key: cord-0787434-79vkkiyc
authors: Fani, Mona; Zandi, Milad; Soltani, Saber; Abbasi, Samaneh
title: Future developments in biosensors for field‐ready SARS‐CoV‐2 virus diagnostics
date: 2020-09-24
journal: Biotechnol Appl Biochem
DOI: 10.1002/bab.2033
sha: 823b6822be50cd0df4689ddbba3972654b298321
doc_id: 787434
cord_uid: 79vkkiyc

According to the evidence, the Coronavirus disease 19 (COVID‐19) is caused by a zoonotic pathogen named respiratory syndrome coronavirus 2 (SARS‐CoV‐2). This virus can spread through personal contact, respiratory droplets, and also through airborne transmission. A rapid, low‐cost, and effective biosensor platform is essential to diagnose patients with COVID‐19 infection, predominantly the asymptomatic individuals, and prevent the spread of the SARS‐CoV‐2 via transmission routes. The objective of this review is to provide a comparative view among current diagnostic methods, focusing on recently suggested biosensors for the detection of SARS‐CoV2 in clinical samples. A capable SARS‐CoV‐2 biosensor can be designed by the holistic insights of various biosensor studies. This article is protected by copyright. All rights reserved

Severe Acute Respiratory Syndrome (SARS) and Middle East Respiratory Syndrome Coronaviruses have a single-stranded RNA and belong to the Coronaviridae family in the Nidovirales order. The subgroups of this family based on genetic properties are alpha (α), beta (ß), gamma (ɣ) and delta (δ) coronavirus. Since the past two decades, Betacoronaviruses (SARS, MERS, and SARS-CoV-2) have been investigated by researchers due to emerging and re-emerging. These infectious agents effect on the upper respiratory tract (URT) and lower respiratory tract (LRT) and also can involve the gastrointestinal system, heart, kidney, liver, and central nervous system resulting in multiple organ failure and generally, the flu-like symptoms of COVID-19 infection include fever, headache, joint pain, rash, and fatigue [2] .

As a member of the Coronavirus genus, SARS-CoV-2 showed over 80% identical to SARS-CoV (CoVZXC21 or CoVZC45) and bat SARS-CoV based on the sequencing of receptor binding spike glycoprotein (S-Spike). The analysis of the nucleic acid sequence confirmed that the SARS-CoV-2 also uses ACE2 (angiotensin-converting enzyme 2) for the cell attachment, as was previously employed by the SARS-CoV. However, the current information indicate that SARS-CoV-2 is more infectious than SARS CoV [3] .

The coronavirus genome encodes several structural and non-structural proteins. S-Spike glycoprotein is viral membrane antigen and consists of two subunits of S1 and S2. The

Receptor Binding Domain (RBD) locates in the S1 subunit and binds to the ACE2, on the other hand, the S2 subunit provides the viral fusion and entry process into the target cell.

Membrane (M) protein promotes the membrane curvature. It plays an essential role in viral assembly, Envelope (E) protein is needed to release the virus, Nucleocapsid (N) protein is interferon antagonistic and can support the viral replication [4] .

The coronaviruses non-structure proteins, such as RNA-dependent RNA polymerase (RdRp), 3C-like proteases (3CLpro, which is a main protease), and papain-like protease (PLpro) are essential for viral replication. These proteins activity result in blocking the host immune system cells expression. Actually, after the SARS-CoV-2 entrance to the host cells, the viral genome is translated into viral polyprotein and subsequently cleaved into effector proteins by viral proteinases 3CLpro and PLpro. On the other hand, PLpro can suppress the immune response via deubiquitinase of interferon factor 3 and NF-κB. RdRp catalyzes the replication of viral genomic from a full-length negative-strand RNA template [5] .

Given the SARS-CoV-2 has been recently discovered, little immunological evidence is available. Previous reports have shown that both humoral and cellular immunity play vital roles in protective responses against SARS-CoV-2. Although antibodies against structure proteins (exclusive N and S proteins) are highly immunogenic, they have a relatively short lifespan. Compared with the humoral immunity specific reaction, the cellular immunity components such as T-helper cells, suppressor T-cells, and cytotoxic T-cells responses can more induce long-lasting protection against SARS-CoV-2 [6] .

Real-time polymerase chain reaction (real-time PCR) is known as an effective and sensitive method [7] . However, the false-negative results can occur that demand the fabrication of an accurate, rapid, and free-PCR technique for diagnosing COVID-19 infection as an alternative and first test compared to current diagnostic techniques.

In recent years, there has been an increasing interest in the biosensor, a transportable analytical device used for detecting various microorganisms and composed of biological molecules with a detector [8] . The device requires the efficient immobilization of antibodies, peptides, aptamers, or nucleic acids on the surface of a transducer responsible for the analyte recognition.

Biosensor introduced new opportunities for reliable, economical, and sensitive detection, particularly for the early detection of infectious diseases. Additionally, the materials (including graphene, gold nanoparticles, polyaniline-multiwall carbon nanotube, Etc.) with the nanometer scale have been used to reach the nano structuration of biosensors. Biosensors This article is protected by copyright. All rights reserved. 4

facilitate the output signal study by cyclic voltammetry (CV), square wave voltammetry (SWV), differential pulse voltammetry (DPV), and electrochemical impedance spectroscopy (EIS) [9] .

In this review, we highlighted the limitation of current techniques. We reviewed the literature on the fabrication of biosensors for the detection of SARS-CoV-2 to encourage researchers to develop further strategies for the detection of COVID-19 disease.

Several inflammation-related parameters increase or decrease in patients with COVID-19infection. They are as screening tests for the prognostics of the COVID-19 infection.

Particularly, C-reactive protein (CRP), lymphocyte count, interleukin-6 (IL-6), interleukin-10 (IL-10), lactate dehydrogenase (LDH), platelet count, D-dimer, and serum-ferritin [10] .

perform SARS-CoV-2 detection: (1) real-time PCR that is a gold-standard method because of its high selectivity and relatively high sensitivity for detection of COVID-19 infection (2) gene sequencing; (3) serological tests, and (4) chest computed tomography (CT) [11] .

Nevertheless, for asymptomatic individuals who have traveled to high-risk areas for COVID-19 infection or contacted with infected people, the preferred detection method is RT-PCR [2] . CT scan is used to confirm the false-negative results real-time PCR from symptomatic patients or as a separate diagnostic tool for the detection of Covid-19 infection. In this method for capturing three-dimensional (3D) images, several X-ray images of the chest are taken to identify SARS-CoV2 infection, which can involve the lower parts of single or both lobes [13] .

According to molecular techniques, laboratories detect the COVID-19 infection using realtime PCR methods within 4−6 h. However, limitations are designing probes-primers, sample preparation, sampling error, the requirement to equipment tools, and their need for multitemperature sample heating for denaturation, annealing, and extension [8] . On the other hand, traveling to a laboratory for testing increases the risk of spreading COVID-19 infection. Also, it was reported several false-positive or -negative results, in the early stages of the COVID-19 infection especially.

The serological assay is rapid and requires minimal equipment, but its efficacy may be This article is protected by copyright. All rights reserved. 6

Many research groups have also focused on sensor methods to delete complicated stages of sample preparation and also reduce the possibility of false-positive and false-negative results and using expensive laboratory equipment.

Asymptomatic patients are a major threat to public health because they are a potential source RdRp, ORF1ab, and E genes [18] .

In parallel work, Murugan et al. proposed both approaches (two plasmonic labeled and labelfree immunoassays) based on the U-bent optical fiber sensor system (P-FAB) for detection of SARS-CoV-2 N gene in the saliva sample. In the label-free bioassay, a U-bent fiber-optic probe as a platform covers with gold nanoparticles, followed by covalent conjugation of anti-N protein monoclonal antibodies via a thiol-PEG-NHS based coupling chemistry. In this way, the results can be obtained within 15 min. The labeled bioassay manner is based on the sandwich immunoassay. The U-bent fiber-optic probe is immobilized with anti-N protein monoclonal antibodies, and gold nanoparticles subsequently is treated with bovine serum albumin (BSA) solution to minimize the non-specific interactions. Then, the modified platform is exposed to saliva samples for 5 min. The signal response can be obtained within 10-15 min. Therefore, the P-FAB system has excellent potential in the detection of COVID-19 infection [19] . The analytical performance of current biosensors for detection of COVID-19 infection were listed in Table 1 .

This review paper overviews the conventional methods and biosensors based techniques that have been recently used to detect the SARS-CoV-2. Currently, real-time PCR is the main and powerful assay for the detection of COVID-19 infection. We found out that conventional methods cannot meet the rapid detection demands and challenges in the viral analysis.

In conjunction with real-time PCR, CT scan significantly increases the sensitivity, facilitating clinical counseling, and improving treatment outcomes. Although the Real-time PCR is an This article is protected by copyright. All rights reserved. 8

accurate method compared with the current assays, it has some limitations. As mentioned above, due to the drawbacks of current diagnostic methods in the early stage infection, the employment of some advanced methods, such as microfluidics, biosensors, and lab-on-a-chip systems, will be recommended as suitable methods for the diagnosis of SARS-CoV-2.

In recent years, biosensor methods have been considered innovative and promising tools for detecting other viruses, which, in contrast to the conventional methods, are less complicated to use and free of prolonged experimentation processes. The biosensor systems are rapid and specific for infection detection, and a physician can quickly decide whether the treatment is needed or not.

Generally, a DNA-based biosensor can detect the pathogens and record the information of them in clinical diagnostics. DNA biosensor is mainly comprised of a bioreceptor and a transducer. A bioreceptor is a DNA probe designed from a conserved sequence to recognize the pathogens DNA by a transducer using converting the biological signal into the desired signal.

DNA biosensor as an alternative method for current techniques can provide rapid response and also is the high sensitivity and low cost. A schematic diagram of the DNA-based biosensor is summarized in Fig 1. It is recommended that further research be undertaken on the developing DNA biosensor in the following subjects. First, choice of a more conserved and specific gene of SARS-CoV-2 (We provide SARS-CoV-2-specific probes and target based on highly conserved regions of the S, E, and N proteins and nonstructural (RdRp and ORF1ab) proteins in Table 2 .) Second, the biosensors' study of performance with the large sample size. Third, a comparison of the sensitivity and specificity of the biosensor with current methods. 

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