Establishment and evaluation of a genome-wide multiplex PCR sequencing method for severe fever with thrombocytopenia syndrome virus based on high-throughput sequencing technology / 中华微生物学和免疫学杂志

Objective:To establish a sequencing method for the genome of severe fever with thrombocytopenia syndrome virus (SFTSV) based on next-generation sequencing (NGS).Methods:SFTSV RNA was extracted from serum samples of patients with severe fever with thrombocytopenia syndrome. SFTSV-specific primers were designed using Primer 5.0 software. A multiplex PCR method was constructed and used to amplify the nucleotide sequence of SFTSV. Whole-genome sequencing was performed on the NGS platform.Results:The whole genes of SFTSV isolates in 28 serum samples were amplified by the multiplex PCR with a coverage over 94%. Sequencing and phylogenetic analysis of those strains revealed that the predominant strains ( n=20) belonged to genotype A, followed by genotypes B ( n=4) and E ( n=3). Conclusions:A high-throughput sequencing method for SFTSV based on multiplex PCR was established in this study. This method was characterized by high specificity and good quality and could improve the sequencing efficiency.

Genome characterization and traceability analysis of SARS-CoV-2 in a local outbreak / 中华微生物学和免疫学杂志

Objective:To analyze the genome characteristics and variations in nucleotides and amino acids of SARS-CoV-2 causing an outbreak in Henan Province in November 2021 and perform the traceability analysis.Methods:In this study, throat swab specimens from cases in the acute phase were collected and tested for the nucleic acids of SARS-CoV-2 by real-time fluorescent RT-PCR. SARS-CoV-2 nucleic acid-positive samples were subjected to high-throughput genome sequencing and whole-genome alignment analysis.Results:The median Ct values of ORF1ab gene and N gene in 70 positive specimens was 26.41 (15.58 to 39.27) and 24.43 (12.04 to 39.74), respectively. Compared with the sequence of Wuhan-Hu(NC_045512) reference strain, 47 to 49 nucleotide mutations sharing 47 nucleotide mutation and 41 amino acid mutations were found in 63 strains of successfully sequenced SARS-CoV-2. Nine nucleotide mutations and 12 amino acid mutations were found in the spike protein. The index case shared 47 mutations with the Russian imported cases in Henan Province on October 14 and the local cases in Jiangxi Province in October. Moreover, their genomes were highly homologous and they all belonged to the Delta variant (AY.122 evolutionary branch).Conclusions:Continuous monitoring of imported COVID-19 cases and prolonging the period of quarantine were needed to reduce the risk of local outbreak and epidemic caused by imported COVID-19 cases. Analysis of the genomic characteristics of SARS-CoV-2 and the variations in nucleotides and amino acids was conducive to trace the origin of COVID-19 outbreak quickly and provide reference for precise control.

Analysis of the dynamic changes of serum IgM and IgG antibodies of the COVID-19 patients / 中华微生物学和免疫学杂志

Objective:To monitor the changes in specific IgM and IgG antibodies in patients diagnosed with COVID-19 after SARS-CoV-2 infection, and analyze their clinical significance.Methods:A total of 168 serum samples were collected from 56 COVID-19 patients with different disease courses who were positive for nucleic acid test at Henan Center for Disease Control and Prevention on January 8, 2020 and February 21, 2020. Serum samples from 25 healthy people excluded from COVID-19 were used as control group. IgM and IgG antibodies against SARS-CoV-2 were detected by chemiluminescence method.Results:IgM antibody increased sharply in 1-3 weeks after onset, and reached the peak value (21.78 AU/ml) in the 3rd week after onset. IgG antibody increased the most in 3-6 weeks after onset, and reached the peak value (81.58 AU/ml) in the 9th week after onset. The levels of IgM and IgG antibodies were closely correlated with age and disease course ( P<0.05). The antibody level of 30-60 years old group was the highest, the IgM antibody positive rate and antibody level of acute stage and previous infection were lower than that of recovery stage, and the IgG antibody positive rate and antibody level of acute stage were lower than that of recovery stage and previous infection. During the whole course of the disease, the levels of IgM and IgG antibodies increased gradually in the acute stage, reached the peak in the recovery stage, and decreased and maintained at a certain level in the past infection. Conclusions:Serum SARS-CoV-2 IgM and IgG antibody detection can be used as auxiliary diagnostic indicators for COVID-19, and its continuous observation is helpful for epidemiological investigation, serological diagnosis and disease course monitoring.

Genomic characteristics of 2019 novel coronavirus from imported cases of coronavirus disease 2019 in Henan Province / 中华传染病杂志

Objective:To analyze the evolutionary characteristics and variations of 2019 novel coronavirus (2019-nCoV) strains imported from abroad in Henan Province.Methods:A total of 16 imported cases of coronavirus disease 2019 (COVID-19) reported in Henan Province from May to December 2020 were enrolled. The throat swab specimens from the patients were collected and sent to the Henan Provincial Center for Disease Control and Prevention for whole genome sequencing. Taking SARS-CoV-2 Wuhan-Hu-1 published in Global Initiative on Sharing All Influenza Data (GISAID) as the reference sequence, the sequences were aligned and analyzed by MEGA X, and the phylogenetic tree was constructed by the maximum likelihood method.Results:Among 16 cases, 13 cases were imported from Russia, two cases were imported from Myanmar, and one case was imported from Ukraine. A total of 16 strains of 2019-nCoV genomes with the lengths of 29 804 bp to 29 882 bp were obtained. A total of 145 nucleotide mutations and 80 amino acid mutations were detected. Nucleotide variations of C241T, C3037T, C14408T, A23403G and the amino acid variation of D614G in spike protein were detected in all sequences. Meanwhile, insertion A at the site of 29704 was found in BetaCov/HEN02/Human/2020, BetaCov/HEN04/Human/2020 and BetaCov/HEN05/Human/2020. Deletion variation was not found. Phylogenetic analysis showed that there was no correlation between the 16 strains and currently epidemic variants of concern (VOC) .Conclusion:From May to December 2020, the detection of viral genome mutations in the imported cases of Henan Province shows randomness and diversity, while the strains are not VOC.

Safety and immunogenicity of seasonal inactivated influenza vaccine (split virion) and cross-reactive antibody responses to the H7N9 avian influenza virus / 中华流行病学杂志

<p><b>OBJECTIVE</b>To evaluate the safety and immunogenicity of seasonal inactivated influenza vaccine (split virion) and to analyze its cross-reactive antibody responses to H7N9 avian influenza virus.</p><p><b>METHODS</b>An open-labeled clinical trial was carried out in infants aged 6-35 months, adults aged 18-60 years and the elderly aged >60 years. After vaccinations (one dose for adults and the elderly and two doses for infants), adverse events were observed. Serum samples were obtained before vaccination and 21 days after vaccination from adults and elderly subjects. Three types of antibody against seasonal influenza virus and antibody against H7N9 avian influenza virus were tested using microhemagglutination inhibition (HI) assay. Immunogenicity of the vaccine was evaluated based on the immunogenicity criteria for adults and the elderly, set by the Committee for Medicinal Products for Human Use (CHMP) for the European Medicines Agency.</p><p><b>RESULTS</b>A total of 202 subjects (65 infants, 69 adults and 68 elderly) were enrolled and injected for at least one dose. The overall rate of adverse events was 12.4% (25/202) and most of them were under systemic reaction. No serious adverse event was reported. Pre- and post-vaccination serum samples were collected from 124 subjects (64 adults, 60 elderly). After 21 days of vaccination, the sero-conversion rate, sero-protection rate, and geometric mean titer (GMT) ratio (post-/pre-vaccination) of antibody against seasonal influenza virus were 78.1%-90.6%, 92.2%-100.0% and 7.9-41.0 among adults while 66.7%-83.3%, 86.7%-100.0% and 5.7-20.4 among the elderly, respectively. However, after vaccination, both sero-conversion rate and sero-protection rate of antibody against H7N9 avian influenza virus among adults and the elderly became zero, with GMT ratio between 1.2 and 1.4.</p><p><b>CONCLUSION</b>This trial vaccine appeared to have good safety and immunogenicity but inducing no cross-reactive antibody response to H7N9 avian influenza virus.</p>

Studies on the safety and immunogenicity of lyophilized live attenuated measles-mumps-rubella combined vaccine / 中华流行病学杂志

<p><b>OBJECTIVE</b>To study the safety and immunogenicity of the measles-mumps-rubella combined vaccine (MMR) produced by Beijing Biological Product Institute.</p><p><b>METHODS</b>Children aged 10-12 years, 2-2.5 years and 8-12 months were selected to be vaccinated with Beijing MMR vaccine (test vaccine). Other groups of children with similar nature were vaccinated with measles vaccine, mumps vaccine and rubella vaccine while using imported MMR vaccine (control vaccine) as controls.</p><p><b>RESULTS</b>The safety of the Beijing MMR vaccine was confirmed after vaccinating 32 children above 2 years old. Among 104 children of 8-12 months were vaccinated with Beijing MMR vaccine, only 6.7% of the children had transient fever and 1.9% had signs of rashes but with no other signs observed. The positive seroconversion rates of measles, rubella and mumps anti-HI were 100%, 100% and 85.7% respectively. GMT also showed high lever.</p><p><b>CONCLUSION</b>The MMR vaccine (Beijing) had good safety and immunogenicity which might be used to be the bases enhance immunization of measles.</p>