本文選題：寄生蟲　切入點：纖毛蟲　出處：《華東師范大學(xué)》2017年博士論文　論文類型：學(xué)位論文

【摘要】：刺激隱核蟲(CryptocaryonirritansBrown,1951)是一種寄生于幾乎所有海水硬骨魚類原生動物纖毛蟲,可引發(fā)宿主魚類刺激隱核蟲病,給水產(chǎn)養(yǎng)殖業(yè)造成巨大的危害。近年來,重新認(rèn)識刺激隱核蟲的基礎(chǔ)生物學(xué)特征和病原特性,又受到了人們的廣泛關(guān)注。刺激隱核蟲具有相對復(fù)雜的生活史,可分為:寄生于宿主組織中、攝食營養(yǎng)的滋養(yǎng)體時期;脫離宿主后在水中短暫自由運動的包囊前體時期;在包囊內(nèi)進(jìn)行細(xì)胞分裂增殖的包囊時期;細(xì)胞分裂產(chǎn)生的幼蟲脫出包囊、再次感染入侵宿主的幼蟲時期。目前,關(guān)于刺激隱核蟲的生活史中各種細(xì)胞器的形態(tài)和結(jié)構(gòu),及其在不同生活史時期間的演化規(guī)律,仍缺乏較為全面深入的認(rèn)識;對一些關(guān)鍵的生活史階段或生命活動過程中所涉及的細(xì)胞器及其生物學(xué)功能,以及相關(guān)的細(xì)胞學(xué)機(jī)制尚不十分清楚。另外,雖然前人的研究己基本確定刺激隱核蟲屬于前口類,但其在前口類中具體的系統(tǒng)分類地位尚存爭議,缺乏必要的形態(tài)學(xué)證據(jù)。本研究選用海水養(yǎng)殖大黃魚作為感染宿主,建立刺激隱核蟲傳代培養(yǎng),對刺激隱核蟲各生活史時期內(nèi)不同生命活動階段的細(xì)胞,采用活體觀察、銀染技術(shù)、熒光標(biāo)記技術(shù)、掃描電鏡技術(shù)、透射電鏡技術(shù)以及酶細(xì)胞化學(xué)技術(shù),從顯微、亞顯微水平進(jìn)行了形態(tài)學(xué)觀察,并通過測序建樹對刺激隱核蟲進(jìn)行了分子系統(tǒng)學(xué)分析。對多種胞器或結(jié)構(gòu)進(jìn)行了全面系統(tǒng)的描述,發(fā)現(xiàn)了較多未報道過的結(jié)構(gòu)或細(xì)胞生命活動現(xiàn)象特征,明確了刺激隱核蟲的分類地位并給出了形態(tài)學(xué)定義,為進(jìn)一步完善并深入闡明刺激隱核蟲及魚類寄生纖毛蟲的細(xì)胞形態(tài)學(xué)和基礎(chǔ)生物學(xué)特征提供了新的資料。主要結(jié)果如下:1刺激隱核蟲各生活史時期細(xì)胞口區(qū)相關(guān)結(jié)構(gòu)或細(xì)胞器除刺激隱核蟲滋養(yǎng)體、包囊前體細(xì)胞外,幼蟲細(xì)胞胞口周緣也具有由口周雙動基纖毛和口乳頭粘聯(lián)而成的三聯(lián)體結(jié)構(gòu);三聯(lián)體結(jié)構(gòu)均與胞咽內(nèi)壁肋嵴一一對應(yīng),但不與口緣外側(cè)體纖毛列一一對應(yīng)。毒絲泡在滋養(yǎng)體和包囊前體細(xì)胞中不存在,在幼蟲細(xì)胞中定位于口乳頭及口區(qū)胞質(zhì)中,可能參與了感染入侵宿主魚的生命活動過程,毒絲泡前體分化形成于包囊內(nèi)細(xì)胞分裂后期仔細(xì)胞中。滋養(yǎng)體、包囊前體細(xì)胞胞口一側(cè)和幼蟲細(xì)胞反口面具有膜性褶皺結(jié)構(gòu),其間分布單動基系體纖毛列,該結(jié)構(gòu)可能由該區(qū)域體纖毛列相互靠近、纖毛列間表膜特化隆起形成,膜性褶皺前端不與胞口直接連通,可能與刺激隱核蟲的寄生特性有關(guān)。滋養(yǎng)體、包囊前體和幼蟲細(xì)胞均具有的口區(qū)纖毛刷為3段雙動基片段,生有短棒狀纖毛,包囊內(nèi)細(xì)胞分裂后期的仔細(xì)胞表面可見口區(qū)纖毛刷原基;刺激隱核蟲口區(qū)纖毛刷,與前口類其他類群該結(jié)構(gòu)特征均不相同。2刺激隱核蟲各生活史時期細(xì)胞表膜及表膜下相關(guān)結(jié)構(gòu)或細(xì)胞器及細(xì)胞核刺激隱核蟲的表膜泡不同于自由生纖毛蟲的表膜泡,結(jié)構(gòu)較為復(fù)雜,不同的生活史階段具有不同的形態(tài)特征:在滋養(yǎng)體攝食營養(yǎng)階段,表膜泡內(nèi)容物豐富,其中一類雙層膜小泡可通過膜融合的形式通過小孔與細(xì)胞外相通,表膜泡可能具有從細(xì)胞外攝取液態(tài)或小顆粒營養(yǎng)物質(zhì)的功能,而攝取的營養(yǎng)物質(zhì)暫時積累于表膜泡中;正在脫離宿主組織的滋養(yǎng)體細(xì)胞,表膜泡中產(chǎn)生大量的絲狀物排出至細(xì)胞外;細(xì)胞形成包囊壁前后,表膜泡及其內(nèi)容物會分階段、團(tuán)塊狀地內(nèi)化進(jìn)入細(xì)胞質(zhì)。包囊內(nèi)細(xì)胞分裂過程中表膜泡內(nèi)容物較少,呈扁平狀態(tài)。幼蟲細(xì)胞表膜泡再次變得腫脹,內(nèi)容物可能來源于胞質(zhì)中高爾基體和轉(zhuǎn)運小泡的分泌活動。滋養(yǎng)體、包囊前體細(xì)胞中不具有黏液囊,成熟的黏液囊定位于幼蟲細(xì)胞表膜下,可能參與了幼蟲細(xì)胞入侵感染宿主魚以及入侵后建立寄生等生命活動,黏液囊前體分化形成于包囊內(nèi)細(xì)胞分裂后期仔細(xì)胞中。刺激隱核蟲具有多個伸縮泡,不同的生活史時期超微結(jié)構(gòu)有所不同,可能具有不同的生物學(xué)功能,可能在滋養(yǎng)體時期發(fā)揮較為重要的功能。刺激隱核蟲大核在不同的生活史時期形態(tài)特征不同,在形成包囊時念珠狀的大核融合變形為球形,染色質(zhì)變得分散。3刺激隱核蟲的包囊現(xiàn)象刺激隱核蟲呈多層形態(tài)的包囊壁,各片層間形態(tài)結(jié)構(gòu)較為相似,可能具有類似的生化本質(zhì)。包囊壁形成過程中未見特異性小泡或特定射出胞器的分泌活動。細(xì)胞形成包囊過程中,體纖毛不發(fā)生退化或脫落,胞口及口周纖毛也不退化消失。包囊內(nèi)進(jìn)行細(xì)胞分裂時,會形成暫時性地長細(xì)胞鏈,細(xì)胞鏈長有正在生長分化的體纖毛,無胞口及原基。新的口原基出現(xiàn)于包囊內(nèi)細(xì)胞分裂后期斷裂開來的仔細(xì)胞表面。繁殖包囊胞質(zhì)胞器的生命活動非�；钴S,可見高爾基體及粘液囊、毒絲泡前體等大量的物質(zhì)和結(jié)構(gòu)的合成和分化。4刺激隱核蟲的系統(tǒng)分類刺激隱核蟲口區(qū)纖毛刷、膜性褶皺、咽微纖絲等結(jié)構(gòu)形態(tài)特征不同于前口類其他科屬纖毛蟲,具有復(fù)雜的生活史和寄生特性,結(jié)合分子系統(tǒng)學(xué)建樹分析,表明刺激隱核蟲具有充足的證據(jù)支持其建立獨立的刺激隱核蟲科(Cryptocaryonidae),甚至具有多項目前分類所屬前管目(Prorodontida)不具有的細(xì)胞形態(tài)學(xué)特征。本研究系統(tǒng)學(xué)分析支持刺激隱核蟲存在種屬分化的觀點。綜上所述,本研究深入系統(tǒng)地揭示了刺激隱核蟲胞口和胞咽、毒絲泡、膜性褶皺結(jié)構(gòu)、口區(qū)纖毛刷、表膜泡、粘液囊、伸縮泡等胞器或結(jié)構(gòu)在各生活史時期的形態(tài)特征及其可能的生物學(xué)功能,初步闡明了一些重要的尤其是與纖毛蟲寄生特性相關(guān)的生命活動現(xiàn)象;首次對刺激隱核蟲的包囊現(xiàn)象進(jìn)行了多方面的描述討論;明確了刺激隱核蟲在前口類中的系統(tǒng)分類地位,并給出了刺激隱核蟲科形態(tài)學(xué)定義。為刺激隱核蟲細(xì)胞形態(tài)學(xué)和基礎(chǔ)生物學(xué)等的研究提供了新的基礎(chǔ)資料。
[Abstract]:Cryptocaryonirritans (CryptocaryonirritansBrown, 1951) is a parasite in almost all marine teleost ciliates, can cause the host fish cryptocaryonirritans disease, causing great harm to the aquaculture industry. In recent years, the basic biological characteristics and pathogenic characteristics of re understanding of cryptocaryonirritans, and has received extensive attention. Cryptocaryonirritans with life history is relatively complex, can be divided into: parasitic in the tissues of the host, the trophozoite feeding nutrition period; from the host in the water after a short free movement of the protomonts period; for cell proliferation in cysts within the cyst; cyst cell division produced from larvae, re infection the invasion of the host larvae period. At present, the morphology and structure of organelles on the life history of cryptocaryonirritans in the evolution and during different life history, is still missing The lack of more comprehensive and in-depth understanding of its biological functions; the organelles involved some life stages or life activities of key process, and the cytological mechanism is not very clear. In addition, although previous research has identified basic cryptocaryonirritans belong to the export category, but in the mouth before class specific the taxonomy is controversial, the lack of morphological evidence necessary. This study used seawater cultured large yellow croaker as host, the establishment of cryptocaryonirritans subcultured on different life stages of cryptocaryonirritans the life history within the cells, the in vivo observation, silver staining, fluorescence labeling technique, scanning electron microscopy transmission electron microscopy, and enzyme cytochemistry, from the microscopic, ultrastructural level were observed, and the cryptocaryonirritans of system analysis through sequencing of molecular contribution. A variety of organelles or structure of a comprehensive description of the system, found life structure or cell features more not reported, the taxonomic status of cryptocaryonirritans and gives the morphological definition, cell morphology and biological characteristics of the foundation to further improve and clarify cryptocaryonirritans and fish parasitic ciliates provides new data. The main results are as follows: 1 cryptocaryonirritans period the life history of cells or organelles related in structure in addition to c.irritans trophozoites, cyst precursor cells, larvae cell opening circumference also has three conjoined structure by mouth double action based adhesive and cilia and mouth nipple the three CIS structure and cell; pharyngeal wall rib ridge correspondence, but not with the edge of the outer body cilia column correspondence. Toxicyst does not exist in the trophozoites and cysts of precursor cells, in larval cells located in the mouth The nipple and mouth area in the cytoplasm of infected fish may be involved in the invasion of host life process, toxicyst precursor differentiation cysts formed in anaphase offspring cells. Trophozoites of the protomonts cells on one side of the opening and mouth mask anti larval cells membrane fold structure, the distribution of single action base body cilia, the structure may be provided by the regional body cilia listed near each other, cilia column membrane specialization uplift formation, membranous fold front end and cell port directly connected, may be related to the parasitic characteristics of c.irritans. Nourish the body, mouth area and brush the protomonts ciliated cells have larvae the 3 section double moving base fragment, having short rod like cilia, ciliated cells visible on the surface area of mouth Zi in anaphase brush primordium cyst; cryptocaryonirritans mouth area fiber brush, and in other groups before the structure characteristics are not the same.2 cryptocaryonirritans life Table table film film history during the period of the cell membrane and membrane associated structures or organelles and the nucleus of cryptocaryonirritans bubble is different from free ciliates in global, more complex structure, with the characteristics of different life history: in the different stages of the trophozoite feeding nutrition stage, form vesicles content rich, including a kind of double membrane vesicles by membrane fusion and cell form through hole is communicated with the outside surface, bubble may be from outside the cell uptake of liquid or small particles of nutrients, and nutrient uptake temporarily accumulated in membrane vesicles; trophozoite cells are separated from the host tissue, membrane vesicles produced in the table a large number of filaments is discharged to the outside of the cell; cell wall before and after the formation of cysts, surface membrane vesicles and its contents in stages, mass internalized into the cytoplasm. In the process of cell division in the cyst membrane bubble content is less, the flat state. Larvae of the cell membrane bubble again become swollen, the contents may originate from the Golgi secretory activity in cytoplasm and transport vesicles. Trophozoite, cyst precursor cells do not have the bursa, located in larvae under the cell membrane of mature bursa, may establish parasitic life activities involved in the cell invasion of infected larvae the host fish and after the invasion, bursa precursor differentiation within the cyst formed in anaphase cells. Zi c.irritans has multiple life history of global expansion, the ultrastructure of different, may have different biological functions, may play a more important function in the trophozoites of cryptocaryonirritans period. Nuclear in different life history of different characteristics, in the formation of cysts when torulose nuclear fusion deformation is spherical, chromatin became encystment dispersed.3 cryptocaryonirritans of c.irritans was The cyst wall multilayer form, each layer structure is similar, may have similar biochemical nature. In the process of no specific vesicles or specific extrusome secretion of cyst wall formation. Cell formation of cysts of ciliary body is not degraded or off, cytosome and perioral cilia did not disappear. Cell division cysts, will form a temporary cell long chain, cell chain length is the growth and differentiation of cells and ciliary body, no mouth primordium. New oral primordium appeared in the surface within the tomont anaphase apart young cells. The propagation of encapsulation of cytoplasmic organelles life is active visible, Golgi apparatus and bursa, classification system toxicyst synthesis and differentiation of.4 precursor material and the structure of a number of cryptocaryonirritans of cryptocaryonirritans mouth area fiber brush, membranous folds, pharyngeal microfibril structure morphology characteristic is not Is the same as the first class in other ciliates, with parasitic life history and complex characteristics, combined with the molecular phylogenetic analysis showed that the contribution that c.irritans has sufficient evidence to support the establishment of an independent cryptocaryonirritans Department (Cryptocaryonidae), even with multi project before the class before the tube (Prorodontida) cells. Morphological features do not have. This study phylogenetic analysis support cryptocaryonirritans species differentiation point of view. In summary, this study systematically reveals c.irritans and cytopharynx, toxicyst, membranous fold structure, mouth area fiber brush, membrane vesicles, bursa, expansion cell organelles or structure in the period of the life history of the morphological characteristics and biological functions of possible, clarify some important especially life phenomena associated with the characteristics of parasitic ciliates; first encystment of c.irritans. Various descriptions and discussions were made, and the taxonomic status of the stimulation of Cryptococcus in the anterior class was clarified. The morphological definition of the stimulation of Cryptococcus was provided. It provided a new basic information for stimulating the study of cell morphology and basic biology of Cryptococcus.

【學(xué)位授予單位】：華東師范大學(xué)
【學(xué)位級別】：博士
【學(xué)位授予年份】：2017
【分類號】：S941.5

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