YMPEV 4919 No. of Pages 14, Model 5G 2 June 2014 Molecular Phylogenetics and Evolution xxx (2014) xxx–xxx 1 Contents lists available at ScienceDirect Molecular Phylogenetics and Evolution journal homepage: www.elsevier.com/locate/ympev 5 6 3 Molecular phylogeny and evolution of the cone snails 4 (Gastropoda, Conoidea) a,⇑ b c,d e f g h 7 Q1 N. Puillandre , P. Bouchet , T.F. Duda , S. Kauferstein , A.J. Kohn , B.M. Olivera , M. Watkins , i 8 C. Meyer 9 a Muséum National d’Histoire Naturelle, Département Systématique et Evolution, ISyEB Institut (UMR 7205 CNRS/UPMC/MNHN/EPHE), 43, Rue Cuvier, 75231 Paris, France 10 b Muséum National d’Histoire Naturelle, Département Systématique et Evolution, ISyEB Institut (UMR 7205 CNRS/UPMC/MNHN/EPHE), 55, Rue Buffon, 75231 Paris, France 11 Q2 c Department of Ecology and Evolutionary Biology and Museum of Zoology, University of Michigan, 1109 Geddes Avenue, Ann Arbor, MI 48109, USA 12 d Smithsonian Tropical Research Institute, Apartado 0843-03092, Balboa, Ancon, Panama 13 e Institute of Legal Medicine, University of Frankfurt, Kennedyallee 104, D-60596 Frankfurt, Germany 14 f Department of Biology, Box 351800, University of Washington, Seattle, WA 98195, USA 15 g Department of Biology, University of Utah, 257 South 1400 East, Salt Lake City, UT 84112, USA 16 h Department of Pathology, University of Utah, 257 South 1400 East, Salt Lake City, UT 84112, USA 17 i Department of Invertebrate Zoology, National Museum of Natural History, Smithsonian Institution, Washington, DC 20013, USA 1819 20 article info abstract 3622 23 Article history: We present a large-scale molecular phylogeny that includes 320 of the 761 recognized valid species of the 37 24 Received 22 January 2014 cone snails (Conus), one of the most diverse groups of marine molluscs, based on three mitochondrial 38 25 Revised 8 May 2014 genes (COI, 16S rDNA and 12S rDNA). This is the first phylogeny of the taxon to employ concatenated 39 26 Accepted 16 May 2014 sequences of several genes, and it includes more than twice as many species as the last published molec- 40 27 Available online xxxx ular phylogeny of the entire group nearly a decade ago. Most of the numerous molecular phylogenies 41 published during the last 15 years are limited to rather small fractions of its species diversity. Bayesian 42 28 Keywords: and maximum likelihood analyses are mostly congruent and confirm the presence of three previously 43 29 Ancestral state reconstruction reported highly divergent lineages among cone snails, and one identified here using molecular data. 44 30 Conidae 31 Conus About 85% of the species cluster in the single Large Major Clade; the others are divided between the Small 45 32 COI Major Clade (12%), the Conus californicus lineage (one species), and a newly defined clade (3%). We also 46 33 16SrRNA define several subclades within the Large and Small major clades, but most of their relationships remain 47 34 12SrRNA poorly supported. To illustrate the usefulness of molecular phylogenies in addressing specific evolution- 48 35 ary questions, we analyse the evolution of the diet, the biogeography and the toxins of cone snails. All 49 cone snails whose feeding biology is known inject venom into large prey animals and swallow them 50 whole. Predation on polychaete worms is inferred as the ancestral state, and diet shifts to molluscs 51 and fishes occurred rarely. The ancestor of cone snails probably originated from the Indo-Pacific; rather 52 few colonisations of other biogeographic provinces have probably occurred. A new classification of the 53 Conidae, based on the molecular phylogeny, is published in an accompanying paper. 54 Ó 2014 Elsevier Inc. All rights reserved. 55 56 57 58 59 1. Introduction tree can help to estimate diversification rates, divergence times, 63 ancestral distributions, and community compositions, and it can 64 60 A molecular phylogeny of a taxon is a hypothesis of its evolu- provide evidence relevant to taxonomic hypotheses. However, 65 61 tionary patterns and processes, and a framework for clarifying its many taxa of considerable evolutionary and practical importance 66 62 classification. A strongly supported molecular-based phylogenetic have very incomplete species-level molecular phylogenies, based 67 on few species with appropriate genes sequenced, not representa- 68 tive of the diversity of the group, or largely unresolved. 69 ⇑ Corresponding author. Address: Puillandre Nicolas, Muséum National d’Histoire The gastropod family Conidae, commonly known as cone snails, 70 Naturelle, Département Systématique et Evolution, ISyEB Institut (UMR 7205 CNRS/ includes the widely distributed, mainly tropical Conus, a relatively 71 UPMC/MNHN/EPHE), 43, Rue Cuvier, 75231 Paris, France. Fax: +33 1 40 79 38 44. young genus first appearing in the Early Eocene. The family Coni- 72 E-mail addresses: [email protected] (N. Puillandre), [email protected] dae is one of the most diverse in the marine environment (Kohn, 73 (P. Bouchet), [email protected] (T.F. Duda), [email protected] (S. Kauferstein), [email protected] (A.J. Kohn), [email protected] (B.M. Olivera), 1990), with 761 valid Recent species currently (21th January 74 [email protected] (M. Watkins), [email protected] (C. Meyer). 2014) recognized in the World Register of Marine Species (WoRMS, 75 http://dx.doi.org/10.1016/j.ympev.2014.05.023 1055-7903/Ó 2014 Elsevier Inc. All rights reserved. Please cite this article in press as: Puillandre, N., et al. Molecular phylogeny and evolution of the cone snails (Gastropoda, Conoidea). Mol. Phylogenet. Evol. (2014), http://dx.doi.org/10.1016/j.ympev.2014.05.023 YMPEV 4919 No. of Pages 14, Model 5G 2 June 2014 2 N. Puillandre et al. / Molecular Phylogenetics and Evolution xxx (2014) xxx–xxx 76 2013) and new species descriptions are published each year. It is species exhibiting more than one feeding mode. In addition, diets 142 77 also the most rapidly diversifying marine molluscan genus (Kohn, tend to be species-specific, especially in areas where multiple spe- 143 78 1990; Stanley, 2008, 1979) and is ecologically important especially cies co-occur (Kohn and Nybakken, 1975; Kohn, 1968, 1959). A 144 79 in coral reef environments where up to 36 species, specialized pre- previous investigation of the evolution of diets of cone snails 145 80 dators on worms, other molluscs, and fishes, co-occur on a single reports that major shifts in diet were relatively rare (Duda et al., 146 81 reef (Kohn, 2001). 2001), although piscivory originated at least twice (Duda and 147 82 These latter attributes all likely relate to the extremely diverse Palumbi, 2004). However, as with all past molecular phylogenetic 148 83 peptide venoms that cone snails use to overcome and capture prey studies of this group, these studies relied on limited taxonomic 149 84 and that also make the Conidae a most promising source for neu- coverage. Analyses of a much larger dataset may provide additional 150 85 robiologic and therapeutic applications (Biass et al., 2009; insights of the evolution of diet that were not available previously. 151 86 Lluisma et al., 2012; Olivera, 2006). Molecular geneticists, evolu- We propose here a molecular phylogeny of the Conidae sensu 152 87 tionary biologists, pharmacologists, and toxicologists thus all Bouchet et al. (2011), based on three mitochondrial genes (COI, 153 88 require a robust phylogeny and taxonomy for this group. New drug 12S, 16S) sequenced for 320 species (>40% of the known species 154 89 discovery is particularly likely to benefit from a clear phylogenetic diversity), and including representatives from the main lineages 155 90 context that permits targeting divergent lineages and thus poten- defined in previous DNA studies: C. californicus, the Small Major 156 91 tial novel toxins (Biggs et al., 2010; Olivera, 2006). Clade and the Large Major Clade (Duda and Kohn, 2005). Tucker 157 92 Since the first published molecular phylogenies for Conus (Duda and Tenorio (2009) classified the Small Major Clade as the Family 158 93 and Palumbi, 1999a; Monje et al., 1999), many others have Conilithidae – it included C. californicus – and the Large Major 159 94 appeared, either for the cone snails and their relatives (Puillandre Clade as the family Conidae (see Table 1 for a comparison of the 160 95 et al., 2011a, 2008), or subgroups (Bandyopadhyay et al., 2008; recent classifications of cone snails and related species). We then 161 96 Biggs et al., 2010; Cunha et al., 2008, 2005; Duda and Kohn, analyse the evolution of three character sets: diet category, biogeo- 162 97 2005; Duda and Palumbi, 2004, 1999b; Duda and Rolán, 2005; graphic province and toxin diversity. Previous molecular phyloge- 163 98 Duda et al., 2008, 2001; Espino et al., 2008; Espiritu et al., 2001; netic studies analysed the main evolutionary diet shifts (from 164 99 Kauferstein et al., 2011, 2004; Kraus et al., 2012, 2011; Nam worms to fishes or molluscs) (Duda and Kohn, 2005; Duda and 165 100 et al., 2009; Pereira et al., 2010; Puillandre et al., 2010; Williams Palumbi, 2004; Duda et al., 2001), but never on such a large data- 166 101 and Duda, 2008). The most comprehensive includes 138 species, set. Disentangling the evolution of these traits throughout this 167 102 ca. 20% of the known diversity of cone snails (Duda and Kohn, hyperdiverse taxon should help to generate and critically examine 168 103 2005). hypotheses of the factors that promoted its exceptional ecological 169 104 Ancestral states of morphological, ecological, and developmen- and evolutionary diversification. 170 105 tal traits have been inferred from some of these phylogenetic stud- 106 ies (Cunha et al., 2005; Duda and Palumbi, 2004, 1999a; Duda et al., 2. Material and methods 171 107 2001; Kohn, 2012) and lineages of toxins with unknown functions 108 identified (Puillandre et al., 2010).