CRISPR-based surveillance for COVID-19 using genomically-comprehensive machine learning design

Abstract

The emergence and outbreak of SARS-CoV-2, the causative agent of COVID-19, has rapidly become a global concern and has highlighted the need for fast, sensitive, and specific tools to surveil circulating viruses. Here we provide assay designs and experimental resources, for use with CRISPR-based nucleic acid detection, that could be valuable for ongoing surveillance. We provide assay designs for detection of 67 viral species and subspecies, including: SARS-CoV-2, phylogenetically-related viruses, and viruses with similar clinical presentation. The designs are outputs of algorithms that we are developing for rapidly designing nucleic acid detection assays that are comprehensive across genomic diversity and predicted to be highly sensitive and specific. Of our design set, we experimentally screened 4 SARS-CoV-2 designs with a CRISPR-Cas13 detection system and then extensively tested the highest-performing SARS-CoV-2 assay. We demonstrate the sensitivity and speed of this assay using synthetic targets with fluorescent and lateral flow detection. Moreover, our provided protocol can be extended for testing the other 66 provided designs. Assay designs are available at https://adapt.sabetilab.org/ .

Overcoming the bottleneck to widespread testing: a rapid review of nucleic acid testing approaches for COVID-19 detection

“…The amplified product RNA activates Cas13a, which in turn cleaves a reporter RNA, liberating a fluorescent dye from a quencher. This method consistently detects synthetic SARS-CoV-2 viral RNA in the range between 10 and 100 copies per L of input, only requires a lateral flow dipstick for visual readout of the detection result, and can be completed in 40 minutes (Hou et al 2020) or 57 minutes Metsky et al 2020) after the RNA extraction step. SHERLOCK (termed "CRISPR-nCoV" in Hou et al 2020) also demonstrated its diagnostic potential by detecting SARS-CoV-2 RNA with 100% sensitivity in 52 patient samples (Hou et al 2020).…”

Shotgun Transcriptome and Isothermal Profiling of SARS-CoV-2 Infection Reveals Unique Host Responses, Viral Diversification, and Drug Interactions

“…As a result, the prevalence of SARS-CoV-2 in the population is mostly unknown, particularly among mild or asymptomatic cases. Though several novel, scalable biotechnological innovations for viral testing have recently emerged (e.g., CRISPR-Cas12a (Broughton et al, 2020) or CRISPR-Cas13 (Metsky et al, 2020) on paper-based detection systems, or loop-mediated isothermal amplification (LAMP) (Tanner et al, 2015, Schmid-Burgk et. al., 2020 these have not been validated against gold-standard clinical assays or nextgeneration sequencing (NGS).…”

SARS-CoV-2 detection with CRISPR diagnostics

“…During this ongoing COVID-19 outbreak, nucleic acid detection has played an important role in early diagnosis [2]. To date, 4 protocols based on CRISPR for detecting SARS-CoV-2 have been published [3][4][5][6]. Using lateral flow protocols, RNA samples harboring more than 1 × 10 4 -1 × 10 5 copies/mL (SHERLOCK) or 1 × 10 4 copies/mL (DETECTR) can be detected within 1 hour.…”