Background: In recent times, there has been an increase in the number of Lassa fever cases resulting from the several episodes of Lassa fever epidemics ravaging Nigeria and other West African countries. The presence o...Background: In recent times, there has been an increase in the number of Lassa fever cases resulting from the several episodes of Lassa fever epidemics ravaging Nigeria and other West African countries. The presence of Lassa virus in rodents other than the major reservoir (Mastomys natalensis) has been a public health concern as to the actual burden of the disease. It is therefore of a public health necessity to explore the LASV RNA habouring potential of several species of rodents in endemic as well as non-endemic areas for proper prevention of emergence of outbreaks in non-endemic areas. Objectives: The aim of this study was to detect the presence of LASV RNA in different species of rodents in Ikorodu, Lagos state and Abeokuta, Ogun state. Methods: A total of ninety one (91) rodents were captured from Ikorodu, Lagos State (61 rodents) and Abeokuta, Ogun State (30 rodents), euthanized, bled, and plasma obtained for the detection of LASV RNA by Reverse Transcriptase Polymerase Chain Reaction. Results: A total of 91 rodents consisting of 77 Rattus rattus and 14 Crocidura spp. The S segment of LASV RNA was not in any of the 91 rodents’ plasma samples. Conclusion: The rodents captured within the rural communities of Ikorodu, Lagos State and Abeokuta, Ogun State were found not to habour the LASV RNA. This study is limited by the relatively small sample size. Similar studies should be encouraged both in endemic and non-endemic areas in order to understand the actual burden of Lassa fever as well as put into check future epidemics.展开更多
Lassa virus(LASV)is an enveloped,negative-sense RNA virus that causes Lassa hemorrhagic fever.Successful entry of LASV requires the viral glycoprotein 1(GP1)to undergo a receptor switch from its primary receptor alpha...Lassa virus(LASV)is an enveloped,negative-sense RNA virus that causes Lassa hemorrhagic fever.Successful entry of LASV requires the viral glycoprotein 1(GP1)to undergo a receptor switch from its primary receptor alpha-dystroglycan(α-DG)to its endosomal receptor lysosome-associated membrane protein 1(LAMP1).A conserved histidine triad in LASV GP1 has been reported to be responsible for receptor switch.To test the hypothesis that other non-conserved residues also contribute to receptor switch,we constructed a series of mutant LASV GP1 proteins and tested them for binding to LAMP1.Four residues,L84,K88,L107,and H170,were identified as critical for receptor switch.Substituting any of the four residues with the corresponding lymphocytic choriomeningitis virus(LCMV)residue(L84 N,K88E,L10F,and H170S)reduced the binding affinity of LASV GP1 for LAMP1.Moreover,all mutations caused decreases in glycoprotein precursor(GPC)-mediated membrane fusion at both pH 4.5 and 5.2.The infectivity of pseudotyped viruses bearing either GPCL84N or GPCK88E decreased sharply in multiple cell types,while L107F and H170S had only mild effects on infectivity.Using biolayer light interferometry assay,we found that all four mutants had decreased binding affinity to LAMP1,in the order of binding affinity being L84 N>L107F>K88E>H170S.The four amino acid loci identified for the first time in this study have important reference significance for the in-depth investigation of the mechanism of receptor switching and immune escape of LASV occurrence and the development of reserve anti-LASV infection drugs.展开更多
The Lassa virus(LASV)is endemic in West Africa and causes severe hemorrhagic Lassa fever in humans.The glycoprotein complex(GPC)of LASV is highly glycosylation-modified,with 11 N-glycosylation sites.All 11 N-linked gl...The Lassa virus(LASV)is endemic in West Africa and causes severe hemorrhagic Lassa fever in humans.The glycoprotein complex(GPC)of LASV is highly glycosylation-modified,with 11 N-glycosylation sites.All 11 N-linked glycan chains play critical roles in GPC cleavage,folding,receptor binding,membrane fusion,and immune evasion.In this study,we focused on the first glycosylation site because its deletion mutant(N79Q)results in an unexpected enhanced membrane fusion,whereas it exerts little effect on GPC expression,cleavage,and receptor binding.Meanwhile,the pseudotype virus bearing GPC_(N79Q)was more sensitive to the neutralizing antibody 37.7H and was attenuated in virulence.Exploring the biological functions of the key glycosylation site on LASV GPC will help elucidate the mechanism of LASV infection and provide strategies for the development of attenuated vaccines against LASV infection.展开更多
<strong>Introduction: </strong>Lassa fever is a disease of public health importance because of the associated morbidity and high case fatality rate among hospitalized patients. Even after recovery, there m...<strong>Introduction: </strong>Lassa fever is a disease of public health importance because of the associated morbidity and high case fatality rate among hospitalized patients. Even after recovery, there may be residual problems such as sensorineural hearing loss. The initial presentation of Lassa fever may be with non-specific symptoms similar to what is seen in the more common febrile illnesses such as malaria or typhoid fever. In such a setting therefore, timely diagnosis of Lassa fever may be difficult. <strong>Case Report: </strong>We report a case of Lassa fever that presented to our institution. She was a middle aged woman who had non-specific symptoms of febrile illness and who died in less than 48 hours of admission. She had a subtle bleeding on the lip just before death which was what raised the suspicion for Lassa fever. Laboratory confirmation of Lassa fever was made retrospectively.<strong> Conclusion: </strong>When the presenting symptoms are non-specific, a high index of suspicion is required for timely recognition of Lassa fever. Early diagnosis is important for prompt therapeutic intervention as well as for limiting the spread of the disease. This is the second case of Lassa fever presenting to our hospital, but the first published case of Lassa fever from our hospital.展开更多
The limited treatment options for the increasing occurrence of Lassa hemorrhagic fever in West Africa poses an urgent need for the discovery and development of novel therapeutics.Dietary supplements,especially natural...The limited treatment options for the increasing occurrence of Lassa hemorrhagic fever in West Africa poses an urgent need for the discovery and development of novel therapeutics.Dietary supplements,especially natural products that are edible and safe for human use,are a good source of drug discovery with potential for uncovering novel applications,In this study,we tested 40 natural products of dietary supplements and identified capsaicin,a common dietary supplement abundant in chili peppers,as an inhibitor of Lassa virus(LASV)entry with EC5,of 6.9-10.0μmol/L using an HIV based pseudo virus platform.Capsaicin inhibits the entry of five LASV strains but not against the Old World arenavirus lymphocytic choriomeningitis virus(LCMV),showing a preferential activity against LASV.Capsaicin inhibits LASV entry by blocking the pH dependent viral fusion through affecting the stable signal peptide(SSP)-GP2 transmembrane(GP2TM)region of the LASV surface glycoprotein.Mutational study revealed the key residues Ala25,Val431,Phe434 and Val435 in SsP-CP2TM region in capsaicin’s antiviral effect.This study for the first time reveals a direct acting antiviral effect of capsaicin against the hemorrhagic fever causing LASV,providing detailed interaction hot spots in the unique SSP-GP2TMinterface of LASV glycoprotein that is crucial in fusion inhibition,and offering a new strategy in discovering and developing antivirals from natural products that are safe for human use.展开更多
Introduction: Lassa viral hemorrhagic fever is common in West Africa. Almost 300,000 persons are affected each year with 5000 deaths. The mice of the genus mastomys is the wild tank. Objective: The aim of our study is...Introduction: Lassa viral hemorrhagic fever is common in West Africa. Almost 300,000 persons are affected each year with 5000 deaths. The mice of the genus mastomys is the wild tank. Objective: The aim of our study is to describe clinic, therapeutic and evolution of the affected patients during February-March 2017 epidemic that occurred in the north of Togo. Methodology: Our study is a record review study from patients’ record, who were hospitalized from February, 1st to March, 31st 2017 at Mango hospital. Lassa diagnosis was performed by PCR. They patients have received Ribavirin and blood transfusion when necessary. Results: We have reported 5 clinical observations of Lassa viral hemorrhagic fever. Patients came from Benin (03 cases), from Burkina-Faso (1 case), from Togo (1 case) and were 25, 34, 60, 52 years old and a premature baby of 13 days. External hemorrhage and abdominal pains were the main symptoms. Fever was observed for all the cases. Complications were marked by hemorrhages and shocks. Only 3 patients had benefitted of Antiviral therapy with Ribavirin. The other 2 patients did not benefit from the treatment because the diagnosis of Lassa fever was done the day they dead before the treatment started. Lethality was 80% (4 cases) with a highly secured burial. Effective management of contacts was done. Conclusion: Diagnostic and therapeutic delays of patients are responsible of the bad prognosis of the disease.展开更多
Introduction: Lassa viral hemorrhagic fever is caused by the Lassa virus. The aim of our study is to describe the therapeutic itinerary of the 4 cases of Lassa virus hemorrhagic of February-March 2017 epidemic that oc...Introduction: Lassa viral hemorrhagic fever is caused by the Lassa virus. The aim of our study is to describe the therapeutic itinerary of the 4 cases of Lassa virus hemorrhagic of February-March 2017 epidemic that occurred in Mango. Methodology: Our study is a transverse retro-prospective and descriptive study from February, 1st to March, 31st 2017 that dealt with 4 confirmed Lassa fever cases declared positive on the PCR basis;hospitalized or deceased at the hospital “Esperance” of Mango;support center of Lassa viral hemorrhagic fever. Results: we reported 4 clinical observations of Lassa viral hemorrhagic fever diagnosed on the PCR basis during the Lassa fever epidemic. Patients came from Benin (2 cases) or from Burkina-Faso (1 case) and were 25, 60, 52 years old and a premature baby of 13 days. The reasons for admission were external hemorrhage, a pultated tonsillitis and abdominal pains. Fever was observed for all the cases. Complications were marked by hemorrhages and shocks. Only two patients benefitted from Antiviral therapy with Ribavirin and were declared healed. The other two patients did not benefit from the treatment due to diagnostic and therapeutic delays. Lethality was 75% (3 cases) with a highly secured burial. Effective management of contacts was established. Conclusion: Diagnostic and therapeutic delays of patients are responsible of the Dark Prognosis of Lassa fever during the epidemic.展开更多
The membrane-proximal external region(MPER)of Lassa virus(LASV)glycoprotein complex(GPC)is critical in modulating its functionality.Till now,the high-resolution structure of the intact GPC,including MPER is not availa...The membrane-proximal external region(MPER)of Lassa virus(LASV)glycoprotein complex(GPC)is critical in modulating its functionality.Till now,the high-resolution structure of the intact GPC,including MPER is not available.In this study,we used alanine substitution to scan all 16 residues located in LASV MPER.Western blotting and quantification fusion assay showed that the residues located at the C terminus of the HR2(M414 and L415)and N terminus of the MPER(K417 and Y419)are critical for GPC-mediated membrane fusion function.Furthermore,cell surface biotinylation experiments revealed that M414 A,K417 A and Y419 A expressed similar levels as WT,whereas L415 A mutant led to a reduction of mature GPC on the cell surface.Moreover,substitution of these residues with the similar residue such as M414 L,L415 I,K417 R and Y419 F would partly compensate the loss of the fusion activity caused by the alanine mutant in these sites.Results from this study showed that several key residues in the MPER region are indispensable to promote the conformational changes that drive fusion events and shed light on the structure analysis of LASV GPC and anti-LASV therapeutics.展开更多
Lassa virus(LASV)belongs to the Mammarenavirus genus(family Arenaviridae)and causes severe hemorrhagic fever in humans.The glycoprotein complex(GPC)contains eleven N-linked glycans that play essential roles in GPC fun...Lassa virus(LASV)belongs to the Mammarenavirus genus(family Arenaviridae)and causes severe hemorrhagic fever in humans.The glycoprotein complex(GPC)contains eleven N-linked glycans that play essential roles in GPC functionalities such as cleavage,transport,receptor recognition,epitope shielding,and immune response.We used three mutagenesis strategies(asparagine to glutamine,asparagine to alanine,and serine/tyrosine to alanine mutants)to abolish individual glycan chain on GPC and found that all the three strategies led to cleavage inefficiency on the 2nd(N89),5th(N119),or 8th(N365)glycosylation motif.To evaluate N to Q mutagenesis for further research,it was found that deletion of the 2nd(N89Q)or 8th(N365Q)glycan completely inhibited the transduction efficiency of pseudotyped particles.We further investigated the role of individual glycan on GPC-mediated immune response by DNA immunization of mice.Deletion of the individual 1st(N79Q),3rd(N99Q),5th(N119Q),or 6th(N167Q)glycan significantly enhanced the proportion of effector CD4+cells,whereas deletion of the 1st(N79Q),2nd(N89Q),3rd(N99Q),4th(N109Q),5th(N119Q),6th(N167Q),or 9th(N373Q)glycan enhanced the proportion of CD8+effector T cells.Deletion of specific glycan improves the Th1-type immune response,and abolishment of glycan on GPC generally increases the antibody titer to the glycan-deficient GPC.However,the antibodies from either the mutant or WT GPC-immunized mice show little neutralization effect on wild-type LASV.The glycan residues on GPC provide an immune shield for the virus,and thus represent a target for the design and development of a vaccine.展开更多
文摘Background: In recent times, there has been an increase in the number of Lassa fever cases resulting from the several episodes of Lassa fever epidemics ravaging Nigeria and other West African countries. The presence of Lassa virus in rodents other than the major reservoir (Mastomys natalensis) has been a public health concern as to the actual burden of the disease. It is therefore of a public health necessity to explore the LASV RNA habouring potential of several species of rodents in endemic as well as non-endemic areas for proper prevention of emergence of outbreaks in non-endemic areas. Objectives: The aim of this study was to detect the presence of LASV RNA in different species of rodents in Ikorodu, Lagos state and Abeokuta, Ogun state. Methods: A total of ninety one (91) rodents were captured from Ikorodu, Lagos State (61 rodents) and Abeokuta, Ogun State (30 rodents), euthanized, bled, and plasma obtained for the detection of LASV RNA by Reverse Transcriptase Polymerase Chain Reaction. Results: A total of 91 rodents consisting of 77 Rattus rattus and 14 Crocidura spp. The S segment of LASV RNA was not in any of the 91 rodents’ plasma samples. Conclusion: The rodents captured within the rural communities of Ikorodu, Lagos State and Abeokuta, Ogun State were found not to habour the LASV RNA. This study is limited by the relatively small sample size. Similar studies should be encouraged both in endemic and non-endemic areas in order to understand the actual burden of Lassa fever as well as put into check future epidemics.
基金supported by the National Key Research and Development Program of China(2023YFC2605504,2022YFC2303300)the National Natural Sciences Foundation of China(82172273 and 31670165)+3 种基金the Open Research Fund Program of the State Key Laboratory of Virology of China(2023JZZD-01)the Health research project of Shaanxi Province(2022D040)the Science and Technology Planning Project of Shaanxi Provincial Education Department(22JK0545)the Natural Science Basic Research Program of Shaanxi(2024JC-YBQN-0922).
文摘Lassa virus(LASV)is an enveloped,negative-sense RNA virus that causes Lassa hemorrhagic fever.Successful entry of LASV requires the viral glycoprotein 1(GP1)to undergo a receptor switch from its primary receptor alpha-dystroglycan(α-DG)to its endosomal receptor lysosome-associated membrane protein 1(LAMP1).A conserved histidine triad in LASV GP1 has been reported to be responsible for receptor switch.To test the hypothesis that other non-conserved residues also contribute to receptor switch,we constructed a series of mutant LASV GP1 proteins and tested them for binding to LAMP1.Four residues,L84,K88,L107,and H170,were identified as critical for receptor switch.Substituting any of the four residues with the corresponding lymphocytic choriomeningitis virus(LCMV)residue(L84 N,K88E,L10F,and H170S)reduced the binding affinity of LASV GP1 for LAMP1.Moreover,all mutations caused decreases in glycoprotein precursor(GPC)-mediated membrane fusion at both pH 4.5 and 5.2.The infectivity of pseudotyped viruses bearing either GPCL84N or GPCK88E decreased sharply in multiple cell types,while L107F and H170S had only mild effects on infectivity.Using biolayer light interferometry assay,we found that all four mutants had decreased binding affinity to LAMP1,in the order of binding affinity being L84 N>L107F>K88E>H170S.The four amino acid loci identified for the first time in this study have important reference significance for the in-depth investigation of the mechanism of receptor switching and immune escape of LASV occurrence and the development of reserve anti-LASV infection drugs.
基金the National Key Research and Development Program(2022YFC2303300,2018YFA0507204)the Strategic Priority Research Program of the Chinese Academy of Sciences(XDB0490000)the Na-tional Natural Science Foundation of China(82172273,31670165).
文摘The Lassa virus(LASV)is endemic in West Africa and causes severe hemorrhagic Lassa fever in humans.The glycoprotein complex(GPC)of LASV is highly glycosylation-modified,with 11 N-glycosylation sites.All 11 N-linked glycan chains play critical roles in GPC cleavage,folding,receptor binding,membrane fusion,and immune evasion.In this study,we focused on the first glycosylation site because its deletion mutant(N79Q)results in an unexpected enhanced membrane fusion,whereas it exerts little effect on GPC expression,cleavage,and receptor binding.Meanwhile,the pseudotype virus bearing GPC_(N79Q)was more sensitive to the neutralizing antibody 37.7H and was attenuated in virulence.Exploring the biological functions of the key glycosylation site on LASV GPC will help elucidate the mechanism of LASV infection and provide strategies for the development of attenuated vaccines against LASV infection.
文摘<strong>Introduction: </strong>Lassa fever is a disease of public health importance because of the associated morbidity and high case fatality rate among hospitalized patients. Even after recovery, there may be residual problems such as sensorineural hearing loss. The initial presentation of Lassa fever may be with non-specific symptoms similar to what is seen in the more common febrile illnesses such as malaria or typhoid fever. In such a setting therefore, timely diagnosis of Lassa fever may be difficult. <strong>Case Report: </strong>We report a case of Lassa fever that presented to our institution. She was a middle aged woman who had non-specific symptoms of febrile illness and who died in less than 48 hours of admission. She had a subtle bleeding on the lip just before death which was what raised the suspicion for Lassa fever. Laboratory confirmation of Lassa fever was made retrospectively.<strong> Conclusion: </strong>When the presenting symptoms are non-specific, a high index of suspicion is required for timely recognition of Lassa fever. Early diagnosis is important for prompt therapeutic intervention as well as for limiting the spread of the disease. This is the second case of Lassa fever presenting to our hospital, but the first published case of Lassa fever from our hospital.
基金financially supported by the National Natural Science Foundation of China(Nos.81473256 and 81273561)the CAMS Innovation Fund for Medical Sciences(No.2016-I2M-1014,China)+3 种基金the Science and Technology Program of Beijing(No.Z151100000115008,China)the Beijing Key Laboratory of New Drug Mechanisms and Pharmacological Evaluation Study(No.BZ0150,China)the Drug Innovation Major Project(Nos.2015ZX09102-023 and 2018ZX09711001-003-002,China)the Disciplines Construction Project(No.201920200802,China)
文摘The limited treatment options for the increasing occurrence of Lassa hemorrhagic fever in West Africa poses an urgent need for the discovery and development of novel therapeutics.Dietary supplements,especially natural products that are edible and safe for human use,are a good source of drug discovery with potential for uncovering novel applications,In this study,we tested 40 natural products of dietary supplements and identified capsaicin,a common dietary supplement abundant in chili peppers,as an inhibitor of Lassa virus(LASV)entry with EC5,of 6.9-10.0μmol/L using an HIV based pseudo virus platform.Capsaicin inhibits the entry of five LASV strains but not against the Old World arenavirus lymphocytic choriomeningitis virus(LCMV),showing a preferential activity against LASV.Capsaicin inhibits LASV entry by blocking the pH dependent viral fusion through affecting the stable signal peptide(SSP)-GP2 transmembrane(GP2TM)region of the LASV surface glycoprotein.Mutational study revealed the key residues Ala25,Val431,Phe434 and Val435 in SsP-CP2TM region in capsaicin’s antiviral effect.This study for the first time reveals a direct acting antiviral effect of capsaicin against the hemorrhagic fever causing LASV,providing detailed interaction hot spots in the unique SSP-GP2TMinterface of LASV glycoprotein that is crucial in fusion inhibition,and offering a new strategy in discovering and developing antivirals from natural products that are safe for human use.
文摘Introduction: Lassa viral hemorrhagic fever is common in West Africa. Almost 300,000 persons are affected each year with 5000 deaths. The mice of the genus mastomys is the wild tank. Objective: The aim of our study is to describe clinic, therapeutic and evolution of the affected patients during February-March 2017 epidemic that occurred in the north of Togo. Methodology: Our study is a record review study from patients’ record, who were hospitalized from February, 1st to March, 31st 2017 at Mango hospital. Lassa diagnosis was performed by PCR. They patients have received Ribavirin and blood transfusion when necessary. Results: We have reported 5 clinical observations of Lassa viral hemorrhagic fever. Patients came from Benin (03 cases), from Burkina-Faso (1 case), from Togo (1 case) and were 25, 34, 60, 52 years old and a premature baby of 13 days. External hemorrhage and abdominal pains were the main symptoms. Fever was observed for all the cases. Complications were marked by hemorrhages and shocks. Only 3 patients had benefitted of Antiviral therapy with Ribavirin. The other 2 patients did not benefit from the treatment because the diagnosis of Lassa fever was done the day they dead before the treatment started. Lethality was 80% (4 cases) with a highly secured burial. Effective management of contacts was done. Conclusion: Diagnostic and therapeutic delays of patients are responsible of the bad prognosis of the disease.
文摘Introduction: Lassa viral hemorrhagic fever is caused by the Lassa virus. The aim of our study is to describe the therapeutic itinerary of the 4 cases of Lassa virus hemorrhagic of February-March 2017 epidemic that occurred in Mango. Methodology: Our study is a transverse retro-prospective and descriptive study from February, 1st to March, 31st 2017 that dealt with 4 confirmed Lassa fever cases declared positive on the PCR basis;hospitalized or deceased at the hospital “Esperance” of Mango;support center of Lassa viral hemorrhagic fever. Results: we reported 4 clinical observations of Lassa viral hemorrhagic fever diagnosed on the PCR basis during the Lassa fever epidemic. Patients came from Benin (2 cases) or from Burkina-Faso (1 case) and were 25, 60, 52 years old and a premature baby of 13 days. The reasons for admission were external hemorrhage, a pultated tonsillitis and abdominal pains. Fever was observed for all the cases. Complications were marked by hemorrhages and shocks. Only two patients benefitted from Antiviral therapy with Ribavirin and were declared healed. The other two patients did not benefit from the treatment due to diagnostic and therapeutic delays. Lethality was 75% (3 cases) with a highly secured burial. Effective management of contacts was established. Conclusion: Diagnostic and therapeutic delays of patients are responsible of the Dark Prognosis of Lassa fever during the epidemic.
基金the National Key Research and Development Program of China (2018YFA0507204)the National Natural Sciences Foundation of China (31670165)+1 种基金Wuhan National Biosafety Laboratory,Chinese Academy of Sciences Advanced Customer Cultivation Project (2019ACCP-MS03)the Open Research Fund Program of the State Key Laboratory of Virology of China (2018IOV001)。
文摘The membrane-proximal external region(MPER)of Lassa virus(LASV)glycoprotein complex(GPC)is critical in modulating its functionality.Till now,the high-resolution structure of the intact GPC,including MPER is not available.In this study,we used alanine substitution to scan all 16 residues located in LASV MPER.Western blotting and quantification fusion assay showed that the residues located at the C terminus of the HR2(M414 and L415)and N terminus of the MPER(K417 and Y419)are critical for GPC-mediated membrane fusion function.Furthermore,cell surface biotinylation experiments revealed that M414 A,K417 A and Y419 A expressed similar levels as WT,whereas L415 A mutant led to a reduction of mature GPC on the cell surface.Moreover,substitution of these residues with the similar residue such as M414 L,L415 I,K417 R and Y419 F would partly compensate the loss of the fusion activity caused by the alanine mutant in these sites.Results from this study showed that several key residues in the MPER region are indispensable to promote the conformational changes that drive fusion events and shed light on the structure analysis of LASV GPC and anti-LASV therapeutics.
基金This work was supported by the National Key Research and Development Program of China(2018YFA0507204)the National Natural Sciences Foundation of China(31670165)+1 种基金Wuhan National Biosafety Laboratory,Chinese Academy of Sciences Advanced Customer Cultivation Project(2019ACCP-MS03)the Open Research Fund Program of the State Key Laboratory of Virology of China(2018IOV001).Author Contributions。
文摘Lassa virus(LASV)belongs to the Mammarenavirus genus(family Arenaviridae)and causes severe hemorrhagic fever in humans.The glycoprotein complex(GPC)contains eleven N-linked glycans that play essential roles in GPC functionalities such as cleavage,transport,receptor recognition,epitope shielding,and immune response.We used three mutagenesis strategies(asparagine to glutamine,asparagine to alanine,and serine/tyrosine to alanine mutants)to abolish individual glycan chain on GPC and found that all the three strategies led to cleavage inefficiency on the 2nd(N89),5th(N119),or 8th(N365)glycosylation motif.To evaluate N to Q mutagenesis for further research,it was found that deletion of the 2nd(N89Q)or 8th(N365Q)glycan completely inhibited the transduction efficiency of pseudotyped particles.We further investigated the role of individual glycan on GPC-mediated immune response by DNA immunization of mice.Deletion of the individual 1st(N79Q),3rd(N99Q),5th(N119Q),or 6th(N167Q)glycan significantly enhanced the proportion of effector CD4+cells,whereas deletion of the 1st(N79Q),2nd(N89Q),3rd(N99Q),4th(N109Q),5th(N119Q),6th(N167Q),or 9th(N373Q)glycan enhanced the proportion of CD8+effector T cells.Deletion of specific glycan improves the Th1-type immune response,and abolishment of glycan on GPC generally increases the antibody titer to the glycan-deficient GPC.However,the antibodies from either the mutant or WT GPC-immunized mice show little neutralization effect on wild-type LASV.The glycan residues on GPC provide an immune shield for the virus,and thus represent a target for the design and development of a vaccine.
