A Brief Review of Graphene-Based Biosensors Developed for Rapid Detection of COVID-19 Biomarkers.

The prevalence of mutated species of COVID-19 antigens has provided a strong impetus for identifying a cost-effective, rapid and facile strategy for identifying the viral loads in public places. The ever-changing genetic make-up of SARS-CoV-2 posts a significant challenfge for the research community to identify a robust mechanism to target, bind and confirm the presence of a viral load before it spreads. Synthetic DNA constructs are a novel strategy to design complementary DNA sequences specific for antigens of interest as in this review's case SARS-CoV-2 antigens. Small molecules, complementary DNA and protein-DNA complexes have been known to target analytes in minimal concentrations. This phenomenon can be exploited by nanomaterials which have unique electronic properties such as ballistic conduction. Graphene is one such candidate for designing a device with a very low LOD in the order of zeptomolar and attomolar concentrations. Surface modification will be the significant aspect of the device which needs to have a high degree of sensitivity at the same time as providing a rapid signaling mechanism.

Management of cardiovascular symptoms after Pfizer-BioNTech COVID-19 vaccine in teenagers in the emergency department.

BACKGROUND: Cardiovascular complications after Pfizer-BioNTech COVID-19 (BNT) vaccination are a concern, especially in adolescents. We analyzed the risk factors for myocarditis after BNT vaccination. METHODS: We used a special evaluation protocol for all patients aged 12-18 years who presented to our emergency department with cardiovascular symptoms after BNT vaccination. RESULTS: A total of 195 patients (109 boys and 86 girls) were enrolled. Eleven (5.6%) patients presented with arrhythmia (arrhythmia group), 14 (7.2%) had a diagnosis of pericarditis/myocarditis (the peri/myocarditis group), and the remaining 170 were controls (no cardiac involvement). Chest pain (77.6%) was the most common symptom. The median time from vaccination to symptom onset was 3 days. In the peri/myocarditis group (13 myocarditis and 1 pericarditis), the median time to the peak troponin T level was 5 days after vaccination. Abnormal electrocardiographic changes, including ST-T changes and conduction blocks, were more commonly detected in the peri/myocarditis group (85.7% vs. 12.4% in the control group, p < 0.01). Echocardiography revealed normal ventricular function in all patients. Symptoms were resolved before discharge in all, with the median duration of hospital stay being 4 days. The electrocardiography was the most appropriate screening tool for myocarditis, with a sensitivity and specificity of 85.7% and 87.6%, respectively. CONCLUSION: Pericarditis or myocarditis was diagnosed in 7.2% of adolescents presenting to the emergency department with cardiovascular symptoms after BNT vaccination. In addition to the troponin T level, ECG change listed above can be used as a screening tool for vaccine-induced cardiac complications.

Construction of infectious bronchitis virsus endoribonucleasensp15 defective recombinant virus by reverse genetic system

Infectious Bronchitis Virus (1BV) belongs to the y coronavirus, however, the function of IBV encoded endoribonuclease (non - structural protein 15, nsp15) has not been determined yet. To explore the function of nsp15 in the process of IBV replication, we mutated the IBV nsp15 endonuclease core residue His238 to Ala, constructed the nsp15-defective recombinant virus rIBV-nsp15-H238A, via in vitro ligation and recombination technology. Plaque assay and TCID50 were performed to measure virus titer, virus plaque size and growth curve. The IBV Beaudette-R genome was cloned as 5 fragments in vectors, BsaI or BsmBI restriction sites were added to the end of each fragment. After plasmid amplification, the cDNA fragments were obtained by enzymatic digestion, followed with in vitro ligation and transcription. Full - length genomic RNA was electroporated into Vero cells, together with N transcript, to rescue the recombinant viruses rIBV and rIBV - nsp15- H238A. Plaque assay was performed to detect and compare the viral titer and plaque size of these two recombinant viruses. Results showed that the virus titer of rIBV -nsp15 -H238A was 2.71x106PFU/mL, 3 times lower than that of rIBV (9.4x106PFU/mL). The plaque size of rIBV-nsp15-H238A was much smaller than that of rIBV, indicating that rIBV-nsp15-11238A replicates and spreads slower than rIBV. The growth curve of rIBV-nsp15-H238A was slower than that of rIBV. Our study demonstrates that nsp15 I-1238 is the key amino acid and plays an important role in the replication and spread of IBV. The construction of nsp15 defective recombinant virus provides a powerful tool for the study of the function of nsp15.

Non-Targeted Metabolomic Analysis of Chicken Kidneys in Response to Coronavirus IBV Infection Under Stress Induced by Dexamethasone.

Stress in poultry can lead to changes in body metabolism and immunity, which can increase susceptibility to infectious diseases. However, knowledge regarding chicken responses to viral infection under stress is limited. Dexamethasone (Dex) is a synthetic glucocorticoid similar to that secreted by animals under stress conditions, and has been widely used to induce stress in chickens. Herein, we established a stress model in 7-day-old chickens injected with Dex to elucidate the effects of stress on IBV replication in the kidneys. The metabolic changes, immune status and growth of the chickens under stress conditions were comprehensively evaluated. Furthermore, the metabolic profile, weight gain, viral load, serum cholesterol levels, cytokines and peripheral blood lymphocyte ratio were compared in chickens treated with Dex and infected with IBV. An LC-MS/MS-based metabolomics method was used to examine differentially enriched metabolites in the kidneys. A total of 113 metabolites whose abundance was altered after Dex treatment were identified, most of which were lipids and lipid-like molecules. The principal metabolic alterations in chicken kidneys caused by IBV infection included fatty acid, valine, leucine and isoleucine metabolism. Dex treatment before and after IBV infection mainly affected the host's tryptophan, phenylalanine, amino sugar and nucleotide sugar metabolism. In addition, Dex led to up-regulation of serum cholesterol levels and renal viral load in chickens, and to the inhibition of weight gain, peripheral blood lymphocytes and IL-6 production. We also confirmed that the exogenous cholesterol in DF-1 cells promoted the replication of IBV. However, whether the increase in viral load in kidney tissue is associated with the up-regulation of cholesterol levels induced by Dex must be demonstrated in future experiments. In conclusion, chick growth and immune function were significantly inhibited by Dex. Host cholesterol metabolism and the response to IBV infection are regulated by Dex. This study provides valuable insights into the molecular regulatory mechanisms in poultry stress, and should support further research on the intrinsic link between cholesterol metabolism and IBV replication under stress conditions.

Characterization of the Protective Efficacy Against QX Strain of a Recombinant Infectious Bronchitis Virus With H120 Backbone and QX Spike Gene.

Infectious bronchitis virus (IBV) has been prevalent in chicken farms for many years, and its control relies on extensive vaccine administration. The continuous emergence of new variants and the low cross-protection efficiency prompt the development of new vaccines. In this study, we develop a reverse genetics technique based on the classical vaccine strain H120 genome, via in vitro ligation method. Using the H120 genome as the backbone, we constructed the recombinant virus rH120-QX(S) by replacing the H120 S gene with the QX S gene, a prevalent strain in China. Biological characteristics of the rH120-QX(S) virus, such as 50% egg lethal dose (ELD50), 50% egg infectious dose (EID50), dwarf embryo, growth curve, and genetic stability, are measured, which are comparable to the parental virus H120. There are no clinical symptoms and tissue lesions in the trachea and kidney in the rH120-QX(S)-infected specific-pathogen-free (SPF) chickens, demonstrating that this recombinant virus does not confer pathogenicity. Furthermore, protection studies show that there is 100% homologous protection of rH120-QX(S) to the virulent QX strain, as shown by the absence of clinical signs and no lethality. Taken together, our results demonstrate that swapping the S gene onto the H120 genetic backbone is a precise and effective way to produce genetically defined IBV vaccine candidates.

Coronavirus Infection and Cholesterol Metabolism.

Host cholesterol metabolism remodeling is significantly associated with the spread of human pathogenic coronaviruses, suggesting virus-host relationships could be affected by cholesterol-modifying drugs. Cholesterol has an important role in coronavirus entry, membrane fusion, and pathological syncytia formation, therefore cholesterol metabolic mechanisms may be promising drug targets for coronavirus infections. Moreover, cholesterol and its metabolizing enzymes or corresponding natural products exert antiviral effects which are closely associated with individual viral steps during coronavirus replication. Furthermore, the coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 infections are associated with clinically significant low cholesterol levels, suggesting cholesterol could function as a potential marker for monitoring viral infection status. Therefore, weaponizing cholesterol dysregulation against viral infection could be an effective antiviral strategy. In this review, we comprehensively review the literature to clarify how coronaviruses exploit host cholesterol metabolism to accommodate viral replication requirements and interfere with host immune responses. We also focus on targeting cholesterol homeostasis to interfere with critical steps during coronavirus infection.

The Interplay Between Coronavirus and Type I IFN Response

In the past few decades, newly evolved coronaviruses have posed a global threat to public health and animal breeding. To control and prevent the coronavirus-related diseases, understanding the interaction of the coronavirus and the host immune system is the top priority. Coronaviruses have evolved multiple mechanisms to evade or antagonize the host immune response to ensure their replication. As the first line and main component of innate immune response, type I IFN response is able to restrict virus in the initial infection stage;it is thus not surprising that the primary aim of the virus is to evade or antagonize the IFN response. Gaining a profound understanding of the interaction between coronaviruses and type I IFN response will shed light on vaccine development and therapeutics. In this review, we provide an update on the current knowledge on strategies employed by coronaviruses to evade type I IFN response.

PABPC4 Broadly Inhibits Coronavirus Replication by Degrading Nucleocapsid Protein through Selective Autophagy.

Emerging coronaviruses (CoVs) can cause severe diseases in humans and animals, and, as of yet, none of the currently available broad-spectrum drugs or vaccines can effectively control these diseases. Host antiviral proteins play an important role in inhibiting viral proliferation. One of the isoforms of cytoplasmic poly(A)-binding protein (PABP), PABPC4, is an RNA-processing protein, which plays an important role in promoting gene expression by enhancing translation and mRNA stability. However, its function in viruses remains poorly understood. Here, we report that the host protein, PABPC4, could be regulated by transcription factor SP1 and broadly inhibits the replication of CoVs, covering four genera (Alphacoronavirus, Betacoronavirus, Gammacoronavirus, and Deltacoronavirus) of the Coronaviridae family by targeting the nucleocapsid (N) protein through the autophagosomes for degradation. PABPC4 recruited the E3 ubiquitin ligase MARCH8/MARCHF8 to the N protein for ubiquitination. Ubiquitinated N protein was recognized by the cargo receptor NDP52/CALCOCO2, which delivered it to the autolysosomes for degradation, resulting in impaired viral proliferation. In addition to regulating gene expression, these data demonstrate a novel antiviral function of PABPC4, which broadly suppresses CoVs by degrading the N protein via the selective autophagy pathway. This study will shed light on the development of broad anticoronaviral therapies. IMPORTANCE Emerging coronaviruses (CoVs) can cause severe diseases in humans and animals, but none of the currently available drugs or vaccines can effectively control these diseases. During viral infection, the host will activate the interferon (IFN) signaling pathways and host restriction factors in maintaining the innate antiviral responses and suppressing viral replication. This study demonstrated that the host protein, PABPC4, interacts with the nucleocapsid (N) proteins from eight CoVs covering four genera (Alphacoronavirus, Betacoronavirus, Gammacoronavirus, and Deltacoronavirus) of the Coronaviridae family. PABPC4 could be regulated by SP1 and broadly inhibits the replication of CoVs by targeting the nucleocapsid (N) protein through the autophagosomes for degradation. This study significantly increases our understanding of the novel host restriction factor PABPC4 against CoV replication and will help develop novel antiviral strategies.

Inhibition of anti-viral stress granule formation by coronavirus endoribonuclease nsp15 ensures efficient virus replication.

Cytoplasmic stress granules (SGs) are generally triggered by stress-induced translation arrest for storing mRNAs. Recently, it has been shown that SGs exert anti-viral functions due to their involvement in protein synthesis shut off and recruitment of innate immune signaling intermediates. The largest RNA viruses, coronaviruses, impose great threat to public safety and animal health; however, the significance of SGs in coronavirus infection is largely unknown. Infectious Bronchitis Virus (IBV) is the first identified coronavirus in 1930s and has been prevalent in poultry farm for many years. In this study, we provided evidence that IBV overcomes the host antiviral response by inhibiting SGs formation via the virus-encoded endoribonuclease nsp15. By immunofluorescence analysis, we observed that IBV infection not only did not trigger SGs formation in approximately 80% of the infected cells, but also impaired the formation of SGs triggered by heat shock, sodium arsenite, or NaCl stimuli. We further demonstrated that the intrinsic endoribonuclease activity of nsp15 was responsible for the interference of SGs formation. In fact, nsp15-defective recombinant IBV (rIBV-nsp15-H238A) greatly induced the formation of SGs, along with accumulation of dsRNA and activation of PKR, whereas wild type IBV failed to do so. Consequently, infection with rIBV-nsp15-H238A strongly triggered transcription of IFN-ß which in turn greatly affected rIBV-nsp15-H238A replication. Further analysis showed that SGs function as an antiviral hub, as demonstrated by the attenuated IRF3-IFN response and increased production of IBV in SG-defective cells. Additional evidence includes the aggregation of pattern recognition receptors (PRRs) and signaling intermediates to the IBV-induced SGs. Collectively, our data demonstrate that the endoribonuclease nsp15 of IBV interferes with the formation of antiviral hub SGs by regulating the accumulation of viral dsRNA and by antagonizing the activation of PKR, eventually ensuring productive virus replication. We further demonstrated that nsp15s from PEDV, TGEV, SARS-CoV, and SARS-CoV-2 harbor the conserved function to interfere with the formation of chemically-induced SGs. Thus, we speculate that coronaviruses employ similar nsp15-mediated mechanisms to antagonize the host anti-viral SGs formation to ensure efficient virus replication.

Estimating the serial interval of the novel coronavirus disease (COVID-19) based on the public surveillance data in Shenzhen, China, from 19 January to 22 February 2020.

The novel coronavirus disease (COVID-19) poses a serious threat to global public health and economics. Serial interval (SI), time between the onset of symptoms of a primary case and a secondary case, is a key epidemiological parameter. We estimated SI of COVID-19 in Shenzhen, China based on 27 records of transmission chains. We adopted three parametric models: Weibull, lognormal and gamma distributions, and an interval-censored likelihood framework. The three models were compared using the corrected Akaike information criterion (AICc). We also fitted the epidemic curve of COVID-19 to the logistic growth model to estimate the reproduction number. Using a Weibull distribution, we estimated the mean SI to be 5.9 days (95% CI: 3.9-9.6) with a standard deviation (SD) of 4.8 days (95% CI: 3.1-10.1). Using a logistic growth model, we estimated the basic reproduction number in Shenzhen to be 2.6 (95% CI: 2.4-2.8). The SI of COVID-19 is relatively shorter than that of SARS and MERS, the other two betacoronavirus diseases, which suggests the iteration of the transmission may be rapid. Thus, it is crucial to isolate close contacts promptly to effectively control the spread of COVID-19.