Correlating Molecular Phylogeny with Venom Apparatus Occurrence in Panamic Auger Snails (Terebridae)

Central to the discovery of neuroactive compounds produced by predatory marine snails of the superfamily Conoidea (cone snails, terebrids, and turrids) is identifying those species with a venom apparatus. Previous analyses of western Pacific terebrid specimens has shown that some Terebridae groups have secondarily lost their venom apparatus. In order to efficiently characterize terebrid toxins, it is essential to devise a key for identifying which species have a venom apparatus. The findings presented here integrate molecular phylogeny and the evolution of character traits to infer the presence or absence of the venom apparatus in the Terebridae. Using a combined dataset of 156 western and 33 eastern Pacific terebrid samples, a phylogenetic tree was constructed based on analyses of 16S, COI and 12S mitochondrial genes. The 33 eastern Pacific specimens analyzed represent four different species: Acus strigatus, Terebra argyosia, T. ornata, and T. cf. formosa. Anatomical analysis was congruent with molecular characters, confirming that species included in the clade Acus do not have a venom apparatus, while those in the clade Terebra do. Discovery of the association between terebrid molecular phylogeny and the occurrence of a venom apparatus provides a useful tool for effectively identifying the terebrid lineages that may be investigated for novel pharmacological active neurotoxins, enhancing conservation of this important resource, while providing supplementary information towards understanding terebrid evolutionary diversification.     

Evolution of the Toxoglossa venom apparatus as inferred by molecular phylogeny of the Terebridae

Toxoglossate marine gastropods, traditionally assigned to thefamilies Conidae, Terebridae, and Turridae, are one of the mostpopulous animal groups that use venom to capture their prey.These marine animals are generally characterized by a venomapparatus that consists of a muscular venom bulb and a tubularvenom gland. The toxoglossan radula, often compared with a hypodermicneedle for its use as a conduit to inject toxins into prey,is considered a major anatomical breakthrough that assistedin the successful initial radiation of these animals in theCretaceous and early Tertiary. The pharmacological success oftoxins from cone snails has made this group a star among biochemistsand neuroscientists, but very little is known about toxins fromthe other Toxoglossa, and the phylogeny of these families islargely in doubt. Here we report the first molecular phylogenyfor the Terebridae and use the results to infer the evolutionof the venom apparatus for this group. Our findings indicatethat most of the genera of terebrids are polyphyletic, and onespecies ("Terebra" (s.l.) jungi) is the sister group to allother terebrids. Molecular analyses combined with mapping ofvenom apparatus morphology indicate that the Terebridae havelost the venom apparatus at least twice during their evolution.Species in the genera Terebra and Hastula have the typical venomapparatus found in most toxoglossate gastropods, but all otherterebrid species do not. For venomous organisms, the dual analysisof molecular phylogeny and toxin function is an instructivecombination for unraveling the larger questions of phylogenyand speciation. The results presented here suggest a paradigmshift in the current understanding of terebrid evolution, whilepresenting a road map for discovering novel terebrid toxins,a largely unexplored resource for biomedical research and potentialtherapeutic drug development.     

Venomous auger snail Hastula (Impages) hectica (Linnaeus, 1758): molecular phylogeny, foregut anatomy and comparative toxinology

The >10,000 living venomous marine snail species [superfamily Conoidea (Fleming, 1822)] include cone snails (Conus), the overwhelming focus of research. Hastula hectica (Linnaeus, 1758), a venomous snail in the family Terebridae (M?rch, 1852) was comprehensively investigated. The Terebridae comprise a major monophyletic group within Conoidea. H. hectica has a striking radular tooth to inject venom that looks like a perforated spear; in Conus, the tooth looks like a hypodermic needle. H. hectica venom contains a large complement of small disulfide-rich peptides, but with no apparent overlap with Conus in gene superfamilies expressed. Although Conus peptide toxins are densely post-translationally modified, no post-translationally modified amino acids were found in any Hastula venom peptide. The results suggest that different major lineages of venomous molluscs have strikingly divergent toxinological and venom-delivery strategies.     

THE TOXOGLOSSAN PROBOSCIS: STRUCTURE AND FUNCTION

The structure of the proboscis and anterior alimentary systemis described and reviewed in the three families (Conidae, Terebridae,Turridae) of the Conoidea (=Toxoglossa). A number of proboscisforms are shown, characterized by the presence of a permanentrhynchodeum, modified alimentary system and a venom apparatus.Despite the diversity of prey types in the Conidae (polychaetes,molluscs and fish) they have a uniform proboscis structure witha long buccal tube and functional buccal and venom apparati.This homogeneity is not present in the Turridae and the Terebridaewhich show a variety of proboscis structures. Terebrids feedonly on polychaetes or hemichordates but appear to have threeprimary feeding modes based on significant difference in thestructure and function of the buccal tube and associated buccalorgans. The greatest variety of proboscis type is found in thediverse Turridae which feed mainly on polychaetes, but includesipunculans, gastropods and nemerteans in their diet.     

A new type of gastropod proboscis: the foregut of Hastula bacillus (Gastropoda: Terebridae)

Hastula bacillus (Deshayes) is a small terebrid gastropod which inhabits sandy surf beaches in southern Thailand, where it feeds upon spionid polychaetes. It possesses a foregut anatomy unlike that of any other gastropod. An elongate arborescent muscular organ, known as the accessory proboscis structure, is extended through the mouth during foraging. When retracted, it is folded into an 's' shape in the permanent rhynchodeum. The accessory proboscis structure bears numerous tufts of short, stiff cilia which are associated with pairs or triplets of dome-like structures. It is suggested that the structures may be chemosensory and concerned with prey location. Hastula bacillus also possesses a retractable labial tube, a long proboscis and buccal tube, dart-shaped radular teeth, an odontophore, an accessory salivary gland, a pair of salivary glands and a well-developed venom gland with muscular bulb. A comparison with other terebrid species suggests that H. bacillus is the most plesiomorphic taxon yet described from the family.     

Brassicaceae phylogeny and trichome evolution

To estimate the evolutionary history of the mustard family (Brassicaceae or Cruciferae), we sampled 113 species, representing 101 of the roughly 350 genera and 17 of the 19 tribes of the family, for the chloroplast gene ndhF. The included accessions increase the number of genera sampled over previous phylogenetic studies by four-fold. Using parsimony, likelihood, and Bayesian methods, we reconstructed the phylogeny of the gene and used the Shimodaira-Hasegawa test (S-H test) to compare the phylogenetic results with the most recent tribal classification for the family. The resultant phylogeny allowed a critical assessment of variations in fruit morphology and seed anatomy, upon which the current classification is based. We also used the S-H test to examine the utility of trichome branching patterns for describing monophyletic groups in the ndhF phylogeny. Our phylogenetic results indicate that 97 of 114 ingroup accessions fall into one of 21 strongly supported clades. Some of these clades can themselves be grouped into strongly to moderately supported monophyletic groups. One of these lineages is a novel grouping overlooked in previous phylogenetic studies. Results comparing 30 different scenarios of evolution by the S-H test indicate that five of 12 tribes represented by two or more genera in the study are clearly polyphyletic, although a few tribes are not sampled well enough to establish para- or polyphyly. In addition, branched trichomes likely evolved independently several times in the Brassicaceae, although malpighiaceous and stellate trichomes may each have a single origin.     

Assembling an arsenal: origin and evolution of the snake venom proteome inferred from phylogenetic analysis of toxin sequences

We analyzed the origin and evolution of snake venom toxin families represented in both viperid and elapid snakes by means of phylogenetic analysis of the amino acid sequences of the toxins and related nonvenom proteins. Out of eight toxin families analyzed, five provided clear evidence of recruitment into the snake venom proteome before the diversification of the advanced snakes (Kunitz-type protease inhibitors, CRISP toxins, galactose-binding lectins, M12B peptidases, nerve growth factor toxins), and one was equivocal (cystatin toxins). In two others (phospholipase A(2) and natriuretic toxins), the nonmonophyly of venom toxins demonstrates that presence of these proteins in elapids and viperids results from independent recruitment events. The ANP/BNP natriuretic toxins are likely to be basal, whereas the CNP/BPP toxins are Viperidae only. Similarly, the lectins were recruited twice. In contrast to the basal recruitment of the galactose-binding lectins, the C-type lectins were shown to be Viperidae only, with the alpha-chains and beta-chains resulting from an early duplication event. These results provide strong additional evidence that venom evolved once, at the base of the advanced snake radiation, rather than multiple times in different lineages, with these toxins also present in the venoms of the "colubrid" snake families. Moreover, they provide a first insight into the composition of the earliest ophidian venoms and point the way toward a research program that could elucidate the functional context of the evolution of the snake venom proteome.     

Sample Limited Characterization of a Novel Disulfide-Rich Venom Peptide Toxin from Terebrid Marine Snail Terebra variegata

Disulfide-rich peptide toxins found in the secretions of venomous organisms such as snakes, spiders, scorpions, leeches, and marine snails are highly efficient and effective tools for novel therapeutic drug development. Venom peptide toxins have been used extensively to characterize ion channels in the nervous system and platelet aggregation in haemostatic systems. A significant hurdle in characterizing disulfide-rich peptide toxins from venomous animals is obtaining significant quantities needed for sequence and structural analyses. Presented here is a strategy for the structural characterization of venom peptide toxins from sample limited (4 ng) specimens via direct mass spectrometry sequencing, chemical synthesis and NMR structure elucidation. Using this integrated approach, venom peptide Tv1 from Terebra variegata was discovered. Tv1 displays a unique fold not witnessed in prior snail neuropeptides. The novel structural features found for Tv1 suggest that the terebrid pool of peptide toxins may target different neuronal agents with varying specificities compared to previously characterized snail neuropeptides.     

Molecular phylogenetics and evolutionary diversification of labyrinth fishes (Perciformes: Anabantoidei)

Labyrinth fishes (Perciformes: Anabantoidei) are primary freshwater fishes with a disjunct African-Asian distribution that exhibit a wide variety of morphological and behavioral traits. These intrinsic features make them particularly well suited for studying patterns and processes of evolutionary diversification. We reconstructed the first molecular-based phylogenetic hypothesis of anabantoid intrarelationships using both mitochondrial and nuclear nucleotide sequence data to address anabantoid evolution. The mitochondrial data set included the complete cytochrome b, partial 12S rRNA, complete tRNA Val, and partial 16S rRNA genes (3332 bp) of 57 species representing all 19 anabantoid genera. The nuclear data set included the partial RAG1 gene (1494 bp) of 21 representative species. The phylogenetic analyses of a combined (mitochondrial+nuclear) data set recovered almost fully resolved trees at the intrafamily level with different methods of phylogenetic inference. Phylogenetic relationships at this taxonomic level were compared with previous morphology-based hypotheses. In particular, the enigmatic pike-head (Luciocephalus) was confidently placed within the "spiral egg" clade, thus resolving the long-standing controversy on its relative phylogenetic position. The molecular phylogeny was used to study the evolution of the different forms of parental care within the suborder. Our results suggest that the evolution of breeding behavior in anabantoids is highly correlated with phylogeny, and that brood care evolved three times independently from an ancestral free spawning condition without parental care. Ancestral character state reconstructions under maximum parsimony and maximum likelihood further indicated that both bubble nesting and mouthbrooding have evolved recurrently during anabantoid evolution. The new phylogenetic framework was also used to test alternative biogeographic hypotheses that account for the disjunct African-Asian distribution. Molecular divergence time estimates support either a drift vicariance linked to the breakup of Gondwana or Late Mesozoic Early Tertiary dispersal from Africa to Asia or vice versa.     

THE MORPHOLOGY OF TOXOGLOSSAN GASTROPODS LACKING A RADULA, WITH A DESCRIPTION OF NEW SPECIES AND GENUS OF TURRIDAE

The morphology of some deep-sea Turridae lacking radulae wasstudied. The main features of their digestive system are theabsence of a radula sac, venom and salivary glands and the reductionor absence of a proboscis. A new genus Teretiopsis including3 new species and T. thaumastopsis (Dautzenberg and Fischer,1896) is described. On the basis of differences of the digestivesystem when compared with other turrids lacking radulae themonotypical subfamily Taraninae Casey, 1904 new status (typegenus Taranis Jeffreys, 1870) is considered. It is shown thatthe process of radula reduction has occurred independently indifferent phylogenetic lines of Toxoglossa—the subfamiliesDaphnellinae and Taraninae among Turridae and the family Terebridae. (Received 8 August 1988; accepted 20 December 1988)     

Using 18S rDNA to resolve diaptomid copepod (Copepoda: Calanoida: Diaptomidae) phylogeny: an example with the North American genera

The phylogenetic relationships among the numerous genera of diaptomid copepods remain elusive due to difficulties in obtaining sufficient numbers of phylogenetically informative morphological characters for cladistic analysis. Molecular phylogenetic techniques offer high potential to resolve phylogenetic relationships in the absence of sufficient morphological characters because of the ease in which many characters can be unambiguously coded. I present the first molecular phylogeny for diaptomid copepod genera using 18S rDNA. Specifically, I test Light’s (1939) hypothesis regarding the interrelationships among the North American diaptomid genera. The 18S phylogeny is remarkably consistent with Light’s hypothesis. The endemic North American genera represent a monophyletic group exclusive of the non-endemic genera. Moreover, his hypothesized basal genus for the North America genera, Hesperodiaptomus, is the basal genus in this analysis. However, his Leptodiaptomus group is not reciprocally monophyletic with his Hesperodiaptomus group, but is rather a derived member of the latter group. Finally, the genus Mastigodiaptomus is found to be more closely allied with the non-endemic genera, as Light suggested. This phylogeny contributes heavily to the understanding of phylogenetic relationships among North American diaptomids and has large implications for the systematics of diaptomids in general. The use of 18S rDNA sequences in phylogenetic analyses of diaptomid copepods can be used to confirm the monophyly of recognized genera, the interrelationships among genera, and subsequent biogeographic interpretation of the family’s diversification. The use of molecular data, such as 18S rDNA sequences, to test phylogenetic hypotheses based on a very limited number of morphological characters will be a particularly useful approach to phylogenetic analysis in this system.     

Parallels in the evolution of the two largest New and Old World seed-beetle genera (Coleoptera, Bruchidae)

This study provides the first phylogenetic analysis of a large sample of the two largest genera of seed-beetles, Acanthoscelides Schilsky and Bruchidius Schilsky, which mostly feed on legumes (Fabaceae). The goal of this study was to investigate evolutionary patterns in relation to biogeography and host-plant associations. We used three mitochondrial molecular markers and parsimony and Bayesian inference methods to reconstruct the phylogeny of 76 species. In addition, we critically reviewed host-plant records in the literature for these two bruchid genera. Our results demonstrated the existence of two major clades, one New World and one largely Old World, which generally correspond to the two genera. Yet, current classification of several species is erroneous, so that both genera as currently defined are paraphyletic. We highlighted a strong trend toward specialization (with high taxonomic conservatism in host-plant use) exhibited by the two studied genera. However, we showed the existence of several host shifts during the evolution of this group of bruchids. Our phylogenetic hypotheses and our evaluation of host-plant associations both suggest that the two genera have undergone parallel evolution, as they have independently colonized similar host plants in their respective areas of distribution. Our estimation of divergence times indicated a more ancient origin for bruchids than that suggested by the fossil records. Interestingly, the suggested timing of diversification is consistent with the hypothesis of a radiation that could have occurred contemporaneously with the diversification of their legume hosts.     

The diversification of the northern temperate woody flora – A case study of the Elm family (Ulmaceae) based on phylogenomic and paleobotanical evidence

Ulmaceae is a woody family widespread in northern temperate forests. Despite the ecological importance of this family, its phylogeny and biogeographic history are poorly understood. In this study, we reconstruct phylogenetic relationships within the family and infer spatio-temporal diversification patterns based on chloroplast genome (complete cpDNA) and nuclear ribosomal DNA sequences (nrDNA). The seven Ulmaceae genera are resolved in two main clades (temperate vs. tropical) by both cpDNA and nrDNA sequences. The temperate clade includes four genera, Hemiptelea, Zelkova, Planera, and Ulmus. The relationships among Planera and other genera are controversial because of inconsistent topologies between plastid and nuclear data. The tropical clade includes three genera ((Ampelocera, Phyllostylon), Holoptelea). Molecular dating and diversification analyses show that Ulmaceae originated in the Early Cretaceous (ca. 110–125 Ma) with the main lineages establishing from the Late Cretaceous to the early Eocene. The diversification rate slowed during the middle to the late Paleogene (ca. 23–45 Ma), followed by a rapid diversification of the East Asian temperate group in the Neogene, congruent with a global cooling event. The ancestral state optimization analysis suggests an East Asian origin of the temperate Ulmaceae clade during the Paleocene, which is consistent with the fossil record. Both phylogenomic and fossil evidence support East Asia as a center of origin and diversification for the temperate woody lineages.     

Mitochondrial genome phylogeny reveals the deep-time origin of Gomphomastacinae (Orthoptera: Eumastacidae) and its alpine genera in China

Gomphomastacinae is a grasshopper subfamily in Eumastacidae, with a morphology and distribution distinct from other subfamilies. The alpine genera of Gomphomastacinae that inhabit the Qinghai–Tibet Plateau in China show unique characteristics adapted to high-altitude life. However, their phylogenetic position and biogeographic history remain controversial. Thus, to determine the diversification history of these alpine genera and the origin of the subfamily, we obtained mitochondrial genome sequences from all seven Gomphomastacinae genera distributed in China. The reconstructed phylogeny was well supported and confirmed the phylogenetic position of Gomphomastacinae within Eumastacidae. Time calibration revealed a deep-time origin of the subfamily dating back to the Cretaceous period, and the diversification among alpine genera was also an ancient pre-Miocene event (30–50 Ma). Based on phylogeny and time estimates, the most likely biogeographic scenario is that Gomphomastacinae originated from an ancestral lineage that lived in East Gondwana and dispersed to Central and Western Asia through India. Subsequently, the alpine genera likely diverged along with the uplift of the Qinghai–Tibet Plateau and survived drastic climate change by in situ adaptation to high-altitude dwellings.     

Phylogeny and evolutionary history of diplobathrid crinoids (Echinodermata)

Order Diplobathrida is a major clade of camerate crinoids spanning the Ordovician–Mississippian, yet phylogenetic relationships have only been inferred for Ordovician taxa. This has hampered efforts to construct a comprehensive tree of life for crinoids and develop a classification scheme that adequately reflects diplobathrid evolutionary history. Here, I apply maximum parsimony and Bayesian phylogenetic approaches to the fossil record of diplobathrids to infer the largest tree of fossil crinoids to date, with over 100 genera included. Recovered trees provide a framework for evaluating the current classification of diplobathrids. Notably, previous suborder divisions are not supported, and superfamily divisions will require significant modification. Although numerous revisions are required for families, most can be retained through reassignment of genera. In addition, recovered trees were used to produce phylogeny‐based estimates of diplobathrid lineage diversity. By accounting for ghost lineages, phylogeny‐based richness estimates offer greater insight into diversification and extinction dynamics than traditional taxonomy‐based approaches alone and provide a detailed summary of the ~150 million‐year evolutionary history of Diplobathrida. This study constitutes a major step toward producing a phylogeny of the Crinoidea and documenting crinoid diversity dynamics. In addition, it will serve as a framework for subsequent phylogeny‐based investigations of macroevolutionary questions.     

Phylogeny and biogeography of the freshwater crayfish Euastacus (Decapoda: Parastacidae) based on nuclear and mitochondrial DNA

Euastacus crayfish are endemic to freshwater ecosystems of the eastern coast of Australia. While recent evolutionary studies have focused on a few of these species, here we provide a comprehensive phylogenetic estimate of relationships among the species within the genus. We sequenced three mitochondrial gene regions (COI, 16S, and 12S) and one nuclear region (28S) from 40 species of the genus Euastacus, as well as one undescribed species. Using these data, we estimated the phylogenetic relationships within the genus using maximum-likelihood, parsimony, and Bayesian Markov Chain Monte Carlo analyses. Using Bayes factors to test different model hypotheses, we found that the best phylogeny supports monophyletic groupings of all but two recognized species and suggests a widespread ancestor that diverged by vicariance. We also show that Euastacus and Astacopsis are most likely monophyletic sister genera. We use the resulting phylogeny as a framework to test biogeographic hypotheses relating to the diversification of the genus.     

Polygyrid land snail phylogeny: external sperm exchange, early North American biogeography, iterative shell evolution

A fresh assessment of reproductive behaviour and anatomy, combined with new allozyme data, results in a phylogenetic hypothesis for the 23 polygyrid genera that differs substantially from previous attempts. Analyses based on this phylogeny suggest that (a) evolutionary transition from internal to external sperm exchange occurred only once in polygyrids, with unusual functional intermediates still extant; (b) polygyrid biogeographic history paralleled that of plethodontid salamanders, with hypothesized vicariance events at about 145, 120, 65 and 40 MA, and, dispersal events at about 55 and 40 MA; and (c) iterative shell evolution occurred despite a phylogenetic constraint on ontogenetic whorl-expansion rate.