IntroductionFollicular helper T (TFH) cells are essential for germinal center reactions and the maintenance of long-lived humoral immunity. Transforming growth factor-β (TGF-β) is a multifunctional cytokine implicated in immune regulation, T-cell differentiation, and the maintenance of cellular stemness. Prior studies have shown that TGF-β promotes stemness across a wide range of cell types and facilitates the differentiation of naïve CD4⁺ T cells into various T helper cell subsets. However, its precise effects on TFH effector function and stem-like properties remain poorly understood.MethodsThe dual regulatory roles of TGF-β1 in modulating TFH effector functions and stem-like properties were investigated using flow cytometry-based phenotyping, co-culture assays with memory B cells, proliferation and apoptosis assays, ELISA for antibody production, and bulk RNA sequencing of naïve-derived and blood-derived TFH cells.ResultsWe found that TGF-β1 treatment in vitro promoted human naïve CD4+ T cells differentiation into CXCR3+ TFH, but significantly attenuated their effector molecule expression and TFH-mediated memory B-cell differentiation and antibody production, whereas it enhanced the expression of stemness-associated molecules in TFH cells both differentiated in vitro from naïve CD4+ T cells and isolated from blood. Notably, TGF-β1 promoted proliferation and reduced apoptosis of naïve-derived TFH cells in vitro, but suppressed proliferation and increased early apoptosis in blood-derived mature TFH cells.DiscussionOur findings indicate that TGF-β1 tunes the balance between TFH effector function and stem-like properties, and show differential regulations of the early phase of TFH differentiation and mature TFH cells, which may have implications for TFH-driven immune pathology and disease.

The outbreaks of severe acute respiratory syndrome coronavirus (SARS-CoV-1), Middle East respiratory syndrome coronavirus (MERS-CoV), and SARS-CoV-2 highlight the need for countermeasures to prevent future coronavirus pandemics. Given the unpredictable nature of spillover events, preparing antibodies with broad coronavirus-neutralizing activity is an ideal proactive strategy. Here, we investigated whether SARS-CoV-2 infection and vaccination could provide cross-neutralizing antibodies (nAbs) against zoonotic sarbecoviruses. We evaluated the cross-neutralizing profiles of plasma and monoclonal antibodies constructed from B cells from coronavirus disease 2019 (COVID-19) convalescents and vaccine recipients; against sarbecoviruses originating from bats, civets, and pangolins; and against SARS-CoV-1 and SARS-CoV-2. We found that both SARS-CoV-2 infection and vaccination elicited broad cross-nAbs against multiple sarbecoviruses, and vaccination boosters significantly augmented the magnitude and breadth of nAbs to sarbecoviruses. Of the nAbs, several exhibited neutralization activity against multiple sarbecoviruses by targeting the spike receptor-binding domain (RBD) and competing with angiotensin-converting enzyme 2 (ACE2) binding. SCM12-61 demonstrated exceptional potency, with half-maximal inhibitory concentration (IC 50 ) values of 0.001–0.091 μg/mL, indicating its potential for combating new sarbecovirus outbreaks. Collectively, our findings suggest that both SARS-CoV-2 infection and current vaccination schemes elicit broad cross-neutralizing antibodies against diverse sarbecoviruses, enforcing prevention and therapeutic strategies for future sarbecovirus spillover events.

AbstractAedes albopictusshows a rapid global expansion and dramatic vectorial capacity for various arboviruses. Mosquitoes display distinct sexual dimorphisms, only adult females consume blood meals to complete ovarian follicle development. Therefore, cyclic reproduction in female mosquitoes serves as a foundation for the transmission of numerous disease-causing pathogens. Aedes have an expansion of the piRNA biogenesis genes, indicated that piRNA may play multiple functional roles in mosquitoes. Although the antiviral function of piRNA pathway in mosquitoes has been extensively studied, the role of piRNAs in mosquito reproduction remain to be further understood. In the present study, we first profiled the characteristics of sex-biased piRNAs in adultAe.albopictus. Then, we identified a female biased piRNA (Aalpi18529) in adult females, that was highly expressed in ovaries at blood feeding-dependent termination, and depended on PIWI5 and ago3 mediated biogenesis. Aalpi18529 overexpression suppressed ovarian development, and reduced fertility and fecundity in adult females post-bloodmeal. Furthermore, we demonstrated that Aalpi18529 can effectively repress its direct target, growth arrest and DNA-damage-inducible protein 45a (GADD45A), and eventually regulates ovarian development via theGadd45a-mediated JNK-dependent nurse cell apoptosis pathway. Our study is the first to report an endogenous piRNA, which trigger silencing of an important protein-coding gene by posttranscriptional regulation in mosquitoes, expanding our current understanding of the important and multiple roles of piRNAs in biological processes inAe. albopictus.Author SummaryHere, we conducted high throughput piRNA sequencing and comprehensive analysis of piRNA sex-based expression profiles in adult females and males of Aedes albopictus. Based on several established universal tools for research, we demonstrate an ovary-enriched endogenous piRNA, Aalpi-18529, is involved in the regulation of the apoptosis of nurse cells during vitellogenesis via the GADD45A/phosphorylated JNK (pJNK) axis and ultimately affects ovarian development. In general, uncovering the biological functions of sex-biased piRNAs in Aedes albopictus will enhance the understanding of piRNA roles in mosquito Sexual dimorphism (SD) and will provide provide more information about the high reproductive capacity of Aedes albopictus, which is essential to find alternative control strategies.Classification:Research Reports

IntroductionSince December 2019, the emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causing coronavirus disease 2019 (COVID-19) has presented considerable public health challenges. Multiple vaccines have been used to induce neutralizing antibodies (nAbs) and memory B-cell responses against the viral spike (S) glycoprotein, and many essential epitopes have been defined. Previous reports have identified severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike-reactive naïve B cells and preexisting memory B cells in unexposed individuals. However, the role of these spike-reactive B cells in vaccine-induced immunity remains unknown.MethodsTo elucidate the characteristics of preexisting SARS-CoV-2 S-reactive B cells as well as their maturation after antigen encounter, we assessed the relationship of spike-reactive B cells before and after vaccination in unexposed human individuals. We further characterized the sequence identity, targeting domain, broad-spectrum binding activity and neutralizing activity of these SARS-CoV-2 S-reactive B cells by isolating monoclonal antibodies (mAbs) from these B cells.ResultsThe frequencies of both spike-reactive naïve B cells and preexisting memory B cells before vaccination correlated with the frequencies of spike-reactive memory B cells after vaccination. Isolated mAbs from spike-reactive naïve B cells before vaccination had fewer somatic hypermutations (SHMs) than mAbs isolated from spike-reactive memory B cells before and after vaccination, but bound SARS-CoV-2 spike in vitro. Intriguingly, these germline-like mAbs possessed broad binding profiles for SARS-CoV-2 and its variants, although with low or no neutralizing capacity. According to tracking of the evolution of IGHV4-4/IGKV3-20 lineage antibodies from a single donor, the lineage underwent SHMs and developed increased binding activity after vaccination.DiscussionOur findings suggest that spike-reactive naïve B cells can be expanded and matured by vaccination and cocontribute to vaccine-elicited antibody responses with preexisting memory B cells. Selectively and precisely targeting spike-reactive B cells by rational antigen design may provide a novel strategy for next-generation SARS-CoV-2 vaccine development.

Aedes albopictus shows a rapid global expansion and dramatic vectorial capacity for various arboviruses. Mosquitoes display distinct sexual dimorphisms, only adult females consume blood meals to complete ovarian follicle development. Therefore, cyclic reproduction in female mosquitoes serves as a foundation for the transmission of numerous disease-causing pathogens. Aedes have an expansion of the piRNA biogenesis genes, indicated that piRNA may play multiple functional roles in mosquitoes. Although the antiviral function of piRNA pathway in mosquitoes has been extensively studied, the role of piRNAs in mosquito reproduction remain to be further understood. In the present study, we first profiled the characteristics of sex-biased piRNAs in adult Ae.albopictus . Then, we identified a female biased piRNA (Aalpi18529) in adult females, that was highly expressed in ovaries at blood feeding-dependent termination, and depended on PIWI5 and ago3 mediated biogenesis. Aalpi18529 overexpression suppressed ovarian development, and reduced fertility and fecundity in adult females post-bloodmeal. Furthermore, we demonstrated that Aalpi18529 can effectively repress its direct target, growth arrest and DNA-damage-inducible protein 45a (GADD45A), and eventually regulates ovarian development via the Gadd45a -mediated JNK-dependent nurse cell apoptosis pathway. Our study is the first to report an endogenous piRNA, which trigger silencing of an important protein-coding gene by posttranscriptional regulation in mosquitoes, expanding our current understanding of the important and multiple roles of piRNAs in biological processes in Ae. albopictus. ### Competing Interest Statement The authors have declared no competing interest.

The concern of COVID-19 persists due to the continuous emergence of variants and the potential spillover of animal coronaviruses. The broad-spectrum neutralizing antibodies play a pivotal role in the prevention and treatment of coronavirus (CoV) infections. Here, we constructed 18 bi-specific antibodies (bsAbs) using 9 antibodies isolated from COVID-19 convalescents and vaccinated individuals, designed as dual variable domain immunoglobulin (DVD-Ig). A bsAb 5-HI showed a high binding capability to the S1 subunit of spike and exhibited breadth and potency against pseudotyped SARS-CoV-2 variants of concerns (VOCs) and SARS-related-CoVs (SARSr-CoVs), with half maximal effective concentration (EC50) of 0.028–3.444 nM and 50

Regulatory T (Treg) cells are involved in the antiviral immune response in patients with coronavirus disease 2019 (COVID-19); however, whether Treg cells are involved in the neutralizing antibody (nAb) response remains unclear. Here, we found that individuals who recovered from mild but not severe COVID-19 had significantly greater frequencies of Treg cells and lower frequencies of CXCR3+ circulating T follicular helper (cTfh) cells than healthy controls. Furthermore, the frequencies of Treg and CXCR3+ cTfh cells were negatively and positively correlated with the nAb responses, respectively, and Treg cells was inversely associated with CXCR3+ cTfh cells in individuals who recovered from mild COVID-19 but not in those with severe disease. Mechanistically, Treg cells inhibited memory B-cell differentiation and antibody production by limiting the activation and proliferation of cTfh cells, especially CXCR3+ cTfh cells, and functional molecule expression. This study provides novel insight showing that mild COVID-19 elicits concerted nAb responses, which are shaped by both Treg and Tfh cells.

• Five broad-spectrum mAbs identified from COVID-19 convalescents and vaccinees possessed the ability to recognize spike S2 of pan-coronaviruses. • Five mAbs targeted distinct but conserved epitopes on spike S2 of pan-coronaviruses. • Three of these five mAbs were competitively bound to the fusion peptide epitopes of coronavirus spike protein.

IntroductionAedes albopictus is an aggressive invasive mosquito species widely distributed around the world, and it is also a known vector of arboviruses. Virus metagenomics and RNA interference (RNAi) are important in studying the biology and antiviral defense of Ae. albopictus. However, the virome and potential transmission of plant viruses by Ae. albopictus remain understudied. MethodsMosquito samples of Ae. albopictus were collected from Guangzhou, China, and small RNA sequencing was performed. Raw data were filtered, and virus-associated contigs were generated using VirusDetect. The small RNA profiles were analyzed, and maximum-likelihood phylogenetic trees were constructed. ResultsThe small RNA sequencing of pooled Ae. albopictus revealed the presence of five known viruses, including Wenzhou sobemo-like virus 4, mosquito nodavirus, Aedes flavivirus, Hubei chryso-like virus 1, and Tobacco rattle virus RNA1. Additionally, 21 new viruses that had not been previously reported were identified. The mapping of reads and contig assembly provided insights into the viral diversity and genomic characteristics of these viruses. Field survey confirmed the detection of the identified viruses in Ae. albopictus collected from Guangzhou. DiscussionThe comprehensive analysis of the virus metagenomics of Ae. albopictus in this study sheds light on the diversity and prevalence of viruses in mosquito populations. The presence of known and novel viruses highlights the need for continued surveillance and investigation into their potential impact on public health. The findings also emphasize the importance of understanding the virome and potential transmission of plant viruses by Ae. albopictus. ConclusionThis study provides valuable insights into the virome of Ae. albopictus and its potential role as a vector for both known and novel viruses. Further research is needed to expand the sample size, explore additional viruses, and investigate the implications for public health.

•BF.7 and BQ.1, escaped most neutralizing antibodies isolated from the recovered COVID-19 individuals and vaccine recipients.•Five potent mAbs were identified that showed a broad neutralizing profile to Omicron subvariants and other VOCs.•These broad neutralizing antibodies targeted the RBD of the spike protein, competing with ACE2 for binding.

The initial signals governing sex determination vary widely among insects. Here we show that Armigeres subalbatus M factor (AsuMf), a male-specific duplication of an autosomal gene of the Drosophila behaviour/human splicing (DBHS) gene family, is the potential primary signal for sex determination in the human filariasis vector mosquito, Ar. subalbatus. Our results show that AsuMf satisfies two fundamental requirements of an M factor: male-specific expression and early embryonic expression. Ablations of AsuMf result in a shift from male- to female-specific splicing of doublesex and fruitless, leading to feminization of males both in morphology and general transcription profile. These data support the conclusion that AsuMf is essential for male development in Ar. subalbatus and reveal a male-determining factor that is derived from duplication and subsequent neofunctionalization of a member of the conserved DBHS family.

Aedes albopictus shows a rapid global expansion and dramatic vectorial capacity for various arboviruses, thus posing a severe threat to global health. Although many noncoding RNAs have been confirmed to play functional roles in various biological processes in Ae. albopictus, the roles of circRNA remain a mystery. In the present study, we first performed high-throughput circRNA sequencing in Ae. albopictus. Then, we identified a cysteine desulfurase (CsdA) superfamily gene-originated circRNA, named aal-circRNA-407, which was the third most abundant circRNA in adult females and displayed a fat body highly expressed manifestation and blood feeding-dependent onset. SiRNA-mediated knockdown of circRNA-407 resulted in a decrease in the number of developing follicles and a reduction in follicle size post blood meal. Furthermore, we demonstrated that circRNA-407 can act as a sponge of aal-miR-9a-5p to promote the expression of its target gene Foxl and eventually regulate ovarian development. Our study is the first to report a functional circRNA in mosquitoes, expanding our current understanding of important biological roles in mosquitoes and providing an alternative genetic strategy for mosquito control.

Background Dengue virus (DENV) is a major public health threat, with Aedes albopictus being the confirmed vector responsible for dengue epidemics in Guangzhou, China. Mosquito densoviruses (MDVs) are pathogenic mosquito-specific viruses, and a novel MDV was previously isolated from Ae. albopictus in Guangzhou. This study aims to determine the prevalence of MDVs in wild Ae. albopictus populations and investigate their potential interactions with DENV and impact on vector susceptibility for DENV. Methods The prevalence of MDV in wild mosquitoes in China was investigated using open access sequencing data and PCR detection in Ae. albopictus in Guangzhou. The viral infection rate and titers in MDV-persistent C6/36 cells were evaluated at 12, 24, 48, 72, 96, and 120 h post infection (hpi) by indirect immunofluorescence assay (IFA) and real time quantitative PCR (RT-qPCR). The midgut infection rate (MIR), dissemination rate (DR), and salivary gland infection rate (SGIR) in various tissues of MDV-infected mosquitoes were detected and quantified at 0, 5, 10, and 15 days post infection (dpi) by RT-PCR and RT-qPCR. The chi-square test evaluated dengue virus serotype 2 (DENV-2) and Aedes aegypti densovirus (AaeDV) infection rates and related indices in mosquitoes, while Tukey's LSD and t -tests compared viral titers in C6/36 cells and tissues over time. Results The results revealed a relatively wide distribution of MDVs in Aedes , Culex , and Anopheles mosquitoes in China and an over 68% positive rate. In vitro , significant reductions in DENV-2 titers in supernatant at 120 hpi, and an apparent decrease in DENV-2-positive cells at 96 and 120 hpi were observed. In vivo, DENV-2 in the ovaries and salivary glands was first detected at 10 dpi in both monoinfected and superinfected Ae. albopictus females, while MDV superinfection with DENV-2 suppressed the salivary gland infection rate at 15 dpi. DENV-2 titer in the ovary and salivary glands of Ae. albopictus was reduced in superinfected mosquitoes at 15 dpi. Conclusions MDVs is widespread in natural mosquito populations, and replication of DENV-2 is suppressed in MDV-infected Ae. albopictus , thus reducing vector susceptibility to DENV-2. Our study supports the hypothesis that MDVs may contribute to reducing transmission of DENV and provides an alternative strategy for mosquito-transmitted disease control. Graphical abstract

Long-term humoral immunity to SARS-CoV-2 is essential for preventing reinfection. The production of neutralizing antibody (nAb) and B cell differentiation are tightly regulated by T follicular help (T FH ) cells. However, the longevity and functional role of T FH cell subsets in COVID-19 convalescents and vaccine recipients remain poorly defined. Here, we show that SARS-CoV-2 infection and inactivated vaccine elicited both spike-specific CXCR3 + T FH cell and CXCR3 − T FH cell responses, which showed distinct response patterns. Spike-specific CXCR3 + T FH cells exhibit a dominant and more durable response than CXCR3 − T FH cells that positively correlated with antibody responses. A third booster dose preferentially expands the spike-specific CXCR3 + T FH cell subset induced by two doses of inactivated vaccine, contributing to antibody maturation and potency. Functionally, spike-specific CXCR3 + T FH cells have a greater ability to induce spike-specific antibody secreting cells (ASCs) differentiation compared to spike-specific CXCR3 − T FH cells. In conclusion, the persistent and functional role of spike-specific CXCR3 + T FH cells following SARS-CoV-2 infection and vaccination may play an important role in antibody maintenance and recall response, thereby conferring long-term protection. The findings from this study will inform the development of SARS-CoV-2 vaccines aiming to induce long-term protective immune memory.

目的 了解郴州地区静脉吸毒人群中乙型肝炎病毒(hepatitis B virus,HBV)、丙型肝炎病毒(hepatitis C virus,HCV)和丁型肝炎病毒(hepatitis D virus,HDV)的感染状况及基因型的分布特征;探究共感染患者病毒载量的水平变化以及HBV/HCV共感染中病毒相互作用的影响因素.方法 收集2014～2018年郴州市戒毒所和美沙酮门诊中静脉吸毒志愿者血样,排除艾滋、梅毒等感染性疾病后共纳入样本2 909例.使用酶联免疫吸附实验(enzyme-linked immunosorbent assay,ELISA)检测血清中乙肝表面抗原(hepatitis B surface antigen,HBsAg)、丙型肝炎 IgG抗体(hepatitis C IgG antibody,HCV-IgG)或丁型肝炎 IgG 抗体(hepatitis D IgG antibody,HDV-IgG)的水平.巢式聚合酶链反应(polymerase chain reaction,PCR)扩增感染样本的脱氧核糖核酸(deoxyribo nucleic acid,DNA)或核糖核酸(ribonucleic acid,RNA),测序鉴定其基因型,并通过Real-time PCR对病毒载量进行定量分析.结果 ①HBsAg、HCV-IgG和HDV-IgG 在静脉吸毒人群血清中的阳性率分别为20.49％、34.03％、3.85％.②基因型分析发现HBV基因型以B型(86.74％)为主,C型(11.94％)相对少见;HCV基因6a型为主要感染型(51.33％),其次为3a(20.28％)、3b型(16.98％);HDV基因型均为Ⅱ型.③HCV单感染患者的HCV RNA病毒载量高于HBV/HCV共感染患者(P=0.007);HBV单感染患者的HBV DNA病毒载量高于HBV/HDV共感染患者(P=0.013)和HBV/HCV/HDV三重感染患者(P=0.042);HBV/HDV共感染患者HBsAg水平高于HBV单感染患者(P=0.003)和HBV/HCV共感染患者(P=0.004).④Logistic回归模型分析结果表明HBsAg水平与慢性HCV感染的自发清除显著相关(OR=1.53,95％CI:1.19～1.96),HCV RNA载量可独立并且准确地预测HBV血清清除(OR=1.36,95％CI:1.03～1.80).结论 HBsAg而非HBV DNA是竞争HCV复制的主要因素,高HCV病毒载量可能有益于HBV感染的自发清除.

Aedes albopictus is one of the most invasive insect species in the world and an effective vector for many important arboviruses. We reported previously that Ae . albopictus Nix ( AalNix ) is the male-determining factor of this species. However, whether AalNix alone is sufficient to initiate male development is unknown. Transgenic lines that express each of the three AalNix isoforms from the native promoter were obtained using piggyBac transformation. We verified the stable expression of AalNix isoforms in the transgenic lines and confirm that one isoform, AalNix3&4 , is sufficient to convert females into fertile males (pseudo-males) that are indistinguishable from wild-type males. We also established a stable sex-converted female mosquito strain, AalNix3&4 -♂4-pseudo-male. The pseudo-male mosquitoes can fly and mate normally with wild-type female, although their mating competitiveness is lower than wild-type. This work further clarifies the role of AalNix in the sex determination pathway and will facilitate the development of Ae . albopictus control strategies that rely on male-only releases such as SIT and sex-ratio distortion.

Zika virus (ZIKV) may cause severe microcephaly in newborn babies and Guillain-Barré syndrome in some adults. In recent decades, its range has expanded in 86 countries. There are two ecologically and evolutionarily district cycles: urban cycle and sylvatic cycle. This work aimed to estimate the urban and sylvatic cycle areas of ZIKV throughout the world. The occurrence records of vectors, non-human primate hosts, and ZIKV were collected. We chose historical climate data, predicted vectors distribution, human population density, and elevation data as the variables to fit the maximum entropy model (MaxEnt). Current risk area and future prediction were performed with global climate models (GCMs) and shared socioeconomic pathways (SSPs). Predicting the ZIKV risk area would help tailor related control strategies. The results indicated that 16.6% of the world’s landmass (except Antarctica) is a risk area in the urban cycle. Approximately 6.22 billion people (78.69% of the global population) live in the risk area, with the vast majority in South Asia, tropical Africa, South America, North America, and countries around the Mediterranean Sea. Future climate change decreases the risk area of ZIKV. This study also suggested that the sylvatic cycle happened between the Tropic of Cancer and the Tropic of Capricorn. The overlap region of the urban and sylvatic cycles could be hotpots that ZIKV spill from the sylvatic to the urban cycle. It is indicated that long-term passenger screening, mosquito surveillance, and control are necessary.

Mosquito densoviruses (MDVs) are mosquito-specific viruses that are recommended as mosquito bio-control agents. The MDV Aedes aegypti densovirus (AeDNV) is a good candidate for controlling mosquitoes. However, the slow activity restricts their widespread use for vector control. In this study, we introduced the Bacillus thuringiensis (Bti) toxin Cry11Aa domain II loop α8 and Cyt1Aa loop β6-αE peptides into the AeDNV genome to improve its mosquitocidal efficiency; protein expression was confirmed using nanoscale liquid chromatography coupled to tandem mass spectrometry (nano LC-MS/MS). Recombinant plasmids were transfected into mosquito C6/36 cell lines, and the expression of specific peptides was detected through RT-PCR. A toxicity bioassay against the first instar Aedes albopictus larvae revealed that the pathogenic activity of recombinant AeDNV was significantly higher and faster than the wild-type (wt) viruses, and mortality increased in a dose-dependent manner. The recombinant viruses were genetically stable and displayed growth phenotype and virus proliferation ability, similar to wild-type AeDNV. Our novel results offer further insights by combining two mosquitocidal pathogens to improve viral toxicity for mosquito control.

IntroductionThe Middle East respiratory syndrome coronavirus (MERS-CoV) and the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) are two highly contagious coronaviruses causing MERS and COVID-19, respectively, without an effective antiviral drug and a long-lasting vaccine. Approaches for diagnosis, therapeutics, prevention, etc., particularly for SARS-CoV-2 that is continually spreading and evolving, are urgently needed. Our previous study discovered that >60% of sera from convalescent COVID-19 individuals, but <8% from general population, showed binding activity against the MERS-CoV spike protein, indicating that SARS-CoV-2 infection boosted antibodies cross-reactive with MERS-CoV.MethodsTo generate antibodies specific to both SARS-CoV-2 and MERS-CoV, here we screened 60 COVID-19 convalescent sera against MERS-CoV spike extracellular domain and S1 and S2 subunits. We constructed and characterized monoclonal antibodies (mAbs) from COVID-19 convalescent memory B cells and examined their binding and neutralizing activities against human coronaviruses.Results and DiscussionOf 60 convalescent serum samples, 34 showed binding activity against MERS-CoV S2, with endpoint titers positively correlated with the titers to SARS-CoV-2 S2. By sorting single memory B cells from COVID-19 convalescents, we constructed 38 mAbs and found that 11 mAbs showed binding activity with MERS-CoV S2, of which 9 mAbs showed potent cross-reactivity with all or a proportion of spike proteins of alphacoronaviruses (229E and NL63) and betacoronaviruses (SARS-CoV-1, SARS-CoV-2, OC43, and HKU1). Moreover, 5 mAbs also showed weak neutralization efficiency against MERS-CoV spike pseudovirus. Epitope analysis revealed that 3 and 8 mAbs bound to linear and conformational epitopes in MERS-CoV S2, respectively. In summary, we have constructed a panel of antibodies with broad-spectrum reactivity against all seven human coronaviruses, thus facilitating the development of diagnosis methods and vaccine design for multiple coronaviruses.

Background Zika virus (ZIKV) and dengue virus (DENV) are closely related flaviviruses primarily transmitted by Aedes mosquitoes. Armigeres subalbatus is an emerging and widely distributed mosquito, and ZIKV has been detected and isolated from it. However, it is not clear whether Ar. s ubalbatus could be a vector for ZIKV and DENV or not. In this study, we investigated the infection and transmission of Ar. subalbatus to ZIKV and DENV. Methods A line of Ar. subalbatus was isolated from Guangdong, China, and further identified by the mitochondrial cytochrome oxidase subunit 1 ( COI ) gene. The adults of Ar. subalbatus were fed with blood meal containing ZIKV or DENV-2. At 4, 7, 10, 14, and 21 days post-inoculation (dpi), the infections of ZIKV or DENV-2 in the midguts, ovaries and salivary glands were detected and quantified by RT-PCR and RT-qPCR. To assess the transmissibility, suckling mice were exposed to bites of ZIKV-infected mosquitoes, and ZIKV was detected in brain tissue by RT-qPCR and plaque assays. Furthermore, the larvae of Ar. subalbatus were reared in artificial urine containing ZIKV or DENV-2. The infection rates and viral titers of larvae and adults were analyzed by RT-PCR and RT-qPCR, and the viral distribution in larval tissues was observed by immunohistochemistry. Chi-square test and one-way ANOVA analysis were used for assessing the infection rate and viral titer in varied tissues and different time points, respectively. Results Following oral inoculation, ZIKV but not DENV-2 could be detected in Ar. subalbatus midguts at 4 dpi, ovaries at 7 dpi and salivary glands at 10 dpi. The highest infection rate (IR) of ZIKV was 27.8% in midgut at 7 dpi, 9.7% in ovary and 5.6% in salivary gland at 21 dpi. Eight days after being bitten by ZIKV-positive mosquitoes, ZIKV was detected in three brain tissues out of four suckling mice exposed to bites. ZIKV could be detected in the larvae reared in artificial urine contained ZIKV at a high concentration of 10 5 pfu/ml and various tissues of adults with a low infection rate (0.70–1.35%). ZIKV could be observed in anal papillae and midgut of larvae at 4 dpi under laboratory conditions. Conclusions ZIKV but not DENV-2 can infect Ar. subalbatus by blood meal and artificial urine, and the infected mosquitoes can transmit ZIKV to suckling mice by bite. From these findings, we can conclude that the Ar. subalbatus isolated from Guangdong province, China, is a potential vector for ZIKV and should therefore be considered in vector control programs to prevent and control of Zika virus disease. Graphical Abstract