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Webinar #4-- The Microbiome and IBD

Guy Daniels
Onegevity

Listen and learn as Guy Daniels dives into the research on the microbiome and IBD, the physiology in the gut, and the key bacteria influencers in health outcomes. 

Slide 1 – Welcome back to another webinar, in our in-depth series, on how the microbiome impacts our health. My name is Guy Daniels, and I’m the head of medical education at Onegevity Health. In today’s webinar, we’ll be discussing inflammatory bowel disease, or IBD for short. 

Slide 2 – Now IBD is an umbrella term, which primarily consists of two sister conditions, Crohn’s disease, and ulcerative colitis. We won’t be discussing the array of drugs used in IBD, nor any dietary or supplement considerations, all of which are outside the scope of this presentation, however at the end, I’ll provide you a hotlink to a longer webinar on IBD which address these topics and more.

Slide 3 – For these next few slides, I do want to at least inform you of the limitations of modern medicine, in the treatment of IBD, and to motivate you, essentially through fear, as if left unchecked, both conditions can result in some nasty life-altering outcomes. As we can see from this slide, even with advances in medicine, surgical, morbidity, and mortality rates have not changed throughout the years. That means that up to 80% of Crohn's patients will require some type of intestinal surgery, and up to 30% of ulcerative colitis patients will require colectomy.

Slide 4 – So what undesirable situations can arise from Crohn’s disease? One common possible outcome is a stricture, which is a narrowing of the lumen (the hollow part of the GI tube) as a result of fibrosis through chronic inflammation. Over 1/3 of Crohn’s patients will require this surgery and the subsequent resection. Another possible outcome severely diminishing the quality of life is a fistula, as pictured here. About 35% of Crohn’s patients will suffer from a fistula at some point, more commonly in Crohn’s colitis, which is a bit of a blend of the two. So, what is a fistula? It’s the unwanted tunneling of cells, as a consequence of chronic inflammation, from one location to another. The most common of which is the anal fistula, where the fecal matter will leak through this new tunnel, out of the body in proximity to the anus. Even worse are the recto-vaginal fistulas, which will leak fecal matter into the vagina from the rectum. With all of this in mind, it may be wise to consider your options in addressing the root cause of Crohn’s disease.

Slide 5 – Ulcerative colitis is no less scary than Crohn’s. For example, in patients diagnosed from the ages of 0-19 years of age, their relative risk for colorectal cancer over their lifetime is 44-fold higher than the norm. Forty-four is an astonishingly increased risk. In addition, ulcerative colitis patients run the risk of losing their entire colon. And there are two methods to this so-called surgical cure. Option one is to remove the colon and rectum, sew up the anus, and attach an ostomy bag to the outside of the body to collect what is essentially incomplete fecal matter, which the patient has to empty. Option two also begins with the removal of the colon and rectum and connects the end of the small intestine to the anus, while reconfiguring the last few inches to serve as a new rectum. Of course, the cells that line the distal ileum are not intended to serve as a rectum, and as such, problems arise with inflammation. In addition, the 5-6 bowel movements per day may be better than having an ostomy bag attached to you, but probably are no less than the number you had before surgery.

Slide 6 – So now that I have your attention let’s focus on the big picture. On the right side of this slide, within the two circles, you’ll notice what may seem to be gibberish but is a list of only a handful of the genetic links to IBD. Some have been only associated with Crohn’s, some with both, and some only with ulcerative colitis. There has been a great deal of research done over the years looking into the genetic connection to IBD. And to be clear, there is one. Even from an epidemiological viewpoint, we see that first-degree relatives have 8-10 times higher risk than the norm. However, no genetic associations can be confirmed in 77% of Crohn’s disease patients and up to 84% of ulcerative colitis patients. Also, the remarkable rise of the incidence of IBD over the past few decades cannot be sufficiently explained by only genetic risk, or increased diagnosis, and accessibility of care. In the text to the left of the slide, you’ll see profound disturbances mentioned; we’ll touch on a couple of them in this webinar but take a much deeper dive in future webinars. 

Slide 7 – So let’s take a look at the core of what’s going on behind both conditions, proven in vast amounts of literature over the years. In essence, the primary flaws in Crohn’s center around defects in anti-microbial defenses. The ones for ulcerative colitis center around mucus production and degradation. The result is that bacteria can adhere to and invade the cells that line your GI tract. This results in an overzealous immune system, chronic inflammation, and ultimately cells going wild and doing things they shouldn’t be doing.

Slide 8 – For example, in this study, these researchers looked at classic genetic flaws in Crohn’s. These SNP’s, or single nucleotide polymorphisms, result in defects in antimicrobial defenses, both within and outside the cells. As such, they found that the shifts in the good bacteria Faecalibacterium and the opportunistic pathogens of Escherichia were associated with these genetic flaws. Also, of note, the phyla actinobacteria and proteobacteria were significantly higher in Crohn's vs. healthy controls, and the healthy clusters IV and XIVa were significantly lower. These are all pretty classical findings for the Crohn’s microbiome, and even for ulcerative colitis at times as well.

Slide 9 – For ulcerative colitis, I want to actually read verbatim a couple of these points. Individuals with UC have decreased numbers of goblet cells (these are the cells that produce mucus) and reduced mucus thickness at presentation, and goblet cell abnormalities play an etiological role in UC. And the composition of short-chain fatty acids in the intestine is determined by the composition of the microbiota, and butyrate can mediate MUC2 mRNA. In other words, you can drive success in UC through changing the balance of the microbiome, which alters SCFA production, which positively impacts mucus production, among other things. 

Slide 10 – Here we see from the two diagrams, that in the colon, there are two layers of mucin, the more compact viscous layer, closer to the endothelium, and the looser outer layer, where bacteria can flourish without causing problems. However, it takes a while for mucus to fully mature, and so if you have an environment of reduced production with increased degradation, then normal inhabitants of the gut can adhere to and invade the enterocytes or contribute to dysfunction in another capacity. Of note on this slide, is another important concept, that of the overall balance of the microbiome. We are not looking for a single bacterium, a single smoking gun, in IBS, IBD or any of the other conditions we’ll be getting to. It is the balance, as partially explained here, by the fact that almost all bacteria only produce enzymes to degrade only a part of the mucous, it is the overall balance that leads to health or dysbiosis. And generally, we know who the good guys, and not-so-good guys are, as you’ll see in the slides to follow.

Slide 11 – In this slide, we have our first in-vitro paper. Here the researchers took a look at the mucus-associated bacteria from normal intestinal epithelial cells, from both Crohn's disease and ulcerative colitis. They found a disproportionate increase in some mucolytic bacteria. Mean Ruminococcus gnavus were increased > 4-fold and Ruminococcus torques ~ 100-fold in macroscopically and histologically normal intestinal epithelium, of both Crohn’s and UC. The most abundantly detected mucolytic bacterium in healthy controls, Akkermansia muciniphila, was reduced many folds in Crohn’s and UC. I will tell you that R. gnavus and R. torques, which have since been reclassified to Blautia by the way, are generally speaking, problematic, while A. muciniphila is a superstar species for health. And if you remember back to our previous slide, the protective tips of the mucus, so to speak, are the sialic acid and sulfate groups. Both R. gnavus and R. torques have been shown to possess the enzymatic machinery to cleave these groups. On the right side of the slide, you’ll see that in vitro, R. torques were able to degrade human mucus significantly better than the rest. Also note the ability of Bifidobacteria to degrade mucus, a point we’ll revisit in this webinar.

Slide 12 – In this recent large study from the Netherlands, there are several findings to cover, some of which illustrate the diagnostic ability of shotgun metagenomics, the type of analysis we utilize. For example, it was found that proteins involved in iron uptake were increased for species like yersinia pestis and E Coli, both of which are opportunistic pathogens. This is not a lecture on iron, but when ill, your body tries to sequester iron away from pathogens, which thrive on it. Another interesting metagenomic finding was that patients with IBD or IBS showed an increased abundance of virulence factors in their gut microbiota. Virulence factors contribute to the pathogenic potential of bacteria through several mechanisms, including increased adhesion of bacteria to the gut mucosa, immune system evasion, or suppression of the host immune response. Other more basic findings from here, some of which you’re probably used to seeing by now, was a decrease in butyrate-producing bacteria, and increases in opportunistic pathogens such as Actinomyces, Streptococcus, Blautia, Escherichia, and Shigella. Lastly, as the text to the left of the slide illustrates, we have a couple of species from Bacteroides, who across all data have conflicting results. Here they were increased in both Crohn’s and UC.

Slide 13 – In this large European study, you’ll see some of the taxa with which you’re becoming familiar. As you can see from the figure to the right, those taxa we often see associated with health, are prevalent in the healthy controls and differ significantly from the Crohn’s subjects. These healthy taxa include Ruminococcus, Coprococcus, Roseburia, Faecalibacterium, and Parabacteroides. Conversely, those consistently associated with dysbiosis, are listed at the bottom, including the now very familiar Escherichia genus, as well as Blautia and a newer one for you, Fusobacterium

Slide 14 – Fusobacterium was also high in this European study. It is a genus within the phylum Fusobacteria and is associated with colorectal cancer, as well as being responsible for the majority of cases of acute appendicitis. So as you look at your report, and if you see high levels of Fusobacteria on your phyla report, or high levels of Fusobacterium, or the species F. nucleatum in the individual scores, you’ll now know you have a potential opportunistic pathogen prevailing. This is just one of several taxa, which, when given the right environment, can become problematic. So again, we’re not focusing on looking for one bug, but the overall balance. And in regards to that balance, this study had some of the usual findings, such as a reduction in healthy butyrate and propionate producing bacteria, names like the species Akkermansia mucinaphila and E. rectale, and the genus Roseburia. A key species in diarrheal condition, R. bromii, was also found to be low. Very few of the good bacteria we’ll discuss are available as probiotics, but even when they are, their use is conditional, as you’ll see. We drive beneficial changes, through the use of the appropriate prebiotic, dependent on your results. So, if you are like the subjects here, and you suffer from diarrhea, then we’d increase your R. bromii with resistant starch type 3.

Slide 15 – And here, we have more potential opportunistic pathogens: Actinomyces, Eggerthella, and Streptococcus -- all of which were shown to be significantly increased in ulcerative colitis vs. healthy controls. The genus Streptococcus does contain two species used as probiotics but also contains several others which can behave badly. Of interest, Bifidobacteria was increased in the UC subjects in this study. That’s the second time we’ve seen that in this webinar, so let’s explore it.

Slide 16 – Now, when we think of Bifidobacteria, which is used prolifically in probiotics, we think of health. In fact, in our keystone taxa section of the report, it’s there listed as essentially universally beneficial. With active ulcerative colitis, this appears to be the exception to the rule. As we can see from the figure to the right, among other findings, Bifidobacteria were significantly increased in the mucosal biopsies from active ulcerative colitis patients. The overall data is mixed on this, but this does make sense. You have to realize that mucus contains many of the same compounds as human milk oligosaccharides. If you recall from our webinar entitled, “let’s start at the beginning,” Bifidobacteria is the premier genus in degrading these HMOs. Now, this is an important point, the human milk oligosaccharides and mucus both contain the compounds galactose, sialic acid, fructose, and N-acetylglucosamine (aka NAG). So, in the active UC patient, with compromised mucus structure, it makes sense that Bifidobacteria would be drawn to their favorite fuels. The real question is, do they hurt or help. We sense that if they are degrading an already compromised mucus layer, then that can’t be beneficial. Instead, we want to focus on increasing other bacteria, in particular, ones which increase butyrate.

Slide 17 – Like in this Japanese study in Crohn’s. We see here a very typical finding, not just for IBD, but for many other conditions, a state of reduced butyrate production. We’ll cover butyrate, other short-chain fatty acids, and just about everything we touch on, in more detailed webinars down the road. With that said, butyrate is a very important compound within the gut, and is not only great fuel for the cells that line the colon, it’s also a very beneficial signaling molecule, like we saw earlier in increasing the synthesis of mucin, or in other aspects such as increasing regulatory T cells, which are immune cells that dampen inflammation. And as you can see on this slide, a very familiar face, F. prausnitzii, was once again decreased in disease vs. healthy control. So let’s take a little closer look at this very beneficial bacteria.

Slide 18 – Now in IBD, we usually associate F. prausnitzii with Crohn’s, as there is a tremendous amount of data showing its reduction in that disease. However, there are a good number of studies which show that it’s reduced in ulcerative colitis as well, such as this one. Here you can see its associations with remission, relapse, and ultimately with fecal calprotectin, which is a biomarker of gut inflammation.

Slide 19 – In fact, these researchers went so far as to use an index, which incorporates probably the two most classical species, on opposite sides of the health spectrum, those being F. prausnitzii and E. coli. Using this index, they were even able to distinguish between subsets, purely off of these two bacteria, of the hundreds present in the gut. This is not to say that this is consistently reproducible but highlights the importance of these species.

Slide 20 – On this busy slide, on the bottom right, you’ll see again the index we were just discussing. And here it was able to distinguish between healthy subjects, IBD in remission and active IBD. Note on table 2, that almost all significant differences in this study were in active-IBD vs. healthy controls. So does this mean that if you use our analysis, you should wait to be in a flare? No, although the differences are more pronounced in a flare, in most studies, and with our technology, we’ll be able to identify the microbiome. One other point from this paper is they showed an increase in fecal Lactobacillus in active-IBD. This is another important take-away from this webinar. Because if you recall from the last webinar on IBS, Lactobacillus had mostly negative associations. Well, we just finished our meta-analysis on IBD, and found the controversial finding, that when there was a statistically significant difference found, Lactobacillus almost always had a negative association. This is not to say that there is no positive data on Lactobacillus probiotics in IBD or IBS, but that it is a very mixed bag at best, and we feel that by far, prebiotics should be used, and in general, Bifidobacteria species only in IBD and IBS, with the exception of active ulcerative colitis. The last point from this slide is that of bile acids, which again is a topic for a later webinar. But I just want to point out, that dysbiosis results in significant differences in primary bile acids and their metabolites. This is important, as it has consequences in inflammation, anti-bacterial activity, and for all intents and purposes, bile acids are hormones which initiate an array of functions.

Slide 21 – You’ve probably noticed that some of these studies use fecal data, and some use biopsy data. Here’s a study that used both, which can highlight a point or two. If you recall, there’s split data on whether a fecal analysis is an accurate picture of the mucosal microbiome. Some studies show no difference between fecal and biopsy samples, and some do. It really can depend on where the biopsy occurs, and in whom. For example, here we have both Crohn’s and ulcerative colitis subjects, but biopsies were only done for Crohn’s. So if we look at the figure, in what’s labeled as ICD – which stands for ileal Crohn's disease, which is your typical case, you’ll see two classic markers, F. prausnitzii, and Enterobacteriaceae, which is a family that contains many opportunistic pathogens such as E. coli. You’ll see, significantly lower F. prausnitzii in the feces, and the ileal biopsy, but no difference in the colon. The same is also true for Roseburia, a very beneficial butyrate-producing genus. And the same is true for the bad family, Enterobacteriaceae, they are significantly higher in the feces of Crohn’s subjects, and at ileal biopsies, but not in colonic biopsies. So what does this tell us? That essentially, if you have classical ileal Crohn’s, we don’t need an invasive biopsy to pick up on dysbiosis. And now with better technology, and as is also the case in several other studies for ulcerative colitis, IBS and more, we’ll pick up on what’s needed from fecal samples.

Slide 22 – So let’s start to put together the taxa we keep encountering. We have here a great figure illustrating the taxa with the positive and negative association in this pediatric Crohn’s cohort. All subjects were newly diagnosed, before treatment, and fecal, and biopsy samples were taken. What we see are some classic results, which we keep seeing in the literature over the past 15 or so years. Up top in red, you’ll see the negative associations with two you know, the genera Escherichia and Fusobacterium, and two new ones, Haemophilus, and Veillonella. Haemophilus, along with Escherichia, Salmonella, Citrobacter, Enterobacter, Klebsiella, Shigella, and Yersinia are all members of the family Enterobacteriaceae, which is a subset of Gammaproteobacteria, which is a class within Proteobacteria. So, if on your report, you see any high levels of Proteobacteria in your phyla report, or high levels of the class Gammaproteobacteria, or high levels of the family Enterobacteriaceae, then you most likely have high levels of one or more of the many opportunistic pathogens within their classification. Looking at this is one of the most efficient ways to judge your microbiome. Beyond those, we also show here Viellonella, which is in Firmicutes, and generally has a lot of negative associations in the data, like here. The last of the ones in red are Fusobacterium, which we covered earlier. The positive associations shown in green contain some beneficial taxa we’ve covered before, such as Faecalibacterium, Coprococcus, Ruminococcus, and Parabacteroides. And it contains some genera with conflicting data such as Dialister and Dorea

Slide 23 – On our last working slide, we’ll look at things at a phylum level on the left side of the slide. In your report, one section is much like this, but looks just a little different, but tells the same information. How your microbiome looks at a birds-eye view, at the phylum level. Here I have a couple of key takeaways for you. One is in regards to the Firmicutes: Bacteroidetes ratio. If you’ve never heard of this, then perfect-- because it’s a source of misinformation. After personally reading well over 1,000 research articles on the microbiome alone, I can tell you that there is about a 50/50 split in the ratio being skewed in one direction, or the other, in disease vs. health. 

What’s much more important is to take take a deeper dive within these two phyla, as they both contain healthy bacteria and potential trouble makers. My second key point is that there are phyla from this birds-eye view which do provide information of value. Both Proteobacteria and Fusobacteria are two phyla we don’t want a lot of, and after this webinar, I think you can understand why a bit more. So if you look at the bars, you’ll see that Proteobacteria are increased in ulcerative colitis vs. health control, and even more so for Crohn’s. In Fusobacteria, the increase in Crohn’s is more subtle, but that’s to be expected. Most of your trouble makers are in Proteobacteria. So when I look at someone’s report, in this section, I look straight-away at these two phyla, as the others don’t tell me anything at that level. And to the right, you’ll see once again many of the usual suspects, and so I’ll throw two new names at you, Prevotella and Alistipes. These two genera have positive associations in disease most of the time. You may recall Prevotella from IBS, where if you have constipation, we want to increase it. Well, it’s also good for IBD, which are classically conditions involving diarrhea. There is no supplement available for either of these two genera nor for most all taxa I’ll mention. They are a function of what you ingest and other lifestyle factors.

Slide 24 – So in review, we covered a few concepts, and shared some novel findings. We discussed how, if left unchecked, both Crohn’s disease and ulcerative colitis can result in some terrible manifestations. The basic underlying flaws behind each condition are known, through extensive research over the years, as are the basis on which bacteria in the gut are generally good for health and those not so good for health. Science has connected many the dots between the bacteria, their by-products, their actions, their source of fuel, and how they impact our cells biochemically and ultimately our health. I think you’re now beginning to understand that we’re looking at the overall balance within the gut, not simplifying it to one or two bad-players, but a look at the bigger picture. And most of the beneficial bacteria, do not exist as probiotics; thus we have to drive their abundance by providing them the fuel they need to thrive, via diet, but more importantly in dysbiosis, prebiotics. For those that do exist as probiotics, we have to reconsider their use and be somewhat more discriminate. As we have seen now with both IBS and IBD, species from Lactobacillus may not be a good choice, as the overall microbiome data is not pretty. Bifidobacteria, on the other hand, are almost always universally beneficial, except active ulcerative colitis. So we really have to be more precise with our interventions, and that’s why we are here. 

If you’re interested in more information on IBD, use this hotlink to connect to the webinar I mentioned at the beginning.

Next, we’ll dive into the connection between the microbiome and mood, which will encompass primarily anxiety and depression. So until next time.