A patient died from an unknown reason, and the doctors suspected that the causative agent was a virus. However, the determination of the virus was not successful. Because we are an NGS laboratory, we were asked to determine it with DNA sequencing. Unfortunately, if we do not enrich the viral DNA, we will detect the human DNA mostly. It would be like searching a haystack for the needle. Could anyone recommend me an enrichment strategy (either on nucleic acid or at the whole virus level)? Of course, I know also other reasons why this task can be hardly solved, but the first point would be the enrichment.
ResearchGate link for the article mentioned by Michael Lauck:
https://www.researchgate.net/publication/51980539
Article A Viral Discovery Methodology for Clinical Biopsy Samples Ut...
Thank you Michael and David. I just read the article, it could be very useful.
You have three options:
* Human DNA depletion
* Viral DNA enrichment
* Extremely deep sequencing
To expand on that third one, as an alternative to enrichment/depletion you can genotype on a platform where even 1% of the sample containing viral DNA is going to produce meaningful results (e.g. a 600Gbp-1000Gbp run on an illumina HiSeq).
I did a bit of web searching, and came up with a couple more specific solutions:
Here's someone else who has done cheap human DNA depletion with some success, testing CF11 cellulose columns against Lymphoprep (a sucrose density gradient filter):
http://www.malariajournal.com/content/pdf/1475-2875-11-41.pdf
I guess the idea behind the Lymphoprep method [http://www.stemcell.com/en/Products/All-Products/Lymphoprep.aspx] is that you filter out the human cells by size, and hope that there's enough extracellular virus floating round for subsequent genotyping.
There's also a NEBNext Microbiome DNA enrichment kit, which filters out host DNA. It apparently works by capturing methylated DNA (essentially absent in viral / bacterial populations):
https://www.neb.com/products/e2612-nebnext-microbiome-dna-enrichment-kit
Presumably there are similar kits from other manufacturers.
Another approach is the Virochip, e.g.:
https://www.researchgate.net/publication/51114072
While this will not identify completely uncharacterized viruses, it will detect many known and closely-related viruses. There are always potential pitfalls with this or any other approach, though...see the sad story of XMRV.
Article Using a Pan-Viral Microarray Assay (Virochip) to Screen Clin...
Thank you David, Kenneth and Rohan! All the suggested solutions were interesting for me, especially the NEB kit. Thank you very much.
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