ABSTRACT
The coronavirus disease 2019 (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) threatens global public health. The development of a vaccine is urgently needed for the prevention and control of COVID-19. Here, we report the pilot-scale production of an inactivated SARS-CoV-2 vaccine candidate (BBIBP-CorV) that induces high levels of neutralizing antibodies titers in mice, rats, guinea pigs, rabbits, and nonhuman primates (cynomolgus monkeys and rhesus macaques) to provide protection against SARS-CoV-2. Two-dose immunizations using 2 µg/dose of BBIBP-CorV provided highly efficient protection against SARS-CoV-2 intratracheal challenge in rhesus macaques, without detectable antibody-dependent enhancement of infection. In addition, BBIBP-CorV exhibits efficient productivity and good genetic stability for vaccine manufacture. These results support the further evaluation of BBIBP-CorV in a clinical trial.
Subject(s)
Betacoronavirus/immunology , Coronavirus Infections/prevention & control , Drug Evaluation, Preclinical/methods , Pandemics/prevention & control , Pneumonia, Viral/prevention & control , Vaccines, Inactivated/therapeutic use , Viral Vaccines/therapeutic use , Animals , Antibodies, Neutralizing/immunology , Antibodies, Viral/immunology , Betacoronavirus/genetics , COVID-19 , COVID-19 Vaccines , Chlorocebus aethiops , Coronavirus Infections/virology , Disease Models, Animal , Female , Guinea Pigs , Immunogenicity, Vaccine , Macaca fascicularis , Macaca mulatta , Male , Mice , Mice, Inbred BALB C , Phylogeny , Pneumonia, Viral/virology , Rabbits , Rats , Rats, Wistar , SARS-CoV-2 , Vaccines, Inactivated/adverse effects , Vero Cells , Viral Vaccines/adverse effectsABSTRACT
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the cause of coronavirus disease 2019 (COVID-19), which has become a public health emergency of international concern1. Angiotensin-converting enzyme 2 (ACE2) is the cell-entry receptor for severe acute respiratory syndrome coronavirus (SARS-CoV)2. Here we infected transgenic mice that express human ACE2 (hereafter, hACE2 mice) with SARS-CoV-2 and studied the pathogenicity of the virus. We observed weight loss as well as virus replication in the lungs of hACE2 mice infected with SARS-CoV-2. The typical histopathology was interstitial pneumonia with infiltration of considerable numbers of macrophages and lymphocytes into the alveolar interstitium, and the accumulation of macrophages in alveolar cavities. We observed viral antigens in bronchial epithelial cells, macrophages and alveolar epithelia. These phenomena were not found in wild-type mice infected with SARS-CoV-2. Notably, we have confirmed the pathogenicity of SARS-CoV-2 in hACE2 mice. This mouse model of SARS-CoV-2 infection will be valuable for evaluating antiviral therapeutic agents and vaccines, as well as understanding the pathogenesis of COVID-19.
Subject(s)
Betacoronavirus/pathogenicity , Coronavirus Infections/pathology , Coronavirus Infections/virology , Lung/pathology , Peptidyl-Dipeptidase A/genetics , Peptidyl-Dipeptidase A/metabolism , Pneumonia, Viral/pathology , Pneumonia, Viral/virology , Transgenes , Angiotensin-Converting Enzyme 2 , Animals , Antigens, Viral/immunology , Antigens, Viral/metabolism , Betacoronavirus/immunology , Betacoronavirus/metabolism , Bronchi/pathology , Bronchi/virology , COVID-19 , Coronavirus Infections/immunology , Disease Models, Animal , Epithelial Cells/pathology , Epithelial Cells/virology , Female , Humans , Immunoglobulin G/immunology , Lung/immunology , Lung/virology , Lymphocytes/immunology , Macrophages, Alveolar/immunology , Macrophages, Alveolar/virology , Male , Mice , Mice, Transgenic , Pandemics , Pneumonia, Viral/immunology , Receptors, Complement 3d/genetics , Receptors, Complement 3d/metabolism , SARS-CoV-2 , Virus Replication , Weight LossABSTRACT
Bats are responsible for the zoonotic transmission of several major viral diseases, including those leading to the 2003 SARS outbreak and likely the ongoing COVID-19 pandemic. While comparative genomics studies have revealed characteristic adaptations of the bat innate immune system, functional genomic studies are urgently needed to provide a foundation for the molecular dissection of the viral tolerance in bats. Here we report the establishment of genome-wide RNA interference (RNAi) and CRISPR libraries for the screening of the model megabat, Pteropus alecto. We used the complementary RNAi and CRISPR libraries to interrogate P. alecto cells for infection with two different viruses: mumps virus and influenza A virus, respectively. Independent screening results converged on the endocytosis pathway and the protein secretory pathway as required for both viral infections. Additionally, we revealed a general dependence of the C1-tetrahydrofolate synthase gene, MTHFD1, for viral replication in bat cells and human cells. The MTHFD1 inhibitor, carolacton, potently blocked replication of several RNA viruses, including SARS-CoV-2. We also discovered that bats have lower expression levels of MTHFD1 than humans. Our studies provide a resource for systematic inquiry into the genetic underpinnings of bat biology and a potential target for developing broad-spectrum antiviral therapy.
Subject(s)
Aminohydrolases/genetics , COVID-19/genetics , Formate-Tetrahydrofolate Ligase/genetics , Methylenetetrahydrofolate Dehydrogenase (NADP)/genetics , Multienzyme Complexes/genetics , Pandemics , Aminohydrolases/antagonists & inhibitors , Animals , Antiviral Agents/therapeutic use , COVID-19/virology , Cell Line , Chiroptera/genetics , Chiroptera/virology , Formate-Tetrahydrofolate Ligase/antagonists & inhibitors , Humans , Methylenetetrahydrofolate Dehydrogenase (NADP)/antagonists & inhibitors , Minor Histocompatibility Antigens , Multienzyme Complexes/antagonists & inhibitors , RNA Viruses/genetics , SARS-CoV-2/pathogenicity , Virus Replication/genetics , COVID-19 Drug TreatmentABSTRACT
The increasing global popularity of ready-to-eat (RTE) foods for their convenience simultaneously brings along a risk, as these products can be contaminated with various microorganisms, including potentially harmful pathogens. We aimed to investigate the food contamination of Staphylococcus aureus (S. aureus) in RTE foods in Guangdong, South China. All S. aureus isolates were subjected to characterization through antimicrobial susceptibility tests, multilocus sequence typing (MLST), and PCR analysis for detecting mec and blaZ genes. A total of 824 RTE food samples were collected from 2017 to 2022, of which 73 (8.9%) were found to be contaminated with S. aureus. Contamination levels were mostly in the range of 0.3-1.0 most probable number (MPN)/g, with 10 samples exceeding 110 MPN/g. Of the 73 S. aureus isolates, 10 were identified as methicillin-resistant S. aureus (MRSA). In MRSA, resistance was most frequently observed to penicillin (100%, 10/10), followed by erythromycin (80.0%, 8/10) and tetracycline (70%, 7/10). And in methicillin-sensitive S. aureus (MSSA), resistance was most frequently observed to penicillin (98.4%, 62/63), followed by tetracycline (30.2%, 19/63) and erythromycin (23.8%, 15/63). Overall, 98.6% (72/73) of the isolates demonstrated resistance to at least one antimicrobial agent, whereas 31.5% (23/73) were resistant to three or more antimicrobials. Fifty-seven S. aureus isolates harbored the penicillin-resistant gene blaZ, and 10 isolates carried the mec gene. In addition, 30.1% of the isolates harbored genes for classical staphylococcal enterotoxins (SEs), with seb being the most frequently detected SE gene. MLST revealed that the 73 isolates belonged to 14 different sequence types (STs), the most prevalent of which was ST7. In MRSA, the most common prevalent clone is ST6, and in MSSA, ST7 was the most common isolates. The prevalent multidrug resistance indicates that the resistance situation of foodborne S. aureus in Guangdong is severe, posing a potential threat to consumer safety and health.
ABSTRACT
In December 2019, a cluster of patients with pneumonia of unknown cause was linked to a seafood wholesale market in Wuhan, China. A previously unknown betacoronavirus was discovered through the use of unbiased sequencing in samples from patients with pneumonia. Human airway epithelial cells were used to isolate a novel coronavirus, named 2019-nCoV, which formed a clade within the subgenus sarbecovirus, Orthocoronavirinae subfamily. Different from both MERS-CoV and SARS-CoV, 2019-nCoV is the seventh member of the family of coronaviruses that infect humans. Enhanced surveillance and further investigation are ongoing. (Funded by the National Key Research and Development Program of China and the National Major Project for Control and Prevention of Infectious Disease in China.).
Subject(s)
Betacoronavirus/isolation & purification , Coronavirus Infections/virology , Lung/diagnostic imaging , Pneumonia, Viral/virology , Adult , Betacoronavirus/genetics , Betacoronavirus/ultrastructure , Bronchoalveolar Lavage Fluid/virology , COVID-19 , Cells, Cultured , China , Coronavirus Infections/diagnostic imaging , Coronavirus Infections/pathology , Epithelial Cells/pathology , Epithelial Cells/virology , Female , Genome, Viral , Humans , Lung/pathology , Lung/virology , Male , Microscopy, Electron, Transmission , Middle Aged , Phylogeny , Pneumonia, Viral/diagnostic imaging , Pneumonia, Viral/pathology , Radiography, Thoracic , Respiratory System/pathology , Respiratory System/virology , SARS-CoV-2ABSTRACT
BACKGROUND: Several COVID-19 vaccines are in widespread use in China. Few data exist on comparative immunogenicity of different COVID-19 vaccines given as booster doses. We aimed to assess neutralizing antibody levels raised by injectable and inhaled aerosolized recombinant adenovirus type 5 (Ad5)-vectored COVID-19 vaccine as a heterologous booster after an inactivated COVID-19 vaccine two-dose primary series. METHODS: Using an open-label prospective cohort design, we recruited 136 individuals who had received inactivated vaccine primary series followed by either injectable or inhaled Ad5-vectored vaccine and measured neutralizing antibody titers against ancestral SARS-CoV-2 virus and Omicron BA.1 and BA.5 variants. We also measured neutralizing antibody levels in convalescent sera from 39 patients who recovered from Omicron BA.2 infection. RESULTS: Six months after primary series vaccination, neutralizing immunity against ancestral SARS-CoV-2 was low and neutralizing immunity against Omicron (B.1.1.529) was lower. Boosting with Ad5-vectored vaccines induced a high immune response against ancestral SARS-CoV-2. Neutralizing responses against Omicron BA.5 were ≥ 80% lower than against ancestral SARS-CoV-2 in sera from prime-boost subjects and in convalescent sera from survivors of Omicron BA.2 infection. Inhaled aerosolized Ad5-vectored vaccine was associated with greater neutralizing titers than injectable Ad5-vectored vaccine against ancestral and Omicron SARS-CoV-2 variants. CONCLUSIONS: These findings support the current strategy of heterologous boosting with injectable or inhaled Ad5-vectored SARS-CoV-2 vaccination of individuals primed with inactivated COVID-19 vaccine.
Subject(s)
COVID-19 Vaccines , COVID-19 , Humans , Antibodies, Neutralizing , Antibodies, Viral , Prospective Studies , SARS-CoV-2ABSTRACT
BACKGROUND: Control of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic needs effective vaccines. METHODS: In a phase 2 randomized, double-blind, placebo-controlled trial, 500 adults aged 18-59 years or ≥60 years were randomized in 2:2:1 ratio to receive 3 doses of 5 µg or 10 µg of a SARS-CoV-2 inactivated vaccine, or placebo separated by 28 days. Adverse events (AEs) were recorded through day 28 after each dosing. Live virus or pseudovirus neutralizing antibodies, and receptor binding domain immunoglobulin G (RBD-IgG) antibody were tested after the second and third doses. RESULTS: Two doses of the vaccine elicited geometric mean titers (GMTs) of 102-119, 170-176, and 1449-1617 for the 3 antibodies in younger adults. Pseudovirus neutralizing and RBD-IgG GMTs were similar between older and younger adults. The third dose slightly (<1.5 fold) increased GMTs. Seroconversion percentages were 94% or more after 2 doses, which were generally similar after 3 doses. The predominant AEs were injection-site pain. All the AEs were grade 1 or 2 in intensity. No serious AE was deemed related to study vaccination. CONCLUSIONS: Two doses of this vaccine induced robust immune response and had good safety profile. A third dose given 28 days after the second dose elicited limited boosting antibody response.
Subject(s)
COVID-19 Vaccines , COVID-19 , Immunogenicity, Vaccine , Adolescent , Adult , Antibodies, Neutralizing/blood , Antibodies, Viral/blood , COVID-19/prevention & control , COVID-19 Vaccines/adverse effects , COVID-19 Vaccines/immunology , Double-Blind Method , Humans , Immunoglobulin G/blood , Middle Aged , SARS-CoV-2 , Vaccines, Inactivated/adverse effects , Vaccines, Inactivated/immunology , Young AdultABSTRACT
BACKGROUND: The cesarean delivery (CD) rate has been increasing globally. Trial of labor after cesarean delivery (TOLAC) has been used as a key method for the reduction of the CD rate. Little is known, however, about the association between the second-stage duration of TOLAC and adverse maternal and neonatal outcomes. This study evaluated the association between perinatal outcomes and the duration of second-stage labor in women undergoing TOLAC. METHODS: A 10-year retrospective cohort study was performed at the Department of Obstetrics and Gynecology, Third Affiliated Hospital of Guangzhou Medical University, between January 2010 and January 2020. Women undergoing TOLAC who reached the second stage of labor were included in this study. Duration of the second stage of labor was examined as a categorical variable (group I: <0.5 h, group II: 0.5-2 h and group III: ≥2 h) and as a continuous variable to evaluate the association with adverse perinatal outcomes by using multivariable regression models and a Cox proportional hazards regression model adjusting for potential confounders. RESULTS: Of the 1,174 women who met the inclusion criteria, the median (interquartile range) length of the second stage was 0.5 h (0.3-0.9 h). Among them, 1,143 (97.4%) delivered vaginally and 31 underwent an unplanned CD. As the second-stage duration increased, operative vaginal delivery (OVD), CD, and postpartum hemorrhage (PPH) rates increased. Women in group III had higher risks of OVD (aOR = 11.34; 95% CI [5.06-25.41]), CD (aOR = 4.22; 95% CI [1.32-13.43]), and PPH (aOR = 2.43; 95% CI [1.31-4.50]) compared with group I. Correspondingly, blood loss and the oxytocin used to treat PPH increased significantly, while the postpartum hemoglobin reduced significantly in group III compared with group I. The incidence of uterine rupture, uterine atony, cervical laceration, red blood cell transfusion, and intensive care unit admission were similar in all three groups. Neonatal outcomes were not affected by the second-stage duration. CONCLUSIONS: Women undergoing TOLAC with second-stage duration of ≥2 h have higher odds of OVD, unplanned intrapartum CD, and PPH.
Subject(s)
Postpartum Hemorrhage , Trial of Labor , Cesarean Section , Female , Humans , Infant, Newborn , Labor Stage, Second , Parturition , Postpartum Hemorrhage/epidemiology , Postpartum Hemorrhage/etiology , Pregnancy , Retrospective StudiesABSTRACT
With increasing demand for customized commodities and the greater insight and understanding of olfaction, the synthesis of fragrances with diverse structures and odor characters has become a core task. Recent progress in organic synthesis and catalysis enables the rapid construction of carbocycles and heterocycles from readily available unsaturated molecular building blocks, with increased selectivity, atom economy, sustainability and product diversity. In this review, synthetic methods for creating cyclic fragrances, including both natural and synthetic ones, will be discussed, with a focus on the key transformations of alkenes, alkynes, dienes and enynes. Several strategies will be discussed, including cycloaddition, catalytic cyclization, ring-closing metathesis, intramolecular addition, and rearrangement reactions. Representative examples and the featured olfactory investigations will be highlighted, along with some perspectives on future developments in this area.
Subject(s)
Alkenes , Alkynes , Alkenes/chemistry , Alkynes/chemistry , Catalysis , Cyclization , Cycloaddition Reaction , OdorantsABSTRACT
BACKGROUND: In late December, 2019, patients presenting with viral pneumonia due to an unidentified microbial agent were reported in Wuhan, China. A novel coronavirus was subsequently identified as the causative pathogen, provisionally named 2019 novel coronavirus (2019-nCoV). As of Jan 26, 2020, more than 2000 cases of 2019-nCoV infection have been confirmed, most of which involved people living in or visiting Wuhan, and human-to-human transmission has been confirmed. METHODS: We did next-generation sequencing of samples from bronchoalveolar lavage fluid and cultured isolates from nine inpatients, eight of whom had visited the Huanan seafood market in Wuhan. Complete and partial 2019-nCoV genome sequences were obtained from these individuals. Viral contigs were connected using Sanger sequencing to obtain the full-length genomes, with the terminal regions determined by rapid amplification of cDNA ends. Phylogenetic analysis of these 2019-nCoV genomes and those of other coronaviruses was used to determine the evolutionary history of the virus and help infer its likely origin. Homology modelling was done to explore the likely receptor-binding properties of the virus. FINDINGS: The ten genome sequences of 2019-nCoV obtained from the nine patients were extremely similar, exhibiting more than 99·98% sequence identity. Notably, 2019-nCoV was closely related (with 88% identity) to two bat-derived severe acute respiratory syndrome (SARS)-like coronaviruses, bat-SL-CoVZC45 and bat-SL-CoVZXC21, collected in 2018 in Zhoushan, eastern China, but were more distant from SARS-CoV (about 79%) and MERS-CoV (about 50%). Phylogenetic analysis revealed that 2019-nCoV fell within the subgenus Sarbecovirus of the genus Betacoronavirus, with a relatively long branch length to its closest relatives bat-SL-CoVZC45 and bat-SL-CoVZXC21, and was genetically distinct from SARS-CoV. Notably, homology modelling revealed that 2019-nCoV had a similar receptor-binding domain structure to that of SARS-CoV, despite amino acid variation at some key residues. INTERPRETATION: 2019-nCoV is sufficiently divergent from SARS-CoV to be considered a new human-infecting betacoronavirus. Although our phylogenetic analysis suggests that bats might be the original host of this virus, an animal sold at the seafood market in Wuhan might represent an intermediate host facilitating the emergence of the virus in humans. Importantly, structural analysis suggests that 2019-nCoV might be able to bind to the angiotensin-converting enzyme 2 receptor in humans. The future evolution, adaptation, and spread of this virus warrant urgent investigation. FUNDING: National Key Research and Development Program of China, National Major Project for Control and Prevention of Infectious Disease in China, Chinese Academy of Sciences, Shandong First Medical University.
Subject(s)
Betacoronavirus/genetics , Coronavirus Infections/epidemiology , Coronavirus Infections/virology , Genome, Viral , Pneumonia, Viral/epidemiology , Pneumonia, Viral/virology , Receptors, Virus/metabolism , Betacoronavirus/metabolism , Bronchoalveolar Lavage Fluid/virology , COVID-19 , China/epidemiology , Coronavirus Infections/diagnosis , Coronavirus Infections/transmission , DNA, Viral/genetics , Disease Reservoirs/virology , Genomics/methods , High-Throughput Nucleotide Sequencing/methods , Humans , Phylogeny , Pneumonia, Viral/diagnosis , Pneumonia, Viral/transmission , SARS-CoV-2 , Sequence AlignmentABSTRACT
COVID-19 has become a global pandemic and there is an urgent call for developing drugs against the virus (SARS-CoV-2). The 3C-like protease (3CLpro) of SARS-CoV-2 is a preferred target for broad spectrum anti-coronavirus drug discovery. We studied the anti-SARS-CoV-2 activity of S. baicalensis and its ingredients. We found that the ethanol extract of S. baicalensis and its major component, baicalein, inhibit SARS-CoV-2 3CLpro activity in vitro with IC50's of 8.52 µg/ml and 0.39 µM, respectively. Both of them inhibit the replication of SARS-CoV-2 in Vero cells with EC50's of 0.74 µg/ml and 2.9 µM, respectively. While baicalein is mainly active at the viral post-entry stage, the ethanol extract also inhibits viral entry. We further identified four baicalein analogues from other herbs that inhibit SARS-CoV-2 3CLpro activity at µM concentration. All the active compounds and the S. baicalensis extract also inhibit the SARS-CoV 3CLpro, demonstrating their potential as broad-spectrum anti-coronavirus drugs.
Subject(s)
Antiviral Agents/pharmacology , COVID-19 Drug Treatment , Coronavirus 3C Proteases/antagonists & inhibitors , Flavanones/pharmacology , Plant Extracts/pharmacology , Protease Inhibitors/pharmacology , SARS-CoV-2/drug effects , Virus Replication/drug effects , Animals , COVID-19/enzymology , COVID-19/virology , Chlorocebus aethiops , Drug Discovery , Enzyme Inhibitors/pharmacology , Humans , In Vitro Techniques , Models, Molecular , SARS-CoV-2/enzymology , Scutellaria baicalensis , Vero CellsABSTRACT
BACKGROUND: The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) first broke out in 2019 and subsequently spread worldwide. Chloroquine has been sporadically used in treating SARS-CoV-2 infection. Hydroxychloroquine shares the same mechanism of action as chloroquine, but its more tolerable safety profile makes it the preferred drug to treat malaria and autoimmune conditions. We propose that the immunomodulatory effect of hydroxychloroquine also may be useful in controlling the cytokine storm that occurs late phase in critically ill patients with SARS-CoV-2. Currently, there is no evidence to support the use of hydroxychloroquine in SARS-CoV-2 infection. METHODS: The pharmacological activity of chloroquine and hydroxychloroquine was tested using SARS-CoV-2-infected Vero cells. Physiologically based pharmacokinetic (PBPK) models were implemented for both drugs separately by integrating their in vitro data. Using the PBPK models, hydroxychloroquine concentrations in lung fluid were simulated under 5 different dosing regimens to explore the most effective regimen while considering the drug's safety profile. RESULTS: Hydroxychloroquine (EC50 = 0.72 µM) was found to be more potent than chloroquine (EC50 = 5.47 µM) in vitro. Based on PBPK models results, a loading dose of 400 mg twice daily of hydroxychloroquine sulfate given orally, followed by a maintenance dose of 200 mg given twice daily for 4 days is recommended for SARS-CoV-2 infection, as it reached 3 times the potency of chloroquine phosphate when given 500 mg twice daily 5 days in advance. CONCLUSIONS: Hydroxychloroquine was found to be more potent than chloroquine to inhibit SARS-CoV-2 in vitro.
Subject(s)
Antiviral Agents/pharmacology , Betacoronavirus/drug effects , Coronavirus Infections/drug therapy , Hydroxychloroquine/pharmacology , Pneumonia, Viral/drug therapy , Severe Acute Respiratory Syndrome/drug therapy , Animals , Antiviral Agents/pharmacokinetics , COVID-19 , Cell Line , Chlorocebus aethiops , Chloroquine/pharmacokinetics , Chloroquine/pharmacology , Hydroxychloroquine/pharmacokinetics , Lung/drug effects , Pandemics , SARS-CoV-2 , Vero Cells , COVID-19 Drug TreatmentABSTRACT
Coronaviruses (CoVs) act as cross-species viruses and have the potential to spread rapidly into new host species and cause epidemic diseases. Despite the severe public health threat of severe acute respiratory syndrome coronavirus and Middle East respiratory syndrome CoV (MERS-CoV), there are currently no drugs available for their treatment; therefore, broad-spectrum inhibitors of emerging and endemic CoVs are urgently needed. To search for effective inhibitory agents, we performed high-throughput screening (HTS) of a 2,000-compound library of approved drugs and pharmacologically active compounds using the established genetically engineered human CoV OC43 (HCoV-OC43) strain expressing Renilla luciferase (rOC43-ns2Del-Rluc) and validated the inhibitors using multiple genetically distinct CoVs in vitro We screened 56 hits from the HTS data and validated 36 compounds in vitro using wild-type HCoV-OC43. Furthermore, we identified seven compounds (lycorine, emetine, monensin sodium, mycophenolate mofetil, mycophenolic acid, phenazopyridine, and pyrvinium pamoate) as broad-spectrum inhibitors according to their strong inhibition of replication by four CoVs in vitro at low-micromolar concentrations. Additionally, we found that emetine blocked MERS-CoV entry according to pseudovirus entry assays and that lycorine protected BALB/c mice against HCoV-OC43-induced lethality by decreasing viral load in the central nervous system. This represents the first demonstration of in vivo real-time bioluminescence imaging to monitor the effect of lycorine on the spread and distribution of HCoV-OC43 in a mouse model. These results offer critical information supporting the development of an effective therapeutic strategy against CoV infection.IMPORTANCE Currently, there is no approved therapy to treat coronavirus infection; therefore, broad-spectrum inhibitors of emerging and endemic CoVs are needed. Based on our high-throughput screening assay using a compound library, we identified seven compounds with broad-spectrum efficacy against the replication of four CoVs in vitro Additionally, one compound (lycorine) was found to protect BALB/c mice against HCoV-OC43-induced lethality by decreasing viral load in the central nervous system. This inhibitor might offer promising therapeutic possibilities for combatting novel CoV infections in the future.
Subject(s)
Coronavirus OC43, Human/drug effects , Coronavirus/drug effects , Amaryllidaceae Alkaloids/pharmacology , Animals , Antiviral Agents/pharmacology , Cell Line , Coronavirus/pathogenicity , Coronavirus OC43, Human/metabolism , Drug Evaluation, Preclinical/methods , Emetine/pharmacology , High-Throughput Screening Assays/methods , Humans , Mice , Mice, Inbred BALB C , Middle East Respiratory Syndrome Coronavirus/drug effects , Phenanthridines/pharmacology , Severe acute respiratory syndrome-related coronavirus/drug effectsABSTRACT
BACKGROUND: The prevalence of migraines in the She population, a minority in China, is significantly higher than that in Han Chinese and other Asian populations. Two single nucleotide polymorphisms (SNPs) have been found to be associated with migraine susceptibility in the She population. PURPOSE: This study investigated four SNPs, identified in genome-wide association studies, within migraine-susceptible loci in Han Chinese for their association with migraine susceptibility in the She population. METHODS: Two-hundred unrelated migraine patients and 200 healthy controls were recruited. The SNPs examined included rs2651899 (PRDM16 ), rs2274316 (MEF2D ), rs7577262 (TRPM8) and rs11172113 (LRP1). Genotyping of the SNPs was performed by allele-specific polymerase chain reaction and direct sequencing. RESULTS: No significant differences between the participants with migraines and controls (participants without migraines) were demonstrated in genotypes, alleles and allele carriage frequencies for the four SNPs. A subgroup analysis found that migraine with aura had a lower frequency of C allele positivity in rs2651899 than in healthy controls (59.6% vs. 74.5%, respectively; P < 0.034). Univariate analyses indicated that no genotype of the four SNPs had a significant association with migraines. Males had a lower risk of migraines, and advanced age was a significant risk factor for migraines in females. CONCLUSION: The SNPs in four migraine susceptible loci in Han Chinese were not risk factors for migraines in a relatively small sample of the She population.
Subject(s)
Genome-Wide Association Study/methods , Polymorphism, Single Nucleotide/genetics , Adult , Arabidopsis Proteins/genetics , Asian People , Case-Control Studies , China/epidemiology , DNA-Binding Proteins/genetics , Female , Genetic Predisposition to Disease/genetics , Genotype , Humans , MEF2 Transcription Factors/genetics , Male , Middle Aged , Risk Factors , TRPM Cation Channels/genetics , Transcription Factors/geneticsABSTRACT
Background: The Middle East respiratory syndrome coronavirus (MERS-CoV) causes severe respiratory infection with a high (~35%) mortality rate. Neutralizing antibodies targeting the spike of MERS-CoV have been shown to be a therapeutic option for treatment of lethal disease. Methods: We describe the germline diversity and neutralizing activity of 13 potent human monoclonal antibodies (mAbs) that target the MERS-CoV spike (S) protein. Biological functions were assessed by live MERS-CoV, pseudotype particle and its variants, and structural basis was also determined by crystallographic analysis. Results: Of the 13 mAbs displaying strong neutralizing activity against MERS-CoV, two with the immunoglobulin heavy-chain variable region (IGHV)1-69-derived heavy chain (named MERS-GD27 and MERS-GD33) showed the most potent neutralizing activity against pseudotyped and live MERS-CoV in vitro. Mutagenesis analysis suggested that MERS-GD27 and MERS-GD33 recognized distinct regions in S glycoproteins, and the combination of 2 mAbs demonstrated a synergistic effect in neutralization against pseudotyped MERS-CoV. The structural basis of MERS-GD27 neutralization and recognition revealed that its epitope almost completely overlapped with the receptor-binding site. Conclusions: Our data provide new insights into the specific antibody repertoires and the molecular determinants of neutralization during natural MERS-CoV infection in humans. This finding supports additional efforts to design and develop novel therapies to combat MERS-CoV infections in humans.
Subject(s)
Antibodies, Neutralizing/immunology , Antibodies, Viral/immunology , Coronavirus Infections/immunology , Coronavirus Infections/virology , Middle East Respiratory Syndrome Coronavirus/immunology , Spike Glycoprotein, Coronavirus/immunology , Adult , Antibodies, Monoclonal/immunology , Antibody Specificity , Enzyme-Linked Immunosorbent Assay , Epitope Mapping , Humans , Male , Models, Molecular , Protein ConformationABSTRACT
OBJECTIVE: To investigate the association between alanine aminotransferase (ALT) levels and risk of osteopenia in middle-aged and elderly Chinese with ALT within the normal range. METHODS: This was a cross-sectional study. A total of 4,890 men and women (pre- and postmenopausal) aged 40 years or older were randomly recruited from Fujian, China. Each participant was required to complete a questionnaire and then undergo anthropometric, biochemical, and bone mineral density measurements. RESULTS: The odds ratio of osteopenia decreased significantly with increasing ALT level at baseline. The three groups (men, pre- and postmenopausal women) were divided by ALT quartiles. In multiple logistic regression models using the first quartile as the reference, after adjusting for corresponding confounding factors, the odds ratios of osteopenia across the other ALT quartiles were 0.576 (95% confidence interval [CI], 0.390 to 0.851), 0.654 (95% CI, 0.460 to 0.930), and 0.629 (95% CI, 0.427 to 0.926) for premenopausal women, and 0.949 (95% CI, 0.699 to 1.289), 0.733 (95% CI, 0.540 to 0.995), and 0.692 (95% CI, 0.508 to 0.943) for postmenopausal women (not significant for quartile 2). However, no significantly different results were found in men. Multiple linear regression models showed that serum ALT concentrations were positively associated with the homeostasis model assessment of insulin resistance. CONCLUSION: Our study of middle-aged and elderly Chinese men and women demonstrates that the prevalence of osteopenia is inversely associated with ALT level when ALT is within the normal range.
Subject(s)
Alanine Transaminase/blood , Bone Diseases, Metabolic/etiology , Aged , Bone Diseases, Metabolic/enzymology , Cross-Sectional Studies , Female , Humans , Logistic Models , Male , Middle AgedABSTRACT
A global outbreak of monkeypox (mpox) caused by the mpox virus (MPXV) has posed a serious threat to public health worldwide, thus calling for the urgent development of antivirals and vaccines to curb its further spread. In this study, we screened 41 anhydride-modified proteins and found that 3-hydroxyphthalic anhydride-modified ß-lactoglobulin (3HP-ß-LG), a clinically used anti-HPV agent, was highly effective in inhibiting infection of vaccinia virus Tiantan strain (VACV-VTT) and MPXV. Mechanistic studies demonstrated that 3HP-ß-LG bound to the virus, not the host cell, by targeting the early stage of virus entry, possibly through the interaction between the amino acids with negatively charges in 3HP-ß-LG and the key amino acids with positive charges in the target region of A29L, a key surface protein of MPXV. A synergistic effect was observed when 3HP-ß-LG was combined with tecovirimat, a small-molecule antiviral drug approved by the United States Food and Drug Administration and the European Medicine Agency for the treatment of smallpox and mpox. Because of its clinically proven safety and stability, 3HP-ß-LG shows promise for further development as a prophylactic agent to prevent the sexual transmission of MPXV.
ABSTRACT
Since May 2022, the multi-country outbreak of monkeypox (mpox) has raised a great concern worldwide. Early detection of mpox virus infection is recognized as an efficient way to prevent mpox transmission. Mpox specific detection methods reported up to now are based on the SNPs among mpox virus and other orthopoxviruses. We have therefore developed a real-time PCR based mpox detection method targeting mpox virus specific sequences (N3R and B18Rplus). We have also optimized an orthopoxvirus detection system which targets the highly conserved E9L and D6R genes. The mpox and orthopoxvirus real-time PCR assays have a high sensitivity (1 copy/reaction) and specificity. Mpox viral DNA and clinical samples from mpox patients are detected with the mpox detection system. Furthermore, we have established a multiplex real-time PCR detection system allowing simultaneous and efficient detection of mpox and orthopoxvirus infections.
Subject(s)
Monkeypox virus , Mpox (monkeypox) , Multiplex Polymerase Chain Reaction , Orthopoxvirus , Poxviridae Infections , Real-Time Polymerase Chain Reaction , Sensitivity and Specificity , Orthopoxvirus/genetics , Orthopoxvirus/isolation & purification , Humans , Real-Time Polymerase Chain Reaction/methods , Multiplex Polymerase Chain Reaction/methods , Monkeypox virus/genetics , Monkeypox virus/isolation & purification , Poxviridae Infections/diagnosis , Poxviridae Infections/virology , Poxviridae Infections/veterinary , Mpox (monkeypox)/diagnosis , Mpox (monkeypox)/virology , Molecular Diagnostic Techniques/methodsABSTRACT
The monkeypox virus (MPXV) has triggered a current outbreak globally. Genome sequencing of MPXV and rapid tracing of genetic variants will benefit disease diagnosis and control. It is a significant challenge but necessary to optimize the strategy and application of rapid full-length genome identification and to track variations of MPXV in clinical specimens with low viral loads, as it is one of the DNA viruses with the largest genome and the most AT-biased, and has a significant number of tandem repeats. Here we evaluated the performance of metagenomic and amplicon sequencing techniques, and three sequencing platforms in MPXV genome sequencing based on multiple clinical specimens of five mpox cases in Chinese mainland. We rapidly identified the full-length genome of MPXV with the assembly of accurate tandem repeats in multiple clinical specimens. Amplicon sequencing enables cost-effective and rapid sequencing of clinical specimens to obtain high-quality MPXV genomes. Third-generation sequencing facilitates the assembly of the terminal tandem repeat regions in the monkeypox virus genome and corrects a common misassembly in published sequences. Besides, several intra-host single nucleotide variations were identified in the first imported mpox case. This study offers an evaluation of various strategies aimed at identifying the complete genome of MPXV in clinical specimens. The findings of this study will significantly enhance the surveillance of MPXV.
Subject(s)
Monkeypox virus , Mpox (monkeypox) , Humans , Monkeypox virus/genetics , Mpox (monkeypox)/diagnosisABSTRACT
Abnormal trophoblast self-renewal and differentiation during early gestation is the major cause of miscarriage, yet the underlying regulatory mechanisms remain elusive. Here, we show that trophoblast specific deletion of Kat8, a MYST family histone acetyltransferase, leads to extraembryonic ectoderm abnormalities and embryonic lethality. Employing RNA-seq and CUT&Tag analyses on trophoblast stem cells (TSCs), we further discover that KAT8 regulates the transcriptional activation of the trophoblast stemness marker, CDX2, via acetylating H4K16. Remarkably, CDX2 overexpression partially rescues the defects arising from Kat8 knockout. Moreover, increasing H4K16ac via using deacetylase SIRT1 inhibitor, EX527, restores CDX2 levels and promoted placental development. Clinical analysis shows reduced KAT8, CDX2 and H4K16ac expression are associated with recurrent pregnancy loss (RPL). Trophoblast organoids derived from these patients exhibit impaired TSC self-renewal and growth, which are significantly ameliorated with EX527 treatment. These findings suggest the therapeutic potential of targeting the KAT8-H4K16ac-CDX2 axis for mitigating RPL, shedding light on early gestational abnormalities.