Targeting the Conserved Stem Loop 2 Motif in the SARS-CoV-2 Genome.

RNA structural elements occur in numerous single-stranded positive-sense RNA viruses. The stem-loop 2 motif (s2m) is one such element with an unusually high degree of sequence conservation, being found in the 3' untranslated region (UTR) in the genomes of many astroviruses, some picornaviruses and noroviruses, and a variety of coronaviruses, including severe acute respiratory syndrome coronavirus (SARS-CoV) and SARS-CoV-2. The evolutionary conservation and its occurrence in all viral subgenomic transcripts imply a key role for s2m in the viral infection cycle. Our findings indicate that the element, while stably folded, can nonetheless be invaded and remodeled spontaneously by antisense oligonucleotides (ASOs) that initiate pairing in exposed loops and trigger efficient sequence-specific RNA cleavage in reporter assays. ASOs also act to inhibit replication in an astrovirus replicon model system in a sequence-specific, dose-dependent manner and inhibit SARS-CoV-2 replication in cell culture. Our results thus permit us to suggest that the s2m element is readily targeted by ASOs, which show promise as antiviral agents. IMPORTANCE The highly conserved stem-loop 2 motif (s2m) is found in the genomes of many RNA viruses, including SARS-CoV-2. Our findings indicate that the s2m element can be targeted by antisense oligonucleotides. The antiviral potential of this element represents a promising start for further research into targeting conserved elements in RNA viruses.

Rapid and sensitive detection of SARS-CoV-2 antibodies by biolayer interferometry.

Serological testing to evaluate antigen-specific antibodies in plasma is generally performed by rapid lateral flow test strips that lack quantitative results or by high complexity immunoassays that are time- and labor-intensive but provide semi-quantitative results. Here, we describe a novel application of biolayer interferometry for the rapid detection of antigen-specific antibody levels in plasma samples, and demonstrate its utility for quantification of SARS-CoV-2 antibodies. Our biolayer interferometry immunosorbent assay (BLI-ISA) utilizes single-use biosensors in an automated "dip-and-read" format, providing real-time optical measurements of antigen loading, plasma antibody binding, and antibody isotype detection. Complete semi-quantitative results are obtained in less than 20 min. BLI-ISA meets or exceeds the performance of high complexity methods such as Enzyme-Linked Immunosorbent Assay (ELISA) and Chemiluminescent Immunoassay. Importantly, our method can be immediately implemented on existing BLI platforms for urgent COVID-19 studies, such as serosurveillance and the evaluation of vaccine candidates. In a broader sense, BLI-ISA can be developed as a novel diagnostic platform to evaluate antibodies and other biomolecules in clinical specimens.