673. Unbiased Microbial Cell-free DNA Next-Generation Sequencing Resolves HHV6 Variant Specificity and Demonstrates a Predominance of HHV6B over HHV6A in Real-World Clinical Experience
Authors: Alexander Jose , Bercovici Sivan , Degner Nicholas R , Castillo-Galvan Ricardo , Arun Aparna , de Vries Christiaan R , Macintyre Ann , Perkins Bradley , Ahmed Asim A , Smollin Matthew 
Published: December 2021

Abstract

Background
Human herpesvirus 6 (HHV6) has been classified in two distinct variants, HHV6A and HHV6B. Although distinct epidemiology, disease association, biological and immunological properties, their genomes are 90% homologous. Less is known about HHV6A which is typically asymptomatic but can cause severe infection in patients with neurological disorders or HIV. HHV6B infection occurs during childhood, but can reactivate after solid organ and stem cell transplantation. Antibody assays may indicate previous, recent or current infection. Quantitative PCR cannot differentiate active from latent infections and some available qualitative PCR assays are unable to differentiate the two variants. A single open-ended test through plasma-based microbial cell-free DNA (mcfDNA) metagenomic next-generation sequencing (NGS) may overcome these limitations.

Methods
Karius TestTM (KT) developed and validated in Karius’s CLIA certified/CAP accredited lab, detects mcfDNA from plasma. mcfDNA is extracted, NGS performed, human sequences removed and remaining sequences aligned to a curated pathogen database of &gt; 1500 organisms. Organisms present above a statistical threshold are reported and quantified. For &gt; 85% of tests the time to result reporting is the next day from sample receipt. KT results were reviewed from November-2018 to May-2021 for detections of HHV6A and HHV6B. The comparative incidence of HHV6A and HHVB detections, their age distributions and their quantitative viral concentration in molecules/µL (MPM) were assessed.

Results
KT detected 322 cases of HHV6; 10% (n=32) were HHV6A and 90% (n=290) HHV6B (Table 1). HHV6B had a higher relative abundance in children (with a distribution into adolescence) compared to adults (Figure 1). The average HHV6A MPM was 860 (range 27 - 10,472); the average HHV6B MPM was 3,361 (range 21 - 131,518).
Table 1. Summary of HHV-6 Detections

Figure 1. Distribution of HHV6A and HHV6B by Age Group

Conclusion
The distribution of the HHV6 variants detected through KT shows an overwhelming 9:1 predominance of HHV6B to HHV6A. The application of mcfDNA metagenomic sequencing for open-ended detection, variant determination and quantification of HHV6 provides more specific resolution than serological and PCR methods. KT may lend important insights into the association of specific HHV6 variants with clinical syndromes affecting vulnerable populations.

Disclosures
Jose Alexander, MD, D(ABMM), FCCM, CIC, SM, MB(ASCP), BCMAS, Karius (Employee) Sivan Bercovici, PhD, Karius (Employee) Nicholas R. Degner, MD, MPH, MS, Karius Inc. (Employee, Shareholder) Ricardo Castillo-Galvan, MD MPH, Karius Inc. (Consultant) Aparna Arun, MD, Karius (Employee) Ann Macintyre, DO, Karius, Inc. (Employee) Bradley Perkins, MD, Karius, Inc. (Employee) Asim A. Ahmed, MD, Karius, Inc. (Employee) Matthew Smollin, PharmD, Karius, Inc. (Employee)