Brahmachary M, Schönbach C, Yang L, Huang E, Tan SL, Chowdhary R, Krishnan SP, Lin CY, Hume DA, Kai C, Kawai J, Carninci P, Hayashizaki Y, Bajic VB
BMC Bioinformatics7 Suppl 5pS8(2006 Dec 18)
Type: Conference presentation
Presenter: Prof. Trevor Marshall
Conference: Workshop on Chlamydial infection
Location: Prague, Czech Republic
Date: April 18, 2009
See also: Transcript with slides
Well thank you very much for inviting me. It’s great to be here in Prague. It’s my first visit to Prague. And I must say I’m enjoying the city.
What I’m going to talk about is a new concept called the “metagenome.”
As we understand more about the human genome and the pathogenic genomes we are starting to understand more about how the various pathogens – be they Chlamydia, Mycobacteria, and Mycoplasma – how the various pathogens interact with our own genome in order to cause disease.
And so my title is based on “Infectious Disease transitions to an understanding of the Metagenome.”
There are three types of biology that are pretty common these days. The first type, in vivoA type of scientific study that analyzes an organism in its natural living environment., of course in animal or human models. In vitroA technique of performing a given procedure in a controlled environment outside of a living organism - usually a laboratory. is where a lot of the work on antibiotics is being done – in cell culture, in the lab.
In silicoExperiment technique performed on computer or via computer emulation. is very new. The first time I came across in silico was at this gathering here in Toronto back in 1981. Human insulin had just been synthesized using mathematical formulae – using the IBM supercomputer that could simulate the insulin molecule at the level of the mathematics. And that’s really what I’ve been doing over the last decade.
There’s a new push going on at the moment. The NIH in the USA has started the Microbiome Project. The goal of the Microbiome Project is to characterize all of the places in the human body where genomes—other than the human genome—are also present. The NIH has estimated that about 10% of the cells in the body are human cells, and about 90% of the cells—in a normal healthy individual’s body—are bacterial cells. Now remember that bacteria cells are very, very small—and in most cases bacterial cells—many, many hundreds of bacterial cells—can live within infected human cells.
We are starting to get to an understanding now that the human body does not work—the human genome does not work—in isolation. It works in concert with a symbiosis of other genomes that have gathered throughout a person’s lifetime, and indeed throughout the ages.
This study has just been published. It’s a metagenome of human saliva.
It was a study done by Max Planck Institute in Germany. They took samples of saliva from 10 different places in the world—all over the world geographically—and individuals in those 10 places—and then they sequenced the genes. Because now, with in silico technology, we don’t rely on being able to culture the organism any more, we can actually sequence the sample and then find these little fragments of DNA we can match up with our known database of 800 or more identified pathogens.
What they found was that there was more variation individual-to-individual, than between geographic locations. But they found that there are a hundred bacterial genomes—including some previously unknown genera—64 unknown genera—in this human saliva from healthy human individuals.
Streptococcus was most common of course at 22%. Haemophilus at 6%. Neisseria at 8%. But look, Treponema and Yersinia were in trace amounts. It is interesting actually that Chlamydia was not found. The body can apparently deal with the Chlamydia organism and reject it—or at least a healthy body can.
A similar study on prosthetic hip joints. This is hip joints that were being replaced during surgery. They used a special procedure with ultrasound to shake the biofilm A structured community of microorganisms encapsulated within a self-developed protective matrix and living together. off the hip joints, and see what species were present. Of course, you’ve got all the normal ones. You’ve got staph. You’ve got the gliding bacteria you’d expect to find in a biofilm, Lysobacter. But look here, down here we’ve got hydrothermal-vent Eubacterium at about 5% of the samples. And that’s the same level as staph. Staph aureus, a very common biofilm pathogen which is at about 4%. The hydrothermal-vent Eubacterium–the genome for that had been isolated in bacteria that had been extracted from hydrothermal vents at the bottom of the ocean—-What’s it doing in man? Well, that is what we are about to find out in the next decade. the next decade we will see a huge change in how we understand how the human organism interacts with the other organisms that are within the human body.
Now there’s two ways of looking at disease. One way of looking at disease is saying, “Well, this person is sick. She can’t put her weight on her leg. She has to lie down all the time.” In other words, focus on the symptoms. And categorize diseases based on symptoms. And that’s the way that medicine has done it for the last century.
But now we’re starting to view diseases based on the genes. On the common genes that affect the various diseases. This [referring to slide] is a gene map that was produced a year or two ago now, 2007, and it has all of the diseases: neurological diseases, deafness; the autoimmune diseases; cancers. It has them all linked together based on common genes.
If we look at a close-up of the area around the autoimmune diseases, you can see that this one gene here, called ACE—which is involved in the progression of SARS, the infectious disease SARS—it’s involved in myocardial infarction, it’s involved in Alzheimer’s, it’s involved in kidney disease, and it’s also involved in Sarcoidosis and some of the other granulomatous diseases.
What’s interesting about this common gene, ACE, it’s not only involved in so many of what we would think of as different diseases—kidney disease, cardiac disease, Alzheimer’s–but also that this particular gene we know is affected by some of the bacteria which are present in every person’s body. In particular, Bifidobacterium and Lactobacillus, which are species which you can find in yogurt. You certainly find them in probiotics. Those effect the expression, by the human genome, of the gene ACE. They also effect all the diseases, or the disease states, whose balance is dependent on ACE. A healthy body is healthy because everything is in balance. When certain proteins start being downregulated, others start being upregulated, you start to get a disease state setting in.
Down here we have another gene, PTPN22, and that one’s associated with Lupus-SLE, Rheumatoid arthritis and Diabetes. That one is one of the body’s primary responses to Mycobacteria. When the body senses mycobacterial infection, one of the first things it does is upregulate this gene, PTPN22.
So by looking at the genes in disease we can get a totally different picture from trying to work basically on symptoms, on differentiating symptoms as we have done for the last century.
Well, Chlamydia is not a very well-studied genome. I’ll give you some data on Chlamydia in a little while, but the one that I want to focus on—because it illustrates the problem that we have; we, being, Homo sapiens, as a species have—and that is the HIV genome.
The HIV genome is very small. It’s a small strand of RNA. From that genome 17 proteins (19 including cleavage), but 17 proteins that are generated from this one strand of RNA, which is the HIV genome. So, HIV does all it’s damage by generating only 17 proteins. You compare that typically with a bacterium which certainly generates hundreds of proteins, usually close to a thousand proteins.
The HIV genome transcribes for 17 proteins. And with the billions of dollars we’ve spent analyzing HIV, finding out what it does, we have identified—we being science—has identified that there are over 3,000 interactions between those 17 proteins and the human metabolome, the human genome.
So if we take the saliva genome, which we saw on slide —which has got more than a hundred species, and they transcribe for approximately 50,000 protein products—we compare 50,000 here to 17 here [referring to slide]. And then we look at 3,000 [interactions with HIV proteins]… how large is that 3,000 going to rise if you’ve got that many more proteins being generated by the salivary metagenome?
The answer is, it’s imponderable. You get to a point where you are just looking at noise. You’re looking at stochastics. The body is trying desperately to deal with the species that it’s got there—the DNA at the level of the transcription—trying to ignore all of the proteins and the enzymes that are coming from the pathogenic genomes and still produce good quality proteins from the human DNA and it’s just an imponderably complex problem.
Why is it so complex? This is a slide courtesy of Professor Peter Wright at the Scripps Institute. This is a human transcription factor called CBP/P300. This transcribes some very important enzymes and proteins—I’ll be dealing with one of them later in the presentation.
What happens is, you’ve got a strand of DNA here [referring to slide] and the transcription factors look for areas of that DNA where they’re attracted—where their active areas are attracted—and then the DNA is transcribed, split into RNA strands and transcribed by the normal transcription mechanism. But the reason that the viruses affect so many different human gene transcription is because they are disordered. They have disordered loops.
This loop here (referring to loop in CBP/P300 on slide) will bind to other substances, it binds to other proteins. Depending on whether that loop is squashed up, or stretched out, you’ll be decoding a totally different region of the DNA. This one transcription factor actually transcribes thousands of human genes.
The important thing to note is where is this human protein has got areas that are very well structured—and these are the colored areas that are defined—very well structured, and the disordered loops are relatively small in size and number.
In a virus, the viral proteins are almost all totally disordered. So they have no shape until they wrap themselves around a protein or an enzyme from their host—from the human body. Then, depending on which atoms attract each other, they take on the shape of the host protein. They can change very quickly. They can mutate very quickly. As you all know that is one of the biggest problems with HIV.
Even though the plethora of interactions is imponderable, even though there are so many of them that it’s imponderable, even though the microbiotaThe bacterial community which causes chronic diseases - one which almost certainly includes multiple species and bacterial forms.—the communities of microbes—affect gene expression from the brain to the toes, we can say one thing, and that is:
The catastrophic failure of the human metabolism we see in chronic disease, which at first glance appears so diverse—and so different between the various disease diagnosis–is actually due to the same underlying mechanism—a ubiquitous microbiota which has evolved to persist in the cytoplasm of nucleated cells.
It’s very important that these bacteria persist by overcoming the innate immune responseThe body's first line of defense against intracellular and other pathogens. According to the Marshall Pathogenesis the innate immune system becomes disabled as patients develop chronic disease.. The very cells that are supposed to kill the bacterial pathogens, they actually overcome, and they live within those phagocytes–and live within those phagocytes for quite a long time.
The same cause is behind Hashimoto’s hypothyroiditis and Multiple Sclerosis. The same cause is behind Chronic fatigue and Rheumatoid Arthritis. Diseases that we wouldn’t normally associate if we are looking at them as symptoms. But when we look at them as genes, we can see how they’re associated.
In fact, when you do a genome-wide study—this is one of apoptosis in Sarcoidosis, 2001—what you find is basically there’s nothing that jumps out at you. There are no genes up here fully on [referring to slide], there are no genes down here that are fully off. There’s just a whole lot noise around the expected region [referring to the mid-line in the diagram]. Yes, there’s some like interleukin-8 that stick out, but basically it’s just noise. It’s the total sum of all that noise, each one of those proteins, that is affected by the infection—each one of those proteins, that is affected by the microbiota—together, the sum total of them is what gives rise to the symptoms of chronic disease.
If we take Mycobacteria; that’s been fairly well studied; there are about 4,000 genes in mycobacteria. There’s a very good study, published in the Chinese Medical Journal back in 2003, where they infected cells in culture—in vitro—with Mycobacterium tuberculosis, and they tracked which genes actually changed in the cell when they infected it with this one strain of pathogen, Mycobacterium.
They are not talking about one toxin. There’s not one toxin. There were 463 genes whose expression were changed. 366 of them were known genes. The other genes were unknown —mutations. And the genes function in various cellular processes including intracellular signaling, cytoskeletal rearrangement, apoptosis, transcriptional regulation, cell surface receptors, cell-mediated immunity as well as other cellular metabolic pathways.
In fact everything, effectively, that determines how the human body operates is affected —in that cell— when it’s infected with Mycobacteria tuberculosis. 25 were up-regulated and 341 were down-regulated, Two that I’m going to point out were: the CD14 receptor was downregulated 2.3 fold, and the VDR receptor was downregulated 3.3 fold. Downregulated three times in size. That is a very significant effect of Mycobacterium on the VDR.
Chlamydia has about a 1,000 genes. It’s a smaller genome. There have been a few studies. The first I cite here only examined cytokinesAny of various protein molecules secreted by cells of the immune system that serve to regulate the immune system., chemokines and is really of very little interest. Then there was another one that was published in 2006 which found that of the 194 mycobacterial genes in that previous study that I was talked about, 35% of them have similar coding sequences in the Chlamydia trachomatis genome. In other words, of the genes in mycobacterium, which are responsible for it being persistent, it being able to become chronic, about 35% of them have very, very similar genes in the Chlamydia genome. But the interesting thing about Chlamydia is it didn’t posses a gene set whose sole function is the maintenance of persistent infection. Mycobacteria does. Mycobacteria has two defined states. Active and latent. But Chlamydia doesn’t have genes which are solely responsible for that.
Gene Expression During CPN Infective Cycle was published in PLoS Pathology in 2007.
Why is the cytoplasm of nucleated cells—that means cells with a nucleus, the phagocytes in particular—why is that so important? Once again, we’ll go back to our old friend HIV, because it’s pretty well studied.
HIV infects through the cell membrane—through the CD4 receptors typically—and then the RNA of the HIV is reverse transcripted into DNA, double stranded DNA. Which then goes into the nucleus where it becomes integrated with the human genome, and then transcription occurs in the normal way. The viral RNA leaves the nucleus, is assembled once again, reconstructed and then leaves the cell. The important thing to notice is a lot of the work is done in the cytoplasm—in the region around the nucleus. In the nucleus you have the integration, you have the DNA repair, you have a number of mechanisms. But the cytoplasm is very important. That’s were all the proteins are generated, that’s where most the enzymatic activity takes place, so the cytoplasm is very, very important.
Here we have a picture from a transmissionAn incident in which an infectious disease is transmitted. electron microscopy study at Columbia University back in the 1980's by Emil Wirostko. Emil’s group studied lymphocytes, monocytes, macrophages, and neutrophils, from patients with Juvenile rheumatoid arthritis, Sarcoidosis, and Lupus. And they found the same thing. In all of those diseases there were infectious colonies of bacteria—which stained as bacteria—that were living within the cytoplasm of these phagocytic cells. The very cells, the lymphocytes that are supposed to get rid of the pathogens from our body are actually being parasitized in these chronic diseases.
If we look at it with an optical microscope, we can see we have a nucleus here [referring to video on the screen] and a cytoplasm which is just swelled and exploded as a result of these small colonies of biofilm-like pathogens. These huge long tubules are being thrown out from the degrading cell. These are very, very thin tubules, caused by bacterial protein. This one’s about 20 cell diameters long, extremely long.
That’s what happens with the cytoplasm of the cell becomes so infected that the pathogens start to break out—the biofilm starts to break out to try and find more suitable hosts—other than that particular cell. Here we can see [referring to image on the screen], zoomed out, the length of this huge long biofilm tubule that is put out.
In the three decades that have gone past since my early research in Perth, Western Australia, and Toronto, Canada, the number of symptomatic similarities which existed between the various chronic diseases become more and more obvious to me.
It became pretty clear to me that all the chronic inflammatory diseases were arising from a common pathogenesis. Which I figured had to be a failure of the innate immune system. And we found that it had to be a Th1-dominant cytoplasmic, metagenomic microbiotaThe community of bacterial pathogens including those in an intracellular and biofilm state which cause chronic disease.. And in particular that persistent phagocytic infection had to be the cause. How did I figure that out? I just did. Sorry. [smiling]
There’s no other way that the biochemistry all fits together. If you look at the number of changes to the molecular chemistry which occurs in these chronic disease states it is imponderable, it is just huge. There’s no other way that the human genome could go that wrong. People are not born with that many mutations in their genome. Basically, the genome of Homo sapiens is fairly uniform. It had to come from somewhere else, and in fact, it had to come from pathogenic genomes.
But the thing that’s really important is, for decades, chronic disease patients have been given antibiotics and have responded to the antibiotics differently from the way healthy people responded to the antibiotics. One of the reasons for that is because the postulates of Koch, from 1897, said basically, “Look, you’ve got to be able to examine the bacterium out of the body, in the lab.”
The moment you take it out of the body, you get rid of a whole lot of things that happen inside the cells of the human body. For example, if you take the antibiotics clindamycin, minocycline, and rifampin—-rifampin was the primary antibiotic used against tuberculosis–minocycline you know, and clindamycin you know. All of those activate a nuclear receptorIntracellular receptor proteins that bind to hydrophobic signal molecules (such as steroid and thyroid hormones) or intracellular metabolites and are thus activated to bind to specific DNA sequences which affects transcription. in the human body call the PXR nuclear receptor, which is right at the heart of the human immune system. We’ll get to that in a future slide. So when those antibiotics are in the human body they have additional actions to what they have in a petri dish. And this is something that we were really only able to understand once we could understand the genes and how the genes interacted. And what I figured they had to be doing is knocking out gene expression by the VDR Nuclear Receptor. The VDR Nuclear Receptor—in man—is responsible for some key endogenous antimicrobials. That means, antimicrobials that are produced in the human body itself. There are 24, approximately, families that have been identified and about 17 of them are affected by the VDR directly or indirectly. So it’s absolutely key. In particular, the Cathelicidin Family of antimicrobial peptides found primarily in immune cells and transcribed by the Vitamin D Receptor. antimicrobial peptide, the receptor TLR-2—that’s the one that’s been on the previous slides of all the other speakers as the one recognizing Chlamydia—that gets knocked out, when you knock out the VDR. You knock out out Cathelicidin and you knock out beta-defensins. At that point, the cells' immune defenses have been virtually knocked out. Just by the bacteria figuring out how to to knock out that one nuclear receptor, out of the thousands and thousands of proteins that are in the human body.
In Homo sapiens — and this is different from animals, animals have different function in their DNA transcription—but in Homo sapiens the VDR Nuclear Receptor transcribes genes for TLR2A receptor which is expressed on the surface of certain cells and recognizes native or foreign substances and passes on appropriate signals to the cell and/or the nervous system., as well as the Cathelicidin and beta-defensin An antimicrobial peptide found primarily in immune cells and transcribed by the Vitamin D Receptor. antimicrobial peptidesBody’s naturally produced broad-spectrum antibacterials which target pathogens., all of which are essential to intraphagocytic innate immune defenses.
A microbiota has evolved, which we’ve called a Th1 microbiota because one of the common factors is interferon-gammaAn inflammatory cytokine which causes extra mast cells to differentiate to monocytes and then to further differentiate into macrophages and dendritic cells. These phagocytes are the most active cells of the immune system and are charged with digesting bacterial pathogens. which is produced when the innate immune system is attacked by these persistent pathogens.
The Th1 microbiota evades the human immune system by blocking DNA transcription by the VDR, which consequently blocks expression of these endogenous antimicrobials.
So it comes as no surprise then that we now know that HIV totally disables VDR. HIV takes VDR and actually uses it to help transcribe it’s own RNA genome, its LTR transcription-repeat. The tat protein from HIV actually takes away one of the human body’s main innate defenses and uses it as part of the viral replication. We saw earlier that Mycobacterium tuberculosis downregulates the VDR by 3.3 fold. So these pathogens know how to get around human innate defenses.
Unfortunately during the 20th century, Homo sapiens changed their lifestyle in several ways that has resulted in further downregulation of gene expression by the VDR and which made their bodies more susceptible when infections came along, like HIV or TB; that required weak VDRs in order to become persistent.
The VDR is called the VDR because it’s short for vitamin D receptor. Now, vitamin D is not a nutrient. Despite what we’ve thought for 100 years, certainly back to the 1930’s. Which is what? 80 years. For the last 80 years we’ve thought vitamin D was a nutrient, but it’s not a nutrient. The body manufactures vitamin D. There’s been no human study on whether any vitamin D is necessary. There has certainly been studies in other other animals. A very elegant study in fish showed that the body manufactures all the vitamin D it needs.
Vitamin D is not a nutrient. It’s a transcriptional activator. It’s a secosteriod hormone. There’s a very complex control system here which involves the P300/CBP that I was talking about earlier, as well as the VDR to synthesize from 7-dehydrocholesterolA cholesterol precursor manufactured by humans. When exposed to ultraviolet light converted into vitamin D3. Also known as previtamin-D3.–the cathelicidin, beta-defensins antimicrobial peptides, and the toll-like receptor 2. In fact, the VDR is responsible for at least 913 confirmed genes, which range everywhere from Down’s syndrome to cancers to the calcium sensing receptor and PTH downregulation. Very, very important receptor indeed.
Now, what we found was that only the active metabolite [1,25-dihydroxyvitamin-D], produced by the body, activates the VDR, while the 25-hydroxyvitamin-DThe vitamin D metabolite widely (and erroneously) considered best indicator of vitamin D "deficiency." Inactivates the Vitamin D Nuclear Receptor. Produced by hydroxylation of vitamin D3 in the liver. , which is produced when vitamin D is ingested through food, or through supplements, or through vitamins, that actually stops transcription. All exogenous forms of vitamin D have to be removed if you want to overcome chronic infections. Because all you are doing is aiding the pathogens in their ability to overcome the innate immune system.
We’ve got the various forms of vitamin D docked here [referring to slide] as they exist within the molecules (this is some in silico work) and only one of them has got the 1-alpha hydroxylation needed to activate the VDR. All the others will take up space in the VDR, but they won’t activate genes. They just get in the way.
So we can luckily restore innate immunityThe body's first line of defense against intracellular and other pathogens. According to the Marshall Pathogenesis the innate immune system becomes disabled as patients develop chronic disease. by using a VDR agonist. I identified one called olmesartan. And here we have a protein [referring to animation on screen] this is a human VDR—it’s moving all the time—all proteins are in motion all the time. In the binding pocket, there is the olmesartan drug
This is the olmesartan drug here in yellow [referring to animation on screen]. This is actually the rat VDR shown on this particular slide.
I’ll show you the human and the rat side by side, and show you that the two are not the same. The drug behaves differently in the rat and in the human. If you look at this tetrazole ring [referring to slide], you can see it’s in different orientations in the rat, and in the human. In fact there’s two less hydrogen bonds and that’s stabilizing, or not stabilizing, a critical helix in the structural assembly.
So this drug does not perform in the same way in the rat as it does in the human. Big problem of course, because most our studies are still being performed in animals, even though we have the capability of examining in silico, at the molecular level, there are very few people doing that at this point.
So what happens when you address the problem of the VDR and remove exogenous—that means from outside the body— remove exogenous vitamin D and just allow the body to make the vitamin D it needs, and then at that point the body becomes susceptible to very small amounts of antibiotics—the antibiotics which really did very little when the VDR was overcome by the pathogens, once the VDR has been reactivated again, then the patient starts to respond to antibiotics in the same way that a normal, healthy individual would do. And I’ll be talking more about that this afternoon, when I talk about the protocol that we have put in place to reverse the mechanisms that the bugs have used to overcome our immune systems.
This is the data that was reported at the Autoimmunity Congress in Portugal last September. It shows a small portion of our clinical cohort, observational cohort. Diseases from Rheumatoid Arthritis, Hashimoto’s Thyroiditis, Uveitis, Psoriasis, Type 2 Diabetes, Sjogren’s, Celiac, SLE, right down to Undifferentiated Connective Tissue Disease, Myasthenia Gravis, Diabetes Insipidus—all responded favorably—in all cases with reversal. 81% of the cohort experienced reduced disease and symptoms between 18 and 53 months, and the trial is ongoing at this point.
What was surprising was it wasn’t just the autoimmune conditions that responded to the antibacterial therapy, but also Chronic Fatigue Syndrome—Myalgic Encephalomyelitis, osteoporosis, periodontal disease, cardiovascular disease, Uveitis, cognitive deficiencies, Obsessive Compulsive Disorder, Bipolar, and memory loss, they also disappeared as the chronic inflammationThe complex biological response of vascular tissues to harmful stimuli such as pathogens or damaged cells. It is a protective attempt by the organism to remove the injurious stimuli as well as initiate the healing process for the tissue. disappeared.
So as these people got better, all of these things that we never really associated with pathogens—osteoporosis, cardiovascular disease—well, actually, this is a Chlamydial infection conference, so you would know that Chlamydia has been implicated in cardiovascular disease. There is a study going on at the moment—one of our collaborating physicians is doing a study where he is actually tracking the reduction in arterial wall thickness, year by year, with patients as they recover on the antibacterial therapy. That is unheard of, even when you’re using statins, there is still some gradual increase in the wall thickness. When you actually combat the pathogens, the walls go back to normal.
The homeostasis of other Type 1 Nuclear Receptors is also indirectly upset by the pathogens. And in particular the thyroid receptors. You’ll notice that we had a lot of people with Hashimoto’s Thyroiditis diagnosis, partly that’s because a lot of people with chronic disease have thyroid problems. Because one of the first things that happens when the VDR stops working, the level of 1,25-dihydroxyvitamin-D rises in the body and that shuts down the thyroid metabolism. It also shuts down the adrenal axis, the glucocorticoid receptor is also overcome by these high levels of 1,25-DPrimary biologically active vitamin D hormone. Activates the vitamin D nuclear receptor. Produced by hydroxylation of 25-D. Also known as 1,25-dihydroxycholecalciferol, 1,25-hydroxyvitamin D and calcitirol..
The reason you can’t get rid of chronic disease by giving people vitamin D is because that vitamin D collects in the body and it starts to hit the other receptors at the same time as it’s overcoming the bacterial effect on the VDR.
So when people get sick to a certain level, they’ll no longer respond to conventional therapy. At that point the bacteria are in control. And usually that occurs late in life—except with people that come down with chronic disease—and tragically these days we are seeing even kids coming down with the chronic diseases that they use not to get.
But typically they just cause the ‘diseases of aging’—dementia, osteoporosis, muscular problems—you know them. Normally the microbiota just stays dormant and pops up in late life and people say, “oh yes, I’m getting old.” Well, no, the bacteria are starting to take charge.
I’ve already mentioned that the loss of Glucocorticoid and Thyroid homeostasis leads to diagnosable disease states.
The genomes accumulate gradually during life. Very important—you were born with this microbiota. Depending on what you come in contact with during life, from food, from saliva, from aerosols through the air, and from infections of course. Actual acute infections. Your metagenome will gradually be built up. And so at any point in life it will be different than it was a decade earlier.
The microbiota have access to the DNA transcription machinery. And that’s what I’ve been talking about—taking a strand of DNA and turning it into proteins that can actually do something.
But what’s more important, the DNA repair mechanisms become susceptible to all of this plethora of imponderable effects from the bacterial DNA. So you get modification of the human DNA repair mechanisms by ‘junk’ from the metagenome. HIV integrates itself into the human genome. HHV6 integrates itself into the human genome. There’s a lot of work being done to show that bacteria do the same thing. But you saw in the mycobacteria study that there were a whole lot of genes that were identified that were not previously documented as being in mycobacteria or in Homo sapiens. And that’s because the interaction between the genomes.
This came out just last week on Reuters, Wednesday, April the 1st, and no, it’s not an April 1st joke. It was reporting a study on heart attacks. I love this word, “germy,” mouths linked to heart attacks, study finds. But what they found was, “people who had the most bacteria of all types in their mouths were the most likely to have had heart attacks.” That’s not a surprise to us. If the body can’t keep the number of species down to the so-called friendly, or at least, mostly friendly bacteria, then the disease mechanisms are going to take over and people will get sick. And that’s exactly what this study on cardiovascular disease found. It wasn’t that Chlamydia was present. It wasn’t that Mycobacteria were present. It wasn’t that mycoplasma were present. The total number of species that could be identified in the saliva was the best indicator that somebody was likely to have had a heart attack.
We are talking about a metagenomic–very many genomes–microbiota.
So, the last question. Really, second to last is, “Why have murine models failed?” Now what’s a murine modelA model of disease which uses rats or mice to mimic human conditions.? That means, mice and rats. When we test these diseases out in mice and rats. Which we nearly always do, before we give it to mankind.
Unfortunately, the immune system of mice and rats is very different to the human immune system. It’s evolved totally differently. Mice and rats live in a different environment to us. And that seems to have been forgotten by science for the last 50 years.
In particular, if you look at the VDR homology–and that means the […] in the VDR, the shape in the VDR that’s produced–the VDR of Homo sapiens transcribes different genes from the VDR of other mammals. Even the VDR of higher primates.
And in particular, the VDR from the murine and canine (or dog) genomes doesn’t transcribe Cathelicidin, or the Defensins. So if a bacteria evolves to knock out the VDR, it makes human beings sick. But it’s not going to make rats and mice sick, because it doesn’t knock out their primary defenses. It doesn’t knock out Cathelicidin, or Defensins, like it does in man.
So the human metagenomic microbiota will not survive when transfected into a mouse. If you take the human microbiotaThe bacterial community in the human body. Many species in the microbiota contribute to the development of chronic disease., put it into a mouse, the mouse will be able to deal with it because, it’s VDR isn’t really as important as man’s is.
Different species and different mutations are necessary if the microbiota was to knock out the different gene pathways needed for survival in a mouse. That is one of the fundamental reasons why chronic disease has remained ‘of unknown cause’ for the last 50 years, in my opinion, is the reliance on mouse and rat models, and to a lesser extent on other mammal models. Without really questioning whether a mouse has the same immune system as a human being.
Until the genome was cracked–until we could crack genomes, until we could sequence the DNA, and figure out exactly what we were dealing with, what made this organism tick, and whether the organism is Homo sapiens, or bacteria, or a virus like HIV, something makes that organism tick–and now, we have just started to figure it out. We have decades, maybe a century of figuring out this imponderable problem that I talked about earlier.
But we’ve just started. And until the genome was cracked, we only had the postulates of Koch as a guide. The postulates of Koch have formed the basis for infectious disease–clinical infectious disease at least–for a century. They caused us to look for a single species that was causing the disease process. Koch basically said, you have polio virus, and it causes, polio. Koch basically said, you have single pathogen, and it causes a singular disease. And that’s not what we’re finding. We’re finding is that the human body is a whole pelethora of pathogens, a whole pelethora of genomes, a metagenome.
So science became fixated on the co-infections–Chlamydia, Bartonella, Borellia, HHV6, EBV–have all been implicated in widely varying diseases. If you look at the literature on EBV, you’ll find that just about every disease has been blamed on EBV at some point in time–Epstein-Bar virus. And the same with Chlamydia. So many different diagnosis are supposed due to Chlamydia. Because these are the pathogens—that when the body’s own immune system becomes weak–these are the pathogens that the body can’t get rid of. So they stay there. And they can be seen, and observed, and measured. And people say, “Ah! That person has a Chlamydia infection! The Chlamydia is making them sick.”
Well yes, the Chlamydia might very well be making them sick, because Chlamydia has got some very nasty toxins itself. But in order for the immune system to allow that Chlamydia to flourish, they first had to have a suppressed immune system from the metagenomic microbiota.
So for the last century, science became fixated on what are predominantly co-infections and have missed the primary disease mechanism. The primary disease mechanism being the intraphagocytic, intracytoplasmic metagenomic microbiota.
The antimicrobial peptide work that I am relying upon was done by Brahmachary’s group in Singapore. I cite it in all of our papers, actually, I think you’ll find it cited.1) And they used an in silico analysis. What they did was look at the DNA and they figured out what activated the DNA. Whether it’s a VDR receptor, whether it was a P300 receptor, and from that they figured out what was responsible for each of the antimicrobial peptides. Cathelicidin, beta-Defensins, and TLR2 have all been confirmed in vitro as coming from the VDR. So the 24 families in silico and the actual Cathelicidin, the key metabolites have been confirmed in vitro by various groups, more than one group.
This is a slide that just came out last week from a group studying HIV. And what we’ve got here [referring to the animation on the screen] is we’ve got two cells–well a number of cells–but there’s a cell which has been infected with HIV. We have a cell which is brushing up against it. And at the junction of the two cells, the HIV is trying to get through the cell membranes into that [uninfected] cell. And in fact, that’s exactly what’s going to happen. This HIV infection is able to cross the membrane. You can see it frozen there as it’s starting to cross. In a little while you’ll find there will be fluorescent staining inside that cell, or fluorescent artifacts inside that cell to indicate that the tat protein has–there it is, it’s broken off, it’s now inside the other cell. One of the reasons that we don’t find these pathogens in the blood is because they don’t need to be in the blood. They can pass from cell to cell. That’s now been shown in HIV with this very elegant study from March 2009, “Microscopy of of HIV Transfer Across T-cell Virological Synapses”2) (which means across the membranes). And of course that cell now also will become infected and will eventually bud into virions.
All of this without the bacteria having to have to deal with what’s in the bloodstream. It hasn't had to deal with any antibiotics the patient’s been taking. Because it’s existing totally inside the cytoplasm of the cells.
Firstly, I don’t follow them. But there have been studies done. The two that come to mind, are the study out of Imperial College in London, where they looked at the urine, they looked at the various proteins that were found in the urine that could not be produced by the human body itself, but were being produced by bacteria which were present in the urine–present in the kidneys, one assumes. And what they found was that there were very distinct grouping—the Japanese population, the Chinese population, and the US population—were all different. And when Japanese moved to the US, their microbiota changed to the US population. Because the food changed, their environment changed, everything changes. So we very much are a product of the bacteria that make us. Now, that was not specifically looking at saliva.
The study from Max Plank that looked at saliva did in fact look at 10 locations throughout the world to make sure that they had good geographic diversity. And if you look up the paper that I cited there you’ll find the detailed data.3)
Right. Well, for example, Helicobacter Pylori, has been used to track migrations in the South Pacific. Because it has been isolated from very ancient skeletons, and other DNA fragments, in various regions of the South Pacific. And depending on the particular genome that they found at these various points, they were able to figure out how the populations migrated in history based on the Helicobacter Pylori genomes that they found.
Borellia was recently located in an Egyptian mummy. They were able to isolate some Borellia genome.
So we have an incredible opening up of discovery over the next decade or two as we start to get our minds around this concept of a microbiota, a community of pathogens
Yes. Many people are exposed to Borellia. And to Chlamydia. But only a fraction of them become very ill. Only a fraction of them become ill, in fact. Some of them can get rid of the organism immediately. And that’s because it’s the innate immune system that is key.
The status of the immune system when that person gets challenged by the acute pathogen that is so important in determining what’s ultimately going to happen during their lives.
And therefore it’s very important to study the family, to study the maternal line–because these bugs are primarily passed down the maternal line–to study the maternal line, and you will find that a kid who’s got Lupus, had a mother who had arthritis, and grandmother that had thyroiditis. You can see the diseases. Once you realize that all these diseases, the chronic diseases, are inter-related from the same cause, you can track them from within a family, and also horizontally within families as well.