both on the dendritics and on the T-cell, is important
for these anti-inflammatory effects.
And so to summarize the potential of PSA as therapeutic for inflammatory bowel disease,
I think the way that this mechanism is working is that PSA is secreted by outer membrane
vesicles from bacteroidetes fragilis, so we've shown that PSA is packaged into these vesicles,
taken up by dendritic cells, and, in a very unique process, presented by MAC Class 2,
two naïve T-cells induces the differentiation of these regulatory T-cells and their expansion.
Regulatory T-cells then produce a cytokine called Interleukin-10, which then ameliorates
intestinal inflammation. And Lloyd Casper [spelled phonetically] has also shown this
same process, oral feeding of PSA, is protective in models of MS, and can ameliorate inflammation
of the central nervous system. So perhaps there are effects beyond just colitis as well.
And so then we got really interested in how this organism colonizes, and, in fact, how
bacteroides colonize, and, once again, many of the talks have alluded to this notion of
who's there and what happens during perturbation. And so we approach this from a, once again,
a very different angle of using model organisms to understand what people like David Relman
and many, many others have shown, is that there is a unique pattern in mammals for the
speciation of organisms. And so even those in the back row can see that this pattern
among humans and other animals is quite similar. So there is some specificity, so some factors
that are mediating that colonization event, and selecting for particular organisms which
can inhabit the guts, whereas all the other organisms in the ecosystem in terrestrial
and aquatic environments are not permanent residents of our intestines. There's clearly
specificity in what organisms colonize as, and there is stability.
And once again, we've heard about this concept as well. And here also is data from David,
showing in either all phyla [spelled phonetically], but more particularly in bacteroides, is these
are three subjects, and these are the different organisms in those subjects, and you can see
that before antibiotic treatment, there's a particular pattern; these blue bars designate
a timeframe where they are treated with Cipro, and you can see that there is a decrease in
most organisms. But very quickly after the cessation of the antibiotic, those organisms
come back. So, clearly, these organisms are stable on a global level. And once again,
multiple other studies have shown this. And we became really interested in what are the
mechanisms that mediate this.
And so a very talented student in the laboratory, Melanie Lee, took on this very ambitious project
of trying to establish of trying to assemble a microbiome from scratch. And the way she
started doing this is, you know, working with organisms that we can culture and genetically
manipulate, and adding them sequentially to mice, and looking at colonization. And so,
initially, if we take a mouse, and start it as a germ-free animal, and we give him an
inoculum of bacteroides fragilis, we see a very nice colonization of this organism, because
germ-free animals are amenable to colonization by almost anything. But if then we, a few
days later, come back with bacteroides vulgatus, a very highly-related species, different from
bacteroides fragilis, you seen that there is really no competition between these two
organisms. We can do this with bacteroides fragilis and bacteroides data out of omicron,
once again see no difference. And this makes sense, because many of us are colonized by
multiple bacteroides species. We can reverse the order of colonization. There are really
no effects there.
But the surprise came when we took an animal and monosociated with bacteroides fragilis.
And then challenged that same animal with the isogenic strain, the same exact strain
of bacteroides fragilis, just marked by an antibiotic resistance gene. And now, in a
germ-free, or at least in a monosociated mouse, we can colonize or challenge with bacteroides
fragilis, and this organism does not colonize. And this was, you know, once seemed very surprising
to us. And initially we thought it was the antibiotic resistance gene, so we swapped
those around. We've done this by other methods as well. Every single time, bacteroides fragilis,
when challenged to a bacteroides fragilis monocolonized animal, does not colonize. The
same thing with bacteroides vulgatus, and now we've shown this with four difference
species, that there is competition between -- within an organism, but not between organisms.
We don't see the same effect with E. coli. We don't see this with enterococcus faecalis.
There's something specific about the bacteroides that mediate this phenotype.
And once again, in data I'm not going to show you, if we were to take the animal that's
monosociated with either of these bacteroides, and then give an antibiotic at the time of
challenge to which the initial strain is sensitive but the challenge strain is resistant, we
can actually displace the initial strain and get the challenge strain to colonize. And
I think what this suggested to us, that there's some limiting nutrients, or some limiting
space, that the initial organism occupies, and then excludes or resists the colonization
by the challenge strain.
And so what we did, instead of taking a larger metagenomic approach, is that we took an experimentomics
approach, and I think Fred stole a little of my thunder yesterday by introducing this
concept, and so this is what we do in the laboratories: When we have a problem, we design
experiments to tackle this. And the way Melanie devises this very ingenious screen is that
she took genome fragments from bacteroides fragilis and transected them into bacteroides
vulgatus. Remember, these two organisms do not compete with each other.
And so imagine if there was locus or piece of DNA that conferred bacteroides fragilis,
specific colonization phenotype, perhaps here expressed in red, that we would have some
clones that contained this element, whereas most of the clones of bacteroides vulgatus
do not contain this element, they act like bacteroides vulgatus. And then we took germ-free
animals and monosociated them with bacteroides vulgatus, and then challenged them with pools
of these clones that contained fragments of bacteroides fragilis. And so imagine most
of these in green are bacteroides vulgatus, and as I showed you before, they will get
cleared over time because they cannot compete with the indigenous strains. But if there
was one isolate, shown in red, that contained a bacteroides fragilis colonizing factor,
then it should be able to dual associate this animal, and then, after time, we can isolate
that organism, and sequence the genes and see what we find.
So she did this for 2,100 clones, all in mice, and what we discovered were two clones out
of the entire pool. The rest were all cleared. Both of those clones contained the same genetic
region, and that's shown here. And then when we sequence this, to our surprise, maybe not
to our surprise, is that the genes contain polysaccharide utilization loci. And so they
had these characteristics, SusC, SusD homologs, and I'll talk a little about those as well,
and regulatory elements up front, so the sigma factor and anti-sigma factor, and we've shown
that they control expression of these genes, and we started to characterize this. And so,
once again, we worked with organisms that we can genetically manipulate so we can make
mutants in these organisms and then test colonization factors.
And so here what you're looking at is the 30 of the exact same graph I showed you earlier.
So the challenge strain -- or, so the initial strain, with the challenge strain, once again
by Day 30, everything is cleared. So you can see very clear phenotypes. But if we take
animals and monosociate them with the SusD homolog, which we call CCFC, or the SusD homolog,
CCFD, that we lose this phenotype, so if the initial strain does not have this factor,
then now the challenge strain can colonize. The CCFE does not have a phenotype, and I
think we know why that is, because there is redundancy in that gene. And if we knock out
the entire operon, we see that there's a loss of function, and, classical microbiology,
we can take the mutant, and then complement the mutant on a plaza [spelled phonetically]
with the entire operon, and, once again, restore this colonization resistance phenotype. So
we can do this with bacteroides fragilis. We did the exact same thing with bacteroides
vulgatus, and saw a similar phenotype, where bacteroides vulgatus will compete against
itself. But it you make a mutant of bacteroides vulgatus, it can no longer exclude a challenge
by the wild-type strain.
And so, as I mentioned, this polysaccharide utilization loci have been studied for many
years; one of the pioneers of this is Abigail Salyers, who really figured out much of the
biochemistry of this, and the way these systems were, in a generalized fashion, is that SusC
is a poor, SusD is an outer membrane lipoprotein, and they take complex sugars, channel them
through, break them down, channel them through SusC, and then the organism uses it for nutrients.
And so these operons are widely dispersed in the bacteroides. So thetaiotaomicron has
88 of them, fragilis has 36 of these, and what we've shown is that the CCF homologs,
those genes that were isolated, are very, very unique on many characteristic levels.
And they are only found once in each of these genomes, if you look at their homology.
And so, once again, as I mentioned, these genes have been studied before, but they have
been studied in the context of foraging. So this is very nice work by Eric Martens and
Jeff Gordon, where they knocked out a sigma factor that controls expression of five of
these polysaccharide utilization loci, and if they colonized mice on a rich diet, they
see no competition. But if they switched to a restricted diet, they can see that the wilds
had colonized, but the mutant cannot.
And so that's where this concept of foraging comes from, is if the diet is restricted,
that the bacteria then use host mucous, likely [spelled phonetically] host mucous, as an
energy source because they can't use the more abundant dietary fibers. And so, once again,
you get this colonization phenotype, once again, only upon switching to this restricted
diet, and then there's a defect in vertical transmission. I know this is hard to see,
but this is this 2008 [unintelligible] micropaper, if you want to look at this defect in vertical
transmission. In fact, we don't see a defect in vertical transmission. I'll get to that
in a second.
But our -- the operon that we're studying is clearly different from this phenotype,
and Justin Sonnenburg also showed something similar, where if he took two organisms, and,
in this case, bacteroides thetaiotaomicron, bacteroides caccae, put them on a rich diet,
then you see if they could both colonize to different levels. But if you switched them
to a saccharide inulin that caccae prefers and theta doesn't, then you can see that only
during the restricted diet do you see this competition, and, in some cases, you cannot
see competition.
And so we became interested in this concept of long-term colonization on a rich diet,
and so the way we approached this is gavaging animals with laboratory-grown bacteria is
artificial, so we looked at horizontal transmission. So we took two different groups of animals,
one colonized with wild type, one colonized with a mutant, and then co-housed these animals
and looked for transfer of the wild type to the mutant colonized, and vice versa. So here
you're looking at the wild-type animals that were -- after co-housing for 14 days. So these
animals were initially colonized with the wild-type bacteria. You
can see that here. But if we then co-housed them with the mutant bacteria, the mutant
never transfers into a mouse that has already been established with the wild type. But when
we do the converse, and the initial organism is the mutant, then the wild type can very
efficiently transfer over, and, in fact, every animal can now colonize. So even during horizontal
transmission, there's a defect for the CCF genes.
Where these genes are expressed? So they are expressed in very, very low abundance in culture
and various degrees throughout the intestine, but most highly associated with the ascending
colon. And so Melanie became interested in what's going on this ascending colon, and
started looking down at the colonization of these organisms in intestinal crypts. And
so here, looking at images of a confocal micrograph of a whole mount microscopy, looking at one
particular crypt, but of course we've looked at many of them, and when you look in the
center of this crypt, only wild-type bacteroides fragilis can colonize this mouse. So once
again, these are germ-free animals, and we're looking in the intestinal crypts only in the
ascending colon, where if we mono-associate a mouse with the mutant, even though the luminal
contents are identical, none of these bacteria wind up in the intestinal crypt. Here's the
control for germ-free animals.
And another way of looking at this is through two-photon microscopies. So this is a crypt.
So the dark area is the void space of a crypt, and, as you can see, only the wild-type bacteria
can penetrate into this crypt. The mutant is at the epithelial surface, but never really
gets into the crypt. And we can quantify hundreds of different crypts, and you can see here
we're looking at the distance from the epithelium, and the wild type penetrates further into
these crypts than the mutant. And so, what we conclude is that these -- this particular
clade of polysaccharide utilization loci, the CCF, are required for colonization of
these crypts.
And so finally, what does this mean in terms of resilience or stability? And so here what
we did is we took SPF mice, and we used a per call by Thaddeus Stappenbeck to be able
to colonize these SPF mice with bacteroides so we can initially treat them with an antibiotic
to displace the indigenous microbes, and then colonize them with either bacteroides fragilis
wild type or the CCF mutant. And here you can see very, very similar levels of colonization.
But if we then challenge those animals with Ciprofloxacin, so very similar to the experiment
that David did in humans, then you can see that the wild type of bacteria is maintained,
but the mutant bacteria is greatly reduced, and thus much sensitive to Ciprofloxacin.
And, in fact, what we've done now with this experiment is we've taken wild-type-associated
mice and given them a very high concentration of Cipro for a very long period of time. We
can clear the luminal bacteria. But if we look in the crypts of these mice, on antibiotic
treatment, the bacteria are still in these reservoirs. They're still in the crypts. And
so it looks like those bacteria in the crypts are resistant to the antibiotic. And another
perturbation that we used was Citrobacter rodentium infection; the paradigm
for colonization is the same. And then we introduced the C. rodentium infection, and,
as you can see, in the dark blue bars, the wild-type bacteria are maintained, but here
we have an even more dramatic phenotype for the mutant. It's completely cleared from the
ecosystem, and you can never recover this organism again. Once again, the CCF genes
are required for resilience. And in the black bars are the Citrobacter. So, all in all,
it looks like these bacteria have evolved mechanisms to stably associate with the host,
and to populate specific regions of the gut in a way that we think is using polysaccharides
and glycans based on homology, and we were really interested in understanding what those
glycans are.
And so, to conclude, I'm going to actually take a quote from Rolf Freter, proposed 30
years ago. And what Rolf did was he took mice, and he looked at transit times of various
organisms, and then used mathematical modeling to come up with this concept that, and I'm
just going to read this, is that "Most indigenous organisms in the gut are controlled by substrate
competition. They're competing with each other for nutrients, particularly nutrients, and
that some species are better than others in acquiring these nutrients, and that these
-- the population level of the species is controlled by the concentration of a few limiting
substrates." And I think this is very important because this nicely explains our data for
how organisms will compete against each other, but not other organisms, because they are
using very, very few of these substrates, but different substrates across different
species.
And what we've done, I think, is extended that to this notion that there are populations
of cells, perhaps microbial stem cells if you will, that allow persistent occupation
of these satripal [spelled phonetically] niches, and once there is a perturbation to this system,
some from environmental stress, that disrupts the luminal bacteria, that these reservoirs
still exist, and these reservoirs can be used to repopulate the gut. And if I were a bacteria
that only lived in the [unintelligible] colon, I would evolve this mechanism to ensure my
long-term colonization. And perhaps this is one of those mechanisms, at least in bacteroides.
And so I would like to acknowledge the people who did the work, so the PSA work was done
by a very talented post-doc, June Round, who now has her own lab at the University of Utah.
As I mentioned, Melanie worked on the crypt occupancy project, with help from a new graduate
student, Greg, in the lab, as well as Silva. These are the other members of the lab, our
collaborators, and I would really like to acknowledge Klaus Ley, who take these wonderful
images for us in the colon. This is an actual crypt of a mouse colon. And so we drew in
the bacteria, but you can see the beautiful architecture here. And he was instrumental
in us understanding the mechanisms by which the CCF genes are working. And, of course,
the funding agencies: We've had two grants from NIDDK, and then one from GM, as well
as funding from the CCFA.
And so then in terms of questions and gaps, and a lot of this really dovetails off some
of the concepts we've already heard, is, maybe on a more teleological level, how do we assemble
the microbiota? Do we choose our own microbiota, or do they select us? Have they, over the
millennia, evolved mechanisms to associate with hosts in a very specific way? And I think
that if we can understand those molecular mechanisms, we can really understand microbial
succession. And so, once again, we have heard a lot about that, and maybe we've heard a
little bit less about biogeography. And so there are different organisms in different
regions of the gut -- I think we know this quite well -- harder to sample in humans,
but I think the mechanisms are coming online soon. Easier in mice, but yet still not that
many people are doing this. And I would like to remind you that there is a different biogeography
if you go longitudinally versus cross-sectionally. I think that's also important as well. And
these molecular mechanisms may really help us understand that.
And the other important question is, can we somehow exploit this colonization or phenotype
to help us resist pathogens? So there is a lot of history going back many years from
Dwayne Savage and Tyrell Conway talking about colonization resistance in bacteria. And if
we can somehow engineer organisms to compete against pathogens, if pathogens use similar
systems, then we could perhaps target pathogenic infections by making better probiotics or
targeted designer probiotics. And ultimately, is there a way to exploit the system to correct
dysbiosis? And so, once again, we have heard about this concept over and over, and if we
could understand what those sugars are, or understand what expresses those sugars, then
perhaps we can pharmacologically induce the substrates for these organisms, and then,
once again, correct dysbiosis by promoting colonization.
And then the needs, I think some of these were already mentioned yesterday, but I think
they're worth repeating because they're quite important, is this incredible heterogeneity
in organisms, or in mice, and their microbiota from different vendors. And so we know this
from the work of Dan Littman and others, that the microbiota is very, very different in
different -- from different commercial vendors. And this really affects our experiments. And
so if you look across the literature, for, let's say, Treg development in the gut, you
see very different numbers on -- from different facilities and different countries, and I
think that having some sort of standardization would really help with that. And perhaps we
can even think about humanizing these mice, and having a cohort of mice that could be
disseminated to the community, and so that we could all be doing experiments on the same
plane.
And then something that I think is quite important, but often overlooked, is whether or not our
germ-free mice are the same. And so we clearly know there is genetic drift in different colonies.
And even our germ-free animals, whether -- how rigorously we've tested them, may have genotypic
differences. In fact, we know this, because if you look a microsatellite mapping of a
black 6 mouse from different vendors, the microsatellites are different. So there has
been genetic drift. All black 6 mice are not the same. So a central repository, I think,
would be terrific.
And finally, just to reiterate some of the concepts that Maria talked about, but perhaps
more in mice because they are feasible, is longitudinal multi-generation studies. So
looking at not just the lifespan of one organism, but the ability of that organism to transfer
its microbiota to its offspring on several generations. But including perturbations that
are affecting our lifestyle, such as diet and antibiotics, we've become quite interested
in environmental antimicrobials. I think many of us already know there are very potent antimicrobials
in all parts of our lifestyle, and we're ingesting these antimicrobials, and perhaps they can
have an effect on our microbiome and our ability to pass that microbiome on to our offspring.
And I think this might be a nice way, at least in mice, to test cause and effect relationships,
and get away from associations, and understand the cause and effect relationships that could
be mediating the increase in allergic and autoimmune, as well as behavioral disorders.
So those are my thoughts. And I'll stop there, and I think we have a minute for questions.
[applause]
Male Speaker: Sarkis, thanks for a wonderful talk, as always.
Is there a quick question from anybody? Right in the center.
Male Speaker: Great talk, Sarkis. Can you comment on the
strain specification? So this question of what keeps the same bug from coming in where
it already may or may not be, and the question is dynamics of different species coming in,
I think you've hit on something that could be an important part of that. So do different
strains of fragilis show the phenotype, or what's the -- how far do you have to go before
you see this?
Sarkis Mazmanian: Yeah, we've never looked at strains, but those
can be important as well. And so we've looked at -- we work with bacteroides fragilis 9343,
and we look at the genome 638R, as well as the others, sequence bacteroides fragilis.
There are some differences in the CCF loci. They made immediate colonization differences,
but we've actually never done the experiment.
Female Speaker: Sarkis, what do you think is going on in the
crypts in the differential colonization? Is that something like substrate availability?
Sarkis Mazmanian: Yeah, so I think that the homology to the
Sus systems and to the polysaccharide utilization loci suggests that there's a limited glycan,
perhaps a host glycan, most likely a host glycan, that is specific to the crypts, and,
in fact, when we look at a field of a monocolonized animals, not every crypt is occupied by bacteria.
And so even though there's only one organism, it doesn't get into every crypt. So I think
that perhaps what this argues is that not all the crypts are the same, and so whether
the bacteria induces its substrate, or was there the entire time, the bacteria figured
out which crypts to occupy and which ones not to. So there might be some developmental
biology that we can understand through these organisms.
And I think, ultimately, that, once again, the way this is going to work is that that
limited substrate, most likely a glycan, is utilized by a specific CCF, and if it's utilized
by an initial strain, then the challenge strain can't compete, but two different species are
not using that same glycan, so therefore, they're not competing against each other.
Male Speaker: One more quick question.
Male Speaker: So you've shown really elegantly that this
works for one strain or one species. How many different species-substrate combinations do
you think there are? There are thousands of different gut species -- bacterial species
in the gut. I mean, is it possible that each one of these has its own separate substrate
that it's working on?
Sarkis Mazmanian: Yeah, so we don't know, but in terms -- so
there may be many other mechanisms for colonization that don't involve polysaccharide utilization
loci. And so the answer is, "We don't know," but when we look at the CCF genes, and what
types of organisms we can find them in, they're only in intestinal bacteroides; they're not
in bacteroides or even bacteroidetes that are not part of the Mammalian gut. So if you
look in other ecosystems, they don't have these CCF genes. And so I think that this
is one colonization mechanism; I think it's a colonization mechanism that's conserved
in bacteroides. Things like Clostridia and proteobacteria are -- probably use different
mechanisms, entirely different mechanisms. And, once again, those are not competing against
each other in a way that we can measure.
Male Speaker: Sarkis, thanks again for a wonderful talk.
[applause]
Please [spelled phonetically] -- Gene Chang from the University of Chicago. The title
of his talk is Ground Zero: The Impact of the Gut Microbiome on Host Epithelial Functions
and Reponses.