Coxsackievirus A16(CA16) is one of the major causes of hand, foot, and mouth disease(HFMD) worldwide, which is a common illness that affects children. The frequent occurrence of HFMD outbreaks has become a serious pub...Coxsackievirus A16(CA16) is one of the major causes of hand, foot, and mouth disease(HFMD) worldwide, which is a common illness that affects children. The frequent occurrence of HFMD outbreaks has become a serious public health problem in Asia. Therefore, it is important to understand the pathogenesis and replication of CA16. In this study, a stable infectious c DNA clone of an epidemic strain of Coxsackievirus A16(CA16) was assembled, and subsequently a reporter virus(e GFP-CA16) was constructed by inserting the e GFP gene between the 5'-UTR and the N-terminus of VP4, with the addition of a 2A protease cleavage site(ITTLG) at its C-terminus. This was transfected into Vero cells to generate infectious recombinant viruses. The growth characteristics and plaque morphology, in vitro, in mammalian cells were found to be indistinguishable between the parental and recombinant viruses. Although the e GFP-CA16 showed smaller plaque size as compared to recombinant CA16, both were found to exhibit similar growth trends and EC50 of NITD008. In summary, this stable infectious c DNA clone should provide a valuable experimental system to study CA16 infection and host response. The e GFP-CA16 is expected to provide a powerful tool to monitor e GFP expression in infected cells and to evaluate the antiviral activity of potential antiviral agents in the treatment of CA16 infections.展开更多
Omsk hemorrhagic fever virus(OHFV) is a tick-borne flavivirus classified as a biosafety level-4(BSL4) pathogen. Studies of OHFV are restricted to be conducted within BSL4 laboratories. Currently, no commercial vaccine...Omsk hemorrhagic fever virus(OHFV) is a tick-borne flavivirus classified as a biosafety level-4(BSL4) pathogen. Studies of OHFV are restricted to be conducted within BSL4 laboratories. Currently, no commercial vaccines or antiviral drugs are available against OHFV infection. In this study, we recovered a replication-deficient OHFV with an NS1 deletion(OHFVDNS1) and reporter virus replacing NS1 with the Gaussia luciferase(Gluc)(OHFV-ΔNS1-Gluc). Both the defective OHFVDNS1 and OHFV-ΔNS1-Gluc virus could only replicate efficiently in the BHK21 cell line expressing NS1(BHK21NS1) but not in na?ve BHK21 cells. The Gluc reporter gene of OHFV-ΔNS1-Gluc virus was maintained stably after serial passaging of BHK21NS1 cells and was used to surrogate the replication of OHFV. Using NITD008, OHFV-ΔNS1-Gluc virus was validated for antiviral screening, and high-throughput screening parameters were optimized in a 96-well plate format with a calculated Z0 value above 0.5. The OHFV-ΔNS1-Gluc reporter virus is a powerful tool for antiviral screening as well as viral replication and pathogenesis studies in BSL2 laboratories.展开更多
COVID-19 has become a global public health crisis since its outbreak in China in December 2019.Currently there are few clinically effective drugs to combat SARS-CoV-2 infection.The main protein(M^(pro)),papain-like pr...COVID-19 has become a global public health crisis since its outbreak in China in December 2019.Currently there are few clinically effective drugs to combat SARS-CoV-2 infection.The main protein(M^(pro)),papain-like protease(PL^(pro))and RNA-dependent RNA polymerase(RdRp)of SARS-CoV-2 are involved in the viral replication,and might be prospective targets for anti-coronavirus drug development.Here,we investigated the antiviral activity of oridonin,a natural small-molecule compound,against SARS-CoV-2 infection in vitro.The time-of-addition analysis showed that oridonin efficiently inhibited SARS-CoV-2 infection by interfering with the genome replication at the post-entry stage.Mechanistically,the inhibition of viral replication by oridonin depends on the oxidation activity ofα,β-unsaturated carbonyl.Further experiments showed that oridonin not only effectively inhibited SARS-CoV-2 Mpro activity,but also had some inhibitory effects on PLpro-mediated deubiquitinating and viral polymerase-catalyzed RNA elongation activities at high concentrations.In particular,oridonin could inhibit the bat SARS-like CoV and the newly emerged SARS-CoV-2 omicron variants(BA.1 and BA.2),which highlights its potential as a pan-coronavirus antiviral agent.Overall,our data provide strong evidence that oridonin is an efficient antiviral agent against SARS-CoV-2 infection.展开更多
We developed a Gaussia luciferase (Gluc) reporter replicon of West Nile virus (WNV) and used it to quantify viral translation and RNA replication. The advantage of the Gluc replicon is that Gaussia luciferase is secre...We developed a Gaussia luciferase (Gluc) reporter replicon of West Nile virus (WNV) and used it to quantify viral translation and RNA replication. The advantage of the Gluc replicon is that Gaussia luciferase is secreted into the culture medium from cells transfected with Gluc replicon RNA, and the medium can be assayed directly for luciferase activity. Using a known Flavivirus inhibitor (NITD008), we demonstrated that the Gluc-WNV replicon could be used for antiviral screening. The Gluc-WNV-Rep will be useful for research in antiviral drug development programs, as well as for studying viral replication and pathogenesis of WNV.展开更多
No samples from the Moon’s far side have been returned to Earth.Thus,lunar meteorite study can deepen the understanding of the Moon’s far side(if we can identify which lunar meteorites came from the Moon’s far side...No samples from the Moon’s far side have been returned to Earth.Thus,lunar meteorite study can deepen the understanding of the Moon’s far side(if we can identify which lunar meteorites came from the Moon’s far side).The Northwest Africa(NWA)15528 meteorite is a lunar feldspathic regolith breccia meteorite and we speculated it might originate from the Feldspathic Highlands Terrane(FHT)on the far side of the Moon.Here,we report detailed petrography,major and trace elements,and noble gas(He,Ne,and Ar)isotopes for the clasts and matrix,respectively.The results show that the NWA 15528 lunar meteorite contains diverse clasts,including anorthosite,granulite,basalt,and impact melt clasts.The coarse,wellcrystallized,uniform chemical composition minerals may come from intrusive plutonic rocks.Among the anorthosite clasts,the norite/olivine clasts originate from the deep lunar crust,whereas the other anorthosite clasts are from lunar highlands.The Sm concentrations in NWA 15528 were similar to those in the fourth group of Apollo 16 melt samples,demonstrating that NWA 15528 has a typical plagioclase highland meteorite composition.Compared with the Apollo sample data and remote sensing results,the chemical composition of NWA 15528 indicated strong affinities with the FHT area and ferroan anorthosite(FAN)material from the far side of the Moon.The noble gas isotopic composition of NWA 15528 is consistent with a binary mixture of solar wind and cosmogenic components;during stepwise pyrolytic extractions,we observed that the abundance of cosmogenic components decreased,whereas that of solar wind components increased with increasing temperature.The average cosmic-ray exposure(CRE)age of the matrix and granulite is 42±6 Ma,with a shielding depth in the same range of 10-20 g cm^(−2).The gas retention age of NWA 15528 is 2.14 Ga,and the antiquity age of NWA 15528 is(0.69-0.74)±0.2 Ga(considering 50%40Arm is 40Artrap)which indicates the different clasts of NWA 15528 are assembled after 0.69-0.74 Ga.展开更多
基金supported by the Science and Technology Plan Projects of Wuhan (grant No. 2013060501010157)
文摘Coxsackievirus A16(CA16) is one of the major causes of hand, foot, and mouth disease(HFMD) worldwide, which is a common illness that affects children. The frequent occurrence of HFMD outbreaks has become a serious public health problem in Asia. Therefore, it is important to understand the pathogenesis and replication of CA16. In this study, a stable infectious c DNA clone of an epidemic strain of Coxsackievirus A16(CA16) was assembled, and subsequently a reporter virus(e GFP-CA16) was constructed by inserting the e GFP gene between the 5'-UTR and the N-terminus of VP4, with the addition of a 2A protease cleavage site(ITTLG) at its C-terminus. This was transfected into Vero cells to generate infectious recombinant viruses. The growth characteristics and plaque morphology, in vitro, in mammalian cells were found to be indistinguishable between the parental and recombinant viruses. Although the e GFP-CA16 showed smaller plaque size as compared to recombinant CA16, both were found to exhibit similar growth trends and EC50 of NITD008. In summary, this stable infectious c DNA clone should provide a valuable experimental system to study CA16 infection and host response. The e GFP-CA16 is expected to provide a powerful tool to monitor e GFP expression in infected cells and to evaluate the antiviral activity of potential antiviral agents in the treatment of CA16 infections.
基金supported by National Science and Technology Major Project on Important Infectious Diseases Prevention and Control (2018ZX10734404-010)National Key Research and Development Program of China (2018YFA0507201)
文摘Omsk hemorrhagic fever virus(OHFV) is a tick-borne flavivirus classified as a biosafety level-4(BSL4) pathogen. Studies of OHFV are restricted to be conducted within BSL4 laboratories. Currently, no commercial vaccines or antiviral drugs are available against OHFV infection. In this study, we recovered a replication-deficient OHFV with an NS1 deletion(OHFVDNS1) and reporter virus replacing NS1 with the Gaussia luciferase(Gluc)(OHFV-ΔNS1-Gluc). Both the defective OHFVDNS1 and OHFV-ΔNS1-Gluc virus could only replicate efficiently in the BHK21 cell line expressing NS1(BHK21NS1) but not in na?ve BHK21 cells. The Gluc reporter gene of OHFV-ΔNS1-Gluc virus was maintained stably after serial passaging of BHK21NS1 cells and was used to surrogate the replication of OHFV. Using NITD008, OHFV-ΔNS1-Gluc virus was validated for antiviral screening, and high-throughput screening parameters were optimized in a 96-well plate format with a calculated Z0 value above 0.5. The OHFV-ΔNS1-Gluc reporter virus is a powerful tool for antiviral screening as well as viral replication and pathogenesis studies in BSL2 laboratories.
基金the Creative Research Group Program of Natural Science Foundation of Hubei Province(2022CFA021)the China Postdoctoral Science Foundation(2022M720895)+1 种基金the National Natural Science Foundation of China(3210011 and 32200132)the Natural Science Foundation of Hubei Province,China(2017CFB535).
文摘COVID-19 has become a global public health crisis since its outbreak in China in December 2019.Currently there are few clinically effective drugs to combat SARS-CoV-2 infection.The main protein(M^(pro)),papain-like protease(PL^(pro))and RNA-dependent RNA polymerase(RdRp)of SARS-CoV-2 are involved in the viral replication,and might be prospective targets for anti-coronavirus drug development.Here,we investigated the antiviral activity of oridonin,a natural small-molecule compound,against SARS-CoV-2 infection in vitro.The time-of-addition analysis showed that oridonin efficiently inhibited SARS-CoV-2 infection by interfering with the genome replication at the post-entry stage.Mechanistically,the inhibition of viral replication by oridonin depends on the oxidation activity ofα,β-unsaturated carbonyl.Further experiments showed that oridonin not only effectively inhibited SARS-CoV-2 Mpro activity,but also had some inhibitory effects on PLpro-mediated deubiquitinating and viral polymerase-catalyzed RNA elongation activities at high concentrations.In particular,oridonin could inhibit the bat SARS-like CoV and the newly emerged SARS-CoV-2 omicron variants(BA.1 and BA.2),which highlights its potential as a pan-coronavirus antiviral agent.Overall,our data provide strong evidence that oridonin is an efficient antiviral agent against SARS-CoV-2 infection.
基金supported by the National Natural Science Foundation of China (grant No. 31170158 and 31000090)the '100 Talents Project' of Chinese Academy of Sciences, China (grant No. Y002171YC1)
文摘We developed a Gaussia luciferase (Gluc) reporter replicon of West Nile virus (WNV) and used it to quantify viral translation and RNA replication. The advantage of the Gluc replicon is that Gaussia luciferase is secreted into the culture medium from cells transfected with Gluc replicon RNA, and the medium can be assayed directly for luciferase activity. Using a known Flavivirus inhibitor (NITD008), we demonstrated that the Gluc-WNV replicon could be used for antiviral screening. The Gluc-WNV-Rep will be useful for research in antiviral drug development programs, as well as for studying viral replication and pathogenesis of WNV.
基金supported by the Strategic Priority Program B of the Chinese Academy of Sciences(Grant No.XDB41010205)the Civil Aerospace Pre-Research Project(Grant No.D020302)+1 种基金the Strategic Priority Program A of the Chinese Academy of Sciences(Grant Nos.XDA17010403 and XDB41010304)the National Natural Science Foundation of China(Grant Nos.42030205 and 41874079).
文摘No samples from the Moon’s far side have been returned to Earth.Thus,lunar meteorite study can deepen the understanding of the Moon’s far side(if we can identify which lunar meteorites came from the Moon’s far side).The Northwest Africa(NWA)15528 meteorite is a lunar feldspathic regolith breccia meteorite and we speculated it might originate from the Feldspathic Highlands Terrane(FHT)on the far side of the Moon.Here,we report detailed petrography,major and trace elements,and noble gas(He,Ne,and Ar)isotopes for the clasts and matrix,respectively.The results show that the NWA 15528 lunar meteorite contains diverse clasts,including anorthosite,granulite,basalt,and impact melt clasts.The coarse,wellcrystallized,uniform chemical composition minerals may come from intrusive plutonic rocks.Among the anorthosite clasts,the norite/olivine clasts originate from the deep lunar crust,whereas the other anorthosite clasts are from lunar highlands.The Sm concentrations in NWA 15528 were similar to those in the fourth group of Apollo 16 melt samples,demonstrating that NWA 15528 has a typical plagioclase highland meteorite composition.Compared with the Apollo sample data and remote sensing results,the chemical composition of NWA 15528 indicated strong affinities with the FHT area and ferroan anorthosite(FAN)material from the far side of the Moon.The noble gas isotopic composition of NWA 15528 is consistent with a binary mixture of solar wind and cosmogenic components;during stepwise pyrolytic extractions,we observed that the abundance of cosmogenic components decreased,whereas that of solar wind components increased with increasing temperature.The average cosmic-ray exposure(CRE)age of the matrix and granulite is 42±6 Ma,with a shielding depth in the same range of 10-20 g cm^(−2).The gas retention age of NWA 15528 is 2.14 Ga,and the antiquity age of NWA 15528 is(0.69-0.74)±0.2 Ga(considering 50%40Arm is 40Artrap)which indicates the different clasts of NWA 15528 are assembled after 0.69-0.74 Ga.
