Variation patterns of the mitochondrial 16S rRNA gene with secondary structure constraints and their application to phylogeny of cyprinine fishes (Teleostei: Cypriniformes)

The mitochondrial 16S ribosomal RNA (rRNA) gene sequences from 93 cyprinid fishes were examined to reconstruct the phylogenetic relationships within the diverse and economically important subfamily Cyprininae. Within the subfamily a biased nucleotide composition (A>T, C>G) was observed in the loop regions of the gene, and in stem regions apparent selective pressures of base pairing showed a bias in favor of G over C and T over A. The bias may be associated with transition-transversion bias. Rates of nucleotide substitution were lower in stems than in loops. Analysis of compensatory substitutions across these taxa demonstrates 68% covariation in the gene and a logical weighting factor to account for dependence in mutations for phylogenetic inference should be 0.66. Comparisons of varied stem-loop weighting schemes indicate that the down-weightings for stem regions could improve the phylogenetic analysis and the degree of non-independence of stem substitutions was not as important as expected. Bayesian inference under four models of nucleotide substitution indicated that likelihood-based phylogenetic analyses were more effective in improving the phylogenetic performance than was weighted parsimony analysis. In Bayesian analyses, the resolution of phylogenies under the 16-state models for paired regions, incorporating GTR + G + I models for unpaired regions was better than those under other models. The subfamily Cyprininae was resolved as a monophyletic group, as well as tribe Labein and several genera. However, the monophyly of the currently recognized tribes, such as Schizothoracin, Barbin, Cyprinion + Onychostoma lineages, and some genera was rejected. Furthermore, comparisons of the parsimony and Bayesian analyses and results of variable length bootstrap analysis indicates that the mitochondrial 16S rRNA gene should contain important character variation to recover well-supported phylogeny of cyprinid taxa whose divergences occurred within the recent 8 MY, but could not provide resolution power for deep phylogenies spanning 10-19 MYA.     

以子囊菌部分类群的rRNA数据为例,评估不同分区策略对贝叶斯分析效果的影响(英文)

在使用rRNA基因进行系统发育分析过程中,不同位点间进化速度的差异性可能是导致系统误差的一个重要原因。以52个真菌为研究对象,利用rRNA二级结构特征构建分区策略,探讨不同分区策略对贝叶斯分析的影响。结果显示各结构分区的最优核酸替代模型及其参数与分区类型密切相关。与传统的贝叶斯方法相比,使用结构环的分区策略对结果没有显著影响,而引入臂元素的方法却导致更高的边际似然值和支持率。此外,不考虑结构特征,简单的增加子分区数量的分区策略尽管也能导致贝叶斯因素值的增加,却没有提高解决亲缘关系的能力,说明一个合理的分区策略应该基于生物学功能(或二级结构特征)而非纯数学因素。     

Ribosomal RNA secondary structure: compensatory mutations and implications for phylogenetic analysis

Using sequence data from the 28S ribosomal RNA (rRNA) genes of selected vertebrates, we investigated the effects that constraints imposed by secondary structure have on the phylogenetic analysis of rRNA sequence data. Our analysis indicates that characters from both base-pairing regions (stems) and non-base-pairing regions (loops) contain phylogenetic information, as judged by the level of support of the phylogenetic results compared with a well-established tree based on both morphological and molecular data. The best results (the greatest level of support of well-accepted nodes) were obtained when the complete data set was used. However, some previously supported nodes were resolved using either the stem or loop bases alone. Stem bases sustain a greater number of compensatory mutations than would be expected at random, but the number is < 40% of that expected under a hypothesis of perfect compensation to maintain secondary structure. Therefore, we suggest that in phylogenetic analyses, the weighting of stem characters be reduced by no more than 20%, relative to that of loop characters. In contrast to previous suggestions, we do not recommend weighting of stem positions by one-half, compared with that of loop positions, because this overcompensates for the constraints that selection imposes on the secondary structure of rRNA.      

Secondary structure models of the nuclear internal transcribed spacer regions and 5.8S rRNA in Calciodinelloideae (Peridiniaceae) and other dinoflagellates

Secondary structure models of the 5.8S rRNA and both internal transcribed spacers (ITS1 and ITS2) are proposed for Calciodinelloideae (Peridiniaceae) and are also plausible for other dinoflagellates. The secondary structure of the 5.8S rRNA corresponds to previously developed models, with two internal paired regions and at least one 5.8S rRNA–28S rRNA interaction. A general secondary structure model of ITS1 for Calciodinelloideae (and other dinoflagellates), consisting of an open multibranch loop with three major helices, is proposed. The homology of these paired regions with those found in other taxa, published in previous studies (e.g. yeast, green algae and Platyhelmithes) remains to be determined. Finally, a general secondary structure model of ITS2 for Calciodinelloideae (and other dinoflagellates) is reconstructed. Based on the 5.8S rRNA–28S rRNA interaction, it consists of a closed multibranch loop, with four major helices. At least helix III and IV have homology with paired regions found in other eukaryotic taxa (e.g. yeast, green algae and vertebrates). Since the secondary structures of both ITS regions are more conserved than the nucleotide sequences, their analysis helps in understanding molecular evolution and increases the number of structural characters. Thus, the structure models developed in this study may be generally useful for future phylogenetic analyses.     

Rate acceleration and long-branch attraction in a conserved gene of cryptic daphniid (Crustacea) species.

The nuclear large subunit (LSU) rRNA gene is a rich source of phylogenetic characters because of its large size, mosaic of slowly and rapidly evolving regions, and complex secondary structure variation. Nevertheless, many studies have indicated that inconsistency, bias, and gene-specific error (e.g., within-individual gene family variation, cryptic sequence simplicity, and sequence coevolution) can complicate animal phylogenies based on LSU rDNA sequences. However, most of these studies sampled small gene fragments from expansion segments--among animals only five nonchordate complete LSU sequences are published. In this study, we sequenced near-complete nuclear LSU genes from 11 representative daphniids (Crustacea). The daphniid expansion segment V6 was larger and showed more length variation (90-351 bp) than is found in all other reported LSU V6 sequences. Daphniid LSU (without the V6 region) phylogenies generally agreed with the existing phylogenies based on morphology and mtDNA sequences. Nevertheless, a major disagreement between the LSU and the expected trees involved a positively misleading association between the two taxa with the longest branches, Daphnia laevis and D. occidentalis. Both maximum parsimony (MP) and maximum likelihood (ML) optimality criteria recovered this association, but parametric simulations indicated that MP was markedly more sensitive to this bias than ML. Examination of data partitions indicated that the inconsistency was caused by increased nucleotide substitution rates in the branches leading to D. laevis and D. occidentalis rather than among-taxon differences in base composition or distribution of sites that are free to vary. These results suggest that lineage-specific rate acceleration can lead to long-branch attraction even in the conserved genes of animal species that are almost morphologically indistinguishable.     

DISCONTINUOUS MITOCHONDRIAL AND CHLOROPLAST LARGE SUBUNIT RIBOSOMAL RNAs AMONG GREEN ALGAE: PHYLOGENETIC IMPLICATIONS1

To provide insights into the occurrence, evolution, and phylogenetic distribution of discontinuous mitochondrial and chloroplast large subunit ribosomal RNAs (LSU rRNAs) among green algae, we surveyed 12 taxa representing three classes of green algae: the Chlorophyceae, Pleurastrophyceae, and Micromonadophyceae (sensu Mattox and Stewart 1984). We present evidence that discontinuous mitochondrial and chloroplast LSU rRNAs are quite widespread among green algae. Mitochondrial LSU rRNAs appear discontinuous in zoosporic chlorophycean lineages displaying a clockwise or directly opposed configuration in their flagellar apparatus, as well as in chlorococcalean autosporic taxa phylogenetically related to them, but are continuous among zoosporic green algal lineages with a counterclockwise flagellar apparatus configuration, as well as among chlorococcalean autosporic taxa phylogenetically related to them. Chloroplast LSU rRNAs appear discontinuous in all of the lineages investigated. Discontinuous mitochondrial LSU rRNA represents a molecular trait that might have originated at or near the base of Chlorophyceae, whereas discontinuous chloroplast LSU rRNA might have developed very early in the evolutionary history of the green algal group itself. We suggest, therefore, that the presence of discontinuous mitochondrial but not chloroplast LSU rRNA can be used as an additional character in assessing phylogenetic affiliations among green algae.     

Molecular phylogenetics and the evolution of labral spines among eastern Pacific ocenebrine gastropods

Labral spines are sharp projections of the apertural lip found in some marine gastropods that are used to penetrate hard-shelled prey. The majority of gastropod genera that contain labral spine-bearing species are found in the subfamily Ocenebrinae (Gastropoda: Muricidae). To reconstruct the evolutionary history of labral spine-bearing and labral spine-lacking gastropods in the eastern Pacific (EP) Ocean, partial sequences of two mitochondrial genes (cytochrome oxidase I and 12S rRNA) were obtained from representative taxa. Despite high nucleotide bias, a variety of phylogenetic reconstruction methods produced the same tree topology. The traditional taxonomic view that all "Nucella-like" spine-bearing taxa in the EP belong to a monophyletic "Acanthina" is rejected due to nonmonophyly of this group. The more recently recognized "Acanthinucella" is also not monophyletic, and we therefore propose the new genus Mexacanthina for two Mexican species formerly assigned to Acanthinucella. The genus Ocinebrina, which first appears in the middle Eocene, is not a stem EP ocenebrine lineage and may also not be a monophyletic clade. Tracing the evolutionary history of labral spines among extant lineages indicates that the absence of a labral spine is ancestral for all EP ocenebrines. Ancestral conditions could not be resolved unambiguously for all nodes of the phylogeny based on extant taxa. However, by jointly considering both molecular phylogenetic relationships and the phylogenetic affinities of several extinct taxa, all remaining character state transformation can be inferred unambiguously. Based on this analysis, a labral spine likely evolved independently in at least four lineages of EP ocenebrines. Although homoplasy appears to characterize labral spine evolution among ocenebrine gastropods, the structural position of a labral spine was evolutionarily altered in one lineage, indicating that different types of labral spines do not necessarily reflect convergent evolution.     

Sequence heterogeneity in the duplicate large subunit ribosomal RNA genes of Tetrahymena pyriformis mitochondrial DNA

In the ciliated protozoan, Tetrahymena pyriformis, the mitochondrial large subunit ribosomal RNA (LSU rRNA) is discontinuous, consisting of two discrete RNA species: a 280-nucleotide LSU alpha (constituting the 5'-portion) and a 2315-nucleotide LSU beta (corresponding to the remaining 3'-portion of this rRNA). The T. pyriformis mitochondrial genome contains two copies of the LSU alpha.beta gene complex, and we have previously provided evidence that both copies are transcribed (Heinonen, T. Y. K., Schnare, M. N., Young, P. G., and Gray, M. W. (1987) J. Biol. Chem. 262, 2879-2887). We now report the complete sequences of the two copies of the LSU alpha.beta gene complex. These are not identical, but differ at 5 out of the 2595 positions by single nucleotide substitutions in one sequence relative to the other. In the secondary structure model we propose here, two of these differences are located in base-paired regions of the LSU rRNA; however, they do not interrupt the complementary interactions in these helices. The other three differences occur in single-stranded regions of the secondary structure. The base substitutions documented here are not localized to those regions of LSU rRNA that are the most highly conserved in global phylogenetic comparisons, and therefore it seems unlikely that they are of fundamental functional significance. Whether they might exert more subtle effects on ribosome function remains to be determined.     

Phylogenetic signal and its decay in mitochondrial SSU and LSU rRNA gene fragments of Anisoptera

The phylogeny of Anisoptera, dragonflies in the strict sense, has proven to be notoriously difficult to resolve. Based on morphological characters, several recent publications dealing with the phylogeny of dragonflies proposed contradicting inter- and intrafamily relationships. We explored phylogenetic information content of mitochondrial large-subunit (LSU) and small-subunit (SSU) ribosomal gene fragments for these systematic problems. Starting at published universal primers, we developed primer sets suitable for amplifying large parts of the LSU and SSU rRNA genes within dragonflies. These fragments turned out to harbor sufficient phylogenetic information to satisfyingly resolve intrafamily relationships, but they contain insufficient phylogenetic structure to permit reliable conclusions about several interfamily relationships. We demonstrate that decay of phylogenetic signal progresses from intrafamily to interfamily to outgroup relationships and is correlated with an increase of genetic distances. As expected, signal decay is most pronounced in fast-changing sites. Additionally, base composition among fast-changing sites significantly deviates from the expected homogeneity. Homogeneity of base composition among all included taxa was restored only after removing fast-changing sites from the data set. The molecular data tentatively support interfamily relationships proposed by the most recent publication based on morphological characters of fossil and extant dragonflies.     

Secondary structure of mitochondrial 12S rRNA among fish and its phylogenetic applications.

The complete 12S ribosomal RNA(rRNA) sequences from 23 gobioid species and nine diverse assortments of other fish species were employed to establish a core secondary structure model for fish 12S rRNA. Of the 43 stems recognized, 41 were supported by at least some compensatory evidence among vertebrates. The rates of nucleotide substitution were lower in stems than in loops. This may produce less phylogenetic information in stems when recently diverged taxa are compared. An analysis of compensatory substitution shows that the percentage of covariation is 68%, and the weighting factor for phylogenetic analyses to account for the dependence of mutations should be 0.66. Different stem-loop weighting schemes applied to the analyses of phylogenetic relationships of the Gobioidei indicate that down-weighting paired regions because of nonindependence could not improve the present phylogenetic analysis. A biased nucleotide composition (adenine% [A%] > thymine% [T%], cytosine% [C%] > guanine% [G%]) in the loop regions was also observed in the mammalian counterpart. The excess of A and C in the loop regions may be because of the asymmetric mechanism of mtDNA replication, which leads to the spontaneous deamination of C and A. This process may also be responsible for a transition-transversion bias and the patterns of nucleotide substitutions in both stems and loops.     

Limitations of Metazoan 18S rRNA Sequence Data: Implications for Reconstructing a Phylogeny of the Animal Kingdom and Inferring the Reality of the Cambrian Explosion

We document the phylogenetic behavior of the 18S rRNA molecule in 67 taxa from 28 metazoan phyla and assess the effects of among-site rate variation on reconstructing phylogenies of the animal kingdom. This empirical assessment was undertaken to clarify further the limits of resolution of the 18S rRNA gene as a phylogenetic marker and to address the question of whether 18S rRNA phylogenies can be used as a source of evidence to infer the reality of a Cambrian explosion. A notable degree of among-site rate variation exists between different regions of the 18S rRNA molecule, as well as within all classes of secondary structure. There is a significant negative correlation between inferred number of nucleotide substitutions and phylogenetic information, as well as with the degree of substitutional saturation within the molecule. Base compositional differences both within and between taxa exist and, in certain lineages, may be associated with long branches and phylogenetic position. Importantly, excluding sites with different degrees of nucleotide substitution significantly influences the topology and degree of resolution of maximum-parsimony phylogenies as well as neighbor-joining phylogenies (corrected and uncorrected for among-site rate variation) reconstructed at the metazoan scale. Together, these data indicate that the 18S rRNA molecule is an unsuitable candidate for reconstructing the evolutionary history of all metazoan phyla, and that the polytomies, i.e., unresolved nodes within 18S rRNA phylogenies, cannot be used as a single or reliable source of evidence to support the hypothesis of a Cambrian explosion. Received: 9 December 1997 / Accepted: 23 March 1998     

Placozoa are not derived cnidarians: evidence from molecular morphology

The phylum Placozoa is represented by a single known species, Trichoplax adhaerens, a tiny marine organism that represents the most simple metazoan bauplan. Because of the latter, placozoans were originally considered the most basal metazoan phylum. A misinterpretation of the life cycle at the turn of the century and some more recent molecular phylogenetic analyses have placed Trichoplax as a derived species within the Cnidaria. The latter hypothesis assumes that the primitive organization of the Placozoa is the result of secondary reduction. Here we compare the molecular morphology of the predicted 16S rDNA structure and the mitochondrial genome between Trichoplax and representatives of all four cnidarian classes. Trichoplax shares a circular mtDNA molecule as a plesiomorphy with all other metazoans except for the derived cnidarian classes Hydrozoa, Scyphozoa, and Cubozoa. The predicted secondary structure of the 16S rRNA molecule differs substantially between Trichoplax and cnidarians, particularly with respect to the number and length of stem and loop regions. The new molecular morphological characters provide compelling evidence that Trichoplax is not a derived (medusozoan) cnidarian. Furthermore, it was found that the mitochondrial genome in Cubozoa consists of four linear molecules instead of a single circular molecule or two linear molecules, suggesting that the cubozoans may represent the most derived cnidarian group.     

Investigation of molluscan phylogeny using large-subunit and small-subunit nuclear rRNA sequences

The Mollusca represent one of the most morphologically diverse animal phyla, prompting a variety of hypotheses on relationships between the major lineages within the phylum based upon morphological, developmental, and paleontological data. Analyses of small-ribosomal RNA (SSU rRNA) gene sequence have provided limited resolution of higher-level relationships within the Mollusca. Recent analyses suggest large-subunit (LSU) rRNA gene sequences are useful in resolving deep-level metazoan relationships, particularly when combined with SSU sequence. To this end, LSU (approximately 3.5 kb in length) and SSU (approximately 2 kb) sequences were collected for 33 taxa representing the major lineages within the Mollusca to improve resolution of intraphyletic relationships. Although the LSU and combined LSU+SSU datasets appear to hold potential for resolving branching order within the recognized molluscan classes, low bootstrap support was found for relationships between the major lineages within the Mollusca. LSU+SSU sequences also showed significant levels of rate heterogeneity between molluscan lineages. The Polyplacophora, Gastropoda, and Cephalopoda were each recovered as monophyletic clades with the LSU+SSU dataset. While the Bivalvia were not recovered as monophyletic clade in analyses of the SSU, LSU, or LSU+SSU, the Shimodaira-Hasegawa test showed that likelihood scores for these results did not differ significantly from topologies where the Bivalvia were monophyletic. Analyses of LSU sequences strongly contradict the widely accepted Diasoma hypotheses that bivalves and scaphopods are closely related to one another. The data are consistent with recent morphological and SSU analyses suggesting scaphopods are more closely related to gastropods and cephalopods than to bivalves. The dataset also presents the first published DNA sequences from a neomeniomorph aplacophoran, a group considered critical to our understanding of the origin and early radiation of the Mollusca.     

Patterns of nucleotide change in mitochondrial ribosomal RNA genes and the phylogeny of piranhas

The patterns and rates of nucleotide substitution in mitochondrial ribosomal RNA genes are described and applied in a phylogenetic analysis of fishes of the subfamily Serrasalminae (Teleostei, Characiformes, Characidae). Fragments of 345 bp of the 12S and 535 bp of the 16S genes were sequenced for 37 taxa representing all but three genera in the subfamily. Secondary-structure models based on comparative sequence analysis were derived to characterize the pattern of change among paired and unpaired nucleotides, forming stem and loop regions, respectively. Base compositional biases were in the direction of A-rich loops and G-rich stems. Ninety-five percent of substitutions in stem regions were compensatory mutations, suggesting that selection for maintenance of base pairing is strong and that independence among characters cannot be assumed in phylogenetic analyses of stem characters. The relative rate of nucleotide substitution was similar in both fragments sequenced but higher in loop than in stem regions. In both genes, C-T transitions were the most common type of change, and overall transitions outnumbered transversions by a factor of two in 16S and four in 12S. Phylogenetic analysis of the mitochondrial DNA sequences suggests that a clade formed by the generaPiaractus, Colossoma, andMylossoma is the sister group to all other serrasalmins and that the generaMyleus, Serrasalmus, andPristobrycon are paraphyletic. A previous hypothesis concerning relationships for the serrasalmins, based on morphological evidence, is not supported by the molecular data. However, phylogenetic analysis of host-specific helminth parasites and cytogenetic data support the phylogeny of the Serrasalminae obtained in this study and provide evidence for coevolution between helminth parasites and their fish hosts.     

Phylogenetic analysis of the mitochondrial genes LSU rRNA and COI suggests early adaptive differentiation of anal teeth in chrysidine cuckoo wasps (Hymenoptera: Chrysididae)

Cuckoo wasps are a morphologically diverse group of Hymenoptera with parasitoid or cleptoparasitic life histories. In the present paper, we explore the phylogenetic signal in fragments of the mitochondrial genes LSU rRNA and COI to resolve the group's phylogeny. We analyzed sequence data of 33 species representing the taxa Cleptinae, Elampini, Parnopini, and Chrysidini. Most of the currently recognized relationships of major cuckoo wasp lineages are supported by the molecular data. A key difference concerns the phylogenetic position of the Euchroeus (=Brugmoia) group within the tribe Chrysidini. It seems likely that an erroneous interpretation of morphological characters has led to inappropriate rooting of that tribe. We suggest that species of the Euchroeus group be interpreted as forming the stem group of the Chrysidini and that the remaining genera of that tribe be united in a subordinated taxon. Our results imply that the evolution of anal dentition, of significance for breaking into sealed host nests otherwise not accessible to cuckoo wasps, already happened at the base of the Chrysidini and that an even number of anal teeth arose prior to an odd number.     

rRNA二级结构序列用于真菌系统学研究的方法初探

本文首次利用核酸二级结构特征代替核酸碱基作为探讨类群之间亲缘关系的信号,构建了基于结构特征的子囊菌部分类群的系统进化树。该方法以S(规范的碱基对),Q(不规范的碱基对),I(单链),B(侧环),M(多分枝环)和H(发卡结构)为代码将二级结构特征区分为6种不同的亚结构类型,然后将二级结构特征转换为结构序列,并进行结构序列分析。该方法使rRNA不只局限于碱基比较,拓展了其应用范围,为揭示分子的功能与进化的关系提供了线索。结果表明,结构序列分析可用于子囊菌的系统学研究;相对于核酸序列分析,结构分析的结果似乎更加清晰地体现子囊果的演化过程。     

Sequence and organization of large subunit rRNA genes from the extrachromosomal 35 kb circular DNA of the malaria parasite Plasmodium falciparum.

The malaria parasite Plasmodium falciparum carries an extrachromosomal 35 kb circular DNA molecule of unknown provenance. A striking feature of the circle is a palindromic sequence of genes for subunit rRNAs and several tRNAs, spanning ca. 10.5 kb. The palindrome has an intriguing resemblance to the inverted repeat of plastid genomes, and the sequence and putative secondary structure of the malarial large subunit (LSU) rRNA described in this report were used as the basis of a phylogenetic study. The malarial rRNA was found to be highly divergent in comparison with a selected group of chloroplast LSU rRNAs but was more closely related to them than to mitochondrial LSU rRNA genes.