Comparison of Sequencing-by-Synthesis and Avidity Base Chemistry Next-Generation Sequencing Platforms in Identifying Somatic Variants of Hematological Malignancies.

Abstract

Background: This study evaluates the performance of the Illumina NextSeq™ 550 and the Element Biosciences AVITI™ next-generation sequencing (NGS) system, in detecting single nucleotide variants (SNVs) and gene fusions.

Methods: A set of 66 NGS libraries, consisting of 33 DNA, 24 cDNA, and triplicates of 3 control libraries, were prepared from bone marrow samples targeting 63 genes and related fusions, and initially sequenced using the NextSeq 550 in the Cleveland Clinic's molecular diagnostic laboratory. The same libraries were subsequently sequenced on the AVITI. The resulting data were analyzed using a combination of Cleveland Clinic developed pipelines and ArcherDx virtual machine software.

Results: The study found that all 105 SNVs and 39 gene fusions identified by the NextSeq 550 were also detected in the AVITI, demonstrating a high degree of concordance between the platforms. The analyses revealed R2 values of 0.86 for read depth and 0.96 for VAF of the 105 DNA variants, and 0.95 for read depth and 0.97 for fusion percentage of the 39 fusion variants. In the reproducibility studies, the VAF and fusion percentage of all variants were within 2 standard deviations of the mean when the same positive controls were sequenced 3 times on the AVITI.

Conclusions: These results indicate that the NextSeq 550 and the AVITI provide comparable performance in terms of accuracy and sensitivity for variant detection. Notably, the AVITI chemistry requires substantially lower PhiX input than the NextSeq 550 needs for this application. This results in substantial cost and efficiency benefits.