Coronaviruses infect both animals and humans and lead to various illnesses, including neurological, enteric, and respiratory diseases. There are four distinct genera of coronaviruses: Alphacoronavirus, Betacoronavirus, Gammacoronavirus, and Deltacoronavirus⁸. SARS-CoV, MERS-CoV, and SARS-CoV-2 are three highly pathogenic coronaviruses, capable of infecting humans, that emerged in 2002, 2012, and 2019, respectively^3-4. In this study, the genetic compositions and sequence similarities of nine Indonesian SARS-CoV-2 spike glycoprotein genes were determined using sequences obtained from GenBank and the GISAID EpiCoV database (Table 2-4). Many researchers worldwide have previously reported mutations in the SARS-CoV-2 genome^10,20-21, however, there has not previously been a study of the SARS-CoV-2 genome from Indonesian isolates.

Table (2):
Nucleotide mutation sites in the SARS-CoV-2 spike glycoprotein.

Table (3):
Amino acid mutation sites in the SARS-CoV-2 spike glycoprotein.

Table (4):
Sequence similarity of SARS-CoV-2 spike glycoprotein genes, determined using the LALIGN web server.

The coronavirus spike glycoprotein mediates membrane fusion and viral entry into host cells and is therefore the primary target for many neutralizing antibodies (Fig. 1). The spike glycoprotein has two domains, S1 and S2, where S1 is responsible for binding the virion to ACE2 on the host cell membrane²¹. Several antiviral drugs and vaccines have been developed which target the spike glycoprotein. Du et al. (2009)²² demonstrated the effectiveness of several of these antiviral therapies, including small interfering RNAs, protease inhibitors, ACE2 blockers, fusion blockers, spike glycoprotein inhibitors, neutralizing antibodies, and spike glycoprotein cleavage inhibitors in in vitro studies. In addition, a number of techniques have been used to generate vaccines using all or part of the spike glycoprotein as an antigen. These include the use of recombinant receptor binding domain protein, viral vectors, full-length S protein, recombinant spike glycoprotein, and spike protein DNA-expressing vectors. Thus, it is very important to investigate the genetic composition and sequence similarity of this protein.

Interestingly, despite reporting several mutation sites in this study, we demonstrate that there is no significant change in the genetic composition of SARS-CoV-2 spike glycoprotein genes. Nucleotide variants in SARS-CoV-2 spike glycoprotein genes described in this study include JKT-EIJK2444 (224C>T), EJ-ITD3590NT (347C>G; 1841A>G; 2031G>T), JKT-EIJK01 (414T>C; 1864G>T), and JKT-EIJK04 (1715C>T; 2464C>T) (Table 2). In addition, we also analyzed amino acid changes in the SARS-CoV-2 spike glycoprotein including JKT-EIJK2444 (T76I), EJ-ITD3590NT (S116C; D614G; Q677H), JKT-EIJK01 (V622F), and JKT-EIJK04 (T572I; L822F) (Table 3). Finally, the interval score of similarity between each of the SARS-CoV-2 spike glycoproteins was between 99.9% and 100% (Table 4).

Previous studies investigating gene variability in SARS-CoV-2 samples include the work of Sekizuka et al. (2020), who performed whole-genome sequencing of SARS-CoV-2, directly from PCR-positive clinical specimens¹⁸. This was conducted in order to generate a haplotype network analysis of the Diamond Princess cruise ship outbreak, using the Wuhan-Hu-1 sequence as a reference. Additionally, Castillo et al. (2020) reported a phylogenetic analysis of the first four SARS-CoV-2 cases in Chile, analyzing nucleotide variants, amino acid changes, and sequence similarity²³. While our study complements these previous works, several shortcomings remain, including the relatively small number of isolates studied, the methodology used for whole-genome sequencing, and the quality coverage of SARS-CoV-2 genomes isolated from Indonesia.

As new information on SARS-CoV-2 is published daily, new concepts and frameworks must constantly be adopted. Currently, the GISAID EpiCoV database and Tang et al. (2020) have established three subtypes of SARS-CoV-2 based on nucleotide variants that produce amino acid changes: S, G, and V²⁴. The mutation rate of viruses is considerably higher than most other biological entities, including prokaryotes and eukaryotes. This is especially true of RNA-based viruses such as SARS-CoV-2, Ebola and dengue, due to hydroxyl groups in RNA that act as catalytic sites for mutation. This advanced mutation rate, leads to enhanced virulence and a higher capacity for adaptive evolution^25-26. While Tang et al. (2020) have suggested that SARS-CoV-2 exhibits the characteristic high mutation rate of an RNA virus²⁴, in fact, the mutation rate of SARS-CoV-2 and other coronaviruses might be slightly lower than other RNA viruses because of its genome-encoded exonuclease. Regardless, its high mutation rate increases the potential of this zoonotic viral pathogen to adapt to efficient transmission from human to human and potentially allows it to become more virulent.

The genomic characteristics of SARS-CoV-2 are significantly different than either SARS-CoV or MERS-CoV²⁷. A previous study reported that the homology of SARS-CoV-2 with the bat coronavirus isolate, RaTG13, was 96%²⁸. Interestingly, another study reported that the homology of SARS-CoV-2 with a pangolin coronavirus was 99%¹. From these results, it could be suggested that pangolins act as an intermediate host between bats and humans.

This study of genomic variants of SARS-CoV-2 isolated from Indonesia is crucial for future investigations into the pathogenesis, prevention, and treatment of SARS-CoV-2. Development of this genomic data is vital work that will facilitate vaccine design, epidemiological investigations, viral detection, functional analysis, and evaluation of treatment options²⁷.

Outbreaks of SARS-CoV-2 have led to a state of medical and economic emergency worldwide. Therefore, understanding the characteristics of the SARS-CoV-2 genome and developing systems to monitor SARS-CoV-2 during the pandemic are critical steps for controlling this disease. The identification of genotypes connected to specific geographic and temporal infectious clusters suggests that genomic data can be used to track and monitor the transmission of SARS-CoV-2. Therefore, the rapid discovery of genetic variants of SARS-CoV-2 is necessary for a streamlined response to the COVID-19 outbreak. Similarly, identifying specific SARS-CoV-2 variants and connecting them using a molecular epidemiology approach would allow researchers to determine the origin of a specific variant and monitor its transmission. This could be an important tool in controlling the outbreak²¹.