Species delimitation in poorly known and diverse taxa is usually performed based on monolocus, DNA-­barcoding-­like approaches, while multilocus data are often used to test alternative species hypotheses in well-­studied groups. We combined both approaches to delimit species in the Xenuroturris/Iotyrris complex, a group of venomous marine gastropods from the Indo-P­ acific. First, COI sequences were analysed using three methods of species delimitation to propose primary species hypotheses. Second, RAD sequencing data were also obtained and a maximum-l­ikelihood phylogenetic tree produced. We tested the impact of the level of missing data on the robustness of the phylogenetic tree obtained with the RAD-­seq data. Alternative species partitions revealed with the COI data set were also tested using the RAD-­seq data and the Bayes factor species delimitation method. The congruence between the species hypotheses proposed with the mitochondrial nuclear data sets, together with the morphological variability of the shell and the radula and the distribution pattern, was used to turn the primary species hypotheses into secondary species hypotheses. Allopatric primary species hypotheses defined with the COI gene were interpreted to correspond to intraspecific structure. Most of the species are found sympatrically in the Philippines, and only one is confidently identified as a new species and described as Iotyrris conotaxis n. sp. The results obtained demonstrate the efficiency of the combined monolocus/multilocus approach to delimit species.

ATIMO VATAE, INHACA 2011, MAINBAZA, PAKAIHI I TE MOANA, PAPUA NIUGINI, SANTO 2006, TERRASSES

Transcriptome-based exon capture methods provide an approach to recover several hundred markers from genomic DNA, allowing for robust phylogenetic estimation at deep timescales. We applied this method to a highly diverse group of venomous marine snails, Conoidea, for which published phylogenetic trees remain mostly unresolved for the deeper nodes. We targeted 850 protein coding genes (678,322 bp) in ca. 120 samples, spanning all (except one) known families of Conoidea and a broad selection of non-Conoidea neogastropods. The capture was successful for most samples, although capture efficiency decreased when DNA libraries were of insufficient quality and/or quantity (dried samples or low starting DNA concentration) and when targeting the most divergent lineages. An average of 75.4% of proteins was recovered, and the resulting tree, reconstructed using both supermatrix (IQ-tree) and supertree (Astral-II, combined with the Weighted Statistical Binning method) approaches, are almost fully supported. A reconstructed fossil-calibrated tree dates the origin of Conoidea to the Lower Cretaceous. We provide descriptions for two new families. The phylogeny revealed in this study provides a robust framework to reinterpret changes in Conoidea anatomy through time. Finally, we used the phylogeny to test the impact of the venom gland and radular type on diversification rates. Our analyses revealed that repeated losses of the venom gland had no effect on diversification rates, while families with a breadth of radula types showed increases in diversification rates, thus suggesting that trophic ecology may have an impact on the evolution of Conoidea.

ATIMO VATAE, AURORA 2007, BIOPAPUA, CEAMARC-AA, CONCALIS, ???msg.security.private???, DongSha 2014, EXBODI, GUYANE 2014, ILES DU SALUT, INHACA 2011, KARUBENTHOS 2012, KAVIENG 2014, MAINBAZA, NORFOLK 2, NanHai 2014, PANGLAO 2005, PAPUA NIUGINI, ???msg.security.private???, SALOMONBOA 3, TAIWAN 2013, TERRASSES, ???msg.security.private???

Profundiconus is the most divergent cone snail genus and its unique phylogenetic position, sister to the rest of the family Conidae, makes it a key taxon for examining venom evolution and diversity. Venom gland and foot transcriptomes of Profundiconus cf. vaubani and Profundiconus neocaledonicus were de novo assembled, annotated, and analyzed for differential expression. One hundred and thirty-seven venom components were identified from P. cf. vaubani and 82 from P. neocaledonicus, with only four shared by both species. The majority of the transcript diversity was composed of putative peptides, including conotoxins, profunditoxins, turripeptides, insulin, and prohormone-4. However, there were also a significant percentage of other putative venom components such as chymotrypsin and L-rhamnose-binding lectin. The large majority of conotoxins appeared to be from new gene superfamilies, three of which are highly different from previously reported venom peptide toxins. Their low conotoxin diversity and the type of insulin found suggested that these species, for which no ecological information are available, have a worm or molluscan diet associated with a narrow dietary breadth. Our results indicate that Profundiconus venom is highly distinct from that of other cone snails, and therefore important for examining venom evolution in the Conidae family.

KANACONO

Marine gastropods of the genus Conus are renowned for their remarkable diversity and deadly venoms. While Conus venoms are increasingly well studied for their biomedical applications, we know surprisingly little about venom composition in other lineages of Conidae. We performed comprehensive venom transcriptomic profiling for Conasprella coriolisi and Pygmaeconus traillii, first time for both respective genera. We complemented referencebased transcriptome annotation by a de novo toxin prediction guided by phylogeny, which involved transcriptomic data on two additional ‘divergent’ cone snail lineages, Profundiconus, and Californiconus. We identified toxin clusters (SSCs) shared among all or some of the four analysed genera based on the identity of the signal region—a molecular tag present in toxins. In total, 116 and 98 putative toxins represent 29 and 28 toxin gene superfamilies in Conasprella and Pygmaeconus, respectively; about quarter of these only found by semi-manual annotation of the SSCs. Two rare gene superfamilies, originally identified from fish-hunting cone snails, were detected outside Conus rather unexpectedly, so we further investigated their distribution across Conidae radiation. We demonstrate that both these, in fact, are ubiquitous in Conidae, sometimes with extremely high expression. Our findings demonstrate how a phylogeny-aware approach circumvents methodological caveats of similarity-based transcriptome annotation

KANACONO, KAVIENG 2014

Abstract The molluskan order Neogastropoda encompasses over 15,000 almost exclusively marine species playing important roles in benthic communities and in the economies of coastal countries. Neogastropoda underwent intensive cladogenesis in the early stages of diversification, generating a “bush” at the base of their evolutionary tree, which has been hard to resolve even with high throughput molecular data. In the present study to resolve the bush, we use a variety of phylogenetic inference methods and a comprehensive exon capture dataset of 1817 loci (79.6% data occupancy) comprising 112 taxa of 48 out of 60 Neogastropoda families. Our results show consistent topologies and high support in all analyses at (super)family level, supporting monophyly of Muricoidea, Mitroidea, Conoidea, and, with some reservations, Olivoidea and Buccinoidea. Volutoidea and Turbinelloidea as currently circumscribed are clearly paraphyletic. Despite our analyses consistently resolving most backbone nodes, 3 prove problematic: First, the uncertain placement of Cancellariidae, as the sister group to either a Ficoidea-Tonnoidea clade or to the rest of Neogastropoda, leaves monophyly of Neogastropoda unresolved. Second, relationships are contradictory at the base of the major “core Neogastropoda” grouping. Third, coalescence-based analyses reject monophyly of the Buccinoidea in relation to Vasidae. We analyzed phylogenetic signal of targeted loci in relation to potential biases, and we propose the most probable resolutions in the latter 2 recalcitrant nodes. The uncertain placement of Cancellariidae may be explained by orthology violations due to differential paralog loss shortly after the whole genome duplication, which should be resolved with a curated set of longer loci.

ATIMO VATAE, AURORA 2007, BIOMAGLO, BIOPAPUA, CORSICABENTHOS 1, DongSha 2014, EXBODI, ILES DU SALUT, INHACA 2011, KANACONO, KANADEEP, KARUBENTHOS 2, KAVIENG 2014, KOUMAC 2.1, KOUMAC 2.3, MADEEP, MADIBENTHOS, MAINBAZA, NanHai 2014, PAPUA NIUGINI, TAIWAN 2013, Tuhaa Pae 2013, ZhongSha 2015

ABSTRACT The family Turridae, once considered to encompass the entire diversity of Conoidea, excluding only the cone shells (family Conidae) and the auger shells (family Terebridae), has undergone significant redefinition based on both morphological and molecular data. Prior to this study, it included 15 Recent genera, comprising 209 species. However, a recent comprehensive analysis of an extensive dataset featuring 3,159 cytochrome c oxidase subunit I sequences has unveiled numerous species complexes, new species and previously undiscovered deep lineages. The newly proposed classification of Turridae based on an exon-capture phylogeny recognizes 24 clades, each worthy of genus rank, though only 13 of them had available names. As a result, 11 new genera have been described, namely Thielesyrinx n. gen., Shutogemmula n. gen., Mcleanigemmula n. gen., Pseudogemmula n. gen., Deceptigemmula n. gen., Oliveragemmula n. gen., Powelligemmula n. gen., Kilburnigemmula n. gen., Alisigemmula n. gen., Taylorigemmula n. gen. and Anisogemmula n. gen. With the exception of Thielesyrinx n. gen., species from all of these were previously classified within Gemmula, or Lophiotoma. Notably, some new genera, such as Deceptigemmula n. gen., Kilburnigemmula n. gen., Shutogemmula n. gen. and Taylorigemmula n. gen., cannot be morphologically differentiated, as similar species are found in other genera. Therefore, their distinction relies solely on molecular data. The name Eugemmula is re-instated from the synonymy of Gemmula. Purpuraturris is synonimized with Annulaturris, while Iotyrris with Xenuroturris. In total, we recognize 312 species/primary species hypotheses (PSHs) within the family. Molecular data were obtained for 212 species (among them 15 species complexes comprising in total 45 PSHs), mostly for the first time. A characteristic feature common to the entire family is the position of the relatively narrow anal sinus on the whorl periphery or immediately above it. Radula was examined in representatives of all genera. Although in several cases the radular morphology can aid genus differentiation (e.g. distinguishing Turris from Annulaturris), it is mostly a highly homoplastic structure, with very similar radular types occurring in distantly related genera.

ATIMO VATAE, AURORA 2007, BIOMAGLO, BIOPAPUA, BOA1, CONCALIS, DongSha 2014, EBISCO, EXBODI, GUYANE 2014, INHACA 2011, KANACONO, KANADEEP, KARUBENTHOS 2, KARUBENTHOS 2012, KOUMAC 2.1, MADEEP, MADIBENTHOS, MAINBAZA, MUSORSTOM 9, NORFOLK 2, NanHai 2014, PAKAIHI I TE MOANA, PANGLAO 2004, PANGLAO 2005, PAPUA NIUGINI, SALOMON 2, SALOMONBOA 3, TAIWAN 2004, TAIWAN 2013, TARASOC, TERRASSES, ZhongSha 2015

KANACONO

The exon capture approach allows for sequencing a large number of loci to reconstruct phylogenetic relationships at varying taxonomic levels. In order to efficiently recover the targeted loci, the probes designed to capture the exons need to be genetically sufficiently similar to bind to the DNA, with a proposed limit of 10% of divergence. However, this threshold has never been tested with a specific protocol. We have designed a set of probes to capture 1125 exons in the Neogastropoda (Mollusca, Gastropoda), processed with the same protocol from the field to the DNA sequencing to control for potential bias in DNA quantity and quality. We sequenced 30 different species, including 14 species of Neogastropoda and 16 species of Caenogastropoda non-­Neogastropoda. Each species includes five specimens, for a total of 150 specimens, and for four specimens among the 150, DNA extracts were aliquoted in four samples, sequenced separately, to estimate the intraspecific and intraspecimen variability in capture success. Our results confirm the impact of genetic distance on the success of exon capture with a negative linear correlation between the genetic distance and the number of exons captured for each sample. Consequently, designing new capture probes would allow for capturing exons in genetically more distant groups without the need to redesign a new set of exons.

CORSICABENTHOS 3

AURORA 2007, INHACA 2011, KAVIENG 2014, PANGLAO 2004, PAPUA NIUGINI, SANTO 2006

For over a decade now, High Throughput sequencing (HTS) approaches have revolutionized phylogenetics, both in terms of data production and methodology. While transcriptomes and (reduced) genomes are increasingly used, generating and analyzing HTS datasets remain expensive, time consuming and complex for most nonmodel taxa. Indeed, a literature survey revealed that 74% of the molecular phylogenetics trees published in 2018 are based on data obtained through Sanger sequencing. In this context, our goal was to identify the strategy that would represent the best compromise among costs, time and robustness of the resulting tree. We sequenced and assembled 32 transcriptomes of the marine mollusk family Turridae, considered as a typical non-model animal taxon. From these data, we extracted the loci most commonly used in gastropod phylogenies (cox1, 12S, 16S, 28S, h3 and 18S), full mitogenomes, and a reduced nuclear transcriptome representation. With each dataset, we reconstructed phylogenies and compared their robustness and accuracy. We discuss the impact of missing data and the use of statistical tests, tree metrics, and supertree and supermatrix methods to further improve phylogenetic data acquisition pipelines. We evaluated the overall costs (time and money) in order to identify the best compromise for phylogenetic data sampling in non-model animal taxa. Although sequencing full mitogenomes seems to constitute the best compromise both in terms of costs and node support, they are known to induce biases in phylogenetic reconstructions. Rather, we recommend to systematically include loci commonly used for phylogenetics and taxonomy (i.e. DNA barcodes, rRNA genes, full mitogenomes, etc.) among the other loci when designing baits for capture.

KANACONO, KAVIENG 2014

Abstract The practice of species delimitation using molecular data commonly leads to the revealing of species complexes and an increase in the number of delimited species. In a few instances, however, DNA-based taxonomy has led to lumping together of previously described species. Here, we delimit species in the genus Cryptogemma (Gastropoda: Conoidea: Turridae), a group of deep-sea snails with a wide geographical distribution, primarily by using the mitochondrial COI gene. Three approaches of species delimitation (ABGD, mPTP and GMYC) were applied to define species partitions. All approaches resulted in eight species. According to previous taxonomic studies and shell morphology, 23 available names potentially apply to the eight Cryptogemma species that were recognized herein. Shell morphometrics, radular characters and geographical and bathymetric distributions were used to link type specimens to these delimited species. In all, 23 of these available names are here attributed to seven species, resulting in 16 synonymizations, and one species is described as new: Cryptogemma powelli sp. nov. We discuss the possible reasons underlying the apparent overdescription of species within Cryptogemma, which is shown here to constitute a rare case of DNA-based species lumping in the hyper-diversified superfamily Conoidea.

ATIMO VATAE, AURORA 2007, BIOMAGLO, BIOPAPUA, CONCALIS, DongSha 2014, EBISCO, EXBODI, GUYANE 2014, KANACONO, KANADEEP, KAVIENG 2014, MADEEP, MAINBAZA, MIRIKY, NORFOLK 2, NanHai 2014, PANGLAO 2004, PAPUA NIUGINI, SALOMON 2, SALOMONBOA 3, TAIWAN 2013, TARASOC, TERRASSES, ZhongSha 2015

ATIMO VATAE, AURORA 2007, BIOMAGLO, BIOPAPUA, BOA1, CONCALIS, CORSICABENTHOS 1, DongSha 2014, EBISCO, EXBODI, GUYANE 2014, INHACA 2011, KANACONO, KANADEEP, KARUBENTHOS 2, KARUBENTHOS 2012, KAVIENG 2014, KOUMAC 2.1, KOUMAC 2.3, MADEEP, MADIBENTHOS, MAINBAZA, MIRIKY, MUSORSTOM 9, NORFOLK 2, NanHai 2014, PAKAIHI I TE MOANA, PANGLAO 2004, PANGLAO 2005, PAPUA NIUGINI, SALOMON 2, SALOMONBOA 3, SANTO 2006, TAIWAN 2004, TAIWAN 2013, TARASOC, TERRASSES, ZhongSha 2015