The Strepsiptera problem: phylogeny of the holometabolous insect orders inferred from 18S and 28S ribosomal DNA sequences and morphology

Phylogenetic relationships among the holometabolous insect orders were inferred from cladistic analysis of nucleotide sequences of 18S ribosomal DNA (rDNA) (85 exemplars) and 28S rDNA (52 exemplars) and morphological characters. Exemplar outgroup taxa were Collembola (1 sequence), Archaeognatha (1), Ephemerida (1), Odonata (2), Plecoptera (2), Blattodea (1), Mantodea (1), Dermaptera (1), Orthoptera (1), Phasmatodea (1), Embioptera (1), Psocoptera (1), Phthiraptera (1), Hemiptera (4), and Thysanoptera (1). Exemplar ingroup taxa were Coleoptera: Archostemata (1), Adephaga (2), and Polyphaga (7); Megaloptera (1); Raphidioptera (1); Neuroptera (sensu stricto = Planipennia): Mantispoidea (2), Hemerobioidea (2), and Myrmeleontoidea (2); Hymenoptera: Symphyta (4) and Apocrita (19); Trichoptera: Hydropsychoidea (1) and Limnephiloidea (2); Lepidoptera: Ditrysia (3); Siphonaptera: Pulicoidea (1) and Ceratophylloidea (2); Mecoptera: Meropeidae (1), Boreidae (1), Panorpidae (1), and Bittacidae (2); Diptera: Nematocera (1), Brachycera (2), and Cyclorrhapha (1); and Strepsiptera: Corioxenidae (1), Myrmecolacidae (1), Elenchidae (1), and Stylopidae (3). We analyzed approximately 1 kilobase of 18S rDNA, starting 398 nucleotides downstream of the 5' end, and approximately 400 bp of 28S rDNA in expansion segment D3. Multiple alignment of the 18S and 28S sequences resulted in 1,116 nucleotide positions with 24 insert regions and 398 positions with 14 insert regions, respectively. All Strepsiptera and Neuroptera have large insert regions in 18S and 28S. The secondary structure of 18S insert 23 is composed of long stems that are GC rich in the basal Strepsiptera and AT rich in the more derived Strepsiptera. A matrix of 176 morphological characters was analyzed for holometabolous orders. Incongruence length difference tests indicate that the 28S + morphological data sets are incongruent but that 28S + 18S, 18S + morphology, and 28S + 18S + morphology fail to reject the hypothesis of congruence. Phylogenetic trees were generated by parsimony analysis, and clade robustness was evaluated by branch length, Bremer support, percentage of extra steps required to force paraphyly, and sensitivity analysis using the following parameters: gap weights, morphological character weights, methods of data set combination, removal of key taxa, and alignment region. The following are monophyletic under most or all combinations of parameter values: Holometabola, Polyphaga, Megaloptera + Raphidioptera, Neuroptera, Hymenoptera, Trichoptera, Lepidoptera, Amphiesmenoptera (Trichoptera + Lepidoptera), Siphonaptera, Siphonaptera + Mecoptera, Strepsiptera, Diptera, and Strepsiptera + Diptera (Halteria). Antliophora (Mecoptera + Diptera + Siphonaptera + Strepsiptera), Mecopterida (Antliophora + Amphiesmenoptera), and Hymenoptera + Mecopterida are supported in the majority of total evidence analyses. Mecoptera may be paraphyletic because Boreus is often placed as sister group to the fleas; hence, Siphonaptera may be subordinate within Mecoptera. The 18S sequences for Priacma (Coleoptera: Archostemata), Colpocaccus (Coleoptera: Adephaga), Agulla (Raphidioptera), and Corydalus (Megaloptera) are nearly identical, and Neuropterida are monophyletic only when those two beetle sequences are removed from the analysis. Coleoptera are therefore paraphyletic under almost all combinations of parameter values. Halteria and Amphiesmenoptera have high Bremer support values and long branch lengths. The data do not support placement of Strepsiptera outside of Holometabola nor as sister group to Coleoptera. We reject the notion that the monophyly of Halteria is due to long branch attraction because Strepsiptera and Diptera do not have the longest branches and there is phylogenetic congruence between molecules, across the entire parameter space, and between morphological and molecular data.     

Evolution and phylogeny of the Diptera: a molecular phylogenetic analysis using 28S rDNA sequences

Portions of the large ribosomal subunit RNA gene (28S rDNA) encompassing the D1 and the D7 region were obtained from 16 dipteran species and families to reconstruct early phylogenetic events in the order Diptera. For outgroup comparison, the corresponding sequences were used from representative taxa of the Siphonaptera, Mecoptera, and Lepidoptera. A subset of 488 unambiguously alignable sites was analyzed with respect to important sequence evolution parameters. We found (1) sequence variability is significantly higher in double-stranded sites than in single-stranded sites, (2) transitions are close to saturation in most pairwise sequence comparisons, (3) significant substitution rate heterogeneity exists across sites, and (4) significant substitution rate heterogeneity exists among lineages. Tree reconstruction was carried out with the neighbor joining, maximum parsimony, and maximum likelihood methods. Four major subgroups are consistently and robustly supported: the Brachycera, the Culicomorpha, the Tipulomorpha sensu stricto, and the hitherto controversial Bibionomorpha sensu lato, which includes the families Sciaridae, Mycetophilidae, Cecidomyiidae, Bibionidae, Scatopsidae, and Anisopodidae. The phylogenetic relationships within or among these subclades and the positions of the families Psychodidae and Trichoceridae were not robustly resolved. These results support the view that the mouthparts of extant dipteran larvae evolved from a derived ground state characterized by subdivided and obliquely moving mandibles. Furthermore, sequence divergence and the paleontological record consistently indicate that a period of rapid cladogenesis gave rise to the major dipteran subgroups.     

Phylogenetic significance of the wing-base of the Holometabola (Insecta)

The present knowledge of the wing-base morphology of the holometabolous insects is summarized, and the value of these structures for phylogenetic analysis is demonstrated. An autapomorphy of the Holometabola is a locking mechanism composed of a knob on the basalare and a corresponding cavity on the ventral wing-base. Two synapomorphic hindwing-base characters support a sister-group relationship of Coleoptera and Neuropterida. Only few data are available on the wing-base of the Hymenoptera. An autapomorphy of the taxon is a modification of the wing locking mechanism with reduced size of the basalare and its knob. It is demonstrated that wing-base characters are helpful for the analysis of the relationships between strepsipteran families. However, characters of the wing-base support neither a relationship of Strepsiptera and Coleoptera nor of Strepsiptera and Antliophora.     

Phylogenetic relationships among insect orders based on three nuclear protein-coding gene sequences

Many attempts to resolve the phylogenetic relationships of higher groups of insects have been made based on both morphological and molecular evidence; nonetheless, most of the interordinal relationships of insects remain unclear or are controversial. As a new approach, in this study we sequenced three nuclear genes encoding the catalytic subunit of DNA polymerase delta and the two largest subunits of RNA polymerase II from all insect orders. The predicted amino acid sequences (In total, approx. 3500 amino acid sites) of these proteins were subjected to phylogenetic analyses based on the maximum likelihood and Bayesian analysis methods with various models. The resulting trees strongly support the monophyly of Palaeoptera, Neoptera, Polyneoptera, and Holometabola, while within Polyneoptera, the groupings of Isoptera/"Blattaria"/Mantodea (Superorder Dictyoptera), Dictyoptera/Zoraptera, Dermaptera/Plecoptera, Mantophasmatodea/Grylloblattodea, and Embioptera/Phasmatodea are supported. Although Paraneoptera is not supported as a monophyletic group, the grouping of Phthiraptera/Psocoptera is robustly supported. The interordinal relationships within Holometabola are well resolved and strongly supported that the order Hymenoptera is the sister lineage to all other holometabolous insects. The other orders of Holometabola are separated into two large groups, and the interordinal relationships of each group are (((Siphonaptera, Mecoptera), Diptera), (Trichoptera, Lepidoptera)) and ((Coleoptera, Strepsiptera), (Neuroptera, Raphidioptera, Megaloptera)). The sister relationship between Strepsiptera and Diptera are significantly rejected by all the statistical tests (AU, KH and wSH), while the affinity between Hymenoptera and Mecopterida are significantly rejected only by AU and KH tests. Our results show that the use of amino acid sequences of these three nuclear genes is an effective approach for resolving the relationships of higher groups of insects.     

Single-copy nuclear genes resolve the phylogeny of the holometabolous insects

Background  

Evolutionary relationships among the 11 extant orders of insects that undergo complete metamorphosis, called Holometabola, remain either unresolved or contentious, but are extremely important as a context for accurate comparative biology of insect model organisms. The most phylogenetically enigmatic holometabolan insects are Strepsiptera or twisted wing parasites, whose evolutionary relationship to any other insect order is unconfirmed. They have been controversially proposed as the closest relatives of the flies, based on rDNA, and a possible homeotic transformation in the common ancestor of both groups that would make the reduced forewings of Strepsiptera homologous to the reduced hindwings of Diptera. Here we present evidence from nucleotide sequences of six single-copy nuclear protein coding genes used to reconstruct phylogenetic relationships and estimate evolutionary divergence times for all holometabolan orders.     

18S rDNA sequences and the holometabolous insects

The Holometabola (insects with complete metamorphosis: beetles, wasps, flies, fleas, butterflies, lacewings, and others) is a monophyletic group that includes the majority of the world's animal species. Holometabolous orders are well defined by morphological characters, but relationships among orders are unclear. In a search for a region of DNA that will clarify the interordinal relationships we sequenced approximately 1080 nucleotides of the 5' end of the 18S ribosomal RNA gene from representatives of 14 families of insects in the orders Hymenoptera (sawflies and wasps), Neuroptera (lacewing and antlion), Siphonaptera (flea), and Mecoptera (scorpionfly). We aligned the sequences with the published sequences of insects from the orders Coleoptera (beetle) and Diptera (mosquito and Drosophila), and the outgroups aphid, shrimp, and spider. Unlike the other insects examined in this study, the neuropterans have A-T rich insertions or expansion regions: one in the antlion was approximately 260 bp long. The dipteran 18S rDNA evolved rapidly, with over 3 times as many substitutions among the aligned sequences, and 2-3 times more unalignable nucleotides than other Holometabola, in violation of an insect-wide molecular clock. When we excluded the long-branched taxa (Diptera, shrimp, and spider) from the analysis, the most parsimonious (minimum-length) trees placed the beetle basal to other holometabolous orders, and supported a morphologically monophyletic clade including the fleas+scorpionflies (96% bootstrap support). However, most interordinal relationships were not significantly supported when tested by maximum likelihood or bootstrapping and were sensitive to the taxa included in the analysis. The most parsimonious and maximum-likelihood trees both separated the Coleoptera and Neuroptera, but this separation was not statistically significant.(ABSTRACT TRUNCATED AT 250 WORDS)     

Long-Branch Abstractions

Recent attention has been focused on the sensitivities of various tree reconstructing algorithms to sequence rate heterogeneity (long-branch attraction). Phylogenetic conclusions from two recent empirical studies have been indicted as artifacts attributable to long-branch attraction. Siddall et al. (1995) concluded that Myxozoa are cnidarians and sister group to Polypodium based on 18S rDNA and morphology. Hanelt et al. (1996) argued that this result is due to long-branch attraction. Whiting et al. (1997) concluded that the Strepsiptera are sister group to Diptera based on parsimony analysis of 18S rDNA, 28S rDNA, and morphology. Huelsenbeck (1997) argued that this result also is attributable to long-branch attraction. We demonstrate that the analyses and arguments dismissing these results as the effects of long-branch attraction are fundamentally flawed. The criteria employed by these authors were applied arbitrarily by them to the groups that they did not want, and yet using those same criteria, there is more reason to exclude other taxa besides Polypodium and there is more reason to disbelieve monophyly of Diptera than monophyly of Strepsiptera with Diptera. Moreover, it is asserted, long-branch attraction cannot explain the presence of nematocysts in Myxozoa and halteres in Strepsiptera. For these reasons, and in light of the demonstration that long branches cannot attract each other in their mutual absence, we conclude that the monophyly of Myxozoa + Polypodium and Strepsiptera + Diptera is not due to long-branch attraction. We suggest that maximum likelihood methods are extremely sensitive to taxon and character sampling and that these data sets are demonstrative of the long-branch repulsion problem.     

Phylogenetic position of the Strepsiptera: Review of molecular and morphological evidence

Molecular evidence of the monophyly of the Halteria (Strepsiptera + Diptera) is reviewed. The majority of morphological characters, which have classically been used to establish a Strepsiptera + Coleoptera sister group, are rejected, because they are based on erroneous interpretations of strepsipteran morphology. The scorings of 31 morphological characters, which directly relate to the phylogenetic position of Strepsiptera, are provided, and their distribution and optimization on the molecular + morphological tree is discussed. Of these, 13 characters specifically support the placement of Strepsiptera within the Mecopterida; seven of which are based on the optimization of inapplicable or missing data, and six of which are based on states that can be scored for Strepsiptera. Only a single character (posteromotorism) suggests a sister group relationship with the Coleoptera. The morphological and molecular data are largely congruent, and suggest that the Strepsiptera are sister group to the Diptera.     

Putative secondary structures of unusually long strepsipteran SSU rRNAs and its phylogenetic implications.

We constructed the putative secondary structures of the small subunit rRNAs (SSU rRNA) from three strepsipteran insects. The primary sequences of the strepsipteran SSU rRNAs are unusually long due to unique and long insertions. In spite of these insertions, the basic shapes of their secondary structures are well maintained as shown in those of other eukaryotes, because these insertions appear mainly in the variable regions. The secondary structures for the V1, V3, V5, V8, and V9 regions are well conserved, even though the primary structures of V1, V5, and V8 regions are quite variable. However, the predicted secondary structures for the V2, V4, and V7 regions are quite different from those of other insects. In the V4 and V7 regions, helices specific to the Strepsiptera exist. These helices have not been reported in other organisms so far. Similarly, four eukaryotic specific helices (E8-1, E10-2, E23-4 and E45-1) not reported in insects exist in the V2, V4, and V8 regions. These helices are formed by the inserted sequences. The secondary structures of the expanded segments of the strepsipteran SSU rRNA were applied to infer the phylogenetic position of Strepsiptera, one of the most enigmatic problems in insect phylogeny. Only the secondary structure of the V7 region showed the weak Strepsiptera/Diptera sister-group relationship.     

Is the Felsenstein zone a fly trap?

Although long-branch attraction, the incorrect grouping of long lineages in a phylogeny because of systematic error, has been identified as a potential source of error in phylogenetic analysis for almost two decades, no empirical examples of the phenomenon exist. Here, I outline several criteria for identifying long-branch attraction and apply these criteria to 18S ribosomal DNA (rDNA) sequence data for 13 insects. Parsimony and minimum evolution with p distances group the two longest branches together (those leading to Strepsiptera and Diptera). Simulation studies show that the long branches are long enough to attract. When a tree is assumed in which Strepsiptera and Diptera are separated and many data sets are simulated for that tree (using the parameter estimates for that tree for the original data), parsimony analysis of the simulated data consistently groups Strepsiptera and Diptera. Analyses of the 18S rDNA sequences using methods that are less sensitive to the problem of long-branch attraction estimate trees in which the long branches are separate.     

Evolution and phylogenetic utility of CAD (rudimentary) among Mesozoic-aged Eremoneuran Diptera (Insecta)

We sequenced nearly the entire carbomoylphosphate synthase (CPS) domain of CAD, or rudimentary, (ca. 4 kb) from 29 species of flies representing all major clades within Eremoneura, or higher flies, and several orthorrhaphous brachyceran outgroups. We compared these sequences with orthologs from Anopheles gambiae and Drosophila melanogaster to assess structure, compositional bias, and phylogenetic utility. CAD is large (6.6+ kb), complex (comprised of three major and myriad minor functional domains) and relatively free of introns, extreme nucleotide bias (except third codon positions), and large hypervariable regions. The CPS domain possesses moderate levels of nonsynonymous divergence among taxa of intermediate evolutionary age and conveys considerable phylogenetic signal. Phylogenetic analysis of CPS sequences under varying methods and assumptions resulted in well-resolved, strongly supported trees concordant with many traditional ideas about higher dipteran phylogeny and with prior inferences from 28S rDNA. The most robustly supported major eremoneuran clades were Cyclorrhapha, Platypezoidea, Eumuscomorpha, Empidoidea, Atelestidae, Empidoidea exclusive of Atelestidae, Hybotidae s.l., Microphoridae+Dolichopodidae, and Empididae s. str. Because CAD is ubiquitous, apparently single copy (at least within holometabolous insects), readily obtained from several insect orders using primers described herein, and exhibits considerable phylogenetic utility, it should have wide applicability in insect molecular systematics.     

The structure of the USP/RXR of <Emphasis Type="Italic">Xenos pecki</Emphasis> indicates that Strepsiptera are not closely related to Diptera

The receptor for the insect molting hormone, ecdysone, is a heterodimer consisting of the Ecdysone Receptor and Ultraspiracle (USP) proteins. The ligand binding domain sequences of arthropod USPs divide into two distinct groups. One group consists of sequences from members of the holometabolous Lepidoptera and Diptera, while the other arthropod sequences group with vertebrate retinoid-X-receptors (RXRs). We therefore wondered whether USP/RXR structure could be used to clarify the contentious phylogenetic position of the order Strepsiptera, which has proposed affinities with either Diptera or Coleoptera. We have cloned and sequenced the USP/RXR from the strepsipteran Xenos pecki. Phylogenetic analyses are not consistent with a close affinity between Strepsiptera and Diptera.Electronic Supplementary Material Supplementary material is available for this article at Edited by D. Tautz     

Insufficient resolving power of mitogenome data in deciphering deep phylogeny of Holometabola

The mitochondrial genome (mitogenome) is one of the most widely used markers for phylogenetic analysis. Compared with whole-genome data, mitogenome data are less expensive to obtain and easier to manipulate. However, compositional bias and accelerated evolutionary rate reduce the effectiveness of the mitogenome in determining insect phylogeny. This study shows that mitogenome data are not suitable to reconstruct deep holometabolan evolution, even with a most comprehensive data coding scheme and the more realistic CAT model. For the deep levels of divergence within Holometabola, protein-coding genes only retain weak phylogenetic signals, leading to peculiar interordinal relationships. Consensus relationships in the Holometabola phylogeny, such as the monophyly of Holometabola, the most basal position of Hymenoptera, and the sister group relationship between the Strepsiptera and Coleoptera were rarely resolved in our analyses. The relationships of the holometabolan groups as inferred by mitogenomes are highly vulnerable to gene types, data coding regimes, model choice, and optimality criteria, and no consistent alternative hypothesis of Holometabola's relationships is supported. Thus, we suggest that the slowly evolving nuclear genes or genome-scale approaches may be better options for resolving deep-level phylogeny of Holometabola.     

Phylogenetic analyses of Cornales based on 26S rRNA and combined 26S rDNA-MATK-RBCL sequence data

Nuclear 26S rDNA sequences were used to corroborate and test previously published matK-rbcL-based hypotheses of phylogenetic relationships in Cornales. Sequences were generated for 53 taxa including Alangium, Camptotheca, Cornus, Curtisia, Davidia, Diplopanax, Mastixia, Nyssa, and four families: Grubbiaceae, Hydrangeaceae, Hydrostachyaceae, and Loasaceae. Fifteen taxa from asterids were used as outgroups. The 26S rDNA sequences were initially analyzed separately and then combined with matK-rbcL sequences, using both parsimony and maximum likelihood methods. Eight strongly supported major clades were identified within Cornales by all analyses: Cornus, Alangium, nyssoids (Nyssa, Davidia, and Camptotheca), mastixioids (Mastixia and Diplopanax), Hydrangeaceae, Loasaceae, Grubbia-Curtisia, and Hydrostachys. However, relationships among the major lineages are not strongly supported in either 26S rDNA or combined 26S rDNA-matK-rbcL topologies, except for the sister relationships between Cornus and Alangium and between nyssoids and mastixioids in the tree from combined data. Discrepancies in relationships among major lineages, especially the placement of the long-branched Hydrostachys, were found between parsimony and maximum likelihood trees in all analyses. Incongruence between the 26S rDNA and matK-rbcL data sets was suggested, where Hydrangeaceae was found to be largely responsible for the incongruence. The long branch of Hydrostachys revealed in previous analyses was reduced significantly with more sampling. Maximum likelihood analysis of combined 26S rDNA-matK-rbcL sequences suggested that Hydrostachys might be sister to the remainder of Cornales, that Cornus-Alangium are sisters, that nyssoids-mastixioids are sisters, and that Hydrangeaceae-Loasaceae are sisters, consistent with previous analyses of matK-rbcL sequence data.     

Aligned 18S and insect phylogeny

The nuclear small subunit rRNA (18S) has played a dominant role in the estimation of relationships among insect orders from molecular data. In previous studies, 18S sequences have been aligned by unadjusted automated approaches (computer alignments that are not manually readjusted), most recently with direct optimization (simultaneous alignment and tree building using a program called "POY"). Parsimony has been the principal optimality criterion. Given the problems associated with the alignment of rRNA, and the recent availability of the doublet model for the analysis of covarying sites using Bayesian MCMC analysis, a different approach is called for in the analysis of these data. In this paper, nucleotide sequence data from the 18S small subunit rRNA gene of insects are aligned manually with reference to secondary structure, and analyzed under Bayesian phylogenetic methods with both GTR+I+G and doublet models in MrBayes. A credible phylogeny of Insecta is recovered that is independent of the morphological data and (unlike many other analyses of 18S in insects) not contradictory to traditional ideas of insect ordinal relationships based on morphology. Hexapoda, including Collembola, are monophyletic. Paraneoptera are the sister taxon to a monophyletic Holometabola but weakly supported. Ephemeroptera are supported as the sister taxon of Neoptera, and this result is interpreted with respect to the evolution of direct sperm transfer and the evolution of flight. Many other relationships are well-supported but several taxa remain problematic, e.g., there is virtually no support for relationships among orthopteroid orders. A website is made available that provides aligned 18S data in formats that include structural symbols and Nexus formats.     

Molecular phylogeny of Scaphopoda (Mollusca) inferred from 18S rDNA sequences: support for a Scaphopoda–Cephalopoda clade

The phylogenetic relationships of the Scaphopoda, one of the 'lesser' molluscan classes, with the other conchiferan taxa are far from clear. They appear either as the sister-group to the Bivalvia (Diasoma concept) or to a Gastropoda–Cephalopoda clade or to the Cephalopoda alone (helcionellid concept). We compiled a 18S rDNA sequence dataset of 48 molluscan species containing 17 scaphopods to test these hypotheses and to address questions regarding high-level relationships with the Scaphopoda. Both parsimony and maximum likelihood trees show low branch support at the base of the Conchifera, except for the robust clade uniting Scaphopoda and Cephalopoda. This result is corroborated by spectral analysis and likelihood mapping. We also tested alternative topologies which scored significantly worse both in tree length and in likelihood. The 18S rDNA data thus reject the Diasoma in favour of a Scaphopoda–Cephalopoda clade as proposed in the helcionellid concept. When plotted on the molecular tree, the pivotal morphological characters associated with the burrowing life style of the Bivalvia and Scaphopoda, i.e. mantle/shell enclosure of the body and the burrowing foot with true pedal ganglia, appear convergent in these groups. In contrast, the prominent and tilted dorsoventral body axes, multiple cephalic tentacles and a ring-shaped muscle attachment on the shell are potential synapomorphies of Scaphopoda and Cephalopoda. Most of the higher taxa within the Scaphopoda are supported by the molecular data. However, there is no support for the families Dentaliidae and Gadilidae. The basal position of the Fustiariidae within the Dentaliida is confirmed.     

Weighting indels as phylogenetic markers of 18S rDNA sequences in Diptera and Strepsiptera

Opinions split when it comes to the significance and thus the weighting of indel characters as phylogenetic markers. This paper attempts to test the phylogenetic information content of indels and nucleotide substitutions by proposing an a priori weighting system of non-protein-coding genes. Theoretically, the system rests on a weighting scheme which is based on a falsificationist approach to cladistic inference. It provides insertions, deletions and nucleotide substitutions weights according to their specific number of identical classes of potential falsifiers, resulting in the following system: nucleotide substitutions weight = 3, deletions of n nucleotides weight = (2ⁿ–1), and insertions of n nucleotides weight = (5ⁿ–1). This weighting system and the utility of indels as phylogenetic markers are tested against a suitable data set of 18S rDNA sequences of Diptera and Strepsiptera taxa together with other Metazoa species. The indels support the same clades as the nucleotide substitution data, and the application of the weighting system increases the corresponding consistency indices of the differentially weighted character types. As a consequence, applying the weighting system seems to be reasonable, and indels appear to be good phylogenetic markers.     

Phylogenetic inference regarding Parergodrilidae and Hrabeiella periglandulata ('Polychaeta', Annelida) based on 18S rDNA, 28S rDNA and COI sequences

Parergodrilidae and Hrabeiella periglandulata are Annelida showing different combinations of clitellate-like and aclitellate characters. Similarities between both of these taxa and Clitellata have widely been regarded as the result of convergent evolution due to similar selection pressures. The position of the three taxa in the phylogenetic system of Annelida is still in debate. However, in analyses based on 18S rDNA sequences a close relationship of Parergodrilidae with Orbiniidae and Questidae was suggested. To infer their phylogeny the sequences of the 28S rDNA and of the cytochrome oxidase I (COI) gene of Stygocapitella subterranea , Parergodrilus heideri and H. periglandulata were determined. The data were extended by sequences of various species including species from Clitellata and Orbiniidae. Prior to tree reconstruction the dataset was analysed in detail for phylogenetic content by applying a sliding window analysis, a likelihood mapping and Modeltest V.3.04. Subsequently, generalized parsimony and maximum likelihood methods were employed. Clade robustness was estimated by bootstrapping. In addition, combined analyses of the sequences of 18S rDNA and 28S rDNA as well as of 18S rDNA, 28S rDNA and COI were performed. The combination of the data of the two structure genes and a mitochondrial gene improved the resolution obtained with the single datasets slightly. These analyses support a close relationship of Parergodrilidae and Orbiniidae but cannot resolve the position of H. periglandulata . In every analysis Clitellata cluster within 'Polychaeta', confirming previous investigations.     

Phylogeny of the Platyhelminthes and the evolution of parasitism

Robust phylogenies provide the basis for interpreting biological variation in the light of evolution. Homologous features provide phylogenetically informative characters whereas homoplasious characters provide phylogenetic noise. Both provide evolutionary signal. We have constructed molecular and morphologically based phylogenies of the phylum Platyhelminthes using a recently revised morphological character matrix and complete 18S and two partial 28S rRNA gene sequences in order to evaluate the emergence and subsequent divergence of parasitic forms. In total we examine 65 morphological characters, 97 18S rDNA, 41 Dl domain 28S rDNA, and 49 D3-D6 domain 28S rDNA sequences. For the molecular data there were 748, 132 and 249 phylogenetically informative sites for the 18S, Dl and D3-D6 28S rDNA data sets respectively. Morphological and molecular phylogenetic solutions are incongruent but not incompatible, and using the principles of conditional combination (18S rDNA + morphology passing Templeton's test) they demonstrate: a single and relatively early origin for the parasitic Neodermata (including the cestodes, trematodes and monogeneans); sister-group status between the cestodes and monogeneans, and between these taxa and the trematodes (digeneans and aspidogastreans). The sister-group to the Neodermata is likely to be a large clade of neoophoran turbellarians, based on combined evidence, or a clade consisting of the Fecampiid + Urastomid turbellarians, based on morphological evidence alone. The combined evidence solution for the phylogeny of fiatworms based on 18S rDNA and morphology is used to interpret morphological and life-history data and to support a model for the evolution and radiation of neodermatan parasites in the group.     

Higher level phylogeny and the first divergence time estimation of Heteroptera (Insecta: Hemiptera) based on multiple genes

Heteroptera, or true bugs, are the largest, morphologically diverse and economically important group of insects with incomplete metamorphosis. However, the phylogenetic relationships within Heteroptera are still in dispute and most of the previous studies were based on morphological characters or with single gene (partial or whole 18S rDNA). Besides, so far, divergence time estimates for Heteroptera totally rely on the fossil record, while no studies have been performed on molecular divergence rates. Here, for the first time, we used maximum parsimony (MP), maximum likelihood (ML) and Bayesian inference (BI) with multiple genes (18S rDNA, 28S rDNA, 16S rDNA and COI) to estimate phylogenetic relationships among the infraorders, and meanwhile, the Penalized Likelihood (r8s) and Bayesian (BEAST) molecular dating methods were employed to estimate divergence time of higher taxa of this suborder. Major results of the present study included: Nepomorpha was placed as the most basal clade in all six trees (MP trees, ML trees and Bayesian trees of nuclear gene data and four-gene combined data, respectively) with full support values. The sister-group relationship of Cimicomorpha and Pentatomomorpha was also strongly supported. Nepomorpha originated in early Triassic and the other six infraorders originated in a very short period of time in middle Triassic. Cimicomorpha and Pentatomomorpha underwent a radiation at family level in Cretaceous, paralleling the proliferation of the flowering plants. Our results indicated that the higher-group radiations within hemimetabolous Heteroptera were simultaneously with those of holometabolous Coleoptera and Diptera which took place in the Triassic. While the aquatic habitat was colonized by Nepomorpha already in the Triassic, the Gerromorpha independently adapted to the semi-aquatic habitat in the Early Jurassic.