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1、病毒主题医学知识,病毒主题医学知识,病毒（virus）,是一类非细胞型微生物主要特点体积非常微小，需在电子显微镜观察结构简单，无细胞结构，只含一种类型核酸（DNA或RNA）严格的细胞内寄生，只能在活细胞中增殖对抗生素不敏感，但对干扰素敏感,2,病毒主题医学知识,病毒（virus）是一类非细胞型微生物2病毒主题医学知识,3,病毒主题医学知识,特性病毒细菌真菌滤菌器+ -结 构非细胞原核细胞真核细,形态,大小: （纳米，nm）The largest : （300X100 nm, 痘病毒）The smallest: 10 nm 形态：球形或杆状，也有复合状,4,病毒主题医学知识,形态大小: （纳米，
2、nm）4病毒主题医学知识,5,病毒主题医学知识,5病毒主题医学知识,6,病毒主题医学知识,6病毒主题医学知识,7,病毒主题医学知识,7病毒主题医学知识,8,病毒主题医学知识,8病毒主题医学知识,HIV,9,病毒主题医学知识,HIV9病毒主题医学知识,Bacteriophage T4,10,病毒主题医学知识,Bacteriophage T410病毒主题医学知识,结构,基本结构核心 Viral core 衣壳 Capsid 辅助结构包膜 Envelope其他病毒体 Virion 核衣壳 Nucleocapsid,11,病毒主题医学知识,结构基本结构11病毒主题医学知识,核心,成分：DNA 或 RN
3、A,病毒复制功能 决定病毒的各种特性 具有感染性 编码病毒结构蛋白和功能蛋白,病毒核酸的特性：具有感染性将提纯的病毒核酸（如肠道病毒RNA）如加至活细胞可以启动病毒复制从而感染细胞。从感染细胞中可以获得子代完整病毒颗粒。,12,病毒主题医学知识,核心成分：DNA 或 RNA 病毒复制病,病毒衣壳,成分：蛋白质由壳粒组成。壳粒是衣壳的形态学亚单位，多肽分子是衣壳的化学亚单位功能保护病毒核酸参与感染过程具有抗原性,13,病毒主题医学知识,病毒衣壳成分：蛋白质13病毒主题医学知识,按壳粒排列方式的对称型,螺旋对称型（helical symmetry）20面体立体对称型（icosahedral sym
4、metry）复合对称型（complex symmetry）,14,病毒主题医学知识,按壳粒排列方式的对称型螺旋对称型（helical symme,病毒包膜,是包绕在病毒核衣壳外面的双层膜主要成分是蛋白质、多糖及脂类，常以糖蛋白或脂蛋白形式存在蛋白质是由病毒基因编码，多糖、脂类来自宿主细胞膜、核膜或空泡膜,15,病毒主题医学知识,病毒包膜是包绕在病毒核衣壳外面的双层膜15病毒主题医学知识,包膜特性与功能,有包膜的病毒以“出芽” 方式释放有包膜病毒对脂溶剂（如乙醚、氯仿）敏感，能灭活病毒，乙醚常用于鉴定病毒有无包膜功能维护病毒体结构的完整性具有与宿主细胞膜亲和及融合的性能具有病毒抗原的特异性,16
5、,病毒主题医学知识,包膜特性与功能有包膜的病毒以“出芽” 方式释放16病毒主题医,病毒复制,复制周期吸附、穿入、脱壳、生物合成、装配与释放病毒本身没有独立的蛋白合成酶系统，必须借助宿主细胞的蛋白合成体系合成病毒蛋白病毒复制周期时间因病毒而异腺病毒25h小RNA病毒6-8h流感病毒15-30h,17,病毒主题医学知识,病毒复制复制周期17病毒主题医学知识,dsDNA病毒复制示意图,18,病毒主题医学知识,dsDNA病毒复制示意图18病毒主题医学知识,+ssRNA病毒复制示意图,19,病毒主题医学知识,+ssRNA病毒复制示意图19病毒主题医学知识,+ssRNA逆转录病毒复制示意图,20,病毒主题
6、医学知识,+ssRNA逆转录病毒复制示意图20病毒主题医学知识,吸附,通过病毒体表面的配体蛋白与易感细胞表面特异性受体相结合吸附过程可在几分钟到几十分钟内完成,21,病毒主题医学知识,吸附通过病毒体表面的配体蛋白与易感细胞表面特异性受体相结合2,穿入,吞饮：无包膜病毒被宿主细胞吞噬，进入胞内融合：有包膜病毒的包膜与宿主细胞膜直接融合，病毒核衣壳进入细胞直接穿入：,22,病毒主题医学知识,穿入吞饮：无包膜病毒被宿主细胞吞噬，进入胞内22病毒主题医学,脱壳,在细胞溶酶体酶的作用下，脱去衣壳蛋白释放病毒核酸,23,病毒主题医学知识,脱壳在细胞溶酶体酶的作用下，脱去衣壳蛋白释放病毒核酸23病毒,生物合
7、成,早期蛋白合成阶段转录、翻译而产生病毒生物合成中必需的酶类及某些抑制或阻断细胞核酸和蛋白质合成的非结构蛋白晚期蛋白合成阶段复制病毒核酸，转录、翻译而产生病毒的结构蛋白隐蔽期生物合成阶段用电镜方法在细胞查不到完整病毒，用血清学方法也测不到病毒抗原各病毒隐蔽期长短不一，,24,病毒主题医学知识,生物合成早期蛋白合成阶段24病毒主题医学知识,装配,无包膜病毒先形成空心衣壳，核酸从衣壳裂隙进入形成核衣壳有包膜病毒核衣壳与细胞的膜系统（浆膜或核膜）结合形成包膜包膜的蛋白质（包括糖蛋白）是由病毒基因组编码，故具有病毒的特异性和抗原性装配的部位除痘病毒外，DNA病毒均在细胞核内装配RNA病毒与痘病毒则在细
8、胞浆内装配,25,病毒主题医学知识,装配无包膜病毒25病毒主题医学知识,释放,无包膜病毒均以破胞方式释放有包膜的病毒以出芽方式释放到细胞外通常细胞不死亡，仍能继续分裂增殖,26,病毒主题医学知识,释放无包膜病毒26病毒主题医学知识,病毒增殖异常,顿挫感染缺陷病毒干扰现象,形成包涵体 某些病毒在宿主细胞内增殖，其细胞质或细胞核内会出现一种光学显微镜下可见的斑块结构，称包涵体。它是病毒在细胞内增殖的场所,27,病毒主题医学知识,病毒增殖异常顿挫感染形成包涵体27病毒主题医学知识,顿挫感染,原因宿主细胞不能提供病毒复制所需的酶、能量或成份病毒虽被复制，但不能装配释放,28,病毒主题医学知识,顿挫感染
9、原因28病毒主题医学知识,缺陷病毒,因病毒基因组不完整或有点突变而不能进行正常复制的病毒，当与辅助病毒共同培养，如能为其提供缺乏的物质，则缺陷病毒也能培殖腺病毒相关病毒（缺陷病毒）与腺病毒（辅助病毒）丁型肝炎病毒（缺陷病毒）与乙型肝炎病毒（辅助病毒）,29,病毒主题医学知识,缺陷病毒因病毒基因组不完整或有点突变而不能进行正常复制的病毒,干扰现象,两种病毒感染同一细胞时，一种病毒会干扰另一病毒复制原因与干扰素（IFN）产生有关病毒改变了宿 主细胞代谢途径意义联合使用疫苗,30,病毒主题医学知识,干扰现象两种病毒感染同一细胞时，一种病毒会干扰另一病毒复制3,理化因素对病毒的影响,灭活病毒受理化因素
10、作用后，失去感染性灭活病毒仍保留某些特性，如抗原性、红细胞吸附、细胞融合等,31,病毒主题医学知识,理化因素对病毒的影响灭活31病毒主题医学知识,物理因素,温度病毒耐冷不耐热。-70C长期保存。冻融可杀死病毒60C 30 min或100C数秒即可杀死病毒，但HBV需100C 10 min方可杀死有包膜病毒比无包膜病毒对热更敏感pH多数在pH 59范围稳定肠道病毒耐酸（pH 35）射线 X线、射线可将病毒核酸致死性断裂，从而杀死病毒,32,病毒主题医学知识,物理因素温度32病毒主题医学知识,化学因素,脂溶剂：乙醚、氯仿、去氧胆酸盐有包膜病毒敏感。无包膜病毒无作用（如肠道病毒）消毒剂：过氧乙酸、甲
11、醛、戊二醛、卤素杀死大多数病毒，但病毒对消毒剂抵抗力比细菌强，尤其是无包膜病毒常用甲醛来制备灭活疫苗其他抗生素：对病毒无作用中草药：有一定作用,33,病毒主题医学知识,化学因素脂溶剂：乙醚、氯仿、去氧胆酸盐33病毒主题医学知识,病毒的变异现象感染性变异（毒力变异）条件致死株：减毒活疫苗（脊灰）抗原性变异耐药性变异机制基因突变基因重组,34,病毒主题医学知识,病毒的变异现象34病毒主题医学知识,病毒与其他微生物的比较,35,病毒主题医学知识,病毒与其他微生物的比较特 性病 毒细,小结,病毒的概念病毒的结构与化学组成病毒的复制理化因素对病毒的影响,36,病毒主题医学知识,小结病毒的概念36病毒主题
12、医学知识,14. Techniques used to Study Viruses,Living hosts. Man. Pasteur used rabbits to study and develop rabies vaccines.Walter Reed developed a mouse model of yellow fever.Transgenic animals, particularly mice invaluable,Embryonated eggs used to propagate viruses in the early decades of this century.
13、 Effective for the isolation & culture of many viruses e.g.influenza.,37,病毒主题医学知识,14. Techniques used to Study V,15. Cell Culture methods,Whole organ cultures, progressed to methods involving individual cells; primary cell cultures which can be maintained for a short period in culture); or immortali
14、zed cell lines, which grow in culture indefinitely. Viruses can be grown in them.,38,病毒主题医学知识,15. Cell Culture methodsWhole,16. Quantifying viruses.,The plaque assay - dilutions of the virus are used to infect a cultured cell monolayer, covered with agar to restrict virus diffusion virus. Results in
15、 localized cell killing & the appearance of plaques. The number of plaques directly relates to numbers of infectious virus particles applied to the plate.Many other approaches.,39,病毒主题医学知识,16. Quantifying viruses. The p,17. Other practical approaches,SerologyStructural studies, purification, EM, X-r
16、ay.Biochemical, electrophoresisGeneticMolecular biology, nucleic acid sequencing.,40,病毒主题医学知识,17. Other practical approaches,Replication 14. Exit,Some viruses cause cell lysis. Unenveloped.Other viruses bud through a cell membrane, aquiring an envelope. Can be the plasma membrane e.g. HIV-1. Can be
17、nuclear or golgi membranes.,41,病毒主题医学知识,Replication 14. ExitSome virus,2. Big fleas have little fleas upon their backs to bite them; and little fleas have lesser fleas and so ad infinitum.,Viruses consist of proteins, nucleic acids and sometimes lipids. However there are other infectious agents stud
18、ied by virologists.Viroids small (200-400nt), circular RNAs, possessing no capsid or envelope. Associated with certain plant diseases. They are infectious obligate intracellular parasites. Virusoids are satellite, viroid-like RNAs, larger than viroids (approximately 1000nt). Depend on the presence o
19、f virus replication for multiplication (hence satellite), they are packaged into virus capsids as passengers. Present in animals and plants. Associated with disease.,42,病毒主题医学知识,2. Big fleas have little fleas,3. Prions,Prions. “pree ons” believed to consist of a single type of protein with no nuclei
20、c acid component. The prion protein & the gene which encodes it are also found in normal uninfected cells. These agents are associated with infectious and inherited diseases, such as Creutzfeldt-Jakob disease in humans, scrapie in sheep & bovine spongiform encephalopathy (BSE) in cattle.,43,病毒主题医学知识
21、,3. PrionsPrions. “pree ons” be,ve RNA,-ve RNA,pol,Proteins,-ve RNA,+ve RNA,RI,44,病毒主题医学知识,ve RNA-ve RNApolProteins-ve R,Genomes 14Ambisense Genomes,Some ssRNA viruses are ambisense, since they are part (-)sense & part (+)sense: 5 ends are +ve, 3 ends are ve.,45,病毒主题医学知识,Genomes 14Ambisense Genomes,
22、Genomes 15. Segmented/multipartite.,Can be confusing, ss or ds. Segmented genomes have 2 or more pieces of nucleic acid packaged in the same particle e.g orthomxyo viruses.(7/8 in flu, packaging problem)Multipartite genomes are segmented but each segment is contained in a different particle e.g bipa
23、rtite Comoviruses, both particles infect cell.,segmented,Multipartite, DNA as well,46,病毒主题医学知识,Genomes 15. Segmented/multipar,Genomes 16. DNA.,Small genomes e.g.bacteriophage M136.4 Kb ss circular DNA. 10 genesBy convention genome is +strand.90% genome is coding.,+,RF,strand transcribed to make mRNA
24、and then proteins,+ve strand nickedextended in rolling circle,+,Specifically cleavedAnd recircularised,47,病毒主题医学知识,Genomes 16. DNA.Small genomes,Genomes 17. Phage l.,Linear ds DNA about 50Kb.Cohesive ss DNA termini 12 nucleotides long, the cos site. Facilitates circularisation and replication cycle.
25、 Concatemers formed and then resolved to reconstitute genome. TCCAGCGGCGGG AGGTCGCCGCCC3,important sequences at the end of the linear virus genomes are a very common feature.,48,病毒主题医学知识,Genomes 17. Phage l. Linear ds,Genomes 18. Phage T4,Large linear ds DNA 160KbGenome exhibits terminal redundancy.
26、 Another common feature of linear genomes. A B C D E A B C D E A B C D E A B C D EA B C D E A B C D E A B C D E A B C D EDuring replication concatemers are formed. Endonuclease recognises and cuts at this terminal redundancy, which is then regenerated.,49,病毒主题医学知识,Genomes 18. Phage T4Large line,Geno
27、mes 19.,Transcriptional control in prokaryotes is sophisticated. That said extensive use of polycistronic mRNAs is made.Genomes are denseM13 10 genes, 10 transcripts 90% usageGenomes of eukaryotes are denserPolyoma, ds DNA circular 5kb genomeSix genes, both strands, overlapping.,50,病毒主题医学知识,Genomes
28、19. Transcriptional co,Genomes 20. Adeno viruses,Larger 30-38kb linear ds genomes infecting eukaryotes. Code for about 30-40 proteinsThese viruses are genetically very similar to the host cells which they infect.Terminal sequences are inverted repeatsComplicated functional structures can form at the
29、se points,TGTGTGCACACAACACACGTGTGT,51,病毒主题医学知识,Genomes 20. Adeno virusesLarge,52,病毒主题医学知识,52病毒主题医学知识,Viral Replication.,Attachment and penetrationUncoating, nucleic acid and protein synthesisAssembly and exit.,53,病毒主题医学知识,Viral Replication. Attachment,Replication 2.,Particle/infectivity ratio can be
30、 low. Sometimes only 1 in 1000 virions are infectious.It makes the study of replication difficult because most infections are abortive. Study of synchronously infected cells is useful.,54,病毒主题医学知识,Replication 2.Particle/infecti,病毒主题医学知识培训课件,Replication 4.,Maturation phase:viral material accumulates
31、in cell or surrounding medium. Cells infected with lytic viruses become metabolically disordered and die, viral production ceases. Titres slowly drop. Cells infected with non lytic viruses can continue to produce viral particles indefinitely.,56,病毒主题医学知识,Replication 4. Maturation phas,Replication 5.
32、,Reproductive cycle less than an hour with many bacteriophage, 6-8hrs in picornaviridae and more than 40 hrs in herpesviridae. Cells infected with polio virus can yield more than 100000 copies of virus per infected cell.,57,病毒主题医学知识,Replication 5. Reproductive cy,Replication 6.,Infection may be: pro
33、ductive, i.e. entry into permissive cells followed by virion formation. abortive, i.e. entry into a non permissive cell which does not result in virion formation; there can be many reasons for non-permissiveness e.g. no receptor.restringent or restrictive, cell is transiently permissive and a few vi
34、rus are produced. Virus production stops but the genome remains in the cell, examples include Epstein Barr Virus and herpes simplex virus. This kind of infection may still have serious consequences e.g cell transformation and cancer.,58,病毒主题医学知识,Replication 6.Infection may be,History of Virology,Vir
35、uses are probably as old as life on earth. Ancients were aware of viral diseases,Perhaps the first written record of a virus infection consists of a heiroglyph from Memphis, drawn in approximately 1400BC, which depicts a temple priest called Siptah showing typical clinical signs of paralytic poliomy
36、elitis,59,病毒主题医学知识,History of VirologyViruses are,Smallpox, endemic in China by 1000BC. Recognizing that survivors of smallpox outbreaks were protected from subsequent infection, the practice of variolation developed. Involved inhalation of dried crusts from smallpox lesions, or in later modificatio
37、ns, inoculation of the pus from a lesion into a scratch on the forearm. Practice survived until this century.,60,病毒主题医学知识,Smallpox, endemic in China by,On May 14, 1796, Edward Jenner used cowpox-infected material obtained from the hand of Sarah Nemes, a milkmaid from Berkley in Gloucestershire to va
38、ccinate 8 year old James Phipps. On July 1, 1796, Jenner challenged the boy by deliberately inoculating him with material from a real case of smallpox ! He did not become infected!,61,病毒主题医学知识,On May 14, 1796, Edward Jenne,In 1892, Dmitri Iwanowski, a Russian botanist, showed that extracts from dise
39、ased tobacco plants could transmit disease to other plants after passage through ceramic filters fine enough to retain the smallest known bacteria. Generally recognised as the beginning of Virology In 1898, Martinus Beijerinick confirmed & extended Iwanowskis results on tobacco mosaic virus & was th
40、e first to develop the modern idea of the virus, which he referred to as contagium vivum fluidum (soluble living germ),62,病毒主题医学知识,In 1892, Dmitri Iwanowski, a R,Compiled from Virus Taxonomy, the Sixth Report of the International Committee on Taxonomy of Viruses (ICTV) 1995. N.B: you can search this
41、 document using the Find command of the browser (Edit Menu)63U-11 virus, Bunyaviridae75V-2374 virus, Bunyaviridae75V-2621 virus, Bunyaviridae78V-2441 virus, Bunyaviridae Abadina virus, ReoviridaeAbelson murine leukemia virus, RetroviridaeAbras virus, BunyaviridaeAbraxas grossulariata cypovirus 8, Re
42、oviridaeAbraxas grossulariata NPV, BaculoviridaeAbsettarov virus, FlaviviridaeAbu Hammad virus, BunyaviridaeAbu Mina virus, BunyaviridaeAbutilon mosaic virus, GeminiviridaeAcado virus, ReoviridaeAcalypha yellow mosaic virus, GeminiviridaeAcantholyda erythrocephala NPV, BaculoviridaeAcara virus, Buny
43、aviridaeAcciptrid herpesvirus 1, HerpesviridaeAchaea janata NPV, BaculoviridaeAcherontia atropas virus, TetraviridaeAcheta domestica densovirus, ParvoviridaeAcholeplasma phage Oc1r, InoviridaeAcholeplasma phage 10tur, InoviridaeAcholeplasma phage L2, PlasmaviridaeAcholeplasma phage L51, InoviridaeAc
44、holeplasma phage M1, PlasmaviridaeAcholeplasma phage MV-L1, InovindaeAcholeplasma phage MVG51, InoviridaeAcholeplasma phage 01, PlasmaviridaeAcholeplasma phage vl, PlasmaviridaeAcholeplasma phage v2, PlasmaviridaeAcholeplasma phage v4, PlasmaviridaeAcholeplasma phage v5, PlasmaviridaeAcholeplasma ph
45、age v7, PlasmaviridaeAchroia grisella NPV, BaculoviridaeAcidalia carticcaria NPV, BaculoviridaeAcleris gloverana NPV, BaculoviridaeAcleris variana NPV, BaculoviridaeAcrobasis zelleri entomopoxvirus, PoxviridaeAcronicta aceris NPV, BaculoviridaeActebia fennica NPV, BaculoviridaeActias selene Cypoviru
46、s 4, ReoviridaeActias selene NPV, BaculoviridaeActinomycetes phage 108/016, MyoviridaeActinomycetes phage 119, SiphoviridaeActinomycetes phage A1-Dat, SiphoviridaeActinomycetes phage Bir, SiphoviridaeActinomycetes phage f115-A, SiphoviridaeActinomycetes phage f150A, SiphoviridaeActinomycetes phage f
47、31C, Siphoviridae,Actinomycetes phage M1, SiphoviridaeActinomycetes phage MSP8, SiphoviridaeActinomycetes phage P-a-1, SiphoviridaeActinomycetes phage R1, SiphoviridaeActinomycetes phage R2, SiphoviridaeActinomycetes phage SK1, MyoviridaeActinomycetes phage SV2, SiphoviridaeActinomycetes phage VP5,
48、SiphoviridaeAdelaide River virus, RhabdoviridaeAdeno-associated virus 1, ParvoviridaeAdeno-associated virus 2, ParvoviridaeAdeno-associated virus 3, ParvoviridaeAdeno-associated virus 4, ParvoviridaeAdeno-associated virus 5, ParvoviridaeAdisura atkinsoni NPV, BaculoviridaeAdoxophyes orana NPV, Bacul
49、oviridaeAedes aegypti densovirus, ParvoviridaeAedes aegypti entomopoxvirus, PoxviridaeAedes aegypti NPV, BaculoviridaeAedes albopictus densovirus, ParvoviridaeAedes annandalei NPV, BaculoviridaeAedes atropalpus NPV, BaculoviridaeAedes epactius NPV, BaculoviridaeAedes nigromaculis NPV, BaculoviridaeA
50、edes pseudoscutellaris densovirus, ParvoviridaeAedes scutellaris NPV, BaculoviridaeAedes sollicitans NPV, BaculoviridaeAedes taeniorhynchus NPV, BaculoviridaeAedes tormentor NPV, BaculoviridaeAedes triseriatus NPV, BaculoviridaeAedia leucomelas NPV, BaculoviridaeAeromonas phage 29, MyoviridaeAeromon
