Molecular analysis of a public cross-neutralizing antibody response to SARS-CoV-2

As SARS-CoV-2 variants of concerns (VOCs) continue to emerge, cross-neutralizing antibody responses become key towards next-generation design of a more universal COVID-19 vaccine. By analyzing published data from the literature, we report here that the combination of germline genes IGHV2-5/IGLV2-14 represents a public antibody response to the receptor-binding domain (RBD) that potently cross-neutralizes all VOCs to date, including Omicron and its sub-lineages. Detailed molecular analysis shows that the complementarity-determining region H3 sequences of IGHV2-5/IGLV2-14-encoded RBD antibodies have a preferred length of 11 amino acids and a conserved HxIxxI motif. In addition, these antibodies have a strong allelic preference due to an allelic polymorphism at amino-acid residue 54 of IGHV2-5, which locates at the paratope. These findings have important implications for understanding cross-neutralizing antibody responses to SARS-CoV-2 and its heterogenicity at the population level as well as the development of a universal COVID-19 vaccine.

A large-scale systematic survey of SARS-CoV-2 antibodies reveals recurring molecular features

In the past two years, the global research in combating COVID-19 pandemic has led to isolation and characterization of numerous human antibodies to the SARS-CoV-2 spike. This enormous collection of antibodies provides an unprecedented opportunity to study the antibody response to a single antigen. From mining information derived from 88 research publications and 13 patents, we have assembled a dataset of [~]8,000 human antibodies to the SARS-CoV-2 spike from >200 donors. Analysis of antibody targeting of different domains of the spike protein reveals a number of common (public) responses to SARS-CoV-2, exemplified via recurring IGHV/IGK(L)V pairs, CDR H3 sequences, IGHD usage, and somatic hypermutation. We further present a proof-of-concept for prediction of antigen specificity using deep learning to differentiate sequences of antibodies to SARS-CoV-2 spike and to influenza hemagglutinin. Overall, this study not only provides an informative resource for antibody and vaccine research, but fundamentally advances our molecular understanding of public antibody responses to a viral pathogen.

Human organoid systems reveal in vitro correlates of fitness for SARS-CoV-2 B.1.1.7

A new phase of the COVID-19 pandemic has started as several SARS-CoV-2 variants are rapidly emerging globally, raising concerns for increased transmissibility. As animal models and traditional in vitro systems may fail to model key aspects of the SARS-CoV-2 replication cycle, representative in vitro systems to assess variants phenotypically are urgently needed. We found that the British variant (clade B.1.1.7), compared to an ancestral SARS-CoV-2 clade B virus, produced higher levels of infectious virus late in infection and had a higher replicative fitness in human airway, alveolar and intestinal organoid models. Our findings unveil human organoids as powerful tools to phenotype viral variants and suggest extended shedding as a correlate of fitness for SARS-CoV-2. One-Sentence SummaryBritish SARS-CoV-2 variant (clade B.1.1.7) infects organoids for extended time and has a higher fitness in vitro.

Homologous and heterologous serological response to the N-terminal domain of SARS-CoV-2

The increasing numbers of infected cases of coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) poses serious threats to public health and the global economy. Most SARS-CoV-2 neutralizing antibodies target the receptor binding domain (RBD) and some the N-terminal domain (NTD) of the spike protein, which is the major antigen of SARS-CoV-2. While the antibody response to RBD has been extensively characterized, the antigenicity and immunogenicity of the NTD protein are less well studied. Using 227 plasma samples from COVID-19 patients, we showed that SARS-CoV-2 NTD-specific antibodies could be induced during infection. As compared to the serological response to SARS-CoV-2 RBD, the SARS-CoV-2 NTD response is less cross-reactive with SARS-CoV. Furthermore, neutralizing antibodies are rarely elicited in a mice model when NTD is used as an immunogen. We subsequently demonstrate that NTD has an altered antigenicity when expressed alone. Overall, our results suggest that while NTD offers an alternative strategy for serology testing, it may not be suitable as an immunogen for vaccine development.

Sequence signatures of two IGHV3-53/3-66 public clonotypes to SARS-CoV-2 receptor binding domain

Since the COVID-19 pandemic onset, the antibody response to SARS-CoV-2 has been extensively characterized. Antibodies to the receptor binding domain (RBD) on the spike protein are frequently encoded by IGHV3-53/3-66 with a short CDR H3. Germline-encoded sequence motifs in CDRs H1 and H2 play a major role, but whether any common motifs are present in CDR H3, which is often critical for binding specificity, have not been elucidated. Here, we identify two public clonotypes of IGHV3-53/3-66 RBD antibodies with a 9-residue CDR H3 that pair with different light chains. Distinct sequence motifs on CDR H3 are present in the two public clonotypes that appear to be related to differential light chain pairing. Additionally, we show that Y58F is a common somatic hypermutation that results in increased binding affinity of IGHV3-53/3-66 RBD antibodies with a short CDR H3. Overall, our results advance fundamental understanding of the antibody response to SARS-CoV-2.

Human airway cells prevent SARS-CoV-2 multibasic cleavage site cell culture adaptation

Virus propagation methods generally use transformed cell lines to grow viruses from clinical specimens, which may force viruses to rapidly adapt to cell culture conditions, a process facilitated by high viral mutation rates. Upon propagation in VeroE6 cells, SARS-CoV-2 may mutate or delete the multibasic cleavage site (MBCS) in the spike protein that facilitates serine protease-mediated entry into human airway cells. We report that propagating SARS-CoV-2 on the human airway cell line Calu-3 - that expresses serine proteases - prevents MBCS mutations. Similar results were obtained using a human airway organoid-based culture system for SARS-CoV-2 propagation. Thus, in-depth knowledge on the biology of a virus can be used to establish methods to prevent cell culture adaptation.

Cross-reactive antibody response between SARS-CoV-2 and SARS-CoV infections

The World Health Organization has recently declared the ongoing outbreak of COVID-19, which is caused by a novel coronavirus SARS-CoV-2, as pandemic. There is currently a lack of knowledge in the antibody response elicited from SARS-CoV-2 infection. One major immunological question is concerning the antigenic differences between SARS-CoV-2 and SARS-CoV. We address this question by using plasma from patients infected by SARS-CoV-2 or SARS-CoV, and plasma obtained from infected or immunized mice. Our results show that while cross-reactivity in antibody binding to the spike protein is common, cross-neutralization of the live viruses is rare, indicating the presence of non-neutralizing antibody response to conserved epitopes in the spike. Whether these non-neutralizing antibody responses will lead to antibody-dependent disease enhancement needs to be addressed in the future. Overall, this study not only addresses a fundamental question regarding the antigenicity differences between SARS-CoV-2 and SARS-CoV, but also has important implications in vaccine development.

Authentication of oviductus ranae and its original animals using molecular marker.

Two pairs of diagnostic primers, IHm01-L/IHm01-H and IHm02-L/IHm02-H, for distinguishing the Chinese crude drug Oviductus Ranae from its substitutes were designed based on sequences of Cyt b gene fragment of the original animals of the drug and substitutes. Total DNAs were extracted from crude drugs purchased from five drugstores in different regions, as well as from original animals of the drug, Rana chensinensis, and seven species of related ranid species. Diagnostic polymerase chain reactions (PCRs) were performed using the two pairs of primers with the total DNAs of the original animals as a template. The result showed that a 240 bp DNA segment was clearly amplified from all templates of Rana chensinensis using primers IHmO1-L and IHm01-H, whereas no DNA band appeared from other templates. While using primers IHm02-L and IHm02-H, we got a clear 140 bp DNA band from all the templates of R. huanrenensis and 3 oviducts of the same species, no PCR product was observed from the other samples. A set of PCR reactions was employed to identify crude drugs from the five drugstores using the two pairs of primers together with HsmL1 and HsmH1 reported in our previous study. The results show that only 20% of the Oviductus Ranae currently sold in markets are qualified products and the rest are not.

IDENTIFICATION OF CHINESE CRUDE DRUG SNAKE GALLBLADDER BY DNA MOLECULAR MARKER / 药学学报

AIM　It is difficult to identify the Chinese crude drug snake gallbladder accurately by morphological and microscopical characteristics or chemical components only. In order to solve the problem, the technique based on DNA molecular marker was introduced into the authentication of snake gallbladder. METHODS　DNA templates were extracted from the membrane or the bile of snake gallbladder, and also from the muscle of the original animal Elaphe schrenckii. About 400 bp DNA fragments of 12S rRNA gene were amplified from the templates and sequenced subsequently. RESULTS　Enough amounts of DNA templates could be extracted from a bit of membrane or bile of snake gallbladder. The sequence of amplicons from the membrane, bile and muscle of the same individual were identical completely. CONCLUSION　The technique of DNA molecular marker could be used for the authentication of snake gallbladder and bile. The results indicate that the technique could be used for the identification of crude drugs from other animal secretion. DNA sequence analysis also demonstrated that the origins of commercial snake gallbladder were complicated and more efficient quality control was necessary for supervising the crude drug in the market.

Physical properties of Ni-Cr ceramic alloys after recasts / 实用口腔医学杂志

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Authentication of TCM Carapax Trionycis by allele-specific diagnostic polymeras chain reaction / 中草药

Object\ To develop a convenient and practical method for the identification of Carapax Trionycis Methods\ Based on the sequence variations of 12S rRNA gene between Pelodiscus sinensis and other softshell turtles, a pair of allele specific primers was designed to distinguish P. sinensis from other species of Trionychidae. DNA were extracted and anplified and Carapax Trionycis could be identified accurately by polymerase chain reaction (PCR) using the primers Results\ Ten samples of turtle shell from different sources were indentified by the allele specific PCR with the primers The result indicated that three samples were substitutes of Carapax Trionycis, consilient with the result from DNA sequence analysis The mitochondrial 12S rRNA gene fragment of P. maculatus and a faked imitation had also been sequenced Conclusion\ The primers could be used as key components in Carapax Trionycis identification kit