Publications

2014
Lenihan, J., S. Kvist, R. Fernandez, G. Giribet, and A. Ziegler. 2014. “A dataset comprising four micro-computed tomography scans of freshly fixed and museum earthworm specimens.” Gigascience 3: 6.Abstract

BACKGROUND: ALTHOUGH MOLECULAR TOOLS ARE INCREASINGLY EMPLOYED TO DECIPHER INVERTEBRATE SYSTEMATICS, EARTHWORM (ANNELIDA: Clitellata: 'Oligochaeta') taxonomy is still largely based on conventional dissection, resulting in data that are mostly unsuitable for dissemination through online databases. In order to evaluate if micro-computed tomography (muCT) in combination with soft tissue staining techniques could be used to expand the existing set of tools available for studying internal and external structures of earthworms, muCT scans of freshly fixed and museum specimens were gathered. FINDINGS: Scout images revealed full penetration of tissues by the staining agent. The attained isotropic voxel resolutions permit identification of internal and external structures conventionally used in earthworm taxonomy. The muCT projection and reconstruction images have been deposited in the online data repository GigaDB and are publicly available for download. CONCLUSIONS: The dataset presented here shows that earthworms constitute suitable candidates for muCT scanning in combination with soft tissue staining. Not only are the data comparable to results derived from traditional dissection techniques, but due to their digital nature the data also permit computer-based interactive exploration of earthworm morphology and anatomy. The approach pursued here can be applied to freshly fixed as well as museum specimens, which is of particular importance when considering the use of rare or valuable material. Finally, a number of aspects related to the deposition of digital morphological data are briefly discussed.

Fernandez, R., C. E. Laumer, V. Vahtera, S. Libro, S. Kaluziak, P. P. Sharma, A. R. Perez-Porro, G.D. Edgecombe, and G. Giribet. 2014. “Evaluating topological conflict in centipede phylogeny using transcriptomic data sets.” Mol Biol Evol 31: 1500-13.Abstract

Relationships between the five extant orders of centipedes have been considered solved based on morphology. Phylogenies based on samples of up to a few dozen genes have largely been congruent with the morphological tree apart from an alternative placement of one order, the relictual Craterostigmomorpha, consisting of two species in Tasmania and New Zealand. To address this incongruence, novel transcriptomic data were generated to sample all five orders of centipedes and also used as a test case for studying gene-tree incongruence. Maximum likelihood and Bayesian mixture model analyses of a data set composed of 1,934 orthologs with 45% missing data, as well as the 389 orthologs in the least saturated, stationary quartile, retrieve strong support for a sister-group relationship between Craterostigmomorpha and all other pleurostigmophoran centipedes, of which the latter group is newly named Amalpighiata. The Amalpighiata hypothesis, which shows little gene-tree incongruence and is robust to the influence of among-taxon compositional heterogeneity, implies convergent evolution in several morphological and behavioral characters traditionally used in centipede phylogenetics, such as maternal brood care, but accords with patterns of first appearances in the fossil record.

Lemer, S., B. Buge, A. Bemis, and G. Giribet. 2014. “First molecular phylogeny of the circumtropical bivalve family Pinnidae (Mollusca, Bivalvia): evidence for high levels of cryptic species diversity.” Mol Phylogenet Evol 75: 11-23.Abstract

The family Pinnidae Leach, 1819, includes approximately 50 species of large subtidal and coastal marine bivalves. These commercially important species occur in tropical and temperate waters around the world and are most frequently found in seagrass meadows. The taxonomy of the family has been revised a number of times since the early 20th Century, the most recent revision recognizing 55 species distributed in three genera: Pinna, Atrina and Streptopinna, the latter being monotypic. However, to date no phylogenetic analysis of the family has been conducted using morphological or molecular data. The present study analyzed 306 pinnid specimens from around the world, comprising the three described genera and ca. 25 morphospecies. We sequenced the mitochondrial genes 16S rRNA and cytochrome c oxidase subunit I, and the nuclear ribosomal genes 18S rRNA and 28S rRNA. Phylogenetic analysis of the data revealed monophyly of the genus Atrina but also that the genus Streptopinna is nested within Pinna. Based on the strong support for this relationship we propose a new status for Streptopinna Martens, 1880 and treat it as a subgenus (status nov.) of Pinna Linnaeus, 1758. The phylogeny and the species delimitation analyses suggest the presence of cryptic species in many morphospecies displaying a wide Indo-Pacific distribution, including Pinna muricata, Atrina assimilis, A. exusta and P. (Streptopinna) saccata but also in the Atlantic species A. rigida. Altogether our results highlight the challenges associated with morphological identifications in Pinnidae due to the presence of both phenotypic plasticity and morphological stasis and reveal that many pinnid species are not as widely distributed as previously thought.

Bracken-Grissom, H., A. G. Collins, T. Collins, K. Crandall, D. Distel, C. Dunn, G. Giribet, et al. 2014. “The Global Invertebrate Genomics Alliance (GIGA): developing community resources to study diverse invertebrate genomes.” J Hered 105: 1-18.Abstract

Over 95% of all metazoan (animal) species comprise the "invertebrates," but very few genomes from these organisms have been sequenced. We have, therefore, formed a "Global Invertebrate Genomics Alliance" (GIGA). Our intent is to build a collaborative network of diverse scientists to tackle major challenges (e.g., species selection, sample collection and storage, sequence assembly, annotation, analytical tools) associated with genome/transcriptome sequencing across a large taxonomic spectrum. We aim to promote standards that will facilitate comparative approaches to invertebrate genomics and collaborations across the international scientific community. Candidate study taxa include species from Porifera, Ctenophora, Cnidaria, Placozoa, Mollusca, Arthropoda, Echinodermata, Annelida, Bryozoa, and Platyhelminthes, among others. GIGA will target 7000 noninsect/nonnematode species, with an emphasis on marine taxa because of the unrivaled phyletic diversity in the oceans. Priorities for selecting invertebrates for sequencing will include, but are not restricted to, their phylogenetic placement; relevance to organismal, ecological, and conservation research; and their importance to fisheries and human health. We highlight benefits of sequencing both whole genomes (DNA) and transcriptomes and also suggest policies for genomic-level data access and sharing based on transparency and inclusiveness. The GIGA Web site (http://giga.nova.edu) has been launched to facilitate this collaborative venture.

Murienne, J., S. R. Daniels, T. R. Buckley, G. Mayer, and G. Giribet. 2014. “A living fossil tale of Pangaean biogeography.” Proc Biol Sci 281: 20132648.Abstract

The current distributions of widespread groups of terrestrial animals and plants are supposedly the result of a mixture of either vicariance owing to continental split or more recent trans-oceanic dispersal. For organisms exhibiting a vicariant biogeographic pattern-achieving their current distribution by riding on the plates of former supercontinents-this view is largely inspired by the belief that Pangaea lacked geographical or ecological barriers, or that extinctions and dispersal would have erased any biogeographic signal since the early Mesozoic. We here present a time-calibrated molecular phylogeny of Onychophora (velvet worms), an ancient and exclusively terrestrial panarthropod group distributed throughout former Pangaean landmasses. Our data not only demonstrate that trans-oceanic dispersal does not need be invoked to explain contemporary distributions, but also reveal that the early diversification of the group pre-dates the break-up of Pangaea, maintaining regionalization even in landmasses that have remained contiguous throughout the history of the group. These results corroborate a growing body of evidence from palaeontology, palaeogeography and palaeoclimatic modelling depicting ancient biogeographic regionalization over the continuous landmass of Pangaea.

Garwood, R. J., P. P. Sharma, J. A. Dunlop, and G. Giribet. 2014. “A Paleozoic stem group to mite harvestmen revealed through integration of phylogenetics and development.” Curr Biol 24: 1017-23.Abstract

Successfully placing fossils in phylogenies is integral to understanding the tree of life. Crown-group Paleozoic members of the arachnid order Opiliones are indicative of ancient origins and one of the earliest arthropod terrestrialization events [1, 2]. Opiliones epitomize morphological stasis, and all known fossils have been placed within the four extant suborders [3-5]. Here we report a Carboniferous harvestman species, Hastocularis argusgen. nov., sp. nov., reconstructed with microtomography (microCT). Phylogenetic analysis recovers this species, and the Devonian Eophalangium sheari, as members of an extinct harvestman clade. We establish the suborder Tetrophthalmi subordo nov., which bore four eyes, to accommodate H. argus and E. sheari, the latter previously considered to be a phalangid [6-9]. Furthermore, embryonic gene expression in the extant species Phalangium opilio demonstrates vestiges of lateral eye tubercles. These lateral eyes are lost in all crown-group Phalangida, but are observed in both our fossil and outgroup chelicerate orders. These data independently corroborate the diagnosis of two eye pairs in the fossil and demonstrate retention of eyes of separate evolutionary origins in modern harvestmen [10-12]. The discovery of Tetrophthalmi alters molecular divergence time estimates, supporting Carboniferous rather than Devonian diversification for extant suborders and directly impacting inferences of terrestrialization history and biogeography. Multidisciplinary approaches integrating fossil and neontological data increase confidence in phylogenies and elucidate evolutionary history.

Zapata, F., N. G. Wilson, M. Howison, S. C. Andrade, K. M. Jorger, M. Schrodl, F. E. Goetz, G. Giribet, and C. W. Dunn. 2014. “Phylogenomic analyses of deep gastropod relationships reject Orthogastropoda.” Proc Biol Sci 281: 20141739.Abstract

Gastropods are a highly diverse clade of molluscs that includes many familiar animals, such as limpets, snails, slugs and sea slugs. It is one of the most abundant groups of animals in the sea and the only molluscan lineage that has successfully colonized land. Yet the relationships among and within its constituent clades have remained in flux for over a century of morphological, anatomical and molecular study. Here, we re-evaluate gastropod phylogenetic relationships by collecting new transcriptome data for 40 species and analysing them in combination with publicly available genomes and transcriptomes. Our datasets include all five main gastropod clades: Patellogastropoda, Vetigastropoda, Neritimorpha, Caenogastropoda and Heterobranchia. We use two different methods to assign orthology, subsample each of these matrices into three increasingly dense subsets, and analyse all six of these supermatrices with two different models of molecular evolution. All 12 analyses yield the same unrooted network connecting the five major gastropod lineages. This reduces deep gastropod phylogeny to three alternative rooting hypotheses. These results reject the prevalent hypothesis of gastropod phylogeny, Orthogastropoda. Our dated tree is congruent with a possible end-Permian recovery of some gastropod clades, namely Caenogastropoda and some Heterobranchia subclades.

Fernandez, R., G. Hormiga, and G. Giribet. 2014. “Phylogenomic analysis of spiders reveals nonmonophyly of orb weavers.” Curr Biol 24: 1772-7.Abstract

Spiders constitute one of the most successful clades of terrestrial predators. Their extraordinary diversity, paralleled only by some insects and mites, is often attributed to the use of silk, and, in one of the largest lineages, to stereotyped behaviors for building foraging webs of remarkable biomechanical properties. However, our understanding of higher-level spider relationships is poor and is largely based on morphology. Prior molecular efforts have focused on a handful of genes but have provided little resolution to key questions such as the origin of the orb weavers. We apply a next-generation sequencing approach to resolve spider phylogeny, examining the relationships among its major lineages. We further explore possible pitfalls in phylogenomic reconstruction, including missing data, unequal rates of evolution, and others. Analyses of multiple data sets all agree on the basic structure of the spider tree and all reject the long-accepted monophyly of Orbiculariae, by placing the cribellate orb weavers (Deinopoidea) with other groups and not with the ecribellate orb weavers (Araneoidea). These results imply independent origins for the two types of orb webs (cribellate and ecribellate) or a much more ancestral origin of the orb web with subsequent loss in the so-called RTA clade. Either alternative demands a major reevaluation of our current understanding of the spider evolutionary chronicle.

Sharma, P. P., S. T. Kaluziak, A. R. Perez-Porro, V. L. Gonzalez, G. Hormiga, W.C. Wheeler, and G. Giribet. 2014. “Phylogenomic interrogation of arachnida reveals systemic conflicts in phylogenetic signal.” Mol Biol Evol 31: 2963-84.Abstract

Chelicerata represents one of the oldest groups of arthropods, with a fossil record extending to the Cambrian, and is sister group to the remaining extant arthropods, the mandibulates. Attempts to resolve the internal phylogeny of chelicerates have achieved little consensus, due to marked discord in both morphological and molecular hypotheses of chelicerate phylogeny. The monophyly of Arachnida, the terrestrial chelicerates, is generally accepted, but has garnered little support from molecular data, which have been limited either in breadth of taxonomic sampling or in depth of sequencing. To address the internal phylogeny of this group, we employed a phylogenomic approach, generating transcriptomic data for 17 species in combination with existing data, including two complete genomes. We analyzed multiple data sets containing up to 1,235,912 sites across 3,644 loci, using alternative approaches to optimization of matrix composition. Here, we show that phylogenetic signal for the monophyly of Arachnida is restricted to the 500 slowest-evolving genes in the data set. Accelerated evolutionary rates in Acariformes, Pseudoscorpiones, and Parasitiformes potentially engender long-branch attraction artifacts, yielding nonmonophyly of Arachnida with increasing support upon incrementing the number of concatenated genes. Mutually exclusive hypotheses are supported by locus groups of variable evolutionary rate, revealing significant conflicts in phylogenetic signal. Analyses of gene-tree discordance indicate marked incongruence in relationships among chelicerate orders, whereas derived relationships are demonstrably robust. Consistently recovered and supported relationships include the monophyly of Chelicerata, Euchelicerata, Tetrapulmonata, and all orders represented by multiple terminals. Relationships supported by subsets of slow-evolving genes include Ricinulei + Solifugae; a clade comprised of Ricinulei, Opiliones, and Solifugae; and a clade comprised of Tetrapulmonata, Scorpiones, and Pseudoscorpiones. We demonstrate that outgroup selection without regard for branch length distribution exacerbates long-branch attraction artifacts and does not mitigate gene-tree discordance, regardless of high gene representation for outgroups that are model organisms. Arachnopulmonata (new name) is proposed for the clade comprising Scorpiones + Tetrapulmonata (previously named Pulmonata).

Sharma, P. P., and G. Giribet. 2014. “A revised dated phylogeny of the arachnid order Opiliones.” Front Genet 5: 255.Abstract

Dating the Opiliones tree of life has become an important enterprise for this group of arthropods, due to their ancient origins and important biogeographic implications. To incorporate both methodological innovations in molecular dating as well as new systematic discoveries of harvestman diversity, we conducted total evidence dating on a data set uniting morphological and/or molecular sequence data for 47 Opiliones species, including all four well-known Palaeozoic fossils, to test the placement of both fossils and newly discovered lineages in a single analysis. Furthermore, we investigated node dating with a phylogenomic data set of 24,202 amino acid sites for 14 species of Opiliones, sampling all extant suborders. In this way, we approached molecular dating of basal harvestman phylogeny using different data sets and approaches to assess congruence of divergence time estimates. In spite of the markedly different composition of data sets, our results show congruence across all analyses for age estimates of basal nodes that are well constrained with respect to fossil calibrations (e.g., Opiliones, Palpatores). By contrast, derived nodes that lack fossil calibrations (e.g., the suborders Cyphophthalmi, and Laniatores) have large uncertainty intervals in diversification times, particularly in the total evidence dating analysis, reflecting the dearth of calibration points and undersampling of derived lineages. Total evidence dating consistently produced older median ages than node dating for ingroup nodes, due to the nested placement of multiple Palaeozoic fossils. Our analyses support basal diversification of Opiliones in the Ordovician-Devonian period, corroborating the inferred ancient origins of this arthropod order, and underscore the importance of diversity discovery-both paleontological and neontological-in evolutionary inference.

Fernandez, R., S. Kvist, J. Lenihan, G. Giribet, and A. Ziegler. 2014. “Sine systemate chaos? A versatile tool for earthworm taxonomy: non-destructive imaging of freshly fixed and museum specimens using micro-computed tomography.” PLoS One 9: e96617.Abstract

In spite of the high relevance of lumbricid earthworms ('Oligochaeta': Lumbricidae) for soil structure and functioning, the taxonomy of this group of terrestrial invertebrates remains in a quasi-chaotic state. Earthworm taxonomy traditionally relies on the interpretation of external and internal morphological characters, but the acquisition of these data is often hampered by tedious dissections or restricted access to valuable and rare museum specimens. The present state of affairs, in conjunction with the difficulty of establishing primary homologies for multiple morphological features, has led to an almost unrivaled instability in the taxonomy and systematics of certain earthworm groups, including Lumbricidae. As a potential remedy, we apply for the first time a non-destructive imaging technique to lumbricids and explore the future application of this approach to earthworm taxonomy. High-resolution micro-computed tomography (muCT) scanning of freshly fixed and museum specimens was carried out using two cosmopolitan species, Aporrectodea caliginosa and A. trapezoides. By combining two-dimensional and three-dimensional dataset visualization techniques, we demonstrate that the morphological features commonly used in earthworm taxonomy can now be analyzed without the need for dissection, whether freshly fixed or museum specimens collected more than 60 years ago are studied. Our analyses show that muCT in combination with soft tissue staining can be successfully applied to lumbricid earthworms. An extension of the approach to other families is poised to strengthen earthworm taxonomy by providing a versatile tool to resolve the taxonomic chaos currently present in this ecologically important, but taxonomically neglected group of terrestrial invertebrates.

Andrade, S. C., H. Montenegro, M. Strand, M. L. Schwartz, H. Kajihara, J. L. Norenburg, J. M. Turbeville, P. Sundberg, and G. Giribet. 2014. “A transcriptomic approach to ribbon worm systematics (nemertea): resolving the pilidiophora problem.” Mol Biol Evol 31: 3206-15.Abstract

Resolving the deep relationships of ancient animal lineages has proven difficult using standard Sanger-sequencing approaches with a handful of markers. We thus reassess the relatively well-studied phylogeny of the phylum Nemertea (ribbon worms)-for which the targeted gene approaches had resolved many clades but had left key phylogenetic gaps-by using a phylogenomic approach using Illumina-based de novo assembled transcriptomes and automatic orthology prediction methods. The analysis of a concatenated data set of 2,779 genes (411,138 amino acids) with about 78% gene occupancy and a reduced version with 95% gene occupancy, under evolutionary models accounting or not for site-specific amino acid replacement patterns results in a well-supported phylogeny that recovers all major accepted nemertean clades with the monophyly of Heteronemertea, Hoplonemertea, Monostilifera, being well supported. Significantly, all the ambiguous patterns inferred from Sanger-based approaches were resolved, namely the monophyly of Palaeonemertea and Pilidiophora. By testing for possible conflict in the analyzed supermatrix, we observed that concatenation was the best solution, and the results of the analyses should settle prior debates on nemertean phylogeny. The study highlights the importance, feasibility, and completeness of Illumina-based phylogenomic data matrices.

2013
Bieler, R., Mikkelsen. PM, and Giribet. G. 2013. “Bivalvia-A discussion of known unknowns.” American Malacological Bulletin 31: 123-133.
Giribet, G. 2013. “Book Review: Animal Evolution: Interrelationships of the Living Phyla. 3rd ed..” Integrative and Comparative Biology 53: 532-534.
Guil, N., Jorgensen. A, Giribet. G, and Kristensen. RM. 2013. “Congruence between molecular phylogeny and cuticular design in Echiniscoidea (Tardigrada, Heterotardigrada).” Zoological Journal of the Linnean Society 169: 713-736.
Smith, SA., Wilson. NG, Goetz. FE, Feehery. C, Andrade. SCS, Rouse. GW, Giribet. G, and Dunn. CW. 2013. “Corrigendum: Resolving the evolutionary relationships of molluscs with phylogenomic tools.” Nature 493: 708.
Salvatierra, L., Tourinho. AL, and Giribet. G. 2013. “Description of the male, larva and nymphal stages of Crypocellus iaci (Arachnida, Ricinulei), with an overview of tarsal sensilla and other integumental structures.” Zootaxa 3709: 149-161.
Taboada, S., Junoy. J, Andrade. SCS, Giribet. G, Cristobo. J, and Avila. C. 2013. “On the identity of two Antarctic brooding nemerteans: redescription of Antarctonemertes valida (Burger, 1893) and description of a new species in the genus Antarctonemertes Friedrich, 1995 (Nemertea, Hoplonemertea).” Polar Biology 36: 1415-1430.
Garwood, RJ., Dunlop. JA, Giribet. G, and Sutton. MD. 2013. “Opiliones fosiles. Los aracnidos actuales de origen mas remoto.” Fundamental! 23: 1-58.
Vahtera, V., Edgecombe. GD, and Giribet. G. 2013. “Phylogenetics.” Invertebrate Systematics 27: 578-602.

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