简介：AnnotationofthegenomesequenceoftheSARS-CoV(severeacuterespiratorysyndrome-associatedcoronavirus)isindispensabletounderstanditsevolutionandpathogenesis.WehaveperformedafullannotationoftheSARS-CoVgenomesequencesbyusingannotationprogramspubliclyavailableordevelopedbyourselves.Totally,21openreadingframes(ORFs)ofgenesorputativeuncharacterizedproteins(PUPs)werepredicted.SevenPUPshadnotbeenreportedpreviously,andtwoofthemwerepredictedtocontaintransmembraneregions.EightORFspartiallyoverlappedwithorembeddedintothoseofknowngenes,revealingthattheSARS-CoVgenomeisasmallandcompactonewithoverlappedcodingregions.ThemoststrikingdiscoveryisthatanORFlocatesontheminusstrand.Wehavealsoannotatednon-codingregionsandidentifiedthetranscriptionregulatingsequences(TRS)intheintergenicregions.TheanalysisofTRSsupportstheminusstrandextendingtranscriptionmechanismofcoronavirus.TheSNPanalysisofdifferentisolatesrevealsthatmutationsofthesequencesdonotaffectthepredictionresultsofORFs.

简介：Knowledgeoftheevolutionofpathogensisofgreatmedicalandbiologicalsignificancetotheprevention,diagnosis,andtherapyofinfectiousdiseases.InordertounderstandtheoriginandevolutionoftheSARS-CoV(severeacuterespiratorysyndrome-associatedcoronavirus),wecollectedcompletegenomesequencesofallvirusesavailableinGenBank,andmadecomparativeanalyseswiththeSARSCoV.GenomicsignatureanalysisdemonstratesthatthecoronavirusesalltaketheTGTTastheirrichesttetranucleotideexcepttheSARS-CoV.Adetailedanalysisoftheforty-twocompleteSARS-CoVgenomesequencesrevealedtheexistenceoftwodistinctgenotypes,andshowedthattheseisolatescouldbeclassifiedintofourgroups.OurmanualanalysisoftheBLASTNresultsdemonstratesthattheHE(hemagglutinin-esterase)geneexistsintheSARS-CoV,andmanymutationsmadeitunfamiliartous.

简介：Adatasetof103SARS-CoVisolates(101humanpatientsand2palmcivets)wasinvestigatedondifferentaspectsofgenomepolymorphismandisolateclassification.Thenumberandthedistributionofsinglenucleotidevariations(SNVs)andinsertionsanddeletions,withrespecttoa"profile",weredeterminedanddiscussed("profile"beingasequencecontainingthemostrepresentedletterperposition).Distributionofsubstitutioncategoriespercodonpositions,aswellassynonymousandnon-synonymoussubstitutionsincodingregionsofannotatedisolates,wasdetermined,alongwithaminoacid(a.a.)propertychanges.Similaranalysiswasperformedforthespike(S)proteininalltheisolates(55ofthembeingpredictedforthefirsttime).TheratioKa/KsconfirmedthattheSgenewassubjectedtotheDarwinianselectionduringvirustransmissionfromanimalstohumans.Isolatesfromthedatasetwereclassifiedaccordingtogenomepolymorphismandgenotypes.Genomepolymorphismyieldstotwogroups,onewithasmallnumberofSNVsandanotherwithalargenumberofSNVs,withuptofoursubgroupswithrespecttoinsertionsanddeletions.Weidentifiedthreebasicnine-locusgenotypes:TTTT/TTCGG,CGCC/TTCAT,andTGCC/TTCGT,withfoursubgenotypes.Bothclassificationsproposedareinaccordancewiththenewinsightsintopossibleepidemiologicalspread,bothinspaceandtime.

简介：Thenucleocapsidprotein(Nprotein)hasbeenfoundtobeanantigenicproteininanumberofcoronaviruses.WhethertheNproteininsevereacuterespiratorysyndrome-associatedcoronavirus(SARS-CoV)isantigenicremainstobeelucidated.UsingWesternblotandEnzyme-linkedImmunosorbentAssay(ELISA),therecombinantNproteinsandthesynthesizedpeptidesderivedfromtheNproteinwerescreenedinserafromSARSpatients.AllpatientserainthisstudydisplayedstrongpositiveimmunoreactivitiesagainsttherecombinantNproteins,whereasnormalseragavenegativeimmunoresponsestotheseproteins,indicatingthattheNproteinofSARS-CoVisanantigenicprotein.Furthermore,theepitopesitesintheNproteinweredeterminedbycompetitionexperiments,inwhichtherecombinantproteinsorthesynthesizedpeptidescompetedagainsttheSARS-CoVproteinstobindtotheantibodiesraisedinSARSsera.OneepitopesitelocatedattheC-terminuswasconfirmedasthemostantigenicregioninthisprotein.AdetailedscreeningofpeptidewithELISAdemonstratedthattheaminosequencefromCodons371to407wastheepitopesiteattheC-terminusoftheNprotein.UnderstandingoftheepitopesitescouldbeverysignificantfordevelopinganeffectivediagnosticapproachtoSARS.

简介：TotesttheantigenicactivityofMprotein(Mcprotein)intheinnermembraneofSARS-CoV,SARS-CoVMcprotein'sbaseslocatinginsidethemembranewerecloned,theHis-fusionproteinwasexpressedinE.coliandanalyzedforitsantigenicactivity.Amongthose7clinicallydiagnosedpatients'sera,therewere5positiveand2negativeinreactionwithHis-fusionprotein.Allofthe20healthypersons'seraandrabbitanti-OC43and229EwereofnegativereactionwithHis-fusionprotein.TheanimalsimmunizedwithHis-fusionproteinhaveproducedmulti-clonalantibody.TheHis-fusionproteincouldspeciallyreactwithclinicallydiagnosedSARSpatients'seraandtheanimalsimmunizedwithHis-fusionproteincouldproducespecificallymulti-clonalantibody,butitcouldnotreactwiththeseraofhealthypersonsandtherabbitanti-OC43and229E.

简介：TheE(envelope)proteinisthesmalleststructuralproteininallcoronavirusesandistheonlyviralstructuralproteininwhichnovariationhasbeendetected.WeconductedgenomesequencingandphylogeneticanalysesofSARS-CoV.Basedongenomesequencing,wepredictedtheEproteinisatransmembrane(TM)pro-teincharacterizedbyaTMregionwithstronghydrophobicityandα-helixcon-formation.Weidentifiedasegment(NH2-_L-Cys-A-Y-Cys-Cys-N_-COOH)inthecarboxyl-terminalregionoftheEproteinthatappearstoformthreedisulfidebondswithanothersegmentofcorrespondingcysteinesinthecarboxyl-terminusoftheS(spike)protein.ThesebondspointtoapossiblestructuralassociationbetweentheEandSproteins.OurphylogeneticanalysesoftheEproteinsequencesinallpub-lishedcoronavirusesplaceSARS-CoVinanindependentgroupinCoronaviridaeandsuggestanon-humananimalorigin.

简介：Thecorona-likespikesorpeplomersonthesurfaceofthevirionunderelectronicmicroscopearethemoststrikingfeaturesofcoronaviruses.TheS(spike)proteinisthelargeststructuralprotein,with1,255aminoacids,intheviralgenome.Itsstructurecanbedividedintothreeregions:alongN-terminalregionintheexte-rior,acharacteristictransmembrane(TM)region,andashortC-terminusintheinteriorofavirion.WedetectedfifteensubstitutionsofnucleotidesbycomparisonswiththeseventeenpublishedSARS-CoVgenomesequences,eight(53.3%)ofwhicharenon-synonymousmutationsleadingtoaminoacidalternationswithpredictedphysiochemicalchanges.ThepossibleantigenicdeterminantsoftheSproteinarepredicted,andtheresultisconfirmedbyELISA(enzyme-linkedimmunosorbentassay)withsynthesizedpeptides.AnotherprofoundfindingisthatthreedisulfidebondsaredefinedattheC-terminuswiththeN-terminusoftheE(envelope)pro-tein,basedonthetypicalsequenceandpositions,thusestablishingthestructuralconnectionwiththesetwoimportantstructuralproteins,ifconfirmed.Phyloge-neticanalysisrevealsseveralconservedregionsthatmightbepotentdrugtargets.

简介：TheCoronaviridaefamilyischaracterizedbyanucleocapsidthatiscomposedofthegenomeRNAmoleculeincombinationwiththenucleoprotein(Nprotein)withinavirion.ThemoststrikingphysiochemicalfeatureoftheNproteinofSARS-CoVisthatitisatypicalbasicproteinwithahighpredictedpIandhighhydrophilicity,whichisconsistentwithitsfunctionofbindingtotheribophosphatebackboneoftheRNAmolecule.ThepredictedhighextentofphosphorylationoftheNproteinonmultiplecandidatephosphorylationsitesdemonstratesthatitwouldberelatedtoimportantfunctions,suchasRNA-bindingandlocalizationtothenucleolusofhostcells.SubsequentstudyshowsthatthereisanSR-richregionintheNproteinandthisregionmightbeinvolvedintheprotein-proteininteraction.TheabundantantigenicsitespredictedintheNprotein,aswellasexperimentalevidencewithsynthesizedpolypeptides,indicatethattheNproteinisoneofthemajorantigensoftheSARS-CoV.Comparedwithotherviralstructuralproteins,thelowvariationrateoftheNproteinwithregardstoitssizesuggestsitsimportancetothesurvivalofthevirus.

简介：WestudiedstructuralandimmunologicalpropertiesoftheSARS-CoVM(mem-brane)protein,basedoncomparativeanalysesofsequencefeatures,phylogeneticinvestigation,andexperimentalresults.TheMproteinispredictedtocontainatriple-spanningtransmembrane(TM)region,asingleN-glycosylationsitenearitsN-terminusthatisintheexteriorofthevirion,andalongC-terminalregionintheinterior.TheMproteinharborsahighersubstitutionrate(0.6%correlatedtoitssize)amongviralopenreadingframes(ORFs)frompublisheddata.ThefoursubstitutionsdetectedintheMprotein,whichcausenon-synonymouschanges,canbeclassifiedintothreetypes.OneofthemresultsinchangesofpI(isoelectricpoint)andcharge,affectingantigenicity.ThesecondchangeshydrophobicityoftheTMregion,andthethirdonerelatestohydrophilicityoftheinteriorstructure.PhylogenetictreebuildingbasedonthevariationsoftheMproteinappearstosupportthenon-humanoriginofSARS-CoV.Toinvestigateitsimmunogenicity,wesynthesizedeightoligopeptidescovering69.2%oftheentireORFandscreenedthembyusingELISA(enzyme-linkedimmunosorbentassay)withserafromSARSpatients.Theresultsconfirmedourpredictionsonantigenicsites.

简介：Inthefaceoftheworldwidethreatofsevereacuterespiratorysyndrome(SARS)tohumanlife,someofthemosturgentchallengesaretodevelopfastandaccurateanalyticalmethodsforearlydiagnosisofthisdiseaseaswellastocreateasafeanti-viralvaccineforprevention.Totheseends,weinvestigatedtheantigenicityofthespikeprotein(Sprotein),amajorstructuralproteinintheSARS-coronavirus(SARS-CoV).BaseduponthetheoreticalanalysisforhydrophobicityoftheSprotein,18peptidesweresynthesized.UsingEnzyme-LinkedImmunosorbentAssay(ELISA),thesepeptideswerescreenedintheserafromSARSpatients.Accordingtotheseresults,twofragmentsoftheSgenewereamplifiedbyPCRandclonedintopET-32a.BothSfragmentswereexpressedintheBL-21strainandfurtherpurifiedwithanaffinitychromatography.TheserecombinantSfragmentswereconfirmedtohavepositivecross-reactionswithSARSsera,eitherbyWesternblotorbyELISA.OurresultsdemonstratedthatthepotentialepitoperegionswerelocatedatCodons469-882intheSprotein,andoneepitopesitewaslocatedatCodons599-620.IdentificationofantigenicregionsintheSARS-CoVSproteinmaybeimportantforthefunctionalstudiesofthisvirusorthedevelopmentofclinicaldiagnosis.

简介：ToinvestigatethesignificanceoftheSARS-associatedcoronavirus(SARS-CoV)antibody,detectedbyELISAandindirectimmunofluorescenceassays(IFA)fortheSARS-CoVVeroE6celllysates,innon-SARSsubjects,114serumsamplesfromhealthycontrolsand104serumspecimensfromautoimmunediseasepatientswerecollected.TheresultsofELISAshowedthatamong114serafromhealthycontrols,4(3.5%)werepositiveofSARS-CoV-IgGantibodyand114(100%)wereallnegativeofSARS-CoV-IgMantibody;thespecificityofSARS-CoV-IgGantibodyforSARSpatientswas96.5%,butthespecificityofbothSARS-CoV-IgGand-IgMantibodiesforSARSpatientswas100%.In58caseswithSLE,positiveratesofSARS-CoV-IgGand-IgMantibodieswere32.8%(19/58)and8.6%(5/58),respectively,inwhich11cases(19%)werepositiveofbothSARS-CoV-IgGand-IgMantibodies;in10caseswithSS,positiverateofbothSARS-CoV-IgGand-IgMantibodieswas10%(1/10);in16caseswithMCTD,positiverateofSARS-CoV-IgGwas37.5%(6/16),positiverateofbothSARS-CoV-IgGand-IgMantibodieswas6.3%(1/16);in20caseswithRA,onecasewaspositive(5%)ofSARS-CoV-IgG.However,ofallsampleswithpositiveSARS-CoV-IgGand-IgMantibodiesforautoimmunediseasesandhealthycontrols,SARS-CoVRNAandantibodieswereallnegativebyRT-PCRandIFA.AllserafornegativeorpositiveELISAresultswerealsonegativeorpositiveresultsusingELISAwithVeroE6cellslysates.ThesestudiesshowedthatSARS-CoVVeroE6celllysatesfortheELISAtodetectSARS-CoVantibodiescouldleadtothefalse-positivereactionsorcross-reactionsofSARS-CoVantibodiesinnon-SARSdiseasesandhealthycontrols,andthefalse-positivereactionsorcross-reactionswererelatedtoVeroE6celllysatesandautoantibodiesinnon-SARSpopulation.

简介：InordertodevelopclinicaldiagnostictoolsforrapiddetectionofSARS-CoV(severeacuterespiratorysyndrome-associatedcoronavirus)andtoidentifycandidateproteinsforvaccinedevelopment,theC-terminalportionofthenucleocapsid(NC)genewasamplifiedusingRT-PCRfromtheSARS-CoVgenome,clonedintoayeastexpressionvector(pEGH),andexpressedasaglutathioneS-transferase(GST)andHisx6double-taggedfusionproteinunderthecontrolofaninduciblepromoter.WesternanalysisonthepurifiedproteinconfirmedtheexpressionandpurificationoftheNCfusionproteinsfromyeast.Todetermineitsantigenicity,thefusionproteinwaschallengedwithserumsamplesfromSARSpatientsandnormalcontrols.TheNCfusionproteindemonstratedhighantigenicitywithhighspecificity,andtherefore,itshouldhavegreatpotentialindesigningclinicaldiagnostictoolsandprovideusefulinformationforvaccinedevelopment.

简介：Themultiplexpolymerasechainreaction(PCR)techniquewasappliedtodetecttheSARS-CoV(severeacuterespiratorysyndrome-associatedcoronavirus)specifictargetcDNAfragmentsinthepresentstudy.ThetargetcDNAfragmentsofSARS-CoVweresynthesizedartificiallyaccordingtothegenomesequenceofSARS-CoVinGenBanksubmittedbyTheChineseUniversityofHongKong,andwereusedassimulatedpositivesamples.FiveprimersrecommendedbyWorldHealthOrganization(WHO)wereusedtoamplifythefragmentsbysinglePCRandmultiplexPCR.ThreetargetcDNAfragments(121,182and302bp),aswellasthethreedifferentcombinationsofanytwoofthesefragments,wereamplifiedbysinglePCR.ThecombinationofthesethreefragmentswasamplifiedbymultiplexPCR.TheresultsindicatedthatthemultiplexPCRtechniquecouldbeappliedtodetecttheSARS-CoVspecifictargetcDNAfragmentssuccessfully.

简介：Thesuddenoutbreakofsevereacuterespiratorysyndrome(SARS)in2002promptedtheestablishmentofaglobalscientificnetworksubsumingmostofthetraditionalrivalriesinthecompetitivefieldofvirology.WithinmonthsoftheSARSoutbreak,collaborativeworkrevealedtheidentityofthedisastrouspathogenasSARS-associatedcoronavirus(SARS-CoV).However,althoughtherapididentificationoftheagentrepresentedanimportantbreakthrough,ourunderstandingofthedeadlyvirusremainslimited.Detailedbiologicalknowledgeiscrucialforthedevelopmentofeffectivecountermeasures,diagnostictests,vaccinesandantiviraldrugsagainsttheSARS-CoV.ThisarticlereviewsthepresentstateofmolecularknowledgeaboutSARS-CoV,fromtheaspectsofcomparativegenomics,molecularbiologyofviralgenes,evolution,andepidemiology,anddescribesthediagnostictestsandtheanti-viraldrugsderivedsofarbasedontheavailablemolecularinformation.

简介：BeijinghasbeenoneoftheepicentersattackedmostseverelybytheSARS-CoV(severeacuterespiratorysyndrome-associatedcoronavirus)sincethefirstpatientwasdiagnosedinoneofthecity'shospitals.WenowreportcompletegenomesequencesoftheBJGroup,includingfourisolates(IsolatesBJ01,BJ02,BJ03,andBJ04)oftheSARS-CoV.ItisremarkablethatallmembersoftheBJGroupshareacommonhaplotype,consistingofsevenlocithatdifferentiatethegroupfromotherisolatespublishedtodate.Among42substitutionsuniquelyidentifledfromtheBJgroup,32arenon-synonymouschangesattheaminoacidlevel.Rootedphylogenetictrees,proposedonthebasisofhaplotypesandothersequencevariationsofSARS-CoVisolatesfromCanada,USA,Singapore,andChina,gaverisetodifferentparadigmsbutpositionedtheBJGroup,togetherwiththenewlydiscoveredGD01(GD-Ins29)inthesameclade,followedbytheH-UGroup(fromHongKongtoUSA)andtheH-TGroup(fromHongKongtoToronto),leavingtheSPGroup(Singapore)moredistant.ThisresultappearstosuggestapossibletransmissionpathfromGuangdongtoBeijing/HongKong,thentoothercountriesandregions.

简介：摘要COVID-19的全球大流行使冠状病毒再次引起重视。在本世纪大流行的三种高致病性冠状病毒(SARS-CoV、MERS-CoV、SARS-CoV-2)研究中，已有直接证据证明SARS-CoV及SARS-CoV-2可引起人眼部感染。冠状病毒感染除引起眼部症状外，也可通过眼部感染引发全身多种临床表现。两种高致病性冠状病毒(SARS-CoV和SARS-CoV-2)具有更强的流行性及更高的病死率，两种病毒眼部症状相似，病毒结构及眼部感染过程也具有相似性，主要是通过其特异性S蛋白与细胞表面相关受体结合，使其核酸进入细胞内并借用细胞内的蛋白合成通路对自身蛋白进行转录、组装、折叠并通过其受体蛋白激发多种细胞因子表达。本文就2003年流行的SARS-CoV以及2019年末流行的SARS-CoV-2两种高致病性冠状病毒的特点、眼部感染通路的研究进展及病毒眼部感染相关的研究现状进行综述，阐述眼部病毒防护及患者眼部检查的必要性。(国际眼科纵览，2020, 45: 374-379)

简介：目的通过生物信息学与合成多肽方法,寻找SARS-CoVS蛋白的抗原表位,为抗SARS多肽疫苗的深入研究提供试验依据.方法利用网络共享软件对与S蛋白抗原性相关的参数进行预测分析,从而设计S蛋白的抗原表位多肽;将合成的抗原表位多肽免疫家兔,用ELISA法检测产生的抗体.结果ELISA法检测发现,设计合成的2个抗原表位多肽片段(残基348-380,434-470)在家兔体内均能诱导抗体产生.结论设计合成的2个抗原表位多肽片段中包含SARS-CoVS蛋白的抗原表位,免疫家兔后能产生特异性抗体.

简介：Wereportacompletegenomicsequenceofrareisolates(minorgenotype)oftheSARS-CoVfromSARSpatientsinGuangdong,China,wherethefirstfewcasesemerged.Themoststrikingdiscoveryfromtheisolateisanextra29-nucleotidesequencelocatedatthenucleotidepositionsbetween27,863and27,864(referredtothecompletesequenceofBJ01)withinanoverlappedregioncomposedofBGI-PUP5(BGI-postulateduncharacterizedprotein5)andBGI-PUP6upstreamoftheN(nucleocapsid)protein.Thediscoveryofthisminorgenotype,GD-Ins29,suggestsasignificantgeneticeventanddifferentiatesitfromthepreviouslyre-portedgenotype,thedominantformamongallsequencedSARS-CoVisolates.A17-ntsegmentofthisextrasequenceisidenticaltoasegmentofthesamesizeintwohumanmRNAsequencesthatmayinterferewithviralgenomereplicationandtranscriptioninthecytosoloftheinfectedcells.Itprovidesanewavenuefortheexplorationofthevirus-hostinteractioninviralevolution,hostpathogenesis,andvaccinedevelopment.

简介：TheR(replicase)proteinistheuniquelydefinednon-structuralprotein(NSP)responsibleforRNAreplication,mutationrateorfidelity,regulationoftranscrip-tionincoronavirusesandmanyotherssRNAviruses.Basedonourcompletegenomesequencesoffourisolates(BJ01-BJ04)ofSARS-CoVfromBeijing,China,weanalyzedthestructureandpredictedfunctionsoftheRproteinincomparisonwith13otherisolatesofSARS-CoVand6othercoronaviruses.TheentireORF(open-readingframe)encodesfortwomajorenzymeactivities,RNA-dependentRNApolymerase(RdRp)andproteinaseactivities.TheRpolyproteinunder-goesacomplexproteolyticprocesstoproduce15function-relatedpeptides.Ahydrophobicdomain(HOD)andahydrophilicdomain(HID)arenewlyidentifiedwithinNSP1.ThesubstitutionrateoftheRproteinisclosetotheaverageoftheSARS-CoVgenome.ThefunctionaldomainsinallNSPsoftheRproteingivedifferentphylogeneticresultsthatsuggesttheirdifferentmutationrateunderselectivepressure.ElevenhighlyconservedregionsinRdRpandtwelvecleavagesitesby3CLP(chymotrypsin-likeprotein)havebeenidentifiedaspotentialdrugtargets.Findingssuggestthatitispossibletoobtaininformationaboutthephy-logenyofSARS-CoV,aswellaspotentialtoolsfordrugdesign,genotypinganddiagnosticsofSARS.

简介：AbstractIn December of 2019, several cases of atypical pneumonia caused by an unknown agent were reported in Wuhan, the capital city of Hubei Province in China. In early January 2020, it was announced that these cases were caused by a novel coronavirus. The virus was later named Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), which causes a disease associated with atypical pneumonia termed Corona virus disease 2019 (COVID-19). Several respiratory viruses, including coronaviruses and influenza viruses tend to have prominent peaks of infection during colder seasons, especially in temperate regions. The cold temperatures, along with accompanying dry conditions can drive respiratory tract infections by assisting with viral transmission, weakening the human immune system, and increasing viral molecular stability. Though the topic of SARS-CoV-2 transmission and warm weather has been associated with misinformation campaigns, it is worth investigating since an informative answer may give an indication of the future behavior of SARS-CoV-2.