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

Wong, Chee-Hong; Ngan, Chew Yee; Goldfeder, Rachel L.; Idol, Jennifer; Kuhlberg, Chris; Maurya, Rahul; Kelly, Kevin; Omerza, Gregory; Renzette, Nicholas; Abreu, Francine de; Li, Lei; Browne, Frederick A.; Liu, Edison T.; Wei, Chia‐Lin

doi:10.1038/s43856-021-00034-y

Cited by 15 publications

(21 citation statements)

References 82 publications

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“…Instead, a significantly higher level of sgRNA was identified in patients with mild disease. This is an interesting finding as it has been reported that sgRNA transcripts are reduced in asymptomatic cases of COVID-19 ( Wong et al, 2021 ). However, this may be a result of the lack of intervention in mild cases compared to interventions which would have been received by hospitalised severe disease cases.…”

Section: Discussionsupporting

confidence: 71%

“…It is not well understood role sgRNA expression plays during infections. There have been reports that the detection of sgRNAs in clinical samples indicates an active viral infection, and expression levels correlate strongly to the severity of symptoms ( Wong et al, 2021 ). However, others report that sgRNA expression is not a reliable indicator of viral replication ( Alexandersen et al, 2020 ).…”

Section: Introductionmentioning

confidence: 99%

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SARS-CoV-2 Within-Host and in vitro Genomic Variability and Sub-Genomic RNA Levels Indicate Differences in Viral Expression Between Clinical Cohorts and in vitro Culture

et al. 2022

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BackgroundLow frequency intrahost single nucleotide variants (iSNVs) of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) have been increasingly recognised as predictive indicators of positive selection. Particularly as growing numbers of SARS-CoV-2 variants of interest (VOI) and concern (VOC) emerge. However, the dynamics of subgenomic RNA (sgRNA) expression and its impact on genomic diversity and infection outcome remain poorly understood. This study aims to investigate and quantify iSNVs and sgRNA expression in single and longitudinally sampled cohorts over the course of mild and severe SARS-CoV-2 infection, benchmarked against an in vitro infection model.MethodsTwo clinical cohorts of SARS-CoV-2 positive cases in New South Wales, Australia collected between March 2020 and August 2021 were sequenced. Longitudinal samples from cases hospitalised due to SARS-CoV-2 infection (severe) (n = 16) were analysed and compared with cases that presented with SARS-CoV-2 symptoms but were not hospitalised (mild) (n = 23). SARS-CoV-2 genomic diversity profiles were also examined from daily sampling of culture experiments for three SARS-CoV-2 variants (Lineage A, B.1.351, and B.1.617.2) cultured in VeroE6 C1008 cells (n = 33).ResultsIntrahost single nucleotide variants were detected in 83% (19/23) of the mild cohort cases and 100% (16/16) of the severe cohort cases. SNP profiles remained relatively fixed over time, with an average of 1.66 SNPs gained or lost, and an average of 4.2 and 5.9 low frequency variants per patient were detected in severe and mild infection, respectively. sgRNA was detected in 100% (25/25) of the mild genomes and 92% (24/26) of the severe genomes. Total sgRNA expressed across all genes in the mild cohort was significantly higher than that of the severe cohort. Significantly higher expression levels were detected in the spike and the nucleocapsid genes. There was significantly less sgRNA detected in the culture dilutions than the clinical cohorts.Discussion and ConclusionThe positions and frequencies of iSNVs in the severe and mild infection cohorts were dynamic overtime, highlighting the importance of continual monitoring, particularly during community outbreaks where multiple SARS-CoV-2 variants may co-circulate. sgRNA levels can vary across patients and the overall level of sgRNA reads compared to genomic RNA can be less than 1%. The relative contribution of sgRNA to the severity of illness warrants further investigation given the level of variation between genomes. Further monitoring of sgRNAs will improve the understanding of SARS-CoV-2 evolution and the effectiveness of therapeutic and public health containment measures during the pandemic.

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Section: Discussionsupporting

confidence: 71%

Section: Introductionmentioning

confidence: 99%

SARS-CoV-2 Within-Host and in vitro Genomic Variability and Sub-Genomic RNA Levels Indicate Differences in Viral Expression Between Clinical Cohorts and in vitro Culture

et al. 2022

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“…Functional tests for interference behavior have yet to be performed to provide evidence that these variants are DIPs. In the case of SARS-CoV-2, genomes that harbor large identical deletions were found in multiple patients who had only mild symptoms or were asymptomatic [127,128]. It remains to be seen what role, if any, DVGs may play in the severity of COVID-19.…”

Section: Dips In Vivomentioning

confidence: 99%

Virus-like Particles: Measures and Biological Functions

Bhat

Cao

Yin

2022

Viruses

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Virus-like particles resemble infectious virus particles in size, shape, and molecular composition; however, they fail to productively infect host cells. Historically, the presence of virus-like particles has been inferred from total particle counts by microscopy, and infectious particle counts or plaque-forming-units (PFUs) by plaque assay; the resulting ratio of particles-to-PFUs is often greater than one, easily 10 or 100, indicating that most particles are non-infectious. Despite their inability to hijack cells for their reproduction, virus-like particles and the defective genomes they carry can exhibit a broad range of behaviors: interference with normal virus growth during co-infections, cell killing, and activation or inhibition of innate immune signaling. In addition, some virus-like particles become productive as their multiplicities of infection increase, a sign of cooperation between particles. Here, we review established and emerging methods to count virus-like particles and characterize their biological functions. We take a critical look at evidence for defective interfering virus genomes in natural and clinical isolates, and we review their potential as antiviral therapeutics. In short, we highlight an urgent need to better understand how virus-like genomes and particles interact with intact functional viruses during co-infection of their hosts, and their impacts on the transmission, severity, and persistence of virus-associated diseases.

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“…No reports have been identified for the role of DVGs in IFN responses and viral persistence for SARS-CoV-2 infection so far. Interestingly, a COVID-19 cohort study (Wong, Ngan et al 2021) indicates that the abundance of TRS-independent deletions (>20nts) is significantly more in symptomatic patients than that in asymptomatic patients, suggesting a potential role of DVGs in modulating host responses and symptom development in COVID-19 patients.…”

Section: Introductionmentioning

confidence: 99%

Generation and functional analysis of defective viral genomes during SARS-CoV-2 infection

Zhou

Gilliam

et al. 2022

Preprint

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Defective viral genomes (DVGs) have been identified in many RNA viruses as a major factor influencing antiviral immune response and viral pathogenesis. However, the generation and function of DVGs in SARS-CoV-2 infection are less known. In this study, we elucidated DVG generation in SARS-CoV-2 and its relationship with host antiviral immune response. We observed DVGs ubiquitously from RNA-seq datasets of in vitro infections and autopsy lung tissues of COVID-19 patients. Four genomic hotspots were identified for DVG recombination and RNA secondary structures were suggested to mediate DVG formation. Functionally, bulk and single cell RNA-seq analysis indicated the IFN stimulation of SARS-CoV-2 DVGs. We further applied our criteria to the NGS dataset from a published cohort study and observed significantly higher DVG amount and frequency in symptomatic patients than that in asymptomatic patients. Finally, we observed unusually high DVG frequency in one immunosuppressive patient up to 140 days after admitted to hospital due to COVID-19, first-time suggesting an association between DVGs and persistent viral infections in SARS-CoV-2. Together, our findings strongly suggest a critical role of DVGs in modulating host IFN responses and symptom development, calling for further inquiry into the mechanisms of DVG generation and how DVGs modulate host responses and infection outcome during SARS-CoV-2 infection.

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

Cited by 15 publications

References 82 publications

SARS-CoV-2 Within-Host and in vitro Genomic Variability and Sub-Genomic RNA Levels Indicate Differences in Viral Expression Between Clinical Cohorts and in vitro Culture

SARS-CoV-2 Within-Host and in vitro Genomic Variability and Sub-Genomic RNA Levels Indicate Differences in Viral Expression Between Clinical Cohorts and in vitro Culture

Virus-like Particles: Measures and Biological Functions

Generation and functional analysis of defective viral genomes during SARS-CoV-2 infection

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