缘毛类纤毛虫伸缩泡收缩频率的研究

纤毛虫伸缩泡的主要功能是排出进入体内的多余水分及部分代谢废物(Kudo,1966)。Kitching(1948),Tsukuda et al.(1984)等曾对纤毛虫伸缩泡的收缩频率进行研究,但国内尚未见这方面的报道。鉴于缘毛类纤毛虫对环境的生物监测及污染水体的生物治理中具有重要作用,为探索有关规律,我们测定了53种缘毛类纤毛虫伸缩泡的收缩频率以及盐度和温度对伸缩泡收缩频率的影响。材料和方法缘毛类纤毛虫(简称缘纤虫)多固着在水生动植物及杂质上,可通过采集水生动植物、水中杂质或水中挂片得到。将采得的被固着物直接放于显微镜下观察,鉴定种类,以秒表记录每…     

两种缘毛类纤毛虫胞质Hsp70基因序列的克隆及其系统发育分析

克隆得到2种缘毛类纤毛虫——钟形钟虫(Vorticella campanula)和螅状独缩虫(Carchesium polypinum)的胞质Hsp70基因部分序列,长度均为438bp,编码146个氨基酸。以细菌为外类群,利用最大似然法和邻接法构建包括其他5种纤毛虫在内的共26个物种的Hsp70基因氨基酸序列系统发育树,其拓扑结构显示：V．campanula和C．polypinum聚在一起,并与另2种寡膜纲的嗜热四膜虫(Tetrahymena thermophila)及草履虫(Paramecium tetraurelia)聚为姊妹枝,提示了缘毛类纤毛虫为单系,且隶属于寡膜纲的系统发育地位。     

四种淡水缘毛类纤毛虫的形态学研究

通过活体显微观察及银染法对采自武汉东湖的4种固着缘毛类纤毛虫: 粗茎睫纤虫Ophrydium crassicaule Penard, 1922, 念珠伪钟虫Pseudovorticella monilata (Tatem, 1870) Foissner & Schiffmann, 1974, 垂盖虫Opercularia nutans (Ehrenberg, 1831) Stein, 1854和一个鞘居虫未定种Vaginicola sp.的活体、纤毛图式、银线系以及细胞核的形态学特征进行详细的描述, 补充和完善了缘毛类纤毛虫的形态学资料, 为纤毛虫分类和系统发育学的研究提供重要信息。此外, 首次在伪钟虫中观察到有性生殖现象, 为纤毛虫生殖进化的研究提供了基础资料。     

五株纤毛虫遗传关系的ISSR分析

采用ISSR分子标记技术,尝试对四种五株纤毛虫(褶累枝虫(Epistylis plicatilis)、绿草履虫(Paramecium bursaria)、多态喇叭虫(Stentor polymorphus)、嗜热四膜虫BF1株(Tetrahymena thermophilaBF1)和嗜热四膜虫BF5株(T.ther-mophilaBF5))进行遗传关系研究。用13个ISSR引物对五株纤毛虫进行扩增,六个ISSR引物获得多态片段。根据Nei s遗传距离矩阵构建了五株纤毛虫的遗传关系树状图。UPGMA,NJ聚类图表明:两株嗜热四膜虫最先聚在一起;其次是褶累枝虫和多态喇叭虫聚在一起,然后再与嗜热四膜虫聚在一起;咽膜亚纲的绿草履虫形成独立的一枝。结果显示:①缘毛亚纲纤毛虫可能是寡膜纲中较独特的一个类群,建议提升缘毛亚纲纤毛虫的分类地位;②缘毛亚纲褶累枝虫与膜口亚纲嗜热四膜虫的亲缘关系近于咽膜亚纲绿草履虫,在寡膜纲中绿草履虫处于原始地位;③五株纤毛虫基因组中均含有微卫星DNA序列:(GTG)4(、GACA)4(、AG)8(、CAA)6和(GAA)6。     

六种土壤腹毛类纤毛虫的形态学研究

本研究采用非淹没培养皿法、活体观察法和蛋白银染色法,对隶属于3科5属的6种土壤腹毛类纤毛虫进行了活体形态学和纤毛图式的研究,包括背褶拟片尾虫(Urosomoida dorsiincisura Foissner,1982)、边缘片尾虫[Urosoma emarginata(Stokes,1885)Berger,1999]、巴西戴维虫(Deviata brasiliensis Siqueira-Castro,PaivaSilva-Neto,2009)、刚强殖口虫[Gonostomum strenuum(Engelmann,1862)Sterki,1878]、拟殖口殖口虫[Gonostomum gonostomoidum(Hemberger,1985)Berger,1999]和布氏瓦拉虫[Wallackia bujoreani(Lepsi,1951)BergerFoissner,1989]。其中,边缘片尾虫和拟殖口殖口虫为国内新记录种;巴西戴维虫为我国土壤新记录种。通过对蛋白银染色显示的纤毛排布结果进行主要分类特征的测量和统计,并计算各个种种群主要分类特征的最小值、最大值、平均值、标准差和变异系数。结果显示,本文所研究6个种的种群与国内外同种种群略有差异,但总体在种内范围内波动。本文丰富了我国纤毛虫的物种多样性,同时丰富了该类群纤毛虫的地理分布研究。     

十种海洋寡毛类纤毛虫:形态学及分类学修订

利用活体观察和蛋白银染色技术对近年来采自青岛、大亚湾、湛江沿岸水体的10个海洋寡毛类纤毛虫种:侧扁急游虫Strombidium apolatum Wilbert & Song,2005、具头急游虫Strombidium capitatum (Leegaard,1995) Kahl,1932、广东急游虫Strombidium guangdongense Liu,et al.,2016、拟卡氏急游虫Strombidiumparacalkinsi (Lei,et al.,1999) Agatha,2004、拟楔尾急游虫Strombidium parastylifer Song,et al.,2009、铃木急游虫Strombidium suzukii Song,et al.,2009、束腰旋游虫Spirostrombidium cinctum (Kahl,1932) Petz,et al.,1995、杨科夫平游虫Parallelostrombidium jankowski (Song,et al.,2009) Song,et al.,2018、卡尔平游虫Parallelostrombidium kahli (Song,et al.,2009) Song,et al.,2018、最小拟盗虫Strombidinopsis minima (Gruber,1884) Song & Bradbury,1998的形态学开展了比较研究,补充和厘定了有关形态特征、纤毛图式以及性状变异等分类学新信息。     

青岛沿海7种异毛类纤毛虫形态学描述

借助活体观察及银染法等对采集于青岛沿海的7种异毛类纤毛虫(绿色爽口虫Climacostomum virens、蓝菌丽克虫Licnophora lyngbycola、盐蚕豆虫Fabrea salina、佛氏环须虫Peritromus faurei、简单瓶囊虫Folliculina simplex、延长类瓶囊虫Folliculinopsis producta、弯曲突口虫Condylostoma curva)进行了活体形态与纤毛图式水平的分类学研究, 其中绿色爽口虫为国内新记录。对迄今缺乏详细研究的简单瓶囊虫与延长类瓶囊虫补充了大量活体形态描述, 如皮层颗粒、色素颗粒的分布与颜色变化等, 同时运用现代银染技术首次报道了该两种游泳体的纤毛图式。       

一种淡水缘毛类纤毛虫————沟钟虫的形态学研究

利用活体观察及蛋白银染色技术对一淡水缘毛目纤毛虫——沟钟虫（Vorticella convallaria）的形态学和表膜下纤维系进行了研究。结果表明：沟钟虫的活体个员自然伸展时外形较稳定,并呈明显的倒置钟状,长宽约为50—85μm×40—75μm;口围缘完全外展时为虫体最宽处;伸缩泡一个,较大,位于口围唇下方及口前庭的左侧。胞质均匀而透明,无脂肪滴存在,游泳体呈圆柱形。细胞表面横纹从口围唇到反口纤毛环为74—78条,自反口纤毛环到帚胚为18—24条。大核呈大幅度盘绕的长肠状,两端高度弯曲,纵贯于细胞内。蛋白银制片后表膜下纤维系特征为：虫体纵向纤维稀疏而粗壮,37—42条,似灯笼状;口围盘纤维呈典型的倾斜态分布,并呈放射状排列。第三咽膜（P3）在近口末端处呈明显的分离态,可视为该物种重要的分类学依据。     

青岛沿海的自由生聚缩虫属纤毛虫(原生动物,纤毛门,缘毛目)

涉及近年来采集自青岛沿海的自由生聚缩虫属纤毛虫14种,包括9个国内新纪录:交替聚缩虫Zoothamnium alternans Claparède & Lachmann,1859,弗氏聚缩虫Z. foissneri Ji et al.,2005,黑凯聚缩虫 Z. hiketes Precht,1935,海洋聚缩虫 Z. marinum Mereschkowski, 1879,霉聚缩虫Z. mocedo Entz,1884,倪氏聚缩虫 Z. nii Ji et al.,2005,拟树状聚缩虫 Z. pararbuscula Ji et al., 2005,王氏聚缩虫Z.wangi Ji et al., 2005,许氏聚缩虫 Z. xuianum Sun et al.,2005;以及其他5个已知种:群栖聚缩虫Z. commune Kahl, 1933,双缘聚缩虫Z. duplicatum Kahl,1933,巨大聚缩虫Z.maximum Song,1986,羽状聚缩虫Z.plumula Kahl,1933,中国聚缩虫Z.sinense Song,1991.对以上各种作了简要的综合性描述,并给出了种检索表.     

腹毛类纤毛虫杆形戴维虫的形态学和细胞发生

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利用RAPD标记构建木耳属种间关系的研究

摘 要: 利用RAPD标记以银耳属的2个种为外类群研究了木耳属8个种的分子系统发育关系。根据UPGMA 构建的树状图结果表明木耳属(Auricularia)和银耳属(Tremella)分别为单一的分类单元。木耳属的8个种被分成3个明显不同的组,第一组包括盾形木耳(A.peltata Lloyd)和角质木耳(A.cornea Spreng.),第二组包括所有供试的黑木耳(A.auricula Underw.),第三组包括皱木耳(A.delicata Henn.)、琥珀木耳(A.fuscosuccinea Farl.)、毛木耳(A.polytricha Sacc.)、大木耳(A.maxima Y. R. Guo)和网膜木耳(A.reticulata L. J. Li)五个形态种。种间发育关系表明角质木耳和毛木耳应是两个不同的种而非同种异名,子实体横切面中髓层的有无并不表明不同种间亲缘关系的远近。     

大叶藻居群微卫星遗传多样性研究

采用4对微卫星引物对大叶藻的7个地理居群进行了遗传多样性与遗传结构分析。扩增148株大叶藻得到57个等位基因, 每个位点平均等位基因数为6, 大叶藻居群的平均期望杂合度(He)为0.687, 平均观测杂合度(Ho)为0.417。青岛湾居群的遗传多样性最高(A=7.750, AR=7.043), 俚岛居群最低(A=4.750, AR=4.543)。从F_st值来看, 7个大叶藻居群间属于中度分化。UPGMA系统发育树显示, 中国4个大叶藻居群聚类到一起, 其遗传分化可能是由于历史大海草场的遗留小片段居群产生, 而中国、韩国、日本和爱尔兰居群间的遗传分化则主要是由于地理隔离造成的。自由交配估计结果支持海草的东亚起源说。青岛湾居群遗传多样性较高, 可优先作为大叶藻移植修复的材料和基因库, 并进行重点保护。     

IDENTIFICATION OF PORPHYRA HAITANENSIS (BANGIALES,RHODOPHYTA) MEIOSIS BY SIMPLE SEQUENCE REPEAT MARKERS1

The simple sequence repeat (SSR) marks were employed to identify the stage at which meiosis occurs in the life cycle of Porphyra haitanensis T. J. Chang et B. F. Zheng. More than 90% of F1 blades of heterozygous conchocelis produced by the cross between a red mutant (R, ♀) and the wildtype (W, ♂) were color sectored. Two parental colors (R and W) and two new colors (R′ and W′) appeared in linear sectors in the color‐sectored F1 blades. Two SSR primer pairs selected from a total of 52 primer pairs generated a specific paternal and maternal fragment, respectively. Co‐occurrence of these two bands was detected in heterozygous conchocelis and in the color‐sectored F1 blades with two to four sectors, such as R + W, R′ + W′, and R′ + R + W + W′. However, the single‐colored F1 blades exhibited only one band. In the sectors isolated from the color‐sectored F1 blades, R and R′ were the same, showing the maternal pattern, whereas W and W′ were the same, showing the paternal pattern. These data suggested that the two different bands from heterozygous conchocelis originated from the parents and segregated in the F1 blades, whereas the two new colors, R′ and W′, in the F1 blades were produced by the exchange and recombination of alleles of the parental colors during meiosis. These results indicated that meiosis of P. haitanensis occurs during the first two cell divisions of a germinating conchospore, and, therefore, the initial four cells constitute a linear genetic tetrad, leading to the formation of a color‐sectored blade.     

用RAPD技术对特化等级裂腹鱼类亲缘关系的探讨

为确定特化等级裂腹鱼类的遗传分化关系 ,对 7种 2 2个裂腹鱼个体进行随机扩增多态DNA(RAPD)研究。从所使用的 40个随机引物中 ,选择了 37个扩增带谱清晰的引物进行分析。根据构建的分子系统树显示 ,特化等级裂腹鱼类划分为 3个属级分类单元较为合适 ,这与采用形态学特征进行系统分析所得出的结论一致。在 2 2个个体之间遗传距离矩阵中 ,最大的遗传距离指数达到了 90 38% ,此外 ,还有较多的遗传距离指数也在 6 0 %～ 90 %之间。通过分析 ,认为用RAPD标记技术来分析属级或属级以上分类单元的亲缘关系 ,具有一定的适用性。对于一些遗传差异较大的类群 (如裂腹鱼类 ) ,应用属级或属级以上分类单元的亲缘关系 ,很可能会出现个体之间遗传距离接近或达到 10 0 %的情况 ,从而无法探讨其相互之间的亲缘关系。     

用ISSR标记技术分析山药品种遗传多样性

利用ISSR标记技术对28个山药品种的遗传多样性进行分析。结果表明,从44条ISSR引物中可筛选出7条能够扩增出清晰、稳定条带的引物;这7条ISSR引物对28个山药品种扩增条带间存在较大差异,多态性条带比率为83.01%,Shannon多样性指数为0.3191;构建的分子树状图将28个山药品种划分为4组:第一组含有日本白、花山药和日本园3个品种;第二组为小叶山药;第三组为嵩野1号;其余23个品种归入第四组。而且主成分分析结果支持上述的聚类分析结果。这为利用ISSR标记技术鉴定山药品种,为有效地利用山药种质资源提供了依据。     

PHYLOGENETIC RELATIONSHIPS WITHIN THE COLONIAL VOLVOCALES (CHLOROPHYTA) INFERRED FROM rbcL GENE SEQUENCE DATA

The chloroplast-encoded large subunit of the ribulose-1, 5-bisphosphate carboxylase / oxygenase (rbcL) gene was sequenced from 20 species of the colonial Volvocales (the Volvacaceae, Goniaceae, and Tetrabaenaceae) in order to elucidate phylogenetic relationships within the colonial Volvocales. Eleven hundred twenty-eight base pairs in the coding regions of the (rbcL) gene were analyzed by the neighbor-joining (NJ) method using three kinds of distance estimations, as well as by the maximum parsimony (MP) method. A large group comprising all the anisogamous and oogamous volvocacean species was resolved in the MP tree as well as in the NJ trees based on overall and synonymous substitutions. In all the trees constructed, Basichlamys and Tetrabaena (Tetrabaenaceae) constituted a very robust phylogenetic group. Although not supported by high bootstrap values, the MP tree and the NJ tree based on nonsynonymous substitutions indicated that the Tetrabaenaceae is the sister group to the large group comprising the Volvocaceae and the Goniaceae. In addition, the present analysis strongly suggested that Pandorina and Astrephomene are monophyletic genera whereas Eudorina is nonmonophyletic. These results are essentially consistent with the results of the recent cladistic analyses of morphological data. However, the monophyly of the Volvocaceae previously supported by four morphological synapomorphies is found only in the NJ tree based on nonsynonymous substitutions (with very low bootstrap values). The genus Volvox was clearly resolved as a polyphyletic group with V. rousseletii Pocock separated from other species of Volvox in the rbcL gene comparisons, although this genus represents a monophyletic group in the previous morphological analyses. Furthermore, none of the rbcL gene trees supported the monophyly of the Goniaceae; Astrephomene was placed in various phylogenetic positions .     

DEVELOPMENT OF SEQUENCE CHARACTERIZED AMPLIFIED REGION MARKERS FROM INTERSIMPLE SEQUENCE REPEAT FINGERPRINTS FOR THE MOLECULAR DETECTION OF TOXIC PHYTOPLANKTON ALEXANDRIUM CATENELLA (DINOPHYCEAE) AND PSEUDO‐NITZSCHIA PSEUDODELICATISSIMA (BACILLARIOPHYCEAE) FROM FRENCH COASTAL WATERS1

Harmful algal blooms are a serious threat to shellfish farming and human health all over the world. The monitoring of harmful algae in coastal waters originally involved morphological identification through microscopic examinations, which was often difficult unless performed by specialists and even then often did not permit identification of toxic species. More recently, specific molecular markers have been used to identify specific phytoplankton species or strains. Here we report on the use of the intersimple sequence repeat (ISSR) technique to develop specific sequence characterized amplified region markers (SCAR) and to identify with these tools two toxic species in French coastal waters, the diatom Pseudo‐nitzschia pseudodelicatissima (Hasle) Hasle and the dinoflagellate Alexandrium catenella (Whedon and Kofoid 1936), Balech 1985. Six polymorphic ISSR regions were selected among amplified fingerprints of a representative sample of phytoplankton species. After cloning and sequencing the selected polymorphic ISSR regions, pairs of internal primers were designed to amplify a unique and specific sequence designed as a SCAR marker. Of the six selected SCAR markers, three were specific to P. pseudodelicatissima and one for A. catenella. The SCAR marker specificity was confirmed by using basic local alignment search tool comparison, by experimental assays on different strains from 11 countries, and by checking that the sequence amplified was the expected one. When tested on water samples collected along the French shores, the four specific SCAR markers proved to be efficient tools for fast and low‐cost detection of toxic phytoplankton species.     

PHYLOGENETIC RELATIONSHIPS AMONG ISOLATES OF EUDORINA SPECIES (VOLVOCALES, CHLOROPHYTA) INFERRED FROM MOLECULAR AND BIOCHEMICAL DATA

Phylogenetic analyses of 19 strains representing five species of Eudorina, one strain of Pleodorina indica, and seven strains of Yamagishiella unicocca were carried out by sequencing the internal transcribed spacer region (ITS 1 and ITS 2) of the nuclear ribosomal DNA (rDNA) repeats. The sequence data resolved five phylogenetic groups, one consisting of Y. unicocca and the other four encompassing all the Eudorina species. Two isolates, Eudorina sp. (ASW 05157) and Pleodorina indica (ASW 05153), were of uncertain affiliation. Whereas one monophyletic group included strains of E. elegans only, the other strains of E. elegans appeared alongside E. cylindrica, E. illinoisensis, and E. unicocca var. unicocca in the other Eudorina clades. The distribution pattern of the carotenoid loroxanthin ([3R,3'R,6'R]-β,ε-carotene-3,19,3'-triol), a systematically useful biochemical marker within chlorophycean flagellates, was shown to match the evaluated molecular data. Whereas it was either totally absent or universally present in six of the deduced phylogenetic lines, it occurred randomly in the E. elegans clade containing only E. elegans isolates. The results substantiated the current hypothesis that the unique vegetative morphology of E. elegans has independently arisen at various times during evolution and that it is not a marker of a monophyletic group.     

CONSTRUCTION OF A GENETIC LINKAGE MAP FOR PORPHYRA HAITANENSIS (BANGIALES,RHODOPHYTA) BASED ON SEQUENCE‐RELATED AMPLIFIED POLYMORPHISM AND SIMPLE SEQUENCE REPEAT MARKERS1

Molecular markers and molecular genetic maps are prerequisites for molecular breeding in any plant species. A comprehensive genetic linkage map for cultivated Porphyra haitanensis T. J. Chang et B. F. Zheng has not yet been developed. In this study, 157 double haploid (DH) lines [derived from a YSIII (wildtype) × RTPM (red‐type artificial pigmentation mutant) cross] were used as a mapping population in P. haitanensis. A total of 60 pairs of sequence‐related amplified polymorphism (SRAP) primers and 39 pairs of simple sequence repeat (SSR) primers were used to detect polymorphisms between the two parents. Fifteen SRAP and 16 SSR polymorphic primer pairs were selected to analyze the DH population. A linkage genetic map comprising 67 SRAP markers and 20 SSR markers in five linkage groups, with a total length of 830.6 cM and an average of 10.13 cM between markers, was constructed. The markers were distributed evenly in all linkage groups without clustering. The linkage groups comprised 12–23 markers ranging in length from 134.2 to 197.3 cM. The estimated genome length of P. haitanensis was 942.4 cM, with 88.1% coverage. This is the first report of a comprehensive genetic map in P. haitanensis. The map presented here will provide a basis for the development of high‐density genetic linkage maps and lay the foundation for molecular breeding work in P. haitanensis.