Abstract
Diagnosis of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) infection has primarily been achieved using reverse transcriptase polymerase chain reaction (RT-PCR) for acute infection, and serology for prior infection. Assay with RT-PCR provides data on presence or absence of viral RNA, with no information on virus replication competence, infectivity, or virus characterisation. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) is typically not used in clinical virology, despite its potential to provide supplemental data about the presence of viral proteins and thus the potential for replication-competent, transmissible virus. Using the SARS-CoV-2 as a model virus, we developed a fast ‘bottom-up’ proteomics workflow for discovery of target virus peptides using ‘serum-free’ culture conditions, providing high coverage of viral proteins without the need for protein or peptide fractionation techniques. This workflow was then applied to Coronaviruses OC43 and 229E, Influenza A/H1N1 and H3N2, Influenza B, and Respiratory Syncytial Viruses A and B. Finally, we created an LC-MS/MS method for targeted detection of the eight-virus panel in clinical specimens, successfully detecting peptides from the SARS-CoV-2 ORF9B and nucleoprotein in RT-PCR positive samples. The method provides specific detection of respiratory viruses from clinical samples containing moderate viral loads and is an important further step to the use of LC-MS/MS in diagnosis of viral infection.
【초록키워드】 severe acute respiratory syndrome coronavirus 2, SARS-CoV-2, viral infection, proteomics, mass spectrometry, serology, Influenza, Infection, peptide, Diagnosis, Viral proteins, severe acute respiratory syndrome Coronavirus, RT-PCR, virus, coronavirus 2, Replication, respiratory viruses, Severe acute respiratory syndrome, Protein, polymerase chain reaction, clinical samples, Coverage, Reverse transcriptase polymerase chain reaction, Culture, Liquid chromatography, Viral load, acute infection, peptides, virus replication, H1N1, Viral RNA, Orf9b, nucleoprotein, respiratory, information, influenza B, moderate, specimens, OC43, 229E, Tandem mass spectrometry, reverse transcriptase, Chain Reaction, presence or absence, not used, Viral protein, acute respiratory syndrome coronavirus, H3N2, characterisation, liquid, positive, transcriptase, SARS-CoV-2 ORF9b, Influenza A/H1N1, polymerase chain, clinical sample, respiratory virus, applied, provide, absence, conditions, the SARS-CoV-2, 【제목키워드】 SARS-CoV-2, Simultaneous, eight, human respiratory virus,