呼肠病毒与细胞凋亡

从SARS患者生前咽拭子和尸检肺组织分离病毒阳性的培养细胞超薄切片中,在电子显微镜下不仅发现了呼肠病毒,而且还查见具有超微结构形态特征的凋亡细胞。凋亡早期的感染细胞,其细胞核染色质浓缩、边集显著,胞浆内尚可查见病毒包涵体的残迹。多数凋亡细胞胞浆内未查见呼肠病毒及其包涵体。在早期,凋亡细胞胞核形状变化不大,核染色质呈不规则的边集、浓缩,或呈平台状。继而,凋亡细胞胞核形状高度不规则,染色质浓缩、致密,可见核孔残迹,但细胞轮廓尚保存完好,外周可查见含核质凋亡小体。到后期,凋亡细胞高度皱缩．核固缩并碎裂成细小的片块或颗粒,胞质稀少,胞体呈芽球状,细胞膜外围可查见大小不等、形状多样的含核质凋亡小体。此研究结果提示：与SARS相关呼肠病毒所诱导的细胞凋亡有可能在SARS的发病机制中起重要作用。     

动物病毒疫苗研制及其产品的电镜检测

应用电镜技术检查了某些原代和传代细胞，并且跟踪检查了疱疹病毒、痘病毒、腺病毒、细小病毒、冠病毒、RNA反录病毒、呼肠病毒、动脉炎病毒、小RNA病毒、弹状病毒、杯状病毒、黄病毒、双RNA病毒、正粘病毒、副粘病毒等某些成员的驯化、培育及筛选的目的毒。结果发现了部分培养细胞有支原体污染，有的还有细菌污染；同时在某些培养的原毒、基础种毒、生产种毒及其产品中也发现了一些病毒的污染和联苗中互相感染现象。如此证明了电镜技术能够及时发现问题，及时解决问题，使之科研少走弯路，提高产品质量。     

感染神经元内流行性出血热病毒抗原的免疫金标记

在所有的RNA病毒中,布尼安病毒科(Family Bunyaviridae)的病毒是唯一的具有在感染细胞高尔基复合体及其邻近光面囊泡内芽生成熟的形态发生学特征。我们曾应用常规超薄切片电镜技术,在我国两种出血热(流行性出血热,EHF和新疆出血热,XHF)病毒感染的新生乳鼠大脑海马回神经元内观察到病毒在增生、扩张的高尔基扁囊和囊泡内芽生成熟的图象。本研究应用低温包埋免疫金标记技术进行感染细胞内EHF病毒抗原分子的亚细胞定位,以求阐明高尔基复合体异常增生与病毒抗原分布的关系。     

脊髓灰质炎病毒免疫电镜形态发生学研究

在感染细胞内,脊髓灰质炎病毒进行复制是否需要核的参与至今仍存在争议。而有关该病毒形态发生的系统报道仍不多见。本文使用已建立的低温包埋金标记技术结合常规包埋技术对脊髓灰质炎病毒感染Hep-2细胞后的形态发生学进行了定位研究,探讨了该病毒与感染细胞的相互作用关系,细胞核在病毒复制中的作用以及病毒诱导空泡、核突出和复制复合体之间的关系。感染后4到12小时的细胞中,观察到大量核突出物形成及其由细胞核到细胞质的迁移动态。感染初期,核     

不同血清浓度培养条件对博尔纳病病毒感染滴度的影响比较

目的：探讨不同血清浓度对博尔纳病病毒（BDV）感染少突胶质细胞（OL细胞）感染力的影响,寻找BDV病毒感染细胞最适宜的血清浓度。方法：接种BDV的OL细胞（OL/BDV）于培养皿中,培养24小时后,通过反复冻融、超声破碎的方法获取BDV病毒液,用于感染正常的OL细胞。在保证除血清浓度因素不同而其余因素均相同的条件下,用DMEM、2%FCS、5%FCS、10%FCS四种不同浓度的血清培养基在96孔板中培养BDV新感染的OL细胞,测定病毒感染滴度,比较各组之间是否有统计学差异。结果：DMEM、5%FCS和10%FCS组所测得的BDV病毒滴度两组之间无统计学差异,2%FCS组所测得的病毒滴度和DMEM、5%FCS、0%FCS组之间有明显统计学差异,2%FCS组病毒滴度明显高于其它两组。结论：不同浓度的血清浓度对Boma病毒对细胞的感染力有影响,2%FCS的培养条件可能更适合于BDV感染OL细胞实验。     

猴艾滋病病毒(SRV—1)形态发生学观察

本文报导了猴艾滋病D型逆转录病毒(SRV—1)接种实验猴前后,有相同形态学特征,描述了培养细胞内外,有五种形态的病毒颗粒;(1)偏心位置核心病毒颗粒;(2)锥形或棒形核心病毒颗粒;(3)等电子密度圆形病毒颗粒;(4)圆环形核心病毒颗粒;(5)带尾形病毒颗粒。同时描述了病毒出芽释放连续过程和细胞内A型颗粒形态。带尾形病毒颗粒,以前未见有报导。     

感染番茄斑萎病毒和番茄环纹斑点病毒的植物叶片细胞病理变化观察

应用透射电子显微镜观察比较了番茄斑萎病毒（ TSWV）和番茄环纹斑点病毒（ TZSV）感病植物叶片的细胞超微结构,发现两种病毒在病变细胞内的形态及细胞病理变化特征相似,但病毒含量及细胞受损程度存在明显差异。与TSWV相比,TZSV侵染细胞的病变程度更为严重,叶绿体产生大的囊泡,片层结构被破坏,线粒体也发生囊泡化,而TSWV侵染的细胞内线粒体保存相对完好。高压冷冻－冷冻置换制备的样品细胞病理结构与常规化学固定的相似,但对膜结构的保存更加良好。该结果从细胞病理学上证明了TZSV在田间对农作物造成的危害更加严重。     

三种动物慢病毒形态及形态发生的比较研究

用电镜检查确定了不同感染时间的马传染性贫血病毒 (EIAV)、维斯纳 梅迪病毒 (MVV)和山羊关节炎脑炎病毒 (CAEV)的形态和形态发生。成熟的病毒粒子为电子致密的球形 ,有囊膜 ,直径约 10 0nm ,含有 1个 (有时有 2个或 2个以上 )的核芯。维斯纳 梅迪病毒和山羊关节炎脑炎病毒的核芯为球形 ,马传染性贫血病毒为杆形或锥形。在大部分病毒里能清晰地看到核芯壳 ,成熟的病毒从感染细胞的胞浆膜上出芽 ,在胞浆中还见到了花环状粒子和多板层结构     

细胞内病毒抗原定位的低温包埋及胶体金标记

本文报告使用低温包埋剂Lowicryl K4M对感染细胞进行低温包埋和制作超薄切片,用包被SPA或IgG的胶体金作为第二抗体,采用间接法对两种RNA病毒(脊髓灰质炎病毒和轮状病毒)和两种DNA病毒(单纯疱疹Ⅰ型病毒及SV_(40))在感染细胞中作了定位标记实验。将结果同常规环氧树脂Epon 812包埋的感染细胞包埋前穿透标记和包埋后超薄切片标记进行比较。常规包埋法对细胞超微结构保存良好,但由于高温聚合对病毒抗原活性破坏较大,不能获得理想的金标记结果。使用皂角甙(Saponin)改变细胞膜进行包埋前穿透标记,虽能获得特异性标记,但皂角甙对细胞膜结构破坏严重,不利于进行病毒与细胞相互作用关系的研究,且作用条件及时间不易掌握。以上缺点可通过使用低温包埋法得到解决。     

三种动物慢病毒形态及形态性的比较研究

用电镜检查确定了不同感染时间的马传染性贫血病毒（EIAV）、维斯纳-梅迪病毒（MVV）和山羊关节炎脑炎病毒（CAEV）的形态和形态发生。成熟的病毒粒子为电子致密的球形，有囊膜，直径约100nm，含有1个（有时有2个或2个以上）的核芯。维斯纳-梅迪病毒和山羊关节炎脑炎病毒的核芯为球形，马传染性贫血病毒为杆形或锥形。在大部分病毒里能清晰地看到核芯壳，成熟的病毒从感染细胞的胞浆膜上出芽，在胞浆中还见到了花环状粒子和多板层结构。     

Budding process and maturation of human immunodeficiency virus examined by means of pre- and post-embedding immunocolloidal gold electron microscopy

The techniques of pre- and post-embedding immunocolloidal gold electron microscopy were tested for their usefulness for analyzing the morphogenesis of human immunodeficiency virus (HIV) strain LAV employing U-937 cells persistently infected with the virus (U-937/LAV). By both techniques the following results were obtained. (1) Budding process was restricted to a localized area at the membrane adjacent to the Golgi apparatus. (2) The distribution of viral envelope antigens was restricted to the electron-dense crescent structures on the cell surface. (3) Virions released from the cells could be classified mainly into 2 categories: virions with doughnut-shaped cores or with conical cores at the center of the particles. And p24 proteins localized on the thick electron-dense cores of both types. These findings support the idea that pre- and post-embedding immunogold electron microscopy is very useful for studying the morphogenesis of viruses.     

水生动物病毒的电镜和荧光显微镜观察

应用透射电镜和荧光显微镜对三种水生动物病毒粒子形态、病毒感染细胞的超微结构变化及亚细胞定位进行比较和研究。负染电镜观察显示:鳜鱼弹状病毒(SCRV)粒子呈典型的子弹状形态,长约76～118 nm,直径约29～52 nm;鲈鱼呼肠孤病毒(LJRV)粒子呈正二十面体结构,具有双层衣壳,直径约70～80 nm;蛙虹彩病毒(RGV)粒子呈正二十面体结构,具有囊膜,直径大小约150 nm。超薄切片观察表明,宿主细胞的基本结构遭到不同程度的破坏,成熟病毒粒子分布在胞质中或以出芽的方式释放到胞外,且RGV增殖后在胞质中聚集形成晶格状结构。免疫荧光观察进一步显示,RGV感染细胞后能产生多个直径大小不一的包涵体。本研究结果有助于了解和认识水生动物病毒的超微形态特征、复制过程及致病机理。     

百合病毒病原的检测诊断

应用电镜和ELISA检测查明浙江丽水和杭州栽培的百合受到黄瓜花叶病毒(CMV)及多种线状病毒侵染。寄主反应测定显示侵染百合的植物病毒寄主范围都很窄,不能通过汁液摩擦接种侵染昆诺藜、苋色藜、普通烟、心叶烟等6科12种指示植物。在64个ELISA检测样品中有26个受到CMV侵染,侵染率达40．6％;在75个负染色检测样品中有58个观察到线状病毒,侵染率达77．3％;在40个被线状病毒侵染的样品中有27个观察到细胞质风轮状内舍体,表明受到马铃薯Y病毒属成员的侵染,侵染率达67．5％。从不同线状病毒粒子形态及所引起的细胞病理变化来看,可能分属于马铃薯Y病毒属、麝香石竹潜隐病毒属和马铃薯X病毒属。     

Application of FIB microsampling technique to long-period ordered TiAl single crystal with composition gradient

Metastable long-period superstructures in an L10-TiAl single crystal with a composition gradient were observed successfully by transmission electron microscopy with a focused ion beam (FIB) microsampling technique. The composition gradient from 54.7 to 75 at.% Al with approximately 6 microm width was detected by electron probe microanalysis and foil specimens containing the composition-gradient area were fabricated by the FIB microsampling method. The foil specimens clearly exhibit sequential changes in long-period superstructure depending on the Al concentration.     

猪瘟病毒与辛德毕斯病毒成熟和释放特征的比较研究

电镜下观察了二种有囊膜的正链ＲＮＡ病毒－－猪瘟病毒（ＣＳＦＶ）与辛德毕斯病毒（ＳｂＶ）感染的宿主细胞。在ＳｂＶ ６Ｋ蛋白突变株感染的ＢＨＫ２１细胞中,可清楚地显示病毒从细胞质膜出芽与释放的过程,并在细胞擀中积累了大量的核衣壳。在ＣＳＦＶ感染的ＭＰＫ细胞中首次观察到较多的病毒核衣壳与成熟的病毒粒子,因而有可能对其成熟与释放方式的不同进行比较研究。此外,对病毒诱导的细胞超微病变特征进行了报道     

Annotation of miRNAs in the COVID-19 Novel Coronavirus

The coronavirus disease 2019 (COVID-19) coronavirus is a new strain of coronavirus that had not been previously detected in humans. As its severe pathogenicity is concerned, it is important to study it thoroughly to aid in the discovery of a cure. In this study, the microRNAs (miRNAs) of COVID-19 were annotated to provide a powerful tool for the study of this novel coronavirus. We obtained 16 novel coronavirus genome sequences and the mature sequences of all viruses in the microRNA database (miRbase), and then used the miRNA matures sequences of the virus to perform the Basic Local Alignment Search Tool (BLAST) analysis in the coronavirus genome, extending the matched regions of approximately 20 bp to two segments by 200 bp. Six sequences were obtained after deleting redundant sequences. Then, the hairpin structures of the mature miRNAs were determined using RNAfold. The mature sequence on one hairpin arm was selected into a total of 4 sequences, and finally the relevant miRNA precursor prediction tools were used to verify whether the selected sequences are miRNA precursor sequences of the novel coronavirus. The miRNAs of the novel coronavirus were annotated by our newly developed method, which will lay the foundation for further study of this virus.     

Poxvirus virions: their surface ultrastructure and interaction with the surface membrane of host cells

Virions of vaccinia and orf viruses were examined by ultrahigh-resolution scanning electron microscopy using a non-coating method. Intracellular mature particles of vaccinia virus appeared to be covered with a net and ultrastructurally their surface consists of many fine ridges and globules, while the surfaces of orf virus mature particles recovered from infected cells consist of spirally running protrusions. The ridge-like structures of vaccinia virus were presumed to correspond to surface tubules shown by negative staining of this virus, while the spiral protrusions of orf virus were presumed to correspond to spiral threads having a criss-cross appearance by the same staining. Using scanning electron microscopy in which the samples were prepared by the conventional method, we observed: (i) many virions, i.e. one or two hundreds, or occasionally more reaching about one thousand particles, of the IHD strain of vaccinia virus, (ii) many or a moderate number of virions, i.e. about one hundred or fewer particles, of the 58 strain of cowpox virus and (iii) rather few virions, i.e. several tens or fewer particles, of the Iwate strain of orf virus on the free surface of each cell infected with these viruses. It must be noted that the number of virions detected considerably differed in respective cells examined. Virus budding was frequently observed at the cell surface of monolayer cells infected with vaccinia virus but it was never detected with cowpox or orf virus, indicating a difference in the mechanism of virus release between vaccinia and the other two viruses. When whole cells infected with vaccinia virus were examined by a combination of high-voltage and scanning electron microscopies, virions on the cell surface and those inside the cells were clearly differentiated. All virions on the cell surface had an envelope, and some of the envelopes had a slack and/or one or more bulges.     

Annotation of miRNAs in COVID-19 coronavirus

The COVID-19 coronavirus is a new strain of coronavirus that had not been previously detected in humans. As its severe pathogenicity is concerned, it is important to study it thoroughly to aid in the discovery of a cure. In this study, the microRNAs (miRNAs) of COVID-19 were annotated to provide a powerful tool for the study of this novel coronavirus. We obtained 16 novel coronavirus genome sequences and the mature sequences of all viruses in the microRNA database (miRbase), and then used the miRNA mature sequences of the virus to perform the Basic Local Alignment Search Tool (BLAST) analysis in the coronavirus genome, extending the matched regions of approximately 20 bp to two segments by 200 bp. Six sequences were obtained after deleting redundant sequences. Then, the hairpin structures of the mature miRNAs were determined using RNAfold. The mature sequence on one hairpin arm was selected into a total of 4 sequences, and finally the relevant miRNA precursor prediction tools were used to verify whether the selected sequences are miRNA precursor sequences of the novel coronavirus. The miRNAs of the novel coronavirus were annotated by our newly developed method, which will lay the foundation for further study of this virus.     

先天性大疱性表皮松解症的超微病理诊断

目的：应用电镜观察先天性大疱性表皮松解症（EB）的超微结构特点并进行分类诊断。方法：按电镜常规进行标本制备,应用透射电镜对22例EB的皮肤活检进行超微结构观察。结果：超微病理诊断单纯型（EBS）15例,交界型（JEB）1例,营养不良型（DEB）6例。EBS的表皮基底层见水疱和裂隙样结构,基底层细胞变性及空泡形成,张力丝断裂、缩短及凝聚。JEB的水泡在基底膜的透明板,半桥粒发育不良、数目减少。DEB水疱位于真皮内,锚纤维缩短、减少。结论：电镜超微病理观察对EB的分类诊断有重要的辅助诊断价值。