Phylogenetic analysis of arthropods using two nuclear protein-encoding genes supports a crustacean + hexapod clade

Recent phylogenetic analyses using molecular data suggest that hexapods are more closely related to crustaceans than to myriapods, a result that conflicts with long-held morphology-based hypotheses. Here we contribute additional information to this debate by conducting phylogenetic analyses on two nuclear protein-encoding genes, elongation factor-1 alpha (EF-1 alpha) and the largest subunit of RNA polymerase II (Pol II), from an extensive sample of arthropod taxa. Results were obtained from two data sets. One data set comprised 1092 nucleotides (364 amino acids) of EF-1 alpha and 372 nucleotides (124 amino acids) of Pol II from 30 arthropods and three lobopods. The other data set contained the same EF-1 alpha fragment and an expanded 1038-nucleotide (346-amino-acid) sample of Pol II from 17 arthropod taxa. Results from maximum-parsimony and maximum-likelihood analyses strongly supported the existence of a Crustacea + Hexapoda clade (Pancrustacea) over a Myriapoda + Hexapoda clade (Atelocerata). The apparent incompatibility between the molecule-based Pancrustacea hypothesis and morphology-based Atelocerata hypothesis is discussed.     

A phylogenetic analysis of Myriapoda (Arthropoda) using two nuclear protein-encoding genes

Nucleotide and inferred amino acid sequences from two nuclear protein-encoding genes, elongation factor-aα and RNA polymerase II, were obtained from 34 myriapods and 14 other arthropods to determine phylogenetic relationships among and within the myriapod classes. Phylogenetic analyses using maximum parsimony and maximum likelihood methods recovered all three represented myriapod classes (Chilopoda, Diplopoda, Symphyla) and all multiply sampled chilopod and diplopod orders, often with high node support. In contrast, relationships between classes and between orders were recovered less consistently and node support was typically lower. The temporal structure of phylogenetic diversification in Myriapoda may explain this apparent pattern of the phylogenetic recovery.     

Molecular phylogeny of Zygomycota based on EF-1alpha and RPB1 sequences: limitations and utility of alternative markers to rDNA

Earlier molecular phylogenetic analyses based on nuclear small subunit ribosomal DNA (nSSU rDNA) suggest that the Zygomycota are polyphyletic within the Chytridiomycota. However, these analyses failed to resolve almost all interordinal relationships among basal fungi (Chytridiomycota and Zygomycota), due to lack of sufficient characters within the nSSU rDNA. To further elucidate the higher-level phylogeny of Zygomycota, we have sequenced partial RPB1 (DNA dependent RNA polymerase II largest subunit) and EF-1alpha (translation elongation factor 1 alpha) genes from 10 and 3 zygomycete fungi, respectively. Independent molecular phylogenetic analyses were performed based on each sequence by distance and maximum likelihood methods. Although deep phylogenetic relationships among basal fungi still remain poorly resolved using either gene, the RPB1-based phylogeny identified a novel monophyletic clade consisting of the Dimargaritales, Harpellales, and Kickxellales. This result suggests that regularly formed septa (cross walls that divide hyphae into segments) with a lenticular cavity are plesiomorphic for this clade, and indicates the importance of septal pore ultrastructure in zygomycete phylogeny. In addition, a peculiar mucoralean genus Mortierella, which was considered to be distantly related to the other Mucorales based on previous nSSU rDNA analyses, was resolved as the basal most divergence within the Mucorales, consistent with traditional phenotypic-based taxonomy. Although the taxa included in our analysis are restricted, the monophyly of each order suggested by nSSU rDNA phylogeny is supported by the present RPB1-based analysis. These results support the potential use of RPB1 as an alternative marker for fungal phylogenetic studies. Conversely, the overall fungal phylogeny based on EF-1alpha sequence is poorly resolved. A comparison of numbers of observed substitutions versus inferred substitutions within EF-1alpha indicates that this gene is much more saturated than RPB1. This result suggests that the EF-1alpha gene is unsuitable for resolving higher-level phylogenetic relationships within the Fungi.     

Molecular Phylogeny of Arthropods and the Significance of the Cambrian "Explosion" for Molecular Systematics

SYNOPSIS. Accurate phylogenetic reconstruction requires charactersystems that have evolved fast enough to have kept pace withcladogenesis but slowly enough to have conveyed the resultingphylogenetic signal to the present. Because stratigraphic evidencesuggests that basal arthropod lineages arose rapidly duringan ancient (Cambrian) phylogenetic radiation, the discoveryof molecular sequences capable of resolving arthropod phylogenymay be a significant challenge for molecular systematists. Thischallenge is exemplified by our attempt to resolve arthropodphylogeny using the amino acid sequence of elongation factor-1(EF-1). Our fossil-based assessment of evolutionary rates indicatesthat EF-la should be capable of resolving Cambrian-age divergences.However, phylogenetic analysis using EF-1 fails to establishrelationships among most higher-level groups, although it doesrecover more recently derived clades. Here we propose two modelsto explain this incongruity. The Rapid Radiation Model maintainsthat fossil-based estimates of arthropod diversification areessentially accurate and that diversification occurred so rapidlyduring the Cambrian that few phylogenetically significant changesoccurred in the slowly evolving EF-1 sequence. The EnhancedPreservation Model maintains that fossil-based estimates ofCambrian-age divergences reflect enhanced preservation of pre-existinglineages and that arthropod diversification occurred beforethe Cambrian. This model attributes lack of resolution to degradationof phylogenetic signal within EF-1 by subsequent evolution.Current evidence is more consistent with the Enhanced PreservationModel, which implies that fossil-based methods can be very misleadingwhen attempting to gauge the phylogenetic information contentof molecular sequences for Cambrian- and Precambrian-age divergences.     

Phylogenetic utility and evidence for multiple copies of elongation factor-1alpha in the spider genus Habronattus (Araneae: Salticidae)

In the continuing quest for informative genes for use in molecular systematics, the protein-coding gene Elongation factor-1alpha (EF-1alpha) has rapidly become one of the most prevalent "single-copy" nuclear genes utilized, particularly in arthropods. This paper explores the molecular evolutionary dynamics and phylogenetic utility of EF-1alpha in the salticid spider genus Habronattus. As has been reported for other arthropod lineages, our studies indicate that multiple (two) copies of EF-1alpha exist in Habronattus. These copies differ in intron structure and thus in size, making it possible to easily separate PCR amplification products. We present data for an intronless EF-1alpha copy for three Habronattus species. The presence of nonsense mutations and generally elevated rates of amino acid change suggest that this copy is evolving under relaxed functional constraints in Habronattus. A larger taxon sample (50 species plus outgroups) is presented for an EF-1alpha copy that includes both intron and exon regions. Characteristics of both regions suggest that this is a functional, orthologous copy in the species sampled. Maximum-likelihood relative-rate comparisons show that exon third codon sites are evolving more than 100 times as fast as second codon sites in these sequences and that intron sites are evolving about twice as fast as exon third sites. In combination, the EF-1alpha data provide robust, species-level phylogenetic signal that is largely congruent with morphologically well supported areas of Habronattus phylogeny. The recovery of some novel clades, and the unexpected fragmentation of others, suggests areas requiring further phylogenetic attention.     

An evaluation of elongation factor 1 alpha as a phylogenetic marker for eukaryotes

Elongation factor 1 alpha (EF-1 alpha) is a highly conserved ubiquitous protein involved in translation that has been suggested to have desirable properties for phylogenetic inference. To examine the utility of EF-1 alpha as a phylogenetic marker for eukaryotes, we studied three properties of EF-1 alpha trees: congruency with other phyogenetic markers, the impact of species sampling, and the degree of substitutional saturation occurring between taxa. Our analyses indicate that the EF-1 alpha tree is congruent with some other molecular phylogenies in identifying both the deepest branches and some recent relationships in the eukaryotic line of descent. However, the topology of the intermediate portion of the EF-1 alpha tree, occupied by most of the protist lineages, differs for different phylogenetic methods, and bootstrap values for branches are low. Most problematic in this region is the failure of all phylogenetic methods to resolve the monophyly of two higher-order protistan taxa, the Ciliophora and the Alveolata. JACKMONO analyses indicated that the impact of species sampling on bootstrap support for most internal nodes of the eukaryotic EF-1 alpha tree is extreme. Furthermore, a comparison of observed versus inferred numbers of substitutions indicates that multiple overlapping substitutions have occurred, especially on the branch separating the Eukaryota from the Archaebacteria, suggesting that the rooting of the eukaryotic tree on the diplomonad lineage should be treated with caution. Overall, these results suggest that the phylogenies obtained from EF-1 alpha are congruent with other molecular phylogenies in recovering the monophyly of groups such as the Metazoa, Fungi, Magnoliophyta, and Euglenozoa. However, the interrelationships between these and other protist lineages are not well resolved. This lack of resolution may result from the combined effects of poor taxonomic sampling, relatively few informative positions, large numbers of overlapping substitutions that obscure phylogenetic signal, and lineage-specific rate increases in the EF-1 alpha data set. It is also consistent with the nearly simultaneous diversification of major eukaryotic lineages implied by the "big-bang" hypothesis of eukaryote evolution.     

Elongation factor-1 alpha occurs as two copies in bees: implications for phylogenetic analysis of EF-1 alpha sequences in insects

We report the complete sequence of a paralogous copy of elongation factor-1 alpha (EF-1 alpha) in the honeybee, Apis mellifera (Hymenoptera: Apidae). This copy differs from a previously described copy in the positions of five introns and in 25% of the nucleotide sites in the coding regions. The existence of two paralogous copies of EF-1 alpha in Drosophila and Apis suggests that two copies of EF-1 alpha may be widespread in the holometabolous insect orders. To distinguish between a single, ancient gene duplication and parallel, independent fly and bee gene duplications, we performed a phylogenetic analysis of hexapod EF-1 alpha sequences. Unweighted parsimony analysis of nucleotide sequences suggests an ancient gene duplication event, whereas weighted parsimony analysis of nucleotides and unweighted parsimony analysis of amino acids suggests the contrary: that EF-1 alpha underwent parallel gene duplications in the Diptera and the Hymenoptera. The hypothesis of parallel gene duplication is supported both by congruence among nucleotide and amino acid data sets and by topology-dependent permutation tail probability (T-PTP) tests. The resulting tree topologies are also congruent with current views on the relationships among the holometabolous orders included in this study (Diptera, Hymenoptera, and Lepidoptera). More sequences, from diverse orders of holometabolous insects, will be needed to more accurately assess the historical patterns of gene duplication in EF-1 alpha.      

A new nuclear gene for insect phylogenetics: dopa decarboxylase is informative of relationships within Heliothinae (Lepidoptera: Noctuidae)

The lack of a readily accessible roster of nuclear genes informative at various taxonomic levels is a bottleneck for molecular systematics. In this report, we describe the first phylogenetic application of the sequence that encodes the enzyme dopa decarboxylase (DDC). For 14 test species within the noctuid moth subfamily Heliothinae that represent the previously best-supported groupings, a 690-bp fragment of DDC resolved relationships that are largely concordant with prior evidence from elongation factor-1 alpha (EF-1 alpha), morphology, and allozymes. Although both synonymous and nonsynonymous changes occur in DDC substantially more rapidly than they do in EF-1 alpha, DDC divergences within Heliothinae are below saturation at all codon positions. Analysis of DDC and EF-1 alpha in combination resulted in increased bootstrap support for several groupings. As a first estimate of previously unresolved relationships, DDC sequences were analyzed from 16 additional heliothines, for a total of 30 heliothine species plus outgroups. Previous relationships based on DDC were generally stable with increased taxon sampling, although a two- to eightfold downweighting of codon position 3 was required for complete concordance with the 14-species result. The weighted strict consensus trees were largely resolved and were congruent with most although not all previous hypotheses based on either morphology or EF-1 alpha. The proposed phylogeny suggests that the major agricultural pest heliothines belong to a single clade, characterized by polyphagy and associated life history traits, within this largely host-specific moth subfamily. DDC holds much promise for phylogenetic analysis of Tertiary-age animal groups.     

Molecular phylogenetic evidence refuting the hypothesis of Batoidea (rays and skates) as derived sharks

Early morphological studies regarding the evolutionary history of elasmobranchs suggested sharks and batoids (skates and rays) were respectively monophyletic. More modern morphological cladistic studies, however, have tended to suggest that batoids are derived sharks, closely related to sawsharks and angelsharks, a phylogenetic arrangement known as the Hypnosqualea hypothesis. Very few molecular studies addressing interordinal relationships of elasmobranchs have been published; the few that do exist, are very limited in terms of both taxon representation and/or aligned sequence positions, and are insufficient to answer the question of whether batoids are derived sharks. The purpose of this study was to address this issue with more complete taxon representation, concomitant with a reasonable number of aligned sequence positions. The data set included a 2.4-kb segment of the mitochondrial 12S rRNA-tRNA valine-16S rRNA locus, and in terms of taxa, representatives of two orders of Batoidea, at least one representative of all orders of sharks, and as an outgroup, the widely recognized sister group to elasmobranchs-Holocephali. The results provide the first convincing molecular evidence for shark monophyly and the rejection of the Hypnosqualea hypothesis. Our phylogenetic placement of batoids as a basal elasmobranch lineage means that much of the current thinking regarding the evolution of morphological and life history characteristics in elasmobranchs needs to be re-evaluated.     

Phylogenetic Relationships of the Order Insectivora Based on Complete 12S rRNA Sequences from Mitochondria

Despite numerous studies, there is no single accepted hypothesis of eutherian ordinal relationships. Among the least understood mammalian orders is the group Insectivora. Currently, molecular and morphological data are in conflict over the possible monophyly of the living members of Insectivora (lipotyphlans), and the relationships within the group remain largely unresolved. One of the primary criticisms concerning molecular analyses is the noticeable lack of data from a well-sampled group of lipotyphlan insectivores. The mitochondrial 12S rRNA gene has been widely used to resolve interordinal and intraordinal relationships across a variety of mammalian taxa. This study compares 118 complete mammalian 12S rRNA sequences, representing all of the 18 eutherian orders and 3 metatherian orders, and includes as well taxa from each of the six families of lipotyphlan insectivores. Insectivoran lineages are thought to have diverged concurrently with the general radiation of mammalian orders. This study suggests that the 12S rRNA sequences lack the ability to resolve relationships extending into this period. This would explain the polyphyly, unusual affinities, and low support derived in this and other studies employing 12S rRNA sequences to diagnose relationships among eutherian orders. The results of these analyses suggest that even extensive taxon sampling is insufficient to provide well supported groups among eutherian orders. Additional genes and species sampling will be necessary to elucidate whether the Insectivora form a monophyletic group.     

More taxa or more characters revisited: combining data from nuclear protein-encoding genes for phylogenetic analyses of Noctuoidea (Insecta: Lepidoptera)

A central question concerning data collection strategy for molecular phylogenies has been, is it better to increase the number of characters or the number of taxa sampled to improve the robustness of a phylogeny estimate? A recent simulation study concluded that increasing the number of taxa sampled is preferable to increasing the number of nucleotide characters, if taxa are chosen specifically to break up long branches. We explore this hypothesis by using empirical data from noctuoid moths, one of the largest superfamilies of insects. Separate studies of two nuclear genes, elongation factor-1 alpha (EF-1 alpha) and dopa decarboxylase (DDC), have yielded similar gene trees and high concordance with morphological groupings for 49 exemplar species. However, support levels were quite low for nodes deeper than the subfamily level. We tested the effects on phylogenetic signal of (1) increasing the taxon sampling by nearly 60%, to 77 species, and (2) combining data from the two genes in a single analysis. Surprisingly, the increased taxon sampling, although designed to break up long branches, generated greater disagreement between the two gene data sets and decreased support levels for deeper nodes. We appear to have inadvertently introduced new long branches, and breaking these up may require a yet larger taxon sample. Sampling additional characters (combining data) greatly increased the phylogenetic signal. To contrast the potential effect of combining data from independent genes with collection of the same total number of characters from a single gene, we simulated the latter by bootstrap augmentation of the single-gene data sets. Support levels for combined data were at least as high as those for the bootstrap-augmented data set for DDC and were much higher than those for the augmented EF-1 alpha data set. This supports the view that in obtaining additional sequence data to solve a refractory systematic problem, it is prudent to take them from an independent gene.     

The evolutionary history of Trichoptera (Insecta): A case of successful adaptation to life in freshwater

The insect order Trichoptera (caddisflies) forms the second most species‐rich monophyletic group of animals in freshwater. So far, several attempts have been made to elucidate its evolutionary history with both morphological and molecular data. However, none have attempted to analyse the time frame for its diversification. The order is divided into three suborders – Annulipalpia, Integripalpia and ‘Spicipalpia’. Historically, the most problematic taxon to place within the order is ‘Spicipalpia’, whose larvae do not build traditional cases or filtering nets like the majority of the caddisflies. They have previously been proposed to be the sister group of all other Trichoptera or more advanced within the order, with equivocal monophyly and with different interordinal placements among various studies. In order to resolve the evolutionary history of the caddisflies as well as timing their diversification, we utilized fragments of three nuclear (carbamoylphosphate synthethase, isocitrate dehydrogenase and RNA polymerase II) and one mitochondrial (cytochrome oxidase I) protein coding genes, with 16 fossil trichopteran taxa used for time calibration. The ‘spicipalpian’ families are recovered as ancestral to all other caddisflies, though paraphyletic. We recover stable relationships among most families and superfamilies, resolving many previously unrecognized phylogenetic affinities amongst extant families. The origin of Trichoptera is estimated to be around 234 Ma, i.e. Middle – Late Triassic.     

Phylogenomic analysis resolves the interordinal relationships and rapid diversification of the laurasiatherian mammals

Although great progress has been made in resolving the relationships of placental mammals, the position of several clades in Laurasiatheria remain controversial. In this study, we performed a phylogenetic analysis of 97 orthologs (46,152 bp) for 15 taxa, representing all laurasiatherian orders. Additionally, phylogenetic trees of laurasiatherian mammals with draft genome sequences were reconstructed based on 1608 exons (2,175,102 bp). Our reconstructions resolve the interordinal relationships within Laurasiatheria and corroborate the clades Scrotifera, Fereuungulata, and Cetartiodactyla. Furthermore, we tested alternative topologies within Laurasiatheria, and among alternatives for the phylogenetic position of Perissodactyla, a sister-group relationship with Cetartiodactyla receives the highest support. Thus, Pegasoferae (Perissodactyla + Carnivora + Pholidota + Chiroptera) does not appear to be a natural group. Divergence time estimates from these genes were compared with published estimates for splits within Laurasiatheria. Our estimates were similar to those of several studies and suggest that the divergences among these orders occurred within just a few million years.     

Myriapod monophyly and relationships among myriapod classes based on nearly complete 28S and 18S rDNA sequences

Myriapods play a pivotal position in the arthropod phylogenetic tree. The monophyly of Myriapoda and its internal relationships have been difficult to resolve. This study combined nearly complete 28S and 18S ribosomal RNA gene sequences (3,826 nt in total) to estimate the phylogenetic position of Myriapoda and phylogenetic relationships among four myriapod classes. Our data set consists of six new myriapod sequences and homologous sequences for 18 additional species available in GenBank. Among the six new myriapod sequences, those of the one pauropod and two symphylans are very important additions because they were such difficult taxa to classify in past molecular-phylogenetic studies. Phylogenetic trees were constructed with maximum parsimony, maximum likelihood, and Bayesian analyses. All methods yielded moderate to strong support for the monophyly of Myriapoda. Symphyla grouped strongly with Pauropoda under all analytical conditions. The KH test rejected the traditional view of Dignatha and Progoneata, and the topology obtained here, though not significantly supported, was Diplopoda versus ((Symphyla + Pauropoda) + Chilopoda).     

Phylogenetic utility of elongation factor-1 alpha in noctuoidea (Insecta: Lepidoptera): the limits of synonymous substitution

To test its phylogenetic utility, nucleotide sequence variation in a 1,240-bp fragment of the elongation factor-1 alpha (EF-1 alpha) gene was examined in 49 moth species representing the major groups of the superfamily Noctuoidea. Both parsimony and distance analyses supported the monophyly of nearly all groups for which there are clear morphological synapomorphies. Clades of subfamily rank and lower, probably mid-Tertiary and younger, were strongly supported. The third codon position contains 88% of variable sites, and approaches saturation at approximately 20% sequence divergence, possibly due to among-site rate heterogeneity and composition bias; higher divergences occur only in association with shifts in composition. Surprisingly, the few nonsynonymous changes appear no more phylogenetically reliable than synonymous changes. Signal strength for basal divergences is weak and fails to improve with character weighting; thus, dense taxon sampling is probably needed for strong inference from EF-1 alpha regarding deeper splits in Noctuoidea (probably early Tertiary). EF-1 alpha synonymous changes show promise for phylogeny reconstruction within Noctuidae and other groups of Tertiary age.      

Intraordinal phylogenetics of treeshrews (Mammalia: Scandentia) based on evidence from the mitochondrial 12S rRNA gene

Despite their traditional and continuing prominence in studies of interordinal mammalian phylogenetics, treeshrews (order Scandentia) remain relatively unstudied with respect to their intraordinal relationships. At the same time, significant morphological variation among living treeshrews has been shown to have direct relevance to higher-level interpretations of character state change as reconstructed in traditional interordinal studies, which have often included only a single species of treeshrew. Therefore, the importance of resolving relationships among treeshrews extends well beyond a better understanding of patterns of diversification within the order. A recent review highlighted several shortcomings in published studies of treeshrew phylogenetics based on morphology. Here we present the first investigation of treeshrew phylogenetics based on DNA sequences, utilizing previously published sequences from the mitochondrial 12S rRNA gene and combining them with newly generated sequence data from 15 species. Parsimony, likelihood, and Bayesian analyses all strongly support a sister relationship between Ptilocercus and the remaining species, further substantiating its recent elevation to familial status. Dendrogale is consistently recovered as the next taxon to diverge, but relationships among the remaining taxa are poorly supported by these data. We provide evidence for a relatively rapid radiation within the genera Tupaia and Urogale, but limited resolution precludes more than a cursory interpretation of biogeographic patterns.     

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.     

A highly conserved nuclear gene for low-level phylogenetics: elongation factor-1 alpha recovers morphology-based tree for heliothine moths

Molecular systematists need increased access to nuclear genes. Highly conserved, low copy number protein-encoding nuclear genes have attractive features for phylogenetic inference but have heretofore been applied mostly to very ancient divergences. By virtue of their synonymous substitutions, such genes should contain a wealth of information about lower-level taxonomic relationships as well, with the advantage that amino acid conservatism makes both alignment and primer definition straightforward. We tested this postulate for the elongation factor-1 alpha (EF-1 alpha) gene in the noctuid moth subfamily Heliothinae, which has probably diversified since the middle Tertiary. We sequenced 1,240 bp in 18 taxa representing heliothine groupings strongly supported by previous morphological and allozyme studies. The single most parsimonious gene tree and the neighbor-joining tree for all nucleotides show almost complete concordance with the morphological tree. Homoplasy and pairwise divergence levels are low, transition/transversion ratios are high, and phylogenetic information is spread evenly across gene regions. The EF-1 alpha gene and presumably other highly conserved genes hold much promise for phylogenetics of Tertiary age eukaryote groups.      

Molecular phylogeny of the Eichhorni group of Delias Hübner, 1819 (Lepidoptera: Pieridae)

The eichhorni group lies within the genus Delias (Lepidoptera: Pieridae) which has markedly diversified aposematic wing markings. The phylogenetic relationships among all species of the eichhorni group, representatives of each of the other 21 species groups of Delias butterflies, and some related genera were analyzed based on nucleotide sequences of the mitochondrial NADH dehydrogenase subunit 5 gene. A supplemental study using the nuclear elongation factor-1alpha (EF-1alpha) gene was also carried out. The results are compared with those of morphological studies. Our results confirm the monophyly of the eichhorni group and suggest the monophyly of the genus Delias. They also indicate phylogenetic intragroup relationships, particularly the division of the eichhorni complex into groups I and II. Moreover, they also indicate that the initial diversification of the eichhorni group involved separation of the D. catisa + D. toxopei clade, followed by the divergence of other species including the eichhorni complex. Based on these findings, it is supposed that this group first appeared close to or within the western mountain range of New Guinea Island (135 degrees 30(')-140 degrees E) where D. catisa, D. toxopei, and representatives of other species cohabit.     

A phylogeny of Vetigastropoda and other “archaeogastropods”: re-organizing old gastropod clades

Abstract. The phylogenetic relationships among the “archaeogastropod” clades Patellogastropoda, Vetigastropoda, Neritimorpha, and Neomphalina are uncertain; the phylogenetic placement of these clades varies across different analyses, and particularly among those using morphological characteristics and those relying on molecular data. This study explores the relationships among these groups using a combined analysis with seven molecular loci (18S rRNA, 28S rRNA, histone H3, 16S rRNA, cytochrome c oxidase subunit I [COI], myosin heavy-chain type II, and elongation factor-1α [EF-1α]) sequenced for 31 ingroup taxa and eight outgroup taxa. The deep evolutionary splits among these groups have made resolution of stable relationships difficult, and so EF-1α and myosin are used in an attempt to re-examine these ancient radiation events. Three phylogenetic analyses were performed utilizing all seven genes: a single-step direct optimization analysis using parsimony, and two-step approaches using parsimony and maximum likelihood. A single-step direct optimization parsimony analysis was also performed using only five molecular loci (18S rRNA, 28S rRNA, histone H3, 16S rRNA, and COI) in order to determine the utility of EF-1α and myosin in resolving deep relationships. In the likelihood and POY optimal phylogenetic analyses, Gastropoda, Caenogastropoda, Neritimorpha, Neomphalina, and Patellogastropoda were monophyletic. Additionally, Neomphalina and Pleurotomariidae fell outside the remaining vetigastropods, indicating the need for further investigation into the relationship of these groups with other gastropods.