微生物抗药性及生态效应ppt课件

微生物抗药性及生态效应,1,微生物抗药机制基因水平转移环境中的抗性基因,2,Mycobacteriumtuberculosis,Enterococcusfaecalis,Streptococcuspneumoniae,E.coli,3,4,抗生素抗药性已经成为全球性的重大问题,Resistance1998-2004-%,%,Resistance1998-2004-%,StreptococcuspneumoniaeinUSA,5,6,S.aureus,Penicillin,1950s,Penicillin-resistant,S.aureus,EvolutionofDrugResistanceinS.aureus,7,EscherichiacoliinAustralia201011,KlebsiellapneumoniaeIndia200912,Kpneumoniae，EcoliIndia,Pakistan,andtheUK20108,CitrobacterfreundiiFrenchreturningfromIndia201010,屎肠球菌，鲍曼不动杆菌201010,超级细菌（Superbug）,NDM-1NewMetallo-Lactamase,NDM-1超级细菌在世界各地被发现。,8,我国发现NDM-1超级细菌,9,10,超级细菌对多种抗生素均有抗性,11,Tigecycline新批准的新型抗生素简介：Tigecycline(商品名为Tygacil；惠氏公司生产)2005年6月获得FDA批准，用于治疗细菌感染。这是一种称为glycylcyclines的新型抗生素第一个获得批准的产品。该类抗生素用于克服早期四环素类抗生素的耐药性。除了Tigecycline，在过去的40年里，只有两个新型抗生素上市，分别是辉瑞公司的linezolid(Zyvox)和Cubist公司的daptomycin(Cubicin)。多粘菌素Colistin(polymyxinE)是是混合物多聚氨基酸，主要针对革兰氏阴性杆菌。,12,13,超级细菌（Superbug）,PlasmidsChennai50kbto350kb；Haryana118kb(54%)or50kb(36%)；UKisolates80kbtogreaterthan500kb.MostblaNDM-1positiveplasmidswerereadilytransferableandpronetorearrangement,losingor(morerarely)gainingDNAontransfer.,质粒的可移动性和可塑性可能意味着在细菌中的广泛传播和不断变异。,14,加拿大魁北克省舍布鲁克大学附属医院的专家日前透露，从2003年年初至今，一种普通的肠道细菌梭状芽孢杆菌历经两年变异，已成为致命的“超级病菌”，它可引起65岁以上老年人和服用抗生素的病人产生严重痢疾，并最终致死。该病菌已使这家医院的100名病人死亡，如不采取紧急行动，这种“超级病菌”很可能会引发一场致命传染病的蔓延。,15,2010年9月5日，日本媒体近日披露，有46名患者在日本帝京大学医学部附属医院住院期间感染了几乎可以抵抗所有抗生素的超级细菌多重耐药鲍曼不动杆菌，其中9名患者的死亡因感染此种病菌所致。该医院曾刻意隐瞒感染事件，导致日本首次出现大规模不动杆菌感染，日本警方遂决定传唤该院医生，展开调查。经检测发现，患者感染的不动杆菌能抵抗多种抗生素，目前只有先锋霉素族的一部分抗生素对这一病菌有效。,16,-MRSAinfectionofhipscrew-VnMIC1mg/LVntreatment-d55,developedEnterobacterinfection-ciproaddedtoVnfor6weeks-d99,VnMIC=4mg/L-d159,Vndiscontinued-d174,readmittedtohospital,Vntreatment-VnMIC=4mg/L-d188,linezolid+rifampin-d198,infectioncleared-12surgeriesincludingremovalofinfectedbone,AcaseofresistantSaureusinpatient,Canada-Edmonton-Websteretal.,2007,17,细菌耐药机制,DrugImpermeability,DrugEfflux,Targetmodification,DrugInactivation,OnetypeofantibioticcanbeovercomebydifferentmechanismsDifferentclassesofantibioticscanhavethesamemechanismsofresistance,18,孔蛋白改变，细胞壁/膜通透性改变,万古霉素（vancomycin）：由一种链霉菌产生的、结构复杂的糖肽类抗生素，专一地抑制肽聚糖的生物合成。万古霉素通过抑制细菌的生长和繁殖来杀死细菌。这种药物通过干扰细菌细胞壁结构中的一种关键组分来干扰细胞壁的合成，抑制细胞壁中磷脂和多肽的生成，只是针对革兰氏阳性菌，对厌氧菌和革兰氏阴性细菌无效。万古霉素的药力较强，且有肾毒性，在其他抗生素对病菌无效时才会被使用，也就是所谓的最后一线药物。,19,Howdenetal.2006,VISAstrainshavethickercellwalls,20,-adsorptionofVnbycell-wallD-ala-D-alaresiduesincreased,diffusiondecreased-drugispreventedfromreachingitsactivesite,Vnsensitive,Vnresistant,VISAstrainshavethickercellwalls,21,TheCaseofCMRSA-1,MRSA,VISA(4mg/L),potentialhVISA(64)mef(specificmacrolideeffulxpump)lowlevelresistance(MIC4-8),Ermbmethylatingasingleadenineresidueofthe23SrRNA,Bussiereetal.Biochemistry1998,35,SensitivityTestof185StreptococcuspneumoniaeIsolates,Shenetal.2008,分离自北京儿童的Streptococcuspneumoniae对大环内酯抗生素的耐药情况,36,erm(B)-carryingelements,37,38,39,40,微生物抗药机制基因水平转移环境中的抗性基因,41,SusceptibleBacteria,EmergenceofAntimicrobialResistance,42,基因水平转移是微生物进化的重要动力,基因水平转移HorizontalGeneTransferThemovementofgeneticmaterialbetweentwoorganisms.Onceincorporateditisthenverticallyinherited.,43,DelwicheandPalmer(1996)-hypothesesofHGTs,1-Cyanobacteria-Proteobacteria2-ProteobacteriaRedandbrownalgae3-Proteobacteria-Proteobacteria4-Proteobacteria-Proteobacteria,44,Pan-genome泛基因组,Thepan-genomeincludesthe“coregenome”containinggenespresentinallstrains,a“dispensablegenome”containinggenespresentintwoormorestrains,andfinally“uniquegenes”specifictosinglestrains。,(Touchonetal.,2009).,Inmolecularbiologyapan-genomedescribesthefullcomplementofgenesinaspecies(typicallyappliedtobacteriaandarchaea,whichcanhavelargevariationingenecontentamongcloselyrelatedstrains).Itisasupersetofallthegenesinallthestrainsofaspecies。,45,基因水平转移的方式,Transformation-nakedDNA,shortpieces,commoninbacteriathattransform.Transductionphage,donor/recipientsharereceptors,closelyrelatedbacteria,DNA:amountinphagehead.Conjugation-plasmids/transposons,celltocellcontact,distantrelations,longDNA.,46,转化Transformation,CelldeathCollis,C.andHall,R.(1992)MolMicro6:2875;Recchia,G.andHall(1995)Microbiol141:3015,allcontainanattCsite(59-baseelement)-integrasetargettheICSisalwayscomplementarytotheCSofthecircularizedcassette;theyformimperfectinvertedrepeats,mostcontainasingleORFpromoterless,60,r,p,l,T,r,p,m,I,i,n,f,C,L,2,0,L,3,5,I,F,3,i,n,t,e,g,r,a,s,e,intI4orVchintIA,TheV.choleraesuperintegron,Mazel,Detal(1998)Science280:605;Heidelberg,Jetal.(2000)Nature406:477;Rowe-Magnusetal.(1999)ResMic150:641,61,微生物抗药机制基因水平转移环境中的抗性基因,62,spore-formingbacteria20JANUARY2006VOL311SCIENCE,自然环境是抗性基因的储存库,63,Antibiotic-resistanceprofilingof75clonalisolatescapableofsubsistingonantibiotics.,GautamDantas,etal.Science2008,64,GlycopeptideAntibioticResistanceGenesinGlycopeptide-ProducingOrganisms,MARSHALLetal.1998,65,Geneticexchangeamongbacterialspecies,Thisprocessdemonstratestheimportanceofbacterialreservoirsofresistance,includingbothpathogenicandnonpathogenicorganisms.Silbergeldetal.2008,66,WW,wastewatereffluent;RWD,downstreamriverwater;RWU,upstreamriverwater.,APPLIEDANDENVIRONMENTALMICROBIOLOGY,June2010,p.34443451,抗生素污染加速了耐药性的传播和蔓延,67,养殖业是抗生素重要的污染源,WorldHealthOrganizationestimateshalfoftotalamountofantimicrobialsproducedgloballyareusedinfoodanimals.InUS,70-80%ofallantimicrobialssoldareforlivestockand85%oflivestockantimicrobialuseisfornon-therapeuticfeedaddition.Diseasecontrolandgrowthpromotion,68,69,WasteTreatmentandAntimicrobials,Antimicrobialsarecomplexcompoundsthatresistbiologicaldecompositionwastetreatment.Anaerobicdigestiondestroyedonly59%ofoxytetracyclineinmanuresin64days.Methaneproductionwasreducedfrom20-80%whenmanurescontainantibiotics,dependingontheconcentrationofantibioticsinthemanures.*Compostingdestroyed95%ofoxytetracylineinmanureswithinfirstweek.Antibioticsfoundintactintreatedsewagesludgewereciprofloxacin,doxycycline,norfloxacin,ofloxacin,andtriclosan.*,*J.Fick,et.al.,AntivialOsetimiverisnotRemovedorDegradedinNormalSewageTreatment,2007*O.A.Arikan,et.al.,FateandEffectofOxytetracyclineduringAnaerobicDigestionofManurefromTherapeuticallyTreatedCalves.,2006*E.Z.Harrison,et.al.,OrganicChemicalsinSewageSludges,2006*O.A.Arikan,et.al,CompostingRapidlyReducesLevelsofExtractableOxytetracyclineinManurefromTherapeuticallyTreatedBeefCalves,2005.,70,WasteTreatmentandAntimicrobials,Antimicrobialsarecomplexcompoundsthatresistbiologicaldecompositionwastetreatment.Anaerobicdigestiondestroyedonly59%ofoxytetracyclineinmanuresin64days.Methaneproductionwasreducedfrom20-80%whenmanurescontainantibiotics,dependingontheconcentrationofantibioticsinthemanures.*Compostingdestroyed95%ofoxytetracylineinmanureswithinfirstweek.Also,levelsofoxytetracyclineresistantbacteriawere10-foldlower.*Antibioticsfoundintactintreatedsewagesludgewereciprofloxacin,doxycycline,norfloxacin,ofloxacin,andtriclosan.*,*J.Fick,et.al.,AntivialOsetimiverisnotRemovedorDegradedinNormalSewageTreatment,2007*O.A.Arikan,et.al.,FateandEffectofOxytetracyclineduringAnaerobicDigestionofManurefromTherapeuticallyTreatedCalves.,2006*E.Z.Harrison,et.al.,OrganicChemicalsinSewageSludges,2006*O.A.Arikan,et.al,CompostingRapidlyReducesLevelsofExtractableOxytetracyclineinManurefromTherapeuticallyTreatedBeefCalves,2005.,71,TetracyclineResistomeoftheOrganicPigGut,Katarzynaetal.AEM,2009,Of9,000bacterialartificialchromosomeclonesanalyzed,10wereidentifiedascarryingtheknowntet(C),tet(W),andtet(40)genes,aswellasnovelgenesencodingresistancetothetetracyclinesminocyclineanddoxycycline.,72,TetracyclinesandTetracyclineResistanceinAgriculturalSoils,Schmittetal.2006,73,象山港网箱养殖区沉积物中细菌生态分布,采样地图,Figure1Diversityoftheisolates,74,象山港网箱养殖区沉积物中细菌抗药情况严重,Figure2Antibiotics-resistanceoftheisolates,-proteobacteria-proteobacteriaActinobacteriaFirmicutesBacteroidetes,75,分离到的耐药细菌所含质粒多样,1D15000DNALadder2,9Comamonassp.CNB-13,8GemoneDNA4Pseudoalteromonashaloplanktis5Exiguobacteriumsp.6Pseudoclavibacterhelvolus7Exiguobacteriumaestuarii10Roseobactergallaeciensis11Roseobactergallaeciensis12Brevundimonasnasdae13Phaeobactercaeruleus14Labrenziaaggregata,76,PlanococcuscitreusBacteria;Firmicutes;Bacillales;Planococcaceae;Planococcus,Planococcuscitreus,bacillussubtilisMIC提高了8倍,EnterococcusFaecom,结合转移,77,plasmidsisolatedfromtheabovestrainswereusedfornaturaltransformationoftheoralcariogenicpathogenStreptococcusmutansunderlaboratoryconditions.ThetetS/MgenewassuccessfullytransferredtoS.mutansUA159atfrequenciesrangingfrom1.910-7to2.810-5,4.710-7to2.310-6,and3.810-7to2.110-6transformantsperrecipientcellusingCZ-T4,CZ-T8andRMKT14plasmidextracts,respectively.,食品中也存在抗性基因，并可通过基因水平转移传递给口腔致病菌,FEMSMicrobiolLett254(2006)226231,78,Mostoftheresistancegenesweidentifiedusingculture-independentsamplinghavenotbeenpreviouslyidentifiedandareevolutionarilydistantfromknownresistancegenes.Nearlyhalfoft