Arachnid relationships based on mitochondrial genomes: asymmetric nucleotide and amino acid bias affects phylogenetic analyses

Phylogenetic analyses based on mitochondrial DNA have yielded widely differing relationships among members of the arthropod lineage Arachnida, depending on the nucleotide coding schemes and models of evolution used. We enhanced taxonomic coverage within the Arachnida greatly by sequencing seven new arachnid mitochondrial genomes from five orders. We then used all 13 mitochondrial protein-coding genes from these genomes to evaluate patterns of nucleotide and amino acid biases. Our data show that two of the six orders of arachnids (spiders and scorpions) have experienced shifts in both nucleotide and amino acid usage in all their protein-coding genes, and that these biases mislead phylogeny reconstruction. These biases are most striking for the hydrophobic amino acids isoleucine and valine, which appear to have evolved asymmetrical exchanges in response to shifts in nucleotide composition. To improve phylogenetic accuracy based on amino acid differences, we tested two recoding methods: (1) removing all isoleucine and valine sites and (2) recoding amino acids based on their physiochemical properties. We find that these methods yield phylogenetic trees that are consistent in their support of ancient intraordinal divergences within the major arachnid lineages. Further refinement of amino acid recoding methods may help us better delineate interordinal relationships among these diverse organisms.     

Comparative morphology,functions, and homologies of the coxal glands in oribatid mites (Arachnida: Acari)

1. Forty-eight species of oribatids in 37 families representing most of the superfamilies were collected from various environments (littoral, salt marsh, litter, sod, and freshwater) and sectioned. 2. The coxal gland is composed of a sacculus and a labyrinth in all stages of all oribatid species. Muscles, originating on the body wall, insert at several points on the thin-walled sacculus which opens into the labyrinth. The labyrinth has an internal, chitinous supporting skeleton. The type A labyrinth has 3–180° bends, producing four parallel regions, and occurs in all inferior oribatids. The type B labyrinth has 1–180° bend, producing two parallel regions, and occurs in all superior oribatids. The coxal gland duct and the lateral gland duct join, penetrate the body wall, and empty into the posterior end of the podocephalic canal. All oribatids have lateral accessory glands, but only inferior oribatids have rostral and medial glands. Three ductless coxendral bodies are always present. 3. The labyrinth length in oribatids is correlated with body size and the environment of the species. Oribatids from sod, leaflitter, or moss show a simple correlation of labyrinth length (X) to total body length (Y) where Y = 4.64X. Freshwater species have a labyrinth length greater than that of comparably sized terrestrial species and salt water (littoral) species have a labyrinth length less than that of comparably sized terrestrial species. There is a greater reduction in labyrinth length in species restricted to salt marshes than in species not restricted to salt marshes. 4. The probable function of oribatid coxal glands is osmoregulation. Hemolymph filtration would occur across the sacculus by positive hemolymph pressure and contraction of the sacculus muscles. Resorption of ions would occur in the labyrinth, which is noncollapsible due to the internal skeleton. The hypothesis is that in freshwater species the rate of filtration is high and resorption of ions would have to be very efficient, therefore they have an elongated labyrinth; but in salt water species water loss must be minimized and preservation of ions would be a disadvantage, therefore they have a shortened labyrinth. Excre ion may also be a function of the coxal glands. The lateral gland may possibly function as an endocrine gland involved with production of a molting hormone. The rostral glands in inferior oribatids may have a salivary function. 5. The coxal glands of Peripatus, some millipedes, apterygote insects, decapod crustaceans, and all arachnid orders are homologous. The Tetrastigmata, Notostigmata, Cryptostigmata, and soft ticks have typical arachnid coxal glands. The coxal glands of higher Prostigmata may be modified into salivary, silk, or venom glands. The coxal glands in Mesostigmata, Astigmata, and hard ticks are lacking or highly modified.     

Morphology of locomotor appendages in Arachnida: evolutionary trends and phylogenetic implications

The morphological diversity of locomotor appendages in Arachnida is surveyed lo reconstruct phylogenetic relationships and discover evolutionary trends in form and function. The appendicular skeleton and musculature of representatives from the ten living arachnid orders ate described, and a system of homology is proposed. Character polarities are established through comparison with an outgroup. Limulus polyphemus Xiphosura). Cladistic analysis suggests that Arachnida is monophyletic and that absence of extensor muscles is a primitive condition. Extensors are primitively absent in Araneae. Amblypygi, Uropygi, Palpigradi, Ricinulei and Acari. Most similarities in the appendages of these orders are symplesiomorphic so that phylogenetic relationships among the 'extensorless' groups cannot be resolved solely on the basis of appendicular characters. Extensor muscles appear to have evolved once, and their presence is considered a synapomorphic feature of Opiliones, Scorpiones, Pseudoscorpiones and Solifugae. Solifugae lack extensors, but a parsimonious interpretation of other characters indicates that this is a secondary, derived condition. The phylogenetic relationships among these four orders are clarified by modifications of the patellotibial joint. Cladistic analysis indicates that Opiliones may be the sister group of the other three orders and that Scorpiones is the sister group of Pseudoscorpiones and Solifugae. Conclusions concerning phylogenetic relationships and evolutionary morphology presented here differ substantially from those of earlier studies on the locomotor appendages of Arachnida.     

Morphology of locomotor appendages in Arachnida: evolutionary trends and phylogenetic implications

The morphological diversity of locomotor appendages in Arachnida is surveyed lo reconstruct phylogenetic relationships and discover evolutionary trends in form and function. The appendicular skeleton and musculature of representatives from the ten living arachnid orders ate described, and a system of homology is proposed. Character polarities are established through comparison with an outgroup. Limulus polyphemus Xiphosura). Cladistic analysis suggests that Arachnida is monophyletic and that absence of extensor muscles is a primitive condition. Extensors are primitively absent in Araneae. Amblypygi, Uropygi, Palpigradi, Ricinulei and Acari. Most similarities in the appendages of these orders are symplesiomorphic so that phylogenetic relationships among the ‘extensorless’ groups cannot be resolved solely on the basis of appendicular characters. Extensor muscles appear to have evolved once, and their presence is considered a synapomorphic feature of Opiliones, Scorpiones, Pseudoscorpiones and Solifugae. Solifugae lack extensors, but a parsimonious interpretation of other characters indicates that this is a secondary, derived condition. The phylogenetic relationships among these four orders are clarified by modifications of the patellotibial joint. Cladistic analysis indicates that Opiliones may be the sister group of the other three orders and that Scorpiones is the sister group of Pseudoscorpiones and Solifugae. Conclusions concerning phylogenetic relationships and evolutionary morphology presented here differ substantially from those of earlier studies on the locomotor appendages of Arachnida.     

Evolution of locomotion in arachnida: The hydraulic pressure pump of the giant whipscorpion,Mastigoproctus Giganteus (Uropygi)

Many arachnids lack extensor muscles at the femoropatellar (knee) joint of their legs and extend this joint with hydraulic pressure during locomotion. Pressure is generated through compression of the prosoma, but there is disagreement about which muscles are involved in this process. Many arachhnologists consider contraction of the musculi laterales, a group of modified extrinsic leg muscles, as the cause of high prosomal pressure and regard hydraulic extension as a derived feature. However, integration of results from phylogenetic and comparative anatomical studies supports the view that hydraulic extension is primitive in Arachnida and that fluid pressure is generated by contraction of endosternal suspensor muscles. The functional predictions of the musculi laterales and endosternite hypotheses were tested by measuring muscle activity and prosomal pressure during unrestrained locomotion in a primitively “extensorless” arachnid, the giant whipscorpion. The results corroborate the endosternite model and refute the musculi laterales model. Changes in the prosomal pressure baseline were correlated with changes in endosternal muscle activity, while the musculi laterales fired in a step-coupled pattern of discrete bursts that appeared to be incapable of generating the pressure observed during locomotion. Step-coupled fluctuations in prosomal pressure were observed but were apparently caused by rapid flexing of the femoropatellar joints of the fourth leg pair rather than contraction of the musculi laterales.     

A trigonotarbid arachnid from the Lower Devonian of Tredomen, Wales

A new trigonotarbid (Arachnida: Trigonotarbida) Arianrhoda bennetti gen. et sp. nov. is described from the Lower Devonian (Lochkovian) of a quarry near Tredomen, Powys, mid Wales, UK. This relatively complete specimen is the first record of a pre-Carboniferous arachnid from Wales, one of only a handful of early Devonian arachnids, and the second oldest trigonotarbid recorded. Based on the rounded prosomal dorsal shield and the relatively narrow, elongate opisthosoma we refer this new fossil to the family Anthracosironidae. A distinct flange-like ornament on the leg 4 tibia in the new fossil is unique among trigonotarbids and is the primary autapomorphy for the new genus.     

Underlying Synapomorphies and Anagenetic Analysis

Evaluations of holomorphological similarities are based on synapomorphies, symplesiomorphies, convergence, and parallelisms as results of parallel selection and of underlying synapomorphies respectively. Only synapomorphies and underlying synapomorphies can show genealogical relationships. Distinctions between parallel selections and underlying synapomorphies are of major phylogenetic importance, while distinctions between different evolutionary histories (eu-parallelisms and pseudo-parallelisms) are not. The circumstance when underlying synapomorphies are of special phylogenetic importance has been termed unique inside-parallelism. Three such unique inside-parallelisms are found in the female genitalia of the Chironomidae, where they are shown necessary for the understanding of subfamily relationships. — The first minimum criterion for recognizing synapomorphy (Schlee 1971) is corrected to: It should be present within the whole group or clearly secondarily reduced an apomorphic taxa. It should not be present in the same formation in any taxon outside the group which can be regarded as a possible sister group. — The anagenetic component of the evolutionary processes can, following a cladistic analysis, be calculated by means of the adjusted evolution index assigning the different recognizable steps of trends or morphocline number from 1 to 2 and calculating the arithmetic mean of all numbers. Examples from Chaoboridae and Chironomidae support the cladistic diagrams and point out that different stages belong to differing "grades". Methods of numerical taxonomy may give a finer gradation of anagenetic levels.     

The affinities of mites and ticks: a review

Mites and ticks can be divided into two well-defined clades, Anactinotrichida and Actinotrichida, for which a recent work formalized a suite of putative autapomorphies and reciprocal differences. Whether they are sister-taxa – forming a monophyletic Acari – is more controversial. Earlier supporters of two independent origins for mites largely failed to demonstrate convincing synapomorphies between either of the two lineages and other arachnid orders; although recent work on reproductive biology revealed explicit characters of this nature. Furthermore, some of the characters proposed in support of a monophyletic Acari do not stand up to detailed scrutiny when compared with Arachnida in general. Effective morphological comparisons between mites and other arachnids are hindered by incompatible nomenclature and long-standing, mite-specific characters which are difficult to score for other arachnids. Furthermore, taxon-specific characters restricted to individual mite groups have sometimes been treated erroneously as 'typical' for all Acari. Here, previous hypotheses of mite affinities are reviewed. Historically, authors have debated whether mites are basal arachnids or highly derived. Excluding weakly supported early hypotheses, mites have been resolved – in whole or in part – as sister-group of all other Arachnida (based on tagmosis), closely related to Opiliones (based mostly on genital morphology), Palpigradi (based on controversial interpretations of limb morphology), Solifugae (based mostly on the mouthparts, but now perhaps also reproductive characters) and Ricinulei (based on hexapodal larvae and perhaps mouthparts). We cannot provide a final resolution here, but we aim to highlight important character sets which should be included in subsequent phylogenetic analyses, as well as useful areas for future investigations: particularly tagmosis and the nature of the gnathosoma.     

Evolutionary conservation and versatility of a new set of primers for amplifying the ribosomal internal transcribed spacer regions in insects and other invertebrates

The evolutionary conservation and versatility of a new set of nuclear primers for the amplification of the ribosomal internal transcribed spacer regions in insects and other invertebrates have been studied. These primers, conserved across Insecta and other invertebrates, are based on a comprehensive taxonomic survey of the current DNA sequence databases. Their versatility was demonstrated by extensive polymerase chain reaction assays in 16 species from two arachnid orders, eight insect orders, three invertebrate and vertebrate chordate orders and by direct sequencing of the amplified products. The availability of these primers to the insect research community should facilitate the use of internal transcribed spacer regions in intraspecific studies as well as phylogenetic analysis of closely related taxa.     

A phylogenetic analysis of the arachnid orders based on morphological characters

Morphological evidence for resolving relationships among arachnid orders was surveyed and assembled in a matrix comprising 59 euchelicerate genera (41 extant, 18 fossil) and 202 binary and unordered multistate characters. Parsimony analysis of extant genera recovered a monophyletic Arachnida with the topology (Palpigradi (Acaromorpha (Tetrapulmonata (Haplocnemata, Stomothecata nom. nov. )))), with Acaromorpha containing Ricinulei and Acari, Tetrapulmonata containing Araneae and Pedipalpi (Amblypygi, Uropygi), Haplocnemata (Pseudoscorpiones, Solifugae) and Stomothecata (Scorpiones, Opiliones). However, nodal support and results from exploratory implied weights analysis indicated that relationships among the five clades were effectively unresolved. Analysis of extant and fossil genera recovered a clade, Pantetrapulmonata nom nov. , with the topology (Trigonotarbida (Araneae (Haptopoda (Pedipalpi)))). Arachnida was recovered as monophyletic with the internal relationships (Stomothecata (Palpigradi, Acaromorpha (Haplocnemata, Pantetrapulmonata))). Nodal support and exploratory implied weights indicated that relationships among these five clades were effectively unresolved. Thus, some interordinal relationships were strongly and/or consistently supported by morphology, but arachnid phylogeny is unresolved at its deepest levels. Alternative hypotheses proposed in the recent literature were evaluated by constraining analyses to recover hypothesized clades, an exercise that often resulted in the collapse of otherwise well-supported clades. These results suggest that attempts to resolve specific nodes based on individual characters, lists of similarities, evolutionary scenarios, etc., are problematic, as they ignore broader impacts on homoplasy and analytical effects on non-target nodes.  © 2007 The Linnean Society of London, Zoological Journal of the Linnean Society , 2007, 150 , 221–265.     

Multi-gene analysis provides a well-supported phylogeny of Rosales

Despite many attempts to resolve evolutionary relationships among the major clades of Rosales, some nodes have been extremely problematic and have remained unresolved. In this study, we use two nuclear and 10 plastid loci to infer phylogenetic relationships among all nine families of Rosales. Rosales were strongly supported as monophyletic; within Rosales all family relationships are well-supported with Rosaceae sister to all other members of the order. Remaining Rosales can be divided into two subclades: (1) Ulmaceae are sister to Cannabaceae plus (Urticaceae+Moraceae); (2) Rhamnaceae are sister to Elaeagnaceae plus (Barbeyaceae+Dirachmaceae). One noteworthy result is that we recover the first strong support for a sister relationship between the enigmatic Dirachmaceae and Barbeyaceae. These two small families have distinct morphologies and potential synapomorphies remain unclear. Future studies should try to identify nonDNA synapomorphies uniting Barbeyaceae with Dirachmaceae.     

Phylogeny and a revised classification of the Monogenoidea Bychowsky, 1937 (Platyhelminthes)

A hypothesis (CI=57.3%) on the evolutionary relationships of families comprising the class Monogenoidea is proposed based on 141 character states in 47 homologous series and employing phylogenetic systematics. Based on the analysis, three subclasses, the Polyonchoinea, Polystomatoinea and Oligonchoinea, are recognised. The analysis supports independent origins of the Montchadskyellidae within the Polyonchoinea and of the Neodactylodiscidae and Amphibdellatidae within the order Dactylogyridea (Polyonchoinea); the suborder Montchadskyellinea is raised to ordinal status and new suborders Neodactylodiscinea and Amphibdellatinea are proposed to reflect these origins. The Gyrodactylidea (Polyonchoinea) is supported by three synapomorphies and comprises the Gyrodactylidae, Anoplodiscidae, Tetraonchoididae and Bothitrematidae. The analysis supports recognition of the Polystomatoinea comprising Polystomatidae and Sphyranuridae. Evolutionary relationships within the Oligonchoinea indicate independent origins of three ordinal taxa, the Chimaericolidea (monotypic), Diclybothriidea (including Diclybothriidae and Hexabothriidae) and Mazocraeidea (with five suborders). The suborder Mazocraeinea comprises the Plectanocotylidae, Mazocraeidae and Mazoplectidae, and is characterised by two synapomorphies. The suborder Gastrocotylinea, characterised by presence of accessory sclerites in the haptoral sucker, is divided into two infraorders, the monotypic Anthocotylina infraorder novum and Gastrocotylina. Two superfamilies of the Gastrocotylina are recognised, the Protomicrocotyloidea and Gastrocotyloidea; the Pseudodiclidophoridae is considered incertae sedis within the Gastrocotylina. The suborder Discocotylinea comprises the Discocotylidae, Octomacridae and Diplozoidae and is supported by four synapomorphies. The monotypic Hexostomatinea suborder novum is proposed to reflect an independent origin of the Hexostomatidae within the Mazocraeidea. The terminal suborder Microcotylinea comprises four superfamilies, the Microcotyloidea, Allopyragraphoroidea, Diclidophoroidea and Pyragraphoroidea. The analysis supports incorporation of the Pterinotrematidae in the Pyragraphoroidea and rejection of the monotypic order Pterinotrematidea. The following taxa are also rejected for reasons of paraphyly and/or polyphyly: Articulonchoinea, Bothriocotylea, Eucotylea, Monoaxonematidea, Tetraonchidea, Gotocotyloidea, Anchorophoridae and Macrovalvitrematidae. The Sundanonchidae, Iagotrematidae and Microbothriidae were not included in the analysis because of lack of pertinent information regarding character states.     

Phylogenetic relationships among the Notharctinae of North America

Study of over 1,000 specimens representing all notharctine genera and species leads to the conclusion that current concepts about the relationships of genera within the Notharctinae are incorrect. The following describes the more probable relationships among these genera. 1) Smilodectes and Notharctus are more closely related to each other than either is to any known early Eocene notharctine. Synapomorphies linking these genera include relatively narrow upper and lower molars, a relatively low-crowned P4, and paraconid size reduction on M1-3. 2) Among known Wasatchian notharctines, a clade consisting of Copelemur tutus and Cop. praetutus shares several lower molar synapomorphies with the Notharctus-Smilodectes clade, and therefore appears to form the Wasatchian sister group of Bridgerian notharctines. Synapomorphies documenting this relationship include well-developed entoconid notches on P4-M2, an anteriorly placed paraconid on M2, and a long premetacristid on M2. 3) Copelemur and Pelycodus are independently derived from early North American Cantius. Recent suggestions that the European adapine taxa Leptadapis priscus and Microadapis sciureus share special phylogenetic relationships with Smilodectes are rejected. The reduced (or lack of a) paraconid and morphology of the paracristid and other features identified as synapomorphies linking these adapines with Smilodectes are also characteristic of most other adapines as well (e.g., other species of Leptadapis, Adapis, Europolemur, and Anchomomys). Such traits developed independently in Smilodectes, which is clearly a notharctine on the basis of many synapomorphies and thus are not evidence of a close phylogenetic relationship between Smilodectes and L. priscus or M. sciureus.     

Comparative ultrastructure of coxal glands in unfed larvae of Leptotrombidium orientale (Schluger, 1948) (Trombiculidae) and Hydryphantes ruber (de Geer, 1778) (Hydryphantidae)

Coxal glands of unfed larvae Leptotrombidium orientale (Schluger, 1948) (Trombiculidae), a terrestrial mite parasitizing vertebrates, and Hydryphantes ruber (de Geer, 1778) (Hydryphantidae), a water mite parasitizing insects were studied using transmission electron microscopy. In both species, the coxal glands are represented by a paired tubular organ extending on the sides of the brain from the mouthparts to the frontal midgut wall and are formed of the cells arranged around the central lumen. As in other Parasitengona, the coxal glands are devoid of a proximal sacculus. The excretory duct, joining with ducts of the prosomal salivary glands constitutes the common podocephalic duct, opening into the subcheliceral space. The coxal glands of L. orientale are composed of a distal tubule with a basal labyrinth, an intermediate segment without labyrinth, and a proximal tubule bearing tight microvilli on the apical cell surface and coiled around the intermediate segment. The coxal glands of H. ruber mainly consist of the uniformly organized proximal tubule with apical microvilli of the cells lacking the basal labyrinth. This tubule shows several loops running backward and forward in a vertical plane on the side of the brain. In contrast to L. orientale , larvae of H. ruber reveal a terminal cuticular sac/bladder for accumulation of secreted fluids. Organization of the coxal glands depends on the ecological conditions of mites. Larvae of terrestrial L. orientale possess distal tubule functioning in re‐absorption of ions and water. Conversely, water mite larvae H. ruber need to evacuate of the water excess, so the filtrating proximal tubule is prominent.     

The mitochondrial genome of the water spider Argyroneta aquatica (Araneae: Cybaeidae)

We sequenced nearly the entire mitochondrial genome of Argyroneta aquatica, a wholly underwater‐living spider, thereby enhancing the available genomic information for Arachnida. The confirmed sequences contained the complete set of known genes present in other metazoan mitochondrial genomes. However, the mitochondrial gene order of A. aquatica was distinctly different from that of the most distant Chelicerata Limulus polyphemus (Xiphosura), probably because of a series of gene translocations and/or inversions. Comparison of arachnid mitochondrial gene orders for the purpose of phylogenetic inference is only minimally useful, but provides a strong signal in closely related lineages. To test the basal relationships and the evolutionary pattern of tRNA gene rearrangements among Arachnida, phylogenetic analyses using amino acid sequences of the 13 protein‐coding genes were performed. An interesting feature, the five 135‐bp tandem repeats and two 363‐bp tandem repeats, was identified in the putative control region. Although control region tandem repeats have been reported in many other arachnid and metazoan species, this is the first time it has been described in spiders.     

High efficiency transformation of E. coli by high voltage electroporation.

E. coli can be transformed to extremely high efficiencies by subjecting a mixture of cells and DNA to brief but intense electrical fields of exponential decay waveform (electroporation). We have obtained 10(9) to 10(10) transformants/micrograms with strains LE392 and DH5 alpha, and plasmids pUC18 and pBR329. The process is highly dependent on two characteristics of the electrical pulse: the electric field strength and the pulse length (RC time constant). The frequency of transformation is a linear function of the DNA concentration over at least six orders of magnitude; and the efficiency of transformation is a function of the cell concentration. Most of the surviving cells are competent with up to 80% transformed at high DNA concentration. The mechanism does not appear to include binding of the DNA to the cells prior to entry. Possible mechanisms are discussed and a simple procedure for the practical use of this technique is presented.     

Chemosystematics in the Opiliones (Arachnida): a comment on the evolutionary history of alkylphenols and benzoquinones in the scent gland secretions of Laniatores

Large prosomal scent glands constitute a major synapomorphic character of the arachnid order Opiliones. These glands produce a variety of chemicals very specific to opilionid taxa of different taxonomic levels, and thus represent a model system to investigate the evolutionary traits in exocrine secretion chemistry across a phylogenetically old group of animals. The chemically best‐studied opilionid group is certainly Laniatores, and currently available chemical data allow first hypotheses linking the phylogeny of this group to the evolution of major chemical classes of secretion chemistry. Such hypotheses are essential to decide upon a best‐fitting explanation of the distribution of scent‐gland secretion compounds across extant laniatorean taxa, and hence represent a key toward a well‐founded opilionid chemosystematics.