*Purpose: Blood virome of healthy individuals is frequently predominated by member of families such as Herpesvirus, Anellovirus and Polyomaviridae. The viral load of commensal species such as Torque Teno virus (TTV) is maintained below a pathogenic level through active immune clearance. The prevalence of anelloviridae in blood is reported to be associated with immunosuppressant dosage and may correlate to the risk of antibody mediated kidney rejection. Ergo, plasma viral abundance may help monitor the degree of immunosuppression in solid-organ transplant (SOT) recipients. Here we present a computational tool (Vira-ome) which identifies the presence of a targeted set of DNA viruses following NGS sequencing of plasma cell-free DNA (cfDNA) with accuracy and sensitivity.

*Methods: We developed a pipeline in which non-host NGS data (reads not mapped to the human genome) undergo a reference-assisted assembly operation and then taxonomic annotation using KrakenUneq (a unique K-mer based classifier). We trained the KrakenUneq on a database of ~12,000 viral genomes (from NCBI) and curated, in-house, on the basis of length and genomic-similarities. We used 3 different K-mer values (16, 21, 31) to train KrakenUneq, and final predictions are made by applying a majority-wins rule. We tested our method on 30 simulated and 29 clinical samples obtained from a biorepository.

*Results: The Vira-ome tool currently screens for the presence of fifteen viral species: human adenovirus, human herpesvirus 1, 2, 4, 6, 7 and 8, BK polyomavirus, human parvovirus B19, JC polyomavirus, KI polyomavirus, WU polyomavirus, TTV, human cytomegalovirus, and varicella-zoster. On a simulated set of viral sample mixes, our protocol had 100% accuracy with no false-positive predictions. For 29 clinical samples, our pipeline predicted: (A) TTV in 13/29 samples, (B) pathogenic viruses in 11/15 cases, predominated by BK polyomavirus (7/15 samples) and (C) at least one viral species in 22/29 cases. Selected samples were then analyzed by qPCR to confirm the presence of BKV, JCV, HHV7 and EBV in 8/9, 3/4, 1/1 and 2/2 computationally predicted samples, respectively. Viral loads ranged from 6 – 10⁶ copies/mL. We have also observed (i) a lower viral diversity in humoral rejection cases and (ii) prevalence of human betaherpesvirus 5 & 7 in non-rejection cases.

*Conclusions: The Vira-ome tool extends the application of cfDNA data to detect pathogenic viruses in SOT patients. And majority of predictions were confirmed by qPCR.

« Back to 2020 American Transplant Congress