posted on 2021-11-15, 16:08authored byAnthony Maus, Santosh Renuse, Jennifer Kemp, Anil K. Madugundu, Patrick M. Vanderboom, Joseph Blommel, Calvin Jerde, Surendra Dasari, Benjamin R. Kipp, Ravinder J. Singh, Stefan K. Grebe, Akhilesh Pandey
COVID-19 vaccines
are becoming more widely available, but accurate
and rapid testing remains a crucial tool for slowing the spread of
the severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) virus.
Although the quantitative reverse transcription-polymerase chain reaction
(qRT-PCR) remains the most prevalent testing methodology, numerous
tests have been developed that are predicated on detection of the
SARS-CoV-2 nucleocapsid protein, including liquid chromatography-tandem
mass spectrometry (LC-MS/MS) and immunoassay-based approaches. The
continuing emergence of SARS-CoV-2 variants has complicated these
approaches, as both qRT-PCR and antigen detection methods can be prone
to missing viral variants. In this study, we describe several COVID-19
cases where we were unable to detect the expected peptide targets
from clinical nasopharyngeal swabs. Whole genome sequencing revealed
that single nucleotide polymorphisms in the gene encoding the viral
nucleocapsid protein led to sequence variants that were not monitored
in the targeted assay. Minor modifications to the LC-MS/MS method
ensured detection of the variants of the target peptide. Additional
nucleocapsid variants could be detected by performing the bottom-up
proteomic analysis of whole viral genome-sequenced samples. This study
demonstrates the importance of considering variants of SARS-CoV-2
in the assay design and highlights the flexibility of mass spectrometry-based
approaches to detect variants as they evolve.