Biotechnological Perspectives to Combat the COVID-19 Pandemic: Precise Diagnostics and Inevitable Vaccine Paradigms

Abstract

The outbreak of the novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the cause for the ongoing global public health emergency. It is more commonly known as coronavirus disease 2019 (COVID-19); the pandemic threat continues to spread aroundthe world with the fluctuating emergence of its new variants. The severity of COVID-19 ranges from asymptomatic to serious acute respiratory distress syndrome (ARDS), which has led to a high human mortality rate and disruption of socioeconomic well-being. For the restoration of pre-pandemic normalcy, the international scientific community has been conducting research on a war footing to limit extremely pathogenic COVID-19 through diagnosis, treatment, and immunization. Since the first report of COVID-19 viral infection, an array of laboratory-based and point-of-care (POC) approaches have emerged for diagnosing and understanding its status of outbreak. The RT-PCR-based viral nucleic acid test (NAT) is one of the rapidly developed and most used COVID-19 detection approaches. Notably, the current forbidding status of COVID-19 requires the development of safe, targeted vaccines/vaccine injections (shots) that can reduce its associated morbidity and mortality. Massive and accelerated vaccination campaigns would be the most effective and ultimate hope to end the COVID-19 pandemic. Since the SARS-CoV-2 virus outbreak, emerging biotechnologies and their multidisciplinary approaches have accelerated the understanding of molecular details as well as the development of a wide range of diagnostics and potential vaccine candidates, which are indispensable to combating the highly contagious COVID-19. Several vaccine candidates have completed phase III clinical studies and are reported to be effective in immunizing against COVID-19 after their rollout via emergency use authorization (EUA). However, optimizing the type of vaccine candidates and its route of delivery that works best to control viral spread is crucial to face the threatening variants expected to emerge over time. In conclusion, the insights of this review would facilitate the development of more likely diagnostics and ideal vaccines for the global control of COVID-19.

Keywords: ARDS; COVID-19; CRISPR; SARS-CoV-2; coronavirus; detection methods; nucleic acid test (NAT); vaccine.

Figure 1

Flow chart of RT-PCR--based detection of SARS-CoV-2 infection: (1) collection of the test sample from anindividual; (2) mixing the collected sample with viral transport media and storageat 4 °C until use; (3) isolation of viral RNAs from the collected sample; (4) synthesis of cDNA from viral RNA by reverse transcription; (5) RT-PCR with specific primers/fluorescent markers demonstrates either positive or negative results for SARS-CoV-2 detection.

Figure 2

RT-LAMP technique-based detection of SARS-CoV-2 infection: (1) collection ofhuman samples; (2) mixing of the collected sample with viral transport media and storageat 4 °C until use; (3) single-stranded RNA genome of SARS-CoV-2 with the targeted genes; (4) specific primers binding atthe targeted genes; (5) synthesizing the first-strand cDNA by reverse transcription; (6) DNA polymerization for second-strand cDNA synthesis; (7) dumbbell-shaped DNA formation during the process of cDNA synthesis; (8) RT-PCR with specific primers/fluorescent markers demonstrates either positive or negative results for SARS-CoV-2 detection.

Figure 3

Transcription-mediated amplification (TMA)-based detection of SARS-CoV-2 infection: (1) reverse transcription-based first-strand cDNA synthesis; (2) RNA strand of hybrid RNA–cDNA is degraded by the RNase H activity of the enzyme reverse transcriptase; (3) DNA polymerization for the synthesis of second-strand cDNA; (4) PCR amplification with specific primers will demonstrate either positive or negative results for SARS-CoV-2 virus detection (multiple arrows indicate multiple cycles of amplification).

Figure 4

CRISPR/Cas-based detection of SARS-CoV-2 infection: (1) reverse transcription of isolated viral RNA; (2) reverse transcription for the synthesis of cDNA; (3) CRISPR-mediated genome editing (identify and cleave) actat the precise sequence of viral RNA SARS-CoV-2 genome. Both Sherlock and Detector of the CRISPR tool convert viral RNA to DNA (isothermal amplification), which activates nuclease enzyme activity (Cas-12/13) to cleave the target sequence (pink colored symbols indicate DNA polymerase;red colored symbols indicates primer); (4) loading of the sample (fluorescence RNA reporter) on to strip for detection of the specific viral RNA sequence; (5) the number of bands visible on the strip (lateral flow) represents whether the test is positive or negative for SARS-CoV-2 infection.

Figure 5

Genome sequencing for detection of SARS-CoV-2 variants: (1) swab/test sample collection from an individual; (2) mixingthe collected sample with viral transport media and storage at 4 °C; (3) isolation of viral RNA from the collected sample; (4) cDNA synthesis from viral RNA by reverse transcription; (5) PCR-aided amplification; (6) computer-aided library preparation and collection of nucleotide databases; (7) further data analysis: RNA/whole-genome sequencing via NGS to identify SARS-CoV-2 variants assist in redesigning novel molecular-based diagnostics therapies to combat COVID-19.

Figure 6

Diagnostic flow chart for COVID-19 disease detection.The probable flow chart used to rule out positive or negative test results forSARS-CoV-2infections in asymptomatic and symptomatic individuals with COVID-19 [103,104,105,106,107,108,109,110,111,112,113].

Figure 7

Rapid antigen test (RAT) kit-based detection of SARS-CoV-2 infection: (1) test samples are collected in the form of swabs; (2) mixing the collected swab samples with buffer; (3) loading collected sample into the well of the antigen-coated strip; (4) formation of one band representsa negative test, and two bands represent a positive test for COVID-19 infection.

Figure 8

Structure of the SARS-CoV-2 virus. The structure of the COVID-19 virus contains the spike glycoprotein (SP), membrane protein (MP), nucleocapsid protein (NP), an envelope protein (EP), and single-stranded +RNA as the genome.

Figure 9

Various types of vaccine platforms used to mitigate COVID-19 pandemic. Major types of vaccine platforms: viral vector vaccines of non-replicating andreplicating types; nucleic acid vaccines based on DNA and mRNA; vaccines based on recombinant proteins/subunit/VLPs virus-like particle;vaccines based on virus-based on inactivated and live attenuated viral components.

Figure 10

Viral vector-based vaccines for the treatment of COVID-19: (1) modification of adenovirus by removing pathogenic genes to make them non-infectious; (2) insertion of a gene of interest encoding the SARS-CoV-2 virus’s spike protein into an adenovirus system; (3) making a modified adenovirus-based vaccine and their administration into individuals; (4) adhesion of a viral vector to human cells and delivery of the antigenic determinant; (5) expression of a viral vector antigenic determinant (spike protein) and its display on the surface of human cells; (6) recognition of viral spike protein by immune cells; (7) immune cells’ production of antibodies (NABs) against viral spike protein; (8) immune responses with elicited antibodies neutralize the SARS-CoV-2 virus in vaccinated people.

Figure 11

Mechanism of mRNA-based vaccine for developing immunity against COVID-19 infection: (1) intramuscular injection of mRNA-based vaccine; (2) entry and delivery of spike protein-encoding mRNA into human cells; (3) decoding of viral mRNA into spike protein and its display on the surface of human cells; (4) recognition of viral spike protein as antigenic determinant by immune cells; (5) immune cells’ production of antibodies (NABs) against viral spike proteins; (6) immune responses elicit antibodies to neutralize the SARS-CoV-2 virus in vaccinated people.

Figure 12

Mechanism of spike protein-based vaccines for developing immunity against COVID-19 infection: (1) spike proteins of the SARS-CoV-2 virus is enclosed inside a capsule; (2) spike proteins are mixed with adjuvants; (3) antigen-presenting structures are made with spike proteins; (4) virus-like particles are made with native spike proteins of the SARS-CoV-2 virus. The spike protein-based vaccines arecreated using one of the above four represented protein formulations. Upon vaccination with thespike protein-based vaccine, the antigen-presenting cells recognize the virus’s spike protein and present it to immune cells (i.e., Tcells and Bcells), resulting in both cell and antibody-mediated immunity.

Figure 13

Mechanism of whole virus-based vaccine for inducing immunity against COVID-19: (1) whole or native virus-based vaccines are prepared by inactivating the whole virus; (2) whole or native virus-based vaccines are also prepared by attenuating the virus. Whole virus-based vaccines aremade with one of the above two represented protein formulations. Upon vaccination of the whole virus-based vaccine, the antigen-presenting cells recognize the inactivated or attenuated SARS-CoV-2 virus and present it to immune cells (i.e., T cells and B cells) to mediate both cell and antibody-mediated immunity.