Reduced subgenomic RNA expression is a molecular indicator of asymptomatic SARS-CoV-2 infection

It is estimated that up to 80% of infections caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) are asymptomatic and asymptomatic patients can still effectively transmit the virus and cause disease. While much of the effort has been placed on decoding single nucleotide variation in SARS-CoV-2 genomes, considerably less is known about their transcript variation and any correlation with clinical severity in human hosts, as defined here by the presence or absence of symptoms. To assess viral genomic signatures of disease severity, we conducted a systematic characterization of SARS-CoV-2 transcripts and genetic variants in 81 clinical specimens collected from symptomatic and asymptomatic individuals using multi-scale transcriptomic analyses including amplicon-seq, short-read metatranscriptome and long-read Iso-seq. Here we show a highly coordinated and consistent pattern of sgRNA expression from individuals with robust SARS-CoV-2 symptomatic infection and their expression is significantly repressed in the asymptomatic infections. We also observe widespread inter- and intra-patient variants in viral RNAs, known as quasispecies frequently found in many RNA viruses. We identify unique sets of deletions preferentially found primarily in symptomatic individuals, with many likely to confer changes in SARS-CoV-2 virulence and host responses. Moreover, these frequently occurring structural variants in SARS-CoV-2 genomes serve as a mechanism to further induce SARS-CoV-2 proteome complexity. Our results indicate that differential sgRNA expression and structural mutational burden are highly correlated with the clinical severity of SARS-CoV-2 infection. Longitudinally monitoring sgRNA expression and structural diversity could further guide treatment responses, testing strategies, and vaccine development. Wong and Ngan et al. characterize the expression and structural variation of SARS-CoV-2 subgenomic RNAs (sgRNAs) in diagnostic specimens from symptomatic and asymptomatic COVID-19 patients. In a series of genomic and transcriptomic analyses, the authors observe reduced sgRNA expression and a distinct set of structural deletions in asymptomatic infections compared with symptomatic infections. Infection with SARS-CoV-2 can lead to symptoms of different severity, with some individuals remaining entirely asymptomatic but still responsible for much of the viral transmission. Here, we sought to identify markers of the severity of symptoms of SARS-CoV-2 infection, as defined by the presence or absence of symptoms. We used a combination of methods to study SARS-CoV-2 genes and their readouts, known as transcripts, in clinical samples from people with symptomatic and asymptomatic infections. We demonstrate that transcripts responsible for making viral proteins are seen at lower levels in asymptomatic infections. We also identify structural changes in the viral genes and transcripts that potentially influence the host’s response to the virus. Our study defines potential markers of symptom severity that may ultimately guide risk mitigation and testing strategies.