Hemocyanin gene family evolution in spiders (Araneae), with implications for phylogenetic relationships and divergence times in the infraorder Mygalomorphae

Hemocyanins are multimeric copper-containing hemolymph proteins involved in oxygen binding and transport in all major arthropod lineages. Most arachnids have seven primary subunits (encoded by paralogous genes a–g), which combine to form a 24-mer (4 × 6) quaternary structure. Within some spider lineages, however, hemocyanin evolution has been a dynamic process with extensive paralog duplication and loss. We have obtained hemocyanin gene sequences from numerous representatives of the spider infraorders Mygalomorphae and Araneomorphae in order to infer the evolution of the hemocyanin gene family and estimate spider relationships using these conserved loci. Our hemocyanin gene tree is largely consistent with the previous hypotheses of paralog relationships based on immunological studies, but reveals some discrepancies in which paralog types have been lost or duplicated in specific spider lineages. Analyses of concatenated hemocyanin sequences resolved deep nodes in the spider phylogeny and recovered a number of clades that are supported by other molecular studies, particularly for mygalomorph taxa. The concatenated data set is also used to estimate dates of higher-level spider divergences and suggests that the diversification of extant mygalomorphs preceded that of extant araneomorphs. Spiders are diverse in behavior and respiratory morphology, and our results are beneficial for comparative analyses of spider respiration. Lastly, the conserved hemocyanin sequences allow for the inference of spider relationships and ancient divergence dates.     

Transpiration from forest dwelling and woodland Mygalomorphae (Araneae)

Rates of water loss in dry air at room temperature (21±2°C) have been measured from seven species of forest-dwelling mygalomorph spiders. They range from 0.260% body wt h^?1 in the tube-dwelling trapdoor spiderDyarcyops sp. (from the humid coastal regions of eastern Australia) to 0.030% wt h^?1 inBrachypelma smithi (from drier environments in Mexico). There is a tendency for rates of transpiration to be related to the humidities of the spiders' normal environments and micro-habitats.     

Two Trichosporon species isolated from Central-European mygalomorph spiders (Araneae: Mygalomorphae)

Trichosporon (Dikarya: Basidiomycota) is a genus of anamorphic yeasts typically associated with soil and water, although many species are causative agents of diseases in animals and man. Here we provide the first compelling evidence that spiders can be occasionally colonized by at least two Trichosporon species. Trichosporon dulcitum (Berkhout) Weijman 1979 was isolated from the exoskeleton of purse-web spider Atypus piceus, while Trichosporon porosum (Stautz) Middelhoven, Scorzetti & Fell 2001 was isolated from the exoskeleton of purse-web spider Atypus affinis. Both of the species were identified based on DNA sequence analysis of the host specimens displaying macroscopic signs of the superficial white mycosis on their exoskeleton. Only two specimens with macroscopic signs of superficial yeast growth were identified among the 125 individuals of A. affinis, A. piceus and Atypus muralis examined that were collected at various sites throughout the Czech Republic. The consistent burrow microclimate, uninterrupted occupancy of the single burrow for several subsequent years, and presence of prey remnants in the burrow below the purse-web may play a role in the course of infection of the mygalomorphs examined. The phylogenetic relationships of Trichosporon species are analyzed, concluding that association with invertebrates clusters predominantly among four groups of closely related species in independent Trichosporon clades.     

Two new species of Chaco Tullgren from the Atlantic coast of Uruguay (Araneae,Mygalomorphae, Nemesiidae)

We describe two new species of the nemesiid spider genus Chaco from Rocha Province, Uruguay. These new species are diagnosed based on genital morphology, male tibial apophysis spination, and burrow entrance. We test cospecificity of one species, Chaco costai,via laboratory mating experiments. The new species are diagnosed and illustrated and habitat characteristics, and capture behavior are described. We conduct a cladistic analysis based on a previously published morphological character matrix that now includes the newly described species.     

Four new Mouse Spider species (Araneae,Mygalomorphae, Actinopodidae,Missulena) from Western Australia

Four new species of the Mouse Spider genus Missulena Walckenaer, 1805 (family Actinopodidae) are described from Western Australia based on morphological features of adult males. Missulena leniae sp. n.(from the Carnarvon and Yalgoo biogeographic regions), Missulena mainae sp. n. (Carnarvon), Missulena melissae sp. n. (Pilbara) and Missulena pinguipes sp. n. (Mallee) represent a broad spectrum of morphological diversity found in this genus and differ from other congeners by details of the male copulatory bulb, colour patterns, eye sizes, leg morphology and leg spination. Two of the species, M. pinguipes sp. n. and M. mainae sp. n., are characterised by swollen metatarsi of the fourth legs in males, a feature not previously recorded in the family. A key to males of all named Missulena species from Australia is presented and allows their identification based on external morphology.     

Taxonomic revision of the tarantula genus Aphonopelma Pocock, 1901 (Araneae,Mygalomorphae, Theraphosidae) within the United States

This systematic study documents the taxonomy, diversity, and distribution of the tarantula spider genus Aphonopelma Pocock, 1901 within the United States. By employing phylogenomic, morphological, and geospatial data, we evaluated all 55 nominal species in the United States to examine the evolutionary history of Aphonopelma and the group’s taxonomy by implementing an integrative approach to species delimitation. Based on our analyses, we now recognize only 29 distinct species in the United States. We propose 33 new synonymies (Aphonopelma apacheum, Aphonopelma minchi, Aphonopelma rothi, Aphonopelma schmidti, Aphonopelma stahnkei = Aphonopelma chalcodes; Aphonopelma arnoldi = Aphonopelma armada; Aphonopelma behlei, Aphonopelma vogelae = Aphonopelma marxi; Aphonopelma breenei = Aphonopelma anax; Aphonopelma chambersi, Aphonopelma clarum, Aphonopelma cryptethum, Aphonopelma sandersoni, Aphonopelma sullivani = Aphonopelma eutylenum; Aphonopelma clarki, Aphonopelma coloradanum, Aphonopelma echinum, Aphonopelma gurleyi, Aphonopelma harlingenum, Aphonopelma odelli, Aphonopelma waconum, Aphonopelma wichitanum = Aphonopelma hentzi; Aphonopelma heterops = Aphonopelma moderatum; Aphonopelma jungi, Aphonopelma punzoi = Aphonopelma vorhiesi; Aphonopelma brunnius, Aphonopelma chamberlini, Aphonopelma iviei, Aphonopelma lithodomum, Aphonopelma smithi, Aphonopelma zionis = Aphonopelma iodius; Aphonopelma phanum, Aphonopelma reversum = Aphonopelma steindachneri), 14 new species (Aphonopelma atomicum sp. n., Aphonopelma catalina sp. n., Aphonopelma chiricahua sp. n., Aphonopelma icenoglei sp. n., Aphonopelma johnnycashi sp. n., Aphonopelma madera sp. n., Aphonopelma mareki sp. n., Aphonopelma moellendorfi sp. n., Aphonopelma parvum sp. n., Aphonopelma peloncillo sp. n., Aphonopelma prenticei sp. n., Aphonopelma saguaro sp. n., Aphonopelma superstitionense sp. n., and Aphonopelma xwalxwal sp. n.), and seven nomina dubia (Aphonopelma baergi, Aphonopelma cratium, Aphonopelma hollyi, Aphonopelma mordax, Aphonopelma radinum, Aphonopelma rusticum, Aphonopelma texense). Our proposed species tree based on Anchored Enrichment data delimits five major lineages: a monotypic group confined to California, a western group, an eastern group, a group primarily distributed in high-elevation areas, and a group that comprises several miniaturized species. Multiple species are distributed throughout two biodiversity hotspots in the United States (i.e., California Floristic Province and Madrean Pine-Oak Woodlands). Keys are provided for identification of both males and females. By conducting the most comprehensive sampling of a single theraphosid genus to date, this research significantly broadens the scope of prior molecular and morphological investigations, finally bringing a modern understanding of species delimitation in this dynamic and charismatic group of spiders.     

Species delimitation and phylogeography of Aphonopelma hentzi (Araneae, Mygalomorphae, Theraphosidae): cryptic diversity in North American tarantulas

Background

The primary objective of this study is to reconstruct the phylogeny of the hentzi species group and sister species in the North American tarantula genus, Aphonopelma, using a set of mitochondrial DNA markers that include the animal “barcoding gene”. An mtDNA genealogy is used to consider questions regarding species boundary delimitation and to evaluate timing of divergence to infer historical biogeographic events that played a role in shaping the present-day diversity and distribution. We aimed to identify potential refugial locations, directionality of range expansion, and test whether A. hentzi post-glacial expansion fit a predicted time frame.

Methods and Findings

A Bayesian phylogenetic approach was used to analyze a 2051 base pair (bp) mtDNA data matrix comprising aligned fragments of the gene regions CO1 (1165 bp) and ND1-16S (886 bp). Multiple species delimitation techniques (DNA tree-based methods, a “barcode gap” using percent of pairwise sequence divergence (uncorrected p-distances), and the GMYC method) consistently recognized a number of divergent and genealogically exclusive groups.

Conclusions

The use of numerous species delimitation methods, in concert, provide an effective approach to dissecting species boundaries in this spider group; as well they seem to provide strong evidence for a number of nominal, previously undiscovered, and cryptic species. Our data also indicate that Pleistocene habitat fragmentation and subsequent range expansion events may have shaped contemporary phylogeographic patterns of Aphonopelma diversity in the southwestern United States, particularly for the A. hentzi species group. These findings indicate that future species delimitation approaches need to be analyzed in context of a number of factors, such as the sampling distribution, loci used, biogeographic history, breadth of morphological variation, ecological factors, and behavioral data, to make truly integrative decisions about what constitutes an evolutionary lineage recognized as a “species”.     

Testing species boundaries in the Antrodiaetus unicolor complex (Araneae: Mygalomorphae: Antrodiaetidae): "paraphyly" and cryptic diversity

The inability to correctly identify species has far reaching implications in nearly all areas of biology, yet few studies investigate methods for delineating species boundaries. Moreover, once these boundaries have been hypothesized, little thought has been given to how these constructs can be further evaluated. We employ a molecular phylogenetic approach using nuclear 28S rRNA and mitochondrial cytochrome c oxidase subunit I genes to test the general efficacy of species boundaries in the Antrodiaetus unicolor spider species complex. Our analyses provide evidence that An. unicolor is "paraphyletic" with respect to An. microunicolor, indicating that morphological criteria used to delineate species boundaries undersplits actual species-level diversity in this group of spiders. These analyses also demonstrate that individuals from geographically proximate populations sometimes exhibit considerable molecular divergence, strongly suggesting that An. unicolor is a cryptic species complex. Finally, this molecular approach has provided the phylogenetic framework that is necessary to begin interpreting the vast amount of morphological variation observed in these spiders based upon findings from previous studies. Our approach using multiple genes appears to be a rigorous method to critically examine species boundaries originally based on traditional morphological approaches to spider taxonomy.     

Molecular phylogenetics of the spider infraorder Mygalomorphae using nuclear rRNA genes (18S and 28S): conflict and agreement with the current system of classification

Mygalomorph spiders, which include the tarantulas, trapdoor spiders, and their kin, represent one of three main spider lineages. Mygalomorphs are currently classified into 15 families, comprising roughly 2500 species and 300 genera. The few published phylogenies of mygalomorph relationships are based exclusively on morphological data and reveal areas of both conflict and congruence, suggesting the need for additional phylogenetic research utilizing new character systems. As part of a larger combined evidence study of global mygalomorph relationships, we have gathered approximately 3.7 kb of rRNA data (18S and 28S) for a sample of 80 genera, representing all 15 mygalomorph families. Taxon sampling was particularly intensive across families that are questionable in composition-Cyrtaucheniidae and Nemesiidae. The following primary results are supported by both Bayesian and parsimony analyses of combined matrices representing multiple 28S alignments: (1) the Atypoidea, a clade that includes the families Atypidae, Antrodiaetidae, and Mecicobothriidae, is recovered as a basal lineage sister to all other mygalomorphs, (2) diplurids and hexathelids form a paraphyletic grade at the base of the non-atypoid clade, but neither family is monophyletic in any of our analyses, (3) a clade consisting of all sampled nemesiids, Microstigmata and the cyrtaucheniid genera Kiama, Acontius, and Fufius is consistently recovered, (4) other sampled cyrtaucheniids are fragmented across three separate clades, including a monophyletic North American Euctenizinae and a South African clade, (5) of the Domiothelina, only idiopids are consistently recovered as monophyletic; ctenizids are polyphyletic and migids are only weakly supported. The Domiothelina is not monophyletic. The molecular results we present are consistent with more recent hypotheses of mygalomorph relationship; however, additional work remains before mygalomorph classification can be formally reassessed with confidence-increased taxonomic sampling and the inclusion of additional character systems (more genes and morphology) are required.     

Searching for gene flow from cultivated to wild strawberries in Central Europe

Background and Aims

Experimental crosses between the diploid woodland strawberry (Fragaria vesca L.) and the octoploid garden strawberry (F. × ananassa Duch.) can lead to the formation of viable hybrids. However, the extent of such hybrid formation under natural conditions is unknown, but is of fundamental interest and importance in the light of the potential future cultivation of transgenic strawberries. A hybrid survey was therefore conducted in the surroundings of ten farms in Switzerland and southern Germany, where strawberries have been cultivated for at least 10 years and where wild strawberries occur in the close vicinity.

Methods

In 2007 and 2008, 370 wild F. vesca plants were sampled at natural populations around farms and analysed with microsatellite markers. In 2010, natural populations were revisited and morphological traits of 3050 F. vesca plants were inspected. DNA contents of cell nuclei of morphologically deviating plants were estimated by flow cytometry to identify hybrids. As controls, 50 hybrid plants from interspecific hand-crosses were analysed using microsatellite analysis and DNA contents of cell nuclei were estimated by flow cytometry.

Key Results

None of the wild samples collected in 2007 and 2008 contained F. × ananassa microsatellite markers, while all hybrids from hand-crosses clearly contained markers of both parent species. Morphological inspection of wild populations carried out in 2010 and subsequent flow cytometry of ten morphologically deviating plants revealed no hybrids.

Conclusions

Hybrid formation or hybrid establishment in natural populations in the survey area is at best a rare event.     

Three new species of Melloleitaoina Gerschman & Schiapelli, 1960 (Araneae,Mygalomorphae, Theraphosidae) from northern Argentina

Three new species of the monotypic genus Melloleitaoina Gerschman & Schiapelli, 1960 are described from northern Argentina: M. mutquina sp. n., M. uru sp. n. and M. yupanqui sp. n. The female specimen originally described as M. crassifemur is not conspecific with the male holotype and thus is removed from this species and described as M. uru sp. n.; M. crassifemur is redescribed. All species are diagnosed, illustrated and a key to species is provided.     

Central projections of cheliceral mechanoreceptors in the spider Cupiennius salei (Arachnida,Araneae)

Central projections of lyriform organs and tactile hairs on the chelicerae of the wandering spider Cupiennius salei were traced using anterograde cobalt fills. Different fibers arising from both mechanoreceptor types arborize in the cheliceral ganglia, which are part of the tritocerebrum, and in sensory longitudinal tracts in the center of the suboesophageal nerve mass together with afferent fibers arising from mechanoreceptors on the walking legs and the pedipalps. This convergence of sensory projections in the sensory longitudinal tracts might provide the anatomical basis for the coordination of the movements of different extremities during prey capture and feeding. The findings also support the hypothesis that the tritocerebrum originally was a preoral ganglion in spiders. © 1993 Wiley-Liss, Inc.     

An alu-based phylogeny of lemurs (infraorder: lemuriformes)

LEMURS (INFRAORDER: Lemuriformes) are a radiation of strepsirrhine primates endemic to the island of Madagascar. As of 2012, 101 lemur species, divided among five families, have been described. Genetic and morphological evidence indicates all species are descended from a common ancestor that arrived in Madagascar ～55-60 million years ago (mya). Phylogenetic relationships in this species-rich infraorder have been the subject of debate. Here we use Alu elements, a family of primate-specific Short INterspersed Elements (SINEs), to construct a phylogeny of infraorder Lemuriformes. Alu elements are particularly useful SINEs for the purpose of phylogeny reconstruction because they are identical by descent and confounding events between loci are easily resolved by sequencing. The genome of the grey mouse lemur (Microcebus murinus) was computationally assayed for synapomorphic Alu elements. Those that were identified as Lemuriformes-specific were analyzed against other available primate genomes for orthologous sequence in which to design primers for PCR (polymerase chain reaction) verification. A primate phylogenetic panel of 24 species, including 22 lemur species from all five families, was examined for the presence/absence of 138 Alu elements via PCR to establish relationships among species. Of these, 111 were phylogenetically informative. A phylogenetic tree was generated based on the results of this analysis. We demonstrate strong support for the monophyly of Lemuriformes to the exclusion of other primates, with Daubentoniidae, the aye-aye, as the basal lineage within the infraorder. Our results also suggest Lepilemuridae as a sister lineage to Cheirogaleidae, and Indriidae as sister to Lemuridae. Among the Cheirogaleidae, we show strong support for Microcebus and Mirza as sister genera, with Cheirogaleus the sister lineage to both. Our results also support the monophyly of the Lemuridae. Within Lemuridae we place Lemur and Hapalemur together to the exclusion of Eulemur and Varecia, with Varecia the sister lineage to the other three genera.     

A total evidence assessment of the phylogeny of North American euctenizine trapdoor spiders (Araneae, Mygalomorphae, Cyrtaucheniidae) using Bayesian inference

North American trapdoor spiders of the subfamily Euctenizinae (Cyrtaucheniidae) are among the most diverse mygalomorph spiders (trapdoor spiders, tarantulas, and their relatives) on the continent in terms of species numbers and ecological habits. We present a generic level phylogenetic study of the subfamily based on a total evidence approach. Our dataset comprises approximately 3.7 kb of molecular characters (18S and 28S rRNA gene sequences) and 71 morphological characters scored for 32 taxa. When analyzed independently, these data sets, particularly the morphology, depict very different views of mygalomorph and euctenizine relationships, albeit with weak support. However, when these data are combined we recover a tree topology that is supported by high posterior probability for most nodes. The combined data recover a phylogenetic pattern for euctenizines different than previously published and indicate the presence of a narrowly endemic new genus from central California. While euctenizine monophyly is unequivocal, the monophyly of a number of other mygalomorph groups is questionable (e.g., Cyrtaucheniidae, Mecicobothriodina, Rastelloidina). This non-monophyly is noteworthy, as our analysis represents the first employing a total evidence approach for mygalomorphs, a group known to be morphologically conservative.     

Checklist of the infraorder Tipulomorpha (Trichoceridae,Tipuloidea) (Diptera) of Finland

A checklist of the infraorder Tipulomorpha: families Trichoceridae, Pediciidae, Limoniidae, Cylindrotomidae and Tipulidae (Diptera) recorded from Finland.     

Some rare desmids (Zygnematophyceae) from Central Europe

The morphology and taxonomy of eight rare desmids (Zygnematophyceae) from Central Europe are studied, i.e. six taxa from Slovakia [Tortitaenia bahusiensis (Nordstedt et Lütkemüller) Coesel, Closterium limneticum Lemmermann var. fallax R??i?ka, Euastrum sublobatum de Brébisson in Ralfs, morpha, Cosmarium simplicius (W. et G.S. West) Grönblad, Pachyphorium canadense (Irénée-Marie) G. H. Tomaszewicz et Hindák, stat. et comb. nova, Staurastrum bloklandiae Coesel et Joosten], one species from Poland (Spirotaenia bacillaris Lütkemüller) and Austria (Cosmarium geminatum Lundell). All desmids taxa are new records for the country of their origin.