Combating the SARS-CoV-2 Omicron variant with non-Omicron neutralizing antibodies

The highly mutated and transmissible Omicron variant has provoked serious concerns over its decreased sensitivity to the current coronavirus disease 2019 (COVID-19) vaccines and evasion from most anti-severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) neutralizing antibodies (NAbs). In this study, we explored the possibility of combatting the Omicron variant by constructing bispecific antibodies based on non-Omicron NAbs. We engineered ten IgG-like bispecific antibodies with non-Omicron NAbs named GW01, 16L9, 4L12, and REGN10987 by fusing the single-chain variable fragments (scFvs) of two antibodies through a linker and then connecting them to the Fc region of IgG1. Surprisingly, eight out of ten bispecific antibodies showed high binding affinity to the Omicron receptor-binding domain (RBD) and exhibited extreme breadth and potency against pseudotyped SARS-CoV-2 variants of concern (VOCs) including Omicron, as well as authentic Omicron(+R346K) variants. Six bispecific antibodies containing the cross-NAb GW01 neutralized Omicron variant and retained their abilities to neutralize other sarbecoviruses. Bispecific antibodies inhibited Omicron infection by binding to the ACE2 binding site. A cryo-electron microscopy (cryo-EM) structure study of the representative bispecific antibody FD01 in complex with the Omicron spike (S) revealed 5 distinct trimers and one unique bi-trimer conformation. The structure and mapping analyses of 34 Omicron S variant single mutants elucidated that two scFvs of the bispecific antibody synergistically induced the RBD-down conformation into 3-RBD-up conformation, enlarged the interface area, accommodated the S371L mutation, improved the affinity between a single IgG and the Omicron RBD, and hindered ACE2 binding by forming bi-trimer conformation. Our study offers an important foundation for anti-Omicron NAb design. Engineering bispecific antibodies based on non-Omicron NAbs may provide an efficient solution to combat the Omicron variant.

Analysis of trends and hot spots in the research of atrial fibrillation and cognitive impairment / 中华全科医师杂志

Objective:To explore the research trends and hot spots of atrial fibrillation and cognitive impairment.Methods:Research papers and review articles related to atrial fibrillation and cognitive impairment was retrieved from the Web of Science core collection database from inception to November 17, 2021. The bibliometric and visualization analysis on the publishing times,journals and countries,related disciplines and key words was conducted with Excel and VOSviewer software.Results:A total of 1 496 articles were collected,showing a parabolic rise in the number of published articles in recent years. During this period,journal Stroke published more high quality articles in the field than others. The academic areas with the most publications were neurology, cardiovascular research, peripheral vascular disease and geriatrics. "Stroke" and "risk factors" were the keywords with the highest frequency. The research hot spots of atrial fibrillation and cognitive impairment mainly focused on anticoagulant drugs, mechanism research, epidemiology catheter ablation and status quo investigation. Conclusion:Through visualization, the development trend and research hot spots on atrial fibrillation and cognitive dysfunction are displayed intuitively, to provide reference for future studies.

Clinical features of patients with acute myocardial infarction presenting de Winter pattern on electrocardiogram / 中华全科医师杂志

Objective:To analysis the clinical features of patients with acute myocardial infarction (AMI) presenting de Winter pattern on electrocardiogram.Methods:A total of 1 287 patients with AMI admitted to Beijing Luhe Hospital between June 2017 and January 2019 were enrolled in the study. Electrocardiogram and clinical features of 13 patients with AMI presenting de Winter pattern on electrocardiogram were analyzed and compared with anterior wall ST-segment elevation myocardial infarction(STEMI, n=206). Results:Among the 13 patients, 12 were males, aged (52.23±12.55) years old. Compared to patients with anterior wall STEMI, the age in the de Winter group was younger [(52.23±12.55)years vs. (59.79±12.46)years; t=-2.12, P=0.03], and the time from onset to appearing a typical ECG was shorter [109.0 (71.5, 152.0)min vs. 200.5 (120.0, 397.5)min; Z=-3.38, P<0.01]. Three cases showed a shifting between de Winter pattern and typical STEMI ECG: the de Winter ECG pattern progressed to STEMI in 2 cases, 1 case changed from STEMI to de Winter,then converted to STEMI again. The emergency angiography was performed in all 13 patients, angiography showed that proximal left anterior descending branch (LAD) was involved in 11 cases, mid LAD was involved in 1 case, and diffuse spasm occurred in all vessels in 1 case. The de Winter ECG pattern vanished in all patients after primary percutaneous coronary intervention or emergency angiography. Conclusions:The de Winter ECG pattern suggests an acute proximal or mid LAD artery occlusion, and the de Winter ECG pattern can be alternated with STEMI. The de Winter pattern should be recognized and revascularization should be given early.

Antibody-dependent enhancement (ADE) of SARS-CoV-2 infection in recovered COVID-19 patients: studies based on cellular and structural biology analysis

Antibody-dependent enhancement (ADE) has been reported in several virus infections including dengue fever virus, severe acute respiratory syndrome (SARS) and Middle East respiratory syndrome (MERS) coronavirus infection. To study whether ADE is involved in COVID-19 infections, in vitro pseudotyped SARS-CoV-2 entry into Raji cells, K562 cells, and primary B cells mediated by plasma from recovered COVID-19 patients were employed as models. The enhancement of SARS-CoV-2 entry into cells was more commonly detected in plasma from severely-affected elderly patients with high titers of SARS-CoV-2 spike protein-specific antibodies. Cellular entry was mediated via the engagement of Fc{gamma}RII receptor through virus-cell membrane fusion, but not by endocytosis. Peptide array scanning analyses showed that antibodies which promote SARS-CoV-2 infection targeted the variable regions of the RBD domain. To further characterize the association between the spike-specific antibody and ADE, an RBD-specific monoclonal antibody (7F3) was isolated from a recovered patient, which potently inhibited SARS-Cov-2 infection of ACE-2 expressing cells and also mediated ADE in Raji cells. Site-directed mutagenesis the spike RBD domain reduced the neutralization activity of 7F3, but did not abolish its binding to the RBD domain. Structural analysis using cryo-electron microscopy (Cryo-EM) revealed that 7F3 binds to spike proteins at a shift-angled pattern with one "up" and two "down" RBDs, resulting in partial overlapping with the receptor binding motif (RBM), while a neutralizing monoclonal antibody that lacked ADE activity binds to spike proteins with three "up" RBDs, resulting in complete overlapping with RBM. Our results revealed that ADE mediated by SARS-CoV-2 spike-specific antibodies could result from binding to the receptor in slightly different pattern from antibodies mediating neutralizations. Studies on ADE using antibodies from recovered patients via cell biology and structural biology technology could be of use for developing novel therapeutic and preventive measures for control of COVID-19 infection.

Neutralizing antibody responses to SARS-CoV-2 in a COVID-19 recovered patient cohort and their implications

BackgroundThe COVID-19 pandemic caused by SARS-CoV-2 coronavirus threatens global public health. Currently, neutralizing antibodies (NAbs) versus this virus are expected to correlate with recovery and protection of this disease. However, the characteristics of these antibodies have not been well studied in association with the clinical manifestations in patients. MethodsPlasma collected from 175 COVID-19 recovered patients with mild symptoms were screened using a safe and sensitive pseudotyped-lentiviral-vector-based neutralization assay. Spike-binding antibody in plasma were determined by ELISA using RBD, S1, and S2 proteins of SARS-CoV-2. The levels and the time course of SARS-CoV-2-specific NAbs and the spike-binding antibodies were monitored at the same time. FindingsSARS-CoV-2 NAbs were unable to cross-reactive with SARS-CoV virus. SARS-CoV-2-specific NAbs were detected in patients from day 10-15 after the onset of the disease and remained thereafter. The titers of NAb among these patients correlated with the spike-binding antibodies targeting S1, RBD, and S2 regions. The titers of NAbs were variable in different patients. Elderly and middle-age patients had significantly higher plasma NAb titers (P<0.0001) and spike-binding antibodies (P=0.0003) than young patients. Notably, among these patients, there were ten patients whose NAb titers were under the detectable level of our assay (ID50: < 40); while in contrast, two patients, showed very high titers of NAb, with ID50 :15989 and 21567 respectively. The NAb titers were positive correlated with plasma CRP levels but negative correlated with the lymphocyte counts of patients at the time of admission, indicating an association between humoral response and cellular immune response. InterpretationThe variations of SARS-CoV-2 specific NAbs in recovered COVID-19 patients may raise the concern about the role of NAbs on disease progression. The correlation of NAb titers with age, lymphocyte counts, and blood CRP levels suggested that the interplay between virus and host immune response in coronavirus infections should be further explored for the development of effective vaccine against SARS-CoV-2 virus. Furthermore, titration of NAb is helpful prior to the use of convalescent plasma for prevention or treatment. FundingMinistry of Science and Technology of China, National Natural Science Foundation of China, Shanghai Municipal Health Commission, and Chinese Academy of Medical Sciences

Clinical study on new seizure types in epilepsy patients / 中华神经科杂志

Objective To study the percentage and cause of new seizure types in epilepsy patients and attempt to evaluate the value of new seizure types.Methods All 1074 eapilepsy patients were envolled in the retrospectively study.The types of seizure were diagnosed according to International League Against Epilepsy seizure classification and neuroimaging results of the patients.The Wilcoxon signed rank sum test was used to assess the difference of age at first seizure and mean duration of epilepsy patients with and without new seizure types.The chi-square test was used to compare the abnormal proportion of EEG and neuroimaging results of epilepsy patients.Results Two hundred and thirty-one(21.5%)of 1074 epilepsy patients experienced new seizure type.Five hundred and eighty-four patients had partial seizure with 132 (22.6%)experiencing new types of seizure while 490 patients had generalized seizure with 99(20.2%)experiencing new types of seizure.Five hundred and seventy-six(53.6%)had abnormal EEG and 237 (22.1%)had abnormal neuroimaging in 1074 epilepsy patients.A hundred and twenty-three(52.3%)of 231 patients with new seizure types had abnormal EEG,and 75(32.5%)of them had abnormal neuroimaging. The possible causes for the new types of seizure could be found in 41 patients.There were no significant difference in the age of the first onset between the patients with and without the new types of seizure while there was significant difference in the average course of disease between them(χ~2=18.511-23.836.P＜0.05).There were significant differences in the rates of abnormal results of examination of nervous system and imaging outcomes.Conclusions There may be different causes of new seizure types.The study of new seizure types is helpful in diagnosis,treatment and prevention of epilepsy.

Structural and functional analysis of 2.2 kb spliced variant of hepatitis B virus genomes isolated from liver tissues from hepatocellular carcinoma patients / 中华实验和临床病毒学杂志

<p><b>BACKGROUND</b>To study the structure and function of 2.2 kb spliced variant of HBV genome from liver tissues of hepatocellular carcinoma patients.</p><p><b>METHODS</b>HBV genomes were amplified by using PCR from paired hepatocellular carcinoma tissues and peritumor tissues. The 3.2 kb full-length HBV genome and 2.2 kb spliced variant were separately cloned and sequenced. Hep G2 cells were co-transfected with full-length HBV DNA and 2.2 kb spliced variant, and after transfection, HBV DNAs from intracellular core particles were harvested and specific primers were used in PCR to evaluate the interactions between spliced variant and full-length counterpart in replication.</p><p><b>RESULTS</b>Semi-quantification by scanning density showed that 2.2 kb spliced variant was present in all tumor and peri-tumor samples studied. Sequence analyes revealed that the 5 terminus packaging signal for pregenomic and X and PreC/C genes were retained. When full-length HBV DNA was co-transfected with 2.2 kb, the replication signal of 3.2 kb HBV genome was increased 3-7 times.</p><p><b>CONCLUSIONS</b>The 2.2 kb HBV spliced variant was present in liver tissues, and relative content was higher in tumor tissues than that in the peri-tumor tissues. This spliced variant could enhance the replication of full-length HBV genome, which suggested the possible role of the variant in the pathogenesis of development of hepatocellular carcinoma.</p>

Association between NS5A gene sequence and response to interferon therapy in chronic hepatitis C patients in Shanghai / 中华实验和临床病毒学杂志

<p><b>BACKGROUND</b>To elucidate relationship between amino acid sequence of non-structural protein 5A (NS5A) and outcome of HCV (1 b) patients after interferon (IFNa) therapy.</p><p><b>METHODS</b>Sera of 24 patients were collected before, during and after IFNa therapy. Pretreatment RNA levels and the sequences of HCV NS5A interferon sensitivity determining region (ISDR) were determined. NS5A full-length sequences of 5 HCV isolates from 3 patients with different response types were also analyzed. Phylogenetic tree analysis and protein secondary structure prediction were undertaken.</p><p><b>RESULTS</b>Pretreatment RNA levels of sustained response group were significantly lower than that of non-response group and relapse group (4.50X104 copies/ml versus 1.82X107 copies/ml, P < 0.01).ISDR sequences of NS5A from pretreatment sera were compared with HCV-J strain (prototype). Thirteen of 24 isolates were wild type,11 of 24 were intermediate type and none of them was mutant type. 3 of 6 sustained responders were infected with wild-type isolates, the rest with intermediate type isolates. Phylogenetic tree based on NS5A full-length sequences classified 5 isolates with 3 different response types into 3 groups. Non-response isolates belonged to the same group as HCV-J. Secondary structure prediction of 5 isolates revealed significant differences existing in 2 255- 2 289. This region was partly overlapped with PKR-binding domain.</p><p><b>CONCLUSIONS</b>Low HCV RNA levels in serum are associated with favorable outcome of IFNa therapy. ISDR sequence alone could not predict outcome of IFN treatment. Combination of determination of HCV RNA levels in serum with sequence analysis of PKR-binding domain may be helpful in predicting the efficacy of IFN therapy.</p>

Studies on virulence mediated by drug resistant Salmonella typhi R plasmid / 中华微生物学和免疫学杂志

Objective　To explore the possibility of R plasmid (pRST98) encoding resistance to antimicrobial agents in S. typhi mediate virulence to its host bacteria. Methods　pRST98 was transferred into a less virulent strain of S. typhimurium RIA for creating a transconjugant pRST98/RIA. The standard S. typhimurium virulence strain SR-11 which carries a 100kb virulence plasmid was used as a positive control, and RIA as a negative one. Infection with S. typhimurium was employed to assess the effect of pRST98 on the virulence of host bacteria by LD50 of peroral (p.o.) and intraperitoneal (i.p.) infection of BALB/c mice. After oral inoculation of the bacteria into BALB/c mice, mesenteric lymph nodes, spleen and liver were examined for quantity of bacteria and for the histopathological changes. The three strains were also studied for their in vitro adhesion and invasion of HEp-2, CHO and HeLa cells. Three chromosomally isogenic strains, i.e., plasmid-containing wild-type S. typhi strain, plasmid artificially cured strain and plasmid-reintroduced into the cured strain were compared for their resistance to the bactericidal activity present in human, rabbit and guinea pig sera. Results　The p.o. and i.p. LD50 of pRST98/RIA was 700 and 75 folds less than that of RIA. The bacteria counting in mesenteric lymph nodes, spleen and liver were more (P<0.05) and more severe histopathological changes were observed in pRST98 harboring S. typhimurium. However, pRST98 did not affect the adhesion and invasion of S. typhimurium to the cells. Experiments showed that pRST98 mediated resistance to serum bactericidal effect in host bacteria (P<0.05). Conclusion　This is the first report about a plasmid carrying genes coding for drug resistance and virulence in S. typhi. The data presented here suggested that novel virulence gene or related sequences, which are important in the pathogenesis of Salmonella infection could exist on pRST98.

STUDY ON AFFINITY OF ANTI-HBc IN VIRAL HEPATITIS B PATIENTS AND ASYMPTOMATIC HBsAg CARRIERS / 中华传染病杂志

An assay for the affinity of serum anti-HBc in chronic hepatitis B patients and asymptomatic HBsAg carriers by 125I,22Na double isotope radioimmunoassay is described.In 7 asymmptomatic HBsAg carriers and 9 chronic hepatitis B patients.the affinity of anti-HBc was 3.25±2.13×106L/Mole in the former,whereas 0.75±0.56×106L/Mole in the latter.The titers of anti-HBc did not correlate to the affinity of anti-HBc and its possib]e role in the pathogenesis of hepatitis B is discussed.