八种海洋盾纤类纤毛虫的形态学研究

对采自山东、广东和香港沿海的八种盾纤类纤毛虫: 冠帆口虫、维亚可夫帆口虫、蠕形康纤虫、贪食迈阿密虫、异海洋尾丝虫、中华后阿脑虫和两种蟹栖异阿脑虫相似种进行了形态学研究。其中, 中国南海新记录种维亚可夫帆口虫种群与Wang, et al.所描述种群以及中华后阿脑虫新种群与Song Wilbert记述的青岛种群和南极种群相比, 在个体大小和体动基列数目上均有所差异; 贪食迈阿密虫潍坊种群的体动基列恒为12列, 与Song Wilbert的报道有较大差异。两种异阿脑虫与蟹栖异阿脑虫在口纤毛器结构和体形上完全相似, 但在体动基列数目上有明显差异, 因数据不足, 暂定为蟹栖异阿脑虫相似种。       

青海湖裸鲤和黄河裸裂尻鱼感染多子小瓜虫的病理学比较研究

为探讨不同裂腹鱼类感染多子小瓜虫后的病理学差异, 利用多子小瓜虫对青海湖裸鲤指名亚种(Gymnocypris przewalskii przewalskii)和黄河裸裂尻鱼(Schizopygopsis pylzovi Kessler)实施感染实验, 并对2种鱼进行了深入的病理学研究。研究结果显示: (1) 2种鱼的死亡量均呈现先激增后明显回落的趋势, 青海湖裸鲤死亡高峰在感染后第3至第4天, 黄河裸裂尻鱼死亡高峰在第3至第5天, 青海湖裸鲤的死亡比黄河裸裂尻鱼急剧。(2)感染后2种鱼体表均出现大量肉眼可见的白点。青海湖裸鲤皮肤黏液分泌量明显增加, 体表形成胶状黏液层, 黏液层中见不同细胞期小瓜虫包囊。黄河裸裂尻鱼鳍出现蛀鳍现象, 皮肤出现细菌感染样溃烂。(3)解剖发现, 感染组青海湖裸鲤和黄河裸裂尻鱼肝脏发生病理改变呈淡黄色, 胆有不同程度肿大。(4)组织切片和电镜观察显示, 小瓜虫在鳃部位的寄生导致青海湖裸鲤和黄河裸裂尻鱼鳃丝黏连, 鳃小片和鳃丝上皮细胞萎缩、脱落, 鳃丝结构被严重破坏。小瓜虫在2种鱼皮肤的寄生使寄生部位组织突起, 周围组织塌陷。青海湖裸鲤表皮下层出现空隙, 表皮结构被严重破坏。黄河裸裂尻鱼皮肤表皮细胞出现空泡化病理改变, 失去原有紧密结构, 表皮层和固有层间界限变模糊。综上所述, 小瓜虫的感染对青海湖裸鲤和黄河裸裂尻鱼的鳃和皮肤组织造成严重损伤, 但2种鱼表现的症状和造成的组织损伤类型有明显差异, 这与2种鱼长期适应不同水体环境密切相关。     

正常与感染粘孢子虫鲫鱼主要组织可溶性蛋白质电泳比较

采用SDS-聚丙烯酰胺凝胶电泳技术,对正常和感染粘孢子虫的鲫鱼动脉球、肝脏、鳃、脑、肠、肌肉、眼、鳍8种组织可溶性蛋白质进行分析比较。结果表明：病鱼的动脉球、肝脏、鳃、脑、肠5种组织的蛋白质电泳谱带与正常鱼相比有较大变化,动脉球缺失1条谱带同时又诱导出1条新谱带;肝脏有9条谱带缺失;鳃有2条谱带缺失的同时诱导出3条新谱带;脑中新增3条谱带;肠中有5条谱带缺失同时产生3条新谱带。这些变化可作为辅助疾病诊断的生化指标,为从生化水平上揭示该病的致病机理及开展有效的早期防治研究奠定基础。     

方斑东风螺单孢子虫病的研究

文章对方斑东风螺(Babylonia areolata Link, 1807)单孢子虫病进行了首次报道, 对其病理学特征、流行季节和诊断方法进行了研究。结果认为方斑东风螺被单孢子虫侵袭后, 吻管、足肌、肠、消化腺、鳃、胃和肝等器官引起一系列病变。病理组织切片观察到肠上皮黏膜组织细胞布满该虫营养体和各发育期的多核质体, 造成肠结缔组织呈现炎症反应。寄生部位病灶肿胀、混浊、坏死、崩解。       

草鱼出血病病毒人工感染稀有Ju鲫出血病鱼主要器官组织的超薄…

用电子显微镜观察了稀有Ｊｕ鲫出血病鱼的病理切片及对照鱼的超薄切片。在对照鱼的鳃、肠道、肾脏、肌肉和肝脏中没有见到任何病毒颗粒。在病鱼鳃血管内皮细胞质中观察到聚集的直径７０ｎｍ左右的ＧＣＨＶ颗粒,说明鳃是ＧＣＨＶ侵袭的主要器官之一,并讨论了鳃对ＧＣＨＶ敏感在ＧＣＨＶ传播的意义。还在病鱼肠道,肾脏中观察到成片的病毒颗粒,它们也是ＧＣＨＶ侵袭的主要组织。在病鱼脾脏在电子密度低在细胞质中观察到散在的病毒可     

草鱼出血病病毒人工感染稀有(鱼句)鲫出血病鱼主要器官组织的超薄切片观察

用电子显微镜观察了稀有(鱼句)鲫出血病(Hemorrhage of Gobiocypris rarus)鱼的病理切片及对照鱼的超薄切片。在对照鱼的鳃、肠道、肾脏、脾脏、肌肉和肝脏中没有见到任何病毒颗粒。在病鱼鳃血管内皮细胞质中观察到聚集的直径70nm左右的GCHV颗粒,说明鳃是GCHV侵袭的主要器官之一,并讨论了鳃对GCHV敏感在GCHV传播中的意义。还在病鱼肠道,肾脏中观察到成片的病毒颗粒,它们也是GCHV侵袭的主要组织。在病鱼脾脏的电子密度低的细胞质中观察到散在的病毒颗粒。很少见到病毒聚集体,脾脏中的病毒可能是脾脏的吞噬细胞吞噬而来。作者认为在病鱼组织中观察到的大小不同的病毒颗粒是处于不同成熟阶段的GCHV。     

养殖梭子蟹血卵涡鞭虫感染的初步研究

2004年7—9月,浙江舟山人工养殖梭子蟹相继发生一种被当地渔民称为"牛奶病"的流行性疾病,并引起大量死亡。病蟹蟹体消瘦,肌肉白浊,蟹盖内可见大量乳白色液体。对带有典型病症的濒死病蟹进行细菌分离,未分离到细菌。组织病理学观察发现,病蟹血淋巴急剧下降,代之以大量的寄生原虫,该寄生虫在病蟹的肝胰腺、鳃、心脏、肌肉等部位大量侵袭,并引起这些组织发生以坏死为主的变质性病变,其病理特征主要表现为:细胞肿胀、变性、坏死,某些细胞的细胞核固缩、碎裂或崩解。电镜观察乳白色体液及病变组织,可见大量寄生原虫,该寄生虫大小约5—7μm,多数呈卵圆形,单核或多核,具有2根鞭毛。进一步通过分子生物学方法,确定该寄生虫为血卵涡鞭虫Hematodinium sp.,初步认定其是引起养殖梭子蟹"牛奶病"的一种重要病原。     

鲤疱疹Ⅱ型病毒(CyHV-2)感染对异育银鲫(Carassius auratus gibelio)组织器官的损伤作用

为分析感染Cy HV-2异育银鲫(Carassius auratus gibelio)体内不同器官组织结构和功能的损伤,采集健康和患病异育银鲫头肾、脾脏、体肾、肠道、肝胰脏、鳃各组织样本,通过石蜡切片,HE染色对各器官组织进行组织病理学观察并对肠道和鳃进行扫描电镜观察。病鱼主要表现为,食欲减弱、离群独游、全身发黑、背鳍、尾鳍末端发白、眼球突出、肌肉充血,鳃呈鲜红色。在鳃丝末端由红色斑块,经解剖有红色腹水流出,组织器官广泛性损伤。组织病理学变化为,头肾和脾脏细胞溶解、免疫细胞侵润、免疫和造血系统严重破坏;体肾中肾小球萎缩、肾小管上皮细胞脱落、肾间质出现空泡、免疫细胞侵润;肠道绒毛增生、绒毛和微绒毛长度显著减少、粘膜下层显著增厚;肝胰脏中肝细胞出现溶解、中央静脉受损;鳃中毛细血管破坏、血管破裂或扩大、红细胞溢出、鳃小片基部细胞显著减少。结果表明,病毒对异育银鲫具有明显的致病性,可导致各器官组织发生病变,同时机体处于缺血、缺氧、营养不足、废物积累的内环境,这进一步加剧对异育银鲫主要的器官组织的损伤。     

中华绒螯蟹肝胰腺坏死综合症病原及病理学研究

为了探讨中华绒螯蟹“水瘪子”病病因, 对病蟹进行了寄生虫检查、病原生物分离、回接攻毒及电镜观察等病原学研究, 同时采用常规石蜡切片技术, 以健康蟹为对照, 对病蟹的不同组织进行了病理学观察。结果表明, 病原分析未见致病性生物; 病蟹的鳃、肌肉和肝胰腺发生了不同程度的病变, 主要的病理特征为鳃组织增厚, 鳃腔增大, 血细胞增多, 细胞核边缘化; 肝胰腺组织中单层上皮细胞空泡化, 并出现转运泡, 随着病情的加重, 肝细胞排列紊乱, 转运泡和空泡的数量增多, 体积增大, 且转运泡内内容物增多, 更甚者肝小管基膜破裂、内容物外流, 细胞核解体, 肝细胞出现坏死; 肌细胞的病变特征主要是肌丝松弛变性、细胞核固缩深染。病原学研究和发病情况调查表明该病为非生物性疾病, 依据该病的临床症状、病理变化以及发病原因, 将该病命名为中华绒螯蟹肝胰腺坏死综合症。     

嘉陵江8种鱼类不同组织微量元素含量分析

测定了黄颡鱼（Pelteobagrus fulvidraco）、中华倒刺钯（Spinibarbus sinensis）、大鳍鲠（Mystus macropterus）、长吻鲍（Leiocassis longirostris）、大眼鳜（Siniperca kneri）、小口鲇（Silurus asotus）、岩原鲤（Procypris rabaudi）和圆口铜鱼（Coreius guichenoti）8种嘉陵江名贵鱼不同组织微量元素含量及肌肉和脑中Ca、P含量,结果表明,鱼脑中各类矿物质元素显著高于肌肉;肌肉5种微量元素总量由低到高依次为：圆口铜鱼、中华倒刺鳃、长吻鲍、大鳍鲮、黄颡鱼、大眼鳜、岩原鲤、小口鲇;而脑由低到高依次为：大眼鳜、岩原鲤、小口鲇、圆口铜鱼、大鳍鲢、中华倒刺鳃、长吻鲍、黄颡鱼。微量元素在各内脏器官的分布：铁和铜以肝（胰）脏最高,肾脏其次,胃肠最低;锰和锌的含量肾脏〉肝（胰）脏〉胃肠;镁的分布各种鱼差异较大。     

Scuticociliate infection and pathology in cultured turbot Scophthalmus maximus from the north of Portugal

During the years 2004 and 2005 high mortalities in turbot Scophthalmus maximus (L.) from a fish farm in the north of Portugal were observed. Moribund fish showed darkening of the ventral skin, reddening of the fin bases and distended abdominal cavities caused by the accumulation of ascitic fluid. Ciliates were detected in fresh mounts from skin, gill and ascitic fluid. Histological examination revealed hyperplasia and necrosis of the gills, epidermis, dermis and muscular tissue. An inflammatory response was never observed. The ciliates were not identified to species level, but the morphological characteristics revealed by light and electronic scanning microscopes indicated that these ciliates belonged to the order Philasterida. To our knowledge this is the first report of the occurrence of epizootic disease outbreaks caused by scuticociliates in marine fish farms in Portugal.     

Histopathology of experimental scuticociliatosis in turbot Scophthalmus maximus

A scuticociliate strain (B-2), originally isolated from an outbreak in a turbot Scophthalmus maximus (= Psetta maxima) farm in Galicia (northwestern Spain) and maintained in axenic culture, was injected intracoelomically (lethal dose 80 equivalent, LD80) in healthy turbot (50 g). Ciliate-injected fish were kept under controlled conditions in a recirculating seawater system and sampled on Days 1 through 8, 10, 12 and 14 postinfection (PI). Necropsies were conducted and included blood collection from the caudal vein and samples of liver, spleen, heart, digestive tract, kidney, gills, abdominal wall and neurocranium taken for routine histology. Mortality occurred from Day 6 until Day 12 PI and reached 66.7% by the end of the experiment. Presence of ciliates in the coelomic fluid was scarce until Day 4 PI. Parasitaemia was only observed from Day 5 until Day 10 PI and its incidence was always low. Presence of scuticociliates in tissue sections followed a progressive pattern of diffusion, with ciliates showing preference for loose connective tissue and also a clear haematophagous activity. The most severely affected organs were the pancreas and digestive tract. No special tropism for nervous tissues was observed in this study. The inflammatory reaction was variable depending on the tissue. After 3 wk, survivors had apparently managed to extinguish the infection.     

Characterization of an iridovirus detected from cultured turbot Scophthalmus maximus in Korea

Juvenile turbot Scophthalmus maximus that became sick during an outbreak of disease at mariculture facilities at Go-Chang, Korea, in 2003, were examined to identify the cause of the disease. The fish had pale body color, an enlarged abdomen, protruding eyes, an enlarged spleen and kidney, and pale gills and/or liver. Histopathogical examination revealed basophilic enlarged cells in the kidney, spleen, gills, heart, stomach, intestine, liver, pancreas and skin. Hexagonal viral particles with a diameter of 136 to 159 nm were observed in the enlarged cells. A specific 1299 bp fragment of the major capsid protein (MCP) gene of the turbot iridovirus (TBIV) was amplified by PCR. Sequence homology was greater than 93.76% between the MCP gene in TBIV and the same gene in 5 viruses in the tentatively proposed genus Tropivirus (family Iridoviridae): red sea bream iridovirus, sea bass iridovirus, grouper sleepy disease iridovirus, African lampeye iridovirus and dwarf gourami iridovirus. These results suggest that the virus detected from turbot is similar to the proposed genus Tropivirus.     

Experimental susceptibility of turbot Scophthalmus maximus to viral haemorrhagic septicaemia virus isolated from cultivated turbot

Juvenile pathogen-free turbot were infected with a viral haemorrhagic septicaemia virus (VHSV) isolate recovered from turbot cultivated on the island of Gigha, West Scotland. Mortality of 100% was recorded in fish infected via the intra-peritoneal (i.p.) route. Horizontal transmission of VHSV in sea water was demonstrated by cohabitation of naive fish with i.p. infected fish at a ratio of 1:1. The total cumulative average mortality in cohabiting fish was 60% by 60 d post-infection. Turbot infected via an immersion route exhibited a cumulative average mortality of 71% by the end of the experiment. VHSV identified by enzyme-linked immunosorbent assay (ELISA) was recovered from both organ (kidney and spleen) and brain samples of individual fish that died following infection by all experimental routes. These findings pose significant implications regarding the persistence of VHSV and its role in limiting natural populations of marine fish species. In addition, the establishment of infection models for the transmission of VHSV in sea water is of fundamental importance to the development of anti-VHSV vaccines in important commercial species such as turbot.     

The ontogeny of complement component C3 in Atlantic cod (Gadus morhua L.)--an immunohistochemical study

The complement system in fish is well developed and plays an important role in the immune response. Very little is known about the ontogeny of C3 in fish and no study has previously been done on the development of C3 in teleosts. In this study we have detected the presence of C3 in cod larvae from the age of 1 day post hatching (p.h.) till 57 days p.h., using immunohistochemistry. The specific primary antibodies used, were produced against the beta-chain of cod C3. Immunostaining on cod larvae sections revealed that C3 is detectable in the yolksac membrane from day 1 p.h., and in liver, brain, kidney and muscle from day 2 p.h. C3 was also detected in other organs such as eye, notochord, stomach, intestines, pancreas, heart and gills at different stages of cod larval development. These findings suggest that complement is not only important in immune defence against invading pathogens but may also play a role in the formation and generation of different organs.     

Colonization of turbot tissues by virulent and avirulent Aeromonas salmonicida subsp. salmonicida strains during infection

Preventing disease outbreaks in cultured turbot Psetta maxima L. caused by Aeromonas salmonicida subsp. salmonicida (ASS) requires a better understanding of how this pathogen colonizes its host. Distribution of 1 virulent and 2 avirulent ASS strains in turbot tissues was investigated during early and late stages of infection following an immersion challenge. To track bacteria within the turbot, the ASS strains were tagged with green fluorescent protein (GFP). Both virulent and avirulent strains colonized the epidermal mucus, gills, and intestine within the first 12 h post challenge, suggesting that these sites may serve as points of entry into turbot. Although the avirulent strains colonized these initial sites in the turbot tissues, they were rarely found in the internal organs and were cleared from the host 4 d post challenge. In contrast, the virulent ASS strain was found in the liver and kidney as early as 12 h post challenge and was found in the muscle tissue at very late stages of infection. The virulent strain persisted in all tested host tissues until death occurred 7 d post challenge, suggesting that ASS must colonize and survive within the turbot tissues for an infection to result in death of the fish. Comparisons of the distribution profiles of both virulent and avirulent strains during early and late stages of an infection in turbot has provided important information on the route and persistence of an ASS infection in this host.     

Quantitative and qualitative evaluation of iNOS expression in turbot (Psetta maxima) infected with Enteromyxum scophthalmi

Enteromyxum scophthalmi is the causative agent of turbot enteromyxosis, an intestinal parasitisation that produces severe desquamative enteritis leading to a cachectic syndrome and eventually the death. It is well known the importance of the innate immune response against parasites in fish, with the release of antimicrobial substances such as reactive oxygen and nitrogen species, produced by the inducible nitric oxide synthase (iNOS). This enzyme is mainly found in phagocytes, but also in structural cells from the intestinal mucosa. The aim of this study was to characterize iNOS in intestine and lymphohaematopoietic organs (spleen and anterior kidney) of turbot by means of immunohistochemistry in order to assess the possible changes of this enzyme through the infection. The presence of the enzyme was evaluated in control and E. scophthalmi-infected turbot. The results showed immunoreactivity in the apical border of enterocytes and mild staining of goblet cells in both control and infected turbot although it was more evident and widespread in infected turbot compared to control. Moderate numbers of iNOS+ cells were present in the lamina propria-submucosa of fish which presented moderate and severe inflammatory infiltrates at this level. In spleen and kidney, iNOS+ cells were scattered through the parenchyma and, in severely infected fish, tended to be allocated near the vascular structures and melano-macrophage centres. The number of positive cells at the lymphohaematopoietic organs was significantly higher in infected turbot and increased as infection progressed. The increase in the expression of iNOS in the tissues of E. scophthalmi-infected turbot was more evident in individuals with severer lesions. The measurement of the levels of iNOS during turbot enteromyxosis reveals a possibly delayed response that would not able to eliminate the parasites but would exacerbate mucosal injury.     

Quantitative investigation of antigen and immune response in nervous and lymphoid tissues of Atlantic halibut (Hippoglossus hippoglossus) challenged with nodavirus

The present study reports the quantitative analysis of the spatio-temporal development of nodavirus infection and corresponding immune response in juvenile Atlantic halibut (Hippoglossus hippoglossus) challenged by intramuscular injection of nodavirus. Novel quantitative real-time RT-PCR protocols were applied to evaluate the absolute copy numbers of nodavirus RNA2 (RNA2) and secretory-IgM mRNA (sec-igmicro) in the eye, brain, mid/posterior kidney and spleen sampled over a period of 81 days. In the eye and brain, levels of both RNA2 and sec-igmicro increased significantly early in the infection. In the spleen and mid/posterior kidney, both RNA2 and sec-igmicro were detected but the levels remained unchanged during the experimental period. The levels of RNA2 and sec-igmicro in the eye and brain were strongly correlated (P<0.0001). Nodavirus antigen was demonstrated by immunohistochemistry (IHC) in the retina of eyes from a relatively few fish from day 34 post challenge (brain not examined), but not at any time in the spleen and anterior kidney. By IHC, IgM+ cells were observed in conjunction with nodavirus positive IHC labelling in the retina. In both the spleen and anterior kidney, the number of IgM+ cells increased from day 3 post challenge. By conventional real-time RT-PCR, RNA2 was only sporadically demonstrated in the posterior intestine, heart and gills. ELISA analysis revealed a nodavirus specific antibody response in serum that was significant from day 18 post challenge. No clinical signs or mortality related to nodavirus infection were observed in the challenged halibut. The results suggest that the nodavirus infection induced a significant antibody response through activation of B-cells in the kidney and spleen, and involved a substantial migration of antibody-secreting cells to infected peripheral tissues.