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In a phase I clinical trial, the chimeric hemagglutinin (cHA) immunogen induced antibody responses against the conserved HA stalk domain. However, the landscape of the B cell specificities and subsets induced by this vaccine remain undetermined. Here, we paired single cell RNA-sequencing and B cell receptor repertoire sequencing to analyze the relationship between transcriptome and B cell specificity following cHA immunization. We show that the cHA inactivated vaccine with a squalene-based adjuvant induced a robust activated B cell and memory B cell phenotype against two broadly neutralizing epitopes of the stalk domain. The overall specificities of the acute plasmablast and memory B cell responses were distinct, with the plasmablast compartment largely targeting non-neutralizing epitopes of the HA stalk. Altogether, these data indicate the B cell landscape following cHA vaccination includes diverse B cell subsets that are differentially induced by distinct vaccine formulations, including memory and de novo B cell responses against diverse broadly conserved epitopes.

Bats are reservoir hosts for many zoonotic viruses. Despite this, relatively little is known about the diversity and abundance of viruses within bats at the level of individual animals, and hence the frequency of virus co-infection and inter-species transmission. Using an unbiased meta-transcriptomics approach we characterised the mammalian associated viruses present in 149 individual bats sampled from Yunnan province, China. This revealed a high frequency of virus co-infection and species spillover among the animals studied, with 12 viruses shared among different bat species, which in turn facilitates virus recombination and reassortment. Of note, we identified five viral species that are likely to be pathogenic to humans or livestock, including a novel recombinant SARS-like coronavirus that is closely related to both SARS-CoV-2 and SARS-CoV, with only five amino acid differences between its receptor-binding domain sequence and that of the earliest sequences of SARS-CoV-2. Functional analysis predicts that this recombinant coronavirus can utilize the human ACE2 receptor such that it is likely to be of high zoonotic risk. Our study highlights the common occurrence of inter-species transmission and co-infection of bat viruses, as well as their implications for virus emergence.

As one of the most rapidly evolving proteins of the genus Betacoronavirus, open reading frames (ORF8's) function and potential pathological consequence in vivo are still obscure. In this study, we show that the secretion of ORF8 is dependent on its N-terminal signal peptide sequence and can be inhibited by reactive oxygen species scavenger and endoplasmic reticulum-Golgi transportation inhibitor in cultured cells. To trace the effect of its possible in vivo secretion, we examined the plasma samples of coronavirus disease 2019 (COVID-19) convalescent patients and found that the patients aged from 40 to 60 had higher antibody titers than those under 40. To explore ORF8's in vivo function, we administered the mice with ORF8 via tail-vein injection to simulate the circulating ORF8 in the patient. Although no apparent difference in body weight, food intake, and vitality was detected between vehicle- and ORF8-treated mice, the latter displayed morphological abnormalities of testes and epididymides, as indicated by the loss of the central ductal lumen accompanied by a decreased fertility in 5-week-old male mice. Furthermore, the analysis of gene expression in the testes between vehicle- and ORF8-treated mice identified a decreased expression of Col1a1, the loss of which is known to be associated with mice's infertility. Although whether our observation in mice could be translated to humans remains unclear, our study provides a potential mouse model that can be used to investigate the impact of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection on the human reproductive system.

The frequently emerging SARS-CoV-2 variants have weakened the effectiveness of existing COVID-19 vaccines and neutralizing antibody therapy. Nevertheless, the infections of SARS-CoV-2 variants still depend on angiotensin-converting enzyme 2 (ACE2) receptor-mediated cell entry, and thus the soluble human ACE2 (shACE2) is a potential decoy for broadly blocking SARS-CoV-2 variants. In this study, we firstly generated the recombinant AAVrh10-vectored shACE2 constructs, a kind of adeno-associated virus (AAV) serotype with pulmonary tissue tropism, and then validated its inhibition capacity against SARS-CoV-2 infection. To further optimize the minimized ACE2 functional domain candidates, a comprehensive analysis was performed to clarify the interactions between the ACE2 orthologs from various species and the receptor binding domain (RBD) of SARS-CoV-2 spike (S) protein. Based on the key interface amino acids, we designed a series of truncated ACE2 orthologs, and then assessed their potential affinity to bind to SARS-CoV-2 variants RBD in silico. Of note, we found that the 24-83aa fragment of dog ACE2 (dACE2

Objective To determine whether the signaling activation of bone morphogenetic protein 2(BMP2)can induce myeloid-derived suppressor cells(MDSC)to secret transforming growth factor β(TGF-β),further enhancing the differentiation and infiltration of regulatory T lymphocytes(Treg)into tumor tissue. Methods The BMP2-induced mRNA and protein expression of TGF-β in MDSC was detected by quantitative real-time polymerase chain reaction and enzyme-linked immunosorbent assay(ELISA),respectively.The effect of BMP2-induced TGF-β secretion by MDSC on Treg differentiation was then determined by flow cytometry.Finally,we implanted the recombined human bone morphogenetic protein 2(rhBMP2)collagen gels into tumor-burdened mice to examine the role of BMP2 in Treg differentiation via MDSC-secreted TGF-β

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Background Understanding factors associated with post-discharge sleep quality among COVID-19 survivors is important for intervention development. Aims This study investigated sleep quality and its correlates among COVID-19 patients 6 months after their most recent hospital discharge. Method Healthcare providers at hospitals located in five different Chinese cities contacted adult COVID-19 patients discharged between 1 February and 30 March 2020. A total of 199 eligible patients provided verbal informed consent and completed the interview. Using score on the single-item Sleep Quality Scale as the dependent variable, multiple linear regression models were fitted. Results Among all participants, 10.1% reported terrible or poor sleep quality, and 26.6% reported fair sleep quality, 26.1% reported worse sleep quality when comparing their current status with the time before COVID-19, and 33.7% were bothered by a sleeping disorder in the past 2 weeks. After adjusting for significant background characteristics, factors associated with sleep quality included witnessing the suffering (adjusted B = −1.15, 95% CI = −1.70, −0.33) or death (adjusted B = −1.55, 95% CI = −2.62, −0.49) of other COVID-19 patients during hospital stay, depressive symptoms (adjusted B = −0.26, 95% CI = −0.31, −0.20), anxiety symptoms (adjusted B = −0.25, 95% CI = −0.33, −0.17), post-traumatic stress disorders (adjusted B = −0.16, 95% CI = −0.22, −0.10) and social support (adjusted B = 0.07, 95% CI = 0.04, 0.10). Conclusions COVID-19 survivors reported poor sleep quality. Interventions and support services to improve sleep quality should be provided to COVID-19 survivors during their hospital stay and after hospital discharge.

Background Many COVID-19 survivors reported stigmatization after recovery. This study investigated the association between stigma (discrimination experiences, self-stigma and perceived affiliate stigma) and sleep quality among COVID-19 survivors six months after hospital discharge. Methods Participants were recovered adult COVID-19 survivors discharged between February 1 and April 30, 2020. Medical staff of five participating hospitals approached all discharged COVID-19 period during this period. A total of 199 participants completed the telephone interview during July to September, 2020. Structural equation modeling was performed to test the hypothesize that resilience and social support would mediate the associations between stigma and sleep quality. Results The results showed that 10.1% of the participants reported terrible/poor sleep quality, 26.1% reported worse sleep quality in the past week when comparing their current status versus the time before COVID-19. After adjusting for significant background characteristics, participants who had higher number of discrimination experience, perceived stronger self-stigma and stronger perceived affiliate stigma reported poorer sleep quality. Resilience and social support were positively and significantly associated with sleep quality. The indirect effect of self-stigma on sleep quality through social support and resilience was significant and negative. Perceived affiliate stigma also had a significant and negative indirect effect on sleep quality through social support and resilience. Conclusions Various types of stigma after recovery were associated with poor sleep quality among COVID-19 survivors, while social support and resilience were protective factors. Resilience and social support mediated the associations between self-stigma/perceived affiliate stigma and sleep quality.

Summary Background Influenza viruses cause substantial annual morbidity and mortality globally. Current vaccines protect against influenza only when well matched to the circulating strains. However, antigenic drift can cause considerable mismatches between vaccine and circulating strains, substantially reducing vaccine effectiveness. Moreover, current seasonal vaccines are ineffective against pandemic influenza, and production of a vaccine matched to a newly emerging virus strain takes months. Therefore, there is an unmet medical need for a broadly protective influenza virus vaccine. We aimed to test the ability of chimeric H1 haemagglutinin-based universal influenza virus vaccine candidates to induce broadly cross-reactive antibodies targeting the stalk domain of group 1 haemagglutinin-expressing influenza viruses. Methods We did a randomised, observer-blinded, phase 1 study in healthy adults in two centres in the USA. Participants were randomly assigned to one of three prime–boost, chimeric haemagglutinin-based vaccine regimens or one of two placebo groups. The vaccine regimens included a chimeric H8/1, intranasal, live-attenuated vaccine on day 1 followed by a non-adjuvanted, chimeric H5/1, intramuscular, inactivated vaccine on day 85; the same regimen but with the inactivated vaccine being adjuvanted with AS03; and an AS03-adjuvanted, chimeric H8/1, intramuscular, inactivated vaccine followed by an AS03-adjuvanted, chimeric H5/1, intramuscular, inactivated vaccine. In this planned interim analysis, the primary endpoints of reactogenicity and safety were assessed by blinded study group. We also assessed anti-H1 haemagglutinin stalk, anti-H2, anti-H9, and anti-H18 IgG antibody titres and plasmablast and memory B-cell responses in peripheral blood. This trial is registered with ClinicalTrials.gov , number NCT03300050 . Findings Between Oct 10, 2017, and Nov 27, 2017, 65 participants were enrolled and randomly assigned. The adjuvanted inactivated vaccine, but not the live-attenuated vaccine, induced a substantial serum IgG antibody response after the prime immunisation, with a seven times increase in anti-H1 stalk antibody titres on day 29. After boost immunisation, all vaccine regimens induced detectable anti-H1 stalk antibody (2·2–5·6 times induction over baseline), cross-reactive serum IgG antibody, and peripheral blood plasmablast responses. An unsolicited adverse event was reported for 29 (48%) of 61 participants. Solicited local adverse events were reported in 12 (48%) of 25 participants following prime vaccination with intramuscular study product or placebo, in 12 (33%) of 36 after prime immunisation with intranasal study product or placebo, and in 18 (32%) of 56 following booster doses of study product or placebo. Solicited systemic adverse events were reported in 14 (56%) of 25 after prime immunisation with intramuscular study product or placebo, in 22 (61%) of 36 after immunisation with intranasal study product or placebo, and in 21 (38%) of 56 after booster doses of study product or placebo. Disaggregated safety data were not available at the time of this interim analysis. Interpretation The tested chimeric haemagglutinin-based, universal influenza virus vaccine regimens elicited cross-reactive serum IgG antibodies that targeted the conserved haemagglutinin stalk domain. This is the first proof-of-principle study to show that high anti-stalk titres can be induced by a rationally designed vaccine in humans and opens up avenues for further development of universal influenza virus vaccines. On the basis of the blinded study group, the vaccine regimens were tolerable and no safety concerns were observed. Funding Bill & Melinda Gates Foundation.

Summary Respiratory syncytial virus (RSV) is a major cause of serious acute lower respiratory tract infection in infants and the elderly. The lack of a licensed RSV vaccine calls for the development of vaccines with other targets and vaccination strategies. Here, we construct a recombinant protein, designated P-KFD1, comprising RSV phosphoprotein (P) and the E.-coli-K12-strain-derived flagellin variant KFD1. Intranasal immunization with P-KFD1 inhibits RSV replication in the upper and lower respiratory tract and protects mice against lung disease without vaccine-enhanced disease (VED). The P-specific CD4+ T cells provoked by P-KFD1 intranasal (i.n.) immunization either reside in or migrate to the respiratory tract and mediate protection against RSV infection. Single-cell RNA sequencing (scRNA-seq) and carboxyfluorescein succinimidyl ester (CFSE)-labeled cell transfer further characterize the Th1 and Th17 responses induced by P-KFD1. Finally, we find that anti-viral protection depends on either interferon-γ (IFN-γ) or interleukin-17A (IL-17A). Collectively, P-KFD1 is a promising safe and effective mucosal vaccine candidate for the prevention of RSV infection.

Although millions of patients have clinically recovered from COVID-19, little is known about the immune status of lymphocytes in these individuals. In this study, the peripheral blood mononuclear cells of a clinically recovered (CR) cohort were comparatively analyzed with those of an age- and sex-matched healthy donor cohort. We found that CD8+ T cells in the CR cohort had higher numbers of effector T cells and effector memory T cells but lower Tc1 (IFN-γ+), Tc2 (IL-4+), and Tc17 (IL-17A+) cell frequencies. The CD4+ T cells of the CR cohort were decreased in frequency, especially the central memory T cell subset. Moreover, CD4+ T cells in the CR cohort showed lower programmed cell death protein 1 (PD-1) expression and had lower frequencies of Th1 (IFN-γ+), Th2 (IL-4+), Th17 (IL-17A+), and circulating follicular helper T (CXCR5+PD-1+) cells. Accordingly, the proportion of isotype-switched memory B cells (IgM−CD20hi) among B cells in the CR cohort showed a significantly lower proportion, although the level of the activation marker CD71 was elevated. For CD3−HLA-DR− lymphocytes in the CR cohort, in addition to lower levels of IFN-γ, granzyme B and T-bet, the correlation between T-bet and IFN-γ was not observed. Additionally, by taking into account the number of days after discharge, all the phenotypes associated with reduced function did not show a tendency toward recovery within 4‒11 weeks. The remarkable phenotypic alterations in lymphocytes in the CR cohort suggest that severe acute respiratory syndrome coronavirus 2 infection profoundly affects lymphocytes and potentially results in dysfunction even after clinical recovery.

Dengue is a significant public health issue, affecting hundreds of millions of people worldwide. As it is spreading from tropical and subtropical zones, some regions previously recognised as non-endemic are at risk of becoming endemic. However, the global circulation of dengue is not fully understood and quantitative measurements of endemicity levels are lacking, posing an obstacle in the precise control of dengue spread. In this study, a sequence-based pipeline was designed based on random sampling to study the transmission of dengue. The limited intercontinental transmission was identified, while regional circulation of dengue was quantified in terms of importation, local circulation and exportation. Additionally, hypo- and hyper-endemic regions were identified using a new metric, with the former characterised by low local circulation and increased importation, whereas the latter by high local circulation and reduced importation. In this study, the global circulation pattern of dengue was examined and a sequence-based endemicity measurement was proposed, which will be helpful for future surveillance and targeted control of dengue.

SUMMARYRespiratory syncytial virus (RSV) is a major cause of serious acute lower respiratory tract infection in infants and the elderly. No licensed RSV vaccine available thus far calls for the development of vaccines with new target(s) and vaccination strategies. Here, we constructed a recombinant protein, designated P-KFD1, comprised of RSV phosphoprotein (P) and E. coli K12 strain-derived flagellin variant KFD1. Intranasal (i.n.) immunization with P-KFD1 inhibits RSV replication in both upper and lower respiratory tract, and protects mice against lung disease without vaccine-enhanced disease (VED). The P-specific CD4+ T cells provoked by P-KFD1 i.n. immunization, either reside in or migrate to respiratory tract, mediate protection against RSV infection. Sc-RNA seq and carboxyfluorescein succinimidyl ester (CFSE) labeled cell transfer further characterized the Th1 and Th17 responses induced by P-KFD1. Finally, we found the anti-viral protection depends on either IFN-γ or IL-17A. Collectively, P-KFD1 is promising as a safe and effective mucosal vaccine candidate to prevent RSV infection.HIGHLIGHTSA new subunit RSV vaccine candidate with new target and vaccination strategy, P-KFD1, is designed and generatedIntranasal immunization with P-KFD1 protects mice against RSV infection and averts vaccine-enhanced diseaseSc-RNA seq and CFSE-labelled cell transfer identified characteristics of the protective CD4+ T cellsLocal and peripheral CD4+ T cells provide protection against RSV infection dependent on either IFN-γ or IL-17A

The COVID-19 pandemic, caused by the SARS-CoV-2 infection, is a global health crisis. While many patients have clinically recovered, little is known about long-term alterations in T cell responses of COVID-19 convalescents. In this study, T cell responses in peripheral blood mononuclear cells of a long-time COVID-19 clinically recovered (20–26 weeks) cohort (LCR) were measured via flow cytometry and ELISpot. The T cell responses of LCR were comparatively analyzed against an age and sex matched short-time clinically recovered (4–9 weeks) cohort (SCR) and a healthy donor cohort (HD). All volunteers were recruited from Wuhan Jinyintan Hospital, China. Phenotypic analysis showed that activation marker PD-1 expressing on CD4+ T cells of LCR was still significantly lower than that of HD. Functional analysis indicated that frequencies of Tc2, Th2 and Th17 in LCR were comparable to those of HD, but Tc17 was higher than that of HD. In LCR, compared to the HD, there were fewer IFN-γ producing T cells but more IL-2 secreting T cells. In addition, the circulating Tfh cells in LCR were still slightly lower compared to HD, though the subsets composition had recovered. Remarkably, SARS-CoV-2 specific T cell responses in LCR were comparable to that of SCR. Collectively, T cell responses experienced long-term alterations in phenotype and functional potential of LCR cohort. However, after clinical recovery, SARS-CoV-2 specific T cell responses could be sustained at least for six months, which may be helpful in resisting re-infection. Electronic supplementary material The online version of this article (10.1007/s12250-021-00348-0) contains supplementary material, which is available to authorized users.

RSV is a major cause of serious acute lower respiratory tract infection in infants and the elderly. No licensed RSV vaccine available thus far calls for the development of vaccines with new target(s) and vaccination strategies. Here, we constructed a recombinant protein, designated P-KFD1, comprised of RSV phosphoprotein (P) and E. coli K12 strain-derived flagellin variant KFD1. Intranasal (i.n.) immunization with P-KFD1 inhibits RSV replication in both upper and lower respiratory tract, and protects mice against lung disease without vaccine-enhanced disease (VED). The P-specific CD4+ T cells provoked by P-KFD1 i.n. immunization, either reside in or migrate to respiratory tract, mediate protection against RSV infection. Sc-RNA seq and CFSE-labeled cell transfer further characterized the Th1 and Th17 responses induced by P-KFD1. Finally, we found the anti-viral protection depends on either IFN-γ or IL-17A. Collectively, P-KFD1 is promising as a safe and effective mucosal vaccine candidate to prevent RSV infection.

B cell response plays a critical role against SARS-CoV-2 infection. However, little is known about the diversity and frequency of the paired SARS-CoV-2 antigen-specific BCR repertoire after SARS-CoV-2 infection. Here, we performed single-cell RNA sequencing and VDJ sequencing using the memory and plasma B cells isolated from five convalescent COVID-19 patients, and analyzed the spectrum and transcriptional heterogeneity of antibody immune responses. Via linking BCR to antigen specificity through sequencing (LIBRA-seq), we identified a distinct activated memory B cell subgroup (CD11chighCD95high) had a higher proportion of SARS-CoV-2 antigen-labeled cells compared with memory B cells. Our results revealed the diversity of paired BCR repertoire and the non-stochastic pairing of SARS-CoV-2 antigen-specific immunoglobulin heavy and light chains after SARS-CoV-2 infection. The public antibody clonotypes were shared by distinct convalescent individuals. Moreover, several antibodies isolated by LIBRA-seq showed high binding affinity against SARS-CoV-2 receptor-binding domain (RBD) or nucleoprotein (NP) via ELISA assay. Two RBD-reactive antibodies C14646P3S and C2767P3S isolated by LIBRA-seq exhibited high neutralizing activities against both pseudotyped and authentic SARS-CoV-2 viruses in vitro. Our study provides fundamental insights into B cell response following SARS-CoV-2 infection at the single-cell level.

Although human antibodies elicited by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) nucleocapsid (N) protein are profoundly boosted upon infection, little is known about the function of N-reactive antibodies. Herein, we isolated and profiled a panel of 32 N protein-specific monoclonal antibodies (mAbs) from a quick recovery coronavirus disease-19 (COVID-19) convalescent patient who had dominant antibody responses to the SARS-CoV-2 N protein rather than to the SARS-CoV-2 spike (S) protein. The complex structure of the N protein RNA binding domain with the mAb with the highest binding affinity (nCoV396) revealed changes in the epitopes and antigen’s allosteric regulation. Functionally, a virus-free complement hyper-activation analysis demonstrated that nCoV396 specifically compromises the N protein-induced complement hyper-activation, which is a risk factor for the morbidity and mortality of COVID-19 patients, thus laying the foundation for the identification of functional anti-N protein mAbs.

Although human antibodies elicited by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) nucleocapsid (N) protein are profoundly boosted upon infection, little is known about the function of N-reactive antibodies. Herein, we isolate and profile a panel of 32 N protein-specific monoclonal antibodies (mAbs) from a quick recovery coronavirus disease-19 (COVID-19) convalescent patient who has dominant antibody responses to the SARS-CoV-2 N protein rather than to the SARS-CoV-2 spike (S) protein. The complex structure of the N protein RNA binding domain with the highest binding affinity mAb (nCoV396) reveals changes in the epitopes and antigen’s allosteric regulation. Functionally, a virus-free complement hyperactivation analysis demonstrates that nCoV396 specifically compromises the N protein-induced complement hyperactivation, which is a risk factor for the morbidity and mortality of COVID-19 patients, thus laying the foundation for the identification of functional anti-N protein mAbs.

Background The novel COVID-19 disease has spread worldwide, resulting in a new pandemic. The Chinese government implemented strong intervention measures in the early stage of the epidemic, including strict travel bans and social distancing policies. Prioritizing the analysis of different contributing factors to outbreak outcomes is important for the precise prevention and control of infectious diseases. We proposed a novel framework for resolving this issue and applied it to data from China. Objective This study aimed to systematically identify national-level and city-level contributing factors to the control of COVID-19 in China. Methods Daily COVID-19 case data and related multidimensional data, including travel-related, medical, socioeconomic, environmental, and influenza-like illness factors, from 343 cities in China were collected. A correlation analysis and interpretable machine learning algorithm were used to evaluate the quantitative contribution of factors to new cases and COVID-19 growth rates during the epidemic period (ie, January 17 to February 29, 2020). Results Many factors correlated with the spread of COVID-19 in China. Travel-related population movement was the main contributing factor for new cases and COVID-19 growth rates in China, and its contributions were as high as 77% and 41%, respectively. There was a clear lag effect for travel-related factors (previous vs current week: new cases, 45% vs 32%; COVID-19 growth rates, 21% vs 20%). Travel from non-Wuhan regions was the single factor with the most significant impact on COVID-19 growth rates (contribution: new cases, 12%; COVID-19 growth rate, 26%), and its contribution could not be ignored. City flow, a measure of outbreak control strength, contributed 16% and 7% to new cases and COVID-19 growth rates, respectively. Socioeconomic factors also played important roles in COVID-19 growth rates in China (contribution, 28%). Other factors, including medical, environmental, and influenza-like illness factors, also contributed to new cases and COVID-19 growth rates in China. Based on our analysis of individual cities, compared to Beijing, population flow from Wuhan and internal flow within Wenzhou were driving factors for increasing the number of new cases in Wenzhou. For Chongqing, the main contributing factor for new cases was population flow from Hubei, beyond Wuhan. The high COVID-19 growth rates in Wenzhou were driven by population-related factors. Conclusions Many factors contributed to the COVID-19 outbreak outcomes in China. The differential effects of various factors, including specific city-level factors, emphasize the importance of precise, targeted strategies for controlling the COVID-19 outbreak and future infectious disease outbreaks.

Objective SARS‐CoV‐2 has caused a worldwide pandemic of COVID‐19. The existence of prolonged SARS‐CoV‐2 positivity (PP) has further increased the burden on the health system. Since T cells are vital for viral control, we aimed to evaluate the characteristics of T‐cell responses associated with PP. Methods We established a PP cohort and two age‐ and sex‐matched control cohorts: a regular clinical recovery (CR) cohort and a healthy donor (HD) cohort. The mean time for RNA negativity conversion in the PP cohort was markedly longer than that in the CR cohort (66.2 vs 25.3 days), while the time from illness onset to sampling was not significantly different. T‐cell responses in the PP cohort were assayed, analysed and compared with those in the CR and HD cohorts by flow cytometry and ELISpot analysis of peripheral blood mononuclear cells. Results Compared with the CR cohort, the proliferation, activation and functional potential of CD8+ and CD4+ T cells in the PP cohort were not significantly different. However, the frequencies and counts of Teff and Tem in CD8+ but not in CD4+ T cells of the PP cohort were prominently lower. Moreover, a weaker SARS‐CoV‐2 N protein‐specific IFN‐γ+ T‐cell response and a higher frequency of Tregs were detected in the PP cohort. Conclusion Suppressed CD8+ T‐cell differentiation is associated with PP and may be an indicator for the prediction of prolonged SARS‐CoV‐2 positivity in COVID‐19 patients. The association between suppressed CD8+ T‐cell differentiation and elevated Tregs warrants studies in the future., In this study, the characteristics of T‐cell responses of peripheral blood mononuclear cells in a COVID‐19 cohort with prolonged SARS‐CoV‐2‐positive (Pht P) were comparatively assayed and analysed with those in clinically recovered (CR) and healthy donor (HD) cohorts by FACS and ELISpot analysis of peripheral blood mononuclear cells. The results showed that the frequencies of Teff and Tem in CD8+ but not in CD4+ of the PP cohort were prominently lower than those of the CR cohort. Moreover, lowered SARS‐CoV‐2 N protein‐specific IFN‐γ+ T‐cell response and higher frequency of Tregs were detected in the PP cohort.

Although human antibodies elicited by severe acute respiratory distress syndrome coronavirus-2 (SARS-CoV-2) nucleocapsid (N) protein are profoundly boosted upon infection, little is known about the function of N-directed antibodies. Herein, we isolated and profiled a panel of 32 N protein-specific monoclonal antibodies (mAb) from a quick recovery coronavirus disease-19 (COVID-19) convalescent, who had dominant antibody responses to SARS-CoV-2 N protein rather than to Spike protein. The complex structure of N protein RNA binding domain with the highest binding affinity mAb nCoV396 reveals the epitopes and antigen’s allosteric changes. Functionally, a virus-free complement hyper-activation analysis demonstrates that nCoV396 specifically compromises N protein-induced complement hyper-activation, a risk factor for morbidity and mortality in COVID-19, thus paving the way for functional anti-N mAbs identification.One Sentence SummaryB cell profiling, structural determination, and protease activity assays identify a functional antibody to N protein.

BACKGROUND The novel COVID-19 disease has spread worldwide, resulting in a new pandemic. The Chinese government implemented strong intervention measures in the early stage of the epidemic, including strict travel bans and social distancing policies. Prioritizing the analysis of different contributing factors to outbreak outcomes is important for the precise prevention and control of infectious diseases. We proposed a novel framework for resolving this issue and applied it to data from China. OBJECTIVE This study aimed to systematically identify national-level and city-level contributing factors to the control of COVID-19 in China. METHODS Daily COVID-19 case data and related multidimensional data, including travel-related, medical, socioeconomic, environmental, and influenza-like illness factors, from 343 cities in China were collected. A correlation analysis and interpretable machine learning algorithm were used to evaluate the quantitative contribution of factors to new cases and COVID-19 growth rates during the epidemic period (ie, January 17 to February 29, 2020). RESULTS Many factors correlated with the spread of COVID-19 in China. Travel-related population movement was the main contributing factor for new cases and COVID-19 growth rates in China, and its contributions were as high as 77% and 41%, respectively. There was a clear lag effect for travel-related factors (previous vs current week: new cases, 45% vs 32%; COVID-19 growth rates, 21% vs 20%). Travel from non-Wuhan regions was the single factor with the most significant impact on COVID-19 growth rates (contribution: new cases, 12%; COVID-19 growth rate, 26%), and its contribution could not be ignored. City flow, a measure of outbreak control strength, contributed 16% and 7% to new cases and COVID-19 growth rates, respectively. Socioeconomic factors also played important roles in COVID-19 growth rates in China (contribution, 28%). Other factors, including medical, environmental, and influenza-like illness factors, also contributed to new cases and COVID-19 growth rates in China. Based on our analysis of individual cities, compared to Beijing, population flow from Wuhan and internal flow within Wenzhou were driving factors for increasing the number of new cases in Wenzhou. For Chongqing, the main contributing factor for new cases was population flow from Hubei, beyond Wuhan. The high COVID-19 growth rates in Wenzhou were driven by population-related factors. CONCLUSIONS Many factors contributed to the COVID-19 outbreak outcomes in China. The differential effects of various factors, including specific city-level factors, emphasize the importance of precise, targeted strategies for controlling the COVID-19 outbreak and future infectious disease outbreaks.

SummaryBackgroundLymphopenia is a typical symptom in the COVID-19 patients. While millions of patients are clinical recovered, little is known about the immune status of lymphocytes in these individuals.MethodsA clinical recovered cohort (CR) of 55 COVID-19 individuals (discharged from hospital 4 to 11 weeks), and 55 age and sex matched healthy donors cohort (HD) were recruited. Detailed analysis on phenotype of the lymphocytes in peripheral blood mononuclear cells (PBMCs) was performed by flow cytometry.FindingsCompared with cohort HD, the CD8+T cells in cohort CR had higher Teff and Tem, but lower Tc1 (IFN-γ+), Tc2 (IL-4+) and Tc17 (IL-17A+) frequencies. The CD4+T cells of CR had decreased frequency, especially on the Tcm subset. Moreover, CD4+T cells of CR expressed lower PD-1 and had lower frequencies of Th1 (IFN-γ+), Th2 (IL-4+), Th17 (IL-17A+) as well as circulating Tfh (CXCR5+PD-1+). Accordingly, isotype-switched memory B cell (IgM-CD20hi) in CR had significantly lower proportion in B cells, though level of activation marker CD71 elevated. For CD3-HLA-DRlolymphocytes of CR, besides levels of IFN-γ, Granzyme B and T-bet were lower, the correlation between T-bet and IFN-γ became irrelevant. In addition, taken into account of discharged days, all the lowered function associated phenotypes showed no recovery tendency within whole observation period.InterpretationThe CR COVID-19 individuals still showed remarkable phenotypic alterations in lymphocytes after clinical recovery 4 to 11 weeks. This suggests SARS-CoV-2 infection imprints profoundly on lymphocytes and results in long-lasting potential dysfunctions.FundingKunming Science and Technology Department (2020-1-N-037)

Humans are repeatedly exposed to variants of influenza virus throughout their lifetime. As a result, preexisting influenza-specific memory B cells can dominate the response after infection or vaccination. Memory B cells recalled by adulthood exposure are largely reactive to conserved viral epitopes present in childhood strains, posing unclear consequences on the ability of B cells to adapt to and neutralize newly emerged strains. We sought to investigate the impact of preexisting immunity on generation of protective antibody responses to conserved viral epitopes upon influenza virus infection and vaccination in humans. We accomplished this by characterizing monoclonal antibodies (mAbs) from plasmablasts, which are predominantly derived from preexisting memory B cells. We found that, whereas some influenza infection-induced mAbs bound conserved and neutralizing epitopes on the hemagglutinin (HA) stalk domain or neuraminidase, most of the mAbs elicited by infection targeted non-neutralizing epitopes on nucleoprotein and other unknown antigens. Furthermore, most infection-induced mAbs had equal or stronger affinity to childhood strains, indicating recall of memory B cells from childhood exposures. Vaccination-induced mAbs were similarly induced from past exposures and exhibited substantial breadth of viral binding, although, in contrast to infection-induced mAbs, they targeted neutralizing HA head epitopes. Last, cocktails of infection-induced mAbs displayed reduced protective ability in mice compared to vaccination-induced mAbs. These findings reveal that both preexisting immunity and exposure type shape protective antibody responses to conserved influenza virus epitopes in humans. Natural infection largely recalls cross-reactive memory B cells against non-neutralizing epitopes, whereas vaccination harnesses preexisting immunity to target protective HA epitopes.