An in-depth investigation of the safety and immunogenicity of an inactivated SARS-CoV-2 vaccine

BACKGROUNDIn-depth investigations of the safety and immunogenicity of inactivated SARS-CoV-2 vaccines are needed. METHODIn a phase I randomized, double-blinded, and placebo-controlled trial involving 192 healthy adults 18-59 years of age, two injections of three different doses (50 EU, 100 EU and 150 EU) of an inactivated SARS-CoV-2 vaccine or the placebo were administered intramuscularly with a 2- or 4-week interval between the injections. The safety and immunogenicity of the vaccine were evaluated within 28 days. FINDINGIn this study, 191 subjects assigned to three doses groups or the placebo group completed the 28-day trial. There were 44 adverse reactions within the 28 days, most commonly mild pain and redness at the injection site or slight fatigue, and no abnormal variations were observed in 48 cytokines in the serum samples of immunized subjects. The serum samples diluted from 1:32 to 1:4096 and incubated with the virus did not show antibody-dependent enhancement effects (ADEs) with regard to human natural killer cells, macrophages or dendritic cells. At day 14, the seroconversion rates had reached 92%, 100% and 96% with geometric mean titers (GMTs) of 18.0, 54.5 and 37.1, and at day 28, the seroconversion rates had reached 80%, 96% and 92% with GMTs of 10.6, 15.4 and 19.6in 0, 14 and 0, 28 procedures, respectively. Seroconversion was associated with the synchronous upregulation of ELISA antibodies against the S protein, N protein and virion and a cytotoxic T lymphocyte (CTL) response. Transcriptome analysis shaped the genetic diversity of immune response induced by the vaccine. INTERPRETATIONIn a population aged 18-59 years, this inactivated SARS-CoV-2 vaccine was safe and immunogenic. Trial registrationNCT04412538 FUNDINGThe National Key R&D Program of China (2020YFC0849700), the Program of Chinese Academy of Medicine Science and the Major Science and Technology Special Projects of Yunnan Province.

A valid protective immune response elicited in rhesus macaques by an inactivated vaccine is capable of defending against SARS-CoV-2 infection

With the relatively serious global epidemic outbreak of SARS-CoV-2 infection, public concerns focus on not only clinical therapeutic measures and public quarantine for this disease but also the development of vaccines. The technical design of our SARS-CoV-2 inactivated vaccine provides a viral antigen that enables the exposure of more than one structural protein based upon the antibody composition of COVID-19 patients convalescent serum. This design led to valid immunity with increasing neutralizing antibody titers and a CTL response detected post-immunization of this vaccine by two injections in rhesus macaques. Further, this elicited immunoprotection in macaques enables not only to restrain completely viral replication in tissues of immunized animals, compared to the adjuvant control and those immunized by an RBD peptide vaccine, but also to significantly alleviate inflammatory lesion in lung tissues in histo-pathologic detection, compared to the adjuvant control with developed interstitial pneumonia. The data obtained from these macaques immunized with the inactivated vaccine or RBD peptide vaccine suggest that immunity with a clinically protective effect against SARS-CoV-2 infection should include not only specific neutralizing antibodies but also specific CTL responses against at least the S and N antigens.

Biological characteristics of coxsackievirus A6 clinical isolates / 中华微生物学和免疫学杂志

Objective To analyze the biological characteristics of clinical isolates of coxsackievir-us A6 (CVA6), a pathogen of hand,foot and mouth disease (HFMD), and to provide reference for vaccine development. Methods CVA6 strains were isolated from 21 stool and throat swab specimens of patients with HFMD in Yunnan Province and then identified. Their growth characteristics, plaque morphology and virulence to suckling mice were analyzed. Results Five CVA6 strains, named CVA6-129, CVA6-113, CVA6-57, CVA6-94 and CVA6-162, were isolated and all belonged to D3 subtype. Only the CVA6-129 strain could proliferate rapidly in Vero and KMB17 cells and the proliferation peaked 30 h after inoculation. The infectious titer of the CVA6-129 strain was 7. 54 lgCCID50 (50％ cell culture infective dose) / ml in KMB17 cells. Different morphologies of plaques were formed by the CVA6-129 strain in Vero and KMB17 cells at the same time points, which were small and round with clear edges in Vero cells, and large and irregular with blurry edges in KMB17 cells. Suckling mice were susceptible to CVA6 via intramuscular and intraperito-neal injection. The most common symptoms in infected suckling mice were reduced mobility, hind limb pa-ralysis and quadriplegia. CVA6 infection could result in death in severe cases. Conclusions This study isolated five CVA6 strains from a number of clinical samples of suspected HFMD cases, of which the CVA6-129 strain showed potential as a vaccine candidate.

Biological characteristics of different epidemic enterovirus 71 strains and their pathogeneses in neonatal mice and rhesus monkeys.

Hand, foot and mouth disease (HFMD) has been prevalent in China since 2008. Enterovirus 71 (EV71) is a common causative agent of HFMD, and various strains of EV71 are prevalent worldwide. The EV71C4 subgenotype is the most endemic strain in China. However, few studies investigating the biological characteristics and pathogeneses of different C4 strains have been reported. Therefore, the current study investigated 19 clinical EV71 strains in neonatal ICR mice and neonatal rhesus monkeys by comparing pathogenicity; the virulence of different viral passages, dosages, and routes of infection; and the effects produced by subject animal age. These 19 clinical EV71 strains, which were of the same subtype, displayed varying pathogenic effects. Three strains (HE31, 231 and 262) induced limb paralysis in neonatal ICR mice. In addition, the degree of virulence was largely dependent upon the dose, route of infection, and number of passages of the challenge virus, as well as the ages of the infected animals. The present study provides valuable basic data to enable further research into EV71 pathogenesis and to facilitate the development of new drugs and vaccines.

Optimization of an assay method for Simian virus 40 nucleic acid sequence / 中华微生物学和免疫学杂志

Objective To optimize the PCR primer sets for Simian virus 40 (SV40) detection and establish an assay method for SV40 which is of high sensitivity, strong specificity, broad applicability. Methods Two pairs of PCR primers were designed of based on 21 different SV40 strains genome by Primer Premier 5.00 software, and the features of two pairs of PCR primers were analyzed by Oligo software (version 6.71), conservative nucleotide of two pairs of PCR primers and the PCR amplification product were analyzed by DNAMAN software (version 6.0.40). Two pairs of new-built PCR primers were compared with those derived from China pharmacopoeia (Clip) in these aspects. The detection sensitivity of four pairs of PCR primers were analyzed using different SV40 DNA diluent as PCR template. The detection specificity of four pairs of PCR primers were analyzed using sterile water, Vero cell DNA, SV40 DNA as PCR template, respectively. Results The sequences of the new PCR primer sets VP1 and T are conservative for 21 Strains. The sequences of PCR primer sets GCVP1 and GCT are conservative for SV40 strains whose accession No. is J02400, NC_001669, AF316139 and AF316141. As far as the same diluent SV40 DNA template is concerned, the PCR amplification efficiency of PCR primer set VP1 and T is higher than that of PCR primer set GCVP1 and GCT. There are non-specific band in nucleic acid electrophoresis for amplification products of PCR primer sets GCVP1 and GCT, whereas there are no non-specific band in nucleic acid electrophoresis for amplification products of PCR primer sets VP1 and T. Conclusion The new assay method for SV40 nucleic acid sequence has many better qualities than those in Chp such as high sensitivity, strong specificity, broad applicability, conservation of primers and their amplification products and so on.

Application Effect of Protein A/G Affinity Chromatograph in Heapatitis C Virus Antibody Purification / 医学研究杂志

Objective To analyze the factors of Protein A/G affinity column,influencing purification effect,and get the best condition to improve application effect of Protein A/G affinity column. Methods We Purified anti-HCV-IgG serum with different disposal modality,different sample input modality and different application number of affinity column before detecting and analyzing the purified samples. Results The Protein A/G affinity column had the best purified effect after using saturated ammonium sulfate to first purification,which increased the affinity column adsorption effect within 30 minutes adsorption. Conclusion Using antibody with first purification and adding the adsorption time could improve utilization rate of affinity column and prolong the service life.

Differentiation of Bone Marrow Mesenchymal Cells to Neural Cells / 华中科技大学学报（医学）（英德文版）

To explore the possibility and condition of differentiation of bone marrow mesenchymal cells (BMSCs) to neural cells in vitro, BMSCs from whole bone marrow of rats were cultured. The BMSCs of passage 3 were identified with immunocytochemical staining of CD44 ( + ), CD71 ( + )and CD45(-). There were type Ⅰ and type Ⅱ cells in BMSCs. Type Ⅰ BMSCs were spindleshaped and strong positive in immunocytochemical staining of CD44 and CD71, whereas flat and big type Ⅱ BMSCs were lightly stained. The BMSCs of same passage were induced to differentiate into neural cells by β-mercaptoethanol (BME). After induction by BME, the type Ⅰ BMSCs withdrew to form neuron-like round soma and axon-like and dendrite-like processes, and were stained positively for neurofilament (NF). The type Ⅱ BMSCs did not change in the BME medium and were negatively or slightly stained of NF.