MALDI-TOF MS analysis for detection of bovine coronavirus with tryptic peptides from viral proteins

Bovine coronavirus (BCoV）, a significant cattle pathogen causing enteric and respiratory diseases, is primarily detected using reverse transcription-polymerase chain reaction. Our objective was to develop a novel detection method for BCoV by matrix-assisted laser desorption/ionization‒time-of-flight mass spectrometry (MALDI-TOF MS）. Peptide mass fingerprint analysis revealed that nucleocapsid (N）, membrane (M）, and hemagglutinin-esterase (HE) were three main BCoV proteins. Their tryptic peptides were used as target molecules for BCoV detection. When the tryptic digest of 10^7.0 viral copies was analyzed by MALDI-TOF MS, five peptides with relatively strong peaks were detected. The detection limit was between 10^5.0 and 10^6.0 copies per test for BCoV alone. To detect BCoV in the swab eluate, ultrafiltration purification achieved a detection limit between 10^6.0 and 10^7.0 copies per test, sufficient to detect BCoV-infected calves. Our findings offer valuable insights for BCoV detection by MALDI-TOF MS.