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Eric Brown: Hello. So, as some of you might know, I'm
not Dr. Finlay. [laughs] But it's okay. So he sends his regards. So he couldn't make
it last minute today, so I'm here. I'm a graduate student in the lab working in this area. So
I'd just like to first thank the organizers, Lita and others, for the invitation. And also,
I'm one of the ones working here in microbiota and vaccines. And we're just still in the
early days in this area. But I'd just like to present more of a conceptual talk of, you
know, where the field is, and, you know, possibly where it's going.
So I don't really think I need to convince this audience of this. But unfortunately,
there are some people that still need to be convinced of this, that vaccines work, quite
simply. So I think this is best viewed through when you look at disease incidence of things
such as measles, mumps, rubella before vaccines and now after vaccines, there's just a, you
know, a dramatic decrease.
But I'd just like to point out, you know, there's a relative lack of license, effective
oral vaccines, particularly for bacterial infections. So we do have some oral vaccines
for rotavirus. You know, so there's Rotarix and RotaTeq; of course, the polio vaccine.
But in terms of developing oral vaccines for diarrheal diseases, such as salmonella, E.
coli, Shigella, these diseases remain, like, a significant worldwide health issue. There
are still tens of millions of people in the developing world that die of these diseases.
And I think that this is best viewed here through disability life years lost. So the
amount of -- the burden in hospitals of treating these patients, and that accounts for around
15 percent of that, is diarrheal diseases.
And the -- so the gold standard for this is the development of a vaccine. Now we know
for these diarrheal diseases, particularly, that we can use antibiotics. But as Julian
Davies nicely pointed out, with the increasing incidence of antibiotic resistance, you know,
we're really trying to develop, you know, oral vaccines that are easy to use and effective
for these diseases.
But, really, the problem in the field -- and many people have been looking at this -- is
there's really a poor understanding of the determinants of protective gut immunity. And
there's another phenomenon, which I'll get into, which David Mills kind of alluded to
as well, is vaccines have reduced efficacy and immunogenicity in developing countries,
specifically areas with poor sanitation.
Now we've had three days of great background into everything that microbiota does to the
host. And I think we know now there are specific bacterial species that actually influence
our immune system. So I don't need to convince you of this. So these are the three, you know,
best characterized examples in the field. There's bacteroides fragilis secreting polysaccharide
A, so Sarkis Mazmanian's group has looked at that. Inducing T-regulatory cells, Kenya
Honda has done some great work on specific clostridial species that can induce T-regulatory
cells. And segmented filamentus bacteria can lead to proliferation of T17 cells.
I don't think I need to convince you that shifts in the microbiota can actually influence
immune development. And this is -- also can be viewed through germ-free mice, which we
know have reduced numbers and size of pairs patches, and really just a lack of gut-associated
lymphoid tissue, reduced levels of class switching to IGG and IGA antibodies, decreased number
of CD4-positive T cells. And these mice don't develop tolerance. But if you come into a
germ-free mouse with a complex host-adapted microbiota, we can now restore the intestinal
immune system, so we need our microbiota for immune development.
So our lab posed the question a few years back, you know, given this, the profound impact
the microbiota has on the development of our immune system, especially early in life, should
human microbiome be considered when we're developing these vaccines? So our lab has
been particularly interested in developing oral vaccines for diarrheal pathogens, such
as E. coli. We have a vaccine for cattle. We're currently working on a oral non-typhoidal
vaccine for salmonella.
So given that we're working on this, and we're also researching the microbiome field, we
posed this question. And still is the early days, which there's really not too much data,
but I know there are groups working on this. And we wanted to ask the question, could our
newfound knowledge of the impacts of the gut microbiota be a missing link to improve oral
vaccine efficacy and develop more effective oral vaccines?
So this is phenomena, there's really a large body of evidence in the literature suggesting
that oral vaccines, specifically polio, rotavirus, cholera, ETEC, show a greatly-reduced efficacy
in developing countries, specifically regions with poor sanitation compared to developed
countries.
A couple examples, using the oral cholera vaccine as a tool to show this, is that Nicaraguan
children have blunted antibody responses compared to those in Sweden. And there's a study by
Myron Levine's [spelled phonetically] group a few years back where they showed that kids
with small intestinal bacterial overgrowth also had blunted antibody responses.
So -- but we also know that, you know, these -- this phenomena, blunted vaccine responses
to oral vaccines, you know, could be due to a wide variety of issues for people living
in these areas of the world. You know, they live in these areas where they have increased
antigen exposure. So if you come in with an oral antigen, the immune system might react
differently than it would here. That's sort of the hygiene hypothesis idea.
Of course, you know, a lot of these people are malnourished. And we know that, you know,
this is -- leads to nutrient deficiencies, such as vitamin A. So what David Mills talked
about, you know, there's hundreds of millions of people who are vitamin A deficient in many
areas of the world. And, you know, that's an important pathway in many immune pathways,
the retinoic acid pathway. And also zinc is quite important.
You know, these people use less antibiotics, and I think the elephant in the room as well
is the parasite. So they really have a greater parasite burden in their gut compared to here,
in more developed countries. However, I'd say that all of these changes can actually
feed into your microbiota composition. So -- and this is a study that's actually been
cited numerous times over the conference. It's one of the first studies to really look
at the microbiota composition in comparing a African cohort to a cohort in the European
Union.
So what they saw is in Burkina Faso, there's actually an increase in prevotella, and actually
microbes that can degrade carbohydrates. And then in the European Union, there's really
an increase in the amount of, let's say, clostridiales or bacteroidetes that are involved in lipid
metabolism.
So we know now that diet can actually shift microbial assemblages, and there's really
a difference in microbial assemblage in some of these areas of the world. And this sort
of phenomenon of more prevotella in the gut has been seen in other studies. And we also
know from some great work by Jeff Gordon's group that malnutrition can actually shift
from the microbial community as well. So could the microbiota be implicated as to why vaccines
shall reduce efficacy in developing countries? And that's still an open question that I think
many people are interested in.
What -- there have been some studies. You know, how can we solve this, right? So we're
modulating the flora to try to use, you know, prebiotics or probiotics as adjuvants, per
se, to, you know, actually help some of these vaccine responses. So there's been some small
studies, mostly in mice, where they used lactobacillus and strains of bifidobacterium to actually
improve rotavirus, cholera vaccine, and salmonella typhi vaccine efficacy, as well as prebiotics,
fructooligosaccharide mixes, to improve salmonella vaccine and influenza vaccine responses. So
there is some evidence that this might be important.
So the question is, you know, can we manipulate the microbiota to improve vaccine responses?
And similarly, there -- I've been talking mostly about oral vaccine. But there is some
evidence that in parental vaccines, probiotics can be important as well, particularly early
in life. So I think there's been a few great presentations that have shown the difference
in microbiota after a C-section, or -- versus a vaginal delivery, or formula feeding versus
breast milk. And this can actually decrease your probiotic microorganisms, and, you know,
what the effect this has on the vaccine responses remains to be seen. However, as David just
suggested, you know, this could be an important thing to realize in these kits.
But there's sort of -- still in the early days -- there's really a lack of long-term
follow-up studies. And some studies do show variable effectiveness. You know, it's important
to choose the right probiotic strain. And some mechanisms are unclear. But I think David
gave a great talk and -- about, you know, possibly the need for synbiotics. So this
could help with the colonization efficiency. And as I've seen, there's no studies as of
yet where they've actually used more of a symbiotic approach in terms of improving vaccine
responses. So this could be an interesting area.
So another sort of paradigm I'd like to touch on is this idea that, you know, we could actually
maybe use probiotics to deliver vaccine antigens themselves. So there is some talk yesterday
about, you know, there's this need to genetically manipulate some of these anaerobes. And we
need to kind of develop the tools of that. And one of the translational applications
of that could be actually getting these probiotics to actually display some vaccine antigens
so we could get a lactobacillus, possibly, and, you know, deliver some vaccine antigens
and have it colonize in the host.
So there's a proof of concept study where they did this in a lactobacillus lactis strain.
So this was just proof of concept, but they made it express a listeria internalin. And
they showed that it actually internalizes and delivers the gene in the small intestines.
So this could be, you know, a way to deliver some sort of a DNA-based vaccine. However,
there's no real data on the efficacy in animals or humans as of yet. So this is just sort
of a concept that could be thought out.
But I think it's also important to point out with this concept that it could be relevant
to what we now know about IGA responses. So Andrew MacPherson's group has done some great
work looking at IGA. And what they've seen is that you actually constant exposure of
oral antigen to elicit lifelong IGA responses. So if you come in with an oral antigen and
look at specific IGA years later, it will be gone. But if you actually constantly expose
the host to that antigen, it'll help mucosal IGA responses.
So there was a couple of studies I also want to highlight. And these are actually from
groups here in Maryland: Marcelo Sztein and Claire Fraser. So they've done some stuff.
This is work in macaques, where they've looked at the microbiota in vaccines. And they looked
at macaques from different geographical regions, and they had different assemblages of microbiota.
And they came in with an oral shigella vaccine. And they showed that this vaccine actually
really didn't have a change in the microbiota. However, after vaccination, different macaques
responded differently to infection. So there's -- actually got shigellosis only in certain
macaques compared to others. So I won't really dive into the details of the study. I think
Claire is here today, so if you want to ask here, but I just wanted to point out that,
you know, maybe we need to take the impact of the microbiota into consideration in some
of these vaccine trials. You know, not all macaques are the same, of course, and not
all humans are the same.
And so another study from the same group is looking in humans at oral typhoid vaccination.
So there's an oral typhoid strain, Ty21A. And this is a great study, but just one thing
I wanted to point out is they saw the same thing: no real change in the microbiota due
to the vaccine. However, they split up the cell-mediated response into a multi-phasic
response, or late responders. And what they did see is if you had a more diverse microbiota,
you actually had a greater multi-phasic cell-mediated response to this vaccine. And this was actually
shown more in the clostridiales groups.
So this kind of, you know -- these are correlative studies, of course, but they kind of hint
to the importance of our microbiota in oral vaccine responses.
So some of the work our lab is doing, we're actually working in MIRI models. And we're
testing different antibiotics to mice early in life or adult mice. And we're coming in
with different -- either salmonella peptide vaccine, or ovalbumin type mock vaccination
orally. And then looking at the microbiota and looking at specific antibody and T-cell-mediated
readouts to see whether different microbiota shifts can actually shift our immune response
to vaccines.
So we're still in the early days of this work. But some of the changes that we've seen have
been in IGA responses. So we know we need diverse microbial exposure to get proper levels
of IGA. So in mice treated with Vancomycin from birth, they have reduced amount of colonic
T-regulatory cells, which have been known to be helper cells for IGA responses. So we
see a reduction in IGA response to these -- the vaccine. And -- but interestingly, when we
took the adult mice and we treated with Vancomycin, we actually saw an increase in the amount
of IGA specific to the vaccine, and we think this is possibly a permeability issue, so
that, you know, from adulthood, they do have a developed immune system. But then the energy
can actually cross the barrier and be seen by systemic immune system, which is also important
in oral vaccination.
But, you know, we're still in the early days. And, you know, I think this conference has
done a fabulous job of really addressing a lot of the sort of gaps, needs, and challenges
in the field, and definitely in the area of microbial vaccines, such as such in early
days, there's a ton of challenges. And this is something that a lot of people have touched
on, but I'll just reiterate, is that, of course, people working with vaccines know that, you
know, you want to get the vaccine to human as quickly as possible. There are so many
examples of vaccines that fail, or that work in animal models, that actually turn out to
not work in humans.
So humans are much more relevant. But, of course, when we're looking at shifts in the
microbiota, in correlating it to vaccine responses, as I -- it's correlative. And you can't actually
dive into the mechanism, whereas if you're using an animal model, whether it be a murine
or macaques, you know, this might have more poor translation to humans. However, we can
be more mechanistic with the function. So I think we still need a balance of both.
And something that I know is an issue in these vaccine trials in humans is that, you know,
we take the feces and we look at the microbiota composition, and then we correlate that to
vaccine responses. But fecal microbiota is just really the flow-through of all the microbiota,
and you really lose that sort of spatial distribution and longitudinal distribution from the small
intestine to the colon that some people have been talking about. And you just kind of get
this community, but where we're not really sure, you know, which ones are actually adhering
to the mucosal surfaces. And I would argue that these ones would be more immunologically
relevant.
And just basically the same point, so it's difficult in humans to study these mucosal-associated
microbiota. You can take biopsies, but there are, like, ethical considerations there. So
there's a significant gap between animal and human studies. And what we need -- and I know
there are some groups interested in this -- is mice with humanized immune systems. So you
can actually take a mouse and give it sort of a more human-based immune system. And now
maybe we can come in with more of a human microbiota, and this might be more translational
to vaccine research. Now, you know, I think some scientists or groups would pay a pretty
penny for actually, maybe, a germ-free mouse with a humanized immune system, and then come
in with a humanized microbiota. And this could be sort of a need in the field for the future.
So for the last part of the talk, I'm going to shift gears and talk about something completely
different, still related to vaccines and microbiota but this sort of idea is can we actually use
vaccines to target specific species in the microbiota? And this might be a need -- has
to begin to identify some of the keystone species or pathobionts, or troublemakers,
per se, in the gut. So we have, you know, probiotics that come in and try to tame those.
But we could also maybe start to target them specifically because right now we really only
have blunt tools at our disposal to do so, with lots of side effects. So that's such,
of course, like dietary changes, antibiotics, or prebiotics, or even phage therapy.
So I think this is one study that explains this is that in periodontitis, there's one
species, porphyromonas gingivalis, that just the presence of this organism is kind of a
troublemaker, per se. So it influences the microbiota in the oral cavity around it to
become more virulent. And it's quite a low abundance organism, but it seems to have profound
effects in this disease. And so, you know, can we target this? And there have been groups
-- there's one that looked at a periodontal vaccine. So can we target, you know, parts
and constituents of the oral cavity to try to get rid of these, you know, pathobionts?
So they looked at an outer membrane porin (FomA) from the species fusobacterium nucleatum.
And they, by targeting it, they kind of got rid of this bridge between how porphyromonas
enables itself to form biofilms and cause gingivitis. So this worked in a mouse model
of gingivitis.
So -- and there's some more examples where we can maybe target pathobionts with vaccines,
like the haemophilus influenzae B vaccine, which has virtually eliminated this haemophilus
from the pharyngeal microbiota. So before the vaccine, there was asymptomatic [spelled
phonetically] carriers in about 3 to 5 percent. But now it's replaced by a less virulent haemophilus
strains. Also, streptococcal pneumonia, the valiant vaccine, has seen a 77 percent decrease
in disease. And this has been replaced by other non-vaccine strains that appear to be
less virulent.
So this opens up some questions, you know, can we target specific microbiota with vaccines?
And, you know, maybe they'll be replaced with sort of less virulent, closely-related species.
Now I realize that, you know, anytime you see sort of autism and vaccines together in
a sentence, it makes me cringe, it makes scientists cringe. But, you know, this is sort of a more
of a provocative slide to say that, you know, there's some evidence coming out now that
actually autism is a GI disease. So 90 percent of autistic kids have some sort of GI irritation.
And there's been studies that showed that there's this clostridium bolteae, or there's
other clades of clostridium that actually produce toxins that could go systemic in the
bloodstream and possibly have neurological effects. So could we actually, you know, target
some of these pathobionts with some sort of vaccine and sort of quell those effects?
Now there is some talk that this autism could also be sort of a gut permeability issue as
well. So can we sort of target some of those metabolites that are produced, and, you know,
try to get rid of some of those effects. But this is something that's sort of very new.
And there's also a group at Guelph who is looking into this. And so try to make a vaccine
specific for a cell wall polysaccharide-immunogen from these clostridia, and to see if we can
actually, you know, get rid of them.
So -- but I have to say, you know, given everything that's been talked about, you know, about
-- I think -- which is great about the ecosystem of the microbiota, you know, what could be
the consequences of actually, you know, playing God and targeting these microbiota with vaccines?
So this is sort of virtually unexplored. But if you're targeting specific constituents
in the microbiota, you know, what are the ripple-down effects in the community. And
this is very difficult to predict. And I think David Relman has done some great work in this
area, looking at the importance and interactions and ecosystem dynamics of the microbiota across
the planes, and how they actually turn dysbiotic or not.
So we need to know -- and still, there is still a need for this basic science, you know,
the contribution of each species to the community, because we've got to remember this microbial
ecosystem is a complex, adaptive system. It's nonlinear, so very small changes can have
profound effects on the community. And I think this is well-known for microbial ecologists
in the field.
So I'd just like to leave you with some future challenges and questions for the field. So
one is, you know, can the microbiota be altered to improve vaccine responses? Can you actually
use microbiota to deliver vaccine antigens, and possibly, can specific vaccines be designed
to target particular troublemaker microbiota strains?
And with that, I'd just like to acknowledge Brett and the lab. And thanks for a great
conference. Thanks.
[applause]
Male Speaker:
So now we'll invite for the open floor all those keynote speakers here.
And we'll be around for taking questions. And those questions will be from this session
or other sessions as well. Thank you.
Female Speaker: Before we get started on the open floor discussion,
you recall we had a poster competition. It turns out --