When we talk about the microbiome, let's step back a bit and think about what we have learned and how. The presence of microbes of many types has been known for a long time, but it wasn't until "supercomputers" that we could really flesh out the substance of who is where and in what amounts, This can be called the taxonomy or phylogenetics, terms that relate to such maps and relationships. Groundbreaking, complicated work out of brilliant labs such as that by leading expert Rob Knight PhD looked like this:
These "high throughput" systems lead to widespread efforts to profile the microbial community. A link to a description of the process circa 2009 is included below. Physicians and scientists leaped at the opportunity to infer and confer around this data, and wild speculation and theory abounded. The Human Microbiome Project, similar to the Human Genome Project, was a massive undertaking that led to more questions and a deeper understanding of the complexity of the system. The work by these scientists was groundbreaking. Collaboration on the Human Gut Project included public participation. Yes, you mailed them your poop. But data about age, exercise, diet also were recorded to begin to untangle this hidden world. Kudos to these noble explorers.
The diversity seen was astounding. The ability to alter or use this information clinically was not. Slow, plodding clinical studies with mixed results and unexpected outcomes became the norm for over a decade and really continue today. We had some early success in the treatment of Clostridium Difficile, though for reasons we still don't fully understand! We saw relationships in digestive diseases, but no clear pathogenic causality such as Crohn's disease or ulcerative colitis.
But this also made intuitive sense to medical scientists, as we have understood molecular roles in human physiology. Let's step back once again to understand what the genetic testing for a microbiome tells us. To do so, we will need to review how DNA becomes proteins first. This may seem complicated, so send questions and I can explain in follow-up!
Here is a simplification before you watch this animation:
1. Unzip the DNA,
2. Make a copy strand called “messenger” RNA.
3. Rearrange mRNA for “transfer” RNA and “ribosomal” RNA to use to make proteins.
The microbiome has largely been studied using “16S” ribosomal RNA microbial profiling for studying bacterial phylogeny and taxonomy. The 16S rRNA gene is the most established genetic marker used for bacterial identification and classification, mainly because it consists of both highly conserved and hypervariable regions. Most organisms have 5 – 10 copies of this gene which have been mapped for known organisms:
(Scientists: The conserved regions can serve as universal primer binding sites for the amplification of the whole gene or fragments of the gene, whereas the hypervariable regions contain species-specific sequences that can discriminate between different bacteria and archaea. Similarly, the internal transcribed spacer (ITS) region is utilized for the taxonomic profiling of fungi.)
With the sequencing data available in public databases, we can semi-quantitatively describe bacteria, archaea and fungi present in complex biological samples. This method is considered by many as the gold standard for analysis of whole bacterial, archaeal and fungal communities and their members.
Determining the DNA sequence of the 16S rRNA gene and ITS region is a preferred method for studying the taxonomy of microbiome and microbiota members as it can be used to:
Characterize bacteria that cultivation misrepresents
Profile hundreds of microorganisms from a single analysis
Semi-quantify the relative abundance of microbiome members
Study complex microbiomes
Provide faster and more accurate classification than traditional identification methods like cloning and culturing
Identify low-abundance bacteria
So when you now read "microbiome", you can understand better how it has been explored. We look for specific genes known to have persisted through evolution, that are specific to species. You should also now have an idea of the limitations of this information. It really doesn't exactly tell us what is going on in there! But knowing what is alive in the ecosystem is the first step to understanding how it works. And there are measures of regulation and expression from genetics, termed metagenomics, that can help us begin to explore function.
This also helps to explain why there is so much misinformation out there. Hopefully, armed with this knowledge you will begin to explore data and findings about the function of this ecosystem and how it impacts our human physiology. It should be noted that "functional" medicine would really be the study of such physiology and not an alternative to it!
In my next post, we'll jump into how the DNA data has been used and misused. What are the promises and failings? Why does it all seem so contradictory or unsure? How are we beginning to use proteomics and molecular interactions to learn more? How does the presence of these microbes, our own bodies mechanisms, disease states and how we alter both with the molecules we ingest as food impact our health? That's what you really want to know, right?
There are emerging data that has high strength of support, data that counter prior assumptions and a whole lot of assertions based on limited data and highly subjective measurements of outcomes. For sure!
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