SARS-CoV-2 variants show a gradual declining pathogenicity and pro-inflammatory cytokine stimulation and an increasing antigenic and anti-inflammatory cytokine induction (preprint)

Hyper-transmissibility with decreased disease severity are typical characteristics of Omicron variant. To understand this phenomenon, we used various bioinformatics approaches to analyze randomly selected genome sequences (one each) of the Gamma, Delta, and Omicron variants submitted to NCBI from 15 to 31 December 2021. We show that: (i) Pathogenicity of SARS-CoV-2 variants decreases in the order: Wuhan > Gamma > Delta > Omicron; however, the antigenic property follows the order: Omicron > Gamma > Wuhan > Delta. (ii) Omicron Spike RBD has lower pathogenicity but higher antigenicity than that of other variants. (iii) Decreased disease severity by Omicron variant may be due to its decreased pro-inflammatory and IL-6 stimulation and increased IFN-{gamma} and IL-4 induction efficacy. (iv) Mutations in N protein are associated with decreased IL-6 induction and human DDX21-mediated increased IL-4 production in Omicron. (v) Due to mutations, the stability of S, M, N, and E proteins decrease in the order: Omicron > Gamma > Delta > Wuhan. (vi) Stronger Spike-hACE2 binding in Omicron is associated with its increased transmissibility. However, the lowest stability of the Omicron Spike protein makes Spike-hACE2 interaction unstable for systemic infection and for causing severe disease. Finally (vii), the highest instability of Omicron E protein may also be associated with decreased viral maturation and low viral load leading to less severe disease and faster recovery. Our method may be used for other similar viruses, and these findings will contribute to the understanding of the dynamics of SARS-CoV-2 variants and the management of emerging variants.

Potential anti-tumor immune response mechanisms of SARS-CoV-2 in lymphomas: can the devil be converted into a boon? (preprint)

Recently, two cases of complete remission of classical Hodgkin lymphoma (cHL) and follicular lymphoma (FL) after SARS-CoV-2 infection were reported. However, the precise molecular mechanism of this rare event is yet to be understood. Here, we hypothesize a potential anti-tumor immune response of SARS-CoV-2 and based on computational approach show that (i) SARS-CoV-2 Spike-RBD may bind to extracellular domains of CD15, CD27, CD45, and CD152 receptors of cHL or FL, (ii) upon internalization, SARS-CoV-2 membrane (M) protein and Orf3a may bind to gamma-tubulin complex component 3 (GCP3) at its tubulin gamma-1 chain (TUBG1) binding site, (iii) M protein may also interact with TUBG1 blocking its binding to GCP3, (iv) both M and Orf3a may render the GCP2-GCP3 lateral binding where M possibly interacts with GCP2 at its GCP3 binding site and Orf3a to GCP3 at its GCP2 interacting residues, (v) interactions of M and Orf3a with these gamma-tubulin ring complex components potentially block the initial process of microtubule nucleation, leading to cell cycle arrest and apoptosis, (vi) Spike-RBD may also interact with and block PD-1 signaling similar to pembrolizumab and nivolumab like monoclonal antibodies and may induce B-cell apoptosis and remission, (vii) finally, the TRADD interacting PVQLSY motif of Epstein-Barr virus LMP-1, that is responsible for NF-kB mediated oncogenesis, potentially interacts with SARS-CoV-2 Mpro, nsp7, nsp10, and Spike proteins and may regulate the LMP-1 mediated cell proliferation. Taken together, our results suggest a possible therapeutic potential of SARS-CoV-2 in proliferative disorders.

Emergence of Unique SARS-CoV-2 ORF10 Variants and Their Impact on Protein Structure and Function (preprint)

The devastating impact of the ongoing coronavirus disease 2019 (COVID-19) on public health, caused by the Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) has made fighting of the COVID-19 pandemic is a top priority in medical research and pharmaceutical development. Surveillance of SARS-CoV-2 mutations is essential for the comprehension of SARS-CoV-2 variant diversity and their impact on virulence and pathogenicity. The SARS-CoV-2 open reading frame 10 (ORF10) protein interacts with multiple human proteins CUL2, ELOB, ELOC, MAP7D1, PPT1, RBX1, THTPA, TIMM8B, and ZYG11B expressed in the lung tissues. Mutations and co-mutations in the emerging SARS-CoV-2 ORF10 variants are expected to impact the severity of the virus and its associated consequences. In this article, We highlight 128 single mutations and 35 co-mutations in the unique SARS-CoV-2 ORF10 variants in this article. The possible predicted effects of these mutations and co-mutations on the secondary structure of ORF10 variants and host protein interactomes are presented. The findings highlight the possible effects of mutations and co-mutations on the emerging 140 ORF10 unique variants from secondary structure and intrinsic protein disorder perspectives.

Non-Uniform Aspects of SARS-CoV-2 Intraspecies Evolution Reopen Questions on Its Origin (preprint)

Several hypotheses have been presented on the origin of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) from its identification as the agent causing the current coronavirus disease 19 (COVID-19) pandemic. So far, no hypothesis has managed to identify the origin, and the issue has resurfaced. Here we have unfolded a pattern of distribution of several mutations in the SARS-CoV-2 proteins across different continents comprising 24 geo-locations. The results showed an evenly uneven distribution of unique protein variants, distinct mutations, unique frequency of common conserved residues, and mutational residues across the 24 geo-locations. Furthermore, ample mutations were identified in the evolutionarily conserved invariant regions in the SARS-CoV-2 proteins across almost all geo-locations we have considered. This pattern of mutations potentially breaches the law of evolutionary conserved functional units of the beta-coronavirus genus. These mutations may lead to several novel SARS-CoV-2 variants with a high degree of transmissibility and virulence. A thorough investigation on the origin and characteristics of SARS-CoV-2 needs to be conducted in the interest of science and to be prepared to meet the challenges of potential future pandemics.

An Issue of Concern: Unique Truncated ORF8 Protein Variants of SARS-CoV-2 (preprint)

Open reading frame 8 (ORF8) protein is one of the most evolving accessory proteins in severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causative agent of coronavirus disease 2019 (COVID-19). It was previously reported that the ORF8 protein inhibits presentation of viral antigens by the major histocompatibility complex class I (MHC-I) and interacts with host factors involved in pulmonary inflammation. The ORF8 protein assists SARS-CoV-2 to evade immunity and replication. Among many contributing mutations, Q27STOP, a mutation in the ORF8 protein defines the B.1.1.7 lineage of SARS-CoV-2, which is engendering the second wave of COVID-19. In the present study, 47 unique truncated ORF8 proteins (T-ORF8) due to the Q27STOP mutations were identified among 49055 available B.1.1.7 SARS-CoV-2 sequences. The results show that only one of the 47 T-ORF8 variants spread to over 57 geo-locations in North America, and other continents which includes Africa, Asia, Europe and South America. Based on various quantitative features such as amino acid homology, polar/non-polar sequence homology, Shannon entropy conservation, and other physicochemical properties of all specific 47 T-ORF8 protein variants, a collection of nine possible T-ORF8 unique variants were defined. The question of whether T-ORF8 variants work similarly to ORF8 has yet to be investigated. A positive response to the question could exacerbate future COVID-19 waves, necessitating severe containment measures.

Implications Derived from S-Protein Variants of SARS-CoV-2 from Six Continents (preprint)

Spike (S) proteins of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) are critical determinants of the infectivity and antigenicity of the virus. Several mutations in the spike protein of SARS-CoV-2 have already been detected, and their effect in immune system evasion and enhanced transmission as a cause of increased morbidity and mortality are being investigated. From pathogenic and epidemiological perspectives, spike proteins are of prime interest to researchers. This study focused on the unique variants of S proteins from six continents Asia, Africa, Europe, Oceania, South America, and North America. In comparison to the other five continents, Africa (29.065%) had the highest percentage of unique S proteins. Notably, only North America had 87% (14046) of the total (16143) specific S proteins available in the NCBI database(across all continents). Based on the amino acid frequency distributions in the S protein variants from all the continents, the phylogenetic relationship implies that unique S proteins from North America were significantly different from those of the other five continents. Overtime, the unique variants originating from North America are most likely to spread to the other geographic locations through international travel or naturally by emerging mutations. Hence it is suggested that restriction of international travel should be considered, and massive vaccination as an utmost measure to combat the spread of COVID-19 pandemic. It is also further suggested that the efficacy of existing vaccines and future vaccine development must be reviewed with careful scrutiny, and if needed, further re-engineered based on requirements dictated by new emerging S protein variants.

Variability of Accessory Proteins Rules the SARS-CoV-2 Pathogenicity (preprint)

The coronavirus disease 2019 (COVID-19) is caused by the Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) which is pandemic with an estimated fatality rate less than 1% is ongoing. SARS-CoV-2 accessory proteins ORF3a, ORF6, ORF7a, ORF7b, ORF8, and ORF10 with putative functions to manipulate host immune mechanisms such as interferons, immune signaling receptor NLRP3 (NOD-, LRR-, and pyrin domain-containing 3) inflammasome, inflammatory cytokines such as interleukin {beta} (IL-1{beta}) are critical in COVID-19 pathology. Outspread variations of each of the six accessory proteins of all complete proteomes (available as of October 26, 2020, in the National Center for Biotechnology Information depository) of SARS-CoV-2, were observed across six continents. Across all continents, the decreasing order of percentage of unique variations in the accessory proteins was found to be ORF3a>ORF8>ORF7a>ORF6>ORF10>ORF7b. The highest and lowest unique variations of ORF3a were observed in South America and Oceania, respectively. This finding suggests that the wide variations of accessory proteins seem to govern the pathogenicity of SARS-CoV-2, and consequently, certain propositions and recommendations can be made in the public interest.

A zebrafish model for COVID-19 recapitulates olfactory and cardiovascular pathophysiologies caused by SARS-CoV-2 (preprint)

The COVID-19 pandemic has prompted the search for animal models that recapitulate the pathophysiology observed in humans infected with SARS-CoV-2 and allow rapid and high throughput testing of drugs and vaccines. Exposure of larvae to SARS-CoV-2 Spike (S) receptor binding domain (RBD) recombinant protein was sufficient to elevate larval heart rate and treatment with captopril, an ACE inhibitor, reverted this effect. Intranasal administration of SARS-CoV-2 S RBD in adult zebrafish recombinant protein caused severe olfactory and mild renal histopathology. Zebrafish intranasally treated with SARS-CoV-2 S RBD became hyposmic within minutes and completely anosmic by 1 day to a broad-spectrum of odorants including bile acids and food. Single cell RNA-Seq of the adult zebrafish olfactory organ indicated widespread loss of expression of olfactory receptors as well as inflammatory responses in sustentacular, endothelial, and myeloid cell clusters. Exposure of wildtype zebrafish larvae to SARS-CoV-2 in water did not support active viral replication but caused a sustained inhibition of ace2 expression, triggered type 1 cytokine responses and inhibited type 2 cytokine responses. Combined, our results establish adult and larval zebrafish as useful models to investigate pathophysiological effects of SARS-CoV-2 and perform pre-clinical drug testing and validation in an inexpensive, high throughput vertebrate model.

Forecasting COVID-19 cases at the Amazon region: a comparison of classical and machine learning models (preprint)

BACKGROUNDSince the first reports of COVID-19, decision-makers have been using traditional epidemiological models to predict the days to come. However, the enhancement of computational power, the demand for adaptable predictive frameworks, the short past of the disease, and uncertainties related to input data and prediction rules, also make other classical and machine learning techniques viable options. OBJECTIVEThis study investigates the efficiency of six models in forecasting COVID-19 confirmed cases with 17 days ahead. We compare the models autoregressive integrated moving average (ARIMA), Holt-Winters, support vector regression (SVR), k-nearest neighbors regressor (KNN), random trees regressor (RTR), seasonal linear regression with change-points (Prophet), and simple logistic regression (SLR). MATERIAL AND METHODSWe implement the models to data provided by the health surveillance secretary of Amapaa, a Brazilian state fully carved in the Amazon rainforest, which has been experiencing high infection rates. We evaluate the models according to their capacity to forecast in different historical scenarios of the COVID-19 progression, such as exponential increases, sudden decreases, and stability periods of daily cases. To do so, we use a rolling forward splitting approach for out-of-sample validation. We employ the metrics RMSE, R-squared, and sMAPE in evaluating the model in different cross-validation sections. FINDINGSAll models outperform SLG, especially Holt-Winters, that performs satisfactorily in all scenarios. SVR and ARIMA have better performances in isolated scenarios. To implement the comparisons, we have created a web application, which is available online. CONCLUSIONThis work represents an effort to assist the decision-makers of Amapa in future decisions to come, especially under scenarios of sudden variations in the number of confirmed cases of Amapa, which would be caused, for instance, by new contamination waves or vaccination. It is also an attempt to highlight alternative models that could be used in future epidemics.

Possible transmission flow of SARS-CoV-2 based on ACE2 features (preprint)

Angiotensin-converting enzyme 2 (ACE2) is the cellular receptor for the Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) that is engendering the severe coronavirus disease 2019 (COVID-19) pandemic. The spike (S) protein receptor-binding domain (RBD) of SARS-CoV-2 binds to the three sub-domains viz. amino acids (aa) 22-42, aa 79-84, and aa 330-393 of ACE2 on human cells to initiate entry. It was reported earlier that the receptor utilization capacity of ACE2 proteins from different species, such as cats, chimpanzees, dogs, and cattle, are different. A comprehensive analysis of ACE2 receptors of nineteen species was carried out in this study, and the findings propose a possible SARS-CoV-2 transmission flow across these nineteen species.

Notable sequence homology of the ORF10 protein introspects the architecture of SARS-COV-2 (preprint)

The global public health is endangered due to COVID-19 pandemic, which is caused by Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2). Despite having similar pathology to MERS and SARS-CoV, the infection fatality rate of SARS-CoV-2 is likely lower than 1%. SARS-CoV-2 has been reported to be uniquely characterized by the accessory protein ORF10, which contains eleven cytotoxic T lymphocyte (CTL) epitopes of nine amino acids length each, across various human leukocyte antigen (HLA) subtypes. In this study, all missense mutations found in sequence databases were examined across twnety-two unique SARS-CoV-2 ORF10 variants that could possibly alter viral pathogenicity. Some of these mutations decrease the stability of ORF10, e.g. I4L and V6I were found in the MoRF region of ORF10 which may also possibly contribute to Intrinsic protein disorder. Furthermore, a physicochemical and structural comparative analysis was carried out on SARS-CoV-2 and Pangolin-CoV ORF10 proteins, which share 97.37% amino acid homology. The high degree of physicochemical and structural similarity of ORF10 proteins of SARS-CoV-2 and Pangolin-CoV open questions about the architecture of SARS-CoV-2 due to the disagreement of these two ORF10 proteins over their sub-structure (loop/coil region), solubility, antigenicity and change from the strand to coil at amino acid position 26, where tyrosine is present. Altogether, SARS-CoV-2 ORF10 is a promising pharmaceutical target and a protein which should be monitored for changes which correlate to change pathogenesis and clinical course of COVID-19 infection.

A unique view of SARS-CoV-2 through the lens of ORF8 protein (preprint)

Immune evasion is one of the unique characteristics of COVID-19 attributed to the ORF8 protein of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). This protein is involved in modulating the host adaptive immunity through downregulating MHC (Major Histocompatibility Complex) molecules and innate immune responses by surpassing the interferon mediated antiviral response of the host. To understand the immune perspective of the host with respect to the ORF8 protein, a comprehensive study of the ORF8 protein as well as mutations possessed by it, is performed. Chemical and structural properties of ORF8 proteins from different hosts, that is human, bat and pangolin, suggests that the ORF8 of SARS-CoV-2 and Bat RaTG13-CoV are very much closer related than that of Pangolin-CoV. Eighty-seven mutations across unique variants of ORF8 (SARS-CoV-2) are grouped into four classes based on their predicted effects. Based on geolocations and timescale of collection, a possible flow of mutations was built. Furthermore, conclusive flows of amalgamation of mutations were endorsed upon sequence similarity and amino acid conservation phylogenies. Therefore, this study seeks to highlight the uniqueness of rapid evolving SARS-CoV-2 through the ORF8.