|Ahead of print publication
Genome Sequencing of SARS-CoV-2: Outcomes, Predictions, and Their Effects on Therapeutic Options
Shashank M Patil, Ramith Ramu
Department of Biotechnology and Bioinformatics, School of Life Sciences, JSS Academy of Higher Education and Research, Mysuru, Karnataka, India
|Date of Submission||10-Jul-2020|
|Date of Decision||08-Sep-2020|
|Date of Acceptance||11-Oct-2020|
|Date of Web Publication||09-Jun-2021|
Department of Biotechnology and Bioinformatics, School of Life Sciences, JSS Academy of Higher Education and Research, Mysuru 570015, Karnataka
Source of Support: None, Conflict of Interest: None
|How to cite this URL:|
Patil SM, Ramu R. Genome Sequencing of SARS-CoV-2: Outcomes, Predictions, and Their Effects on Therapeutic Options. MAMC J Med Sci [Epub ahead of print] [cited 2021 Jun 13]. Available from: https://www.mamcjms.in/preprintarticle.asp?id=318089
The outbreak of novel coronavirus SARS-CoV-2 has resulted in the development of COVID-19, a currently emerging human infectious disease. Affecting several countries around the globe, COVID-19 has been declared a pandemic by the World Health Organization due to the soaring death rates and its unmatchable expansion around the world. In the absence of specific therapeutic options, the disease may further escalate to new heights. This is attributed to the possible upgradation of the viral mechanisms to enhance the chances of survivability against host innate immune responses and currently available therapeutic options. In light of these developments, several studies have performed genome sequencing of the virus to decipher the enigma of molecular pathogenesis. These studies have reported the occurrence of several mutations in the genome of SARS-CoV-2 concerning its pathogenic factors. This commentary is written on such reported mutations and their outcomes, and concerns their effect on pathogenesis and the possible effects on therapeutic options.
Researchers have hypothesized the fact that mutations occurring at the genomic level are responsible for the remarkable modification of the virus compared to its counterparts., Thus, deciphering the structure and the role of the genome in infection, replication, and transmission was essential to shed light on the discovery of possible and development of specific therapeutic agents. Although the initial efforts in genomic sequencing of the SARS-CoV-2 revealed notable facts about the structure and arrangement of the genomic constituents, phylogeny, infection, and transmission,, the majority of the studies showed mutations that occurred in course of the viral life cycle in the host cells, that is, replication.
The life cycle of SARS-CoV-2 begins with the adherence of the virus to the host cell. The virus uses spike (S) glycoprotein for binding with host cells that are believed to play a pivotal role in the invasion and further pathogenic mechanisms, which are yet to be depicted. In case of S protein, the occurrence of 12 variations in the N’-terminal domain, six variations in the receptor-binding domain (RBD), and >22 amino acid insertions have completely updated the binding efficiency of SARS-CoV-2 in comparison with SARS-CoV-1., These results depict the extension of the viral modification even from the first stage of the life cycle. This is supported by a study, which reported multiple amino acid residue insertions at S1/S2 cleavage of S protein in RmYN02, a novel bat-coronavirus. RmYN02 was identified from a collection of genome samples from 227 bats collected from Yunnan Province in China. It shares 93.3% complete nucleotide homology with SARS-CoV-2 and 97.2% homology with 1aboratory gene, although with lower homology with SARS-CoV-2 RBD (61.3%), with possible inability to bind with angiotensin-converting enzyme 2. However, this provided the evidence that insertion mechanisms could occur naturally in animal coronaviruses. Thus, it becomes evident that SARS-CoV-2 modifies itself to invade the host cell with effective binding, even resulting in the emergence of new strains. These changes can lead to drug resistance towards those drugs like chloroquine and hydroxychloroquine, which target the viral binding and adsorption.,
Subsequently, when the viral genome enters the host, it tends to replicate with the help of helicase that unfurls the viral RNA, and RNA-dependent RNA polymerase (RdRp) aids in the replication–transcription process. Studies have found mutations even in RdRp. The presence of eight novel recurring mutations in nucleotides has modified the activity of RdRp, and it has resulted in enhancing the pathogenicity predominantly in Europe, North America, and Asia. This shows the efficacy of the virus to modify itself after entering the host body, in turn, to infect a greater number of individuals further. Also, it could make things difficult for the currently employed drugs (RdRp inhibitors such as remdesivir and ribavirin), to track down and inhibit the RdRp and rest of the replication machinery.
In the case of successful replication, the genome undergoes frequent recombination like any other RNA viruses, resulting in the rapid evolution with respect to host specificity and drug sensitivity. For example, the presence of modified shorter poly (A) tails could alter 3‘ tail, and frequent fusions could make the genome stable and less susceptible to innate immune response as well as drugs such as ribavirin, favipiravir, ivermectin, and remdesivir that inhibit the viral transcription., In addition to structural proteins, genes encoding the nonstructural proteins known as open reading frames (ORFs) have also been reported to undergo mutation. These are considered to play an important role in the viral transcription alongside structural proteins. Changes detected in ORF1ab and ORF3a could result in possible alterations in the host immune response, allowing them to replicate even in CD4+T immune cells. ORF8 is reported to be the principle gene responsible for the effective transmission of the virus. Because of the modifications, it has been found showing high homology with a membrane protein, ORF7, thus making it one of the fastest evolving proteins. In support of this, a ∼430 bp region is found overlapping with the ORF8 gene, though being susceptible to nucleotide substitutions and deletions. Such details of the ability of the virus indicate the coding potential of SARS-CoV-2 with modified genome endeavors to leave a clear picture of the tremendous production of viral particles inside the host cell. As a result, the viral load increases inside the host body, ultimately affecting the immune system of the host.
Furthermore, comprehensive approaches in deciphering the protein synthesis showed that the rate of protein synthesis depends on the type of codon sequences present. In the process of protein synthesis, the presence of two successive slow codons (slow di-codons) could result in a slower translation rate. Calculating the composition of slow di-codons in the SARS-CoV-2 genome could predict the rate of protein synthesis. In support of this, both the genomes of SARS-CoV-1 and SARS-CoV-2 showed a higher rate of protein synthesis compared to the other coronaviruses, indicating the presence of a lower number of slow di-codons. The faster replication thus can produce the particulate matter, that is, aerosols, droplets with high viral loads that increase the survival rate of viruses in the environment. Also, the rate of infection also escalates infecting more persons even with the lower dose. These remain a proof of the fast pathogenicity of the virus. Infection with SARS-CoV-2 also increases the chance of co-infection, where the infected person may get infected with viral and bacterial diseases.,
In the case of transmission, enhanced transmissibility of the virus is attributed to some sequences known as “super spreaders” (SS). Researchers have identified four such genetic clusters that could be responsible for the viral outbreaks around the world. SS1 was found widely distributed in the United States and Asia, whereas SS4 is believed to have affected Europe. SS4 dominates 90% of the genomes sequenced. In addition, genomic analysis of infected persons from different countries also reveals the same. For example, genome sequencing of the virus isolated from a patient in Bangladesh showed high homology, revealing nine mutations shared with the virus isolated from Wuhan patient. Similarly, a genome with eight nucleotide mutations isolated from a cat in France was found to be identical to the Wuhan virus. This indicates the stability of the virus even after the transmission to the other host. Therefore, based on phylogenetic analyses, viral genomes are categorized into 10 clades, known as A1, A1a, A2, A2a, A3, A5, B, B1, B2, and B4. Out of these, A2a is believed to be the most pathogenic clade, responsible for the viral outbreaks in different countries. Being genetically polymorphic with a unique genetic constitution, these clads could ultimately interfere with vaccine development. This has been reflected in vaccine trials conducted in China and England, where volunteers from different countries, races, and ethnic groups were not involved, arising issues on the safety of the vaccine.,
As mutations occur randomly at different stages of the life cycle, it becomes more difficult for therapeutic agents to reach and inhibit the specific target. Ultimately, the virus confronts these drugs to become a resistant strain, which can cause unimaginable loss of life. To avoid this, either the current therapeutic agents can be modified according to the genomic variations or novel agents can be designed. Through this commentary, we suggest that it is better to modify the currently available drugs and other therapeutic agents according to the detected variations of the virus, with the usage of bioinformatic tools like homology modeling. We also emphasize on the usage of CRISPR-Cas13 technology that is being used to detect the mutations as well as to develop a therapy with no drugs. For example, the employment of Cas13 with combinatorial arrayed reactions for multiplexed evaluation of nucleic acids effectively detected SARS-CoV-2. Also, the usage of prophylactic antiviral CRISPR in human cells strategy along with CRISPR-Cas13 has shown effective degradation of both influenza A and SARS-CoV-2 viral genomes. Designing and using of CRISPR-RNAs (crRNAs) targeting SARS-CoV-2 has revealed that these crRNAs efficiently target the viral genome of SARS-CoV-2. In total, the genome sequencing has revealed that it is essential to combat the pandemic along with its modifying variants. It has also given us the potential druggable targets, which can be manipulated for the betterment of life.
Financial support and sponsorship
Conflicts of interest
The authors reported no conflicts of interest.
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