广东大亚湾六种海洋刺钩类纤毛虫的形态学研究

利用活体观察及银染法首次对采自广东大亚湾沿岸的盐菲阿虫Phialina salinarum Kahl,1928,海洋长吻虫Lacrymaria marina Kahl,1933,惊扰伪颈毛虫Pseudotrachelocerca trepida(Kahl,1928)Song,1990,贪食纤口虫Chaenea vorax Quennerstedt,1867,柱核柱毛虫Cyclotrichium cyclokaryon Meunier,1907和加冈栉毛虫Didinium gargantua Meunier,1910六种刺钩类纤毛虫进行了形态学研究,补充了活体形态学以及纤毛图式等分类学新信息。首次观察到了海洋长吻虫体纤毛基部的皮层颗粒和柱核柱毛虫大亚湾种群较青岛种群具有更多的体动基列等若干新信息。研究表明,刺钩类纤毛虫在大亚湾近岸水域具有较高的物种多样性。       

蟹栖异阿脑虫寄生大菱鲆及其组织病理学研究

论文首次报道了蟹栖异阿脑虫(Mesanophrys carcini)在养殖大菱鲆体内的寄生情况以及所引起宿主的组织病理学变化。观察结果表明,感染的苗期大菱鲆的大部分器官组织包括皮肤、鳍、鳃、眼、脑、心脏、肝脏、肾脏、脾脏、胰腺和消化道等以及器官组织间隙中都发现了寄生的蟹栖异阿脑虫,属于全身性感染;而对于成鱼,蟹栖异阿脑虫主要寄生于皮肤、鳍和鳃等器官组织;严重感染者,蟹栖异阿脑虫可侵入其脑、肝脏、肾脏和心脏等器官组织,但数量不多。蟹栖异阿脑虫在大菱鲆体内的寄生能够引起病鱼各器官组织发生不同程度的组织病理学改变。该病在幼鱼和成鱼身上所表现出较一致的病理学特征为:鳃小叶呼吸上皮细胞增生;脑膜炎和脑组织液化性坏死;肾脏和脾脏造血组织坏死;肝脏中央静脉和窦状隙扩张;肌纤维发生凝固性坏死和眼的脉络膜充血、水肿;单核巨噬细胞浸润。研究认为,蟹栖异阿脑虫寄生引起的鳃损害使病鱼呼吸困难,窒息是病鱼发生死亡的主要原因,而虫体寄生导致的各器官组织变性,坏死,使得病鱼器官功能衰竭可加速鱼的死亡。另外,文中还探讨了蟹栖异阿脑虫入侵大菱鲆的可能途径,认为破损的皮肤和鳃是入侵的主要途径。研究结果告诉我们,在大菱鲆养殖过程中防止鱼体擦伤,保持池底清洁对于预防疾病的发生非常重要;对该病的治疗应采取外部杀虫和内部口服药物相结合的方式。     

三种危害性纤毛虫的形态学研究

三种分离自海水养殖水体中的纤毛虫原生动物,含一新亚种及两种国内新记录：相似瓜形膜袋虫,有唇蓝口虫和贪食拟阿脑虫;本文对其纤毛图式及重要的形态学特征做了综合的描述。有唇蓝口虫为可导致水质恶化的赤潮生物,其余两种则有明显的嗜污习性,或附着养殖动物的浅表倍位从而有可能构成对宿主的危害。     

尖前口虫的口器发生研究(纤毛门, 膜口目)

研究了咽膜类纤毛虫尖前口虫无性生殖期的核器及口器的演化.其发生特征为;1)新的口原基形成于原前庭动基列与口侧膜间,表观为原口侧膜分裂而致;2)随着形态发生的进行,由口原基依次演化出后仔虫的三片咽膜、口侧膜和三条前庭动基列;3)原口器完全被前仔虫所继承;4)体纤毛器在整个形态发生过程中一直保持双动基列结构.       

松花江四种纤毛虫的形态学研究

利用活体观察, 氨银染色和蛋白银染色的方法, 对采自松花江的4种广布种纤毛虫: 贻贝棘尾虫Stylonychia mytilus (Müller, 1773) Ehrenberg, 1830、鬃异源棘尾虫Tetmemena pustulata (Müller, 1786) Eigner, 1999、尾草履虫Paramecium caudatum (Ehret, 1963)和刚毛榴弹虫Coleps hirtus的形态学进行研究。依据新采集种群, 对4个种活体形态和纤毛图式特征, 以及鬃异源棘尾虫的发生学特征进行补足性描述。研究对贻贝棘尾虫做了重要重描述, 可清晰观察到该种蛋白银染后6列背触毛的排列特征、左右缘棘毛数的具体统计, 结果发现, 贻贝棘尾虫哈尔滨种群与前人报道的种群相比, 形态特征基本吻合, 区别在于前者小膜数目多于后者。鬃异源棘尾虫哈尔滨种群与漠河种群相比, 形态和纤毛图式方面十分相似, 发生模式基本一致, 细微区别在于前者左右缘棘毛数目普遍多于后者, 前者虫体大小波动范围较小, 小核数目多于后者。尾草履虫哈尔滨种群与其青岛种群相比, 二者显著差异在于收集管数目前者比后者少。本文榴弹虫与前人报道的刚毛榴弹虫种群相比, 盔板带的数目、排列方式、窗口类型等形态学基本一致, 细微区别在于本文报道种群体型普遍较前人报道种群略大。     

钩刺亚册虫的光镜和扫描电镜观察

作者对钩刺亚册虫（Ａｃｉｎｅｒｉａｕｎｃｉｎａｔａ）的纤毛器和形态结构做了光镜和电镜水平的研究,结果表明：口侧动基索均为单动基列结构,其中ＰＫ１仅后行至胞口后方;背触毛含约１０对紧密排列的毛基体,后端不与其它体动基列相联;射出体除沿胞口分布外,在虫体顶端丛集成盾片状;虫体前部自右向左扭转约１８０°。     

环境因子对旋毛蟹栖拟阿脑虫（Paranophrys carcini spiralis）生长繁殖的影响

在实验室条件下观察研究培养液的浓度、盐度、ｐＨ值及环境温度对旋毛蟹栖拟阿脑虫在单因子和综合因子作用下生长繁殖的影响,结果证明：在单因子作用条件下,随着培养液浓度的增加,其虫体种群的内禀增长率（ｒｍ）随之增加,达一定浓度时则停止增加,盐度在８—５５ｇ·Ｌ－１时,虫体可繁殖生长,３５ｇ·Ｌ－１时,ｒｍ最大;温度在４—３２℃范围内可生长繁殖,２５℃时ｒｍ最大;ｐＨ为５—９时,虫体可生长繁殖,ｐＨ为７时,ｒｍ最大。综合因子作用分析表明：温度与盐度对ｒｍ的影响显著,ｐＨ值在实验范围（５—９）无显著影响。当ｐＨ＝５—９、温度２２．１℃、盐度２２．４ｇ·Ｌ－１时,旋毛蟹栖拟阿脑虫的ｒｍ最大,而盐度和温度间具明显的交互作用。     

崇明岛潮间带夏季大型底栖动物多样性

2006年6月,在上海市崇明岛潮间带设置21个断面,进行了大型底栖动物定量定性采样调查.采集到大型底栖动物63 种,其中甲壳动物29种,占总种数的46.03%;软体动物20种,占31.75%;环节动物10种,占15.87%;底栖鱼类 2种;其它2种.优势种为无齿相手蟹、天津厚蟹、谭氏泥蟹、中华拟蟹守螺、堇拟沼螺、绯拟沼螺、光滑狭口螺和丝异蚓虫.大型底栖动物总平均丰度为138.28ind/m2,平均生物量为 79.11g/m2.系统聚类分析和MDS分析结果基本吻合,可以把21个断面分为3组,崇明北部断面群、崇明东部断面群和崇明南部(含西部)断面群.Shannon-Weiner指数H′＝1.85±0.528(范围0.946～2.783),Pielou均匀度指数J＝0.306±0.098(范围0.142～0.489),Simpson优势度指数D＝0.622±0.141(范围0.303～0.797).底栖动物的种类组成、丰度、生物量、多样性指数在各断面的差异主要与盐度分布、底质及人为干扰有关.与同期的历史资料相比,底栖动物总物种数变化不大,但种类组成有所变化;优势种有所变化;丰度和生物量均有所下降;多样性指数近年来变化不大.围垦减缓使崇明潮间带底栖动物多样性有所恢复,但在人为干扰和其他因素综合影响下,原为优势种的河蚬、彩虹明樱蛤、麂眼螺、小头虫和疣吻沙蚕等的密度下降,已不再是优势种.     

车轮虫齿体特征的量化及其种内种间系统发育的研究

齿体特征是车轮虫物种鉴定的一个重要依据。本文首次提出一种齿体结构面积比例量化法,即通过对齿钩、齿锥、齿棘、齿体环各部分反口面观面积的测量,分别计算出前三者与所在齿体的面积比以及所有齿体总面积与齿体环的面积比,从而将齿体特征转换为量化指标依据所测量化值,采用相似性聚类法,我们首先探讨了显著车轮虫(Trichodina nobbillis chen,1963)的三个种群的亲缘关系,然后以劳牧小车轮虫(Trichodinella lomi Xu,Song ＆ Warren,1999)为外类群,选择显著车轮虫(Trichodina nobillis chen,1963)、异齿车轮虫(Trichodina heterodentata Duncan,1977)、杜氏车轮虫(Trichodina dormergui Wallengren,1897)和Trchodina fahaka(Khaled A．S．Al-Rasheid,2000)为研究对象,对车轮虫种间亲缘关系进行了分析。结果表明,Pdr值可以有效地表现种群间的差异;同时还发现齿棘面积比例较高的杜氏车轮虫和异齿车轮虫进化较晚,而齿棘面积比例较低的显著车轮虫进化较早,并得出了齿棘越发达,车轮虫越进化这一结论,支持了徐奎栋等所提出的游走亚目六种基本齿型的进化顺序。这种量化研究方法不但可对车轮虫齿体的各部分结构以及它们之间的相互关系进行定量描述,而且还可以这些数据为基础对车轮虫种群间及物种间亲缘关系进行研究。     

海洋纤毛虫--水滴伪康纤虫的口器发生及形态学重描述

利用蛋白银法对采自山东胶州育虾池的一种海洋盾纤类纤毛虫,水滴伪康纤虫(Pseudocohnilembus persalinus Evans&Thompson,1964)的口器发生过程进行了详细的观察和研究,并对其形态学做了补足性描述。文章通过对该青岛种群发生过程的研究,认为前人所报道的种群(Evans&Thompson,1964;Pomp&Wilbert,1988)缺乏对某个发生关键时期的观察而存在着错误,即:后仔虫的小膜2明确来自老的口侧膜,而不是前人报道的盾片。此外,文章还发现该种的发生与本属另一哈氏伪康纤虫的发生过程几乎完全相同。主要细胞发生过程为:盾片最先增殖,形成初级原基区,然后分裂成前后两部分,前部分最终消失,而后部分最终形成后仔虫的小膜3。继盾片增殖之后口侧膜的锯齿状结构沿细胞纵轴方向分裂成两列,右侧的一列增殖形成次级原基区,之后分裂成前后两部分,前部分迁移形成后仔虫的口侧膜和盾片,后部分形成后仔虫的小膜1和小膜2;老口侧膜的残余部分形成前仔虫的口侧膜及盾片。老的小膜1、小膜2和小膜3则完全为前仔虫所继承。     

Insights into the phylogenetic and taxonomy of philasterid ciliates (Protozoa, Ciliophora, Scuticociliatia) based on analyses of multiple molecular markers

Scuticociliates are a rich assemblage of species with mostly unresolved phylogenetic relationships, especially in the order Philasterida. In the present work, 48 new sequences for three linked genes are characterized and phylogenetic trees are constructed to assess the inter- and intra-generic relationships of philasterids. Results reveal the following: (1) the combined three-gene tree provides more resolution in nodes than in the SSU-rDNA topologies; (2) the family Orchitophryidae is non-monophyletic as it is split into two parts and Paranophrys magna, Metanophrys sp. and Metanophrys sinensis are designated incertae sedis at the familial level; (3) Uronematidae is non-monophyletic and Homalogastra setosa is designated incertae sedis; (4) Parauronematidae becomes a junior synonym of Uronematidae and the clade containing A. haemophila, Miamiensis avidus, and Glauconema trihymene might stand for a new family; (5) Parauronema being a junior synonym of Uronema is supported and P. longum should be removed from the genus Parauronema; (6) Uronema is not monophyletic and molecular analyses reveal that Uronema sp. QD shares a more recent common ancestor with Uronemella species than with other Uronema species; (7) Metanophrys is polyphyletic; (8) multiple samples of two highly controversial species, viz., Mesanophrys pugettensis and M. chesapeakensis have identical ITS1-5.8S-ITS2 region sequence and we propose they should be synonymous with M. carcini, and (9) there may be cryptic species in M. carcini and M. avidus.     

Complete small subunit rRNA gene sequence of the scuticociliate Miamiensis avidus pathogenic to olive flounder Paralichthys olivaceus

Eight isolates of Miamiensis avidus (scuticociliates) were collected from olive flounder Paralichthys olivaceus with symptoms of severe ulcers and haemorrhages at several culture farms in 1999 and 2003. Cloned strains were produced and the complete small subunit ribosomal RNA gene (SSU rRNA) of each strain was sequenced for classification and phylogenic study. The SSU rRNA is 1759 bp in length and the sequence was deposited in the GenBank under accession number AY550080. All 8 strains exhibited the same sequence, but this sequence did not match any previously deposited scuticociliate SSU rRNA sequence. Phylogenetic analysis placed Miamiensis avidus in a sister lineage to Cohnilembus verminus, Pseudocohnilembus hargisi and P. marinus.     

Analysis of the internal transcribed spacer 2 (ITS2) region of scuticociliates and related taxa (Ciliophora, Oligohymenophorea) to infer their evolution and phylogeny

The ITS2 (ITS--internal transcribed spacer) region of the rDNA in 11 representative scuticociliates and two ambiguously related genera was analyzed. In common with other eukaryotes, the putative ITS2 folding pattern consists of a closed loop with four helices supported by minimum free energy and compensatory base changes (CBCs), although two of these helices are variable and sometimes absent. Three topologies were obtained on the basis of traditional primary sequence analysis, "string" strategy of secondary structure and analysis of the combined data. It was found that the secondary structure information could help to improve alignment and utilize appropriately phylogenetic strategies. The proposed phylogenies, though differing between sequence- and structure-based results, provide consistent support for high-level clades: the systematically questionable genera Dexiotrichides and Cardiostomatella always cluster together in a clade basal to the scuticociliates s.s., whereas Pleuronema branches from other uronematids at a deep level, and is hence a divergent taxon. Within the well-supported monophyletic philasterids, a sister relationship exists between Orchitophrya and Mesanophrys, while Uronema shows a close relationship with the group including Paranophrys and Parauronema. The positions of Metanophrys, Pseudocohnilembus and Anophryoides among the philasterids remain poorly resolved. Our findings firmly support the proposed evolutionary scenario inferred previously both from morphological and molecular data.     

Morphological studies indicate that Pleuronema grolierei nov. spec. and P. coronatum Kent, 1881 represent different sections of the genus Pleuronema (Ciliophora: Scuticociliatida)

The morphology, infraciliature and silverline system of two marine scuticociliates, Pleuronema grolierei nov. spec. and Pleuronema coronatum Kent, 1881, isolated from the sand beaches along the coast of Qingdao, China, were investigated using live observation and silver impregnation methods. Ciliates of the genus Pleuronema are normally easily recognized by their large sail-like paroral membrane. In this respect, P. coronatum is a typical member of the genus, whereas P. grolierei is unique in having much shorter cilia in the paroral membrane, which, in consequence, is rather inconspicuous. Nevertheless, details of the infraciliature confirm that P. grolierei belongs to the genus Pleuronema. Within the genus Pleuronema, the posterior end of the anterior fragment of the second membranelle (M2a) may be either straight, as in P. grolierei, or hooked-shaped, as in P. coronatum, providing a criterion for recognizing two sections within the genus. Based on the current study and previous reports, we propose that three nominal forms, Pleuronema balli Small and Lynn, 1985, Pleuronema smalli Dragesco, 1968 and Pleuronema borrori Dragesco, 1968 should be synonymized with P. coronatum.     

Morphology and Phylogeny of Three Pleuronema Species (Ciliophora,Scuticociliatia) from Hangzhou Bay,China, with Description of Two New Species,P. binucleatum n. sp. and P. parawiackowskii n. sp.

The morphology and phylogeny of Pleuronema binucleatum n. sp., P. parawiackowskii n. sp., and P. marinum Dujardin 1841, collected from Hangzhou Bay estuary, China, were investigated using standard methods. Pleuronema binucleatum n. sp. can be identified by possessing about 90–120 × 35–50 μm cell size in vivo, reniform body outline, two macronuclei, six to eight preoral kineties, 32–41 somatic kineties, and posterior end of the anterior fragment of membranelle 2 (M2a) hook‐like. Pleuronema parawiackowskii n. sp. is characterized by the combination of the following characters: body size about 60–90 × 20–40 μm in vivo, elliptical in outline, four to eight preoral kineties, 20–29 somatic kineties, posterior portion of the M2a slightly curved but nonhooked, and single macronucleus sausage‐like. After comparison with other populations of P. marinum, it is suggested that many misidentifications exist in previous studies. And an improved diagnosis of P. marinum was supplied: cell about 95–180 μm long, elliptical in outline; 2–4 preoral kineties and 53–70 somatic kineties; both membranelle 1 and membranelle 3 three‐rowed; posterior end of the M2a straight; single contractile vacuole characteristically positioned near mid‐body. The small subunit rRNA genes of three forms were sequenced. Phylogenetic analyses indicate that the monophyly of the genus Pleuronema is still not supported.     

Descriptions of two new marine scuticociliates,Pleuronema sinica n. sp. and P. wilberti n. sp. (Ciliophora: Scuticociliatida), from the Yellow Sea,China

Two new marine scuticociliates, Pleuronema sinica n. sp. and P. wilberti n. sp., collected from the sand beach of Qingdao, China, were investigated in vivo and following protargol impregnation. Ciliates of the genus Pleuronema are normally recognizable by their large sail-like paroral membrane although one species, P. grolierei, has shorter cilia in the paroral membrane. Neither of the new forms has the conspicuous paroral membrane in vivo so in this respect they are not typical members of this genus. Pleuronema sinica is characterized by its large, conspicuously flattened body, the possession of only one preoral kinety, the irregular-shaped macronucleus and the rather unusual structure of the oral apparatus. By contrast P. wilberti has a medium-size broad-oval body, six to eight preoral kineties and a highly differentiated membranelle 3 that is five- or six-rowed. An identification key is supplied for the 15 species of Pleuronema for which the infraciliature is known.     

The Cultivation of Symbiote-free Marine Ciliates in Axenic Medium

SYNOPSIS. Media have been developed for axenic cultivation of 10 strains belonging to 7 species of small marine ciliates. A medium containing cerophyl extract, proteose peptone, trypticase, yeast nucleic acid, biotin, calcium pantothenate folic acid, nicotinamide, pyridoxal HCl, riboflavin, thiamine HCl, and DL-thioctic acid in sea water supports the growth of Uronema nigricans strains Pc and 34/2, Parauronema virginiatum strains 2/1 and 19/1, Miamiensis avidus strains Ma and 19/3, Miamiensis sp. strain 1/1, a strain of "G" ciliate, and strains 33/8 and 34/7 of 2 unidentified species. By substituting a mixture of asolecithin, cephalin, and Tween 80 for cerophyl in the medium, luxurious growth of all except the strains of the 2 unidentified species can be obtained. A defined medium consisting of 18 amino acids, 5 purine derivatives, 8 vitamins, asolecithin, cephalin, and Tween 80 in synthetic sea water also has been developed for 6 of the strains: M. avidus Ma and 19/3, Miamiensis sp. 1/1, P. virginiatum 2/1 and 19/1, and U. nigricans Pc. In general the ciliates grow best at pH 7.2 in the dark at 27 C in media containing sea water of density = 1.015. Under these conditions maximum populations are reached in 4–5 days and range from several hundred thousand to 3 or 4 × 10⁶ depending upon the strain. Electronmicroscopic observations for the presence of endosymbiotes gave negative results.     

Miamiensis avidus (Ciliophora: Scuticociliatida) causes systemic infection of olive flounder Paralichthys olivaceus and is a senior synonym of Philasterides dicentrarchi

The scuticociliate Miamiensis avidus was isolated from olive flounder Paralichthys olivaceus showing typical symptoms of ulceration and hemorrhages in skeletal muscle and fins. In an infection experiment, olive flounder (mean length: 14.9 cm; mean weight: 26.8 g) were immersion challenged with 2.0 x 10(3), 2.0 x 10(4) and 2.0 x 10(5) ciliates ml-1 of the cloned YS1 strain of M. avidus. Cumulative mortalities were 85% in the 2.0 x 10(3) cells ml-1 treatment group and 100% in the other 2 infection groups. Many ciliates, containing red blood cells in the cytoplasm, were observed in the gills, skeletal muscle, skin, fins and brains of infected fish, which showed accompanying hemorrhagic and necrotic lesions. Ciliates were also observed in the lamina propria of the digestive tract, pharynx and cornea. The fixed ciliates were 31.5+/-3.87 pm in length and 18.5+/-3.04 microm in width, and were ovoid and slightly elongated in shape, with a pointed anterior and a rounded posterior, presenting a caudal cilium. Other morphological characteristics were as follows: 13 to 14 somatic kineties, oral ciliature comprising membranelles M1, M2, M3, and paroral membranes PM1 and PM2, contractile vacuole at the posterior end of kinety 2, shortened last somatic kinety and a buccal field to body length ratio of 0.47+/-0.03. In addition, continuous PM1 and PM2, lack of M3 and variable kinetosome numbers in M2 and M3 were frequently observed. Specimens in the current study were compared with previous reports on M. avidus and Philasterides dicentrarchi and confirmed consistently that these 2 taxa are conspecific.     

A parasitic scuticociliate infection in the Norway lobster (Nephrops norvegicus)

A histophagous ciliate infection was discovered in a number of Norway lobsters (Nephrops norvegicus) from the Clyde Sea Area, Scotland. Silver-carbonate staining of cultured ciliates revealed an oral apparatus and additional structural features that are morphologically similar to scuticociliates in the genus Mesanophrys, which are known to parasitize crustaceans. However, ribosomal DNA sequences (ITS1/5.8S/ITS2) of the ciliate were identical to Orchitophyra stellarum, a parasitic scuticociliate of sea stars with a different morphology from Mesanophrys spp. and to the ciliate from N. norvegicus. Associated pathology included degeneration and necrosis of the myocardial heart muscle, and large numbers of ciliates in the gill filaments.