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1.
Viruses ; 14(6)2022 May 26.
Article in English | MEDLINE | ID: covidwho-1869813

ABSTRACT

BACKGROUND: Immune response assessed by the quantification of neutralizing antibodies (nAbs) and predictors associated with immunogenicity after the prime-boost ChAdOx1 (Oxford-AstraZeneca) COVID-19 vaccine in hemodialysis (HD) patients remains unclear. METHODS: This prospective study enrolled 174 HD patients and 67 healthy subjects to evaluate antibodies against the spike protein 1 and receptor-binding domain of severe acute respiratory syndrome coronavirus type 2 after prime-booster vaccination, by using enzyme-linked immunosorbent assay and applied spline-based generalized additive model regression analysis to predict 50% neutralization titer (NT50). The correlation between HD parameters and NT50 was analyzed. RESULTS: NT50 was lower in HD patients compared with healthy controls after the prime-boost dose (p < 0.001). The geometric mean titer ratios were higher in first-dose seronegative than in the seropositive subgroup in HD patients and healthy controls (6.96 vs. 2.36, p = 0.002, and 9.28 vs. 1.26, p = 0.011, respectively). After two doses of ChAdOx1, one-way ANOVA showed that Ca × P was positively associated with NT50 (p trend = 0.043) and multiple linear regression showed the similar results (p = 0.021). Kt/V (a quantification of dialysis adequacy) (OR = 20.295, p = 0.005) could independently predict seroconversion (NT50 ≥ 35.13 IU/mL). CONCLUSION: Adequacy of hemodialysis could independently predict seroconversion in HD subjects vaccinated with prime-boost doses of ChAdOx1.

2.
Infect Dis Ther ; 2022 May 17.
Article in English | MEDLINE | ID: covidwho-1850469

ABSTRACT

INTRODUCTION: MVC-COV1901 is a protein subunit COVID-19 vaccine based on the stable prefusion spike protein S-2P adjuvanted with CpG 1018 and aluminum hydroxide. Interim results of a phase 2 clinical trial demonstrated favorable safety profile and immunogenicity and the vaccine has been authorized for use in Taiwan. However, waning antibody levels after immunization and variants of concern (VoC) could negatively impact vaccine-induced neutralization of virus. In this extension to the phase 1 clinical study we investigated a three-dose regimen of MVC-COV1901 for durability of antibody levels and virus neutralization capacity, including neutralization of the Omicron variant. METHODS: Forty-five healthy adults from 20 to 49 years of age were divided into three groups of 15 participants receiving two doses of either low dose (LD), medium dose (MD), or high dose (HD) of MVC-COV1901. Six months after the second dose (day 209), a third MD dose of MVC-COV1901 was administered to the LD and MD groups and a HD dose was given to the HD group. Safety was followed for up to 28 days after the booster dose by monitoring incidences of adverse events (AE). Immunogenicity and antibody persistence for up to 6 months after the booster dose were assessed by neutralizing assay with the wild-type (Wuhan) SARS-CoV-2 virus. To examine the immunogenicity of booster dose against variants, neutralizing assays were carried out with the Alpha, Beta, and Delta variant viruses and the Omicron variant pseudovirus using samples from 4 weeks after the booster dose. RESULTS: Adverse reactions after the booster dose were mostly mild and comparable to that of the first two doses. Compared to day 209, neutralizing antibodies were increased by 10.3-28.9 times at 4 weeks after the booster. During the 6-month follow-up after the booster, the rate of decline of neutralizing antibody level was much less than that after the second dose. Three doses of MVC-COV1901 also improved antibody-mediated neutralization of Alpha, Beta, and Delta variants as well as the Omicron variant pseudovirus. CONCLUSION: Our data showed increased persistence of neutralizing antibodies and enhancement of immunogenicity against VoCs offered after a third dose of MVC-COV1901. TRIAL REGISTRATION: ClinicalTrials.gov identifier NCT04487210.

3.
Methods Mol Biol ; 2452: 3-18, 2022.
Article in English | MEDLINE | ID: covidwho-1844256

ABSTRACT

A novel coronavirus (CoV) that emerged in Wuhan, Hubei province in China, in December 2019, has rapidly spread worldwide. Named as the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), this virus has been responsible for infecting about 153 million people and causing 3 million deaths by May 2021. There is obvious interest in gaining novel insights into the epidemiologic evolution of this virus; however, inappropriate application and interpretation of genomic and phylogenetic analyses has led to dangerous outcomes and misunderstandings. This chapter focuses on not only introducing this virus, its genomic characteristics and molecular mechanisms but also describing the application and interpretation of phylogenetic tree analyses, in order to provide useful information to better understand the evolution and epidemiology of this virus. In addition, recombinant region and genetic ancestry of SARS-CoV-2 remain unknown. It is urgently required to collect samples and obtain related viral genetic data from animal sources for identifying the intermediate host of SARS-CoV-2 that is responsible for its cross-species transmission.


Subject(s)
COVID-19 , SARS-CoV-2 , Animals , COVID-19/epidemiology , China/epidemiology , Humans , Phylogeny , SARS-CoV-2/genetics
4.
Microbiol Mol Biol Rev ; 86(2): e0002621, 2022 Jun 15.
Article in English | MEDLINE | ID: covidwho-1765086

ABSTRACT

Coronavirus disease 2019 (COVID-19) is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The global COVID-19 pandemic continues to threaten the lives of hundreds of millions of people, with a severe negative impact on the global economy. Although several COVID-19 vaccines are currently being administered, none of them is 100% effective. Moreover, SARS-CoV-2 variants remain an important worldwide public health issue. Hence, the accelerated development of efficacious antiviral agents is urgently needed. Coronavirus depends on various host cell factors for replication. An ongoing research objective is the identification of host factors that could be exploited as targets for drugs and compounds effective against SARS-CoV-2. In the present review, we discuss the molecular mechanisms of SARS-CoV-2 and related coronaviruses, focusing on the host factors or pathways involved in SARS-CoV-2 replication that have been identified by genome-wide CRISPR screening.


Subject(s)
COVID-19 , SARS-CoV-2 , Antiviral Agents/pharmacology , Antiviral Agents/therapeutic use , COVID-19 Vaccines , Humans , Pandemics/prevention & control , SARS-CoV-2/genetics
5.
EuropePMC; 2022.
Preprint in English | EuropePMC | ID: ppcovidwho-329648

ABSTRACT

The outbreak of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has substantially impacted human health globally. Spike-specific antibody response plays a major role in protection against SARS-CoV-2. Here, we demonstrated that acute SARS-CoV-2 infection elicits rapid and robust spike-binding and ACE2-blocking antibody responses, which wane approximately 11 months after infection. Serological responses were found to be correlated with the frequency of spike-specific memory B cell responses to natural infections. Further, significantly higher spike-binding, ACE2-blocking, and memory B cell responses were detected in patients with fever and pneumonia. Spike-specific antibody responses were found to be greatly affected by spike mutations in emerging variants, especially the Beta and Omicron variants. These results warrant continued surveillance of spike-specific antibody responses to natural infections and highlight the importance of maintaining functional anti-spike antibodies through immunization. Importance As spike protein-specific antibody responses play a major role in protection against SARS-CoV-2, we examined the spike-binding and ACE2-blocking antibody responses in SARS-CoV-2 infection at different time points. We found robust responses following acute infection, which waned approximately 11 months after infection. Further, the serological responses were correlated with the frequency of spike-specific memory B cell responses to natural infections. Patients with fever and pneumonia showed significantly stronger spike-binding, ACE2-blocking antibody, and memory B cell responses. Moreover, the spike-specific antibody responses were substantially affected by the emerging variants, especially the Beta and Omicron variants. These results warrant continued surveillance of spike-specific antibody responses to natural infections and highlight the importance of maintaining functional anti-spike antibodies through immunization.

7.
mSphere ; 7(1): e0088321, 2022 02 23.
Article in English | MEDLINE | ID: covidwho-1673356

ABSTRACT

Considering the urgent demand for faster methods to quantify neutralizing antibody titers in patients with coronavirus (CoV) disease 2019 (COVID-19), developing an analytical model or method to replace the conventional virus neutralization test (NT) is essential. Moreover, a "COVID-19 immunity passport" is currently being proposed as a certification for people who travel internationally. Therefore, an enzyme-linked immunosorbent assay (ELISA) was designed to detect severe acute respiratory syndrome CoV 2 (SARS-CoV-2)-neutralizing antibodies in serum, which is based on the binding affinity of SARS-CoV-2 viral spike protein 1 (S1) and the viral spike protein receptor-binding domain (RBD) to antibodies. The RBD is considered the major binding region of neutralizing antibodies. Furthermore, S1 covers the RBD and several other regions, which are also important for neutralizing antibody binding. In this study, we assessed 144 clinical specimens, including those from patients with PCR-confirmed SARS-CoV-2 infections and healthy donors, using both the NT and ELISA. The ELISA results analyzed by spline regression and the two-variable generalized additive model precisely reflected the NT value, and the correlation between predicted and actual NT values was as high as 0.917. Therefore, our method serves as a surrogate to quantify neutralizing antibody titer. The analytic method and platform used in this study present a new perspective for serological testing of SARS-CoV-2 infection and have clinical potential to assess vaccine efficacy. IMPORTANCE Herein, we present a new approach for serological testing for SARS-CoV-2 antibodies using innovative laboratory methods that demonstrate a combination of biology and mathematics. The traditional virus neutralization test is the gold standard method; however, it is time-consuming and poses a risk to medical personnel. Thus, there is a demand for methods that rapidly quantify neutralizing antibody titers in patients with COVID-19 or examine vaccine efficacy at a biosafety level 2 containment facility. Therefore, we used a two-variable generalized additive model to analyze the results of the enzyme-linked immunosorbent assay and found the method to serve as a surrogate to quantify neutralizing antibody titers. This methodology has potential for clinical use in assessing vaccine efficacy.


Subject(s)
Antibodies, Neutralizing/blood , COVID-19/immunology , Enzyme-Linked Immunosorbent Assay , Models, Immunological , Models, Statistical , Neutralization Tests/methods , SARS-CoV-2/immunology , Biomarkers/blood , COVID-19/blood , COVID-19/diagnosis , Case-Control Studies , Humans , Regression Analysis
8.
mBio ; : e0271721, 2022 Jan 18.
Article in English | MEDLINE | ID: covidwho-1634330

ABSTRACT

Enterovirus infections can cause severe complications, such as poliomyelitis, encephalitis, myocarditis, meningitis, neurological pulmonary edema, and even death. Here, we used genome-wide CRISPR screens to gain new insight into the mechanism by which enteroviruses co-opt host pathways to potentiate replication and propagation. We found that acyl-coenzyme A synthetase long-chain family member 4 (ACSL4) is involved in viral replication organelle formation. ACSL4 is a key component of ferroptosis, an iron-dependent, nonapoptotic programmed cell death. Our results indicated that enteroviruses and coronaviruses can induce ferroptosis via ACSL4. Most importantly, ferroptosis inhibitors, including two FDA-approved drugs, rosiglitazone (ROSI; ACSL4 inhibitor) and pioglitazone (PIO; ACSL4 inhibitor), decreased the viral load of human enteroviruses and coronaviruses, suggesting that ACSL4 is a target for counteracting viral infection. IMPORTANCE We provide the first evidence for the role of ACSL4 in enterovirus replication organelle formation. Moreover, both enteroviruses and coronaviruses induce ferroptosis via ACSL4. These findings establish a novel regulatory mechanism for viral replication. The inhibition of ACSL4 by ferroptosis inhibitors can reduce viral yields of enteroviruses and coronaviruses, including SARS-CoV-2, implying that ACSL4-mediated ferroptosis is a promising therapeutic target for viral diseases.

9.
Lancet Respir Med ; 9(12): 1396-1406, 2021 12.
Article in English | MEDLINE | ID: covidwho-1621134

ABSTRACT

BACKGROUND: MVC-COV1901, a recombinant protein vaccine containing pre-fusion-stabilised spike protein S-2P adjuvanted with CpG 1018 and aluminium hydroxide, has been shown to be well tolerated with a good safety profile in healthy adults aged 20-49 years in a phase 1 trial, and provided a good cellular and humoral immune responses. We present the interim safety, tolerability, and immunogenicity results of a phase 2 clinical trial of the MVC-COV1901 vaccine in Taiwan. METHODS: This is a large-scale, double-blind, randomised, placebo-controlled phase 2 trial done at ten medical centres and one regional hospital in Taiwan. Individuals aged 20 years or older who were generally healthy or had stable pre-existing medical conditions were eligible for enrolment. Exclusion criteria included (but were not limited to) travel overseas within 14 days of screening, intention to travel overseas within 6 months of the screening visit, and the absence of prespecified medical conditions, including immunosuppressive illness, a history of autoimmune disease, malignancy with risk to recur, a bleeding disorder, uncontrolled HIV infection, uncontrolled hepatitis B and C virus infections, SARS-CoV-1 or SARS-CoV-2 infections, an allergy to any vaccine, or a serious medical condition that could interfere with the study. Study participants were randomly assigned (6:1) to receive two doses of either MVC-COV1901 or placebo, administered via intramuscular injection on day 1 and day 29. MVC-COV1901 contained 15 µg of S-2P protein adjuvanted with 750 µg CpG 1018 and 375 µg aluminium hydroxide in a 0·5 mL aqueous solution, and the placebo contained the same volume of saline. Randomisation was done centrally by use of an interactive web response system, stratified by age (≥20 to <65 years and ≥65 years). Participants and investigators were masked to group assignment. The primary outcomes were to evaluate the safety, tolerability, and immunogenicity of MVC-COV1901 from day 1 (the day of the first dose) to day 57 (28 days after the second dose). Safety was assessed in all participants who received at least one dose. Immunogenicity was assessed by measuring geometric mean titres (GMTs) and seroconversion rates of neutralising antibody and antigen-specific IgG in the per-protocol population. This study is registered with ClinicalTrials.gov, NCT04695652. FINDINGS: Of 4173 individuals screened between Dec 30, 2020, and April 2, 2021, 3854 were enrolled and randomly assigned: 3304 to the MVC-COV1901 group and 550 to the placebo group. A total of 3844 participants (3295 in the MVC-COV1901 group and 549 in the placebo group) were included in the safety analysis set, and 1053 participants (903 and 150) had received both doses and were included in the per-protocol immunogenicity analysis set. From the start of this phase 2 trial to the time of interim analysis, no vaccine-related serious adverse events were recorded. The most common solicited adverse events in all study participants were pain at the injection site (2346 [71·2%] of 3295 in the MVC-COV1901 group and 128 [23·3%] of 549 in the placebo group), and malaise or fatigue (1186 [36·0%] and 163 [29·7%]). Fever was rarely reported (23 [0·7%] and two [0·4%]). At 28 days after the second dose of MVC-COV1901, the wild-type SARS-CoV-2 neutralising antibody GMT was 662·3 (95% CI 628·7-697·8; 408·5 IU/mL), the GMT ratio (geometric mean fold increase in titres at day 57 vs baseline) was 163·2 (155·0-171·9), and the seroconversion rate was 99·8% (95% CI 99·2-100·0). INTERPRETATION: MVC-COV1901 has a good safety profile and elicits promising immunogenicity responses. These data support MVC-COV1901 to enter phase 3 efficacy trials. FUNDING: Medigen Vaccine Biologics and Taiwan Centres for Disease Control, Ministry of Health and Welfare.


Subject(s)
Adjuvants, Immunologic , Aluminum Hydroxide , COVID-19 Vaccines/immunology , COVID-19 , HIV Infections , Oligodeoxyribonucleotides , Adult , Aged , Antibodies, Neutralizing/blood , Antibodies, Viral/blood , Double-Blind Method , Humans , Middle Aged , SARS-CoV-2 , Taiwan , Young Adult
10.
J Pers Med ; 12(1)2022 Jan 07.
Article in English | MEDLINE | ID: covidwho-1613865

ABSTRACT

BACKGROUND: Data are lacking regarding predictors of quantification of neutralizing antibodies (nAbs) based on severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) 50% neutralization titer (NT50) after a single dose of COVID-19 vaccine in hemodialysis (HD) patients. METHODS: This prospective single-center study enrolled 200 HD patients and 82 healthy subjects to estimate antibodies against the SARS-CoV-2 viral spike protein 1 and receptor-binding domain after a first dose of a COVID-19 vaccine (ChAdOx1 or mRNA-1273), measured by enzyme-linked immunosorbent assay and applied spline-based generalized additive model regression analysis to predict NT50 converted to international units. RESULTS: After the first dose of ChAdOx1, multiple linear regression showed that age (p = 0.011) and cardiothoracic ratio (p = 0.002) were negatively associated with NT50. Older age (OR = 0.958, p = 0.052) and higher cardiothoracic ratio (OR < 0.001, p = 0.037) could predict negative humoral response (NT50 < 35.13 IU/mL). NT50 was lower in HD patients compared with healthy controls receiving ChAdOx1 (10.68 vs. 43.01 IU/m, p < 0.001) or mRNA-1273 (36.39 vs. 262.2 IU/mL, p < 0.001). ChAdOx1 elicited lower GMTs than mRNA-1273 in the HD cohort (10.68 vs. 36.39 IU/mL, p < 0.001) and in healthy controls (43.01 vs. 262.22 IU/mL, p < 0.001). CONCLUSION: High cardiothoracic ratio and old age could independently predict a decline in nAb titers in an HD cohort vaccinated with a single dose of ChAdOx1.

12.
2021.
Preprint in English | Other preprints | ID: ppcovidwho-296105

ABSTRACT

Summary Background We have assessed the safety and immunogenicity of the COVID-19 vaccine MVC-COV1901, a recombinant protein vaccine containing prefusion-stabilized spike protein S-2P adjuvanted with CpG 1018 and aluminium hydroxide. Methods This is a phase 2, prospective, randomised, double-blind, placebo-controlled, and multi-centre study to evaluate the safety, tolerability, and immunogenicity of the SARS-CoV-2 vaccine candidate MVC-COV1901. The study comprised 3,844 participants of ≥ 20 years who were generally healthy or with stable pre-existing medical conditions. The study participants were randomly assigned in a 6:1 ratio to receive either MVC-COV1901 containing 15 μg of S-2P protein or placebo containing saline. Participants received two doses of MVC-COV1901 or placebo, administered 28 days apart via intramuscular injection. The primary outcomes were to evaluate the safety, tolerability, and immunogenicity of MVC-COV1901 from Day 1 (the day of first vaccination) to Day 57 (28 days after the second dose). Immunogenicity of MVC-COV1901 was assessed through geometric mean titres (GMT) and seroconversion rates (SCR) of neutralising antibody and antigen-specific immunoglobulin. This clinical trial is registered at ClinicalTrials.gov: NCT04695652 . Findings From the start of this phase 2 trial to the time of interim analysis, no vaccine-related Serious Adverse Events (SAEs) were recorded. The most common solicited adverse events across all study participants were pain at the injection site (64%), and malaise/fatigue (35%). Fever was rarely reported (<1%). For all participants in the MVC-COV1901 group, at 28 days after the second dose against wild type SARS-CoV-2 virus, the GMT was 662·3 (408 IU/mL), the GMT ratio was 163·2, and the seroconversion rate was 99·8%. Interpretation MVC-COV1901 shows good safety profiles and promising immunogenicity responses. The current data supports MVC-COV1901 to enter phase 3 efficacy trials and could enable regulatory considerations for Emergency Use Authorisation (EUA). Funding Medigen Vaccine Biologics Corporation and Taiwan Centres for Disease Control.

13.
EuropePMC; 2021.
Preprint in English | EuropePMC | ID: ppcovidwho-294956

ABSTRACT

Abstract: In this extension of the phase 1 clinical study, we report the immunogenicity and reactogenicity of the booster dose of a COVID-19 vaccine, MVC-COV1901, administered six months after the completion of the primary two dose schedule. Antibody persistence was detected at 6 months after the second dose of MVC-COV1901, albeit at reduced levels. At 28 days after the booster dose, the neutralizing antibody titer was 1.7-fold higher compared to the previous peak at 2 weeks after the second dose. These data demonstrated the safety and immunogenicity of booster shot of MVC-COV1901 after the primary schedule of the vaccine.

14.
EuropePMC; 2021.
Preprint in English | EuropePMC | ID: ppcovidwho-293592

ABSTRACT

Abstract: In this extension of the phase 1 clinical study, we report the immunogenicity and reactogenicity of the booster dose of a COVID-19 vaccine, MVC-COV1901, administered six months after the completion of the primary two dose schedule. Antibody persistence was detected at 6 months after the second dose of MVC-COV1901, albeit at reduced levels. At 28 days after the booster dose, the neutralizing antibody titer was 1.7-fold higher compared to the previous peak at 2 weeks after the second dose. These data demonstrated the safety and immunogenicity of booster shot of MVC-COV1901 after the primary schedule of the vaccine.

15.
Mol Cell ; 81(21): 4467-4480.e7, 2021 11 04.
Article in English | MEDLINE | ID: covidwho-1473419

ABSTRACT

Viral RNA-dependent RNA polymerases (RdRps) are a target for broad-spectrum antiviral therapeutic agents. Recently, we demonstrated that incorporation of the T-1106 triphosphate, a pyrazine-carboxamide ribonucleotide, into nascent RNA increases pausing and backtracking by the poliovirus RdRp. Here, by monitoring enterovirus A-71 RdRp dynamics during RNA synthesis using magnetic tweezers, we identify the "backtracked" state as an intermediate used by the RdRp for copy-back RNA synthesis and homologous recombination. Cell-based assays and RNA sequencing (RNA-seq) experiments further demonstrate that the pyrazine-carboxamide ribonucleotide stimulates these processes during infection. These results suggest that pyrazine-carboxamide ribonucleotides do not induce lethal mutagenesis or chain termination but function by promoting template switching and formation of defective viral genomes. We conclude that RdRp-catalyzed intra- and intermolecular template switching can be induced by pyrazine-carboxamide ribonucleotides, defining an additional mechanistic class of antiviral ribonucleotides with potential for broad-spectrum activity.


Subject(s)
Pyrazines/chemistry , RNA Viruses/genetics , RNA, Viral/genetics , RNA-Dependent RNA Polymerase/genetics , Recombination, Genetic , Ribonucleotides/chemistry , Animals , Antiviral Agents , Catalysis , Cells, Cultured , Genetic Techniques , Genome , Genome, Viral , Homologous Recombination , Humans , Kinetics , Mice , Mice, Transgenic , Molecular Dynamics Simulation , Mutagenesis , Nucleotides/genetics , Protein Conformation , RNA/chemistry , RNA-Dependent RNA Polymerase/metabolism , RNA-Seq , Transgenes , Virulence
17.
EClinicalMedicine ; 38: 100989, 2021 Aug.
Article in English | MEDLINE | ID: covidwho-1375926

ABSTRACT

BACKGROUND: This was a phase 1, dose-escalation open-label trial to evaluate the safety and immunogenicity of MVC-COV1901, a SARS-CoV-2 S-2P protein vaccine adjuvanted with aluminum hydroxide and CpG 1018. METHODS: Between September 28 and November 13 2020, 77 participants were screened. Of these, 45 healthy adults from 20 to 49 years of age were to be administered two doses of MVC-COV1901 in doses of 5 µg, 15 µg, or 25 µg of spike protein at 28 days apart. There were 15 participants in each dose group; all were followed for 28 days after the second dose at the time of the interim analysis. Adverse events and laboratory data were recorded for the safety evaluation. Blood samples were collected for humoral, and cellular immune response at various time points. Trial Registration: ClinicalTrials.gov NCT04487210. FINDINGS: Solicited adverse events were mostly mild and similar. No subject experienced fever. After the second dose, the geometric mean titers (GMTs) for SARS-CoV-2 spike-specific immunoglobulin G were 7178.2, 7746.1, 11,220.6 in the 5 µg, 15 µg, and 25 µg dose groups, respectively. The neutralizing activity were detected in both methods. (Day 43 GMTs, 538.5, 993.1, and 1905.8 for pseudovirus; and 33.3, 76.3, and 167.4 for wild-type virus). The cellular immune response induced by MVC-COV1901 demonstrated substantially higher numbers of IFN-γ- producing cells, suggesting a Th1-skewed immune response. INTERPRETATION: The MVC-COV1901 vaccine was well tolerated and elicited robust immune responses and is suitable for further development. FUNDING: Medigen Vaccine Biologics Corporation.

18.
Pathogens ; 10(6)2021 Jun 13.
Article in English | MEDLINE | ID: covidwho-1270098

ABSTRACT

A total of 15 RT-PCR confirmed COVID-19 patients were admitted to our hospital during the in-itial outbreak in Taiwan. The average time of virus clearance was delayed in seven patients, 24.14 ± 4.33 days compared to 10.25 ± 0.56 days post-symptom onset (PSO) in the other eight pa-tients. There was strong antibody response in patients with viral persistence at the pharynx, with peak values of serum antibody 677.2 ± 217.8 vs. 76.70 ± 32.11 in patients with delayed versus rapid virus clearance. The patients with delayed viral clearance had excessive antibodies of compromised quality in an early stage with the delay in peak virus neutralization efficacy, 34.14 ± 7.15 versus 12.50 ± 2.35 days PSO in patients with rapid virus clearance. Weak antibody re-sponse of patients with rapid viral clearance was also effective, with substantial and comparable neutralization efficacy, 35.70 ± 8.78 versus 41.37 ± 11.49 of patients with delayed virus clearance. Human Cytokine 48-Plex Screening of the serial sera samples revealed elevated concentrations of proinflammatory cytokines and chemokines in a deceased patient with delayed virus clear-ance and severe disease. The levels were comparatively less in the other two patients who suf-fered from severe disease but eventually survived.

19.
mSphere ; 6(2)2021 03 31.
Article in English | MEDLINE | ID: covidwho-1166378

ABSTRACT

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) carrying the D614G mutation on the spike protein is the predominant circulating variant and is associated with enhanced infectivity. However, whether this dominant variant can potentially spread through the cold chain and whether the spike protein affects virus stability after cold storage remain unclear. To compare the infectivity of two SARS-CoV-2 variants, namely, SARS-CoV-2 variants with spike protein with the D614 mutation (S-D614) and G614 mutation (S-G614), after different periods of refrigeration (4°C) and freezing (-20°C). We also determined the integrity of the viral RNA and the ability of the spike protein to bind angiotensin-converting enzyme 2 (ACE2) after storage at these conditions. The results showed that SARS-CoV-2 was more stable and infectious after storage at -20°C than at 4°C. Particularly, the S-G614 variant was found to be more stable than the S-D614 variant. The spike protein of the S-G614 variant had better binding ability with the ACE2 receptor than that of the S-D614 variant after storage at -20°C for up to 30 days. Our findings revealed that SARS-CoV-2 remains stable and infectious after refrigeration or freezing, and their stability and infectivity up to 30 days depends on the spike variant. Stability and infectivity are related to each other, and the higher stability of S-G614 compared to that of S-D614 may contribute to rapid viral spread of the S-G614 variant.IMPORTANCE It has been observed that variants of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) are more stable and infectious after storage at -20°C than at 4°C. A SARS-CoV-2 S-D614G variant is currently the most dominant variant in circulation and is associated with enhanced infectivity. We compared the stability of two SARS-CoV-2 variants: the early S-D614 variant carrying the D614 spike protein and the new S-G614 variant carrying the G614 spike protein, stored at both 4°C and -20°C for different periods. We observed that SARS-CoV-2 remains stable and infectious after refrigeration or freezing, which further depends on the spike variant, that is, the ability of the spike protein to bind with the ACE2 receptor with higher efficiency. The high stability of the S-G614 variant also explains its rapid spread and infectivity. Therefore, precautions should be taken during and after handling food preserved under cold conditions.


Subject(s)
COVID-19 , SARS-CoV-2/genetics , Spike Glycoprotein, Coronavirus/genetics , Cold Temperature , Genetic Fitness/genetics , Humans , Mutation , Protein Stability
20.
Biosens Bioelectron ; 183: 113213, 2021 Jul 01.
Article in English | MEDLINE | ID: covidwho-1163433

ABSTRACT

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) enters the cells through the binding of its spike protein (S-protein) to the cell surface-expressing angiotensin-converting enzyme 2 (ACE2). Thus, inhibition of S-protein-ACE2 binding may impede SARS-CoV-2 cell entry and attenuate the progression of Coronavirus disease 2019 (COVID-19). In this study, an electrochemical impedance spectroscopy-based biosensing platform consisting of a recombinant ACE2-coated palladium nano-thin-film electrode as the core sensing element was fabricated for the screening of potential inhibitors against S-protein-ACE2 binding. The platform could detect interference of small analytes against S-protein-ACE2 binding at low analyte concentration and small volume (0.1 µg/mL and ~1 µL, estimated total analyte consumption < 4 pg) within 21 min. Thus, a few potential inhibitors of S-protein-ACE2 binding were identified. This includes (2S,3aS,6aS)-1-((S)-N-((S)-1-Carboxy-3-phenylpropyl)alanyl)tetrahydrocyclopenta[b] pyrrole-2-carboxylic acid (ramiprilat) and (2S,3aS,7aS)-1-[(2S)-2-[[(2S)-1-Carboxybutyl]amino]propanoyl]-2,3,3a,4,5,6,7,7a-octahydroindole-2-carboxylic acid (perindoprilat) that reduced the binding affinity of S-protein to ACE2 by 72% and 67%; and SARS-CoV-2 in vitro infectivity to the ACE2-expressing human oral cavity squamous carcinoma cells (OEC-M1) by 36.4 and 20.1%, respectively, compared to the PBS control. These findings demonstrated the usefulness of the developed biosensing platform for the rapid screening of modulators for S-protein-ACE2 binding.


Subject(s)
Biosensing Techniques , COVID-19 , Dielectric Spectroscopy , Humans , Protein Binding , SARS-CoV-2 , Spike Glycoprotein, Coronavirus
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