长江上游圆口铜鱼幼鱼肠道内寄生小吻虫种群生态的研究

对长江上游圆口铜鱼幼鱼肠道内寄生小吻虫种群生态的研究表明,长江上游圆口铜鱼幼鱼体内小吻虫感染率为69．88％,感染强度8．18(1-47),相对密度5．72。雌鱼的感染率和感染强度均高于雄鱼,体长4．0～5．9cm的圆口铜鱼被小吻虫的感染率最高,且染虫对肥满度有显著影响。小吻虫在圆口铜鱼幼鱼种群中呈负二项分布,66％的虫体集丛分布在圆口铜鱼肠道的前段,其种群主要由5mm以下个体组成,雌虫明显多于雄虫。     

铜鱼和圆口铜鱼的DNA含量

有关鱼类细胞核DNA含量的研究,国外有较多的报道,国内自八十年代初陆续开展了这方面的工作,并通过测量DNA含量来探讨鱼类种群关系〔2〕、倍性关系〔1〕和杂交育种等。现采用血涂片,Feulgen染色,显微吸收光度法对长江中的铜鱼Coreius heterodon和圆口铜鱼Coreius guichenoti的红细胞核DNA含量进行了初步测定。1材料与方法材料于1989年5月取自长江上游的四川省泸州市,每种鱼各取5尾。1.1制片用肝素作抗凝剂,用注射器从鱼尾静脉取血,涂布于洁净载玻片的一端,用公鸡血细胞作对照标准,即从公鸡腋静脉取血,涂布于该载玻片上的另一端。涂片空…     

铜鱼和圆口铜鱼的早期发育

本文比较系统地观察了长江和汉江铜鱼和圆口铜鱼鱼卵、鱼苗的胚胎发育,记述了两种铜鱼胚胎发育的各个阶段的形态特征,比较两种铜鱼早期发育的相似性状和主要差别。这些差别主要是：雏形胚体出现的迟早、眼基和肌节出现的先后、眼睛大小、胸鳍长短以及颌须出现的迟早和长短等,依据这些差异,可以把两种铜鱼在早期发育阶段准确地区分开来。本文还对长江兴修水利枢纽与两种铜鱼资源的关系进行了调查和分析。     

蛇(鱼句)感染多子小瓜虫病一例

2003年6月18日从长江重庆木洞段购回十几条活泼蛇(鱼句)Saurogobio dabryi Bleeker.均由罾网捕捞,健康无损伤.放在室外水泥池饲养.室外水温白天26℃左右 ,夜晚24℃左右, 6月26日在常规检查时又发现蛇(鱼句)身上出现小白点,病鱼体色发黑、消瘦、反应迟钝、体表粘液增多、浮头、体表和鳃遍布小白点;感染后期表皮有脱落现象.取病鱼体表一个白点镜检,可见虫体全身密布短而均匀的纤毛;大核呈马蹄形,确定为多子小瓜虫Ichthyophthirius multifiliis Fouquet.感染小瓜虫病的蛇(鱼句)7月6日死亡.蛇(鱼句)是否能大面积发病还有待进一步研究.     

基于微卫星标记的圆口铜鱼亲子鉴定技术

为快速有效地鉴别不同的圆口铜鱼家系及来源, 研究从已发表的40个微卫星标记中筛选出20个多态性较高且稳定扩增的微卫星位点, 通过对8个圆口铜鱼家系339尾个体进行微卫星基因分型检测, 建立了圆口铜鱼荧光微卫星标记与多重毛细管电泳相结合的亲子鉴定技术。遗传多样性分析结果显示, 圆口铜鱼8个家系群体的平均等位基因数(N_a)为9个, 平均多态信息含量(PIC)为0.616, 平均期望杂合度(H_e)为0.659, 平均观测杂合度(H_o)为0.691, 其中子一代群体的遗传多样性水平明显低于亲本群体。亲子鉴定分析结果显示, 当双亲基因型未知时其单亲累积排除概率(CE-1P)为0.99954473, 当单亲基因型已知时其累积排除概率(CE-2P)为0.99999825, 当双亲基因型未知时其双亲累积排除概率(CE-PP)为1.00000000, 当使用20个微卫星位点进行亲子鉴定时, 297尾子一代均能正确找到其父母本, 亲子鉴定准确率为100%。由此可见, 研究建立的圆口铜鱼亲子鉴定技术是可靠的, 能为圆口铜鱼的家系管理、种群遗传管理和增殖放流效果评估提供科学依据     

蛇(鱼旬)感染多子小瓜虫病一例

20 0 3年 6月 1 8日从长江重庆木洞段购回十几条活泼蛇SaurogobiodabryiBleeker。均由罾网捕捞 ,健康无损伤。放在室外水泥池饲养。室外水温白天2 6℃左右 ,夜晚 2 4℃左右 ,6月 2 6日在常规检查时又发现蛇身上出现小白点 ,病鱼体色发黑、消瘦、反应迟钝、体表粘液增多、浮头、体表和鳃遍布小白点 ;感染后期表皮有脱落现象。取病鱼体表一个白点镜检 ,可见虫体全身密布短而均匀的纤毛 ;大核呈马蹄形 ,确定为多子小瓜虫IchthyophthiriusmultifiliisFouquet。感染小瓜虫病的蛇 7月 6日死亡。蛇是否能大面积发病还有待进一步研究蛇(鱼…     

葛洲坝水利枢纽兴建后长江干流铜鱼和圆口铜鱼的繁殖生态

本文报道了葛洲坝水利枢纽兴建后,长江干流铜鱼和圆口铜鱼的产卵场分布、产卵规模、产卵季节以及促使产卵的主要外界条件等。指出葛洲坝水利枢纽截流后,大坝附近江段铜鱼产卵场的变化情况。分析和讨论了大坝对两种铜鱼的影响程度、两种铜鱼产卵与水文条件的关系及其资源的保护等问题。     

长江上游圆口铜鱼生长方程的分析

根据2005～2007年在长江上游鱼类资源调查中采集到的476尾圆口铜鱼Coreius guichenoti Sauvage et Dabry标本的数据,对其生长方程进行了分析.圆口铜鱼鳞片半径-体长的函数关系在雌雄群体间不存在显著性差异并符合方程L=85.429S0.8125 (R2=0.9635),推算体长表现出Lee氏现象;运用非线性混合模型方法,对基于4种备选生长方程(von Bertalanffy生长方程(VBG)、Richards生长方程、Gompertz生长方程和Robertson生长方程)的36种不同模型分别以生长推算数据进行拟合,并根据AIC选择最优生长模型,得到不同备选方程对圆口铜鱼生长描述的优劣顺序为:VBG、Richards、Gompertz、Robertson方程,其中最优生长模型为VBG Model 6:Lt=602.9(1-e-0.1693(t+0.024)) (雌雄群体间在0.01水平上不存在显著性差异).圆口铜鱼的生长特征指数φ=4.7891±0.01612.对于存在Lee氏现象的鱼类,在使用生长推算数据构建生长方程时,建议使用非线性混合模型以对参数值进行较合理的估计.本文为圆口铜鱼种群动态及资源保护提供了基础资料,并为不同种鱼类、同种鱼类不同生活史阶段表现出的不同生长模式的识别提供了一个范例.     

三峡水库蓄水后铜鱼和圆口铜鱼肌肉和肝脏中重金属水平

为了解三峡水库蓄水后鱼体重金属富集现状及其潜在的生态风险和食品安全, 测定了三峡水库上、中、下游不同年龄组铜鱼(Coreius heterodon)和圆口铜鱼(C. guichenoti)肌肉和肝脏中重金属含量。利用等离子吸收光谱法、石墨炉原子吸收光谱法、原子荧光光谱法检测样品中Cu、Zn、Cr、Pb、Cd、Hg、As 等 7种重金属含量。结果表明: 7 种重金属在铜鱼和圆口铜鱼体内的含量水平基本一致, 重金属在铜鱼和圆口铜鱼肌肉中含量大小均为Zn Cu Cr Hg AsPb Cd, 在铜鱼肝脏中含量大小为Zn Cu Pb Cd Cr As Hg, 而在圆口铜鱼肝脏中含量大小为Zn Cu Pb Cr Cd Hg As。铜鱼和圆口铜鱼肝脏中重金属含量显著高于肌肉(P0.05)。两种鱼类间大部分重金属在库区上、中、下游无显著差异(P 0.05)。铜鱼和圆口铜鱼肌肉(可食用部分)中7 种重金属含量均未超过国家食品安全卫生标准, 属于安全食用范围;肝脏中除Cd 和Pb 外的其他元素含量均未超过国家标准。相关结果反映了三峡水库175 m 蓄水后底栖土著经济鱼类重金属的污染状况, 对了解该地区水产品质量安全状况及水产品安全评价提供了参考依据。       

长薄鳅感染多子小瓜虫一例

2004年12月5日从长江重庆木洞段购回活泼长薄鳅(Leptobobitia elongata Bleeker)8尾,健康无伤。放在室内水泥池和圆口铜鱼混合饲养。室内水温白天14℃左右,夜晚9℃左右,2005年1月13日在常规检查时发现3尾长薄鳅身上出现小白点(封3,图版),反应迟钝、体表粘液增多;白点出现在鱼体的头部、鳃片、背鳍、胸鳍、腹鳍、臀鳍、尾鳍,身体两侧和尾柄部白点较多;感染后期表皮有脱落现象。取病鱼体表白点镜检,可见虫体呈圆形或椭圆形,作滚动运动,身体遇障碍物可变形,虫体全身密布短而均匀的纤毛。经虫体固定、染色制片测量和观察,     

Ichthyophthirius multifiliis Has Membrane-Associated Immobilization Antigens

Sera from fish that survive infections with the ciliated protozoon, Ichthyophthirius multifiliis , immobilize the parasite in vitro. In order to identify ceil surface antigens involved in the immobilization response, integral membrane proteins were extracted from tomites with Triton X-l14 and used to immunize rabbits. The rabbit antisera immobilized the parasite in vitro and antigens were localized to cell and ciliary plasma membranes by indirect immunofluorescent microscopy. The membrane protein fractions from both whole cells and tomite cilia were characterized by 1 - and 2-dimensiona! SDS-PAGE. A 43,000-dalton (D) glycoprotein with an isoelectric point of 7.0 is the predominant protein in these fractions, comprising 12% and 60% of the total protein of whole cell and ciliary membranes, respectively. Western blot analysis of ciliary proteins with immune rabbit sera indicated that the 43,000-D glycoprotein is the principal antigen.     

Two year study on the infectivity of Ichthyophthirius multifiliis in channel catfish Ictalurus punctatus

Ichthyophthirius multifiliis Fouquet (Ich) is a fish parasite that causes serious economic loss for aquaculture. A major difficulty in the maintenance of single isolates of Ich for research purposes is the loss of infectivity. After an unknown number of passages or infection cycles the Ich isolate loses its infectivity. This study determined the infectivity of an Ich isolate during 105 infection cycles in channel catfish Ictalurus punctatus over a 2 yr period. The mean percentage of fish infected by Ich, the infection levels and the time to trophont emergence were each compared after 4 cyclic periods: 1-25, 26-60, 61-90 and 91-105 Ich cycles. Results of this study demonstrated that Ich was significantly more infective (p < 0.05) at 1-25 than 26-105 cycles. Channel catfish were infected at a ratio of 1 infected fish to 8 naive fish at 1-25 and 26-60 cycles. A higher infection ratio occurred at 61-90 and 91-105 cycles. Trophont emergence was noted to be significantly longer at 91-105 compared to 1-25 cycles, during 7 and 5 d respectively, at 23.4 +/- 1.1 degrees C. The results of the present study indicate that the infectivity of I. multifiliis started to decrease after 25 infection cycles and was predominant in the single Ich isolate at 61-90 and 91-105 cycles.     

The I-antigens of Ichthyophthirius multifiliis are GPI-Anchored Proteins

The parasitic ciliate Ichthyophthirius multifiliis has abundant surface membrane proteins (i-antigens) that when clustered, trigger rapid, premature exit from the host. Similar antigens are present in free-living ciliates and are GPI-anchored in both Paramecium and Tetrahymena. Although transmembrane signalling through GPI-anchored proteins has been well-documented in metazoan cells, comparable phenomena have yet to be described in protists. Since premature exit of Ichthyophthirius is likely to involve a transmembrane signalling event, we sought to determine whether i-antigens are GPI-anchored in these cells as well. Based on their solubility properties in Triton X-114, the i-antigens of Ichthyophthirius are amphiphilic in nature and partition with the detergent phase. Nevertheless, following treatment of detergent lysates with phospholipase C, the same proteins become hydrophilic. Concomitantly, they are recognized by antibodies against a cross-reacting determinant exposed on virtually all GPI-anchored proteins following cleavage with phospholipase C. Finally, when expressed in recombinant form in Tetrahymena thermophila, full-length i-antigens are restricted to the membrane, while those lacking hydrophobic C-termini are secreted from the cell. Taken together, these observations argue strongly that the i-antigens of Ichthyophthirius multifiliis are, in fact, GPI-anchored proteins.     

Differences in virulence between two serotypes of Ichthyophthirius multifiliis

Naive channel catfish Ictalurus punctatus were infected by 2 isolates of the parasitic ciliate Ichthyophthirius multifiliis that differed in virulence. The isolates, NY1 and G5, Serotypes A and D, respectively, express different surface immobilization-antigens. The virulence of the 2 isolates was compared using tail-fin infections to quantitate parasite numbers and by analysis of the survival of infected fish. Although NY1 infected fish at a lower level than G5, all NY1-infected fish died, but 51% of G5-infected fish survived. The greater virulence of NY1 is apparently a consequence of its shorter life cycle, which results in overwhelming reinfection of fish before they can develop a protective immune response. This report represents the first experimental evidence for differences in virulence between serotypes of I. multifiliis.     

Endosymbiotic Bacteria in the Parasitic Ciliate Ichthyophthirius multifiliis

Endosymbiotic bacteria were identified in the parasitic ciliate Ichthyophthirius multifiliis, a common pathogen of freshwater fish. PCR amplification of DNA prepared from two isolates of I. multifiliis, using primers that bind conserved sequences in bacterial 16S rRNA genes, generated an ∼1,460-bp DNA product, which was cloned and sequenced. Sequence analysis demonstrated that 16S rRNA gene sequences from three classes of bacteria were present in the PCR product. These included Alphaproteobacteria (Rickettsiales), Sphingobacteria, and Flavobacterium columnare. DAPI (4′,6-diamidino-2-phenylindole) staining showed endosymbionts dispersed throughout the cytoplasm of trophonts and, in most, but not all theronts. Endosymbionts were observed by transmission electron microscopy in the cytoplasm, surrounded by a prominent, electron-translucent halo characteristic of Rickettsia. Fluorescence in situ hybridization demonstrated that bacteria from the Rickettsiales and Sphingobacteriales classes are endosymbionts of I. multifiliis, found in the cytoplasm, but not in the macronucleus or micronucleus. In contrast, F. columnare was not detected by fluorescence in situ hybridization. It likely adheres to I. multifiliis through association with cilia. The role that endosymbiotic bacteria play in the life history of I. multifiliis is not known.The ciliate Ichthyophthirius multifiliis is an obligate parasite of freshwater fish that infects epithelia of the skin and gills. The life cycle of I. multifiliis consists of three stages: an infective theront, a parasitic trophont, and a reproductive tomont. Infection is initiated by invasion of the skin and gills by free-swimming, 40-μm-long, pyriform-shaped theronts that burrow several cell layers deep into epithelial tissue of the skin and gills and rapidly differentiate into trophonts. Trophonts feed on epithelial cells and grow into 500- to 800-μm-diameter cells, causing extensive damage to skin and gills, which in severe infections results in mortality (10-12). After feeding for 5 to 7 days, trophonts leave the host, form encysted tomonts, and undergo up to 10 cell divisions over 18 to 24 h, producing as many as 10³ daughter cells, which exit the cyst as infective theronts to reinitiate the life cycle. I. multifiliis is ciliated at all stages (9).DNA sequencing of the I. multifiliis genome at the J. Craig Venter Institute unexpectedly revealed that bacterial DNA sequences, including sequences with homology to Rickettsia, were present in the DNA preparations (R. S. Coyne, 2009 [http://www.jcvi.org/cms/research/projects/ich/overview]). The origin of these sequences was unclear, but they represented evidence for either horizontal gene transfer into the I. multifiliis genome (17, 27) or the presence of intracellular bacteria. No previous evidence suggested the presence of intracellular bacteria in I. multifiliis, even though the fine structure of I. multifiliis theronts and trophonts has been examined by transmission electron microscopy (10-12). Intracellular or endosymbiotic bacteria, however, are commonly found in protists, and about 200 ciliate species are known to harbor intracellular bacteria (13, 15). Sonneborn and Preer in their classic studies on endosymbionts in Paramecium characterized a number of different endosymbionts, including “killers,” named for their ability to kill uninfected strains of Paramecium. Cytoplasmic endosymbionts in Paramecium now include Caedibacter taeniospiralis (Gammaproteobacteria), and Pseudocaedibacter conjugates, Tectibacter vulgaris, and Lyticum flagellatum (Alphaproteobacteria). Macronuclear endosymbionts include the Alphaproteobacteria, Holospora caryophila, and Caedibacter caryophila, which can also infect the cytoplasm (4, 16, 22, 26). The roles these endosymbionts play in protists are not well understood.The presence of sequences with homology to bacterial genomes prompted us to determine if I. multifiliis contained endosymbionts, or if these sequences represented evidence for horizontal gene transfer into the I. multifiliis genome. Our identification of the same two endosymbionts, in two different isolates of I. multifiliis, suggests that endosymbionts are common in I. multifiliis. However, the physiological relationships between I. multifiliis and its resident endosymbionts are unclear. It is not known if the endosymbionts contribute to the growth of I. multifiliis, if they contribute to the severity or pathogenicity of infection, or if they provide their host with any selective advantage, as occurs with Paramecium containing killer particles (4). It has not been determined if they influence the immune response of fish infected with I. multifiliis. It is possible that they may simply be parasites of this parasitic ciliate.     

Reproduction of Ichthyophthirius multifiliis (Ciliata, Ophryoglenidae)

A case of reproduction of Ichthyophthirius multifiliis in the superficial tissues of larvae and fry of Amur wild carp is described. At a temperature of water from 28 to 29 degrees trophonts of Ichthyophthirius encysted on fishes. Inside cysts repeated cell division occurred but this process did not result in swarm spores formation. Later on with the increase of temperature to 29.5--31.5 degrees cysts degenerated.     

Purification and Partial Characterization of Immobilization Antigens from Ichthyophthirius multifiliis

ABSTRACT Ichthyophthirius multifiliis , a parasitic ciliate of freshwater fishes, was found to have surface antigens (Ag) which elieited immobilizing antibodies (Ab) when injected into rabbits. An effort was made to purify and characterize these Ag (referred to as immobilization Ag) because of their potential role in protective immunity in fishes. Mice immunized with theront cilia were used for production of immobilizing monoclonal antibodies (MAb). Hybridomas were screened by indirect immunofluorescent light microscopy and immobilization of live parasites. Six hybridomas producing immobilizing MAb were cloned. Immobilizing MAb were used to affinity purify Ag solubilized with Triton X-114 and Na deoxycholate. Two membrane protein Ag of approximately 48 and 60 kDa were identified. Immobilizing MAb failed to react with these Ag on Western blots and, conversely, MAb that reacted with the Ag on Western blots did not immobilize live organisms. These results suggest that immobilization required native conformational epitopes which were altered by Western blotting procedures. Individual MAb reactive on Western blots recognized both the 48- and 60-kDa proteins indicating the presence of common epitopes. Affinity purified Ag elicited immobilizing antisera when injected into rabbits, mice, and channel catfish.     

Ichthyophthirius multifiliis (Ciliophora) exit from gill epithelium

The first change in the sequence of morphological events occurring as fully developed Ichthyophthirius multifiliis trophonts spontaneously left gill epithelium or as younger trophonts departed, following experimentally induced death of the fish, was the separation of parasites from overlying host cells. Discharge of contractile vacuoles may have played a role in this process. Spaces then appeared between host cells, and individual epithelial cells became vacuolated. Finally, the epithelium ruptured and the parasites swam free. In induced exit after three days residence in the host, departure of the trophont was evident only after autolysis of epithelium had occurred. Induced departure of trophonts after four days residence was more rapid, suggesting an active role for the parasite in exit. Changes in parasite and epithelium observed in induced exit were similar to those in spontaneous departure after five days residence.