Metaproteomic Analysis of Nasopharyngeal Swab Samples to Identify Microbial Peptides in COVID-19 Patients

Duringthe COVID-19 pandemic, impaired immunity and medical interventionsresulted in cases of secondary infections. The clinical difficultiesand dangers associated with secondary infections in patients necessitatethe exploration of their microbiome. Metaproteomics is a powerfulapproach to study the taxonomic composition and functional statusof the microbiome under study. In this study, the mass spectrometry(MS)-based data of nasopharyngeal swab samples from COVID-19 patientswas used to investigate the metaproteome. We have established a robustbioinformatics workflow within the Galaxy platform, which includes(a) generation of a tailored database of the common respiratory tractpathogens, (b) database search using multiple search algorithms, and(c) verification of the detected microbial peptides. The microbialpeptides detected in this study, belong to several opportunistic pathogenssuch as Streptococcus pneumoniae, Klebsiella pneumoniae, Rhizopus microsporus, and Syncephalastrum racemosum. Microbialproteins with a role in stress response, gene expression, and DNArepair were found to be upregulated in severe patients compared tonegative patients. Using parallel reaction monitoring (PRM), we confirmedsome of the microbial peptides in fresh clinical samples. MS-basedclinical metaproteomics can serve as a powerful tool for detectionand characterization of potential pathogens, which can significantlyimpact the diagnosis and treatment of patients.