Multi-pathogen qPCR assay.

Previously published primers and probes for detecting H. influenzae, C. pneumoniae, P aeruginosa, S. pneumoniae, and S. aureus were used in this assay, with slight modifications to sequences to capture variation present in published reference sequences. In silico PCR was performed against all complete genome sequences available in NCBI’s RefSeq database; the number of reference sequences for each species indicated in the Notes column. In cases where a substantial proportion of the reference genomes contained a base mismatch at a primer or probe site, a degenerate base was added to the primer sequence to improve inclusivity across variants.
(XLSX)