Compositional Bias May Affect Both DNA-Based and Protein-Based Phylogenetic Reconstructions

It is now well-established that compositional bias in DNA sequences can adversely affect phylogenetic analysis based on those sequences. Phylogenetic analyses based on protein sequences are generally considered to be more reliable than those derived from the corresponding DNA sequences because it is believed that the use of encoded protein sequences circumvents the problems caused by nucleotide compositional biases in the DNA sequences. There exists, however, a correlation between AT/GC bias at the nucleotide level and content of AT- and GC-rich codons and their corresponding amino acids. Consequently, protein sequences can also be affected secondarily by nucleotide compositional bias. Here, we report that DNA bias not only may affect phylogenetic analysis based on DNA sequences, but also drives a protein bias which may affect analyses based on protein sequences. We present a striking example where common phylogenetic tools fail to recover the correct tree from complete animal mitochondrial protein-coding sequences. The data set is very extensive, containing several thousand sites per sequence, and the incorrect phylogenetic trees are statistically very well supported. Additionally, neither the use of the LogDet/paralinear transform nor removal of positions in the protein alignment with AT- or GC-rich codons allowed recovery of the correct tree. Two taxa with a large compositional bias continually group together in these analyses, despite a lack of close biological relatedness. We conclude that even protein-based phylogenetic trees may be misleading, and we advise caution in phylogenetic reconstruction using protein sequences, especially those that are compositionally biased. Received: 19 February 1998 / Accepted: 28 August 1998     

Sister-scanning: a Monte Carlo procedure for assessing signals in recombinant sequences

MOTIVATION: To devise a method that, unlike available methods, directly measures variations in phylogenetic signals in gene sequences that result from recombination, tests the significance of the signal variations and distinguishes misleading signals. RESULTS: We have developed a method, that we call 'sister-scanning', for assessing phylogenetic and compositional signals in the various patterns of identity that occur between four nucleotide sequences. A Monte Carlo randomization is done for all columns (positions) within a window and Z-scores are obtained for four real sequences or three real sequences with an outlier that is also randomized. The usefulness of the approach is demonstrated using tobamovirus and luteovirus sequences. Contradictory phylogenetic signals were distinguished in both datasets, as were regions of sequence that contained no clear signal or potentially misleading signals related to compositional similarities. In the tobamovirus dataset, contradictory phylogenetic signals were separated by coding sequences up to a kilobase long that contained no clear signal. Our re-analysis of this dataset using sister-scanning also yielded the first evidence known to us of an inter-species recombination site within a viral RNA-dependent RNA polymerase gene together with evidence of an unusual pattern of conservation in the three codon positions.     

连香树科及其近缘植物matK序列分析和系统学意义

测定和分析了连香树科(Cercidiphyllaeeae)、交让木科(Daplmiphyllaceac)、金缕梅科(Hamamelidaceae)代表植物的叶绿体marK序列(5′端31bps除外),以木兰属作为外类群,应用邻接法构建分子系统树,结果表明：连香树科与水青树科的亲缘关系较远。连香树科、交让木科和金缕梅科形成了一个自展数据支持率(bootstrap)为100％的单系类群,其中金缕梅科枫香属(Liquidambar)、红花荷属(Rhodoleia)和金缕梅属(Hamamelis)虽构成了一个单系类群,但自展数据支持率仅为68％;连香树科与交让木科构成的单系分支自展数据支持率仅为53％。由于连香树科、交让木科、金缕梅科之间的进化距离相当短,表明这3个科之间亲缘关系密切,内部分支的自展数据支持率不高,表明它们之间准确的亲缘关系有待进一步研究。本研究结果与rbcL、aptB、18S rDNA序列分析结果相似,但自展数据支持率更高,表明marK序列分析可应用于较高等级分类群系统发育关系的研究。     

Expanding the known diversity and environmental distribution of an uncultured phylogenetic division of bacteria

Culture-independent molecular phylogenetic methods were used to explore the breadth of diversity and environmental distribution of members of the division-level "candidate" phylogenetic group WS6, recently discovered in a contaminated aquifer and with no cultivated representatives. A broad diversity of WS6-affiliated sequences were cloned from 7 of 12 environments investigated: mainly from anaerobic sediment environments. The number of sequences representing the WS6 candidate division was increased from 3 to 60 in this study. The extent of phylogenetic divergence (sequence difference) in this candidate division was found to be among the largest of any known bacterial division. This indicates that organisms representing the WS6 phylogenetic division offer a broad diversity of undiscovered biochemical and metabolic novelty. These results provide a framework for the further study of these evidently important kinds of organisms and tools, the sequences, with which to do so.     

Expanding the Known Diversity and Environmental Distribution of an Uncultured Phylogenetic Division of Bacteria

Culture-independent molecular phylogenetic methods were used to explore the breadth of diversity and environmental distribution of members of the division-level “candidate” phylogenetic group WS6, recently discovered in a contaminated aquifer and with no cultivated representatives. A broad diversity of WS6-affiliated sequences were cloned from 7 of 12 environments investigated: mainly from anaerobic sediment environments. The number of sequences representing the WS6 candidate division was increased from 3 to 60 in this study. The extent of phylogenetic divergence (sequence difference) in this candidate division was found to be among the largest of any known bacterial division. This indicates that organisms representing the WS6 phylogenetic division offer a broad diversity of undiscovered biochemical and metabolic novelty. These results provide a framework for the further study of these evidently important kinds of organisms and tools, the sequences, with which to do so.     

衣藻属的系统发育分析——基于形态形状和nrDNA ITS序列

通过实验分析莱茵衣藻 ( Chlamydomonas reinhardtii) 1个种和互连网获得衣藻属 1 5个种及丝藻属 1个种 ( Ulothrix zonata) ,共 1 7个种的 nr DNA ITS序列 ,并以 U.zonata为外类群 ,采用计算机分析软件包对其进行分析及构建分子系统发育树图。同时以 1 2个传统分类性状 ,对此 1 6种衣藻构建数据矩阵 ;以 U.zonata动孢子的相应性状为外类群原始性状 ,用Wagner法在计算机上对其进行分枝分析 ;然后比较并分析分子系统树和表征性状分支分析树的异同。初步尝试以 ITS分子序列系统发育分析作为传统性状分析的补充来研究衣藻种间的亲缘关系。     

鹳形目12种鸟类核c-mos 基因和线粒体12S rRNA基因序列分析及其系统发生研究

鹳形目鸟类的传统分类一直存在分歧,而近期的分子系统学研究大多只用单个基因,其结论的可信度需要进一步验证.本文通过核c-mos基因和线粒体12S rRNA基因序列分别和合并分析,采用分子系统学方法探讨了鹳形目6科12种鸟类的系统发生关系.文中测出鹳形目鸟类6种核c-mos基因的片断序列,结合来自Genebank的其他种类的c-mos和12S rRNA基因序列,分别经Clustal W软件对位排列后,以原鸡为外类群用最大似然法、邻接法和最大简约法建立系统树.系统树分析表明, 鹳形目6科之间的系统发生关系总结为：（鹭科,（（鹮科,美洲鹫科）,（鹳科,（鲸头鹳科,锤头鹳科））））.鹭科7个属之间的系统发生关系总结为：（麻（开鸟）属（夜鹭属（池鹭属（苍鹭属（中白鹭属（白鹭属,大白鹭属））））））.分别基于两个单基因的系统树有一定差异,而基于合并数据的系统树支持率和分辨率都高于基于单基因的系统树,表明使用在遗传上相对独立的分子数据合并建立系统树有较高的可信度和分辨率,是一种更好的研究方法.     

Consequences of recombination on traditional phylogenetic analysis

We investigate the shape of a phylogenetic tree reconstructed from sequences evolving under the coalescent with recombination. The motivation is that evolutionary inferences are often made from phylogenetic trees reconstructed from population data even though recombination may well occur (mtDNA or viral sequences) or does occur (nuclear sequences). We investigate the size and direction of biases when a single tree is reconstructed ignoring recombination. Standard software (PHYLIP) was used to construct the best phylogenetic tree from sequences simulated under the coalescent with recombination. With recombination present, the length of terminal branches and the total branch length are larger, and the time to the most recent common ancestor smaller, than for a tree reconstructed from sequences evolving with no recombination. The effects are pronounced even for small levels of recombination that may not be immediately detectable in a data set. The phylogenies when recombination is present superficially resemble phylogenies for sequences from an exponentially growing population. However, exponential growth has a different effect on statistics such as Tajima's D. Furthermore, ignoring recombination leads to a large overestimation of the substitution rate heterogeneity and the loss of the molecular clock. These results are discussed in relation to viral and mtDNA data sets.     

Dinoflagellate phylogeny as inferred from heat shock protein 90 and ribosomal gene sequences

Background

Interrelationships among dinoflagellates in molecular phylogenies are largely unresolved, especially in the deepest branches. Ribosomal DNA (rDNA) sequences provide phylogenetic signals only at the tips of the dinoflagellate tree. Two reasons for the poor resolution of deep dinoflagellate relationships using rDNA sequences are (1) most sites are relatively conserved and (2) there are different evolutionary rates among sites in different lineages. Therefore, alternative molecular markers are required to address the deeper phylogenetic relationships among dinoflagellates. Preliminary evidence indicates that the heat shock protein 90 gene (Hsp90) will provide an informative marker, mainly because this gene is relatively long and appears to have relatively uniform rates of evolution in different lineages.

Methodology/Principal Findings

We more than doubled the previous dataset of Hsp90 sequences from dinoflagellates by generating additional sequences from 17 different species, representing seven different orders. In order to concatenate the Hsp90 data with rDNA sequences, we supplemented the Hsp90 sequences with three new SSU rDNA sequences and five new LSU rDNA sequences. The new Hsp90 sequences were generated, in part, from four additional heterotrophic dinoflagellates and the type species for six different genera. Molecular phylogenetic analyses resulted in a paraphyletic assemblage near the base of the dinoflagellate tree consisting of only athecate species. However, Noctiluca was never part of this assemblage and branched in a position that was nested within other lineages of dinokaryotes. The phylogenetic trees inferred from Hsp90 sequences were consistent with trees inferred from rDNA sequences in that the backbone of the dinoflagellate clade was largely unresolved.

Conclusions/Significance

The sequence conservation in both Hsp90 and rDNA sequences and the poor resolution of the deepest nodes suggests that dinoflagellates reflect an explosive radiation in morphological diversity in their recent evolutionary past. Nonetheless, the more comprehensive analysis of Hsp90 sequences enabled us to infer phylogenetic interrelationships of dinoflagellates more rigorously. For instance, the phylogenetic position of Noctiluca, which possesses several unusual features, was incongruent with previous phylogenetic studies. Therefore, the generation of additional dinoflagellate Hsp90 sequences is expected to refine the stem group of athecate species observed here and contribute to future multi-gene analyses of dinoflagellate interrelationships.     

Evolution of the nad3-rps12 Gene Cluster in Angiosperm Mitochondria: Comparison of Edited and Unedited Sequences

We have analyzed the nad3-rps12 locus for eight angiosperms in order to compare the utility of mitochondrial DNA and edited mRNA sequences in phylogenetic reconstruction. The two coding regions, containing from 25 to 35 editing sites in the various plants, have been concatenated in order to increase the significance of the analysis. Differing from the corresponding chloroplast sequences, unedited mitochondrial DNA sequences seem to evolve under a quasi-neutral substitution process which undifferentiates the nucleotide substitution rates for the three codon positions. By using complete gene sequences (all codon positions) we found that genomic sequences provide a classical angiosperm phylogenetic tree with a clear-cut grouping of monocotyledons and dicotyledons with Magnoliidae at the basal branch of the tree. Conversely, owing to their low nucleotide substitution rates, edited mRNA sequences were found not to be suitable for studying phylogenetic relationships among angiosperms. Received: 24 January 1996 / Accepted: 5 June 1996     

Community structure of arbuscular mycorrhizal fungi in undisturbed vegetation revealed by analyses of LSU rDNA sequences

Arbuscular mycorrhizal fungi (AMF) form a mutualistic symbiosis with plant roots and are found in most ecosystems. In this study the community structure of AMF in a clade of the genus Glomus was examined in undisturbed costal grassland using LSU rDNA sequences amplified from roots of Hieracium pilosella. Roots were sampled from May to November along eight 30-m transects, 30-120 m apart. Phylogenetic analysis of the sequences revealed 11 phylogenetic clusters within the clade of Glomus. The phylogenetic clusters were patchily distributed within the area; time had no influence on the distribution pattern. The dominant cluster covered up to 10 m along the transect, whereas other clusters formed what can be interpreted as small individual mycelia. Four of the phylogenetic clusters included known species; the other clusters, including the dominant sequence types, were unknown. The dominant phylogenetic cluster enclosed nine haplotypes, and analyses of genetic diversity of this phylogenetic cluster showed that the total diversity could be found within single root fragments, suggesting that the multiple sequences were derived from a single individual.     

A comparison of 18s ribosomal RNA and rubisco large subunit sequences for studying angiosperm phylogeny

Summary Partial sequences of 18s rRNA were obtained for 2 gymnosperms and 12 angiosperms from a wide range of families and these were analyzed with 5 other published sequences to form a phylogenetic tree. Using 16 published sequences of the large subunit of rubisco (rbcL), also from a wide range of angiosperm families, another phylogenetic tree was derived and the two approaches were compared. Both phylogenetic trees gave good grouping within families but in neither case was there resolution of the branching order of major taxa. Superficially the long rbcL sequences (whose base composition was homogeneous among all species) seemed very promising, but analysis showed that a large proportion of the variation did not affect the amino acid sequence. Although silent substitution contained some phylogenetic information, at the level required to order major taxa, much of it was random and obfuscating. It was concluded that neither macromolecule alone was likely to yield a solution to the problem of angiosperm phylogeny and therefore that studies of both, at least, will be required. For this reason, a method wa described for obtaining both DNA and RNA of good quality from the same preparation and which had been used successfully with a wide range of species including many with pungent leaves.     

Resolving deep phylogenetic relationships in salamanders: analyses of mitochondrial and nuclear genomic data

Phylogenetic relationships among salamander families illustrate analytical challenges inherent to inferring phylogenies in which terminal branches are temporally very long relative to internal branches. We present new mitochondrial DNA sequences, approximately 2,100 base pairs from the genes encoding ND1, ND2, COI, and the intervening tRNA genes for 34 species representing all 10 salamander families, to examine these relationships. Parsimony analysis of these mtDNA sequences supports monophyly of all families except Proteidae, but yields a tree largely unresolved with respect to interfamilial relationships and the phylogenetic positions of the proteid genera Necturus and Proteus. In contrast, Bayesian and maximum-likelihood analyses of the mtDNA data produce a topology concordant with phylogenetic results from nuclear-encoded rRNA sequences, and they statistically reject monophyly of the internally fertilizing salamanders, suborder Salamandroidea. Phylogenetic simulations based on our mitochondrial DNA sequences reveal that Bayesian analyses outperform parsimony in reconstructing short branches located deep in the phylogenetic history of a taxon. However, phylogenetic conflicts between our results and a recent analysis of nuclear RAG-1 gene sequences suggest that statistical rejection of a monophyletic Salamandroidea by Bayesian analyses of our mitochondrial genomic data is probably erroneous. Bayesian and likelihood-based analyses may overestimate phylogenetic precision when estimating short branches located deep in a phylogeny from data showing substitutional saturation; an analysis of nucleotide substitutions indicates that these methods may be overly sensitive to a relatively small number of sites that show substitutions judged uncommon by the favored evolutionary model.     

Phylogeny of protostome moulting animals (Ecdysozoa) inferred from 18 and 28S rRNA gene sequences

Reliability of reconstruction of phylogenetic relationships within a group of protostome moulting animals was evaluated by means of comparison of 18 and 28S rRNA gene sequences sets both taken separately and combined. Reliability of reconstructions was evaluated by values of the bootstrap support of major phylogenetic tree nodes and by degree of congruence of phylogenetic trees inferred by various methods. By both criteria, phylogenetic trees reconstructed from the combined 18 and 28S rRNA gene sequences were better than those inferred from 18 and 28S sequences taken separately. Results obtained are consistent with phylogenetic hypothesis separating protostome animals into two major clades, moulting Ecdysozoa (Priapulida + Kinorhyncha, Nematoda + Nematomorpha, Onychophora + Tardigrada, Myriapoda + Chelicerata, Crustacea + Hexapoda) and unmoulting Lophotrochozoa (Plathelminthes, Nemertini, Annelida, Mollusca, Echiura, Sipuncula). Clade Cephalorhyncha does not include nematomorphs (Nematomorpha). Conclusion was taken that it is necessary to use combined 18 and 28S data in phylogenetic studies.     

Effect of co-evolving amino acid residues on topology of phylogenetic trees

The presence in proteins of amino acid residues that change in concert during evolution is associated with keeping constant the protein spatial structure and functions. As in the case with morphological features, correlated substitutions may become the cause of homoplasies--the independent evolution of identical non-homological adaptations. Our data obtained on model phylogenetic trees and corresponding sets of sequences have shown that the presence of correlated substitutions distorts the results of phylogenetic reconstructions. A method for accounting for co-evolving amino acid residues in phylogenetic analysis is proposed. According to this method, only a single site from the group of correlated amino acid positions should remain, whereas other positions should not be used in further phylogenetic analysis. Simulations performed have shown that replacement on the average of 8% of variable positions in a pair of model sequences by coordinately evolving amino acid residues is able to change the tree topology. The removal of such amino acid residues from sequences before phylogenetic analysis restores the correct topology.     

基于线粒体COI基因全序列的直翅目部分类群系统发育关系分析

该研究基于直翅目56种昆虫的COI基因全序列构建了该目部分类群间的系统发育关系,同时也分析了COI基因编码的氨基酸序列构建直翅目系统发育关系的可靠性。将COI序列按照密码子一、二、三位点划分,分别计算PBS(partioned Bremer support)值,评估蛋白质编码基因密码子不同位点的系统发生信号强度。分析结果支持螽亚目和蝗亚目的单系性;剑角蝗科、斑腿蝗科、斑翅蝗科、网翅蝗科和槌角蝗科5科均不是单系群,科间的遗传距离在0.107~0.153之间变化,与其他科相比遗传距离较小,符合将这5科合并为一科(即蝗科)的分类系统,瘤锥蝗科和锥头蝗科归为锥头蝗总科,癞蝗科单独成为一科,这也与Otte(1997)系统的划分一致。根据PBS值的大小推断密码子第三、第一位点对系统树分支的贡献比第二位点大,并且较长的序列含有较多的信息位点。研究也证实将各物种COI基因之间的遗传距离作为直翅目划分科级阶元的工具是可行的。     

Amplification of mitochondrial small subunit ribosomal DNA of polypores and its potential for phylogenetic analysis

There has been a systematic need to seek adequate phylogenetic markers that can be applied in phylogenetic analyses of fungal taxa at various levels. The mitochondrial small subunit ribosomal DNA (mt SSU rDNA) is generally considered to be one of the molecules that are appropriate for phylogenetic analyses at a family level. In order to obtain universal primers for polypores of Hymenomycetes, mt SSU rRNA genes were cloned from Bjerkandera adusta, Ganoderma lucidum, Phlebiopsis gigantea, and Phellinus laevigatus and their sequences were determined. Based on the conserved sequences of cloned genes from polypores and Agrocybe aegerita, PCR primers were designed for amplification and sequencing of mt SSU rDNAs. New primers allowed effective amplification and sequencing of almost full-sized genes from representative species of polypores and related species. Phylogenetic relationships were resolved quite efficiently by mt SSU rDNA sequences, and they proved to be more useful in phylogenetic reconstruction of Ganoderma than nuclear internal transcribed spacer (ITS) rDNA sequences.     

Conflicting phylogenetic signals at the base of the metazoan tree

A phylogenetic framework is essential for under-standing the origin and evolution of metazoan development. Despite a number of recent molecular studies and a rich fossil record of sponges and cnidarians, the evolutionary relationships of the early branching metazoan groups to each other and to a putative outgroup, the choanoflagellates, remain uncertain. This situation may be the result of the limited amount of phylogenetic information found in single genes and the small number of relevant taxa surveyed. To alleviate the effect of these analytical factors in the phylogenetic recons-truction of early branching metazoan lineages, we cloned multiple protein-coding genes from two choanoflagellates and diverse sponges, cnidarians, and a ctenophore. Comparisons of sequences for alpha-tubulin, beta-tubulin, elongation factor 2, HSP90, and HSP70 robustly support the hypothesis that choanoflagellates are closely affiliated with animals. However, analyses of single and concatenated amino acid sequences fail to resolve the relationships either between early branching metazoan groups or between Metazoa and choano-flagellates. We demonstrate that variable rates of evolution among lineages, sensitivity of the analyses to taxon selection, and conflicts in the phylogenetic signal contained in different amino acid sequences obscure the phylogenetic associations among the early branching Metazoa. These factors raise concerns about the ability to resolve the phylogenetic history of animals with molecular sequences. A consensus view of animal evolution may require investigations of genome-scale characters.     

Phylogenetic tree of bacterial and eucaryotic FtsZ-proteins based on the homology of their primary sequences

In this report we present phylogenetic tree based on the sequences of FtsZ-proteins of Archaea, Bacteria and Eukaryota. We have analyzed the relative positions of separate sequences and their clusters and their distance to the hypothetical root of phylogenetic tree as well. It has been demonstrated that most of procaryotic and eucaryotic FtsZ-protein sequences are located at the distance of 0.3-0.8 substitutions per site from the hypothetical root of phylogenetic tree. The tubulins are outlined from the root at 7-8 fold larger distances in comparison with the separate sequences. This confirms the slight aminoacid homology between these two groups of cytoskeletal proteins. At the same time the significant similarity between FtsZ-proteins of cyanobacteria and of plant plastids has been shown.