9th Annual BMRP Investigator Meeting - Abstract
Metagenomic Approach to Assess the Virome of Patients with IBD
Weiwei Wang1, Bo Meng1, Mandana Rhabari1, Jun Wang 2, Gane Wong1, 2 and Andrew Mason1,a
1Department of Medicine, University of Alberta (Alberta, Edmonton, Canada); 2 Beijing Genomics Institute (Shenzhen, China)
Our laboratory is working on the hypothesis that certain viruses may be relevant to the etiology and pathogenesis of IBD. To identify the complexity of viral populations, we are investigating the presence of all agents in different bodily compartments in patients with IBD and control subjects that constitute the “Virome” using a metagenomic approach with massively parallel deep sequencing. Our long-term objective is to distinguish potential pathogens from common agents that cause no apparent harm and determine whether specific viral agents are associated with IBD. For the first approach, viral preparations are isolated from plasma or feces from patients with IBD and matched controls; total DNA or RNA is then cloned into libraries and sequenced using the Illumina HiSeq platform to identify viral genomes. For the second approach, total RNA is extracted from PBMC, colon and mesenteric lymph nodes from patients with IBD and control subjects; ribosomal RNA is removed and cDNA libraries are made for high throughput sequencing with the Illumina HiSeq platform. Approximately 4 x 107 sequences of 2 x 75bp paired end data are obtained per run to generate approximately 2 GB data. Both SOAP and BLAST algorithms are used to identify and assemble non-human/non-bacterial sequences and to categorize known and novel viral sequences.
In our preliminary studies of colon cDNA, we observed an order of magnitude higher representation of bacteriaphage and mammalian viruses in pooled UC and CD samples as well as individual UC samples (n=5) compared to CD (n=5). Specifically, a higher frequency and range of herpes virus sequences (HSV2, CMV, EBV and HHV6) were observed in UC colon samples. All samples had a distribution of other common viruses, such as papillomaviruses and adenoviruses as well as plant viruses and bacteriophage. These studies will serve to determine complexity of the virome in patients with IBD and help determine whether specific agents contribute to disease.
a Principal Investigator