Rapid Genomic Characterization of SARS-CoV-2 by Direct Amplicon-Based Sequencing Through Comparison of MinION and Illumina iSeq100TM System

Abstract

Global human health is increasingly challenged by emerging viral threats, especially those observed over the last 20 years with coronavirus-related human diseases, such as the Severe Acute Respiratory Syndrome (SARS) and the Middle East Respiratory Syndrome (MERS). Recently, in late December 2019, a novel Betacoronavirus, SARS-CoV-2, originating from the Chinese city of Wuhan, emerged and was then identified as the causative agent of a new severe form of pneumonia, COVID-19. Real-time genome sequencing in such viral outbreaks is a key issue to confirm identification and characterization of the involved pathogen and to help establish public health measures. Here, we implemented an amplicon-based sequencing approach combined with easily deployable next-generation sequencers, the small and hand-held MinION sequencer and the latest most compact Illumina sequencer, the iSeq100 TM system. Our results highlighted the great potential of the amplicon-based approach to obtain consensus genomes of SARS-CoV-2 from clinical samples in just a few hours. Both these mobile next-generation sequencers are proven to be efficient to obtain viral sequences and easy to implement, with a minimal laboratory environment requirement, providing useful opportunities in the field and in remote areas.

Current and innovative methods for the diagnosis of COVID‑19 infection (Review)

“…As already mentioned, amplicon-based and metagenomic approaches based on the sequence-independent single primer amplification (SISPA) method are able to detect mutations occurring in the sequence of SARS-CoV-2 and potentially associated with vaccine inefficacy or resistance to antiviral therapies ( 110 , 111 ). Examples of this type of sequencing technique are the MinION and IDbyDNA platforms produced by Oxford Nanopore Technologies and Illumina, respectively ( 112 ). As regards the IDbyDNA platform, it ensures the collection of >13 million reads of which >8 million are unique reads with an average length of ~75 bp and an in-depth coverage.…”

Rapid genomic surveillance of SARS-CoV-2 in a dense urban community using environmental (sewage) samples

“…In the past twenty years, several diseases caused by coronavirus have posed significant global health challenges-including Severe Acute Respiratory Syndrome (SARS, 2002), Middle East Respiratory Syndrome (MERS, 2012), as well as the current pandemic of COVID-19 [1,2]. COVID-19 is caused by a single-stranded, positive-sense RNA virus (SARS-CoV-2) which belongs to the Coronaviridae family [2].…”

“…Environmental surveillance (ES) offers a complementary and more feasible approach to clinical disease surveillance. This approach provides unbiased information on a population level and detects viral shedding by symptomatic and asymptomatic patients [1,16], thus providing a snapshot of the outbreak over an entire sewage catchment area and an early indication of clinical cases in the area [17]. Because not all symptomatic patients get tested due to reluctance or lack of access to tests, clinical samples are not a sensitive indicator of the occurrence of cases in a given area [18,19].…”

“…Portable and reliable sequencing tools such as MinION can play a crucial role in conducting genomic surveillance in a developing country like Nepal where access to high throughput DNA sequencing machines is limited. The pocket-sized and field-deployable, the next-generation sequencer has enabled real-time outbreak surveillance of several recent outbreaks of Zika, Ebola, and Lassa viruses [1]. Obtaining whole or partial genome sequences of the virus during outbreak is critical in understanding viral evolution and disease epidemiology, and can help design accurate diagnostic tests for rapidly evolving RNA viruses such as SARS-CoV-2 [25].…”

iSeq100 for metagenomic pathogen screening in ticks

“…NGS can also provide quantitative data on the concentration of organisms in a sample by counting the sequence reads [21]. iSeq100, a recently released instrument, is capable of processing a targeted NGS library with accuracy comparable to that of MiSeq, the most widely used instrument in microbiome studies [22,23]. iSeq100 also features shortened sequencing work ow preparation time and a total sequencing run time of under 24 hours, as it allows the sequencing of a lower read count than the MiSeq [22].…”