Extended Data Fig. 4: Orthogonal testing for Case S31: Klebsiella pneumoniae infection of pleural fluid.
From: Rapid pathogen detection by metagenomic next-generation sequencing of infected body fluids
a, Genomic coverage of K. pneumoniae from Illumina mNGS. Sequencing spanned 36,490 base pairs, or 0.65% of the K. pneumoniae genome. b, Orthogonal confirmation of K. pneumoniae by dPCR of the sequencing library. Nine negative controls from other cases were run in parallel. Out of 10 sequencing libraries, only Case S31 had any positive droplets (n = 43 of 12022 total droplets as circled). c, Orthogonal confirmation of K. pneumoniae by dPCR of the DNA extract. Three positive droplets were detected, indicating a low positive result. d, Orthogonal confirmation of K. pneumoniae by dPCR of contralateral pleural fluid (sample C31). 29 and 24 positive droplets were detected out of 2 replicates. Digital PCR targeting Streptococcus mitis on both pleural fluids did not yield any positive droplets. The positive controls for these experiments were from sheared DNA from Klebsiella pneumoniae and Streptococcus mitis respectively, whereas the negative control was water. e, Sanger sequencing of the K. pneumoniae amplicon from dPCR. Shown are sequencing traces confirming the presence of K. pneumoniae.
