Genomic Epidemiology of the Los Angeles COVID-19 Outbreak (preprint)

Background The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused a global disruption to human health and activity. Being able to trace the early outbreak of SARS-CoV-2 within a locality will inform public health measures and provide insights to contain or prevent the viral transmission to save lives. Investigation of the transmission history requires efficient sequencing methods and analytic strategy, which can be generally useful in the study of viral outbreaks.  Methods Los Angeles (LA) County has sustained a large outbreak of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). To learn about the transmission history, we carried out surveillance viral genome sequencing to determine 142 viral genomes from unique patients seeking care at UCLA Health System. 86 of these genomes are from samples collected before April 19, 2020. Results We found that the early outbreak in LA, as in other international air travel hubs, was seeded by multiple introductions of strains from Asia and Europe. We identified a US-specific strain, B.1.43, which has been found predominantly in California and Washington State. While samples from LA County carry the ancestral B.1.43 genome, viral genomes from neighboring counties in California and from counties in Washington State carry additional mutations, suggesting a potential origin of B.1.43 in Southern California. We quantified the transmission rate of SARS-CoV-2 over time, and found evidence that the public health measures put in place in LA County to control the virus were effective at preventing transmission, but may have been undermined by the many introductions of SARS-CoV-2 into the region. Conclusion Our work demonstrates that genome sequencing can be a powerful tool for investigating outbreaks and informing the public health response. Our results reinforce the critical need for the U.S. to have coordinated inter-state responses to the pandemic.

Genomic epidemiology of the Los Angeles COVID-19 outbreak (preprint)

Los Angeles (LA) County has sustained a large outbreak of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). To learn about the transmission history of SARS-CoV-2 in LA County, we sequenced 142 viral genomes from unique patients seeking care at UCLA Health System. 86 of these genomes are from samples collected before April 19, 2020. We found that the early outbreak in LA, as in other international air travel hubs, was seeded by multiple introductions of strains from Asia and Europe. We identified a US-specific strain, B.1.43, which has been found predominantly in California and Washington State. While samples from LA County carry the ancestral B.1.43 genome, viral genomes from neighbouring counties in California and from counties in Washington State carry additional mutations, suggesting a potential origin of B.1.43 in Southern California. We quantified the transmission rate of SARS-CoV-2 over time, and found evidence that the public health measures put in place in LA County to control the virus were effective at preventing transmission, but may have been undermined by the many introductions of SARS-CoV-2 into the region. Our work demonstrates that genome sequencing can be a powerful tool for investigating outbreaks and informing the public health response. Our results reinforce the critical need for the U.S. to have coordinated inter-state responses to the pandemic.

Rapid cost-effective viral genome sequencing by V-seq (preprint)

ABSTRACT Conventional methods for viral genome sequencing largely use metatranscriptomic approaches or, alternatively, enrich for viral genomes by amplicon sequencing with virus-specific PCR or hybridization-based capture. These existing methods are costly, require extensive sample handling time, and have limited throughput. Here, we describe V-seq, an inexpensive, fast, and scalable method that performs targeted viral genome sequencing by multiplexing virus-specific primers at the cDNA synthesis step. We designed densely tiled reverse transcription (RT) primers across the SARS-CoV-2 genome, with a subset of hexamers at the 3’ end to minimize mis-priming from the abundant human rRNA repeats and human RNA PolII transcriptome. We found that overlapping RT primers do not interfere, but rather act in concert to improve viral genome coverage in samples with low viral load. We provide a path to optimize V-seq with SARS-CoV-2 as an example. We anticipate that V-seq can be applied to investigate genome evolution and track outbreaks of RNA viruses in a cost-effective manner. More broadly, the multiplexed RT approach by V-seq can be generalized to other applications of targeted RNA sequencing.

Swab-Seq: A high-throughput platform for massively scaled up SARS-CoV-2 testing (preprint)

The rapid spread of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is due to the high rates of transmission by individuals who are asymptomatic at the time of transmission. Frequent, widespread testing of the asymptomatic population for SARS-CoV-2 is essential to suppress viral transmission and is a key element in safely reopening society. Despite increases in testing capacity, multiple challenges remain in deploying traditional reverse transcription and quantitative PCR (RT-qPCR) tests at the scale required for population screening of asymptomatic individuals. We have developed SwabSeq, a high-throughput testing platform for SARS-CoV-2 that uses next-generation sequencing as a readout. SwabSeq employs sample-specific molecular barcodes to enable thousands of samples to be combined and simultaneously analyzed for the presence or absence of SARS-CoV-2 in a single run. Importantly, SwabSeq incorporates an in vitro RNA standard that mimics the viral amplicon, but can be distinguished by sequencing. This standard allows for end-point rather than quantitative PCR, improves quantitation, reduces requirements for automation and sample-to-sample normalization, enables purification-free detection, and gives better ability to call true negatives. We show that SwabSeq can test nasal and oral specimens for SARS-CoV-2 with or without RNA extraction while maintaining analytical sensitivity better than or comparable to that of fluorescence-based RT-qPCR tests. SwabSeq is simple, sensitive, flexible, rapidly scalable, inexpensive enough to test widely and frequently, and can provide a turn around time of 12 to 24 hours.