Capturing diverse microbial sequence with comprehensive and scalable probe design

Hayden C. Metsky; Katherine J. Siddle; Adrianne Gladden-Young; James Qu; David K. Yang; Patrick Brehio; Andrew Goldfarb; Anne Piantadosi; Shirlee Wohl; Amber Carter; Aaron E. Lin; Kayla G. Barnes; Damien C. Tully; Björn Corleis; Scott Hennigan; Giselle Barbosa-Lima; Yasmine R. Vieira; Lauren M. Paul; Amanda L. Tan; Kimberly F. Garcia; Leda A. Parham; Ikponmwonsa Odia; Philomena Eromon; Onikepe A. Folarin; Augustine Goba; Etienne Simon-Lorière; Lisa Hensley; Angel Balmaseda; Eva Harris; Douglas Kwon; Todd M. Allen; Jonathan A. Runstadler; Sandra Smole; Fernando A. Bozza; Thiago M. L. Souza; Sharon Isern; Scott F. Michael; Ivette Lorenzana; Lee Gehrke; Irene Bosch; Gregory Ebel; Donald Grant; Christian Happi; Daniel J. Park; Andreas Gnirke; Pardis C. Sabeti; Christian B. Matranga

In addition to measuring enrichment on patient and environmental samples, we sought to evaluate the sensitivity of V ALL on samples with known quantities of viral and background material. To do so, we performed capture with V ALL on serial dilutions of Ebola virus (EBOV) -ranging from 10 6 copies down to single copy -in known background amounts of human RNA. At a depth of 200,000 reads, use of V ALL allowed us to reliably detect viral content (i.e., observe viral reads in two technical replicates) down to 100 copies in 30 ng of background and 1,000 copies in 300 ng (Fig. 3a, Supplementary Table 5) , each at least an order of magnitude fewer than without capture, and similarly lowered the input at which we could assemble genomes ( Supplementary Fig. 7a ). Although we chose a single sequencing depth so that we could compare pre-and post-capture results, higher sequencing depths provide more viral material and thus more sensitivity in detection ( Supplementary Fig. 7b , c).