Single Virus Genomics - the future of virus research?

Bacteriophage Lambda particles - like those used in this study

Viruses are the most common organisms on this planet and also most likely the most diverse. It is therefore unsurprising how ecologically important they are in particular environments and indeed global ecological processes. Just look at the amount of papers published on the subject recently. What is preventing us from extending these studies to more environments and preventing us having a closer look at what viruses are present is our inability to study small amounts of these microorganisms. Our knowledge of global viral diversity has most likely been biased by this lack of single-virus investigations.

Metagenomics?

In order for us to study viruses at the molecular level we have to somehow get the virus. This usually requires infecting certain cells with the virus in order to take advantage of virus replication and produce more and more copies of viral particles. This means that we have to find cells that the virus infects, which is difficult for some viruses found in environmental samples where no knowledge of host is available. This problem has been encountered with bacterial species where it was found that we were only able to grow in the lab a small percent of the total microbes from a single sample. The field of bacterial 'metagenomics' (get your free pdf review here) took off when it became easier and easier to study the molecular biology of those bacteria without the need to grow them in the lab; we are now able to fully sequence bacterial genomes without the need for culturing. These developments are now being applied to virus species in a diverse range of environments.

Single Virus Genomics (SVG)

A recent study in PLoS ONE reports the development of a strategy to overcome such problems; Allen LZ et al (2011) - of the J. Craig Venter Institute - show, in a 'proof-of-principle' study, how they can now separate individual virus particles from a mixture and submit these single viruses to genomic analysis. The results of this paper are likely to enhance many areas of virology including ecology, immunology and basic biology which is probably why the group have patented the process. Although the paper didn't actually deal with environmental samples, a paper by the same group is currently in the works.

SVG strategy: seperate, agarose and PCR

What Allen et al did was make themselves a well defined mixture of bacteriophage (viruses of bacteria) particles. They filtered this suspension in order to only get the viruses in their sample. These viruses were then dyed by the attachment of fluorescent molecules to their surfaces so as they could be separated using what is known as a flow cytometry machine. This machine allowed only single virus particles to pass through and were then 'printed' onto a microscope slide in a 'high-throughput' fashion. To stabilise the particles they embedded each virus in a bead of agarose - a gel-like substance; they were also able to visualise each virus found within the agarose. Once they had separated each particle out they then used a special kind of PCR to amplify the entire virus genome inside each agarose bead. The products of this were then used for DNA sequencing and finally assembled into the whole virus genome using computational methods.

The group are currently attempting to further optimise this protocol to make it easier and more accurate especially considering that this method cannot yet be easily used for RNA viruses nor viruses about which no sequence information is known. Also, the viruses amenable to this technique would have to be pretty stable to survive the whole process. You can read about their plans for this in the paper's discussion.



A single bacteriophage particle

What does this mean for virology in general?

Virologists have generally concentrated on studying populations of viruses in their investigations; animals or cells are infected with one million infectious virus particles and the effects on host functioning are measured, for example. What this research shows is that in some cases it is entirely possible to look at the genome of individual particles and not have to rely upon mixtures of thousands, if not millions, of genetically variable viruses. The major areas of science that may benefit from this development will be those that rely on virus discovery, for example, the assessment of total virus diversity in a given environmental sample. The ability to uncover low concentrations of virus in an infectious disease context in humans may also benefit, especially if there is difficulty in growing the virus in tissue culture. Groups studying the genetic diversity of viruses can now accurately determine the exact genome sequences of individual viruses and also quantify them. This research will hence benefit a wide range of disciplines within virology in general.


 

Allen LZ, Ishoey T, Novotny MA, McLean JS, Lasken RS, & Williamson SJ (2011). Single virus genomics: a new tool for virus discovery. PloS one, 6 (3) PMID: 21436882