DeepSARS: simultaneous diagnostic detection and genomic surveillance of SARS-CoV-2.

BACKGROUND: The continued spread of SARS-CoV-2 and emergence of new variants with higher transmission rates and/or partial resistance to vaccines has further highlighted the need for large-scale testing and genomic surveillance. However, current diagnostic testing (e.g., PCR) and genomic surveillance methods (e.g., whole genome sequencing) are performed separately, thus limiting the detection and tracing of SARS-CoV-2 and emerging variants. RESULTS: Here, we developed DeepSARS, a high-throughput platform for simultaneous diagnostic detection and genomic surveillance of SARS-CoV-2 by the integration of molecular barcoding, targeted deep sequencing, and computational phylogenetics. DeepSARS enables highly sensitive viral detection, while also capturing genomic diversity and viral evolution. We show that DeepSARS can be rapidly adapted for identification of emerging variants, such as alpha, beta, gamma, and delta strains, and profile mutational changes at the population level. CONCLUSIONS: DeepSARS sets the foundation for quantitative diagnostics that capture viral evolution and diversity. DeepSARS uses molecular barcodes (BCs) and multiplexed targeted deep sequencing (NGS) to enable simultaneous diagnostic detection and genomic surveillance of SARS-CoV-2. Image was created using Biorender.com .

Clonally Expanded Virus-Specific CD8 T Cells Acquire Diverse Transcriptional Phenotypes During Acute, Chronic, and Latent Infections

CD8+ T cells play a crucial role in the control and resolution of viral infections and can adopt a wide range of phenotypes and effector functions depending on the inflammatory context and the duration and extent of antigen exposure. Similarly, viral infections can exert diverse selective pressures on populations of clonally related T cells. Technical limitations have nevertheless made it challenging to investigate the relationship between clonal selection and transcriptional phenotypes of virus-specific T cells. We therefore performed single-cell T cell receptor (TCR) repertoire and transcriptome sequencing of virus-specific CD8 T cells in murine models of acute, chronic and latent infection. We observed clear infection-specific populations corresponding to memory, effector, exhausted, and inflationary phenotypes. We further uncovered a mouse-specific and polyclonal T cell response, despite all T cells sharing specificity to a single viral epitope, which was accompanied by stereotypic TCR germline gene usage in all three infection types. Persistent antigen exposure during chronic and latent viral infections resulted in a higher proportion of clonally expanded T cells relative to acute infection. We furthermore observed a relationship between transcriptional heterogeneity and clonal expansion for all three infections, with highly expanded clones having distinct transcriptional phenotypes relative to less expanded clones. Together our work relates clonal selection to gene expression in the context of viral infection and further provides a dataset and accompanying software for the immunological community.

Platypus: an open-access software for integrating lymphocyte single-cell immune repertoires with transcriptomes.

High-throughput single-cell sequencing (scSeq) technologies are revolutionizing the ability to molecularly profile B and T lymphocytes by offering the opportunity to simultaneously obtain information on adaptive immune receptor repertoires (VDJ repertoires) and transcriptomes. An integrated quantification of immune repertoire parameters, such as germline gene usage, clonal expansion, somatic hypermutation and transcriptional states opens up new possibilities for the high-resolution analysis of lymphocytes and the inference of antigen-specificity. While multiple tools now exist to investigate gene expression profiles from scSeq of transcriptomes, there is a lack of software dedicated to single-cell immune repertoires. Here, we present Platypus, an open-source software platform providing a user-friendly interface to investigate B-cell receptor and T-cell receptor repertoires from scSeq experiments. Platypus provides a framework to automate and ease the analysis of single-cell immune repertoires while also incorporating transcriptional information involving unsupervised clustering, gene expression and gene ontology. To showcase the capabilities of Platypus, we use it to analyze and visualize single-cell immune repertoires and transcriptomes from B and T cells from convalescent COVID-19 patients, revealing unique insight into the repertoire features and transcriptional profiles of clonally expanded lymphocytes. Platypus will expedite progress by facilitating the analysis of single-cell immune repertoire and transcriptome sequencing.

Viral nucleoprotein antibodies activate TRIM21 and induce T cell immunity.

Nucleoprotein (N) is an immunodominant antigen in many enveloped virus infections. While the diagnostic value of anti-N antibodies is clear, their role in immunity is not. This is because while they are non-neutralising, they somehow clear infection by coronavirus, influenza and LCMV in vivo. Here, we show that anti-N immune protection is mediated by the cytosolic Fc receptor and E3 ubiquitin ligase TRIM21. Exploiting LCMV as a model system, we demonstrate that TRIM21 uses anti-N antibodies to target N for cytosolic degradation and generate cytotoxic T cells (CTLs) against N peptide. These CTLs rapidly eliminate N-peptide-displaying cells and drive efficient viral clearance. These results reveal a new mechanism of immune synergy between antibodies and T cells and highlights N as an important vaccine target.