A Rapid Point-of-Care Dipstick Assay for Differentiation of Sars-Cov-2 Variants in Covid-19 Patients

Intro: Delta and Omicron variants of SARS-CoV-2 are highly contagious, currently dominating the globe and recognized as variant of concern (VOC). The transmissibility efficiency of viruses, disease symptoms, and severity of COVID- 19 is highly heterogeneous. Therefore, testing at the community level is essential to identify the infected people at an early stage-carrying VOC to reduce the spread of virus and combat the pandemic. Method(s): In this study, we analysed thousands of genome sequences representing 30 different SARS-CoV-2 variants and identified Delta and Omicron variants specific nucleic acid signatures in the spike gene. Based on the variant specific nucleic acid sequences we synthesized different oligos and optimized a mPCR assay that can specifically differentiate the Delta and Omicron variants. We further translated our work into a dipstick assay (Tohoku Bio-array, Japan) by adding tag linker sequence to 5' end of the forward primer and adding biotin in 3' end of the oligos. Finding(s): A total of 250 samples were subjected to WGS using MiSeq platform and these confirmed samples were processed for validation of our specific designed probes using PCR assay and the readout was found to be 100% specific to Delta, BA.1 & BA.2 of SARS-CoV-2 variants which were further confirmed by Sanger sequencing. The dipstick was used to screen these samples, and specific signals were observed. WGS and Sanger sequencing were used to validate our PCR and dipstick assay results, and the readout was found to be 100% specific. The results can be visualised by the naked eye and interpreted easily. Conclusion(s): This study developed a rapid point-of-care test of SARS-CoV-2 patients, which can differentiate Delta, BA.1 and BA.2 variants at the same time of confirmation of the infection in patient. The current nucleic-acid chromatography-based dipstick assay is highly specific and can work even in the case of low viral load as well.Copyright © 2023

Enhanced electrochemiluminescence immunoassay: 2. Enabling signal detection at an early stage of incubation for rapid point-of-care testing

Signal detection in a label-based immunoassay is performed normally when the antigen/antibody binding reaction reaches the equilibrium state during the incubation period of an assay process. Shortening the incubation period in an assay helps reduce the turnaround time and is particularly valuable for point-of-care testing, but the cost is the reduction of signal level and, possibly, measurement precision as well. This work demonstrates that the signal loss could be offset by the stronger emission of an electronically neutral ruthenium(II) complex label, Ru(2, 2′-bipyridine) (bathophenanthroline disulfonate)[4-(2, 2′-bipyridin-4-yl)butanoic acid], used in the electrochemiluminescence (ECL) immunoassay. Combined with the uniquely well-established flow-through washing process in the automated ECL analyzers and the precise control over liquid handling, the assays performed with a 5-minute incubation period showed the same signal level and measurement precision as those of conventional ECL assays. Additionally, the absence of biotin and streptavidin components in the reagent formulation avoids the biotin-streptavidin interaction during assay incubation and fundamentally eliminates the interference of biotin, especially when used in some high-dose therapies. The results obtained from the procalcitonin prototype kit and the supporting evidence from other preliminary reagents (for SARS-CoV-2 N protein and troponin T) are general. The nonequilibrium detection, along with the downsized instrument design, makes the enhanced ECL (EECL) technology a fast high-performance POCT platform that provides the same high-quality data as those generated from the widely deployed [Ru(bpy)3]2+ based laboratorial ECL systems. The anticipated regulatory approval and follow-up clinical implementation will be a significant stride in the decade-long pursuit of novel ECL labels. © 2023 The Author(s)

Electrochemical immunosensor nanoarchitectonics with the Ag-rGO nanocomposites for the detection of receptor-binding domain of SARS-CoV-2 spike protein.

As the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) poses a grave threat to human life and health, it is essential to develop an efficient and sensitive detection method to identify infected individuals. This study described an electrode platform immunosensor to detect SARS-CoV-2-specific spike receptor-binding domain (RBD) protein based on a bare gold electrode modified with Ag-rGO nanocomposites and the biotin-streptavidin interaction system. The Ag-rGO nanocomposites was obtained by chemical synthesis and characterized by electrochemistry and scanning electron microscope (SEM). Cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS) were used to record the electrochemical signals in the electrode modification. The differential pulse voltammetry (DPV) results showed that the limit of detection (LOD) of the immunosensor was 7.2 fg mL-1 and the linear dynamic detection range was 0.015 ~ 158.5 pg mL-1. Furthermore, this sensitive immunosensor accurately detected RBD in artificial saliva with favorable stability, specificity, and reproducibility, indicating that it has the potential to be used as a practical method for the detection of SARS-CoV-2.

Rapid and simple detection of influenza virus via isothermal amplification lateral flow assay.

Respiratory illness caused by influenza virus is a serious public health problem worldwide. As the symptoms of influenza virus infection are similar to those of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, it is essential to distinguish these two viruses. Therefore, to properly respond to a pathogen, a detection method that is capable of rapid and accurate diagnosis in a hospital or at home is required. To satisfy this need, we applied loop-mediated isothermal amplification (LAMP) and an isothermal nucleic acid amplification technique, along with a system to analyze the results without specialized equipment, a lateral flow assay (LFA). Using the platform developed in this study, all processes, from sample preparation to detection, can be performed without special equipment. Unlike existing PCR methods, the nucleic acid amplification can be performed in the field because hot packs do not require electricity. Thus, the designed platform can provide rapid results without the need to transport the samples to a laboratory or hospital. These advantages are not limited to operations in developing countries with poor access to medical systems. In conclusion, the developed technology is a promising tool for infectious disease management that allows for rapid identification of infectious diseases and appropriate treatment of patients.

A new-type HOCl-activatable fluorescent probe and its applications in water environment and biosystems.

The outbreak and spread of Corona Virus Disease 2019 (COVID-19) has led to a significant increase in the consumption of sodium hypochlorite (NaOCl) disinfectants. NaOCl hydrolyzes to produce hypochlorous acid (HOCl) to kill viruses, which is a relatively efficient chlorine-based disinfectant commonly used in public disinfection. While people enjoy the convenience of NaOCl disinfection, excessive and indiscriminate use of it will affect the water environment and threaten human health. Importantly, HOCl is an indispensable reactive oxygen species (ROS) in human body. Whether its concentration is normal or not is closely related to human health. Excessive production of HOCl in the body contributes to some inflammatory diseases and even cancer. Also, we noticed that the concentration of ROS in cancer cells is about 10 times higher than that in normal cells. Herein, we developed a HOCl-activatable biotinylated dual-function fluorescent probe BTH. For this probe, we introduced biotin on the naphthalimide fluorophore, which increased the water solubility and enabled the probe to aggregate in cancer cells by targeting specific receptor overexpressed on the surface of cancer cell membrane. After reacting to HOCl, the p-aminophenylether moiety of this probe was oxidatively removed and the fluorescence of the probe was recovered. As expected, in the PBS solution with pH of 7.4, BTH could give full play to the performance of detecting HOCl, and it has made achievements in detecting the concentration of HOCl in actual water samples. Besides that, BTH had effectively distinguished between cancer cells and normal cells through a dual-function discrimination strategy, which used biotin to enrich the probe in cancer cells and reacted with overexpressed HOCl in cancer cells. Importantly, this dual-function discrimination strategy could obtain the precision detection of cancer cells, thereby offering assistance for improving the accuracy of early cancer diagnosis.

Biotin and thyroid function test

Introduction: Biotin is a vitamin-B supplement and is sold over-the-counter alone or in combination with other vitamins and minerals to improve skin, nail, and hair health. Exogenous biotin intake is becoming a common cause of apparent abnormal thyroid function tests in endocrine practice. We present a case report. Case Description: A 55-year-old lady was referred to endocrinology clinic for low TSH but normal fT4. The only symptom she had was abnormal smell of ‘smoke’. She had no ongoing symptoms of thyrotoxicosis or hypothyroidism apart from hair thinning. She had COVID-19 infection with pyrexia and constitutional symptoms a year ago. There was no other past medical or family history of note. On direct questioning, she admitted taking biotin 300ug every day in preceding six months for hair loss. Examination revealed euthyroidism and no goitre. TFTs prior to commencing biotin were normal. Further TFTs confirmed low TSH but normal fT4 and fT3 and thyroid receptor antibody was negative. TFTs off biotin for three days were normal and she was reassured. Discussion: There is no in vivo relationship between biotin and thyroid hormones, but high dose biotin affects thyroid function test in vitro when assessed with immunoassays using streptavidin-biotin immobilizing system. Apparently low TSH with or without raised T4 suggests thyrotoxicosis or thyroxine over-replacement. All patients presenting with such abnormal thyroid function tests should be questioned about biotin intake along with iodine as many multivitamins contain high dose biotin and patients are often unaware. This can avoid unnecessary investigations, patient anxiety, and healthcare costs. American Thyroid Association advises stopping biotin two days before thyroid function test. Patients can continue to take biotin except temporarily before thyroid blood tests.

The great value of clinical laboratories in the pandemic process

Laboratory Medicine has been showing its great Value during Covid-19 pandemic in the whole care process: diagnosis, screening, follow up and outcome of the disease. The diagnostic tools to date are able to dose viral RNA, and antibodies against viral proteins or viral proteins themselves. Even if the conventional RT-PCR has been the most widely used method, the greatest innovation in molecular diagnosis was reported by the so called CRISPR Community. The first serological tests to be introduced were rapid serological tests with direct reading (first generation) or with fluorescence reading (2nd generation) and microfluidic with fluorescent reading (3rd generation). Then conventional CLIA method have been introduced with better sensitivity and more recently a new kind of serological assay has been proposed able to detect anti SARS-Cov-2 serum antibodies throughout a competitive streptoavidin/biotin assay which utilize RBD as coated antigen and ACE-2 labelled with biotin as competitor molecule for serum antibodies. Until few months ago the harmonization between many methods produced by many manufacturers was impossible. Recently the WHO produced an international standard (pool of sera-total antibodies) able to permit a sort of harmonization. To date NGS return to be widely utilized in the effort to intercept new variants. Finally, pathophysiology and natural course of Post-acute Sequelae of COVID-19 (PASC) also called Long Covidis unclear, meriting further studies. Also in this almost still unknown field, Clinical Laboratory could contribute in diagnosis and monitoring.

Construction of Orthogonal Modular Proteinaceous Nanovaccine Delivery Vectors Based on mSA-Biotin Binding.

Proteinaceous nanovaccine delivery systems have significantly promoted the development of various high-efficiency vaccines. However, the widely used method of coupling the expression of scaffolds and antigens may result in their structural interference with each other. Monovalent streptavidin (mSA) is a short monomer sequence, which has a strong affinity for biotin. Here, we discuss an orthogonal, modular, and highly versatile self-assembled proteinaceous nanoparticle chassis that facilitates combinations with various antigen cargos by using mSA and biotin to produce nanovaccines. We first improved the yield of these nanoparticles by appending a short sugar chain on their surfaces in a constructed host strain. After confirming the strong ability to induce both Th1- and Th2-mediated immune responses based on the plasma cytokine spectrum from immunized mice, we further verified the binding ability of biotinylated nanoparticles to mSA-antigens. These results demonstrate that our biotinylated nanoparticle chassis could load both protein and polysaccharide antigens containing mSA at a high affinity. Our approach thus offers an attractive technology for combining nanoparticles and antigen cargos to generate various high-performance nanovaccines. In particular, the designed mSA connector (mSA containing glycosylation modification sequences) could couple with polysaccharide antigens, providing a new attractive strategy to prepare nanoscale conjugate vaccines.

Technical Note: Bimodal Negative Interference of Biotin in Roche IL-6 Immunoassay

OBJECTIVE: Interleukin -6 (IL-6) is an important diagnostic test in COVID-19 patients to determine whether to initiate tocilizumab therapy or mechanical ventilation. We investigated potential interference of biotin in Roche IL-6 assay which utilizes biotinylated antibody. METHODS: We prepared three serum pools from left-over specimens which showed IL-6 values over 40 pg/mL. Then aliquots of each serum pool were further supplemented with various amounts of biotin expected in patients taking biotin supplement and then IL-6 values were measured again using Roche IL-6 assay on the Cobas e411 analyzer. RESULTS: We observed negative interference of biotin in IL-6 assay but interference was bimodal as maximum negative interference was observed with 100 ng/mL biotin but not with 1000 ng/mL. However, no interference was observed in the presence of 25 ng/mL biotin. CONCLUSIONS: Biotin showed negative interference with IL-6 assay.

Antimicrobial Photodynamic Disinfection (APDT) Destroys SARS-COV-2 Receptor Binding Domains and Spike Protein Independent of Viral Variant

Introduction: We previously reported on the elimination of the RNA signature of laboratory strains as well as wild-type SARS-CoV-2 using a photodynamic disinfection technique. This report extends the work to destruction of receptor binding domains, spike protein and nucleocapsid protein of major SARS-CoV-2 variants including the dominant Delta variant. Objectives: The objective of this work was to evaluate effect of a 2-min cycle of aPDT on receptor binding domains, spike protein and nucleocapsid protein of SARS-CoV-2. Methods: Spike glycoprotein receptor binding domain proteins from SARS-Related Coronavirus 2 included recombinant NR-54004 (United Kingdom Variant), NR-54005 (South African Variant), and NR-52366 (Wuhun-Hu-1 Variant) (BEI Resources, NIAID, ATCC). Recombinant spike proteins included B.1.1.7 Spike Protein, United Kingdom Variant (10748-CV-100), B.1.617 Spike Protein, Indian (Delta) Variant (10861-CV-100), B.1.351 Spike Protein, South African Variant (10777-CV-100), P.1 Spike Protein, Brazilian Variant (10795-CV-100) (all from R&D Systems, Inc.), and the stabilized spike glycoprotein, Wuhun-Hu-1 Variant (NR-52397). The SARS-CoV-2 nucleocapsid protein (NR55344, Avi-Histag, Biotin-Labeled, BEI Resources, NIAID, ATCC) was also tested for susceptibility to damage. aPDT was carried out by exposing each viral component to a photosensitizer formulation containing 320 uM methylene blue in an aqueous adjuvant, immediately followed by 36 J/cm2 (120 s) of non-thermal laser light at 664 nm (Steriwave® aPDT system, Ondine Biomedical Inc., Vancouver, B.C.). Illumination was conducted in a custom thin-film cell replicating the anterior nasal architecture. Control solutions were identically treated but without light exposure. 50 ul samples of treated and control solutions were evaluated by SDSPAGE using SYPRO Ruby Protein Gel Stain. Results: SDS-PAGE evaluation of all treated RBD, spike and SARSCoV-2 nucleocapsid protein samples showed no detectable protein remaining after 2 min exposure to photodynamic disinfection treatment. All no-light control samples remained unaffected. Conclusion: A 2-min photodynamic disinfection procedure was demonstrated to destroy RBD's, spike protein and nucleocapsid protein of major SARS-CoV-2 viral variants in circulation, expanding on previous work evaluating RNA damage in this virus. This outcome supports the use of aPDT as a potential SARS-CoV-2 suppression technique.

Bio-SCAN: A CRISPR/dCas9-Based Lateral Flow Assay for Rapid, Specific, and Sensitive Detection of SARS-CoV-2.

Simple, rapid, specific, and sensitive point-of-care detection methods are needed to contain the spread of SARS-CoV-2. CRISPR/Cas9-based lateral flow assays are emerging as a powerful alternative for COVID-19 diagnostics. Here, we developed Bio-SCAN (biotin-coupled specific CRISPR-based assay for nucleic acid detection) as an accurate pathogen detection platform that requires no sophisticated equipment or technical expertise. Bio-SCAN detects the SARS-CoV-2 genome in less than 1 h from sample collection to result. In the first step, the target nucleic acid sequence is isothermally amplified in 15 min via recombinase polymerase amplification before being precisely detected by biotin-labeled nuclease-dead SpCas9 (dCas9) on commercially available lateral flow strips. The resulting readout is visible to the naked eye. Compared to other CRISPR-Cas-based pathogen detection assays, Bio-SCAN requires no additional reporters, probes, enhancers, reagents, or sophisticated devices to interpret the results. Bio-SCAN is highly sensitive and successfully detected a clinically relevant level (4 copies/µL) of synthetic SARS-CoV-2 RNA genome. Similarly, Bio-SCAN showed 100% negative and 96% positive predictive agreement with RT-qPCR results when using clinical samples (86 nasopharyngeal swab samples). Furthermore, incorporating variant-specific sgRNAs in the detection reaction allowed Bio-SCAN to efficiently distinguish between the α, ß, and δ SARS-CoV-2 variants. Also, our results confirmed that the Bio-SCAN reagents have a long shelf life and can be assembled locally in nonlaboratory and limited-resource settings. Furthermore, the Bio-SCAN platform is compatible with the nucleic acid quick extraction protocol. Our results highlight the potential of Bio-SCAN as a promising point-of-care diagnostic platform that can facilitate low-cost mass screening for SARS-CoV-2.

Biotin-thiamine responsive basal ganglia disease in the era of COVID-19 outbreak diagnosis not to be missed: A case report.

BACKGROUND: Biotin-thiamine-responsive basal ganglia disease (BTRBGD) is a rare treatable autosomal recessive neurometabolic disorder characterized by progressive encephalopathy that eventually leads to severe disability and death if not treated with biotin and thiamine. BTRBGD is caused by mutations in the SLC19A3 gene on chromosome 2q36.6, encoding human thiamine transporter 2 (hTHTR2). Episodes of BTRBGD are often triggered by febrile illness. CASE REPORT: The patient was 2 years 10 months old male child presented with fever and progressive acute encephalopathy associated with severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) virus infection. MRI revealed bilateral symmetrical high signal involving both basal ganglia and medial thalami which is swollen with central necrosis, initially diagnosed as acute necrotizing encephalomyelitis with increased severity. Genetic analysis revealed BTRBGD. CONCLUSION: BTRBGD requires high index of suspicion in any patient presenting with acute encephalopathy, characteristic MRI findings (that are difficult to differentiate from necrotizing encephalopathy), regardless of the existence of a proven viral infection.

2021 ACLPS Annual Meeting Abstracts

The proceedings contain 38 papers. The topics discussed include: longitudinal assessment of SARS-CoV-2 nucleocapsid antigenemia in patients hospitalized with COVID-19;machine learning-based automated selection of regions for analysis on bone marrow aspirate smears;direct detection of beta-lactamase mediated antibiotic resistance in clinical specimens;convalescent plasma does not provide adequate replacement of C1-Inhibitor and complements in COVID-19 patients;mapping cell-to-cell mitochondria transfer in obesity using high-dimensional spectral flow cytometry;capture the tag: mitigation of biotin interference in ELISA and automated immunoassays by pre-conjugating biotinylated antibodies to the streptavidin surface;and assessment of coagulation tests in hospitalized COVID19 patients;challenging coagulopathies.

Equipment-free, salt-mediated immobilization of nucleic acids for nucleic acid lateral flow assays.

As the world has been facing several deadly virus crises, including Zika virus disease, Ebola virus disease, severe acute respiratory syndrome (SARS), Middle East respiratory syndrome (MERS), and Coronavirus disease 2019 (COVID-19), lateral flow assays (LFAs), which require minimal equipment for point-of-care of viral infectious diseases, are garnering much attention. Accordingly, there is an increasing demand to reduce the time and cost required for manufacturing LFAs. The current study introduces an equipment-free method of salt-mediated immobilization of nucleic acids (SAIoNs) for LFAs. Compared to general DNA immobilization methods such as streptavidin-biotin, UV-irradiation, and heat treatment, our method does not require special equipment (e.g., centrifuge, UV-crosslinker, heating device); therefore, it can be applied in a resource-limited environment with reduced production costs. The immobilization process was streamlined and completed within 30 min. Our method improved the color intensity signal approximately 14 times compared to the method without using SAIoNs and exhibited reproducibility with the long-term storage stability. The proposed method can be used to detect practical targets (e.g., SARS-CoV-2) and facilitates highly sensitive and selective detection of target nucleic acids with multiplexing capability and without any cross-reactivity. This novel immobilization strategy provides a basis for easily and inexpensively developing nucleic acid LFAs combined with various types of nucleic acid amplification.

Flex Printed Circuit Board Implemented Graphene-Based DNA Sensor for Detection of SARS-CoV-2.

Since the COVID-19 outbreak was declared a pandemic by the World Health Organization (WHO) in March 2020, ongoing efforts have been made to develop sensitive diagnostic platforms. Detection of viral RNA provides the highest sensitivity and specificity for detection of early and asymptomatic infections. Thus, this work aimed at developing a label-free genosensor composed of graphene as a working electrode that could be embedded into a flex printed circuit board (FPCB) for the rapid, sensitive, amplification-free and label-free detection of SARS-CoV-2. To facilitate liquid handling and ease of use, the developed biosensor was embedded with a user-friendly reservoir chamber. As a proof-of-concept, detection of a synthetic DNA strand matching the sequence of ORF1ab was performed as a two-step strategy involving the immobilization of a biotinylated complementary sequence on a streptavidin-modified surface, followed by hybridization with the target sequence recorded by the differential pulse voltammetric (DPV) technique in the presence of a ferro/ferricyanide redox couple. The effective design of the sensing platform improved its selectivity and sensitivity and allowed DNA quantification ranging from 100 fg/mL to [Formula: see text]/mL. Combining the electrochemical technique with FPCB enabled rapid detection of the target sequence using a small volume of the sample (5-[Formula: see text]). We achieved a limit-of-detection of 100 fg/mL, whereas the predicted value was ~33 fg/mL, equivalent to approximately [Formula: see text] copies/mL and comparable to sensitivities provided by isothermal nucleic acid amplification tests. We believe that the developed approach proves the ability of an FPCB-implemented DNA sensor to act as a potentially simpler and more affordable diagnostic assay for viral infections in Point-Of-Care (POC) applications.

Proximity labeling approaches to study protein complexes during virus infection.

Cellular compartmentalization of proteins and protein complex formation allow cells to tightly control biological processes. Therefore, understanding the subcellular localization and interactions of a specific protein is crucial to uncover its biological function. The advent of proximity labeling (PL) has reshaped cellular proteomics in infection biology. PL utilizes a genetically modified enzyme that generates a "labeling cloud" by covalently labeling proteins in close proximity to the enzyme. Fusion of a PL enzyme to a specific antibody or a "bait" protein of interest in combination with affinity enrichment mass spectrometry (AE-MS) enables the isolation and identification of the cellular proximity proteome, or proxisome. This powerful methodology has been paramount for the mapping of membrane or membraneless organelles as well as for the understanding of hard-to-purify protein complexes, such as those of transmembrane proteins. Unsurprisingly, more and more infection biology research groups have recognized the potential of PL for the identification of host-pathogen interactions. In this chapter, we introduce the enzymes commonly used for PL labeling as well as recent promising advancements and summarize the major achievements in organelle mapping and nucleic acid PL. Moreover, we comprehensively describe the research on host-pathogen interactions using PL, giving special attention to studies in the field of virology.

Proximity Labeling for the Identification of Coronavirus-Host Protein Interactions.

Biotin-based proximity labeling circumvents major pitfalls of classical biochemical approaches to identify protein-protein interactions. It consists of enzyme-catalyzed biotin tags ubiquitously apposed on proteins located in close proximity of the labeling enzyme, followed by affinity purification and identification of biotinylated proteins by mass spectrometry. Here we outline the methods by which the molecular microenvironment of the coronavirus replicase/transcriptase complex (RTC), i.e., proteins located within a close perimeter of the RTC, can be determined by different proximity labeling approaches using BirAR118G (BioID), TurboID, and APEX2. These factors represent a molecular signature of coronavirus RTCs and likely contribute to the viral life cycle, thereby constituting attractive targets for the development of antiviral intervention strategies.

Mapping the SARS-CoV-2-Host Protein-Protein Interactome by Affinity Purification Mass Spectrometry and Proximity-Dependent Biotin Labeling: A Rational and Straightforward Route to Discover Host-Directed Anti-SARS-CoV-2 Therapeutics.

Protein-protein interactions (PPIs) are the vital engine of cellular machinery. After virus entry in host cells the global organization of the viral life cycle is strongly regulated by the formation of virus-host protein interactions. With the advent of high-throughput -omics platforms, the mirage to obtain a "high resolution" view of virus-host interactions has come true. In fact, the rapidly expanding approaches of mass spectrometry (MS)-based proteomics in the study of PPIs provide efficient tools to identify a significant number of potential drug targets. Generation of PPIs maps by affinity purification-MS and by the more recent proximity labeling-MS may help to uncover cellular processes hijacked and/or altered by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), providing promising therapeutic targets. The possibility to further validate putative key targets from high-confidence interactions between viral bait and host protein through follow-up MS-based multi-omics experiments offers an unprecedented opportunity in the drug discovery pipeline. In particular, drug repurposing, making use of already existing approved drugs directly targeting these identified and validated host interactors, might shorten the time and reduce the costs in comparison to the traditional drug discovery process. This route might be promising for finding effective antiviral therapeutic options providing a turning point in the fight against the coronavirus disease-2019 (COVID-19) outbreak.