Analytical Validation of the Karius Test


Analytical validation employed a panel of thirteen representative microbes to characterize performance across a spectrum of potential performance determinants, including GC-content, superkingdom, presence as a commensal on the skin or in the gut, extent of background contamination in reagents, genome length, and high genetic similarity to other organisms. In total, 358 samples containing sheared microbial DNA at various concentrations and combinations, 2,625 in silico generated data sets, and 167 samples from asymptomatic individuals were used to characterize performance. In addition, 25 samples from individuals with confirmed CMV infection were also compared to quantitative blood CMV PCR results.


Microbial cfDNA sequencing of plasma showed 100% sensitivity compared to quantitative CMV PCR testing, with strong correlation between the quantitative measurements of these methods (R2 = 0.88). The median analytical sensitivity was 41 microbe-specific cfDNA fragments/microliter of plasma at typical sequencing depth, with precision ranging from 17.9%-22.2% relative error. Specificity determined using 50 well-characterized plasma aliquots was 98% on a per sample basis, and greater than 99.99% on a per analyte basis in silico.