Exposed... The Secret World of Microbes: Metagenomics at Work

By Dr. Soumya Biswas

Microbes rule the world. It is as simple as that. Even if you cannot usually witness them, they are indispensable for all life on Earth. Everything that is alive on earth makes the biosphere. The collective actions of microbial communities shape the biosphere all over the world. Microbes also have the endless capacity to transform their surroundings.

The operation and stability of the biosphere are dependent on the fantastic abilities of these microbes. They can breathe metals, eat rocks, transform the inorganic to the organic, and break the toughest of chemical compounds. No microbe can do the complex transformations alone. It takes a community of microbes to do so. The supremacy of these communities lies hidden in the metabolic resourcefulness of their constituent species. Acting together, they regulate most of the matter and the energy transformations on our planet.

Take a deep breath. Half of the oxygen in the air, you just breathe in, is produced directly by the microbes. The microbes are present at billions of numbers in a liter of ocean water. They transform 50 billion tons of carbon dioxide to sugar every year. While doing so, they produce oxygen. The plants produce the other half of the oxygen. This is also possible due to the presence of microbes in the soil.

Take a bite of the Apple. Every bite you just ate was possible because microbes have been working every second of every day since time immemorial. Nitrogen is essential for every life forms including plants. Nitrogen is the most abundant component of the atmosphere. Nevertheless, it is not readily accessible to use by any living thing except microbes. Some Bacteria and Archaea take up the unusable nitrogen from the atmosphere and transform it to usable nitrogen compound, ammonia. 193 million tons of nitrogen is fixed by microbes every year. All life on Earth uses this usable form of nitrogen.

Drink a glass of water. Did you smell gasoline? Obviously not. Every drop of the water you drank is potable because of the microbes. We are continually polluting the ground with leaked or spilled gasoline. These gasoline seep down to groundwater. They are broken down by microbes living in the subsurface, below our feet. The complex community of microbes works together until the gasoline is transformed into harmless carbon dioxide and water.

Making the atmosphere breathable, putting food on our table, filling our glass with potable water are just a few examples of countless ways microbes sustain the biosphere. Metagenomics reveals the intricate details of what the microbes are doing and how they are doing it.

Most of the living microbial cells that can be seen under a microscope cannot be grown on Petri plates or cultures in test tubes. This phenomenon is known as the ‘great plate-count anomaly.’ It is estimated that less than 1% of the living bacteria present in soils, and even lesser in case of water, can be cultured.

By the mid-20th century, pure cultures developed as a gold standard for research and formed the foundation of practically all current knowledge of the microbial world. However, culture-dependent techniques fall short of uncovering the environmental processes, biofilms, energy and matter flux, and biogeochemical cycles. Whatever we know about microbes by culture-dependent techniques is ‘laboratory knowledge.’ It is done in the unusual and unnatural conditions of growing them in artificial media in a pure culture deprived of any ecological context. Understanding microbial communities in the real world will require the deployment of metagenomics.

The term metagenome first appears in the October of 1998 in work published by Jo Handelsman and others in the journal Chemistry & Biology. Since 1998 the science of metagenomics had a snowball effect on the understanding of science in a new age context. Although the science of metagenomics is comparatively young, the technique makes it possible to explore microbes in their natural habitat, the complex communities in which they typically live. It now appears that many microbes function in nature as groups of interconnected webs of chains. They are sometimes physically connected and often metabolically connected.

Metagenomics is the study of genetic material, DNA and RNA, extracted directly from the environment. Metagenomics comprises cultivation-independent, genome-level and gene-level account of communities or their members. It is the science of discovering, modeling, understanding and managing at the molecular level the dynamic relationships between the molecules that define living communities and the biosphere.

Let us take a look at how metagenomics work.

1.      The sample processing is the first in any metagenomics venture. The extracted DNA must represent all cells present in the sample. Extraction should be sufficient in the amount and high in quality.

2.      The second step is preparing the DNA for sequencing. Two general approach is there for this step. You can amplify the part of DNA you are interested in, like 16SrRNA gene or you can cut the DNA into small pieces and sequence them all. These ready to sequence DNA fragments are called libraries.

3.      The third step is sequencing the DNA (the metagenomic library). Over the last 10 years, metagenomic sequencing has increasingly shifted from classical Sanger sequencing technology to next-generation sequencing (NGS). Several options come with its pros and cons.

4.      The fourth step is acquiring and processing of sequence data. The approach for analysis will also vary depending on the previous steps.

5.      The last step is obtaining your answer from the analysis. If your library was a PCR product of 16SrRNA, then the result is mostly the microbes present in the sample (biodiversity). If your library was fragments of all DNA, then the answers are the microbes present in the sample (biodiversity), their function in the environment (functional diversity), and their way of performing their function (metabolic pathway).

Every step of this workflow is complex and customizable depending on the question you want to be answered (experimental design).

We require more metagenomic studies to characterize microbial communities better, comprehend microbial dynamics, learn microbial and environmental interrelationships, detect and decrypt microbial diversity, discover functions, and elucidate microbial adaptation and evolution. We still do not know the extent of what we do not know. Now we have the tools and techniques to expose the invisible. I argue the time has come for us to venture even more into the secret world of microbes.