GI Map Tutorial with Dr. Tom Fabian: Rational Wellness Podcast 182

Tom Fabian, PhD discusses How to Interpret the GI Map Stool Test with Dr. Ben Weitz at the Functional Medicine Discussion Group meeting on October 22, 2020.

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Podcast Highlights

2:01  Quantitative PCR vs Whole Genome Sequencing.  The GI Map stool test uses quantitative PCR, which is the best way to detect specific, clinically relevant bacteria or other microorganisms. It also tells you specific quantities, which is important to detect overgrowth and to allow you to see if levels are increasing or decreasing over time.  It can also allow you to detect microorganisms, like H. pylori, that are in pretty low concentrations.  Whole genome sequencing technology is better for getting an overall picture of the microbiome, but it is not good for clinical diagnosis and does not include any physiological markers, like calprotectin.  While quantitative PCR is highly accurate, the ability to detect something comes down to the choice and the design of the primers. Primers are designed for specific, individual strains or organisms.

12:50  The team at Diagnostic Solutions has designed the GI Map stool test with a focus on both research and clinical experience and it helps clinicians to understand the relationship between physiology and the gut.  You can see three common dysbiosis patterns: 1. Insufficiency dysbiosis, 2. Inflammatory dysbiosis, and 3. Digestive dysfunction.  Insufficiency dysbiosis is a lack of beneficial bacteria. And there can be a combination of these three.

15:56  There is some confusion that stool tests do not provide information about the upper gut but only about the lower gut/colon.  But this is not true.  There are a number of bacteria on the GI Map stool test that are more common in the upper gut, including H. pylori, which grows predominantly in the stomach, and the bacilli class of the Firmicutes phylum that includes species like streptococcus, lactobacillus, enterococcus and staphlococcus, which grow predominantly in the small intestine.  These species thrive in higher oxygen environments like the small intestine, whereas the colon is more anaerobic. Bile is present in the small intestine and the species mentioned above are also bile tolerant.  The Proteobacteria phylum, which we tend to think of as inflammatory microbes, are also much more prevalent in the small intestine than the large. Pseudomonas is linked to inflammation in the duodenum and it is also linked to issues with gluten and other food sensitivities.  Also, pseudomonas thrives on simple sugars and amino acids, which are prevalent in the small intestine.  They are also very bile tolerant and you can find these in the bile ducts and in the gall bladder.  Some of the data as to which species thrive in the small intestine comes from Dr. Mark Pimentel’s group, which mapped the microbiome of the small intestine, the Reemagine Study, which was published earlier this year.  [Mapping the Segmental Microbiomes in the Human Small Bowel in Comparison with Stool: A REIMAGINE Study]  

In the colon, the Clostridium class of the Firmicutes phylum tends to dominate and one of the key species is Faecalibacterium Prausnitzii, which is a major butyrate producer.  The Bacteroides group of the Firmicutes phylum is a major group in the colon. Akkermansia is a well known species in the Verrucomicrobia phylum, which is also one of the key groups in the colon. They thrive in an anaerobic environment and tend to thrive on complex fibers, as well as the mucus ecosystem, and the mucus layer in the colon is much thicker than in the small intestine.

22:17  Here are a list of organisms seen on the GI Map that are predominantly in the upper GI Tract: 1. Enterotoxigenic E. coli, 2. Vibrio Cholerae, 3. Yersinia enterocolitica, 4. Cryptosporidium, 5. Giardia, 6. Norovirus, 7. H. pylori, 8. Lactobacillus, 9. Bacillus, 10. Pseudomonas, 11. Staph, 12. Strep, 13. Citrobacter, 14. Klebsiella, 15. Fusobacterium, 16. Candida. 

Here are a list of microorganisms on the GI Map that are predominantly in the lower GI tract/large intestine:  1. C. difficile, 2. Enterohemorrhagic E. coli, 3. E. coli 0157, 4. Enteroinvasive E. coli/Shigella, 5. Entamoeba histolytica, 6. Bacteroides fragilis, 7. Clostridia, 8. Akkermansia mucinophila, 9. Faecalibacterium, 10. Methanobrevibacteriacea, 11. Microsporidium, 12. Blastocystis hominis, 13. Dientamoeba Fragilis.

25:47  H. pylori’s presence can be clinically relevant, esp. if it’s high, even without any virulence factors present.  Tom said that the presence of H. pylori is often associated with reduced stomach acid.  If the virulence factors are present, the two that stand out as most significant with the most research are cagA and vacA.  Virulence factors tend to work together, so the more that are positive, the more likely they will be significant.

33:26  Dr. Sam Rahbar, Integrative Gastroenterologist in Los Angeles, asked if bacteria like Klebsiella and Citrobacter fruendii are found in the stool, should these be seen as markers of dysbiosis or are they potential pathogens that should be targeted for treatment? Dr. Fabian explained that they are opportunistic pathogens that can exist in the gut without any effects or can potentially be part of a pathogenic process. For example, Klebsiella can be associated with certain autoimmune conditions, including inflammatory bowel disease, psoriatic arthritis, and ankylosing spondylitis.  [Here is one article discussing the connection between Klebsiella and Crohn’s and Ankylosing spondylitis:  The Link between Ankylosing Spondylitis, Crohn’s Disease, Klebsiella, and Starch Consumption.]  While a pathogen in the gut may be a trigger for autoimmune disease, it’s not yet clear whether eliminating this bug will improve the autoimmune condition. 

38:58  If a potential pathogen like H. pylori is present as either high in red or a number above the <DL that is black color on the report, whether it creates a problem for a patient depends upon the overall health of the gut and the microbiome.

43:00  If Pseudomonas is high on the stool test, that does not imply that it has colonized the large intestine. This bacteria mostly colonizes the small intestine and the stomach. 

43:57  If the secretory IgA level on the GI Map is low, that is an indicator that the bacteria in the microbiome that produce butyrate, like Faecalibacterium, are at low levels.  While it is a good idea to build up these beneficial butyrate producers in the microbiome with probiotics or prebiotics and more fiber in the diet, that can take some time. In the short term, some clinicians will recommend supplements like colostrum, Saccharomyces boulardii, L-glutamine, and even vitamin A to increase secretory IgA levels.

46:06  Calprotectin is a marker of inflammation in the colon.  If calprotectin is low and the patient appears to have a lot of gut inflammation, the inflammation is probably in the upper GI tract.  For example, Pseudomonas has been shown in research to cause inflammation in the upper GI tract. Candida is inflammatory and if often colonizes anywhere in the upper GI tract, even the oral cavity.  If you have H. pylori, which can cause inflammation in the stomach, you will not see an increase in calprotectin.

52:14  Inflammatory Dysbiosis.  There are particularly inflammatory bacteria in the Protebacteria phylum and some are in the subclass called Gamma Proteobacteria.  These include: E. coli, Salmonella, Shigella, Vibrio cholera, Yersinia enterocolitica, Enterobacter, Morganella, Pseudomonas, Citrobacter, Klebsiella, and Proteus. A lot of them have been shown to secrete the inflammatory type of Lipopolysccharides.  Dysbiosis in the mouth, like with periodontitis, leads to the increase of pathobionts like Klebsiella and Enterobacteria, which can then translocate to the gut.  If bacteria like Klebsiella are able to colonize the gut, this can activate the inflammasome.  If there is a lack of beneficial bacteria in the gut, then bacteria like Klebsiella will be more likely to colonize.  When you see Klebsiella, you should look at the patient’s oral health. You also want to look at indicators of low stomach acid, such as H. pylori or are they taking proton pump inhibitors.

55:48  Digestive Dysfunction Dysbiosis.  Elastase is a marker for pancreatic insufficiency.  If we see H. pylori, then often stomach acid is low. Also if we see overgrowth of Pseudomonas, Enterococcus, or Staph or Strep or if we see Faecalibacterium low, these indicate that stomach acid is low.  Research suggests that if Kebsiella is overgrown, this is a bacteria that often comes from the oral cavity or the respiratory tract, esp. if there is low stomach acid.

1:16:33  Are Blastocystis Hominis and Dientamoeba Fragilis pathogenic organisms (parasites) that should be treated or normal parts of the gut that should not be treated?  I recently interviewed Dr. Jason Hawrelak, well respected Australian Functional Medicine Doctor and expert on gastrointestinal disorders and the microbiome, and he said that these protozoans are extremely common and are generally irrelevant to that person’s symptoms. [Rational Wellness podcast episode 169 on Gut Parasites with Dr. Jason Hawrelak.] Dr. Fabian explained that Blasto and D. Fragilis are not commensal, since they are not present in the majority of patients tested. When we see them present, they are often part of a digestive dysfunction pattern. These protozoans are often seen in the context of things like H. pylori, candida, and general overgrowth. 



Tom Fabian, PhD is an educator and medical consultant with Diagnostic Solutions Laboratory to help clinicians learn to better interpret the GI Map Stool test. Clinicians can sign up for a professional account to order the GI Map stool test by going to DiagnosticSolutionsLab.com

Dr. Ben Weitz is available for nutrition consultations, including remote consults via video or phone, specializing in Functional Gastrointestinal Disorders like IBS/SIBO and Reflux and also specializing in Cardiometabolic Risk Factors like elevated lipids, high blood sugar, and high blood pressure and also weight loss, as well as sports chiropractic work by calling his Santa Monica office 310-395-3111 or go to www.drweitz.com. Phone or video consulting with Dr. Weitz is available.



Podcast Transcript

Dr. Weitz:            Hey, this is Dr. Ben Weitz, host of the Rational Wellness Podcast. I talk to the leading health and nutrition experts and researchers in the field to bring you the latest in cutting edge health information. Subscribe to the Rational Wellness Podcast for weekly updates, and to learn more, check out my website, drweitz.com. Thanks for joining me. And let’s jump into the Podcast. Our topic for tonight is we’re taking a deep dive into the GI MAP stool test.  Tonight, we have Tom Fabian, PhD of Diagnostic Solutions and so I’m sorry for everybody who’s missing the [Biden vs Trump] debate.  And I’m sure there’s going to be a lot of poop discussed, but we’re going to have a much higher level discussion of poop tonight. So Tom, thank you for joining us tonight and I’m sure we all have a lot of questions about our favorite functional medicine stool test, and so we hope to develop some clinical skills to be able to better help our patients.

Tom Fabian:       Absolutely. Well, thanks so much Dr. Ben for inviting me to join your discussion group. And I look forward to interacting with everyone here, and hopefully giving everyone a better understanding of GI MAP and how to better apply it in clinical practice. All right.  So I’m going to go ahead and fire up the PowerPoint here.  So first I’ll just share my screen.  So tonight we’re going to be focusing on a deep dive into GI MAP. So this presentation is meant to be flexible since this is a very interactive group.  So Dr. Ben, if you have any questions in the meantime, definitely feel free to interrupt, or if anyone has anything that I’ve found over, they don’t really understand. I’m happy to stop and we can just cover whatever we can cover in the amount of time.

Dr. Weitz:            Maybe I’ll ask just one quick question since there’s a lot of different types of stool tests that are out there. Can you talk about the difference between the technology used for the GI MAP stool test, which is quantitative PCR, and how this is different than say whole genome sequencing, stool testing?

Tom Fabian:       Sure. Yeah, so they are different technologies for different purposes and you get different types of information. So starting with meta genome sequencing–that’s a DNA based method as is qPCR, at least the type that GI MAP is using. So we’re looking at detecting the DNA, for specific organisms or for genes, specific genes. So for example, in the instance of toxin genes, you can basically detect any aspect of DNA either way with either type of test but that’s kind of one of the areas where the similarity stops. So metogenomics is used, especially if you look at research for kind of a broad overview, a survey of the microbiome and the genes of the microbiome encodes to get a sense of the overall community. So there’s a lot of ongoing research in that area to figure out of the hundreds of species that you can potentially detect with that method. What does that actually mean? So that’s a lot of ongoing research and there’s really not a whole lot of conclusions we can make definitively about most of the species.  A lot of them just have not been well studied yet, of course we’re learning more every day.  So that’s one of the advantages of sequencing as you can potentially get kind of a more high level, bigger picture of the ecosystem.  But the challenge is if you want to measure specific clinically relevant organisms particularly quantitatively, so you can see what their levels are, whether they’re increasing or decreasing over time and with treatment, you really need a more quantitative method.  And especially when you look at sort of the composition overall of the microbiome, so you have very abundant species all the way down to species that are barely detectable. So there’s a really wide range of abundance levels for different species. Some of the species, in fact, quite a few of them on the low end of that range are clinically significant classic example would be H. pylori. H. pylori is mostly in the stomach by the time it’s in a stool sample, it’s usually pretty low concentration if it’s present.

                                So that’s difficult to pick up these low abundance species in typical kind of commercial based sequencing, because the ability to detect these low abundance organisms and to detect them in a way that’s at least reasonably quantitative depends a lot on the number of sequencing reads. So you may have heard the terms, sequencing depth, sequencing reads and that’s expensive to do that. So most of the commercial type testing doesn’t really go that deep. So you get a picture of the more abundant ones at the top, and that may include some of the ones in the middle, but oftentimes cannot identify the ones that are on the low abundance and of things very accurately. So that’s really where qPCR shines. In fact, it’s kind of like a technology that you want to use to find those needles in the haystack. And I’m actually going to go through a few examples here in this presentation. And I do have a couple of slides about qPCR as well, I can elaborate on that just a little bit, but that’s one of the main differences. Also a lot of the metagenome sequencing companies. In fact, I’m not aware of any right now that also include physiological markers. So to really get some insights clinically into what’s going on in the gut, you need to know both sides of the equation, microbiome, as well as what’s going on with digestion inflammation and things like that.  So you get a more complete picture, even though you have fewer markers because these types of tests focus on organisms that have been established to be clinically relevant and also these physiological markers. And then with that careful quantitation, then you can start to really see patterns and trends and things like that. And I kind of like it to say, you’re just looking at blood glucose levels on our standard lab test or a hemoglobin test. If all you got was plus or minus, higher or low, or sort of high, medium, low, that gives you a kind of a rough idea of what the levels are, but hemoglobin A1C of 5.6 is very different clinically from say 7.5. Although, in some measures, both of those might be considered kind of on the high end. So quantitation really is important in that’s part of a really a big part of what makes a marker more clinically relevant. So that’s kind of, sort of at a high level tip of the iceberg. What’s some of the key differences are.

Dr. Weitz:            Okay.

Tom Fabian:       Technologies.

Dr. Weitz:            Good.

Tom Fabian:       Okay.  You can tell from the title here. So we’re going to really focus on clinical insights, how to get some clinical insights out of GI MAP, particularly by connecting the dots.  So that’s really where the more advanced users get the most clinically relevant information out of GI MAP has learning how to connect those dots on the test with what’s going on clinically as well. And then of course, that’s all the point of all that is so you can get better clinical outcomes by really understanding what’s going on in the gut. So really in terms of connecting the dots, you have to know something about the dots, and which ones are most important and then how to connect them. And that really has to do with the microbial ecosystem as well as GI physiology and those two interact heavily. I’m going to go through a couple of examples in this presentation as well for basically illustrating how those dots are connected and can really yield some great clinical insights.

                                So in terms of qPCR which is also known as quantitative PCR or real-time PCR, so sometimes we’ll see ages RT-PCR on the purpose of that, the reason it was developed in research and is also used a lot clinically now is it’s really good for highly accurate identification and quantitation of microbes. I have heard some sort of… When you look at social media and you look at some of the comments that are out there, there’s a lot of misunderstanding of methods. So I’ve heard some sort of competing companies and other prominent folks that are out there in the community say that GI MAP uses 16 sequencing, which is pretty far from what we actually use. So we actually again use qPCR, quantitative PCR, but that it’s ability to identify something accurately comes down to the choice and the design of the primers. You can design primers that are highly specific for an individual strain, for example, you can detect right down to the strain level, for example, that’s how we detect pathogenic strains of H. pylori, C. diff, some of the other pathogens, is based on detecting them at the strain level.  So again, it’s especially good for low abundance microbes, there’s also the issue of absolute versus relative quantitation and it’s a little bit technical. I don’t want to get into that right now, unless anyone has questions, but absolute you’re looking at how much of that organism is present per say, gram of stool versus sequencing, which is all relative to what else is there. That could be a problem because if something goes up, that can make it look like something else is going down, when in fact it may be the same level, it’s just that it’s all relative. So it treats every sample as if it’s the same amount.  I do have one quick figure here to kind of illustrate that. And that’s actually in this figure from this particular article, which is quantitative microbiome profiling links gut community variation to microbial load. So it was published in a prominent journal, Nature, a couple of years ago where they compared relative versus quantitative approaches and they actually found that the absolute quantitation, especially when they looked at a Crohn’s disease patient was extremely important because without looking at absolute quantitation, you miss the whole picture of general decrease in microbes in the gut. And that’s a key feature of for example, Crohn’s disease. So I’ll look at… Show that data here in the next slide. So this is a pretty complicated diagram, but the main point here is on the left is a different samples that basically where the data is expressed in a relative sense. So every sample is treated as if it’s the same amount, starting amount, and all the different taxa indicated by different colors. So all the different types of microbes make up percentage of that pie, but you don’t really know if in person A, that the overall levels have gone down say F after antibiotics or after an infection unless you look at it from an absolute standpoint, and that’s what you see here. This is the breakdown, also expressed in terms of total amount per gram. You can see here on the side, it says per 10, they live in cells per gram.

                                So essentially you can see that they’re some individuals that have quite a bit less overall sort of total microbiome in their gut. And they found that to be very clinically significant. That’s one of the big difference between sequencing and qPCR. A qPCR is basically looking at things from an absolute standpoint. So the big question there is really why is that important? And again, I kind of alluded to some of those important clinical relevance type aspects, but it really allows you to differentiate low, normal and high levels at a much more detailed level.  So you can identify trends and patterns and that’s a big part of the connecting the dots with GI MAP. And that’s also critical for assessing the clinical relevance and your treatment approach, because you want to see if you do a retest, that organism may still be there, but it may be responding well, you may have gone down a couple of orders of magnitude in quantity showing that your approach is working. That’s really where quantitation is important.

The other thing that we bring to the table in terms of kind of a differentiator for GI MAP is, how we present our interpretive and educational information. So our staff has a really wide range of expertise, industry experience in Functional Medicine, clinical testing, several PhDs, et cetera.  And so we bring all that kind of cross disciplinary expertise to the table, particularly with clinical experience mixed with research and we really focus a lot on the latest research so that we can focus on these interactions, and GI MAP is really all about the second part of the equation helping clinicians understand the potential interactions between the microbiome and gut physiology.  So a key thing to understand here is there’s a lot of different aspects of physiology and the gut that really come into play once we understand them at a certain level that can help us understand how to interpret a stool test and what it may mean as far as these underlying issues. So we may, for example, see that, you’ll see here on the list pH is one of the major factors throughout the gut that affects the microbiome and vice versa, same with oxygen, the first nutrients from the diet as well, flow rate digestion, especially stomach acid and enzymes, immune factors, et cetera.

                                So a lot of factors can affect the interaction between the microbiome and our health. And that’s really reflected in GI MAP through three common dysbiosis patterns that we’ve identified over time, just in our overall experience with GI MAP, so those really boil down to 1. Insufficiency dysbiosis, which is a lack of beneficial bacteria, so that’s important to understand how to identify that on GI MAP, 2. Inflammatory dysbiosis, again, very important for a wide variety of diseases, autoimmune inflammatory diseases, and it’s also important to understand how to identify that. And then lastly, 3. Digestive dysfunction dysbiosis, which is kind of a generic term that we use because various aspects of digestion may be compromised or reduced and have an effect on the microbiome and the microbiome in some cases can actually have an effect on digestion. So once again, I’ll go through examples of this and in many patients, they can have a combination of these types of dysbiosis that reflect that interaction between physiological imbalances and the microbiome.

                                So just to note, we do have a resource on our website, a downloadable PDF that lists what markers constitute these different patterns. We will be updating that sometime shortly within the next couple of weeks or so, and then we will make that available on our website. But we do have that as a current resource that was created a couple of years ago, so it’s just a little bit out of date at this point. The other thing that we really kind of weave into this based on research, and how that applies clinically beyond sort of this holistic high-level integrative perspective of looking at the whole gut, a big part of that is recognizing upper versus lower GI MAP microbes. That’s something that you may have heard.

                                Some clinicians claim that with stool testing, you can’t get any insights at all, except into what’s going on in the colon that you basically don’t really understand from a stool sample what’s going on in the small intestine for example, it turns out there’s a lot that you can gain from that. You don’t get necessarily a completely accurate picture of the composition in compartments that are sort of higher up in the GI tract, but sort of the classic example is H. pylori.        We didn’t know that that is detectable in stool, and that certainly yields important clinical information with regards to what may be going on in the stomach, so think of this as kind of a domino effect that you want to get a bit of an understanding of what’s going on higher up in the GI tract, even as high up as the oral cavity.  And one of the examples I’ll go through that’s features one of the markers on GI MAP, for a lot of patients that may actually come from dysbiosis originally in the mouth. And then you want to understand how these physiological imbalances result in its presence in the gut and what it may be doing there.  So to really get into this a little bit more, it’s important to understand a little bit about the composition of the upper GI MAP microbiome.  It’s not as well studied as the lower GI, the colon microbiome, but we’re learning much more about it especially in the last few years.  Mostly what’s dominant in the upper GI from the mouth down to small intestine are a large class in the Firmicutes phylum called the Bacilli.  So that species and groups like streptococcus, lactobacillus, enterococcus and staph. We do have these groups on GI MAP when you see them overgrown, it’s likely that they may reflect imbalances in the upper GI tract. And there’s some just basic information about the physiological conditions that they thrive in, they like higher oxygen environments.  So they don’t like the anaerobic environment in the colon, unless the colon environment’s been disrupted.  They like simple nutrients that are highly available in the small intestine.  Bile is in the small intestine, so they’re bile tolerant, et cetera,.  Also, organisms from the Proteobacteria phylum, which we tend to think of more of the inflammatory microbes, the dysbiotic microbes, they’re much more prevalent as a percentage in a small intestine. So typically they constitute say 20% of the microbiome in the small intestine and only 2 to 5% in the large intestine. So proportionally they’re much smaller in the large intestine.

                                A lot of the research has come out on Pseudomonas, which you do have on our tests, actually going to go back to that, linking Pseudomonas to inflammation in the duodenum, for example, and linking that to issues with gluten and kind of the broader picture of food sensitivities. They also thrive in a higher oxygen environment, also thrive on simple nutrients like sugars and amino acids, highly bile tolerant, you can actually find many of these in the bile ducts and in the gallbladder.  So they’re very bio tolerant, so really important group in the upper small intestine.  Composition of lower GI microbiome at a high level, the most abundant groups in some ways fairly simple, the Clostridium class really dominates and that’s the..in the colon, the largest class or largest group in the Firmicutes phylum, and one of the key species is Faecalibacterium Prausnitzii which we do have on GI MAP as well. That’s a major butyrate producer, again, it’s in the Clostridium class, which is the dominant group in the Firmicutes phylum. And then there’s a Bacteroides group, which is the major group, typically in most people in the bacteria Firmicutes phylum, Akkermansia is a well known species in the Verrucomicrobia phylum, which is also one of the key groups in the colon. They thrive in an anaerobic environment, generally that whole ecosystem, at least in a healthy individual thrives on complex fibers, as well as the mucus ecosystem.  So the mucus layer in the colon is much thicker than in the small intestine. So that’s one of the reasons why there’s a more significant you can see ecosystem in the colon. So this is data from Dr. Mark Pimentel’s lab that was published just earlier this year. This is just basically showing… You can see at the bottom here on the X axis duodenum, jejunum, FD, which is their term for the furthest distance the scope could go in the ileum. So it essentially means ileum for the most part compared to a stool sample. So the take home point here is the different colors represent the different major groups in the different compartments here, but it’s fairly consistent across the small intestine and then suddenly you have this huge change in the large intestine, which is reflected by the fecal sample. So that’s really reflecting that big change in physiological parameters from the small intestine, which again has a higher oxygen content, et cetera, usually lower acid levels, higher pH, then the colon very different environments particularly because of the fact that it’s anaerobic.  So that’s why anaerobic species thrive there that ferment carbohydrates. So a big difference and that’s really key to understanding this sort of localization aspects to interpreting stool tests. And this is the Tyler study mapping, the segmental microbiomes in the human small bowel in comparison with stool, the Reimagine study. And again, this is a study that was just published earlier this year by Mark Pimentel’s lab. [Mapping the Segmental Microbiomes in the Human Small Bowel in Comparison with Stool: A REIMAGINE Study]

Dr. Weitz:           Yeah. We were lucky enough for Dr. Pimentel to join us several months ago.

Tom Fabian:       Oh, you did? Okay, great. I have to go back and make sure I check that out.

Dr. Weitz:           Yeah.

Tom Fabian:       So this is a summary of markers on GI MAP based on research that indicates where they’re predominant in the GI tract. We just sort of broke it down between upper GI and lower GI, with the upper GI being on the left and then the lower GI being on the right. So not all species or groups are known in terms of research that’s available in terms of where their predominance is, and then in other cases, they may be prevalent in both but both of these groups here research supports the predominant primarily either in the upper or the lower GI tract. So once again, there’s quite a few markers on GI MAP that are likely reflecting what’s going on in the upper GI. Again, the classic one being H. pylori, GRD is another, well-known microbe that basically thrives in the upper small intestine. The two that I’m going to focus on here just briefly in this presentation are Klebsiella and C. diff. So one example of an organism that’s present more commonly in the upper GI and also the oral cavity and then less prevalent in the lower GI except in cases of potential inflammation.

                                So as far as stool composition, just to kind of summarize it primarily reflects the colon composition. So that’s sort of intuitive, but also that’s what research shows as well. And of course within the column, there’s a mix of luminol and mucus associated microbes, with the composition primarily dominated and fecal samples with luminol microbes. But definitely you get a sense of the mucus associated microbes such as Akkermansia. It’s also dominated by anaerobic Fermenter bacteria in particular, the Clostridium class in the Firmicutes phylum and then the Bacteroidetes genus in the Bacteroidetes phylum. Those are far and away, the two most dominant groups in the microbiome in most individuals, at least in Western countries. Microbes from the upper GI tract can be present in stool, but usually in really low numbers.

                                So that’s of course where qPCR comes in, because we can still get some good insights based on the presence of some of those low abundance type organisms. So once again, this is just sort of back to the figure from the Pimentel study showing the breakdown of these major groups across the small intestine and stool. Of course, the question is why we’ll get into some of that a bit more as well. So that guy somehow made a mark there. So really, I think it’s important to consider since we can detect potentially some of these upper GI microbes. Some of them are better established as being pretty exclusive to the upper GI tract. Again like H. pylori and Giardia. Pseudomonas, mostly research does indicate that it’s likely thriving in the stomach and the upper small intestine and so far, most studies indicate that it’s not present and are not active, usually not active in the large intestine.

Dr. Weitz:            Are you going to have time to go into H. pylori? And if not, can I ask a couple of questions now about H. pylori?

Tom Fabian:       I’m sure. Yeah. I’m happy to answer questions right now, but it’s H.pylori.

Dr. Weitz:            Okay. So if we see H. pylori present, but there aren’t any virulence factors essentially it’s my understanding that that’s not super important. Correct?

Tom Fabian:       So as far as H. pylori its presence without virulence factors, it is important apparently, we certainly see a potential clinical impact quite often when it’s present and especially if it’s high.

Dr. Weitz:            O.K., so when we look at the numbers, like the normal says 1.0 e3, now what’s considered high?

Tom Fabian:       High is generally that’s what that cutoff level is indicating. So above that number is considered high, below that number is considered detected.

Dr. Weitz:            Like if it’s 5.0 e3, is that a little high or a lot high? Is it a lot high if its e4, e5 that makes it a lot high, right?

Tom Fabian:       Right. Yeah. I mean, because those are orders of magnitude, but definitely clinically in the E three range is high and it’s of course you have to take everything into the context of the patient and how that patient’s presenting symptoms that may be related to that, but also we can see a pattern on GI MAP. And so there’s a lot of research showing, for example, the H. pylori, regardless of virulence factors likely reduces stomach acid in majority of people that have a chronic infection, and so we have an overall digestive dysfunction pattern that I mentioned briefly earlier that is often associated with H. pylori, and it’s one of the ways that we can gauge based on just the test markers and connecting the dots, doesn’t look like this is having an impact in the ecosystem and then of course the clinician has to decide, is that relevant to the patient’s symptoms and how the patient is presenting?

Dr. Weitz:           So you’re saying if we have a patient with elevated H. pylori and it seems like it might correspond with the symptoms, it might be worth treating, even if there’s no virulence factors?

Tom Fabian:       It would be worth considering, a lot of clinicians do, but I think the… When it comes to functional integrative type clinicians, a lot of clinicians will often focus more on antimicrobial approaches versus going straight to antibiotics. So it’s not a scenario that would be kind of in conventional medicine it’s the more looking at the potential virulence in relation to things like potential for stomach cancer ulcers also things like that. Some of the more serious outcomes that are tied, the risk for those types of outcomes are tied to the virulence factors. But the more mild sort of to moderate effects based on just depression of stomach acid still can be clinically relevant for patients, but it may not warrant sort of complete eradication for example.

Dr. Weitz:           And in some of the virulence factors more important than others?

Tom Fabian:       Based on research so far, the answer is yes. The two that stand out the most, partly because they’ve been well-researched, but seem to be especially required for the more significant inflammatory outcomes and that would cagA and vacA.  That’s important to point out though that virulence factors tend to work together and so generally the more you see on the test, so the more that are positive, the more likely than that could be a more significant scenario.

Dr. Weitz:           One more quick question, just in general, in terms of any bacteria or pathogen or potential overgrowth when we see the number there, but it’s less than the normal that could still be significant?

Tom Fabian:       Right. Now, again that’s really where clinical judgment comes into place, so-

Dr. Weitz:           Because it’s really well, it’s has <dL> right? Like super low.

Tom Fabian:       Exactly. Yeah. That either means it’s just not there at all or it’s presence at a low level.

Dr. Weitz:           Okay.

Tom Fabian:       We often do see that based on interacting with clinicians that their judgment is, and also based on what we see on the pattern on the test that [inaudible00:30:31] can be clinically significant for some patients, again, mostly in that context of potential for contributing to dysbiosis and reduce digestion.

Dr. Weitz:           Okay, great. Go ahead.

Tom Fabian:       So that’s really kind of… It was actually a great time to talk about H. pylori because that is kind of the classic example when you hear certain folks out there saying you can’t really get any information about the upper GI from a stool test, that’s simply not true because H. pylori is sort of the case in point. We can certainly get some great insights into what may be going on in the stomach in that case.

Dr. Weitz:            And of course, zonulin gives us some information about the upper bowel as well. Right?

Tom Fabian:       Right, exactly. So qPCR, again, just kind of that analogy that it’s great at finding these needles in the haystack. H. pylori is a good example. There are a lot of the metogenomic type tests, metagenome sequencing that generally aren’t as good at detecting H. pylori let alone quantitating it. So when you really want to know about a clinically relevant organism, qPCR is a very effective way to assess that. So let’s talk a little bit about the insufficiency type of dysbiosis. So this is something that probably a lot of clinicians who worked a lot with GI MAP have seen, you may see a really broad insufficiency like we see here for this patient where there are a lot of markers in the normal bacteria section that are deficient, at the phylum level that’s certainly one of the most important levels to look at because those two groups together make up the bulk of the microbiome, particularly in the colon.  So when they’re both deficient, that’s telling you the microbiome overall is probably deficient and then all those beneficial functions that they serve are probably also deficient. It’s also a great to focus, particularly on Akkermansia and the E. coli bacteria, and those are two Keystone species. So once again, when one or both of those are low that can be pretty significant in terms of indicating that the colon ecosystem is not so healthy. The other thing I want to note about kind of connecting the dots on GI MAP is we do lots of research.

Dr. Weitz:            Let me just see if I could throw a question in Dr. Rahbar who’s joining us tonight is a integrative gastroenterologist. Let Dr. Rahbar ask a question. Let see, I’m going to unmute you Sam, why don’t you go ahead and ask your question. Did I unmute you?

Dr. Rahbar:          Can you hear me?

Dr. Weitz:            Oh yeah. Now we can hear you.

Dr. Rahbar:          Okay.

Tom Fabian:        Yes, I can hear you.

Dr. Rahbar:         Yeah. I have several questions since we use this lab quite frequently, and I kind of use my common sense approach to it.  For example, I see sometimes practitioners wanting to treat Klebsiella and Citrobacter fruendii in the stool. And I always question, are these specific pathogens, or are these markers of dysbiosis?  It means that, do you need to look at it as dysbiotic flora or like if I got Shigella or something in the stool I would say, okay, you’re going to have some specific bug that I have to treat it.  Or campylobacter.  At least from the literature that I see, I don’t recognize these as specific pathogens of the gut, indications of disease.  So I usually respond that maybe you treat SIBO some other dysbiotic scenarios, but I wouldn’t target a specific one, but Dr. Fabian, I wanted to get your opinion on this one without just saying something. 

Tom Fabian:       Absolutely. Yeah. I mean-

Dr. Weitz:           And these are two markers that are listed in the section potential autoimmune triggers.

Tom Fabian:       Right. And that actually is one of the examples I’ll be covering here in this a little bit, so that’s really why it’s considered an opportunistic pathogen. So opportunistic pathogens can exist in the gut without any effects at all or they can potentially be part of a pathogenic process. So they’re not generally recognized at least in the gut as being outright pathogens. That picture is probably going to change a little bit with some of the organisms like Klebsiella, where we do see that it’s more frequently now associated with things like inflammatory bowel disease, et cetera.  So it’s beginning to square where it’s not thought of as sort of an outright infection, a specific infection like salmonella, but it is part of the pathogenic process in some cases. So it’s kind of in that gray area, and that’s a perfect example of where clinical judgment really to come into play in combination with really understanding how to connect the dots with a test like GI MAP.  So you can really get a sense for, does look like this is a problem, or does it look like it’s for this patient probably not much of a problem.  And that’s, well-documented research some people can have relatively high levels of Klebsiella and no evidence of inflammation.  Other people can have high levels and have inflammation, but the big difference often is the ecosystem.  If they don’t have as many of the beneficial bacteria, they may be more likely to have inflammation.

Dr. Rahbar:        Right and if I may say something, obviously we need to evolve into this to understand if we correct that, whether it reverses the immunological abnormality or not.  We can say that this is part of the autoimmune trigger but if you fix it, does it fix your autoimmune problem?  And that answer obviously is not clear.  And so far, I don’t think if you just change that bug, you’re going to see it, but we have to see how that answer is going to evolve.  I wanted to ask you a question regarding the H. pylori.

Dr. Weitz:            So actually, let me just ask that same question again. Tom, is there any data showing that if we have a stool test that shows potential autoimmune triggers and we reduce that particular bacteria, that we may have a positive effect on their autoimmune disease?

Tom Fabian:       That’s a good question. So I’d have to go back and kind of dig through some of the research that I have to see what specific studies there are, mostly to date what I’m aware of is, those are associations and a few cases, there may be some kind of mechanistic insights suggesting there might be a causal link. But mostly to date, a lot of those are based on associations. I’d have to go back and again, dig through the research to see if I do have some studies that specifically talk about whether or not if they’re addressed, does the disease reverse or do some of the symptoms improve?  Not entirely sure.

Dr. Weitz:            Yeah. I just had a patient today who has Ankylosing Spondylitis and he has high levels of Prevotella [I meant Klebsiella] and there’s data showing that that can be directly related to how the Ankylosing Spondylitis is created in the body.

Tom Fabian:       Right. Yeah, definitely for some of them, at least it’s suspected, and some evidence that there might be some kind of molecular mimicry, for example where the immune system reacts to protein on the pathogen or the opportunist that then looks very similar to a protein in the body and then the body develops an immune reaction to those proteins as well. So that’s kind of an ongoing thing. I think it’s mostly well established for one of the oral microbes, which is Porphyromonas gingivalis, but there’s probably a growing list of examples there.

Dr. Rahbar:         Dr. Fabian, may I ask a question regarding H. pylori, we occasionally find this in duodenal aspirate as interesting that even in the stool test sometimes the marker comes up as red and sometimes it’s black. I agree that the best value is below <DL, below the detectable levels. So if any amount is showing would it be abnormal whether it’s in the black or red zone?

Tom Fabian:       That’s a good question. So certainly below detection limit, if you’re aiming for eradication or greatly reducing its levels from a therapeutic goal, then certainly that’s what you’d be looking at. Now there’s with any sort of microbe, I mean, except for outright pathogens and H. pylori is kind of more on the opportunistic pathogen category versus something like enterohemorrhagic E coli, which is not really pathogen. Those you generally don’t want to see those in the gut at all of course.  When there’s H.pylori there’s some debate as to whether or not complete elimination is always desirable. It really depends on kind of looking at all the risk factors, but some data indicates that there may be some increased risk for other conditions by completely eliminating H. pylori. So what we know mostly from research is that most of these types of opportunists in the gut like Klebsiella, H. pylori, as long as they’re kept in check by healthy gut and healthy microbiome, they may not be a problem.   So that’s kind of where that gray area, low levels for some people may be fine, because maybe they have an overall healthy stomach, they have a lot of Lactobacillus, for example, in the stomach, which is definitely one of the known healthy groups in the stomach and a healthy ecosystem. Then they may be fine and there may be no reason to target H. pylori, but if people are having symptoms that could be ascribed to it, plus we see a signature on the test and there may be other considerations as well.  Some clinicians may decide that they want to at least try to reduce that and see if that helps symptoms for that patient.

Dr. Weitz:           Does it make sense to also do a H. pylori breath test or antibody test to confirm it?

Tom Fabian:       Some clinicians feel that because it really depends again, on the scenario and the risks, say if you have lots of virulence factors and there’s a history, family history of stomach cancer or anything like that, there may be scenarios where you really do want to make absolutely sure that it’s eradicated, if you’re trying to eradicate it or even just verifying the levels over time. So there are some clinicians that prefer to do multiple tests. Just to really get a better handle on it. PCR is very sensitive. So it is often a scenario where we’ll see a low level detected where a breath test or a stool antigen tests may be negative.

Dr. Weitz:           By the way, is PCR ever too sensitive?  Is it ever the case that it’s picking up things that maybe are no longer there?

Tom Fabian:       That’s a highly unlikely scenario as far as past infection. So it’s a very different scenario than say antibodies, which can linger for a long time. DNA from organisms, especially in the gut is highly unlikely to linger, partly just from the simple fact that materials moving through within a certain period of time.  So if the microbe isn’t attached and to be attached to it has to be alive, because it has to actively produce those proteins that allow it to attach.  So it’s not likely, that it would be detectable past a certain relatively short period of time on the order of days.

Dr. Weitz:            Okay. And Dr. Greenberg asked if the Pseudomonas is high, does that imply that it’s overgrown in the small intestine or that it has also colonized the large intestine?

Tom Fabian:       So far there is no research supporting that it can colonize or at least colonizes on any sort of frequent basis in the colon. All the research to date supports that it’s mostly in the upper GI and stomach.

Dr. Weitz:            Okay.

Tom Fabian:       Really it’s niche where it thrives. I haven’t yet come across a single study. It can be detected, of course that’s how we pick it up. It really doesn’t seem to thrive in the colon. In fact, specific studies have looked at that based on transcription gene expression, and Pseudomonas is one of those that when you look at gene expression methods is not detected in the gut, meaning it’s not active.

Dr. Weitz:            Okay. Yeah, go ahead.

Tom Fabian:       Okay. So coming back to this section where were talking a little bit about the beneficial bacteria and what to look for when there’s a lack of beneficial bacteria. In addition, of course, to the normal bacteria section you really want to make sure you pay attention to secretory IgA. So we do know that, the main stimulus for normal physiological ranges of secretory IgA is the normal commensal microbiome, through secretion of Butyrate and other products that stimulates the production of secretory IgA. So we see a really strong correlation there.  Typically when there’s a lack of at least, one of the key groups of beneficial bacteria, we often see low secretory IgA.  Clinically it appears that the lower, the number like we see here, the more likely that is to reflect lack of Butyrate producers.  So you’ll often see, for example, really low secretory IgA below 100 when Faecalibacterium, for example, is also very low.  So it’s kind of a great overall marker for lack of beneficial bacteria.

Dr. Weitz:            So, if we see this on a test with a patient who maybe has some other infection or parasite at the same time, would we best be trying to improve their microbiome with probiotics or prebiotics, or would be better to directly try to stimulate their immune system with colostrum or some products like that have an immunostimulatory effect?

Tom Fabian:       A lot of clinicians will focus on both building up, the microbiome can take time depending on what’s going on. So if you need a more immediate effects then those types of supplements, Saccharomyces Boulardii, L-glutamine even Vitamin A, has been shown to be necessary for normal secretory IgA levels. So supplement approaches certainly can help bridge that gap until the beneficial bacteria are recovering. So again, that’s a really important marker to pay attention to that has a very strong correlation with lack of beneficial bacteria.

Dr. Weitz:            I just want to ask about the fecal calprotectin, which is a marker of inflammation. I’ve had a number of tasks where that was normal, and not particularly high, but the patient has a lot of gut symptoms and it just seems like their gut is really inflamed. So, why would that number be low if their gut’s really appears to be inflamed?

Tom Fabian:       That’s a good question. So it’s a good marker for inflammation particularly in the lower GI. So if you look at studies on Calprotectin, for example in relation to H. pylori, so Calprotectin in stool, there’s not much of a correlation, so we know the H. pylori can cause inflammation in the stomach but that doesn’t reflect so much, in ability to detect it in stool. So one possibility is that you may have upper GI inflammation, if they’re inflamed and that’s likely in a lot of cases. So for example, Pseudomonas, lots of new research shows, that when that’s overgrown that’s likely causing inflammation, doesn’t always mean it is. So again, you want to take into account the big picture but if it’s significantly overgrown that’s a significant possibility that could be causing inflammation the upper GI. Candida is known to be inflammatory often colonizes pretty much anywhere in the upper GI tract, even the oral cavity. So that’s one of the major reasons if they seem to be inflamed and Calprotectin looks fine, maybe more on the upper GI.

Dr. Weitz:            One of the patients [I meant practitioners] asked that, she had a patient with Ulcerative colitis. Calprotectin was 660 on the GI MAP a week later, the calprotectin was 150 on a standard lab.

Tom Fabian:       So, it’s difficult to kind of compare among labs due to, we know what our lab does. Our lab certainly does everything that’s required of a CLIA lab and then some, so we know that our markets are internally validates very well, but we can’t speak to other labs in terms of how they perform the test and also do their validation, but also be aware that Calprotectin is a marker that can change over time.

Dr. Weitz:           So if you change, can it change quickly in a week?

Tom Fabian:       But actually, yeah.

Dr. Weitz:           Okay.

Tom Fabian:       Yeah. Because, it’s really responding to what’s going on in the gut. Now, if you have chronic inflammation, that’s not necessarily changing, you shouldn’t expect to see that changed. Generally though, if you’re doing a retest say after treatment and you want to see what’s happening with the levels, it’s always recommended to use the same lab that you started with. So you have that consistency and baseline because otherwise, you’re factoring in possible methodological differences.

Dr. Weitz:           Yeah. I have to say sometimes this sort of question comes up when I’m managing a patient as a Functional Medicine practitioner/Chiropractor, and then the patient goes to see a standard GI doctor and they send them to LabCorp or Quest and get a different result and then they scoff at this test.

Tom Fabian:       Right. Yeah. I mean, that’s always going to be an issue when you’re comparing tests. We see that, like I mentioned with H. pylori where a patient may be negative with a stool antigen test, they do a PCR test and because of the sensitivity level, we pick it up and the other lab with the other method, didn’t pick it up. So, it’s ideally you’re doing the same if you’re kind of comparing time points with the same test.

Dr. Weitz:           Okay, good. Go ahead.

Tom Fabian:       Okay. So on the next slide, it’s just kind of a visual reminder of what we’re looking at. So, we’re so used to looking at tests, the numbers and kind of thinking about things a bit more in the abstract when it comes to the gut, because we can’t see the ecosystem.  It’s important to really kind of come back to sort of the visual understanding that we’re talking about the bugs, and this is the lumen side towards the top, and you can see the bugs here. And if these are largely representing beneficial bacteria, there’s secreting, creating factors, particularly short chain fatty acids that are absorbed into the mucosa. That then basically elicit effects, particularly from the immune cells and the Epithelial cells, certain Epithelial cells, the Goblet cells, respond by producing mucus and then certain immune cells, the Plasma cells respond by producing secretory IgA, and the secretory IgA is secreted in the mucus layer. So just kind of another quick little clinical Pearl is you’ll often see low secretory IgA along with low Akkermansia. And that’s indicating that likely this mucus layer that includes mucus and secretory IgA could be deficient. You can see here, it’s representing that this mucus layer is keeping the microbes away from the mucosa, which is important to keep the reactivity of the immune interactivity down. So that’s part of the whole immune tolerance type, a phenomenon is preventing overreaction of the immune system to harmless microbes.

And so just a little bit more about the Firmicutive bacteria to these phylum, just to reiterate they’re anaerobic and they’re dominant in the colon and the main source of short chain, fatty acids, and those short chain, fatty acids have many important effects. One of which public have a little bit of time to go through here and a little bit when I go through one of the studies, but there are definitely overall key for colon health particularly in terms of protection from pathogens and opportunists. So it’s not common to see some of these opportunists, particularly under Autoimmune triggers, for example. And when we do see them, that suggests possibly the overall beneficial bacteria may be deficient.

So the other information you’ve already talked about, so we’ll move on.  I just want to touch briefly on inflammatory Dysbiosis. So this is a subset of the inflammatory kind of the main inflammatory types of organisms on GI MAP.  So there are particularly inflammatory bacteria in the Proteobacteria phylum. Most of them tend to be in a subclass or subgroup called Gamma Proteobacteria. So they’re all fairly related. A lot of them have been shown to have the more inflammatory type of LPS, for example. But quickly I want to focus on Klebsiella.  So that’s something we’ve talked about a little bit so far here, as an example, there’s this really interesting study that just came out a little while ago.  Actually just this past summer.  So the title is Intermucosal connection between the mouth and the gut and commensal pathobiont-driven colitis.   So this is basically showing that the connection between Dysbiosis in the mouth and Dysbiosis in the gut, and then what can happen in between. So they say here real quick, Periodontitis leads to expansion of oral pathobionts, including Klebsiella and Enterobacteria species, which were both on our tests and the oral cavity amassed oral pathobionts are ingested, then translocate to the gut.

                                So basically swallow pass through the stomach and then end up in the lower GI tract, particularly if there’s not enough stomach acid, where they activate the inflammasome in Colonic mononuclear, Phagocytes triggering inflammation. And I want to go on to a little bit more information here. So they say in the article, this evidence suggests that at least two conditions must be met for oral pathobionts to topically colonize the gut.  First, the colonization resistance of the gut resident microbiota must be disrupted often, meaning that you have a lack of beneficial bacteria enabling the oral microbes to invade the gut.  Hence intestinal inflammation favors the growth of Enterobacteriaceae, that’s kind of a subgroup that includes E-coli, Salmonella, Klebsiella, et cetera in cleaning bacteria transmitted from the oral mark, because one more thing I want to highlight from this article is alternatively neutralization of gastric acid or inhibition of acid secretion could promote ectopic colonization of oral bacteria in the gut.  For example, it has been reported that the use of Proton pump inhibitors, which reduce production of gastric acid leads to increased colonization of mouth resident bacteria in the gut, consistent with his observations been reported that gastric acid inhibitors worsen clinical outcomes in IBD. So definitely this is an important article that kind of follows up on earlier research showing this integrative approach. You’ve got to take the whole GI tract into account.

                                When you see Klebsiella come up on a test, the questions you want to ask are, is there evidence it’s causing inflammation? You may even want to check with the patient, ask the patient about their oral health. You may want to look at indicators of stomach acid because it may be telling you this patient might have insufficient stomach acid. So lots of, kind of connecting the dots there when you see these various organisms. So that’s talked a little bit about the insufficiency Dysbiosis, inflammatory Dysbiosis. Those two are really linked. As you can imagine, when you have a lack of beneficial bacteria, that’s one of the key factors that allows these inflammatory microbes to grow in the lower gut. I just want to touch on the Digestive Dysfunction Dysbiosis. When we think of digestive dysfunctions, most clinicians go straight to this digestion section and look at the markers.  Steatocrit is looking at fat malabsorption in this case, it’s high.  Elastase is a marker for pancreatic enzyme deficiency or our production in this case it’s low. But if you see those normal, does that mean digestion is fine?  And often times the answer is no. Fortunately we can get some additional insights by looking at the patterns of the microbes and what we know about certain microbes, such as H. pylori.

                                So it’s just kind of coming back to what I talked about that this review article basically came to the conclusion that most patients chronically infected with H. pylori have Gastritis and also Hypochlorhydria, not necessarily all.  It’s important to know that not everybody that has H. pylori has low stomach acid but a pretty large proportion of them do seem to have low stomach acid and that low stomach acid for various reasons on this case, they’re looking at Proton pump inhibitors leads to certain types of Dysbiosis that’s really reproducible. So they mentioned here in the highlighted area that Streptococcaceae and Enteroccaceae species are among the most common that you see elevated. Also PPIs are risk proceeded infection. So I’ll get to that. Hopefully we have a little bit time later on here too. Sometimes decrease with E coli bacterium as well. So these are the patterns that we can see, for example, in relation to low stomach acid, there’s a lot of research backing this up, particularly with the Fermicutes phylum. Many studies have replicated that, finding that you see elevated levels of those in stool when patients have been shown to have low stomach acid. And so this is just an example of a patient that had high H. pylori, so suspected low stomach acid, again, not diagnostic for low stomach acid, but something to think about. You’re looking for an overgrowth pattern. We’d see the overgrowth pattern typically in the normal bacteria section.

                                This would be pretty significant. We don’t always see it, this prevalent with so many markers elevated. But we often do see this overgrowth pattern, particularly with the Firmicutes phylum. And then in this case, we also saw Faecalibacterium was low, which really follows that particular study that was found to be low as well. With low stomach acid, your opportunistic bacteria is really where you’re going to see the most common evidence of overgrowth. That’s where the Enterococcus species are and Streptococcus. So if those are high, then suspect that stomach low stomach acid may be an issue. And then some of these others too, like Pseudomonas can also be overgrown and stomach acid is too low, or there’s other.

Dr. Weitz:            Can I ask about Methanobrevibacter?  So we know that methane SIBO is now IMO according to Dr. Pimentel. And so therefore it’s an overgrowth of methanogens, like methanobrevibacter, Methanobrevibacter smithii, and so if the patient has a positive SIBO breath test for methane SIBO, would we necessarily see elevated levels of Methanobrevibacter and, what if they don’t correlate?

Tom Fabian:       That’s a good question. So they generally do seem to correlate as far as we can tell but not always. So that is an important factor to consider that sometimes you can have an increase in bacteria or in this case, microbials because Methano bacteria is they actually are archea, not bacteria kind of closely related, but and so you can have an increase in function, of microbes in some cases without necessarily a significant increase in their numbers. Now that said, on our test and based on looking at literally thousands of tests over the last couple of years then the methanobacteria family on our test tracks very closely with overall evidence of overgrowth on our test. And you’ll see here in a moment, if I get a chance to get to the one of the slides here that talks about that more, it’s, it’s really a direct reflection of the level of fermentation, particularly in the colon.  So when you see methanobacteriaceae in our experience it seems to be clinically relevant anytime it’s in the E nine range, not even above the cutoffs.  It’s highly correlated with this. So generally, yes. I would say when, when patients have also had a breath test, they will have some level of methane detected. But it’s not necessarily a hundred percent correlation for example.

Dr. Weitz:            And somebody asked a question about H. pylori, which is can it also increase stomach acid production, depending upon where the H. pylori is?

Tom Fabian:       Yeah, that’s true. So there is some evidence that in some cases that can be the case during more than acute sort of early infection. And it really does depend, apparently there’s some conflicting research on it. So it’s not really all that well studied, but it does depend on where in the stomach it’s infecting and the extent of the infection. Now, there is some evidence based on a couple of studies I came across that typically the infection can migrate in the stomach over time. And I forget which direction if it’s antrum to corpus or corpus to antrum, but the overall conclusion was that it tends to long-term be more consistent with hypo or Hyper. But some patients may have an increase in acid depending on where the infection is.

Dr. Weitz:           Okay.

Tom Fabian:       So this is just kind of showing you in this example that Klebsiella was elevated which again is something we often see when there’s other evidence suggesting low stomach acid. And that goes along with that research that suggests Klebsiella may often come from the oral cavity or possibly the respiratory tract, which is also another common location for Klebsiella. And then it may get into the GI tract to those routes when stomach acid is insufficient. I’m going to go ahead and skip this here other than, of course, whenever you see an overgrowth of inflammatory species and we had that list higher up on one of the other slides oftentimes, but not always, you’ll see Calprotectin elevated. So there is a correlation there. We see it most commonly in, for example, patients that have already been diagnosed with inflammatory bowel disease we often will see that connection between higher Calprotectin and higher levels of inflammatory species.

Dr. Weitz:           By the way, if pancreatic elastase is low, is the best recommendation is to recommend that they supplement with pancreatic enzymes or are there other recommendations?

Tom Fabian:       That’s a good question. So a lot of clinicians do take that route to help compensate. But of course you want to also ask the question what may be contributing to that.

Dr. Weitz:           Right.

Tom Fabian:       So there may be issues with the pancreas itself. Of course, Pancreatitis, things like that may play a role for some patients, but even low stomach acid can lead to lower release of digestive enzymes. That might be another factor for a lot of patients that can be gut-brain axis issues.

Dr. Weitz:           Interesting. Low acid leads to low pancreatic enzymes?

Tom Fabian:       It’s a contributor. Yeah.

Dr. Weitz:           Okay.

Tom Fabian:       It wouldn’t be considered, if you say you have really low elastase are rate sufficiency. That’s, well below that 200 cutoff it would probably be unlikely that that’s the sole contributor unless for example stomach acid is exceptionally low. You may be wanting to look into some other contributing factors at that point.

Dr. Weitz:           And if steatocrit is high, that’s fat in the stool, is the best recommendation to give them a lipase, fat breaking, fat digestive enzymes, or is bile going to be more beneficial or both?

Tom Fabian:       Good question. So just based on the presence of the excess fat in the stoool you can’t tell directly just from that which upstream process is affected.  Now, if you do see that our last days is really low, particularly if there’s outright insufficiency, meaning well below 200, then it’s more likely that, the highest steatocrit would be caused by lack of lipases from the pancreas as well. But you can also have a scenario where there’s insufficient bile but it’s probably less appreciated is issues in the small intestine, which is where fats are primarily absorbed, where they’re supposed to be absorbed in a healthy gut. So if you have an inflamed, small intestine, and you’re not able to really absorb fats as efficiently.

Dr. Weitz:           Such as if you have SIBO.

Tom Fabian:       Exactly.

Dr. Weitz:           Okay.

Tom Fabian:       Right. Just have a few more things to go through and then based on time, do you want me to stop here or take more questions or do you want me to continue on for a couple more slides?

Dr. Weitz:           Go a couple more quiet slides.

Tom Fabian:       Okay. so again, these are the three main Dysbiosis patterns that learning to recognize them can give you a lot of insights into what’s going on in the overall gut, in terms of physiology and the microbiome, and even some insights into say lower versus upper GI issues, I’ll skip this, but we all know that of course pH is one of the key factors along the GI tract. We mostly think of stomach being the key kind of checkpoint. So when you may unfortunately ingest pathogens, for example, food poisoning if you have good stomach acid, then they’re less likely to survive the transit, but what’s less appreciated is that slightly acidic levels in a healthy colon, especially the ascending colon, are almost as important and helping to keep pathogens and other opportunists from colonizing your colon. So I’m just going to go through a quick example of that here. This is a really interesting paper that was published recently, so the title is Inhibiting antibiotic-resistant Enterobacteriaceae by microbiota-mediated intracellular acidification. So that’s in the Proteobacteria phylum, that’s a particularly inflammatory group.  So inhibiting that group by microbiota mediated, intracellular acidification. So just to kind of cut to the chase, they mentioned Klebsiella and E coli as kind of the examples they’re looking at here. And then the highlighted in orange part towards the bottom says Klebsiella pneumonia, Escherichia coli, Proteus mirabilis by acidifying the proximal colon and triggering short chain, fatty acid mediating intracellular acidification. So essentially what they showed in this study is that normal levels of beneficial microbes that are producing adequate levels of short chain, fatty acids literally inhibit the growth of these pathogens by basically producing the short chain fatty acids that then are taken up by those cells and that satisfies the cells and basically prevents them from growing.  So that’s one of the ways in which these beneficial bacteria, like Clostridia, Bacteroidites can help to keep those pathogens in check. So once again, when you see those at high level particularly Klebsiella or Proteus that’s one of the things we’ll be looking at is evidence for low beneficial bacteria. And they actually showed in this study that just restoring the beneficial bacteria and the short chain fatty acids can be enough to address those pathogens. So that’s kind of an example where you don’t always have to resort to antimicrobials.  Restoring gut balance may be sufficient.

Dr. Weitz:           Can I just ask a question about anti-microbials?  I interviewed Dr. Jason Hawrelak a few months ago, and he mentioned that he thinks that taking anti-microbials like Berberine can significantly damage the microbiome.  Do you think that that is something that is liable to happen?  Do we see evidence of that?  In other words, can natural anti-microbials produce damage in the microbiome?…   The way antibiotics can say, for example.

Tom Fabian:       Generally, no, we have not seen that. Not even close.

Dr. Weitz:           Okay, good.

Tom Fabian:       Mostly when we see a really broad deficiency pattern, it’s related to antibiotics or other pathology like IBD or something like that.

Dr. Weitz:           Good.

Tom Fabian:       Real quick though, I think he was referring to at least the research I’m aware of on Berberine in one study a few years ago, showed that it can long-term or high doses can reduce diversity. But it was in an animal model in mice. So that suggests maybe not taking super high doses for a long time would be-

Dr. Weitz:           By the way what’s a high dosage? Because for example I’m using Berberine not only for gut patients, but we use it to help manage metabolic factors and I also use it as a sort of a substitute for Metformin in anti-aging programs.

Tom Fabian:       Right. Good question.

Dr. Weitz:           It works synergistically with Metformin to manage blood sugars, so.

Tom Fabian:       Right. So that was really just one study and I have not come across any additional studies on that and again it was a mouse study. It was a very high dose though. Something maybe equivalent to a least more than five grams per day for humans. Again, you have to take those kinds of things into account that if it was just an animal study and it was super high dose, maybe it doesn’t apply to real world settings. Right?

Dr. Weitz:           Okay.

Tom Fabian:       But that’s not been our experience. The patient is on strong antimicrobial botanicals. Don’t seem to develop a deficiency pattern based on what we see on GI MAP so far.

Dr. Weitz:           Right. Good then that’s been my experience as well.

Tom Fabian:       Okay. So this actually, it’s kind of interesting that we talked about that, because I won’t go through this full case. It’s not really a full clinical case, but just a quick example of an older female history of bladder cancer was on long-term Amoxicillin antibiotic for five or more years. Main complaint was just Chronic diarrhea. So of course there was a suspicion of antibiotic associated diarrhea in this case.  You can see here very broad deficiency, which would be pretty much expected from such a long-term exposure to an antibiotic.  But just based on what we’ve talked about before with the insufficiency, you would expect that this would potentially lead to overgrowth of opportunists or pathogens.  And sure enough, you see Pseudomonas and Klebsiella overgrown and it turns out that those two are the ones that are known to be resistant typically to Amoxicillin.  So, we’re seeing exactly the pattern you would expect based on that patient being on that antibiotic.  This kind of scenario to me is very important. You’re thinking about, which gut tests you want to go with and how good is a gut test? And you can kind of debate validation approaches and all these things and sequencing versus PCR. But in the end of the day, it really comes down to “Does this clinically validate?” And as you know a patient for example, is on an antibiotic long-term, you would expect to see patterns of this. If you don’t see patterns like this, there may be some individual variation, but when you… So that’s just something to keep in mind as you’re going through some people may try different gut tests. You want to make sure that the test you’re using clinically validates and that it makes sense based on the patterns that you’re seeing.

Dr. Weitz:            Can I just comment your test looks at, if there’s a pathogen what antibiotics might be effective for those pathogens, some of the stool tests, in which in the past, I know used to use natural anti-microbials, how come you don’t include that?

Tom Fabian:       That’s a good question. So I’m not sure of all the reasons why I know part of it is we wanted to kind of keep our test to molecular that’s what our lab does, that other sort of approach to look at potential sensitivity involves culturing. So that’s part of the picture, but also how realistic is it from a couple of different standpoints that what you’re measuring on a Petri plate with one organism really translates to how effective that’s going to be in an actual, vast, complicated ecosystem. You don’t really know that plus most clinicians tend to use common formulations.  Some do use single ingredient approaches.  A lot of clinicians use formulations that they found to be generally effective.  So it’s not necessarily going to affect what they use clinically either.  So it’s not always clinically useful information.

Dr. Weitz:           Okay.

Tom Fabian:       I think I’ll skip all of this here just real quick. And one more thing about Klebsiella and also Morganella they have been linked to excess Histamine production.  So again, be thinking of these things, when you see Klebsiella overgrown you want to kind of look at the context are beneficial bacteria lacking. Does the patient also potentially have Histamine intolerance symptoms, et cetera. So the more you know about these microbes, the more you can potentially clinically connect those dots. This is the only thing I was going to show and I’ll stop at this one. This is a little bit of a complicated diagram, but it’s representing the colon on the left side is the proximal colon and moving on across horizontally to the right towards the distal colon. There’s a lot of research showing basically the dynamics in the colon so that you understand better, why a patient may have say a high Firmicutes, low Bacteroidetes why they might have Sulfur-reducing bacteria and methanogens.

                                So the main point I wanted to make here is generally in the first part of the colon is where most of the Fiber fermentation, Carbohydrate fermentation is going to happen, that can release hydrogen and then basically that part has to kind of start happening first before that hydrogen is available for the methanogens.  And then basically further down the line would be the Sulfur-reducing bacteria, which tend to be more in that latter part of the colon.  Where also bacteroidetes tend to dominate and that’s going to be affected by transit time and other factors too.  But there’s a lot of, sort of modeling that’s being done to understand the ecosystem.  And I think this is a great visual and understand why do we see high methanogens on our test when there’s high Firmicutes that are showing you that there’s a general overgrowth pattern, probably because they’re fermenting, generating H2 and then promoting methanogen growth.  All right.  So that’s in a few additional slides, a lot of material that we could always cover.  Maybe we’ll have an opportunity a part two at some point.

Dr. Weitz:            Okay.

Tom Fabian:       Yeah, so I bring that in, so really it’s all about connecting the dots and just learning. We have a lot of educational information because we offer the consultation. So I encourage everyone to take advantage of those resources because the more you can connect the dots, the more you can get out of the stool tests in terms of good clinical insights that can really help you be more effective in helping your patients and clients.

Dr. Weitz:            I have one more question I’d like to ask and then, I’ll see if anybody else has some additional questions. I’ve interviewed several clinicians. One of whom is very prominent and feels that some protozoa like Blastocystis hominis and D. fragilis are commensal and not something that we should be concerned with. And he’s been talking about that quite a bit.  And I think it’s common for a number of Functional Medicine practitioners to see that on a GI MAP stool tests and say here is likely the cause of your problem and are used to treating it and getting results.  And then I also talked to, in another recent Podcasts another practitioner Ilana Gurevitch and she felt that in patients where correlated with their symptoms, very important to treat blasto and D fragilis. What do you think are these pathogens, commensals or does it depend?

Tom Fabian:       Good question. So they’re not likely to be commensals based on our experience because we don’t see them in the majority of patients. It’s commensals tend to be present in the majority of individuals. I think the real answer is probably somewhere in between. So if you look at the research, most of the research suggests that Blastocystis in particular and possibly Dientamoeba their conclusion and in most of the research is that for the majority of people that have them, they’re not a problem because most of those people are asymptomatic, but I think they’re thinking of them asymptomatic in a conventional sense, not a functional sense. So they may be ignoring SIBO and these other things, mild GI symptoms, et cetera, and not taking those into account. With GI MAP, we only see, I would say probably more than 95% of the time when we see blastocystis and, or Dientamoeba Fragilis, it’s a long with that digestive dysfunction pattern. I don’t think I’ve ever seen blastocyst is just sort of by itself with no other patterns where it’s kind of like the obvious cause of things.   It’s almost always in the context of things like H. pylori, maybe candida, general overgrowth. So the question is what’s causing the symptoms?  Now trying to pin it on an individual organism, some may be more important than others.  H. pylori often is because of its effects on stomach acid.  Candida often is because we know it has, it can cause inflammation like you got Pseudomonas. Blastocystis I think is somewhere in between probably for some patients, if they have a certain subtype and it’s a really high level that may be more significant than if it’s there at a low level.

Dr. Weitz:            Okay. So one question that came in is what causes Insufficiency Dysbiosis? Is it low stomach acid?

Tom Fabian:       That’s a good question. So recent research indicates the antibiotics are often a major cause even long-term because what can happen is antibiotics can actually effect the mitochondria in the cells that line the colon, in a way that makes the gut environment less favorable to those beneficial bacteria. So, that’s at least one example of pharmaceuticals that may have that effect. Certainly lack of fiber in the diet. That’s been correlated with lower levels of these beneficial species, any source of inflammation.  So inflammation is highly detrimental to many of those beneficial bacteria. So whatever gets inflammation going that can really turn the tide.  One of the other contributing factors and it’s related to low digestion is too much protein in the colon.  That could be from just high protein diet or not digesting well.  But this process of sort of breaking down amino acids, which is called protein fermentation, and then breaking down carbs which is carb permutation. So primarily fibers they’re antagonistic.  So the less fiber you have, and the more protein you have in the colon, that can really also be detrimental to those beneficial bacteria.

Dr. Weitz:            So another question is how does the Ketogenic diet, and I would even throw in there, how about the carnivore diet impact the microbiome?

Tom Fabian:       I’ve actually only seen two cases so far and they were all almost identical and they were probably one of the… There were both among the worst cases I’ve seen as far as Dysbiosis. That’s a perfect example of this sort of… The sayings of tests don’t guess, or the opposite of that is treat the patient, not the test, right? Two different philosophies or ends of the spectrum. When you see that, so in both cases, the patient improved in symptoms, they had typical GI symptoms, gas bloating, apparently issues with carbohydrates. So they went full carnival symptoms improved. And yet you look at the gut microbiome and it looks terrible, lots of inflammation in both cases just massive Dysbiosis. So the question is, well, is short-term improvement in symptoms, are you setting the stage for long-term problems? And is there another way to address those symptoms without kind of ruining the microbiome? So good question. I mean, I guess time will tell, from everything we know about the microbiome, those results look terrible. They really would be considered by almost everybody is really bad [inaudible00:40:45]

Dr. Weitz:           Okay. And one more question, is it necessary to stop digestive enzymes before collecting the stool sample? In fact, are there any other recommendations that should be taken before collecting the stool sample?

Tom Fabian:       Not really a lot of us can depend on the clinician goals in terms of what you want to see. Do you want to see how your patient is doing on the treatments or do you want to see them of course, before treatment, et cetera?

Dr. Weitz:           Well, I guess if they’re taking enzymes, would that affect the pancreatic elastase levels?

Tom Fabian:       There’s not much evidence that does now.

Dr. Weitz:           Okay.

Tom Fabian:       Certainly, with acid, that’s typically not in the enzyme formulation, so, you wouldn’t literally affect it from just coming from the antibiotics or the main one, if you do take antibiotics we recommend waiting at least 30 days post antibiotic treatments, maybe 60 days to allow the microbiome to recover somewhat.

Dr. Weitz:           Okay. Excellent. Thank you so much, Thomas.

Tom Fabian:       It was my pleasure. Thank you.

Dr. Weitz:           Great. And thanks everybody. And we’ll see you next month.


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