Metagenomics vs 16S rRNA – How do they compare?

Author: Dr Gregory Crocetti

07 November 2018 Education
Microba's metagenomic gut analysis provides higher resolution picture of the microorganisms in your gut that 16S technology

Growing interest in gut health and advancements in technology have made gut microbiome analysis accessible to individuals through one of two common methods: metagenomic and 16S rRNA gene sequencing. So how do they differ and which one is right for you?

The Evolution of Gut Microbiome Analysis

The human gut contains trillions of microbes, including a variety of fungi, archaea, protists and viruses – and importantly, hundreds of different species of bacteria. When scientists first began exploring the microbiome, they only had one tool available for studying microbes. This was called ‘culturing’ and involved placing a sample into a growth or ‘culture’ medium to see what microorganisms grew. Unfortunately, most microbes don’t like to grow in culture and so for a long while we drastically underestimated the number and types of microorganisms that were present in the gut.

In the last few decades, technology has advanced by leaps and bounds, and it has become possible to sequence the genetic material or DNA of microorganisms. DNA sequencing has revealed that there is greater diversity present in the microbial world than we previously suspected and that these previously unknown microorganisms play an important role in our health and are an essential part of us.

Single Genes (16S ribosomal RNA gene) vs Metagenomics

Two main methods of gut microbiome analysis have emerged as DNA sequencing technology has advanced: 16S rRNA gene sequencing, a method that sequences one single gene present in bacteria; and metagenomic sequencing, a more recent method that sequences all the genes (DNA) in all microorganisms present.

Until recently, 16S ribosomal RNA sequencing was the standard approach used to analyse the gut microbiome. This method sequences the DNA from a single gene common to all bacteria, called the 16S ribosomal gene. This gene acts as a ‘fingerprint’ to identify different groups of bacteria. By comparing all versions of this gene identified in a sample, a picture is painted of which types of bacteria are present and in what proportions1.

However, in the last few years, a better way to study microorganisms has emerged called ‘metagenomics’. Metagenomics sequences all the DNA present in a sample and can tell us not only which microbes are present in a sample and how many there are, but also what they can do. This advancement in DNA sequencing has been driven by two factors. First, the cost of DNA sequencing has plummeted, offering the potential to affordably sequence far more genetic material from a microbiome sample than was previously possible. At the same time, modern supercomputers now can crunch a vast amount of DNA data to assemble the genomes (all the DNA that makes up an organism) from microorganisms that were previously unknown2.

Metagenomics reveals a whole new level of detail

The genome of a microorganism essentially provides scientists with the ‘blueprints’ for that microbe, allowing them to ‘see’ what it can do. For example, the genome can be used to uncover a microorganism’s potential to break down carbohydrates, proteins and fats; produce short-chain fatty acids and essential vitamins important for health; produce metabolites associated with inflammation and more3. This profile of metabolic strengths and weaknesses can then be used to suggest targeted dietary changes to achieve a healthy balance within the gut microbiome.

Understanding small differences makes a big difference

Metagenomics can also accurately measure a broad spectrum of gut microbes down to a species and even strain level, including potentially parasitic or beneficial yeasts, fungi, protists and DNA viruses (e.g. bacteriophages)1.

An example of the importance of distinguishing between strains has recently been demonstrated with the common gut bacteria Faecalibacterium prausnitzii. This species has been widely accepted as a positive indicator for human gut health, due to its ability to convert dietary fibre into the beneficial short-chain fatty acid, butyrate. However, recent metagenomic analyses revealed an important variation in the genome of a strain of F.prausnitzii (L2-6) – where researchers observed it was associated with a chronic skin disorder called atopic dermatitis (due to a critical difference in how it makes butyrate)4.

The potential for metagenomics to resolve subtle differences between similar microbes can have a huge impact on clinical research and how we interpret the microbiome of individuals. In turn, this information can help progress the study of the gut microbiome and how it influences health at a much faster pace.

References

  1. Ranjan, R., Rani, A., Metwally, A., McGee, H. S., & Perkins, D. L. (2016). Analysis of the microbiome: advantages of whole genome shotgun versus 16S amplicon sequencing. Biochemical and biophysical research communications, 469(4), 967-977. https://doi.org/10.1016/j.bbrc.2015.12.083
  2. Deneault, S. (2017). Sequencing the microbiome: Are you getting the full story? Retrieved from http://blog.dnagenotek.com/microbiome/sequencing-the-microbiome-are-you-getting-the-full-story
  3. Jovel, J., Patterson, J., Wang, W., Hotte, N., O'Keefe, S., Mitchel, T., ... & Wong, G. K. S. (2016). Characterization of the gut microbiome using 16S or shotgun metagenomics. Frontiers in microbiology, 7, 459. https://doi.org/10.3389/fmicb.2016.00459
  4. Song, H., Yoo, Y., Hwang, J., Na, Y. C., & Kim, H. S. (2016). Faecalibacterium prausnitzii subspecies–level dysbiosis in the human gut microbiome underlying atopic dermatitis. Journal of Allergy and Clinical Immunology, 137(3), 852-860. https://doi.org/10.1016/j.bbrc.2015.12.083