Lab Rats: IITB student finds giant viruses in Mumbai lakes

In the first article in our series ‘Lab Rats’, Gokul Rajan talks about the ground-breaking research going on in the Department of Biosciences and Bioengineering. If you too would like to use this platform showcase your research, please contact us at insight@iitb.ac.in

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I stand on the sixth floor of the Department of Biosciences and Bioengineering at IIT Bombay. In Lab 601, Professor Kiran Kondabagil, who heads the Molecular Virology laboratory here, is showing me the work of Disha Bhange, an M. Sc. student in the lab. Amrutraj Zade and Avi Shukla, two senior PhD scholars in the lab also guided her in the project.

I am flabbergasted upon seeing the Transmission Electron Microscopy (TEM) images being flashed on the screen – giant viruses of sizes ~400nm!

First Findings of Giant Viruses
Isolation of such large viruses is a relatively new event and is yet another example of serendipity in science. While investigating a pneumonia outbreak in 1992, researchers came across some particles within an Amoeba. Since some pneumonia-causing bacteria do reside inside Amoeba, the researchers thought these to be another pneumonia-causing bacterium and named it Bradford coccus, after the name of the place where it was isolated- Bradford in England. It was only in 2003, that researchers at the Université de la Méditerranée in Marseille, France identified this organism as a giant virus with a diameter of ~400nm, which measures ~600nm with the surface filaments. This was then named as Mimivirus, for mimicking microorganisms.

The average virus size is about 80-100 nm. So, it came as a great surprise, when these were first identified.

While we still do not know if Mimivirus has something to do with pneumonia infection, this serendipitous discovery has opened door to a host of new question on the evolutionary history of viruses.

This event set the stage for the discovery of many new large viruses. Recent additions include the Megavirus (~440nm diameter) discovered in 2011 off the Chile coast and the Pandoravirus which measures 1000 nm. Pithovirus, the most recent discovery (as of 8th May, 2014!) was reported this year and is also the largest in the group, which was isolated from a 30,000 year old sample of permafrost in Siberia, Russia and measures 1.5 µm in length and 0.5 µm in diameter.
Pithovirus

Disha’s Discovery at IIT B
The aim of Disha’s thesis was to hunt for unexplored giant viruses infecting Amoeba and Tetrahymena in the rich and diverse geological and environmental conditions found in India.

Last summer, Disha started by collecting samples from 15 different lakes in Navi Mumbai and Thane, including the Powai Lake.

She processed her samples and screened them for new viruses. All large viruses mentioned previously were also isolated from testing against amoeba. While one can simply write that the samples were processed in a single statement, the task itself was not so simple. Disha describes that standardizing the protocols to process the samples was the most difficult part of the entire experiment and it took her months to finish this step. She also adds that this was also the most useful step and it taught her a lot about the scientific method – from standardization of protocols to the usefulness of controls in any experimental design.

She has now isolated large viruses from three different samples, sample 11, 13 and 18 which correspond to water from Chincholi, Shivaji and Khairane Lakes respectively. The TEM analysis shows that sample 11 is remarkably similar to the Mimivirus with an even larger diameter (~420nm) but lacks filaments on the surface. Sample 11 also showed small sized viruses (<200nm) with a bowtie shaped structure (see the figure 2B) which Disha says, “…our team predicts to be a unique vertex for genome (complete set of an organism’s DNA) delivery”. Sample 13 showed smaller viruses (~127-136nm) which according to Disha, appear to belong to the Marseilleviridae family. There’s nothing Martian about Marseillevirus by the way, it is just named after the French town Marseille!

Disha’s hunch about the virus particles measuring ~450nm in sample 18 was first that they were Mimivirus, but further molecular analysis showed that they’re more closely related to Marseillevirus. They are yet to prove the uniqueness of these viruses, which will be done by further molecular and genomic analysis. When asked what the ‘Eureka!’ moment in the project was, she answers,”The moment when we all saw the image of a virus in the sample 11 having unique bow tie-shaped structure”. The joy of research in science and engineering comes from the excitement of discoveries like this, which keeps one motivated.

Figure 2: [A] shows the TEM image of a purified mimivirus which measures ~400nm. [B] and [C] are the TEM images of viruses isolated from sample 11 measuring <200 nm and ~420nm respectively. [B] Shows the bowtie shaped vertex. [D] is the TEM image of the other large virus isolated from sample 18 measuring ~450nm. The smaller viruses isolated from sample 13 are not shown here. Picture credits: Disha Bhange et. al. at SAIF and BSBE, IIT, Bombay. Figure 2: [A] shows the TEM image of a purified mimivirus which measures ~400nm. [B] and [C] are the TEM images of viruses isolated from sample 11 measuring <200 nm and ~420nm respectively. [B] Shows the bowtie shaped vertex. [D] is the TEM image of the other large virus isolated from sample 18 measuring ~450nm. The smaller viruses isolated from sample 13 are not shown here. Picture credits: Disha Bhange et. al. at SAIF and BSBE, IIT, Bombay.[/caption] The Importance of the discoveries
The size of these viruses certainly contradicts the general perception of viruses, but what interests scientists more is their large genome. Studying the genome of these larger viruses might lead to a complete revision of our understanding of evolution of viruses.

Traditionally, viruses have been best described as organisms at the edge of life, as although they carry and transfer genetic material and reproduce, most of them, so far, did not possess many other qualities characteristic of living cells. And hence, the surprising result from the genome sequencing of such large eukaryotic viruses (which are grouped under NCLDVs – Nucleocytoplasmic Large DNA Viruses) is the finding of genes for processes like replication, translation, DNA repair and metabolism which are not found in most other viruses or the processes are carried out differently. All these processes are a result of higher number of certain gene sequences (involved in making proteins) in these newly discovered viruses, which makes the discovery of these giant viruses important, flying in the face of our understanding of viruses as tiny non-living entities and opens up a debate on a fourth domain of life after eukaryotes, archaea, and bacteria.

Disha’s finding of these large viruses from an Indian lake infecting amoeba is probably the first such report from India and confirms the existence of such viruses in this part of the world. While all her results are certainly intriguing, a lot more remains to be done to characterize these viruses, build upon these discoveries, and further our understanding.

If you too would like to use this platform to showcase your research, please contact us at insight@iitb.ac.in