Dengue virus(DENV)infection is a worldwide public health threat.To date,the knowledge about the pathogenesis and progression of DENV infection is still limited.Combining global profiling based on proteomic analysis to...Dengue virus(DENV)infection is a worldwide public health threat.To date,the knowledge about the pathogenesis and progression of DENV infection is still limited.Combining global profiling based on proteomic analysis together with functional verification analysis is a powerful strategy to investigate the interplay between the virus and host cells.In the present study,quantitative proteomics has been applied to evaluate host responses(as indicated by altered proteins and modifications)in human cells(using K562 cell line)upon DENV-2 infection,as DENV-2 spreads most widely among all DENV serotypes.Comparative analysis was performed to define differentially expressed proteins in the infected cells compared to the mock-control,and it revealed critical pathogen-induced changes covering a broad spectrum of host cellular compartments and processes.We also discovered more dramatic changes(>20%,160 regulated phosphoproteins)in protein phosphorylation compared to protein expression(14%,321 regulated proteins).Most of these proteins/phosphoproteins were involved in transcription regulation,RNA splicing and processing,immune system,cellular response to stimulus,and macromolecule biosynthesis.Western blot analysis was also performed to confirm the proteomic data.Potential roles of these altered proteins were discussed.The present study provides valuable large-scale protein-related information for elucidating the functional emphasis of host cell proteins and their post-translational modifications in virus infection,and also provides insight and protein evidence for understanding the general pathogenesis and pathology of DENV.展开更多
Dengue virus(DENV) and Zika virus(ZIKV) have spread throughout many countries in the developing world and infect millions of people every year, causing severe harm to human health and the economy. Unfortunately, there...Dengue virus(DENV) and Zika virus(ZIKV) have spread throughout many countries in the developing world and infect millions of people every year, causing severe harm to human health and the economy. Unfortunately, there are few effective vaccines and therapies available against these viruses. Therefore, the discovery of new antiviral agents is critical.Herein, a scorpion venom peptide(Smp76) characterized from Scorpio maurus palmatus was successfully expressed and purified in Escherichia coli BL21(DE3). The recombinant Smp76(rSmp76) was found to effectively inhibit DENV and ZIKV infections in a dose-dependent manner in both cultured cell lines and primary mouse macrophages. Interestingly,rSmp76 did not inactivate the viral particles directly but suppressed the established viral infection, similar to the effect of interferon(IFN)-b. Mechanistically, rSmp76 was revealed to upregulate the expression of IFN-b by activating interferon regulatory transcription factor 3(IRF3) phosphorylation, enhancing the type-Ⅰ IFN response and inhibiting viral infection.This mechanism is significantly different from traditional virucidal antimicrobial peptides(AMPs). Overall, the scorpion venom peptide Smp76 is a potential new antiviral agent with a unique mechanism involving type-Ⅰ IFN responses,demonstrating that natural AMPs can enhance immunity by functioning as immunomodulators.展开更多
Dengue fever, caused by dengue viruses(DENVs), is a widespread mosquito-borne zoonotic disease; however, there is no available anti-dengue vaccine for worldwide use. In the current study, a DNA vaccine candidate(pV-D4...Dengue fever, caused by dengue viruses(DENVs), is a widespread mosquito-borne zoonotic disease; however, there is no available anti-dengue vaccine for worldwide use. In the current study, a DNA vaccine candidate(pV-D4 ME) expressing prM-E protein of DENV serotype 4(DENV-4) was constructed, and its immunogenicity and protection were evaluated in immunocompetent BALB/c mice. The pV-D4 ME candidate vaccine induced effective humoral and cellular immunity of mice against DENV-4 in vivo when administered both at 50 μg and 5 μg through electroporation. Two weeks after receiving three immunizations, both doses of pV-D4 ME DNA were shown to confer effective protection against lethal DENV-4 challenge. Notably, at 6 months after the three immunizations, 50 μg, but not 5 μg, of pV-D4 ME could provide stable protection(100% survival rate) against DENV-4 lethal challenge without any obvious clinical signs. These results suggest that immunization with 50 μg pV-D4 ME through electroporation could confer effective and long-term protection against DENV-4, offering a promising approach for development of a novel DNA vaccine against DENVs.展开更多
After dengue virus(DENV)infection,antibody-dependent enhancement(ADE)is easy to occur when the neutralizing antibody(NAb)gradually decreases to a sub-neutralizing concentration.In this cohort surveillance,we utilized ...After dengue virus(DENV)infection,antibody-dependent enhancement(ADE)is easy to occur when the neutralizing antibody(NAb)gradually decreases to a sub-neutralizing concentration.In this cohort surveillance,we utilized sera samples collected from dengue fever patients at different convalescent phases in Jinghong City,to investigate the dynamic change rule of DENV-specific antibodies,and to analyze the risk of ADE caused by secondary infection with heterologous serotypes DENVs.For baseline serosurvey,191 four-year and 99 six-year sera samples during convalescence were collected in 2017 and 2019,respectively.The positive rate of DENVspecific immunoglobulin G was 98.4%in 2017,which significantly decreased to 82.8%in 2019.The geometric mean titer(GMT)of NAb decreased from 1:155.35 to 1:46.66.Among 290 overall samples,73 paired consecutive samples were used for follow-up serosurvey.In four-year sera,the GMTs of NAb against DENV-3 and cross-reactive antibodies against DENV-1,DENV-2 and DENV-4 were 1:167.70,1:13.80,1:18.54 and 1:45.26,respectively,which decreased to 1:53.18,1:10.30,1:14.60 and 1:8.17 in six-year sera.In age-stratified analysis,due to the increasing number of ADE positive samples from 2017 to 2019 in 31–40 and 51–60 years groups,the risk of ADE in DENV-4 infection was positively associated with the extension of convalescent phase,and the odd ratio was higher than other groups.With the recovery period lengthened,the risk of secondary infection with DENV-1 and DENV-2 was reduced.Our results offer essential experimental data for risk prediction of severe dengue in hyper-endemic dengue areas,and provide crucial scientific insight for the development of effective dengue vaccines.展开更多
The Pearl River Delta,where Aedes albopictus(Ae.albopictus)is the only vector for dengue transmission,has exhibited one of the highest dengue burdens in southern China in recent decades.However,whether dengue virus(DE...The Pearl River Delta,where Aedes albopictus(Ae.albopictus)is the only vector for dengue transmission,has exhibited one of the highest dengue burdens in southern China in recent decades.However,whether dengue virus(DENV)can overwinter in Ae.albopictus in the Pearl River Delta has not been determined to date.In this study,300 field-derived Ae.albopictus mosquitoes from Guangzhou that were infected with the predominant endemic DENV-1 strain were investigated under simulated urban balcony environment from October 16,2016,to June 16,2017.The vertical transmission of DENV in the infected overwintering Ae.albopictus was analyzed.The DENV infected overwintering mosquitoes were evaluated for viral load at nine-time points using reverse transcription-quantitative PCR.The vector competence of the infected overwintering Ae.albopictus was also investigated by using suckling mice.Adult mosquitoes and larvae were found during the observation period.The vertical transmission of DENV-1 was documented.The DENV-1-positive rates between overwintering males and females had no difference.The proportion of DENV-1-positive overwintering mosquitoes decreased over time and had no difference beyond three months after the experiment.Overwintering mosquitoes can spread DENV-1 to hosts.No engorged mosquitoes at an ambient temperature below 15℃were observed.The ratio of engorged mosquitoes was positively correlated with the ambient temperature ranging from 15 to 30℃.Our results demonstrated that DENV can overwinter in Ae.albopictus in the Pearl River Delta,Ae.albopictus is the competent vector for DENV,and maintain autochthonous dengue outbreaks in the Pearl River Delta through vertical transmission.展开更多
Dengue virus(DENV)is one of the most important arboviral pathogens in the tropics and subtropics,and nearly one-third of the world's population is at risk of infection.The transmission of DENV involves a sylvatic ...Dengue virus(DENV)is one of the most important arboviral pathogens in the tropics and subtropics,and nearly one-third of the world's population is at risk of infection.The transmission of DENV involves a sylvatic cycle between nonhuman primates(NHP)and Aedes genus mosquitoes,and an endemic cycle between human hosts and predominantly Aedes aegypti.DENV belongs to the genus Flavivirus of the family Flaviviridae and consists of four antigenically distinct serotypes(DENV-1-4).Phylogenetic analyses of DENV have revealed its origin,epidemiology,and the drivers that determine its molecular evolution in nature.This review discusses how phyloge-netic research has improved our understanding of DENV evolution and how it affects viral ecology and improved our ability to analyze and predict future DENV emergence.展开更多
基金supported by the National Natural Science Foundation of China (31870827 to X.Zhao, 31670161 to X.Zhou., and 81873964 to Y.Q.)the Hubei Natural Science Foundation (2018CFB603 to X.Zhao)the Fundamental Research Funds for the Central Universities (2042018kf0247 to X.Zhao)
文摘Dengue virus(DENV)infection is a worldwide public health threat.To date,the knowledge about the pathogenesis and progression of DENV infection is still limited.Combining global profiling based on proteomic analysis together with functional verification analysis is a powerful strategy to investigate the interplay between the virus and host cells.In the present study,quantitative proteomics has been applied to evaluate host responses(as indicated by altered proteins and modifications)in human cells(using K562 cell line)upon DENV-2 infection,as DENV-2 spreads most widely among all DENV serotypes.Comparative analysis was performed to define differentially expressed proteins in the infected cells compared to the mock-control,and it revealed critical pathogen-induced changes covering a broad spectrum of host cellular compartments and processes.We also discovered more dramatic changes(>20%,160 regulated phosphoproteins)in protein phosphorylation compared to protein expression(14%,321 regulated proteins).Most of these proteins/phosphoproteins were involved in transcription regulation,RNA splicing and processing,immune system,cellular response to stimulus,and macromolecule biosynthesis.Western blot analysis was also performed to confirm the proteomic data.Potential roles of these altered proteins were discussed.The present study provides valuable large-scale protein-related information for elucidating the functional emphasis of host cell proteins and their post-translational modifications in virus infection,and also provides insight and protein evidence for understanding the general pathogenesis and pathology of DENV.
基金supported by grants from National Science Fund of China (Nos. 31572289, 31872239 and 81630091)International S&T Cooperation Program of China (No. S2016G3110)+3 种基金Hubei Science Fund (Nos. 2015CFA042 and 2016CFA018)China-Kazakhstan Cooperation Program (No. CK-07-09)Fundamental Research Funds for the Central Universities in China (Nos. 2042017kf0242 and 2042017kf0199)financial support from Higher Education Commission (HEC) of Pakistan
文摘Dengue virus(DENV) and Zika virus(ZIKV) have spread throughout many countries in the developing world and infect millions of people every year, causing severe harm to human health and the economy. Unfortunately, there are few effective vaccines and therapies available against these viruses. Therefore, the discovery of new antiviral agents is critical.Herein, a scorpion venom peptide(Smp76) characterized from Scorpio maurus palmatus was successfully expressed and purified in Escherichia coli BL21(DE3). The recombinant Smp76(rSmp76) was found to effectively inhibit DENV and ZIKV infections in a dose-dependent manner in both cultured cell lines and primary mouse macrophages. Interestingly,rSmp76 did not inactivate the viral particles directly but suppressed the established viral infection, similar to the effect of interferon(IFN)-b. Mechanistically, rSmp76 was revealed to upregulate the expression of IFN-b by activating interferon regulatory transcription factor 3(IRF3) phosphorylation, enhancing the type-Ⅰ IFN response and inhibiting viral infection.This mechanism is significantly different from traditional virucidal antimicrobial peptides(AMPs). Overall, the scorpion venom peptide Smp76 is a potential new antiviral agent with a unique mechanism involving type-Ⅰ IFN responses,demonstrating that natural AMPs can enhance immunity by functioning as immunomodulators.
基金supported by the National Natural Science Foundation of China (81772172, 81471957, 81671971, U1602223)
文摘Dengue fever, caused by dengue viruses(DENVs), is a widespread mosquito-borne zoonotic disease; however, there is no available anti-dengue vaccine for worldwide use. In the current study, a DNA vaccine candidate(pV-D4 ME) expressing prM-E protein of DENV serotype 4(DENV-4) was constructed, and its immunogenicity and protection were evaluated in immunocompetent BALB/c mice. The pV-D4 ME candidate vaccine induced effective humoral and cellular immunity of mice against DENV-4 in vivo when administered both at 50 μg and 5 μg through electroporation. Two weeks after receiving three immunizations, both doses of pV-D4 ME DNA were shown to confer effective protection against lethal DENV-4 challenge. Notably, at 6 months after the three immunizations, 50 μg, but not 5 μg, of pV-D4 ME could provide stable protection(100% survival rate) against DENV-4 lethal challenge without any obvious clinical signs. These results suggest that immunization with 50 μg pV-D4 ME through electroporation could confer effective and long-term protection against DENV-4, offering a promising approach for development of a novel DNA vaccine against DENVs.
文摘目的在急性单核细胞白血病细胞(THP-1)水平建立Ⅲ型登革病毒(Dengue virus typeⅢ,DENV-3,DV-3)感染和抗体依赖性增强(antibody dependent enhancement,ADE)感染模型,探讨胞内长链非编码RNA(long non-coding RNA,LncRNA)的差异性表达,绘制竞争性内源RNA(competitive endogenous RNA,CeRNA)调控网络并进行LncRNA翻译功能预测。方法DENV-3感染C6/36细胞6 d后,收获培养上清,采用CCID50法测定病毒滴度,并通过PCR进行型别以及基因组全长扩增鉴定;扩增DENV-3标准质粒,PCR鉴定,绘制标准曲线;将THP-1细胞分为阴性对照(THP-1)、直接感染(DV-3)、ADE及空白对照[1640(-)]组,感染48 h后提取胞内总RNA,测定病毒拷贝数;通过全转录组测序技术,对THP-1 vs DENV-3、THP-1 vs ADE、DENV-3 vs ADE各组中上调和下调前5个的LncRNA进行CeRNA调控网络构建,并分析其编码蛋白的功能。结果DENV-3感染C6/36细胞3 d后有明显的细胞融合、空泡和脱落;病毒滴度约为1.0×10^(4.64)PFU/mL,PCR特异引物鉴定为DENV-3,获得病毒完整的基因序列;ADE组胞内病毒核酸拷贝数明显高于DV-3组和空白对照组;在THP-1 vs DENV-3中,预测到人细胞黏附蛋白相互作用蛋白(cytohesin interacting protein,CYTIP)的表达量出现上调;在THP-1 vs ADE中,预测到驱动蛋白家族成员5A(kinesin family 5A,KIF5A)的表达量下调;在DENV-3 vs ADE中,预测到簇分化抗原9(cluster differentiation antigen 9,CD9)和胰岛素样生长因子2(insulin like growth factor 2,IGF2)的表达量上调。这些差异性的LncRNA均具有开放阅读框(open reading frame,ORF),除了Lnc-SH3BP1和Lnc-RPL41以外,其余的LncRNA均具有内部核糖体结合位点(internal ribosome binding site,IRES)。结论在DENV-3感染THP-1细胞及其介导的ADE感染中,LncRNA的表达发生了明显的差异性改变,且可能通过多种生物学功能调控感染的进程,有助于更深层次理解ADE感染的发生机制。
基金supported by the National Natural Science Foundation of China under grants 81772172,U1902210,81972979 and 81902048
文摘After dengue virus(DENV)infection,antibody-dependent enhancement(ADE)is easy to occur when the neutralizing antibody(NAb)gradually decreases to a sub-neutralizing concentration.In this cohort surveillance,we utilized sera samples collected from dengue fever patients at different convalescent phases in Jinghong City,to investigate the dynamic change rule of DENV-specific antibodies,and to analyze the risk of ADE caused by secondary infection with heterologous serotypes DENVs.For baseline serosurvey,191 four-year and 99 six-year sera samples during convalescence were collected in 2017 and 2019,respectively.The positive rate of DENVspecific immunoglobulin G was 98.4%in 2017,which significantly decreased to 82.8%in 2019.The geometric mean titer(GMT)of NAb decreased from 1:155.35 to 1:46.66.Among 290 overall samples,73 paired consecutive samples were used for follow-up serosurvey.In four-year sera,the GMTs of NAb against DENV-3 and cross-reactive antibodies against DENV-1,DENV-2 and DENV-4 were 1:167.70,1:13.80,1:18.54 and 1:45.26,respectively,which decreased to 1:53.18,1:10.30,1:14.60 and 1:8.17 in six-year sera.In age-stratified analysis,due to the increasing number of ADE positive samples from 2017 to 2019 in 31–40 and 51–60 years groups,the risk of ADE in DENV-4 infection was positively associated with the extension of convalescent phase,and the odd ratio was higher than other groups.With the recovery period lengthened,the risk of secondary infection with DENV-1 and DENV-2 was reduced.Our results offer essential experimental data for risk prediction of severe dengue in hyper-endemic dengue areas,and provide crucial scientific insight for the development of effective dengue vaccines.
基金This project was supported by the Provincial Science and Technology Projects of Guangdong Province,China(Grant numbers:2014A020219007,2015A020218003,2016A020219006 and 2017A020215101)the Medical Scientific Research Foundation of Guangdong Province,China(Grant numbers:A2018167).
文摘The Pearl River Delta,where Aedes albopictus(Ae.albopictus)is the only vector for dengue transmission,has exhibited one of the highest dengue burdens in southern China in recent decades.However,whether dengue virus(DENV)can overwinter in Ae.albopictus in the Pearl River Delta has not been determined to date.In this study,300 field-derived Ae.albopictus mosquitoes from Guangzhou that were infected with the predominant endemic DENV-1 strain were investigated under simulated urban balcony environment from October 16,2016,to June 16,2017.The vertical transmission of DENV in the infected overwintering Ae.albopictus was analyzed.The DENV infected overwintering mosquitoes were evaluated for viral load at nine-time points using reverse transcription-quantitative PCR.The vector competence of the infected overwintering Ae.albopictus was also investigated by using suckling mice.Adult mosquitoes and larvae were found during the observation period.The vertical transmission of DENV-1 was documented.The DENV-1-positive rates between overwintering males and females had no difference.The proportion of DENV-1-positive overwintering mosquitoes decreased over time and had no difference beyond three months after the experiment.Overwintering mosquitoes can spread DENV-1 to hosts.No engorged mosquitoes at an ambient temperature below 15℃were observed.The ratio of engorged mosquitoes was positively correlated with the ambient temperature ranging from 15 to 30℃.Our results demonstrated that DENV can overwinter in Ae.albopictus in the Pearl River Delta,Ae.albopictus is the competent vector for DENV,and maintain autochthonous dengue outbreaks in the Pearl River Delta through vertical transmission.
基金National Key Research and Development Plan of China,Grant/Award Number:2018YFA0507202,2020YFC1200100 and 2021YFC2300200National Natural Science Foundation of China,Grant/Award Number:31825001,32188101,81730063,81961160737 and 82102389+5 种基金Provincial Innovation Team for the Prevention and Control of Highly Pathogenic Pathogens,Institute of Medical Biology,Chinese Academy of Medical Sciences,Grant/Award Number:202105AE160020Shenzhen San-Ming Project for Prevention and Research on Vector-borne Diseases,Grant/Award Number:SZSM201611064Shenzhen Science and Technology Project,Grant/Award Number:JSGG20191129144225464Tsinghua University Spring Breeze Fund,Grant/Award Number:2020Z99CFG017Young Elite Scientists Sponsorship Program,Grant/Award Number:2021QNRC001Yunnan Cheng Gong Expert Work-Station,Grant/Award Number:202005AF150034。
文摘Dengue virus(DENV)is one of the most important arboviral pathogens in the tropics and subtropics,and nearly one-third of the world's population is at risk of infection.The transmission of DENV involves a sylvatic cycle between nonhuman primates(NHP)and Aedes genus mosquitoes,and an endemic cycle between human hosts and predominantly Aedes aegypti.DENV belongs to the genus Flavivirus of the family Flaviviridae and consists of four antigenically distinct serotypes(DENV-1-4).Phylogenetic analyses of DENV have revealed its origin,epidemiology,and the drivers that determine its molecular evolution in nature.This review discusses how phyloge-netic research has improved our understanding of DENV evolution and how it affects viral ecology and improved our ability to analyze and predict future DENV emergence.
