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SDS-PAGE of reduced G. tridactyla venom. Figure S2. Genomic organization of the glycerotoxin gene analyzed in G. tridactyla. Figure S3. Immunolocalization of GLTx in a cross section through a putative venom gland embedded in paraffin. Figure S4. Anti-serotonin (5-HT) staining and phalloidinâ rhodamine counterstaining on everted G. tridactyla pharynges cut into two halves. Figure S5. Quantitative real-time PCR (qPCR) expression levels (shown as Fold Change, RQ) between biological groups (putative venom glands [A], pharyngeal lobes [B], and posterior body wall [C]) per analyzed specimen (nâ =â 10, biological samples). Figure S6. Tissues analyzed in comparative GLTx expression studies (qPCR experiments and transcriptome analyses) on G. tridactyla. (PDF 2931 kb)

Protocol S1â S3. Protocol S1. Protocol: In situ hybridization on pharynx tissue of G. tridactyla (Glyceridae, Annelida). Protocol S2. Protocol: Anti-GLTx staining on pharynx tissue of G. tridactyla (Glyceridae, Annelida). Protocol S3. Protocol: Fluorescence in situ hybridization (FISH) coupled with antibody staining against GLTx on pharynx tissue of G. tridactyla. (PDF 590 kb)

Animal venoms have evolved many times. Venomous species are especially common in three of the four main groups of arthropods (Chelicerata, Myriapoda, and Hexapoda), which together represent tens of thousands of species of venomous spiders, scorpions, centipedes, and hymenopterans. Surprisingly, despite their great diversity of body plans, there is no unambiguous evidence that any crustacean is venomous. We provide the first conclusive evidence that the aquatic, blind, and cave-dwelling remipede crustaceans are venomous and that venoms evolved in all four major arthropod groups. We produced a three-dimensional reconstruction of the venom delivery apparatus of the remipede Speleonectes tulumensis, showing that remipedes can inject venom in a controlled manner. A transcriptomic profile of its venom glands shows that they express a unique cocktail of transcripts coding for known venom toxins, including a diversity of enzymes and a probable paralytic neurotoxin very similar to one described from spider venom. We screened a transcriptomic library obtained from whole animals and identified a nontoxin paralog of the remipede neurotoxin that is not expressed in the venom glands. This allowed us to reconstruct its probable evolutionary origin and underlines the importance of incorporating data derived from nonvenom gland tissue to elucidate the evolution of candidate venom proteins. This first glimpse into the venom of a crustacean and primitively aquatic arthropod reveals conspicuous differences from the venoms of other predatory arthropods such as centipedes, scorpions, and spiders and contributes valuable information for ultimately disentangling the many factors shaping the biology and evolution of venoms and venomous species., Molecular Biology and Evolution, 31 (1), ISSN:0737-4038, ISSN:1537-1719

There is no consensus on the inter-ordinal relationships of eumalacostracans, despite the recent synthesis of several morphological matrices with data from four molecular markers. Signals from different molecules confl ict with each other, and all are conspicuously at odds with morphology. Can fossils help to resolve the problem? Here, we utilize palaeontological data in two ways. Firstly we coded a selection of fossil taxa into our morphological matrix, and assessed their impact upon inferred phylogeny relative to that of their living counterparts (fi rst order jackknifing). This revealed that our morphological tree is very sensitive to the precise taxon sample (a problem that must be addressed in future studies), but that our fossil groups were not disproportionately infl uential. Secondly, we asked whether the order in which groups appear in the fossil record provides a means to choose between competing trees. The congruence between morphological and stratigraphic signals was extremely weak and non-signifi cant in most cases, precluding the use of fossil dates in this way. Many trees imply ghost ranges of duration near the theoretical maximum, and worse than for the majority of other animal groups so far investigated. An incomplete fossil record and fragile/weakly-supported trees combine with considerable molecular rate heterogeneity to make the Eumalacostraca extremely poorly suited to molecular clock studies. Future insights into their phylogeny are likely to come from the development of new molecular markers, as well as hard-won data on internal anatomy and ultrastructure.

Review of and commentary on: Animal Evolution. Interrelationships of the Living Phyla, by Claus Nielsen. Oxford University Press, 2001, x + 563 pp., ISBN 0-19-850682-1 (Pbk). My initial browsing of the new edition of Claus Nielsen’s Animal Evolution could scarcely have produced a bigger surprise. Opening the book from the back I first read the last line of the last chapter on molecular phylogeny: “The evolutionary scenario pictured in fig. 57.2 [a molecular cladogram based on 18S rDNA sequences] is fully compatible with the phylogeny proposed on morphological evidence in this book...” (p. 519). Imagine the surprise! Anyone even marginally familiar with recent developments in higher-level animal phylogenetics will be aware of some major conflicting phylogenetic signals lurking below the surface of morphological and molecular data sets. Certainly, many may agree with Balter’s (1997) observation that “morphologists learn to live with molecular upstarts,” but in view of Nielsen’s less than enthusiastic embrace of molecular phylogenetic evidence in previous publications (Nielsen, 1997a, b) he appears particularly unlikely to serve as a candidate molecular convert. However, a glance at Nielsen’s molecular ‘tree’ offers immediate clarification. It resembles a three-layered terrace overgrown with a phylogenetic grass reminiscent of the massively polyphyletic scheme advocated a decade ago by Willmer(1990), which enjoyed some notoriety as the then most recent pictorial incarnation of phylogenetic ignorance. Nielsen’s rendition of IBS rDNA sequence data merely yields three phylogenetic plateaus representing the ascending organizational levels of Metazoa, Bilateria, and Deuterostomia forming the substrates of three successive Precambrian radiations. Unsurprisingly, the complete lack of resolution in the molecular phylogeny-guarantees compatibility with any morphological tree. Needless to say, others have reached rather different conclusions about the degree of phylogenetic resolution that 18S data can offer. I