Abstract
In resource-limited countries, early detection of novel pathogens is often challenging, due to financial and technical constraints. This study reports the efficacy of family-wide polymerase chain reaction (PCR) in screening, detecting, and identifying initial cases of the novel SARS-CoV-2 in Thailand. Respiratory secretions were collected from suspected individuals traveling from Wuhan, China to Thailand at the beginning of January 2020. Family-wide PCR assays yielded positive results for coronavirus in one traveler within 12 h on January 8, 2020. Nucleotide sequences (290 bp) showed 100% similarity to SARS-CoV-2. The whole genome sequence was further characterized by Next Generation Sequencing (NGS) for confirmation. Combining family-wide PCR, as a rapid screening tool, with NGS, for full genome characterization, could facilitate early detection and confirmation of a novel pathogen and enable early containment of a disease outbreak.
Highlights
The accurate and timely identification of novel pathogens presents an obvious challenge in resource-limited settings, requiring expensive laboratory infrastructure and equipment along with the associated consumable supplies and reagents, and highly-trained technical staff
Between January 4 and 8, 2020, specimens were collected from five individuals exhibiting fever or signs of respiratory symptoms [patients under investigation (PUIs)] identified by the Department of Disease Control (DDC), Ministry of Public Health Thailand, and sent to laboratories for diagnostic testing
All PUIs were transported to the Bamrasnaradura Infectious Disease Institute (BIDI) and quarantined
Summary
The accurate and timely identification of novel pathogens presents an obvious challenge in resource-limited settings, requiring expensive laboratory infrastructure and equipment along with the associated consumable supplies and reagents, and highly-trained technical staff. Family-wide polymerase chain reaction (PCR) assays have previously been used to detect various pathogens such as paramyxoviruses and MERS-CoV [7,8,9,10]. Our laboratory modified and applied an existing family-wide PCR assay [11,12,13,14] to detect the novel pathogen from individuals suspected of being part of the Wuhan outbreak. Whole genome sequencing (WGS) using NGS technology was performed on respiratory specimens from patients whose family-wide CoV PCR was positive for SARS-CoV-2. Once specific primer was available, real-time PCR (qPCR) assay targeting the receptor-binding domain of the spike gene using SYBR-Green One Step RTPCR was later performed according to the published protocol [17] on respiratory tract specimens from patients whose SARS-CoV-2 testing was previously positive by family-wide PCR
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