Porcine epidemic diarrhea virus(PEDV)is the most common diarrhea-causing pathogen in newborn piglets.The clarifications of the overall antibody repertoire and antigen-specific antibody repertoire are essential to prov...Porcine epidemic diarrhea virus(PEDV)is the most common diarrhea-causing pathogen in newborn piglets.The clarifications of the overall antibody repertoire and antigen-specific antibody repertoire are essential to provide important insights into the B-cell response and reshape new vaccines.Here,we applied next-generation sequencing(NGS)technology to investigate immunoglobulin(Ig)variable(V)gene segment usage of swine B-cells from peripheral blood lymphocytes(PBL)and mesenteric lymph node(MLN)cells following PEDV vaccination.We identified the transcripts of all functional Ig V-genes in antibody repertoire.IgHV1 S2,IgKV1-11,and IgLV3-4 were the most prevalent gene segments for heavy,kappa,and lambda chains,respectively,in PBL and MLN.Unlike previous studies,IgKV1,instead of IgKV2,and IgLV3,instead of IgLV8,were the prevalent Ig V-gene families for kappa and lambda light chains,respectively.We further examined the antibody repertoire of PEDV spike-specific B cells by single-cell RT-PCR.In contrast to the overall antibody repertoire,Ig V-gene segments of PEDV spike-specific B cells preferentially adopted IgHV1-4 and IgHV1-14 for heavy chain,IgKV1-11 for kappa chain,and IgLV3-3 for lambda chain.These results represent a comprehensive analysis to characterize the Ig V-gene segment usage in the overall and PEDV spike-specific antibody repertoire in PBL and MLN.展开更多
Coronaviruses(CoVs)are important human and animal pathogens that cause respiratory and gastrointestinal diseases.Porcine epidemic diarrhoea(PED),characterized by severe diarrhoea and vomiting in pigs,is a highly letha...Coronaviruses(CoVs)are important human and animal pathogens that cause respiratory and gastrointestinal diseases.Porcine epidemic diarrhoea(PED),characterized by severe diarrhoea and vomiting in pigs,is a highly lethal disease caused by porcine epidemic diarrhoea virus(PEDV)and causes substantial losses in the swine industry worldwide.However,currently available commercial drugs have not shown great therapeutic effects.In this study,a fluorescence resonance energy transfer(FRET)-based assay was applied to screen a library containing 1,590 compounds and identified two compounds,3-(aminocarbonyl)-1-phenylpyridinium and 2,3-dichloronaphthoquinone,that target the 3C-like protease(3CL^(pro))of PEDV.These compounds are of low molecular weight(MW)and greatly inhibited the activity of this enzyme(IC_(50) values were obtained in this study).Furthermore,these compounds exhibited antiviral capacity against another member of the CoV family,feline infectious peritonitis virus(FIPV).Here,the inhibitory effects of these compounds against CoVs on Vero cells and feline kidney cells were identified(with EC_(50) values)and cell viability assays were performed.The results of putative molecular docking models indicate that these compounds,labeled compound 1 and compound 2,contact the conserved active sites(Cys144,Glu165,Gln191)of 3CL^(pro) via hydrogen bonds.These findings provide insight into the antiviral activities of compounds 1 and 2 that may facilitate future research on anti-CoV drugs.展开更多
基金supported by the National Natural Science Foundation of China(31772718)the Open Research Fund of State Key Laboratory of Veterinary Biotechnology(SKLVBF2018XX)。
文摘Porcine epidemic diarrhea virus(PEDV)is the most common diarrhea-causing pathogen in newborn piglets.The clarifications of the overall antibody repertoire and antigen-specific antibody repertoire are essential to provide important insights into the B-cell response and reshape new vaccines.Here,we applied next-generation sequencing(NGS)technology to investigate immunoglobulin(Ig)variable(V)gene segment usage of swine B-cells from peripheral blood lymphocytes(PBL)and mesenteric lymph node(MLN)cells following PEDV vaccination.We identified the transcripts of all functional Ig V-genes in antibody repertoire.IgHV1 S2,IgKV1-11,and IgLV3-4 were the most prevalent gene segments for heavy,kappa,and lambda chains,respectively,in PBL and MLN.Unlike previous studies,IgKV1,instead of IgKV2,and IgLV3,instead of IgLV8,were the prevalent Ig V-gene families for kappa and lambda light chains,respectively.We further examined the antibody repertoire of PEDV spike-specific B cells by single-cell RT-PCR.In contrast to the overall antibody repertoire,Ig V-gene segments of PEDV spike-specific B cells preferentially adopted IgHV1-4 and IgHV1-14 for heavy chain,IgKV1-11 for kappa chain,and IgLV3-3 for lambda chain.These results represent a comprehensive analysis to characterize the Ig V-gene segment usage in the overall and PEDV spike-specific antibody repertoire in PBL and MLN.
基金This work was supported by the National Key R&D Plan of China(grant no.2018YFD0500102)the Natural Science Foundation of Hubei Province of China(grant no.2016CFA069)。
文摘Coronaviruses(CoVs)are important human and animal pathogens that cause respiratory and gastrointestinal diseases.Porcine epidemic diarrhoea(PED),characterized by severe diarrhoea and vomiting in pigs,is a highly lethal disease caused by porcine epidemic diarrhoea virus(PEDV)and causes substantial losses in the swine industry worldwide.However,currently available commercial drugs have not shown great therapeutic effects.In this study,a fluorescence resonance energy transfer(FRET)-based assay was applied to screen a library containing 1,590 compounds and identified two compounds,3-(aminocarbonyl)-1-phenylpyridinium and 2,3-dichloronaphthoquinone,that target the 3C-like protease(3CL^(pro))of PEDV.These compounds are of low molecular weight(MW)and greatly inhibited the activity of this enzyme(IC_(50) values were obtained in this study).Furthermore,these compounds exhibited antiviral capacity against another member of the CoV family,feline infectious peritonitis virus(FIPV).Here,the inhibitory effects of these compounds against CoVs on Vero cells and feline kidney cells were identified(with EC_(50) values)and cell viability assays were performed.The results of putative molecular docking models indicate that these compounds,labeled compound 1 and compound 2,contact the conserved active sites(Cys144,Glu165,Gln191)of 3CL^(pro) via hydrogen bonds.These findings provide insight into the antiviral activities of compounds 1 and 2 that may facilitate future research on anti-CoV drugs.
