海湾扇贝一种球形病毒的形态发生及细胞病理学观察

2001年3-5月,青岛海区养殖海湾扇贝发生外套膜"糜烂病"并导致约50%亲贝死亡,主要表现为外套膜糜烂;严重者,约2/3的外套膜溃烂呈胶水状.电镜检测发现病贝体内感染有病毒等病原微生物.本文报道了该病毒粒子的形态、发生及宿主细胞由此所产生的细胞病理学变化.成熟的病毒粒子近球形,直径150～180nm,具囊膜,在细胞核附近的溶酶体内发生和增殖.发生初期溶酶体内形成板层髓样结构,随后形成块状、泡状、絮状等多形态的病毒发生基质及具有正方形花样的蛋白质晶格结构.最后,大量的病毒粒子装配形成,填充在溶酶体内.该病毒粒子主要存在于消化盲囊上皮细胞及结缔组织细胞的胞质中.受感染的宿主细胞线粒体肿胀、嵴溶解,内质网肿胀、核糖体脱落,溶酶体数量增多,核膜膨胀、溶解等,大部分细胞器受损.     

刺参一种囊膜病毒的分离及其超微结构观察

利用电镜负染技术检测发病的养殖刺参[Apostichopus japonicus（Selenka）]组织提取液。观察发现，提取液中存在大量病毒样粒子，该病毒粒子近似球形，具有囊膜。囊膜内可见高电子密度的核心结构。完整的病毒粒子直径为80～100nm，囊膜厚6～10nm，核心结构直径35-45nm，呈六边形。应用超薄切片技术对刺参的触手顶部、触手臂、疣足、呼吸树、背肠血管、肠等组织的病毒感染状况进行观察，发现该种病毒粒子大量存在于所检测各组织内。感染细胞超微结构表现为大量细胞器崩解形成空泡结构，并出现“髓袢样”结构等病理变化。根据观测结果，该病毒是一种无包涵体病毒。     

养殖扇贝几种常见疾病及防治

1 栉孔扇贝的球形病毒病 1.1 病原 球形病毒。王崇明等(2002)报告了栉孔扇贝(Chlamys farreri)的球形病毒病。病毒粒子近似圆形,大小为130～170nm,核衣壳直径为90～140nm。具有囊膜,厚约7～10nm,囊膜与核衣壳之间的间距为13～16nm,囊膜表面覆有长20～25nm的纤突,囊膜纤突致密地镶嵌成规则的毛边样。无包涵体。     

中国对虾淋巴细胞培养中的病毒及病理观察

通过对中国对虾的淋巴器官和经过培养的养殖,海捕中国对虾的淋巴组织细胞进行电镜观察,均发现在细胞内存在一种形状为球形,有囊膜,平均直径为１３６ｎｍ的病毒。病毒分布于细胞质内,或成团存在,似为一种虹彩病毒;在养殖对虾淋巴器官及培养过的组织细胞中还发现另一种病毒,病毒粒子为正二十面体,无囊膜,平均直径为３３ｎｍ,分布于细胞质内,似为一种小ＲＮＡ病毒。     

栉孔扇贝大规模死亡致病病原的研究

对2000-2002年山东沿海6个疫区患病栉孔扇贝进行电镜观察，在消化腺、外套膜、肾和肠的结缔组织细胞和间质细胞中发现一种球形病毒．并引起相直的病理学变化。该病毒具囊膜，直径为130～170nm，核衣壳直径为90～140nm。病毒分离纯化后．观察到的病毒囊膜表面覆有长20nm的纤突。病毒在细胞质中的囊泡样结构内完成装配，其内未发现包涵体存在。从发病疫区栉孔扇贝组织中分离出病毒毒种对键康扇贝进行人工感染。结果显示，各感染实验组发病扇贝表现出与自然海区发病扇贝相同的临床症状。病毒注射组死r二率为?5％，病毒浸浴组死亡率为68．7％．灭活病毒注射组和空白对照组死亡率皆为12．5％。病毒注射、浸浴组与灭活病毒注射组、空白对照组死亡率差异显著。电镜复检结果显示，各感染实验组发病扇贝组织中分布有大量病毒粒子，与自然海区发病扇贝组织所观察到的病毒粒子在形态特征和病理学特征上完全一致。以上结果证明，病毒是导致栉孔扇贝大规模死亡的直接病原。     

养殖中华鳖出血病病毒的电镜观察

本文应用电镜超薄切片技术，观察了福建省四个主要养鳖县市现场采集的鳖出血病样品，首次发现了一种感染养殖中华鳖的球形病毒（Trionyx sinensis spherovirus,TSSV）。该病毒分布在肺、胃、咽喉粘膜和腹甲皮层。主要靶细胞是这些组织的小血管和毛细血管的内皮细胞。TSSV粒子近球形，直径35-39nm，无囊膜包被，有的成群聚集在内皮细胞质中，有的由单位膜包裹在包涵体中，病毒感染的内皮细胞病变明显。本文还讨论了TSSV与出血病的关系。     

南美白对虾的病害防(二)

3.黄头病【病因】该病病原是黄头病病毒,呈杆状,有囊膜,病毒粒子(50～95)纳米×(130～380)纳米,属单链RNA病毒。【症状】病虾早期游动迟缓,头胸甲呈黄色或发白,膨大,鳃变成淡黄色到棕色,肝胰腺变成淡黄色。主要感染鳃组织、淋巴器官、血细胞和结缔组织等。该病毒毒力较强,对虾被感染后3～5天内可全军覆灭。患病虾组织多处坏死。胞核缩小或破裂,核周围细胞质内有嗜碱性的球形包涵体。染毒组织的血窦中有一种独特的数目不等的球形细胞。其胞质嗜碱性、球形胞核居中。【流行及危害】该病首先在印度斑节对虾的精养塘中发现,后在东亚地区的其它…     

中国对虾1种球形病毒及肝胰腺病变的电镜观察↑（*）

在１９９４年山东地区对虾暴发性流行病研究中,在未检出杆状病毒的中国对虾样品中首次发现１种球形病毒,电镜下观察到该病毒外有囊膜,直径２００ｎｍ左右,在细胞质中形成球形包涵体,以宿主肝胰腺和心脏为主要靶器官;受其感染的肝胰腺细胞的细胞器出现一系列病理变化：线粒体从增生、肿胀、内嵴溶解到出现纤维化,最后崩解;糙面内质网也明显增生、脱颗粒、排列紊乱,严重感染时,肿胀成泡状,甚至分解;细胞核出现病变较晚且较轻。人工感染实验证明该病毒可引起中国对虾暴发性流行病的症状,１３ｄ累积死亡率可达９０％。     

牡蛎疱疹病毒对魁蚶的致病性

为探明牡蛎疱疹病毒(Os HV-1)对魁蚶的致病性,本研究使用发病魁蚶组织制作病毒悬液进行感染实验。感染实验分为空白组、阴性悬液注射组和病毒悬液注射组,并使用实时定量PCR法对感染后魁蚶体内病毒的时空分布进行检测。实验结果显示,空白组和阴性悬液组魁蚶感染后未检测到病毒粒子,病毒悬液注射组魁蚶经人工感染后,各部位病毒含量均呈先上升再下降随后又上升的趋势,最终达到106拷贝/ng DNA左右。通过电镜观察,在感染魁蚶的鳃、肝胰腺、外套膜中出现染色质边缘化甚至消失,细胞核肿胀、溶解,核仁消失,核膜扩张、不清晰,线粒体肿大,脊崩解,核糖体脱落等一系列细胞病理变化。在其细胞核和细胞质中均能发现大量直径为90~110 nm球形病毒粒子,该病毒粒子具囊膜,囊膜内可见均匀高电子密度的核衣壳,与自然发病魁蚶负染电镜中的病毒粒子形态相同。研究结果表明,Os HV-1可以感染魁蚶并与魁蚶大规模死亡有直接相关关系;魁蚶感染Os HV-1后机体产生应激反应,对Os HV-1有一定抑制作用,但其作用机制还有待进一步实验验证。     

中国对虾淋巴组织培养中的病毒及病理观察

通过对中国对虾的淋巴器官和经过培养的养殖、海捕中国对虾的淋巴组织细胞进行电镜观察 ,均发现在细胞内存在一种形状为球形 ,有囊膜 ,平均直径为 136nm的病毒 ,病毒分布于细胞质内 ,或成团存在 ,似为一种虹彩病毒 ;在养殖对虾淋巴器官及培养过的组织细胞中还发现另一种病毒 ,病毒粒子为正二十面体 ,无囊膜 ,平均直径为 33nm ,分布于细胞质中 ,似为一种小RNA病毒。在体外培养淋巴组织过程中 ,病毒在细胞内没有明显增殖迹象。感染病毒的细胞呈现细胞核固缩 ,细胞质空泡化 ,线粒体内嵴模糊 ,粗面内质网水肿等一系列病理变化。对淋巴组织切片进行光镜观察 ,发现淋巴器官组织部分坏死 ,有些细胞核肿大 ,苏木精深染 ,严重的细胞核结构已经被破坏。     

Morphogenesis of koi herpesvirus observed by electron microscopy

Koi herpesvirus (KHV or cyprinid herpesvirus 3) was inoculated onto three fish cell lines derived from carp, Cyprinus carpio L., and the process of virion formation observed with electron microscopy. Essentially, similar features of virus particles were observed in all three cell lines. The nucleus of infected cells was characterized by margination of chromatin and often contained many capsids of about 110 nm in diameter with varying morphology. The morphological variation of the capsids was very similar to that of mammalian herpesviruses. Some capsids protruded from the inner nuclear membrane, and others, with envelopes, were located in the perinuclear space between the inner and outer nuclear membrane, suggesting budding of capsids at the inner nuclear membrane. Naked capsids and envelopment of capsids by budding into vesicles were also observed in the cytoplasm. Mature, enveloped virions 170-200 nm in diameter were seen in cytoplasmic vesicles or outside the cell. These observations suggest KHV virions mature through a complex morphological pathway including two distinct envelopments, which have been found only in herpesviruses. These observations support the inclusion of KHV in the family Herpesviridae.     

A dual infection by infectious cuticular epithelial necrosis virus and a Chlamydia-like organism in cultured Litopenaeus vannamei (Boone) in Ecuador

During 1996, microscopic examinations of post larvae and juveniles of moribund Litopenaeus vannamei showed multifocal necrosis in the cuticular epithelial tissues. In addition to these severe degenerative alterations in the epithelial cells typical of infectious cuticular epithelial necrosis virus (ICENV), columnar cells of the epithelium displayed small round intracytoplasmic inclusions in the necrotic tissue. Examination by electron microscopy of affected tissues demonstrated prokaryotic organisms in the cytoplasm of epithelial cells delineated by a distinct cytoplasmic vesicle; the prokaryotic organisms were morphologically similar to the genus Chlamydia. The necrotic tissue also showed the presence of particles of ICENV; the double infection by two different organisms in cuticular epithelial cells has not been reported previously. Two distinct stages in the intracellular development of a Chlamydia‐like organism were recognized: (1) pleomorphic elementary bodies (EBs) that were spherical to oval were often observed in the process of division or in forming a common chain of three cells, the cells were surrounded by a rigid cell envelope and the presence of a cap or plaque hexagonally arrayed; (2) the reticular bodies (RBs) were forms often in the process of division. These cells had an electron‐dense cytoplasm and contained a loose network of nuclear fibrils and a more fragile cell envelope. Regardless of the development stages of the Chlamydia‐like organism within the cytoplasmic vesicles, ICENV particles were observed, either dispersed or in clusters, surrounded or inside the vesicles. The potential adverse impact of this dual infection on shrimp culture should be considered, especially in high‐density operations.     

一种真鲷球形病毒的形态及细胞病理学电子显微镜观察

对１９９５年青岛地区患病真鲷肝脏、肠上皮组织、贤脏、鳃等组织进行电镜观察,发现一种球形病毒,该病毒具有囊腊,直径为８０￣１００ｎｍ,在核和细胞质中均形成不规则形态包涵体;细胞病理变化表现为核内染色质变性,核膜水肿变形且部分溶解,内质网膨胀,胞质内有大量游离核糖体分布,线粒体外膜及内嵴变形且有少量溶解。     

Studies of the ultrastructure and morphogenesis of fish pathogenic viruses grown in cell culture

A birnavirus (infectious pancreatic necrosis virus, IPNV), three rhabdoviruses (viral haemorrhagic septicaemia virus, VHSV; infectious haematopoietic necrosis virus, IHNV; and spring viraemia of carp virus, SVCV) and an iridovirus (isolate from a sheatfish) were investigated with regard to their morphogenetic interactions with cells in culture. In cells infected with birnavirus, a granular viromatrix, single virions randomly distributed in the cytoplasm, viral particles aggregated in pseudocrystals and cytoplasmic tubuli similar in diameter to that of the virus were found. Rhabdoviruses entered the cells by viropexis and replicated within the cytoplasm. Maturation occurred predominantly at the cell membrane and sporadically at membranes of the Golgi cisternae. Inclusion bodies were found partially consisting of viral nucleocapsids. After budding, new virions were found adsorbed to the cell membrane. Viral haemorrhagic septicaemia virus, known to exhibit an atypical shape because of preparative procedures, could be identified by immunostaining using two monoclonal antibodies directed against G- and N-proteins and colloidal gold. Iridoviruses entered the cells by viropexis. Viral particles were found in coated vesicles. Subsequently, vesicles without a clathrin coat were detected. Replication occurred within prominent cytoplasmic inclusion bodies. Isometric viral nucleocapsids were transported in an unknown manner to the cell membrane and matured by budding.     

“红肉病”文蛤中寄生类立克次体的超微结构与细胞病理学

报道了患“红肉病”文蛤的消化盲囊上皮中所发现的类立克次体以及所引起的受感染细胞的病理学变化。该病原包涵体呈球形或椭球形,有强嗜碱性和嗜酸性两种,电镜下观察到上皮细胞内充满了大量的类立克次体,其形态结构及大小表现了极大的多样性,受感染的细胞具有明显的病理学变化,内质网膨大为潴泡状,核糖体脱落;线粒体溶解或固缩,核膜肿胀,染色质固缩并出现液泡结构,高度感染时,细胞核与大部分的细胞器消失,在上皮细胞中,观察到类立克次体以二分裂和出芽方式进行繁殖。     

Ultrastructural morphogenesis of salmonid alphavirus 1

Studies on the ultrastructural morphogenesis of viruses give an insight into how the host cell mechanisms are utilized for new virion synthesis. A time course examining salmonid alphavirus 1 (SAV 1) assembly was performed by culturing the virus on Chinook salmon embryo cells (CHSE‐214). Different stages of viral replication were observed under electron microscopy. Virus‐like particles were observed inside membrane‐bound vesicles as early as 1 h following contact of the virus with the cells. Membrane‐dependent replication complexes were observed in the cytoplasm of the cells, with spherules found at the periphery of late endosome‐like vacuoles. The use of intracellular membranes for RNA replication is similar to other positive‐sense single‐stranded RNA (+ssRNA) viruses. The number of Golgi apparatus and associated vacuoles characterized by ‘fuzzy’‐coated membranes was greater in virus‐infected cells. The mature enveloped virions started to bud out from the cells at approximately 24 h post‐infection. These observations suggest that the pathway used by SAV 1 for the generation of new virus particles in vitro is comparable to viral replication observed with mammalian alphaviruses but with some interesting differences.     

Lymphocystis infection in snakeskin gourami, Trichogaster pectoralis (Regan), (Anabantidae)

Abstract. A lymphocystis infection is described from aquarium-held snakeskin gourami, Trichogaster pectoralis (Regan). The cytoplasm of the hypertrophic cells revealed virus particles of extended hexagonal profiles with thin two-layered capsids. Subsequent examination of the fish 4 months later revealed hypertrophic cells showing varying degrees of collapse and degradation. The virus particles loading such cells were of regular hexagonal profiles with thick three-layered capsids.     

凡纳滨对虾卵子发生的超微结构

利用透射电镜研究凡纳滨对虾卵子发生过程中细胞内部结构的变化。结果显示,卵原细胞结构简单,代谢水平低,核孔稀少,通过核周池来完成核、质之间的物质交换。卵黄发生前晚期和卵黄发生初期的卵母细胞变化显著：核膜凹凸不平,核仁数量多,核孔密集,大量核仁外排物经核孔输送到卵质中;卵质中胞器极为丰富和发达,代谢活性极强。卵黄发生旺盛期是卵黄大量形成的阶段,卵质边缘还呈辐射状排列了一圈椭圆形皮质棒,细胞出现微吞饮活动并形成卵黄膜。卵黄发生晚期卵质中充满了粗大的卵黄粒和脂滴,胞器锐减。另外,探讨了卵细胞内部结构的变化和卯黄形成的关系以及皮质棒的来源与功能。