In December 2019, a large number of Pneumonia like cases of unknown etiology were detected in the Wuhan province of China (Bogoch et al.,2020, Lu et al.,2020). Soon the outbreak was declared as a public health emergency and was termed as severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) by WHO (WHO report,2019). Subsequently, it was named Coronavirus disease-19 (COVID-19) in January 2020.
The most common symptoms of this infection include, mild to severe respiratory infection. It was observed that this virus affects the respiratory system primarily and then spreads systemically to the heart, liver, and kidneys. The patients suffering from conditions like cardiovascular diseases, diabetes, cancer, and immunosuppression usually developed a serious illness as compared to others (Bai et al.,2020). COVID-19 is spread through contact and respiratory droplets and its high transmissibility is supposed to be related to its high viral loads in the upper respiratory tract initiating viral pneumonia(Bai et al.,2020).
Presently, almost all the countries on the globe have been affected by COVID-19 and the impact is frightening. It has resulted in millions of infected cases and about a hundred thousand deaths, this has led the WHO to declare the outbreak as a global pandemic(Dong et al.,2020). Governments are compelled to impose mandatory quarantines and lockdowns( Pulla et al.,2020, Lau et al.,2020). This has also affected the world economy and if the situation is not controlled it is expected to worsen in the future (World Economic Situation And Prospects: September 2020, The global macroeconomic impacts of COVID-19: Seven scenarios,2020). Coronaviruses are a member of subfamily Coronavirinae, the viruses of this group are spherical, enveloped, and have a single-stranded RNA genome (Dhama et al. ,2020, Schoeman et al. ,2020).
It was found that around two-thirds of the RNA of COVID-19 is composed of replicase ORF1a/1b and the rest of the viral genome encodes the spike, envelope, membrane, and nucleocapsid(Cui et al.,2020, Wu et al.,2020, Lu et al. ,2020). Recent studies have shown that the genetic makeup of this virus is similar to bat-SL-CoVZC45, bat-SL-CoVZXC21, and SARS-CoV(Lu et al. ,2020). Both CoV-2 and SARS-CoV infect host cells using the same angiotensin-converting enzyme 2 (ACE2)(Hoffmann et al.,2020, Wan et al.,2020). However, the human to human transmission of SARS-CoV-2 is much faster with ACE2 receptor binding to SARS-CoV-2 with 10 to 20-fold higher capacity(Sciencemag.org, October 2020).
Considerable efforts are being made to improve the methods of detection as the most critical weapon to fight COVID-19 is its early detection. The detection techniques are a powerful tool to combat COVID-19 by playing a monitoring system (figure 1). Molecular techniques have a significant advantage over syndromic detection techniques and CT (computed tomography) scans as they offer specificity. The following are some of the molecular diagnostic techniques being used or being developed.
Nucleic Acid Testing
Nucleic acid testing by PCR (polymerase chain reaction) is popularly being used for the routine diagnosis of patients. RT-PCR (reverse transcription PCR) or qRT-PCR (quantitative PCR) offers qualitative detection of nucleic acids. In this technique, a matrix pooling strategy is used by collecting nasal swabs in transport media(FDA,2020). The sample is then treated to isolate RNA and then synthesize cDNA using enzyme reverse transcriptase. This cDNA is then added to the working mix of PCR. After amplification of cDNA by PCR, the cycle threshold (Ct) values are calculated and compared to control values. This determines the relative expression for the diagnosis of COVID-19. Studies are being conducted on RT-insulated isothermal PCR (RT-iiPCR)(Go et al.,2017), reverse transcription loop-mediated isothermal amplification (RT-LAMP) (Shirato et al. ,2014), and a one-step rRT-PCR assay(Hashemzadeh et al.,2016).
Antibody Detection
An antibody is a Y-shaped protein that binds to antigen as an immune response. IgM antibodies have a potential role for SARS-CoV-2 as a marker. Guo and colleagues have shown using ELISA with recombinant SARS-CoV-2 NP antigen that IgM antibodies can be detected in more than 85% of infected people(Guo et al. ,2020). Similar studies are being performed on IgA and IgG antibodies as well. Although, IgG has a more significant role to play since it lasts longer. A positive SARS-CoV-2 IgG result suggests prior infection with the virus but it does not independently imply protective immunity and its duration. Several surface-modified immuno sensors can also be developed for virus detection(Uzun et al.,2009, Shafiee et al.,2014, Yanik et al.,2010, Lee et al.,2011, Kaushik et al.,2018, Ashiba et al.,2017, Nidzworski et al.,2014, Theel et al.,2020).
Biosensors
A biosensor is a device that converts biological stimulus into a response that can be analyzed. For the diagnosis of this infection optical biosensors are being explored to get it commercialized. Jing Wang and coworkers developed an optical biosensor for SARS-CoV-2 using gold nanoislands on a glass substrate(A new biosensor for the COVID-19 virus: Detection in the environment – ScienceDaily,2020). DNA receptors complementary to the sequence of SARS-CoV-2 were embedded and detected using the optical phenomenon of localized surface plasmon resonance. Localized surface plasmon resonance is the phenomenon in which metal nanoparticles exhibit unique optical properties when the wavelength of light incident on a metal surface is comparable to the wavelength of electrons oscillating in the conduction band and exhibit resonance.
CRISPR-Technology
CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats) technology is developed from the innate immunity of bacteria and archaea. This technology has allowed drastic development in the area of gene editing. Feng Zhang et al. developed CRISPR-based SHERLOCK (Specific High Sensitivity Enzymatic Reporter UnLOCKing) for the detection of COVID-19( Zhang et al.,2020). According to the detection protocol, nucleic acid extraction is followed by isothermal amplification, detection of pre-amplified viral DNA, and visual detection using a paper dipstick.
Nanomaterial based approach
Being the most important step to identify and isolate infected individuals, several nanotechnology-based approaches for detection are now being developed (Cohen et al.,2020, Kumar et al. 2020). Most of the techniques associated with COVID-19 testing are based on the detection of antibodies e.g. enzyme-linked immunosorbent assay (ELISA) and RNA-based techniques e.g. PCR. The basic principle behind all these techniques is surface interactions with a complementary detection ligand or strand in the kit. But these techniques have few disadvantages like poor sensitivity, false-negative results, and take a lot of time to give a response(Vaculovicova et al.,2017). Nanosized materials can be very useful in this detection as these provide faster yet reliable detection to obtain highly efficient surface interactions between the sensor and the analyte (Mokhtarzadeh et al., 2017). Such concepts are being critically accelerated for the development of new techniques to combat the global COVID crisis.
The recent advancement in the field of nanoscience and technology has enabled the researchers to provide solutions to various research and industrial problems. Nanomaterials are classified as materials with at least one dimension in the nano range 1-100 nm. Several nanomaterials including metals, metal oxides, dendrimers, zeolites, etc. find their application in industry. Amidst the COVID-19 pandemic, nanotechnology has yet again proved to be a popular tool. Due to the increased surface area to volume ratio, nanoparticles show unique properties at the nano level. Although nanotechnology holds potential scope in diagnosis, prevention, and treatment the focus of the article is on the role of diagnosis only. Quantum dots are semiconductor nanoparticles with extraordinary optical and electronic properties. These nanoparticles are 2-10 nm and have specifically high quantum yield, fluorescence, tunable light emission, etc. Carbon-based nanosystems have been explored for various viral disease evaluation(Bhattachariya et al.,2017). Along with quantum dots, several organic and inorganic nanoparticles are being explored for the diagnosis of this disease (Bhavana et al.,2020).
Other Developing Techniques
Molecular diagnostic tools play a pivotal role in the early detection of COVID-19. It is essential to diagnose evry suspected cases at hospital, but results take time from few hours to days which slow the treatment and screening process. Thus, there is need of rapid diagnostic tool which can provide the diagnosis result within few seconds and required to be commercialized.
Conclusion
Molecular diagnostic tools play a pivotal role in the early detection of COVID-19. It is essential to diagnose evry suspected cases at hospital, but results take time from few hours to days which slow the treatment and screening process. Thus, there is need of rapid diagnostic tool which can provide the diagnosis result within few seconds and required to be commercialized.
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