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1. Evaporation‐Enhanced Redox Cycling for Rapid Detection of Attomolar SARS‐CoV‐2 Virions Using Nanolithography‐Free Electrochemical Devices

   https://doi.org/10.1002/admt.202401397  

   Soltan_Khamsi, Pouya; Kammarchedu, Vinay; Ebrahimi, Aida (January 2025, Advanced Materials Technologies)

   Abstract In fighting against infectious diseases such as COVID‐19, simple‐to‐use, sensitive, scalable, and rapid diagnostics are crucial for early disease diagnosis. In this regard, electrochemical biosensors are particularly attractive in developing point‐of‐need diagnostics. Importantly, by being compatible with nano‐ and microfabrication methods, they are amenable to miniaturization, which reduces background noise and the required sample volume. However, miniaturization also reduces the signal level, making it challenging to detect low virus counts. In this work, microfabricated electrochemical sensors with a dual signal amplification scheme based on evaporation‐enhanced redox cycling (E2RC) in a generator–collector configuration are developed. A scalable, nanolithography‐free fabrication method is proposed to achieve a controllable sub‐micrometer gap between three dimensional (3D) interdigitated microelectrodes by combining photolithography with template‐driven electrodeposition. Using the optimized electrodes, the sensors achieve rapid detection with a limit of quantification of ≈1.2 × 10³particles mL⁻¹through continuous measurement in evaporating droplets containing SARS‐CoV‐2 virion mimics. Investigating particle charge and size reveals the role of electrophoretic enrichment in the overall response. The sensor performance is also validated using heat‐inactivated SARS‐CoV‐2 virions, with selective response to SARS‐CoV‐2 against HCoV‐299E, SARS‐CoV S1, and MERS‐CoV S1 (captured using antibody‐functionalized magnetic nanoparticles). The proposed sensing method is sensitive, rapid, scalable, and can be extended to broader applications, including detection of bacteria, extracellular vesicles, and other viruses. 

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2. Effect of the SARS-CoV-2 Delta-associated G15U mutation on the s2m element dimerization and its interactions with miR-1307-3p

   https://doi.org/10.1261/rna.079627.123  

   Cunningham, Caylee L.; Frye, Caleb J.; Makowski, Joseph A.; Kensinger, Adam H.; Shine, Morgan; Milback, Ella J.; Lackey, Patrick E.; Evanseck, Jeffrey D.; Mihailescu, Mihaela-Rita (October 2023, RNA)

   The s2m, a highly conserved 41-nt hairpin structure in the SARS-CoV-2 genome, serves as an attractive therapeutic target that may have important roles in the virus life cycle or interactions with the host. However, the conserved s2m in Delta SARS-CoV-2, a previously dominant variant characterized by high infectivity and disease severity, has received relatively less attention than that of the original SARS-CoV-2 virus. The focus of this work is to identify and define the s2m changes between Delta and SARS-CoV-2 and the subsequent impact of those changes upon the s2m dimerization and interactions with the host microRNA miR-1307-3p. Bioinformatics analysis of the GISAID database targeting the s2m element reveals a >99% correlation of a single nucleotide mutation at the 15th position (G15U) in Delta SARS-CoV-2. Based on¹H NMR spectroscopy assignments comparing the imino proton resonance region of s2m and the s2m G15U at 19°C, we show that the U15–A29 base pair closes, resulting in a stabilization of the upper stem without overall secondary structure deviation. Increased stability of the upper stem did not affect the chaperone activity of the viral N protein, as it was still able to convert the kissing dimers formed by s2m G15U into a stable duplex conformation, consistent with the s2m reference. However, we show that the s2m G15U mutation drastically impacts the binding of host miR-1307-3p. These findings demonstrate that the observed G15U mutation alters the secondary structure of s2m with subsequent impact on viral binding of host miR-1307-3p, with potential consequences on immune responses. 

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3. Discovery of SARS‐CoV‐2 Inhibitors Featuring Novel Histidine α ‐Nitrile Motif

   https://doi.org/10.1002/cbdv.202300957  

   Vijay Gone, Nilu; Ghalib Enayathullah, Mohammed; Thomas, Jessie; Rathee, Parth; Prabhakar, Rajeev; Kumar Bokara, Kiran; Sanjayan, Gangadhar J. (November 2023, Chemistry & Biodiversity)

   Abstract As COVID‐19 infection caused severe public health concerns recently, the development of novel antivirals has become the need of the hour. Main protease (M^pro) has been an attractive target for antiviral drugs since it plays a vital role in polyprotein processing and virus maturation. Herein we report the discovery of a novel class of inhibitors against the SARS‐CoV‐2, bearing histidineα‐nitrile motif embedded on a simple dipeptide framework.In‐vitroandin‐silicostudies revealed that the histidineα‐nitrile motif envisioned to target the M^procontributes to the inhibitory activity. Among a series of dipeptides synthesized featuring this novel structural motif, some dipeptides displayed strong viral reduction (EC₅₀=0.48 μM) with a high selectivity index, SI>454.54. These compounds also exhibit strong binding energies in the range of −28.7 to −34.2 Kcal/mol. The simple dipeptide structural framework, amenable to quick structural variations, coupled with ease of synthesis from readily available commercial starting materials are the major attractive features of this novel class of SARS‐CoV‐2 inhibitors. The histidineα‐nitrile dipeptides raise the hope of discovering potent drug candidates based on this motif to fight the dreaded SARS‐CoV‐2. 

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4. A rapid, low-cost, and highly sensitive SARS-CoV-2 diagnostic based on whole-genome sequencing

   https://doi.org/10.1371/journal.pone.0294283  

   Adastra, Per A; Durand, Neva C; Mitra, Namita; Pulido, Saul Godinez; Mahajan, Ragini; Blackburn, Alyssa; Colaric, Zane L; Theisen, Joshua_W M; Weisz, David; Dudchenko, Olga; et al (November 2023, PLOS ONE)

   Giri, Basant (Ed.)

   Early detection of SARS-CoV-2 infection is key to managing the current global pandemic, as evidence shows the virus is most contagious on or before symptom onset. Here, we introduce a low-cost, high-throughput method for diagnosing and studying SARS-CoV-2 infection. Dubbed Pathogen-Oriented Low-Cost Assembly & Re-Sequencing (POLAR), this method amplifies the entirety of the SARS-CoV-2 genome. This contrasts with typical RT-PCR-based diagnostic tests, which amplify only a few loci. To achieve this goal, we combine a SARS-CoV-2 enrichment method developed by the ARTIC Network (https://artic.network/) with short-read DNA sequencing andde novogenome assembly. Using this method, we can reliably (>95% accuracy) detect SARS-CoV-2 at a concentration of 84 genome equivalents per milliliter (GE/mL). The vast majority of diagnostic methods meeting our analytical criteria that are currently authorized for use by the United States Food and Drug Administration with the Coronavirus Disease 2019 (COVID-19) Emergency Use Authorization require higher concentrations of the virus to achieve this degree of sensitivity and specificity. In addition, we can reliably assemble the SARS-CoV-2 genome in the sample, often with no gaps and perfect accuracy given sufficient viral load. The genotypic data in these genome assemblies enable the more effective analysis of disease spread than is possible with an ordinary binary diagnostic. These data can also help identify vaccine and drug targets. Finally, we show that the diagnoses obtained using POLAR of positive and negative clinical nasal mid-turbinate swab samples 100% match those obtained in a clinical diagnostic lab using the Center for Disease Control’s 2019-Novel Coronavirus test. Using POLAR, a single person can manually process 192 samples over an 8-hour experiment at the cost of ~$36 per patient (as of December 7^th, 2022), enabling a 24-hour turnaround with sequencing and data analysis time. We anticipate that further testing and refinement will allow greater sensitivity using this approach. 

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5. Extracellular Vimentin as a Target Against SARS‐CoV‐2 Host Cell Invasion

   https://doi.org/10.1002/smll.202105640  

   Suprewicz, Łukasz; Swoger, Maxx; Gupta, Sarthak; Piktel, Ewelina; Byfield, Fitzroy J.; Iwamoto, Daniel V.; Germann, Danielle; Reszeć, Joanna; Marcińczyk, Natalia; Carroll, Robert J.; et al (December 2021, Small)

   Abstract Infection of human cells by pathogens, including SARS‐CoV‐2, typically proceeds by cell surface binding to a crucial receptor. The primary receptor for SARS‐CoV‐2 is the angiotensin‐converting enzyme 2 (ACE2), yet new studies reveal the importance of additional extracellular co‐receptors that mediate binding and host cell invasion by SARS‐CoV‐2. Vimentin is an intermediate filament protein that is increasingly recognized as being present on the extracellular surface of a subset of cell types, where it can bind to and facilitate pathogens’ cellular uptake. Biophysical and cell infection studies are done to determine whether vimentin might bind SARS‐CoV‐2 and facilitate its uptake. Dynamic light scattering shows that vimentin binds to pseudovirus coated with the SARS‐CoV‐2 spike protein, and antibodies against vimentin block in vitro SARS‐CoV‐2 pseudovirus infection of ACE2‐expressing cells. The results are consistent with a model in which extracellular vimentin acts as a co‐receptor for SARS‐CoV‐2 spike protein with a binding affinity less than that of the spike protein with ACE2. Extracellular vimentin may thus serve as a critical component of the SARS‐CoV‐2 spike protein‐ACE2 complex in mediating SARS‐CoV‐2 cell entry, and vimentin‐targeting agents may yield new therapeutic strategies for preventing and slowing SARS‐CoV‐2 infection. 

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