Microbial GWAS studies revealing combinations of Omicron RBD mutations existed and may contribute to antibody evasion and ACE2 binding

Since Omicron variant of SARS-CoV-2 was first detected in South Africa (SA), it has now dominated in United Kingdom (UK) of Europe and United State (USA) of North America. A prominent feature of this variant is the gathering of spike protein mutations, in particularly at the receptor binding domain (RBD). These RBD mutations essentially contribute to antibody resistance of current immune approaches. During global spillover, combinations of RBD mutations may exist and synergistically contribute to antibody resistance in fact. Using three geographic-stratified genome wide association studies (GWAS), we observed that RBD combinations exhibited a geographic pattern and genetical associated, such as five common mutations in both UK and USA Omicron, six or two specific mutations in UK or USA Omicron. Although the UK specific RBD mutations can be further classified into two separated sub-groups of combination based on linkage disequilibrium analysis. Functional analysis indicated that the common RBD combinations (fold change, -11.59) alongside UK or USA specific mutations significantly reduced neutralization (fold change, -38.72, -18.11). As RBD overlaps with angiotensin converting enzyme 2(ACE2) binding motif, protein-protein contact analysis indicated that the common RBD mutations enhanced ACE2 binding accessibility and were further strengthened by UK or USA-specific RBD mutations. Spatiotemporal evolution analysis indicated that UK-specific RBD mutations largely contribute to global spillover. Collectively, we have provided genetic evidence of RBD combinations and estimated their effects on antibody evasion and ACE2 binding accessibility.

Effect of CYP3A5 and MDR1 gene polymorphism on blood concentration of tacrolimus and creatinine level in uremic patients during the early phase of kidney transplantation / 药学实践杂志

Objective To investigate the effect of CYP3A5 and MDR1 gene polymorphisms on blood concentration of tacrolimus and creatinine level in uremic patients during the early phase after kidney transplantation in real clinical practice. Methods 131 patients who underwent kidney transplantation for the first time with triple immunotherapy based on tacrolimus in single-center from 2013 to 2017 were enrolled for retrospective study. Tacrolimus daily dose, blood concentration, blood concentration-to-dose ratio, and serum level were compared according to the various genotypes of CYP3A5 and MDR1 polymorphisms in renal transplantation recipients, respectively. Results The dosage of tacrolimus in CYP3A5*3/*3 (GG) kidney transplantation recipients within 4 weeks after kidney transplantation was lower than those of CYP3A5*1/*1 (AA) and CYP3A5*1/*3 (AG). The serum creatinine levels of patients whose tacrolimus concentration in the range of 10-13 ng/ml were close to the normal value. Conclusion CYP3A5 gene polymorphism affects the blood concentrations of tacrolimus in renal transplant recipients. No association has been found between the blood concentrations of tacrolimus and MDR1 gene polymorphism. Tacrolimus concentration in the range of 10-13 ng/ml might contribute to restore the early kidney graft function.

Tracing two causative SNPs reveals SARS-CoV-2 transmission in North America population

During the COVID-19 pandemic, precisely tracing the route of the SARS-CoV-2 transmission in human population remains challenging. Because this RNA virus can mutate massively without a specifically tracing maker. Herein, using a geographic stratified genome-wide association study (GWAS) of 2599 full-genome sequences, we identified that two SNPs (i.e., 1059.C>T and 25563.G>T) of linkage disequilibrium were presented in approximately half of North America SARS-CoV-2 population (p = 2.44 x 10-212 and p = 2.98 x 10-261), resulting two missense mutations (i.e., Thr 265 Ile and Gln 57 His) in ORF1ab and ORF3a, respectively. Interestingly, these two SNPs exclusively occurred in the North America dominated clade 1, accumulated during mid to late March, 2020. We did not find any of these two SNPs by retrospectively tracing the two SNPs in bat and pangolin related SARS-CoV-2 and human SARS-CoV-2 from the first epicenter Wuhan or other regions of China mainland. This suggested that the SARS-CoV-2 population of Chinese mainland were different from the prevalent strains of North America. Time-dependently, we found that these two SNPs first occurred in Europe SARS-CoV-2 (26-Feb-2020) which was 3 days early than the occurring date of North America isolates and 17 days early for Asia isolates (Taiwan China dominated). Collectively, this population genetic analysis highlights a well-confidential transmission route of the North America isolates and the two SNPs we newly identified are possibly novel diagnosable or druggable targets for surveillance and treatment.

The role of miR-221 in the injury induced by hydrogen peroxide in rat myocardial cells / 中国比较医学杂志

Objective To explore the role of miR-221 in the injury induced by hydrogen peroxide (H2O2) in rat myocardial cells (H9c2).Methods The viability of H9c2 cell induced by cell different concentrations of H2O2 was determined by MTT.The expression of miR-221 was detected by RT-PCR method.The miR-221 inhibitor and negative control were transferred into H9c2 cells by Lipofectamine 2000, then the cells were divided into normal control group, model control group (H2O2 group), negative control group (H2O2+ negative control group), inhibition group (H2O2+miR-221 inhibitor group).The cell viability was measured by MTT assay.Cell apoptosis was detected by acridine orange staining method.The expression of Bcl-2, Bax, phosphatase and tensin homolog deleted on chromosome ten (PTEN, p-protein kinase B (AKT) were assayed by Western Blot.Results 0,25,50,100,200,400 μmol/L H2O2 inhibited H9c2 cell activity gradually, of which 200 mol/L inhibition of cell viability moderate, so as a subsequent induction dose.Compared with normal control group, cell viability was decreased (P < 0.01), cell apoptotic rat was increased (P < 0.01), the expression of Bax and PTEN was upregulated (P < 0.01), the expression of Bcl-2 and p-AKT was downregulated (P < 0.01) in model control group and negative control group.Compared with model control group and negative control group, inhibition group proves the contrary.Conclusions Down-expression of miR-221 could significantly inhibit oxidative stress damage in H9c2 cells, which related to regulation of PTEN/AKT signal pathway.

Molecular characterization of duck enteritis virus CHv strain UL49.5 protein and its colocalization with glycoprotein M

The UL49.5 gene of most herpesviruses is conserved and encodes glycoprotein N. However, the UL49.5 protein of duck enteritis virus (DEV) (pUL49.5) has not been reported. In the current study, the DEV pUL49.5 gene was first subjected to molecular characterization. To verify the predicted intracellular localization of gene expression, the recombinant plasmid pEGFP-C1/pUL49.5 was constructed and used to transfect duck embryo fibroblasts. Next, the recombinant plasmid pDsRed1-N1/glycoprotein M (gM) was produced and used for co-transfection with the pEGFP-C1/pUL49.5 plasmid to determine whether DEV pUL49.5 and gM (a conserved protein in herpesviruses) colocalize. DEV pUL49.5 was thought to be an envelope glycoprotein with a signal peptide and two transmembrane domains. This protein was also predicted to localize in the cytoplasm and endoplasmic reticulum with a probability of 66.7%. Images taken by a fluorescence microscope at different time points revealed that the DEV pUL49.5 and gM proteins were both expressed in the cytoplasm. Overlap of the two different fluorescence signals appeared 12 h after transfection and continued to persist until the end of the experiment. These data indicate a possible interaction between DEV pUL49.5 and gM.

Apoptosis induced in vivo by new type gosling viral enteritis virus

In this study, apoptosis was induced by new type gosling viral enteritis virus (NGVEV) in experimentally infected goslings is reported in detail for the first time. After 3-day-old goslings were orally inoculated with a NGVEV-CN strain suspension, the time course of NGVEV effects on apoptotic morphological changes of the internal tissues was evaluated. These changes were observed by histological analysis with light microscopy and ultrastructural analysis with transmission electron microscopy. DNA fragmentation was assessed with a terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling (TUNEL) assay and DNA ladder analysis. A series of characteristic apoptotic morphological changes including chromatin condensation and margination, cytoplasmic shrinkage, plasma membrane blebbing, and formation of apoptotic bodies were noted. Apoptosis was readily observed in the lymphoid and gastrointestinal organs, and sporadically occurred in other organs after 3 days post-infection (PI). The presence and quantity of TUNEL-positive cells increased with infection time until 9 days PI. DNA extracted from the NGVEV-infected gosling cells displayed characteristic 180~200 bp ladders. Apoptotic cells were ubiquitously distributed, especially among lymphocytes, macrophages, monocytes, and epithelial and intestinal cells. Necrosis was subsequently detected during the late NGVEV-infection phase, which was characterized by cell swelling, plasma membrane collapse, and rapidly lysis. Our results suggested that apoptosis may play an important role in the pathogenesis of NGVE disease.

Development and application of a reverse transcriptase polymerase chain reaction to detect Chinese isolates of duck hepatitis virus type 1.

We developed a reverse transcriptase polymerase chain reaction (RT-PCR) method for the detection of duck hepatitis virus type 1 (DHV-1) in the tissues of infected and clinically affected ducks and in chick and duck embryos. We found the assay to be effective in detecting the virus in China, where it is being used in studies on the epidemiology of the disease. We applied this simple and rapid diagnostic method to the detection of DHV isolates grown in chick and duck embryos and in tissues obtained from infected birds. The assay also proved useful for the differentiation of DVH from the duck plague virus (DPV), muscovy parvovirus (MPV), gosling parvovirus (GPV), avian influenza virus (AIV/H5N1), Pasteurella multocida (PA/5:A), Riemerella anatipestifer (RA/serotype 1), and Salmonella enteritidis (SE). The limit of the sensitivity of this method for the detection of DHV-1 RNA was 3 pg/10 microl. As compared to Dot-ELISA and virus isolation, the rate of agreement for the detection of experimentally infected livers was 100%; moreover, the RT-PCR method was also capable of detecting DHV-1 RNA from the livers that had been infected and stored at -20 degrees C for 22 years; in contrast, Dot-ELISA and virus isolation method could only detect DHV-1 from the livers that had been infected and stored at -20 degrees C for 13 and 11 years, respectively. The rate of positivity in 185 clinically suspected diseased livers subjected to detection by RT-PCR, Dot-ELISA, and virus isolation was 89.2%, 69.2%, and 55.7%, respectively. These results indicated that the RT-PCR approach is rapid, sensitive, and reliable for the detection and differentiation of DHV-1 from the other clinical samples and suspected isolates.

Development and application of a reverse transcriptase polymerase chain reaction to detect Chinese isolates of duck hepatitis virus type 1.

We developed a reverse transcriptase polymerase chain reaction (RT-PCR) method for the detection of duck hepatitis virus type 1 (DHV-1) in the tissues of infected and clinically affected ducks and in chick and duck embryos. We found the assay to be effective in detecting the virus in China, where it is being used in studies on the epidemiology of the disease. We applied this simple and rapid diagnostic method to the detection of DHV isolates grown in chick and duck embryos and in tissues obtained from infected birds. The assay also proved useful for the differentiation of DVH from the duck plague virus (DPV), muscovy parvovirus (MPV), gosling parvovirus (GPV), avian influenza virus (AIV/H5N1), Pasteurella multocida (PA/5:A), Riemerella anatipestifer (RA/serotype 1), and Salmonella enteritidis (SE). The limit of the sensitivity of this method for the detection of DHV-1 RNA was 3 pg/10 microl. As compared to ELISA and virus isolation, the rate of agreement for the detection of experimentally infected livers was 100%; moreover, the RT-PCR method was also capable of detecting DHV-1 RNA from the livers that had been infected and stored at -20 degrees C for 22 years; in contrast, ELISA and virus isolation method could only detect DHV-1 from the livers that had been infected and stored at -20 degrees C for 13 and 11 years, respectively. The rate of positivity in 185 clinically suspected diseased livers subjected to detection by RT-PCR, ELISA, and virus isolation was 89.2%, 69.2%, and 55.7%, respectively. These results indicated that the RT-PCR approach is rapid, sensitive, and reliable for the detection and differentiation of DHV-1 from the other clinical samples and suspected isolates.

The pathogenesis of duck virus enteritis in experimentally infected ducks: a quantitative time-course study using TaqMan polymerase chain reaction.

Duck virus enteritis is an acute and contagious herpesvirus infection of duck, geese and swans with high morbidity and mortality. The kinetics of viral DNA loads and immunohistochemical localization of virulent duck enteritis virus, as well as histopathological examination in various tissues of ducks following oral infection, were investigated. The time course for the appearance of viral antigen and tissue lesions in various tissues was coincident with the levels of duck enteritis virus at the various sites, suggesting that the levels of duck enteritis virus in systemic organs have a close correlation with the progression of disease. The abundance of target epithelial and lymphoid cells may contribute to the high levels of virus infection and replication in lymphoid and intestinal tissues.