Neutrophil Infiltration Induces Myocardial Injury In COVID-19 Post-Mortem Cases (preprint)

Objectives: The pathological features of severe cardiac injury induced by COVID-19 and relevant clinical features is unknown.Methods: This autopsy cohort study, including hearts from 26 deceased patients hospitalized in intensive care unit due to COVID-19, was conducted at four sites in Wuhan, China. Cases were divided into neutrophil-infiltration group and no-neutrophil group according to histopathological identification of neutrophilic infiltrates or not.Results: Among 26 cases, four cases had active myocarditis with histopathological examination. All cases with myocarditis accompanied with extensive neutrophil infiltration, while cases without myocarditis did not. Detection rates of interleukin-6 (100% vs 4.6%) and tumor necrosis factor-a (100% vs 31.8%) in neutrophil-infiltration group were significantly higher compared to no-neutrophil group (p<0.05 for both). At admission, patients with neutrophil infiltration in myocardium had significantly higher baseline values of aspartate aminotransferase, D dimer and high-sensitivity C reactive protein compared to other 22 patients (p<0.05 for all). During hospitalization, patients with neutrophil infiltration had a significantly higher maximum of creatine kinase (CK)-MB (median 280.0 vs 38.7IU/L, p=0.04), and a quantitatively higher top Troponin I (median 1.112 vs 0.220ng/ml, p=0.56) than patients without neutrophil infiltration. Conclusions: In hearts from deceased patients with severe COVID-19, active myocarditis was commonly infiltrated with neutrophils. Cases with neutrophil-infiltrated myocarditis had a series of severe abnormal laboratory tests at admission, and a high maximum of CK-MB during hospitalization. Role of neutrophil on severe heart injury and even systemic condition in COVID-19 should be emphasized.

Neutrophil Infiltration Induces Myocardial Injury in COVID-19 Post-Mortem Cases (preprint)

Background: The pathological features of severe cardiac injury induced by COVID-19 and relevant clinical features is unknown.Methods: This autopsy cohort study, including hearts from 26 deceased patients hospitalized in intensive care unit due to COVID-19, was conducted at four sites in Wuhan, China. Cases were divided into neutrophil-infiltration group and no-neutrophil group according to histopathological identification of neutrophilic infiltrates or not.Findings: Among 26 cases, four cases had active myocarditis with histopathological examination. All cases with myocarditis accompanied with extensive neutrophil infiltration, while cases without myocarditis did not. Detection rates of interleukin-6 (100% vs 4.6%) and tumor necrosis factor-α (100% vs 31.8%) in neutrophil-infiltration group were significantly higher compared to no-neutrophil group (p<0.05 for both). At admission, patients with neutrophil infiltration in myocardium had significantly higher baseline values of aspartate aminotransferase, D dimer and high-sensitivity C reactive protein compared to other 22 patients (p<0.05 for all). During hospitalization, patients with neutrophil infiltration had a significantly higher maximum of creatine kinase (CK)-MB (median 280.0 vs 38.7IU/L, p=0.04), and a quantitatively higher top Troponin I (median 1.112 vs 0.220ng/ml, p=0.56) than patients without neutrophil infiltration.Interpretation: In hearts from deceased patients with severe COVID-19 , active myocarditis was commonly infiltrated with neutrophils. Cases with neutrophil-infiltrated myocarditis had a series of severe abnormal laboratory tests at admission, and a high maximum of CK-MB during hospitalization. Role of neutrophil on severe heart injury and even systemic condition in COVID-19 should be emphasized.Funding Information: : Emergency Key Program of Guangzhou Laboratory, Grant No. EKPG21-32. Declaration of Interests: None exist.Ethics Approval Statement: Full autopsy was performed after patient death with the approval of the ethics committees and written consent of patient relatives in accordance with regulations issued by the National Health Commission of China and the Helsinki Declaration.

Structural basis for repurpose and design of nucleoside drugs for treating COVID-19 (preprint)

SARS-CoV-2 has caused a global pandemic of COVID-19 that urgently needs an effective treatment. Nucleoside analog drugs including favipiravir have been repurposed for COVID-19 despite of unclear mechanism of their inhibition of the viral RNA polymerase (RdRp). Here we report the cryo-EM structures of the viral RdRp in complex with favipiravir and two other nucleoside inhibitor drugs ribavirin and penciclovir. Ribavirin and the ribosylated form of favipiravir share a similar ribose scaffold that is distinct from penciclovir. However, the structures reveal that all three inhibitors are covalently linked to the primer strand in a monophosphate form despite the different chemical scaffolds between favipiravir and penciclovir. Surprisingly, the base moieties of these inhibitors can form mismatched pairs with the template strand. Moreover, in view of the clinical disadvantages of remdesivir mainly associated with its prodrug form, we designed several orally-available remdesivir parent nucleoside derivatives, including VV16 that showed 5-fold more potent than remdesivir in inhibition of viral replication. Together, these results demonstrate an unexpected promiscuity of the viral RNA polymerase and provide a basis for repurpose and design of nucleotide analog drugs for COVID-19.

The effect of heat on SARS-CoV-2 viability and RNA integrity as determined by plaque assay, virus culture and real-time RT-PCR (preprint)

The effect of heat on SARS-CoV-2/England/2/2020 viability was assessed by plaque assay and virus culture. Heating to 56{degrees}C and 60{degrees}C for 15, 30 and 60 minutes led to a reduction in titre of between 2.1 and 4.9 log 10 pfu/ml but complete inactivation was not observed. At 80{degrees}C plaques were observed after 15 and 30 minutes of heating, however after 60 minutes viable virus was only detected following virus culture. Heating to 80{degrees}C for 90 minutes and 95{degrees}C for 1 and 5 minutes resulted in no viable virus being detected. At 56{degrees}C and 60{degrees}C significant variability between replicates was observed and the titre often increased with heat-treatment time. Nucleic acids were extracted and tested by RT-PCR. Sensitivity of the RT-PCR was not compromised by heating to 56{degrees}C and 60{degrees}C. Heating to 80{degrees}C for 30 minutes or more and 95{degrees}C for 1 or 5 minutes however, resulted in an increase of at least three Ct values. This increase remained constant when different dilutions of virus underwent heat treatment. This indicates that high temperature heat inactivation of clinical samples prior to nucleic acid extraction could significantly affect the ability to detect virus in clinical samples from patients with lower viral loads by RT-PCR.

Photocatalyst under visible light irradiation inactivates SARS-CoV-2 on an abiotic surface (preprint)

There is a worldwide attempt to develop prevention strategies against SARS-CoV-2 transmission. Here we examined the effectiveness of visible light-responsive photocatalyst RENECAT on the inactivation of SARS-CoV-2 under different temperatures and exposure durations. The viral activation on the photocatalyst-coated glass slides decreased from 5.93{mp}0.38 logTCID50/ml to 3.05{mp} 0.25 logTCID50/ml after exposure to visible light irradiation for 6h at 20 degree C. On the other hand, lighting without the photocatalyst, or the photocatalyst-coat without lighting retained viral stability. Immunoblotting and electron microscopic analyses showed the reduced amounts of spike protein on the viral surface after the photocatalyst treatment. Our data suggest a possible implication of the photocatalyst on the decontamination of the SARS-CoV-2 in indoor environments, thereby preventing indirect viral spread.

Genetic diversity analysis of the D614G mutation in SARS-CoV-2 (preprint)

In this work, we evaluated the levels of genetic diversity in 18 genomes of SARS-CoV-2 carrying the D614G mutation, coming from Malaysia and Venezuela and publicly available at the National Center of Biotechnology and Information (NCBI). These haplotypes were previously used for phylogenetic analysis, following the LaBECom protocols. All gaps and unconserved sites were extracted for the construction of a phylogenetic tree. As specific methodologies for paired FST estimators, Molecular Variance (AMOVA), Genetic Distance, mismatch, demographic and spatial expansion analyses, molecular diversity and evolutionary divergence time analyses, 20,000 random permutations were always used. The results revealed the presence of only 57 sites of polymorphic and parsimonium-informative among the 29,827bp analyzed and the analyses based on FST values confirmed the presence of two distinct genetic entities with fixation index of 22% and with a higher component of population variation (78.14%). Tau variations revealed a significant time of divergence, supported by mismatch analysis of the observed distribution ({tau} = 42%). It is safe to say that the small number of existing polymorphisms should not reflect major changes in the protein products of viral populations in both countries and this consideration provides the safety that, although there are differences in the haplotypes studied, these differences are minimal for both regions analyzed geographically and, therefore, it seems safe to extrapolate the levels of polymorphism and molecular diversity found in the samples for other mutant genomes of SARS-CoV-2 in other countries. This reduces speculation about the possibility of large differences between mutant strains of SARS-CoV-2 (D614G) and wild strains, at least at the level of their protein products, although the mutant form has higher transmission speed and infection. The analyses suggest that possible variations in protein products, of the wild virus in relation to its mutant form, should be minimal, bringing peace of mind as to the increased risk of death from the new form of the virus, as well as possible problems of gradual adjustments in some molecular targets for vaccines.

Temporal patterns in the evolutionary genetic distance of SARS-CoV-2 during the COVID-19 pandemic (preprint)

Background: During the pandemic of coronavirus disease 2019 (COVID-19), the genetic mutations occurred in severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) cumulatively or sporadically. In this study, we employed a computational approach to identify and trace the emerging patterns of the SARS-CoV-2 mutations, and quantify accumulative genetic distance across different periods and proteins. Methods: Full-length human SARS-CoV-2 strains in United Kingdom were collected. We investigated the temporal variation in the evolutionary genetic distance defined by the Hamming distance since the start of COVID-19 pandemic. Findings: Our results showed that the SARS-CoV-2 was in the process of continuous evolution, mainly involved in spike protein (S protein), the RNA-dependent RNA polymerase (RdRp) region of open reading frame 1 (ORF1) and nucleocapsid protein (N protein). By contrast, mutations in other proteins were sporadic and genetic distance to the initial sequenced strain did not show an increasing trend.

D614G substitution enhances the stability of trimeric SARS-CoV-2 spike protein (preprint)

SARS-CoV-2 spike protein with D614G substitution has become the dominant variant in the ongoing COVID-19 pandemic. Several studies to characterize the new virus expressing G614 variant show that it exhibits increased infectivity compared to the ancestral virus having D614 spike protein. Here, using in-silico mutagenesis and energy calculations, we analyzed inter-residue interaction energies and thermodynamic stability of the dominant (G614) and the ancestral (D614) variants of spike protein trimer in closed and partially open conformations. We find that the local interactions mediated by aspartate at the 614th position are energetically frustrated and create unfavourable environment. Whereas, glycine at the same position confers energetically favourable environment and strengthens intra- as well as inter-protomer association. Such changes in the local interaction energies enhance the thermodynamic stability of the spike protein trimer as free energy difference ({Delta}{Delta}G) upon glycine substitution is -2.6 kcal/mol for closed conformation and -2.0 kcal/mol for open conformation. Our results on the structural and energetic basis of enhanced stability hint that G614 may confer increased availability of functional form of spike protein trimer and consequent in higher infectivity than the D614 variant.

Structural basis for repurposing a 100-years-old drug suramin for treating COVID-19 (preprint)

The COVID-19 pandemic by non-stop infections of SARS-CoV-2 has continued to ravage many countries worldwide. Here we report the discovery of suramin, a 100-year-old drug, as a potent inhibitor of the SARS-CoV-2 RNA dependent RNA polymerase (RdRp) through blocking the binding of RNA to the enzyme. In biochemical assays, suramin and its derivatives are at least 20-fold more potent than remdesivir, the currently approved nucleotide drug for COVID-19. The 2.6 Å cryo-EM structure of the viral RdRp bound to suramin reveals two binding sites of suramin, with one site directly blocking the binding of the RNA template strand and the other site clash with the RNA primer strand near the RdRp catalytic active site. Furthermore, suramin potently inhibits SARS-CoV-2 duplication in Vero E6 cells. These results provide a structural mechanism for the first non-nucleotide inhibitor of the SARS-CoV-2 RdRp and a rationale for repurposing suramin for treating COVID-19.

IP-10 and MCP-1 as biomarkers associated with disease severity of COVID-19 (preprint)

Background: COVID-19 is a viral respiratory disease caused by the severe acute respiratory syndrome-Coronavirus type 2 (SARS-CoV-2). Patients with this disease may be more prone to venous or arterial thrombosis because of the activation of many factors involved in it, including inflammation, platelet activation and endothelial dysfunction. Interferon gamma inducible protein-10 (IP-10), monocyte chemoattractant protein-1 (MCP-1) and macrophage inflammatory protein 1-alpha (MIP1α) are cytokines related to thrombosis. Therefore, this study focused on these three indicators in COVID-19, with the hope to find biomarkers that are associated with patients’ outcome. Methods: This is a retrospective single-center study involving 74 severe and critically ill COVID-19 patients recruited from the ICU department of the Tongji Hospital in Wuhan, China. The patients were divided into two groups: severe patients and critically ill patients. The serum IP-10, MCP-1 and MIP1α level in both groups was detected using the enzyme-linked immunosorbent assay (ELISA) kit. The clinical symptoms, laboratory test results, and the outcome of COVID-19 patients were retrospectively analyzed. Results: The serum IP-10 and MCP-1 level in critically ill patients was significantly higher than that in severe patients ( P <0.001). However, no statistical difference in MIP1α between the two groups was found. The analysis of dynamic changes showed that these indicators remarkably increased in patients with poor prognosis. Since the selected patients were severe or critically ill, no significant difference was observed between survival and death. Conclusions: IP-10 and MCP-1 are biomarkers associated with the severity of COVID-19 disease and can be related to the risk of death in COVID-19 patients.

Structural basis for repurposing a 100-years-old drug suramin for treating COVID-19 (preprint)

The COVID-19 pandemic by non-stop infections of SARS-CoV-2 has continued to ravage many countries worldwide. Here we report the discovery of suramin, a 100-year-old drug, as a potent inhibitor of the SARS-CoV-2 RNA dependent RNA polymerase (RdRp) through blocking the binding of RNA to the enzyme. In biochemical assays, suramin and its derivatives are at least 20-fold more potent than remdesivir, the currently approved nucleotide drug for COVID-19. The 2.6 [A] cryo-EM structure of the viral RdRp bound to suramin reveals two binding sites of suramin, with one site directly blocking the binding of the RNA template strand and the other site clash with the RNA primer strand near the RdRp catalytic active site, therefore inhibiting the viral RNA replication. Furthermore, suramin potently inhibits SARS-CoV-2 duplication in Vero E6 cells. These results provide a structural mechanism for the first non-nucleotide inhibitor of the SARS-CoV-2 RdRp and a rationale for repurposing suramin for treating COVID-19.

Structure Basis for Inhibition of SARS-CoV-2 by the Feline Drug GC376 (preprint)

The pandemic of SARS-CoV-2 coronavirus disease-2019 (COVID-19) caused by SARS-COV-2 continues to ravage many countries in the world. Mpro is an indispensable protein for viral translation in SARS-CoV-2 and a potential target in high-specificity anti-SARS-CoV-2 drug screening. In this study, to explore potential drugs for treating COVID-19, we elucidated the structure of SARS-CoV-2 Mpro and explored the interaction between Mpro and GC376, an antiviral drug used to treat a range of coronaviruses in Feline via inhibiting Mpro. The availability and safety of GC376 were proved by biochemical and cell experiments in vitro. We determined the structure of an important protein, Mpro, in SARS-CoV-2, and revealed the interaction of GC376 with the viral substrate and inhibition of the catalytic site of SARS-CoV-2 Mpro.

Clinical characteristics and survival analysis in critical and non-critical patients with COVID-19 in Wuhan, China: a single-center retrospective case control study (preprint)

Since the outbreak of COVID-19 in China at the end of 2019, the world has experienced a large-scale epidemic caused by the SARS-CoV-2. Epidemiological and clinical course of COVID-19 patients have been reported, but there have been few analyses about the characteristics, predictive risk factors and outcomes of critical patients. In this single-center retrospective case-control study, 90 adult inpatients hospitalized at Tongji Hospital (Wuhan, China) were included. Demographic, clinical, laboratory test and treatment data were obtained and compared between critical and non-critical patients. We found that compared with non-critical patients, the critical patients had higher SOFA score and qSOFA scores. Critical patients had lower lymphocyte and platelet count, elevated D-dimer, decreased fibrinogen, and elevated high-sensitivity C-reactive protein (hsCRP) and interleukin-6(IL-6). More critical patients received treatment including antibiotics, anticoagulation, corticosteroid and oxygen therapy than non-critical ones. Multivariable regression showed higher qSOFA score and elevation of IL-6 were related to critical patients. Antibiotic usage and anticoagulation were associated with decreased in-hospital mortality. And critical grouping contributed greatly to in-hospital death. Critical COVID-19 patients have a more severe clinical cours. qSOFA score and elevation of IL-6 are risk factors for critical condition. Non-critical grouping, positive antibiotic application and anticoagulation may be beneficial for patient survival.

The performance of Chest CT and Its Imaging Features for Diagnosing Coronavirus Disease 2019: a Meta-analysis (preprint)

Early diagnosis and isolation of cases are particularly crucial for coronavirus disease 2019 (COVID-19) in global pandemic. The aim of this study is to determine the diagnostic performance of chest computed tomography (CT) and imaging features for diagnosing COVID-19. Diagnostic accuracy studies of CT and RT-PCR in patients with clinically suspected COVID-19, which were published up to April 25th, 2020 from MEDLINE, EMBASE, and the Cochrane Library. Twelve studies (n=2,204) were included. The pooled sensitivity, specificity, likelihood ratio (PLR), negative likelihood ratio (NLR), and diagnostic odds ratio (DOR) of chest CT for detecting COVID-19 were 94.5% (95% confidence interval (CI) 89.5 to 97.2%) and 41.8% (95% CI 24.2 to 61.6%), 1.6 (95% CI: 1.6-2.3), 0.13 (95% CI: 0.06-0.31), and 12.4 (95% CI: 4.0-38.5), respectively. Initial RT-PCR revealed a better diagnostic performance. Peripheral lesions, bilateral involvement, multiple lesions, and ground-glass opacities (GGO), revealed to be with better diagnostic value than other CT manifestations. Using chest CT for COVID-19 diagnosis has a high sensitivity and a relatively low specificity. Bilateral multiple peripheral lesions and GGO revealed to be with better diagnostic value. For areas with high prevalence, chest CT could be a good screening test to preliminary screen patients with COVID-19 quickly.

The Role of the Hercules Autonomous Vehicle During the COVID-19 Pandemic: An Autonomous Logistic Vehicle for Contactless Goods Transportation (preprint)

Since early 2020, the coronavirus disease 2019 (COVID-19) has spread rapidly across the world. As at the date of writing this article, the disease has been globally reported in 223 countries and regions, infected over 108 million people and caused over 2.4 million deaths (https://covid19.who.int/, accessed on Feb. 17, 2021). Avoiding person-to-person transmission is an effective approach to control and prevent the pandemic. However, many daily activities, such as transporting goods in our daily life, inevitably involve person-to-person contact. Using an autonomous logistic vehicle to achieve contact-less goods transportation could alleviate this issue. For example, it can reduce the risk of virus transmission between the driver and customers. Moreover, many countries have imposed tough lockdown measures to reduce the virus transmission (e.g., retail, catering) during the pandemic, which causes inconveniences for human daily life. Autonomous vehicle can deliver the goods bought by humans, so that humans can get the goods without going out. These demands motivate us to develop an autonomous vehicle, named as Hercules, for contact-less goods transportation during the COVID-19 pandemic. The vehicle is evaluated through real-world delivering tasks under various traffic conditions.

Structural Basis for the Inhibition of the RNA-Dependent RNA Polymerase from SARS-CoV-2 by Remdesivir (preprint)

The pandemic of Corona Virus Disease 2019 (COVID-19) caused by SARS-CoV-2 has become a global crisis. The replication of SARS-CoV-2 requires the viral RNA-dependent RNA polymerase (RdRp), a direct target of the antiviral drug, Remdesivir. Here we report the structure of the SARS-CoV-2 RdRp either in the apo form or in complex with a 50-base template-primer RNA and Remdesivir at a resolution range of 2.5-2.8 [A]. The complex structure reveals that the partial double-stranded RNA template is inserted into the central channel of the RdRp where Remdesivir is incorporated into the first replicated base pair and terminates the chain elongation. Our structures provide critical insights into the working mechanism of viral RNA replication and a rational template for drug design to combat the viral infection.

Forecasting the Worldwide Spread of COVID-19 based on Logistic Model and SEIR Model (preprint)

Background: With the outbreak of coronavirus disease 2019 (COVID-19), a sudden case increase in late February 2020 led to deep concern globally. Italy, South Korea, Iran, France, Germany, Spain, the US and Japan are probably the countries with the most severe outbreaks. Collecting epidemiological data and predicting epidemic trends are important for the development and measurement of public intervention strategies. Epidemic prediction results yielded by different mathematical models are inconsistent; therefore, we sought to compare different models and their prediction results to generate objective conclusions. Methods: We used the number of cases reported from January 23 to March 20, 2020, to estimate the possible spread size and peak time of COVID-19, especially in 8 high-risk countries. The logistic growth model, basic SEIR model and adjusted SEIR model were adopted for prediction. Given that different model inputs may infer different model outputs, we implemented three model predictions with three scenarios of epidemic development. Results: When comparing all 8 countries short-term prediction results and peak predictions, the differences among the models were relatively large. The logistic growth model estimated a smaller epidemic size than the basic SERI model did; however, once we added parameters that considered the effects of public health interventions and control measures, the adjusted SERI model results demonstrated a considerably rapid deceleration of epidemic development. Our results demonstrated that contact rate, quarantine scale, and the initial quarantine time and length are important factors in controlling epidemic size and length. Conclusions: We demonstrated a comparative assessment of the predictions of the COVID-19 outbreak in eight high-risk countries using multiple methods. By forecasting epidemic size and peak time as well as simulating the effects of public health interventions, the intent of this paper is to help clarify the transmission dynamics of COVID-19 and recommend operation suggestions to slow down the epidemic. It is suggested that the quick detection of cases, sufficient implementation of quarantine and public self-protection behaviors are critical to slow down the epidemic.