2021), and restriction fragment length polymorphism (RFLP) ( Niranji & Al-Jaf 2021). 2022), an amplification refractory mutation system (ARMS) ( Islam et al. These technologies rely on: Probe-based real-time reverse transcriptase PCR (rRT-PCR) ( Chan et al. Low-cost rapid technologies have been put forward to investigate the A23403G (D614G) mutation. Moreover, the number of cases studied by WGS may not necessarily be representative of the number of cases detected by quantitative Polymerase Chain Reaction (qPCR) at these locations.Īs a result, several methodologies have been developed as affordable alternatives that provide rapid detection of not only relevant SNPs in the S protein, but also SNPs in variants of concern ( Hashemi et al. However, this approach is too expensive to be used in massive testing campaigns, especially in countries with limited resources. Studies related to this mutation have used, in the majority, whole genome sequences (WGS). The A23403G (D614G) mutation was first described in early 2020, and rapidly detected in various regions around the globe ( Isabel et al. The aspartate (D) to glycine (G) substitution related to the non-synonymous A23403G SNP (single nucleotide polymorphism) has been associated with the rapid spread of the virus ( Yuan et al. The S protein recognizes and binds to the angiotensin-converting enzyme 2 (ACE2) receptor to gain entry to hosts cells.Įvidently, mutations in the gene encoding the S protein alter the affinity to the ACE2 receptor enhances infectivity ( Watanabe et al. Structural proteins, such as the Spike (S) protein, nucleocapsid (N) protein, membrane (M) protein, and envelope (E) protein are encoded by short sub-genomic RNAs (sgRNAs) ( Kim et al. Two Open Reading Frames (ORFs) (1a and 1b), that cover most of its genome, encode non-structural proteins needed for viral replication as well as an RNA-dependent RNA polymerase (RdRp). The genome of this virus is approximately 30-kb long ( Bar-On et al. SARS-CoV-2 is the agent responsible for the COVID-19 pandemic that has caused almost 4 million deaths worldwide ( World Health Organization (WHO) 2021). This approach will be crucial for producing relevant information for public health management, especially during the ongoing pandemic. The application of this method allowed the production of epidemiological data regarding the A23403G (D614G) mutation in Quito, where no previous reports were available. This technique proved to be reliable, reproducible, and might be expandable to study other mutations without major protocol amendments. From these cases, almost all samples were associated with the G23403 (G614) variant (>98%). In March, a total of 264 out of 1319 samples yielded positive results (20%), while 777 out of 5032 (15%) did so in October. This strategy was tested in SARS-CoV-2 positive samples collected in Quito during March and October of 2020. This report has evaluated an easy cost-effective approach, based on sanger sequencing, for detection of the A23403G (D614G) mutation.
Consequently, developing rapid and accessible protocols will be fundamental for producing epidemiological data linked to this SNP, especially in countries with limited resources. However, this technology is not suited for massive testing as it requires expensive reagents, equipment, and infrastructure. The most utilized and robust approach is the study of whole genome sequences, generally available at public databases. The Single Nucleotide Polymorphism (SNP) A23403G associated with the D to G change in position 614 of the SARS-CoV-2 spike protein has recently become dominant.