Circulation of genetically diverse non-polio enteroviruses in respiratory samples during COVID-19 pandemic period (2021-22)

Enteroviruses, beyond poliovirus, are important pathogens. Several non-polio enteroviruses (NPEVs) are causing epidemics all around the world. Limited data is available on the prevalence and diversity of these viruses from India. Objective(s): Detection and characterization of NPEVs in respiratory samples during the COVID-19 pandemic period. Material(s) and Method(s): COVID-19 negative samples from acute respiratory infections (ARI) [n = 105] and severe-acute respiratory infections (SARI) [n = 148] during the period 2021-22 were screened for NPEVs. Detection was carried out using the one step RT-PCR method targeting the 5'UTR region followed by molecular analysis. Results and Conclusion(s): Total positivity of NPEVs was noted in 35.23% and 31.08% of the ARI and SARI cases, respectively. Comparison within the two groups studied, showed significant difference in the age-wise distribution for cases>18 years of age. Year round seasonality for ARI cases while autumn seasonality for SARI cases was observed. Sequencing of representative samples of ARI cases showed prevalence of Rhinovirus A (RVA), Rhinovirus B (RVB), Rhinovirus C (RVC) and Echovirus, while predominance of RVC followed by RVA was observed for the SARI cases. Phylogenetic analysis of all the strains showed clustering of RVC strains in different clusters. Divergence was also noted in RVA and RVB strains studied. Circulation of a rare Echovirus-29 strain was noted in the ARI cases. The study highlighted significant divergence in the Rhinovirus strains studied. It warrants the need for surveillance of NPEVs, whole-genome sequencing of the circulating strains for better understanding of biodiversity among the NPEVs and the potential health burden.

Thermodynamic Analysis of 1 x 2 Internal Loop Regions of SL1 of SARS-CoV-2 and SL of U6 snRNA

The genomic material of SARS-CoV-2 is a positive-sense single-stranded RNA. SARS-CoV-2 produces non-structural protein 1 (NSP1), which inhibits host cell translation by binding its' N-terminal to the host's 40S ribosomal subunit. Once NSP1 is bound its C-terminal domain folds and binds to the mRNA entry channel. Stem loop 1 (SL1) in the 5'-UTR of the viral mRNA binds to NSP1 to abrogate translation inhibition leading to the expression of viral proteins. SL1 contains a 1 x 2 internal loop that is not seen in other coronaviruses and may be involved in conformational changes that influence SL1-NSP1 interactions. The 1 x 2 internal loop of SL1 contains a putative A*C non-canonical base pair. The U6 snRNA also contains a 1 x 2 internal loop known to undergo conformation changes in response to pH and magnesium ion binding. Here we examine the thermodynamic properties and magnesium binding of the 1 x 2 internal loop of SL1 in varying helical contexts. Thermal denaturation experiments were performed on various DNA and RNA constructs in the presence of 1 M KCl or 10 mM magnesium chloride at a pH of 5.5 and 7. We show that formation of the A+*C base pair and the construct stability in the presence of magnesium ions is dependent on the helical context.Copyright © 2023 The American Society for Biochemistry and Molecular Biology, Inc.

Extracellular vesicles as delivery vectors to enhance killing of bacteria and viruses by airway-recruited human neutrophils

Background: Polymorphonuclear neutrophils (PMNs) recruited to the airway lumen in cystic fibrosis (CF) undergo a rapid transcriptional program, resulting in exocytosis of granules and inhibition of bacterial killing. As a result, chronic infection, feed-forward inflammation, and structural tissue damage occur. Because CF airway PMNs are also highly pinocytic, we hypothesized that we could deliver protein- and ribonucleic acid (RNA)-based therapies to modulate their function to benefit patients. We elected to use extracellular vesicles (EVs) as a delivery vector because they are highly customizable, and airway PMNs have previously been shown by our group to process and use their cargo efficiently [1]. Furthermore, our prior work on CF airway PMNs [2] led to identification of the long noncoding RNA MALAT1, the transcription factor Ehf, and the histone deacetylase/long-chain fatty deacylase HDAC11 as potential targets to modulate CF airway PMN dysfunction. Method(s): H441 human club epithelial cells were chosen for EV production because they efficiently communicate with lung-recruited primary human PMNs [1]. Relevant constructs were cloned into an expression plasmid downstream of a constitutive cytomegalovirus or U6 promoter with an additional puromycin selection cassette. EVs were generated in serumdepleted media and purified by differential centrifugation. Quality and concentration of EVs was determined by electron microscopy and nanoparticle tracking analysis and cargo content by western blot (protein) or qualitative reverse transcription polymerase chain reaction (RNA). Enhanced green fluorescent protein and messenger ribonucleic acid (mRNA) were used as controls. To test delivery to primary human PMNs, generated EVs were applied in the apical fluid of an airway transmigration model [2]. PMN activation was assessed by flow cytometry, and bacterial (PA01 and Staphylococcus aureus 8325-4) killing and viral (influenza Avirus [IAV] H1N1/PR/8/34;SARS-CoV-2/Washington) clearance assays were conducted. Result(s): To package protein, we used EV-loading motifs such as the tetraspanin CD63, Basp1 amino acids 1-9, and the palmitoylation signal of Lyn kinase. To load mRNA, a C'D box motif recognized by the RNA-binding protein L7Ae was included in the 3' untranslated region of the expressed RNA, and CD63-L7Ae was co-expressed. Airway-recruited PMNs treated with EVs containing small interfering RNAs against MALAT1 or HDAC11 showed greater ability to clear bacteria. Conversely, PMNs treated with constructs encasing MALAT1 or HDAC11 efficiently cleared IAV and SARSCoV- 2. PMNs expressing Ehf showed greater clearance of bacteria and viruses. Conclusion(s): Our findings suggest mutually exclusive roles of MALAT-1 and HDAC11 in regulating bacterial and viral clearance by airway-recruited PMNs. Expression of Ehf in airway PMNs may be a pathogen-agnostic approach to enhancing clearance by airway-recruited PMNs. Overall, our study brings proof-of-concept data for therapeutic RNA/protein transfer to airway-recruited PMNs in CF and other lung diseases and for use of EVs as a promising method for cargo delivery to these cells. It is our expectation that, by treating the immune compartment of CF airway disease, pathogentherapies, such as antibiotics will be more effective, and epithelial-targeted therapies, such as CFTR modulators, will have greater penetrance into the cell types of interest.Copyright © 2022, European Cystic Fibrosis Society. All rights reserved

Effects of Various Mrna-Lnp Vaccine Doses on Neuroinflammation in Balb/C Mice

It has been proven that mRNA vaccines are highly effective against the COVID-19 outbreak, and low prevalence of side effects has been shown. However, there are still many gaps in our understanding of the biology and biosafety of nucleic acids as components of lipid nanoparticles (LNPs) most often used as a system for inctracellular delivery of mRNA-based vaccines. It is known that LNPs cause severe injection site inflammation, have broad biodistribution profiles, and are found in multiple tissues of the body, including the brain, after administration. The role of new medications with such pharmacokinetics in inflammation developing in inaccessible organs is poorly understood. The study was aimed to assess the effects of various doses of mRNA-LNP expressing the reporter protein (0, 5, 10, and 20 microg of mRNA encoding the firefly luciferase) on the expression of neuroinflammation markers (Tnfalpha, Il1beta, Gfap, Aif1) in the prefrontal cortex and hypothalamus of laboratory animals 4, 8, and 30 h after the intramuscular injection of LNP nanoemulsion. It was shown that mRNA-LNP vaccines in a dose of 10-20 microg of mRNA could enhance Aif1 expression in the hypothalamus 8 h after vaccination, however, no such differences were observed after 30 h. It was found that the Gfap, l11beta, Tnfalpha expression levels in the hypothalamus observed at different times in the experimental groups were different. According to the results, mRNA-LNPs administered by the parenteral route can stimulate temporary activation of microglia in certain time intervals in the dose-dependent and site specific manner.Copyright © 2022 Pirogov Russian National Research Medical University. All rights reserved.

Viral enteritis in cattle: To well known viruses and beyond

Livestock products supply about 13 percent of energy and 28 percent of protein in diets consumed worldwide. Diarrhea is a leading cause of sickness and death of beef and dairy calves in their first month of life and also affecting adult cattle, resulting in large economic losses and a negative impact on animal welfare. Despite the usual multifactorial origin, viruses are generally involved, being among the most important causes of diarrhea. There are several viruses that have been confirmed as etiological agents (i.e., rotavirus and coronavirus), and some viruses that are not yet confirmed as etiological agents. This review summarizes the viruses that have been detected in the enteric tract of cattle and tries to deepen and gather knowledge about them.Copyright © 2021 by the authors. Licensee MDPI, Basel, Switzerland.

Statistical analysis supports UTR (untranslated region) deletion theory in SARS-CoV-2.

It was noticed that the mortality rate of SARS-CoV-2 infection experienced a significant declination in the early stage of the epidemic. We suspect that the sharp deterioration of virus toxicity is related to the deletion of the untranslated region (UTR) of the virus genome. It was found that the genome length of SARS-CoV-2 engaged a significant truncation due to UTR deletion after a mega-sequence analysis. Sequence similarity analysis further indicated that short UTR strains originated from its long UTR ancestors after an irreversible deletion. A good correlation was discovered between genome length and mortality, which demonstrated that the deletion of the virus UTR significantly affected the toxicity of the virus. This correlation was further confirmed in a significance analysis of the genetic influence on the clinical outcomes. The viral genome length of hospitalized patients was significantly more extensive than that of asymptomatic patients. In contrast, the viral genome length of asymptomatic was considerably longer than that of ordinary patients with symptoms. A genome-level mutation scanning was performed to systematically evaluate the influence of mutations at each position on virulence. The results indicated that UTR deletion was the primary driving force in alternating virus virulence in the early evolution. In the end, we proposed a mathematical model to explain why this UTR deletion was not continuous.

miRNA expression in COVID-19.

Coronavirus disease 2019 (COVID-19) is regarded as a challenge in health system. Several studies have assessed the immune-related aspect of this disorder to identify the host-related factors that affect the course of COVID-19. microRNAs (miRNAs) as potent regulators of immune responses have gained much attention in this regard. Recent studies have shown aberrant expression of miRNAs in COVID-19 in association with disease course. Differentially expressed miRNAs have been enriched in pathways related with inflammation and antiviral immune response. miRNAs have also been regarded as potential therapeutic targets in COVID-19, particularly for management of pathological consequences of COVID-19. In the current review, we summarize the data about dysregulation of miRNAs in COVID-19.

Specific Recognition of the 5'-Untranslated Region of West Nile Virus Genome by Human Innate Immune System.

In the last few years, the sudden outbreak of COVID-19 caused by SARS-CoV-2 proved the crucial importance of understanding how emerging viruses work and proliferate, in order to avoid the repetition of such a dramatic sanitary situation with unprecedented social and economic costs. West Nile Virus is a mosquito-borne pathogen that can spread to humans and induce severe neurological problems. This RNA virus caused recent remarkable outbreaks, notably in Europe, highlighting the need to investigate the molecular mechanisms of its infection process in order to design and propose efficient antivirals. Here, we resort to all-atom Molecular Dynamics simulations to characterize the structure of the 5'-untranslated region of the West Nile Virus genome and its specific recognition by the human innate immune system via oligoadenylate synthetase. Our simulations allowed us to map the interaction network between the viral RNA and the host protein, which drives its specific recognition and triggers the host immune response. These results may provide fundamental knowledge that can assist further antivirals' design, including therapeutic RNA strategies.

Mutational cascade of SARS-CoV-2 leading to evolution and emergence of omicron variant.

BACKGROUND: Emergence of new variant of SARS-CoV-2, namely omicron, has posed a global concern because of its high rate of transmissibility and mutations in its genome. Researchers worldwide are trying to understand the evolution and emergence of such variants to understand the mutational cascade events. METHODS: We have considered all omicron genomes (n = 302 genomes) available till 2nd December 2021 in the public repository of GISAID along with representatives of variants of concern (VOC), i.e., alpha, beta, gamma, delta, and omicron; variant of interest (VOI) mu and lambda; and variant under monitoring (VUM). Whole genome-based phylogeny and mutational analysis were performed to understand the evolution of SARS CoV-2 leading to emergence of omicron variant. RESULTS: Whole genome-based phylogeny depicted two phylogroups (PG-I and PG-II) forming variant specific clades except for gamma and VUM GH. Mutational analysis detected 18,261 mutations in the omicron variant, majority of which were non-synonymous mutations in spike (A67, T547K, D614G, H655Y, N679K, P681H, D796Y, N856K, Q954H), followed by RNA dependent RNA polymerase (rdrp) (A1892T, I189V, P314L, K38R, T492I, V57V), ORF6 (M19M) and nucleocapsid protein (RG203KR). CONCLUSION: Delta and omicron have evolutionary diverged into distinct phylogroups and do not share a common ancestry. While, omicron shares common ancestry with VOI lambda and its evolution is mainly derived by the non-synonymous mutations.

Inhibition of SARS-CoV-2 by Targeting Conserved Viral RNA Structures and Sequences.

The ongoing COVID-19/Severe Acute Respiratory Syndrome CoV-2 (SARS-CoV-2) pandemic has become a significant threat to public health and has hugely impacted societies globally. Targeting conserved SARS-CoV-2 RNA structures and sequences essential for viral genome translation is a novel approach to inhibit viral infection and progression. This new pharmacological modality compasses two classes of RNA-targeting molecules: 1) synthetic small molecules that recognize secondary or tertiary RNA structures and 2) antisense oligonucleotides (ASOs) that recognize the RNA primary sequence. These molecules can also serve as a "bait" fragment in RNA degrading chimeras to eliminate the viral RNA genome. This new type of chimeric RNA degrader is recently named ribonuclease targeting chimera or RIBOTAC. This review paper summarizes the sequence conservation in SARS-CoV-2 and the current development of RNA-targeting molecules to combat this virus. These RNA-binding molecules will also serve as an emerging class of antiviral drug candidates that might pivot to address future viral outbreaks.

The dynamic nature of the coronavirus receptor, angiotensin-converting enzyme 2 (ACE2) in differentiating airway epithelia.

Once inhaled, SARS-CoV-2 particles enter respiratory ciliated cells by interacting with angiotensin converting enzyme 2 (ACE2). Understanding the nature of ACE2 within airway tissue has become a recent focus particularly in light of the COVID-19 pandemic. Airway mucociliary tissue was generated in-vitro using primary human nasal epithelial cells and the air-liquid interface (ALI) model of differentiation. Using ALI tissue, three distinct transcript variants of ACE2 were identified. One transcript encodes the documented full-length ACE2 protein. The other two transcripts are unique truncated isoforms, that until recently had only been predicted to exist via sequence analysis software. Quantitative PCR revealed that all three transcript variants are expressed throughout differentiation of airway mucociliary epithelia. Immunofluorescence analysis of individual ACE2 protein isoforms exogenously expressed in cell-lines revealed similar abilities to localize in the plasma membrane and interact with the SARS CoV 2 spike receptor binding domain. Immunohistochemistry on differentiated ALI tissue using antibodies to either the N-term or C-term of ACE2 revealed both overlapping and distinct signals in cells, most notably only the ACE2 C-term antibody displayed plasma-membrane localization. We also demonstrate that ACE2 protein shedding is different in ALI Tissue compared to ACE2-transfected cell lines, and that ACE2 is released from both the apical and basal surfaces of ALI tissue. Together, our data highlights various facets of ACE2 transcripts and protein in airway mucociliary tissue that may represent variables which impact an individual's susceptibility to SARS-CoV-2 infection, or the severity of Covid-19.

Endothelial exosomal mir-24 predicts cerebrovascular events in covid-19 patients

Introduction: Neuropilin-1 has been recently identified as a co-factor needed for the entry of SARSCoV-2 in host cells and has been linked to neurologic symptoms of COVID-19 (Science 2020). Emerging evidence indicates that exosomal microRNAs (miRNAs) are involved in a number of physiologic and pathologic processes. However, to our knowledge, exosomal miRNAs have not been hitherto investigated in COVID-19. Hypothesis: Since we have recently demonstrated that miR-24 targets the 3'UTR of the gene encoding for Neuropilin-1 and this miRNA is expressed in human brain endothelial cells, we hypothesized an association between plasma levels of CD31 extracellular vesicles (EVs) enriched in miR-24 and the risk of cerebrovascular manifestations in patients hospitalized for COVID-19. Methods and Results: We obtained plasma from >300 COVID-19 patients;as control COVID-19 negative populations, we obtained plasma from healthy donors and patients hospitalized for cerebrovascular disorders. CD31 EVs were isolated from plasma on hospital admission, and miR24 levels were quantified. When comparing patients with vs without cerebrovascular disorders, we found that plasma levels of CD31 EV miR-24 were significantly different between these populations. We did not find any significant difference among groups when assessing circulating free levels of miR-24. Using a multiple regression analysis, adjusting for age, hypertension, and diabetes, the association between EV miR-24 and cerebrovascular complications in COVID-19 patients was confirmed (P<0.05). Conclusions: This is the first study showing a significant association between EV non-coding RNAs and clinical outcome in COVID-19 patients. Our results are relevant for basic researchers, because we identified an unprecedented significant association between EV miR-24 and cerebrovascular disorders, which could be helpful to better understand the molecular mechanisms underlying the pathophysiology of cerebrovascular events in COVID-19, as well as for clinicians, inasmuch as this association may help healthcare professionals in identifying COVID-19 patients who are at high risk of developing cerebrovascular disease.

RNAi-mediated siRNA sequences to combat the COVID-19 pandemic with the inhibition of SARS-CoV2.

The outbreak of the COVID-19 pandemic has cost five million lives to date, and was caused by a positive-sense RNA virus named SARS-CoV2. The lack of drugs specific to SARS-CoV2, leads us to search for an effective and specific therapeutic approach. Small interfering RNA (siRNA) is able to activate the RNA interference (RNAi) pathway to silence the specific targeted gene and inhibit the viral replication, and it has not yet attracted enough attention as a SARS-CoV2 antiviral agent. It could be a potential weapon to combat this pandemic until the completion of full scale, effective mass vaccination. For this study, specific siRNAs were designed using a web-based bioinformatics tool (siDirect2.0) against 14 target sequences. These might have a high probability of silencing the essential proteins of SARS-CoV2. such as: 3CLpro/Mpro/nsp5, nsp7, Rd-Rp/nsp12, ZD, NTPase/HEL or nsp13, PLpro/nsp3, envelope protein (E), spike glycoprotein (S), nucleocapsid phosphoprotein (N), membrane glycoprotein (M), ORF8, ORF3a, nsp2, and its respective 5' and 3'-UTR. Among these potential drug targets, the majority of them contain highly conserved sequences; the rest are chosen on the basis of their role in viral replication and survival. The traditional vaccine development technology using SARS-CoV2 protein takes 6-8 months; meanwhile the virus undergoes several mutations in the candidate protein chosen for vaccine development. By the time the protein-based vaccine reaches the market, the virus would have undergone several mutations, such that the antibodies against the viral sequence may not be effective in restricting the newly mutated viruses. However, siRNA technology can make sequences based on real time viral mutation status. This has the potential for suppressing SARS-CoV2 viral replication, through RNAi technology.

All hands on deck: SARS-CoV-2 proteins that block early anti-viral interferon responses.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection is responsible for the current pandemic coronavirus disease of 2019 (COVID-19). Like other pathogens, SARS-CoV-2 infection can elicit production of the type I and III interferon (IFN) cytokines by the innate immune response. A rapid and robust type I and III IFN response can curb viral replication and improve clinical outcomes of SARS-CoV-2 infection. To effectively replicate in the host, SARS-CoV-2 has evolved mechanisms for evasion of this innate immune response, which could also modulate COVID-19 pathogenesis. In this review, we discuss studies that have reported the identification and characterization of SARS-CoV-2 proteins that inhibit type I IFNs. We focus especially on the mechanisms of nsp1 and ORF6, which are the two most potent and best studied SARS-CoV-2 type I IFN inhibitors. We also discuss naturally occurring mutations in these SARS-CoV-2 IFN antagonists and the impact of these mutations in vitro and on clinical presentation. As SARS-CoV-2 continues to spread and evolve, researchers will have the opportunity to study natural mutations in IFN antagonists and assess their role in disease. Additional studies that look more closely at previously identified antagonists and newly arising mutants may inform future therapeutic interventions for COVID-19.

Emergence, evolution, and vaccine production approaches of SARS-CoV-2 virus: Benefits of getting vaccinated and common questions.

The emergence of coronavirus disease 2019 (COVID-19) pandemic in Wuhan city, China at the end of 2019 made it urgent to identify the origin of the causal pathogen and its molecular evolution, to appropriately design an effective vaccine. This study analyzes the evolutionary background of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2 or SARS-2) in accordance with its close relative SARS-CoV (SARS-1), which was emerged in 2002. A comparative genomic and proteomic study was conducted on SARS-2, SARS-1, and Middle East respiratory syndrome coronavirus (MERS), which was emerged in 2012. In silico analysis inferred the genetic variability among the tested viruses. The SARS-1 genome harbored 11 genes encoding 12 proteins, while SARS-2 genome contained only 10 genes encoding for 10 proteins. MERS genome contained 11 genes encoding 11 proteins. The analysis also revealed a slight variation in the whole genome size of SARS-2 comparing to its siblings resulting from sequential insertions and deletions (indels) throughout the viral genome particularly ORF1AB, spike, ORF10 and ORF8. The effective indels were observed in the gene encoding the spike protein that is responsible for viral attachment to the angiotensin-converting enzyme 2 (ACE2) cell receptor and initiating infection. These indels are responsible for the newly emerging COVID-19 variants αCoV, ßCoV, γCoV and δCoV. Nowadays, few effective COVID-19 vaccines developed based on spike (S) glycoprotein were approved and become available worldwide. Currently available vaccines can relatively prevent the spread of COVID-19 and suppress the disease. The traditional (killed or attenuated virus vaccine and antibody-based vaccine) and innovated vaccine production technologies (RNA- and DNA-based vaccines and viral vectors) are summarized in this review. We finally highlight the most common questions related to COVID-19 disease and the benefits of getting vaccinated.

1H, 13C, and 15N backbone chemical-shift assignments of SARS-CoV-2 non-structural protein 1 (leader protein).

The current COVID-19 pandemic caused by the Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) has become a worldwide health crisis, necessitating coordinated scientific research and urgent identification of new drug targets for treatment of COVID-19 lung disease. The covid19-nmr consortium seeks to support drug development by providing publicly accessible NMR data on the viral RNA elements and proteins. The SARS-CoV-2 genome comprises a single RNA of about 30 kb in length, in which 14 open reading frames (ORFs) have been annotated, and encodes approximately 30 proteins. The first two-thirds of the SARS-CoV-2 genome is made up of two large overlapping open-reading-frames (ORF1a and ORF1b) encoding a replicase polyprotein, which is subsequently cleaved to yield 16 so-called non-structural proteins. The non-structural protein 1 (Nsp1), which is considered to be a major virulence factor, suppresses host immune functions by associating with host ribosomal complexes at the very end of its C-terminus. Furthermore, Nsp1 facilitates initiation of viral RNA translation via an interaction of its N-terminal domain with the 5' untranslated region (UTR) of the viral RNA. Here, we report the near-complete backbone chemical-shift assignments of full-length SARS-CoV-2 Nsp1 (19.8 kDa), which reveal the domain organization, secondary structure and backbone dynamics of Nsp1, and which will be of value to further NMR-based investigations of both the biochemical and physiological functions of Nsp1.

In search of SARS CoV-2 replication inhibitors: Virtual screening, molecular dynamics simulations and ADMET analysis.

Severe acute respiratory syndrome has relapsed recently as novel coronavirus causing a life threat to the entire world in the absence of an effective therapy. To hamper the replication of the deadly SARS CoV-2 inside the host cells, systematic in silico virtual screening of total 267,324 ligands from Asinex EliteSynergy and BioDesign libraries has been performed using AutoDock Vina against RdRp. The molecular modeling studies revealed the identification of twenty-one macrocyclic hits (2-22) with better binding energy than remdesivir (1), marketed SARS CoV-2 inhibitor. Further, the analysis using rules for drug-likeness and their ADMET profile revealed the candidature of these hits due to superior oral bioavailability and druggability. Further, the MD simulation studies of top two hits (2 and 3) performed using GROMACS 2020.1 for 10 ns revealed their stability into the docked complexes. These results provide an important breakthrough in the design of macrocyclic hits as SARS CoV-2 RNA replicase inhibitor.

Computational prediction of potential siRNA and human miRNA sequences to silence orf1ab associated genes for future therapeutics against SARS-CoV-2.

The coronavirus disease 2019 (COVID-19) is an ongoing pandemic caused by an RNA virus termed as severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2). SARS-CoV-2 possesses an almost 30kbp long genome. The genome contains open-reading frame 1ab (ORF1ab) gene, the largest one of SARS-CoV-2, encoding polyprotein PP1ab and PP1a responsible for viral transcription and replication. Several vaccines have already been approved by the respective authorities over the world to develop herd immunity among the population. In consonance with this effort, RNA interference (RNAi) technology holds the possibility to strengthen the fight against this virus. Here, we have implemented a computational approach to predict potential short interfering RNAs including small interfering RNAs (siRNAs) and microRNAs (miRNAs), which are presumed to be intrinsically active against SARS-CoV-2. In doing so, we have screened miRNA library and siRNA library targeting the ORF1ab gene. We predicted the potential miRNA and siRNA candidate molecules utilizing an array of bioinformatic tools. By extending the analysis, out of 24 potential pre-miRNA hairpins and 131 siRNAs, 12 human miRNA and 10 siRNA molecules were sorted as potential therapeutic agents against SARS-CoV-2 based on their GC content, melting temperature (Tm), heat capacity (Cp), hybridization and minimal free energy (MFE) of hybridization. This computational study is focused on lessening the extensive time and labor needed in conventional trial and error based wet lab methods and it has the potential to act as a decent base for future researchers to develop a successful RNAi therapeutic.

A single nucleotide distinguishes the SARS-CoV-2 in the Wuhan outbreak in December 2019 from that in Beijing-Xinfadi in June 2020, China.

Two major locally transmitted outbreaks of coronavirus disease 2019 occurred in China, one in Wuhan from December 2019 to April 2020, another in Beijing-Xinfadi in June 2020. Severe acute respiratory syndrome coronavirus 2 isolated from these two outbreaks can be distinguished by a conserved pyrimidine nucleotide located at nucleotide position 241 in the 5'-untranslated region of the virus genome.