Comparison of Metagenomic Sequencing and the NanoString nCounter Analysis System for the Characterization of Bacterial and Viral Communities in Vaginal Samples

Abstract

ABSTRACT DNA sequencing assays have been used to characterize the vaginal microbiome and to identify associations with clinical outcomes. The purpose of this study was to evaluate the utility of the NanoString nCounter platform, a more efficient assay compared to sequencing, for the characterization of vaginal microbial communities. A panel of NanoString nCounter probes was designed to detect common vaginal bacteria and viruses with relevance to reproductive health. A defined synthetic community of microbes and 43 clinical samples were interrogated with NanoString nCounter assays and compared to known compositions or metagenomic shotgun sequencing (MSS) results. The NanoString nCounter platform and MSS were able to distinguish closely related microbes. In clinical samples, the relative abundance of bacterial species was similar between the two assays. The assays sometimes disagreed when targets were present at low abundance. More viruses were detected by MSS than by nCounter. However, the nCounter assays are able to provide results in about 30 h with minimal hands-on time, whereas MSS requires at least 138 to 178 h with extensive hands-on time. The reagent cost for the two assays was similar, but the overall cost of the nCounter was lower due to the minimal hands-on time. MSS can be used to inform the design of a targeted multiplex panel for the assessment of vaginal microbial communities, thereby allowing for more cost-effective and rapid screening of patient samples for research studies. The sensitivity for low abundance microbes could be improved, possibly by adding additional target amplification cycles before nCounter assessment. This approach has potential as an assay with both research and clinical applications. IMPORTANCE Metagenomic shotgun sequencing can inform the design of a targeted multiplex panel by which the NanoString nCounter platform can assess vaginal microbial communities, thereby allowing for more cost-effective and rapid screening of patient samples.

Keywords

• NanoString nCounter
• microbiome
• sequencing
• vagina
• virome