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

To test whether the performance of the highly complex 356-virus V ALL probe set matches that of focused ssRNA probe sets, we first compared it to the 23-virus V WAFR probe set. We evaluated the 6 viral species we tested from the patient and environmental samples that were present in both the V ALL and V WAFR probe sets, and we found that performance was concordant between them: V WAFR provides almost the same number of unique viral reads as V ALL (1.01× as many; Q 1 = 0.93, Q 3 = 1.34) (Supplementary Table 3 ). The percentage 6 . CC-BY-NC-ND 4.0 International license is made available under a The copyright holder for this preprint (which was not peer-reviewed) is the author/funder. It . https://doi.org/10.1101/279570 doi: bioRxiv preprint x Fraction of positions with At each position across a genome, the post-capture read depth is divided by the pre-capture depth, and the plotted curve is the empirical cumulative distribution of the log of these fold-change values. A curve that rises fully to the right of the black vertical line illustrates enrichment throughout the entirety of a genome; the more vertical a curve, the more uniform the enrichment. Read depth across viral genomes DENV-SM3 (purple) and DENV-SM5 (green) are shown in more detail in (b). (b) Read depth throughout a genome of DENV in two samples. DENV-SM3 (left) has few informative reads before capture and does not produce a genome assembly, but does following capture. DENV-SM5 (right) does yield a genome assembly before capture, and depth increases following capture. (c) Percent of the viral genomes unambiguously assembled in the 30 samples, which had 8 known viral infections across them. Shown before capture (orange), after capture with V WAFR (light blue), and after capture with V ALL (dark blue). Red bars below samples indicate ones in which we could not assemble any contig before capture but, following capture, were able to assemble at least a partial genome (> 50%). (d) Left: Number of reads detected for each species across the 30 samples with known viral infections, before and after capture with V ALL . Reads in each sample were downsampled to 200,000 reads. Each point represents one species detected in one sample. For each sample, the virus previously detected in the sample by another assay is colored. Homo sapiens matches in samples from humans are shown in black. Right: Abundance of each detected species before capture and fold-change upon capture with V ALL for these samples. Abundance was calculated by dividing pre-capture read counts for each species by counts in pooled water controls. Coloring of human and viral species are as in the left panel.