Phylogenetic positions of several amitochondriate protozoa-Evidence from phylogenetic analysis of DNA topoisomerase II

Several groups of parasitic protozoa, as represented by Giardia, Trichomonas, Entamoeba and Microsporida, were once widely considered to be the most primitive extant eukaryotic group―Archezoa. The main evidence for this is their 'lacking mitochondria' and possessing some other primitive features between prokaryotes and eukaryotes, and being basal to all eukaryotes with mitochondria in phylogenies inferred from many molecules. Some authors even proposed that these organisms diverged before the endosymbiotic origin of mitochondria within eukaryotes. This view was once considered to be very significant to the study of origin and evolution of eukaryotic cells (eukaryotes). However, in recent years this has been challenged by accumulating evidence from new studies. Here the sequences of DNA topoisomerase II in G. lamblia, T. vaginalis and E. histolytica were identified first by PCR and sequencing, then combining with the sequence data of the microsporidia Encephalitozoon cunicul and other eukaryotic groups of different evolutionary positions from GenBank, phylogenetic trees were constructed by various methods to investigate the evolutionary positions of these amitochondriate protozoa. Our results showed that since the characteristics of DNA topoisomerase II make it avoid the defect of 'long-branch attraction' appearing in the previous phylogenetic analyses, our trees can not only reflect effectively the relationship of different major eukaryotic groups, which is widely accepted, but also reveal phylogenetic positions for these amitochondriate protozoa, which is different from the previous phylogenetic trees. They are not the earliest-branching eukaryotes, but diverged after some mitochondriate organisms such as kinetoplastids and mycetozoan; they are not a united group but occupy different phylogenetic positions. Combining with the recent cytological findings of mitochondria-like organelles in them, we think that though some of them (e.g. diplomonads, as represented by Giardia) may occupy a very low evolutionary position, generally these organisms are not as extremely primitive as was thought before; they should be polyphyletic groups diverging after the endosymbiotic origin of mitochondrion to adapt themselves to anaerobic parasitic life.     

Phylogenetic positions of several amitochondriate protozoa——Evidence from phylogenetic analysis of DNA topoisomerase II

In the 1980s some special protozoa were gradually recognized: being eukaryotes, they possess some characteristics between prokaryotes and eukaryotes, for example, lack of eukaryotic organelle such as mi-tochondria, and even 70S ribosomes in some species, containing 16S, 23S rRNA like those of prokaryotes. These protozoa contain diplomonads (e.g. intestinal parasite Giardia), parabasalids (e.g. venereal disease pathogen Trichomonas), microsporidia (e.g. the human pathogen Encephalitozoon), a…     

Phylogenetic positions of several amitochondriate protozoa

Several groups of parasitic protozoa, as represented by Giardia, Trichomonas, En-tamoeba and Microsporida, were once widely considered to be the most primitive extant eukaryotic group—Archezoa. The main evidence for this is their ‘lacking mitochondria’ and possessing some other primitive features between prokaryotes and eukaryotes, and being basal to all eukaryotes with mitochondria in phylogenies inferred from many molecules. Some authors even proposed that these organisms diverged before the endosymbiotic origin of mitochondria within eukaryotes. This view was once considered to be very significant to the study of origin and evolution of eukaryotic cells (eukaryotes). However, in recent years this has been challenged by accumulating evidence from new studies. Here the sequences of DNA topoisomerase II in G. lamblia, T. vaginalis and E. histolytica were identified first by PCR and sequencing, then combining with the sequence data of the microsporidia Encephalitozoon cunicul and other eukaryotic groups of different evolutionary positions from GenBank, phylogenetic trees were constructed by various methods to investigate the evolutionary positions of these amitochondriate protozoa. Our results showed that since the characteristics of DNA topoisomerase II make it avoid the defect of ‘long-branch attraction’ appearing in the previous phylogenetic analyses, our trees can not only reflect effectively the relationship of different major eukaryotic groups, which is widely accepted, but also reveal phylogenetic positions for these amitochondriate protozoa, which is different from the previous phylogenetic trees. They are not the earliest-branching eukaryotes, but diverged after some mitochondriate organisms such as kinetoplastids and mycetozoan; they are not a united group but occupy different phylogenetic positions. Combining with the recent cytological findings of mitochondria-like organelles in them, we think that though some of them (e.g. diplomonads, as represented by Giardia) may occupy a very low evolutionary position, generally these organisms are not as extremely primitive as was thought before; they should be polyphyletic groups diverging after the endosymbiotic origin of mitochondrion to adapt themselves to anaerobic parasitic life.     

Identification of Horizontal Gene Transfer from Phylogenetic Gene Trees

We suggest a new procedure to search for the genes with horizontal transfer events in their evolutionary history. The search is based on analysis of topology difference between the phylogenetic trees of gene (protein) groups and the corresponding phylogenetic species trees. Numeric values are introduced to measure the discrepancy between the trees. This approach was applied to analyze 40 prokaryotic genomes classified into 132 classes of orthologs. This resulted in a list of the candidate genes for which the hypothesis of horizontal transfer in evolution looks true.     

饶氏藻系统位置的研究

用18S rDNA基因序列分析饶氏藻属(Jaoa)与相关藻类的亲缘关系。结果表明:饶氏藻18S rDNA的长度为1632 bp,GC%为50.6%,泡状饶氏藻18S rDNA的长度为1639 bp,GC%为50.3%。用最大简约法与饶氏藻上一级分类单元(目)构建的系统树表明有4个大的分支。两种饶氏藻与石莼目的 Ulva curvata、U.rigida、Enteromorpha intestinalis和Monostroma grevillei构成很强的支持分支(分支B),它们之间的核苷酸趋异性最低仅0.041,而与胶毛藻目的 Chaetophora incrassata的趋异性则很显著,达0.112,因此,饶氏藻应属于石莼目的一个类群,且饶氏藻和泡状饶氏藻构成一单系起源的分支,这两个物种的趋异性仅0.002,显示出它们具有非常紧密的亲缘关系,很可能是1种1变种而不是2种。     

基于DNA拓扑异构酶II基因部分序列的侧耳属系统发育分析

对侧耳属（Pleurotus）14个种的14个菌株,亚侧耳属（Hohenbuehelia）1个种和离褶伞属（Lyophyllum）1个种的DNA拓扑异构酶II（EC 5.99.1.3）部分序列进行扩增分析。以灰盖鬼伞（Coprinus cinereus）、新型隐球酵母（Cryptococcus neoformans）和玉米黑粉菌（Puccinia polysora）等担子菌为外群,采用最大距离法、最大简约法和最大似然法构建侧耳属的系统发育树。结果表明：作为外群的三个物种独立于所有供试材料;侧耳属菌株作为一大类与供试的离褶伞属和亚侧耳属分开。侧耳属14个种又可细分成六支：可能为侧耳属起源最早的P. flabellatus、P. salmoneostramineus和P. djamor组成一组;P. tuberregium和P. pulmonarius均自成一组;P. abalonus和P. cystidiosus均产生束梗孢,聚为同组;P. eryngii var. eryngii、P. eryngii var. ferulae和P. nebrodensis为一组;另一组由P. ostreatus、P. columbinus、P. cornucopiae和P. citrinopileatus组成。按照侧耳属物种菌丝类型的不同,构建的系统发育树将该属14个种进行系群划分,单、二系菌丝分属于各个不同的分支中,说明单、二系菌丝均为多系起源,与用28S rDNA序列进行系统分析的结果一致。     

12株酵母菌的亲缘关系分析

克隆了９株假线酵母和１株克鲁维酵母的２５ＳｒＤＮＡ片段,测定其５′端部分苷酸序列与报道的Ｃａｎｄｉｄａａｌｂｉｃａｎｓ及Ｓａｃｃｈａｒｏｍｙｃｅｓｃｅｒｅｖｉｓｉａｅ２５ＳｒＤＮＡ相应区域的核苷酸序列比较,采用ｎｅｉｇｈｂｏｒ－ｊｏｉｎｉｎｇ和ｂｏｏｔ－ｓｔｒａｐ法分析并绘制系统树,结果提示ＣａｎｄｉｄａｋｅｆｙｒＣＢＳ８３４与Ｋｌｕｙｖｅｒｏｍｙｃｅｓ ｃｉｃｅｒｉｓｐｏｒｕｓＣＢＳ４８５７亲缘     

Phylogenetic information improves homology detection.

We present a database search method that is based on phylogenetic trees (treesearch). The method is used to search a protein sequence database for homologs to a protein family. In preparation for the search, a phylogenetic tree is constructed from a given multiple alignment of the family. During the search, each database sequence is temporarily inserted into the tree, thus adding a new edge to the tree. Homology between family and sequence is then judged from the length of this edge. In a comparison of our method to profiles (ISREC pfsearch), two implementations of hidden Markov models (HMMER hmmsearch and SAM hmmscore), and to the family pairwise search (FPS) method on 43 families from the SCOP database based on minimum false-positive counts (min-FPCs), we found a considerable gain in sensitivity. In 69% of the test cases, treesearch showed a min-FPC of at most 50, whereas the two second best methods (hmmsearch and FPS) showed this performance only in 53% cases. A similar impression holds for a large range of min-FPC thresholds. The results demonstrate that phylogenetic information can significantly improve the detection of distant homologies and justify our method as a useful alternative to existing methods.     

Interleukin-2 induces the activities of DNA topoisomerase I and DNA topoisomerase II in HuT 78 cells

The induction by interleukin-2 of DNA topoisomerase I and DNA topoisomerase II activities in the human T cell line HuT 78 was investigated. HuT 78 cells were treated with 1000 U of interleukin-2/ml, and extracts of the HuT 78 nuclei were prepared over a 24 h period. The extracts were assayed quantitatively for the activities of DNA topoisomerase I and DNA topoisomerase II. Three concomitant, transient increases of 3- to 11-fold in the specific activities of both DNA topoisomerase I and DNA topoisomerase II were observed following treatment with IL-2 at 0.5, 4, and 10 h after treatment with interleukin-2. The specific activities of both enzymes returned to base-line values after each of these transient increases. These results reveal that the activities of DNA topoisomerase I and DNA topoisomerase II are highly regulated in HuT 78 cells upon treatment with IL-2.     

The best of both worlds: Phylogenetic eigenvector regression and mapping

Eigenfunction analyses have been widely used to model patterns of autocorrelation in time, space and phylogeny. In a phylogenetic context, Diniz-Filho et al. (1998) proposed what they called Phylogenetic Eigenvector Regression (PVR), in which pairwise phylogenetic distances among species are submitted to a Principal Coordinate Analysis, and eigenvectors are then used as explanatory variables in regression, correlation or ANOVAs. More recently, a new approach called Phylogenetic Eigenvector Mapping (PEM) was proposed, with the main advantage of explicitly incorporating a model-based warping in phylogenetic distance in which an Ornstein-Uhlenbeck (O-U) process is fitted to data before eigenvector extraction. Here we compared PVR and PEM in respect to estimated phylogenetic signal, correlated evolution under alternative evolutionary models and phylogenetic imputation, using simulated data. Despite similarity between the two approaches, PEM has a slightly higher prediction ability and is more general than the original PVR. Even so, in a conceptual sense, PEM may provide a technique in the best of both worlds, combining the flexibility of data-driven and empirical eigenfunction analyses and the sounding insights provided by evolutionary models well known in comparative analyses.     

Tree Reconciliation: Reconstruction of Species Phylogeny by Phylogenetic Gene Trees

It is well known that phylogenetic trees derived from different protein families are often incongruent. This is explained by mapping errors and by the essential processes of gene duplication, loss, and horizontal transfer. Therefore, the problem is to derive a consensus tree best fitting the given set of gene trees. This work presents a new method of deriving this tree. The method is different from the existing ones, since it considers not only the topology of the initial gene trees, but also the reliability of their branches. Thereby one can explicitly take into account the possible errors in the gene trees caused by the absence of reliable models of sequence evolution, by uneven evolution of different gene families and taxonomic groups, etc.     

Phylogenetic analysis of protozoa in the rumen contents of cow based on the 18S rDNA sequences

AIMS: To examine the diversity of protozoa in the rumen contents of cow. METHODS AND RESULTS: Protozoa that inhabit the rumen were detected by PCR using protozoan-specific primers. Libraries of protozoan rDNA sequences were constructed from rumen fluid, solid tissues and epithelium. Twenty-three clones isolated from rumen fluid fell into two genera identified as Entodinium (69.6% of clones) and Epidinium (31.4% of clones). Of the clones isolated from rumen fluid, a moderate number were unidentifiable (30.4%). CONCLUSIONS: The predominant protozoan genus identified in the whole rumen belonged to the Entodinium group (81.1%). Protozoa were not detected in the rumen epithelium. SIGNIFICANCE AND IMPACT OF THE STUDY: These findings suggest that rumen fluid and solid tissues contain different protozoan populations that may play specific roles in rumen function. Quantitative PCR techniques and a more specific set of phylogenetic probes that distinguish between protozoan species are needed to determine the significance of newly identified groups and to determine the distribution of identified protozoan clusters in rumen microbial communities.     

基于rbcL和trnL-trnF序列探讨粤紫萁的系统位置及遗传关系

通过对粤紫萁(Osmunda mildei C.Chr.)及紫萁属(Osmunda L.)植物的rbcL和trnL-trnF(包括trnL内含子和trnL-F间隔区)序列进行了测定,并结合部分已发表的紫萁科(Osmundaceae)植物的相关序列进行系统发育分析。结果表明:(1)所分析的紫萁属植物构成一个单系群;(2)粤紫萁与紫萁属中Sect.Plenasium组亲缘关系相近,聚成一支系,支持率为100%;(3)粤紫萁与Sect.Plenasium组的华南紫萁(O.vachellii Hook.)在碱基变异位点上高度一致,达94%,而与Sect.Osmunda组的紫萁(O.japonica Thunb)在变异位点上仅有2%的一致性。因此,根据叶绿体基因母系遗传的特性,推测华南紫萁为粤紫萁的母本,紫萁为其父本。     

3种杓兰属植物菌根真菌系统发育和多样性分析

兰科植物菌根真菌(Orchid mycorrhizal fungi, OrMF)在兰科植物种子萌发和后续生长发育过程中具有重要作用。该研究采用培养(菌丝团分离)和非培养(克隆文库)2种方法获得同一栖息地3种不同杓兰属植物根中菌根真菌ITS序列并划分可操作分类单元(Operational taxonomic units, OTUs),分析其系统发育关系和多样性。结果表明:(1)所有根段中都有菌丝团定植,共分离出菌根真菌64株,其中63株为胶膜菌科(Tulasnellaceae)真菌,1株为角担菌科(Ceratobasidiaceae)真菌;可划分为7个OUT,每个OTU代表菌株的菌丝都能形成OrMF典型的近球形或椭球形链状排列的念珠状细胞;分离出来的菌根真菌均为无性型菌丝且不产生无性孢子。(2) 非培养法得到的3种杓兰属植物的根中OrMF分别隶属于胶膜菌科(Tulasnellaceae),腊壳菌科(Sebacinaceae)、角担菌科(Ceratobasidiaceae)和革菌科(Thelephoraceae),其中胶膜菌科OTU在种类和数量上占有绝对优势,培养和非培养2种方法得到的OrMF OTU类型和数量均为西藏杓兰(Cypripedium tibeticum)>无苞杓兰(C. flavum)>黄花杓兰(C. bardolphianum),但培养法少于非培养法。(3)对胶膜菌进行系统发育分析显示,优势和非优势OTU均分布在系统发育树的3个不同分支上,这种与多种亲缘关系较远的OrMF共生的现象可能与杓兰属植物对环境的适应性有关,且不同杓兰的OrMF物种丰富度没有显著差异,但群落结构存在差异。     

Phylogenetic dimension of tree communities reveals high conservation value of disturbed tropical rain forests

Aim

The conversion of old‐growth tropical forests into human‐modified landscapes threatens biodiversity worldwide, but its impact on the phylogenetic dimension of remaining communities is still poorly known. Negative and neutral responses of tree phylogenetic diversity to land use change have been reported at local and landscape scales. Here, we hypothesized that such variable responses to disturbance depend on the regional context, being stronger in more degraded rain forest regions with a longer history of land use.

Location

Six regions in Mexico and Brazil.

Methods

We used a large vegetation database (6,923 trees from 686 species) recorded in 98 50‐ha landscapes distributed across two Brazilian and four Mexican regions, which exhibit different degrees of disturbance. In each region, we assessed whether phylogenetic alpha and beta diversities were related to landscape‐scale forest loss, the percentage of shade‐intolerant species (a proxy of local disturbance) and/or the relatedness of decreasing (losers) and increasing (winners) taxa.

Results

Contrary to our expectations, the percentage of forest cover and shade‐intolerant species were weakly related to phylogenetic alpha and beta diversities in all but one region. Loser species were generally as dispersed across the phylogeny as winner species, allowing more degraded, deforested and species‐poorer forests to sustain relatively high levels of evolutionary (phylogenetic) diversity.

Main conclusion

Our findings support previous evidence indicating that traits related to high susceptibility to forest disturbances are convergent or have low phylogenetic signal. More importantly, they reveal that the evolutionary value of disturbed forests is (at least in a phylogenetic sense) much greater than previously thought.

     

RNA interference of a trypanosome topoisomerase II causes progressive loss of mitochondrial DNA

We studied the function of a Trypanosoma brucei topoisomerase II using RNA interference (RNAi). Expression of a topoisomerase II double-stranded RNA as a stem-loop caused specific degradation of mRNA followed by loss of protein. After 6 days of RNAi, the parasites' growth rate declined and the cells subsequently died. The most striking phenotype upon induction of RNAi was the loss of kinetoplast DNA (kDNA), the cell's catenated mitochondrial DNA network. The loss of kDNA was preceded by gradual shrinkage of the network and accumulation of gapped free minicircle replication intermediates. These facts, together with the localization of the enzyme in two antipodal sites flanking the kDNA, show that a function of this topoisomerase II is to attach free minicircles to the network periphery following their replication.