云南腾冲热海三热泉细菌多样性的研究*

应用变性梯度凝胶电泳 (DGGE)对云南腾冲热海 3个高温热泉中菌藻席和泉底沉积物的细菌多样性进行了初步研究。直接从环境样品中提取总DNA ,用两套细菌通用引物进行PCR扩增 ,分别得到包含V₈和V₉高变区的 1 6SrDNA片段 ,进行DGGE分析。结果表明 :菌藻席和泉底沉积物中不仅物种组成差异较大 ,而且都存在丰富的细菌多样性 ;一些关键的生态因子 ,如氧气、温度等会对群落中微生物的物种组成有很大影响。     

云南腾冲热海两热泉菌藻席细菌多样性的研究

应用显微形态观察和变性梯度凝胶电泳(DGGE)对云南腾冲热海两热泉菌藻席的细菌多样性进行了比较分析.直接从环境样品中提取总DNA,用两套细菌通用引物进行PCR扩增,得到包含V8和V9高变区的16S rDNA片段,进行DGGE分析,结合形态观察,结果显示,热泉菌藻席中存在丰富的细菌多样性,且不同温度范围的菌藻席细菌组成差异显著.     

腾冲热海四种不同颜色菌藻席的DGGE图谱比较

研究热泉水化学成分与菌藻席群落结构的关系。直接提取四种菌藻席(mat)总DNA,PER扩增获得16S rDNA的V8高变区片段,进行DGGE分析。结果：四种菌藻席的细菌组成差异很大,DGGE条带数目最低为20,最高为47;4种菌藻席共有的条带为1。结合文献报道的3个热泉的水化学性质分析,表明水化学成分是热泉生态系统中重要的生态因子,直接影响菌藻席的群落结构和物种组成。     

腾冲热海眼镜泉粉红色菌藻席的细菌组成分析

应用免培养法系统研究眼镜泉粉红色菌藻席的细菌组成。经过克隆筛选,测定了23个克隆的16S rDNA插入片段的近全序列。与GenBank的序列进行比对和相似性分析,结果表明,组成该菌藻席的细菌分属于Proteobacteria、Firmicutes、Bacteroidetes、Actinobacter、Deinococcus-thermus、Aquificals 6个类群(phylum),表现出了高度的细菌多样性。结合分析温度相近的5个热泉：Octopus spring、Haegindi and Fluidir spring、Olkelduhals、Grendalur spring中的菌藻席细菌组成,表明生态位相近的不同环境中,其物种组成相近。Aquificales是中性或弱碱性高温热泉菌藻席群落的优势物种。     

宜春温泉水体与泉底沉积物细菌多样性分析

研究宜春富硒温泉水体与泉底沉积物的细菌群落多样性。利用高通量测序技术分析泉水与沉积物中细菌群落结构与多样性。温泉水中主要的细菌类群为变形菌门和拟杆菌门,而在沉积物样品中的主要优势菌群为OP1、蓝细菌、浮霉菌门和绿弯菌门。细菌在属分类水平上,温泉水中优势菌群为不动杆菌属、假单胞菌属、水栖菌属、Thermosynechococcus、鞘脂杆菌属和金黄杆菌属等。沉积物样品细菌中优势菌群属于未知物种,在数据库中并没有相关的注释信息;其中已知的优势菌属为Candidatus acetothermum、Thermosynechococcus、亚热栖菌属、不动杆菌属。宜春温汤富硒温泉水体与沉积物中存在着丰富的微生物群落且组成差异性很大,该研究为了解与发掘温泉微生物菌种资源具有重要价值。     

腾冲热海眼镜泉菌藻席细菌ARDRA指纹图谱分析

对腾冲热海眼镜泉中粉红色丝状菌藻席进行ARDRA指纹图谱分析,获得其细菌多样性的部分信息。直接提取菌藻席总DNA,以之为模板PCR扩增出细菌的16S rDNA,将其克隆、转化进入大肠杆菌,获得120个克隆子。用限制性内切酶(RsaⅠ、HhaⅠ、HapⅡ)筛选出了29个16S rDNA插入片段酶切带型差异明显的克隆,表明眼镜泉菌藻席细菌组成丰富。聚类分析显示这29个克隆聚为A、B两个大类,表明该菌藻席可能主要由两个遗传多样性丰富的分类群组成。     

舌苔细菌PCR-DGGE分析条件的建立与优化

目的针对口腔舌苔细菌16S rDNA序列进行变性梯度凝胶电泳(denaturing gradient gel electrophoresis,DGGE)适用序列的筛选及电泳条件的优化。方法以DGGE图谱的高分辨率为指标,选择舌苔细菌DGGE分离最适的16S rDNA高变区、电泳电压和电泳时间,并采用优化的条件分析健康青年人舌苔细菌群落的分布。结果舌苔细菌16S rDNA V3高变区引物序列能获得更加丰富清晰的DGGE条带;基于该区,当变性剂浓度为30%～60%、电泳温度60℃、电压60 V和电泳时间14 h时能得到分辨率最佳的DGGE图谱。运用此优化条件对12个样本舌苔细菌群落的分析表明,舌苔微生物主要由厚壁菌门、梭杆菌门、拟杆菌门和变形菌门等组成。优化后的DGGE技术对舌苔细菌多样性的分析具有准确性、灵敏性和可重复性。结论 DGGE图谱显示,不同分析条件对图谱类型和细菌多样性指数均有所差异。利用优化的DGGE条件能有效分离舌苔细菌16S rDNA V3区序列,为口腔微生物群落结构分析提供可靠的技术支持,也为其他不同生态细菌的多样性分析提供参考。     

基于PCR-DGGE技术的红树林区微生物群落结构

【目的】为了解红树林沉积物中细菌的群落结构特征。【方法】应用PCR-DGGE技术对福建浮宫红树林的16个采样站位样品细菌的群落结构进行了研究。根据DGGE指纹图谱,对它们的遗传多样性进行了分析。【结果】各站位样品细菌多样性指数(H)、丰度(S)和均匀度(EH)均有所不同,这些差异与它们所处站位的不同有关,红树林区细菌多样性高于非红树林区细菌多样性。对不同站位细菌群落相似性分析,它们的相似性系数也存在一定的规律,同一断面的细菌群落结构相近性较高。对DGGE的优势条带序列分析,同源性最高的微生物分别属于变形菌门(Proteobacteria)、酸菌门(Acidobacteria)和绿菌门(Chlorobi),它们均为未培养微生物,分别来自于河口海岸沉积物。【结论】应用PCR-DGGE技术更能客观地反映红树林沉积物中真实的细菌群落结构信息。另外,研究也表明红树林区微生物多样性丰富,在红树林区研究开发未知微生物资源具有巨大的潜力。     

基于高通量测序对呼伦贝尔陶顺阿尔山细菌群落多样性的研究

对内蒙古呼伦贝尔陶顺阿尔山水体与沉积物的细菌群落结构和多样性，进行16S rRNA基因(V3～V5区)高通量测序。运用Illumina MiSeq高通量测序平台，共获得594 226条优化序列。结果显示，水体中最主要的细菌类群为变形菌门(Proteobacteria),而沉积物样品中的主要优势类群为盐厌氧菌门(Haloanaerobium);细菌在属分类水平上，水体中优势菌群为嗜盐单胞菌属(Halomonas)、冷弯菌属(Psychroflexus)、盐厌氧菌属(Haloanaerobium);沉积物样品中优势菌群为盐厌氧菌属、盐单胞菌属、未分类菌(no-rank-f-unclassified),其中存在大量的未知物种。陶顺阿尔山水体与沉积物中存在着丰富的微生物群落，且差异性大，未知菌种居多。本研究首次阐明了陶顺阿尔山细菌群落的多样性，为今后开发和利用陶顺阿尔山的功能菌群提供参考。     

应用PCR-DGGE技术研究石莼、网地藻藻际微生物多样性

应用变性凝胶梯度电泳(PCR-DGGE)技术对石莼、网地藻藻际微生物多样性进行研究,并对其进行相似性、未加权聚类分析(UPGMA)及微生物多样性(Shannon)指数分析。结果表明,PCR-DGGE图谱显示,石莼、网地藻藻际微生物DGGE图谱有着明显的差异性,具体表现在条带数及密度值的不同。Quantity one图谱分析表明:石莼藻际微生物16S r RNA基因的V3区共分离得到25条DNA片段,网地藻为16条DNA片段。DGGE相似性及未加权聚类分析表明:网地藻藻际微生物间的相似性为77.78%,石莼藻际微生物细菌群落间的相似性为49.25%。Shannon指数分析表明,石莼藻际微生物多样性指数显著高于网地藻(P0.05)。石莼藻际微生物细菌群落较网地藻丰富。     

On the origin of the Hirudinea and the demise of the Oligochaeta

The phylogenetic relationships of the Clitellata were investigated with a data set of published and new complete 18S rRNA gene sequences of 51 species representing 41 families. Sequences were aligned on the basis of a secondary structure model and analysed with maximum parsimony and maximum likelihood. In contrast to the latter method, parsimony did not recover the monophyly of Clitellata. However, a close scrutiny of the data suggested a spurious attraction between some polychaetes and clitellates. As a rule, molecular trees are closely aligned with morphology-based phylogenies. Acanthobdellida and Euhirudinea were reconciled in their traditional Hirudinea clade and were included in the Oligochaeta with the Branchiobdellida via the Lumbriculidae as a possible link between the two assemblages. While the 18S gene yielded a meaningful historical signal for determining relationships within clitellates, the exact position of Hirudinea and Branchiobdellida within oligochaetes remained unresolved. The lack of phylogenetic signal is interpreted as evidence for a rapid radiation of these taxa. The placement of Clitellata within the Polychaeta remained unresolved. The biological reality of polytomies within annelids is suggested and supports the hypothesis of an extremely ancient radiation of polychaetes and emergence of clitellates.     

On the ontogeny of the haptor and the evolution of the Monogenea

Data on the ontogeny of the posterior haptor of monogeneans were obtained from more than 150 publications and summarised. These data were plotted into diagrams showing evolutionary capacity levels based on the theory of a progressive evolution of marginal hooks, anchors and other attachment components of the posterior haptor in the Monogenea (Malmberg, 1986). 5 + 5 unhinged marginal hooks are assumed to be the most primitive monogenean haptoral condition. Thus the diagrams were founded on a 5 + 5 unhinged marginal hook evolutionary capacity level, and the evolutionary capacity levels of anchors and other haptoral attachement components were arranged according to haptoral ontogenetical sequences. In the final plotting diagram data on hosts, type of spermatozoa, oncomiracidial ciliation, sensilla pattern and protonephridial systems were also included. In this way a number of correlations were revealed. Thus, for example, the number of 5 + 5 marginal hooks correlates with the most primitive monogenean type of spermatozoon and with few sensillae, many ciliated cells and a simple protonephridial system in the oncomiracidium. On the basis of the reviewed data it is concluded that the ancient monogeneans with 5 + 5 unhinged marginal hooks were divided into two main lines, one retaining unhinged marginal hooks and the other evolving hinged marginal hooks. Both main lines have recent representatives at different marginal hook evolutionary capacity levels, i.e. monogeneans retaining a haptor with only marginal hooks. For the main line with hinged marginal hooks the name Articulon-choinea n. subclass is proposed. Members with 8 + 8 hinged marginal hooks only are here called Proanchorea n. superord. Monogeneans with unhinged marginal hooks only are here called Ananchorea n. superord. and three new families are erected for its recent members: Anonchohapteridae n. fam., Acolpentronidae n. fam. and Anacanthoridae n. fam. (with 7 + 7, 8 + 8 and 9 + 9 unhinged marginal hooks, respectively). Except for the families of Articulonchoinea (e.g. Acanthocotylidae, Gyrodactylidae, Tetraonchoididae) Bychowsky's (1957) division of the Monogenea into the Oligonchoinea and Polyonchoinea fits the proposed scheme, i.e. monogeneans with unhinged marginal hooks form one old group, the Oligonchoinea, which have 5 + 5 unhinged marginal hooks, and the other group form the Polyonchoinea, which (with the exception of the Hexabothriidae) has a greater number (7 + 7, 8 + 8 or 9 + 9) of unhinged marginal hooks. It is proposed that both these names, Oligonchoinea (sensu mihi) and Polyonchoinea (sensu mihi), will be retained on one side and Articulonchoinea placed on the other side, which reflects the early monogenean evolution. Except for the members of Ananchorea [Polyonchoinea], all members of the Oligonchoinea and Polyonchoinea have anchors, which imply that they are further evolved, i.e. have passed the 5 + 5 marginal hook evolutionary capacity level (Malmberg, 1986). There are two main types of anchors in the Monogenea: haptoral anchors, with anlages appearing in the haptor, and peduncular anchors, with anlages in the peduncle. There are two types of haptoral anchors: peripheral haptoral anchors, ontogenetically the oldest, and central haptoral anchors. Peduncular anchors, in turn, are ontogenetically younger than peripheral haptoral anchors. There may be two pairs of peduncular anchors: medial peduncular anchors, ontogentically the oldest, and lateral peduncular anchors. Only peduncular (not haptoral) anchors have anchor bars. Monogeneans with haptoral anchors are here called Mediohaptanchorea n. superord. and Laterohaptanchorea n. superord. or haptanchoreans. All oligonchoineans and the oldest polyonchoineans are haptanchoreans. Certain members of Calceostomatidae [Polyonchoinea] are the only monogeneans with both (peripheral) haptoral and peduncular anchors (one pair). These monogeneans are here called Mixanchorea n. superord. Polyonchoineans with peduncular anchors and unhinged marginal hooks are here called the Pedunculanchorea n. superord. The most primitive pedunculanchoreans have only one pair of peduncular anchors with an anchor bar, while the most advanced have both medial and lateral peduncular anchors; each pair having an anchor bar. Certain families of the Articulonchoinea, the Anchorea n. superord., also have peduncular anchors (parallel evolution): only one family, the Sundanonchidae n. fam., has both medial and lateral peduncular anchors, each anchor pair with an anchor bar. Evolutionary lines from different monogenean evolutionary capacity levels are discussed and a new system of classification for the Monogenea is proposed.In agreeing to publish this article, I recognise that its contents are controversial and contrary to generally accepted views on monogenean systematics and evolution. I have anticipated a reaction to the article by inviting senior workers in the field to comment upon it: their views will be reported in a future issue of this journal. EditorIn agreeing to publish this article, I recognise that its contents are controversial and contrary to generally accepted views on monogenean systematics and evolution. I have anticipated a reaction to the article by inviting senior workers in the field to comment upon it: their views will be reported in a future issue of this journal. Editor