New insights into the molecular phylogeny, biogeographical history, and diversification of Amblyomma ticks (Acari: Ixodidae) based on mitogenomes and nuclear sequences.

BACKGROUND: Amblyomma is the third most diversified genus of Ixodidae that is distributed across the Indomalayan, Afrotropical, Australasian (IAA), Nearctic and Neotropical biogeographic ecoregions, reaching in the Neotropic its highest diversity. There have been hints in previously published phylogenetic trees from mitochondrial genome, nuclear rRNA, from combinations of both and morphology that the Australasian Amblyomma or the Australasian Amblyomma plus the Amblyomma species from the southern cone of South America, might be sister-group to the Amblyomma of the rest of the world. However, a stable phylogenetic framework of Amblyomma for a better understanding of the biogeographic patterns underpinning its diversification is lacking. METHODS: We used genomic techniques to sequence complete and nearly complete mitochondrial genomes -ca. 15 kbp- as well as the nuclear ribosomal cluster -ca. 8 kbp- for 17 Amblyomma ticks in order to study the phylogeny and biogeographic pattern of the genus Amblyomma, with particular emphasis on the Neotropical region. The new genomic information generated here together with genomic information available on 43 ticks (22 other Amblyomma species and 21 other hard ticks-as outgroup-) were used to perform probabilistic methods of phylogenetic and biogeographic inferences and time-tree estimation using biogeographic dates. RESULTS: In the present paper, we present the strongest evidence yet that Australasian Amblyomma may indeed be the sister-group to the Amblyomma of the rest of the world (species that occur mainly in the Neotropical and Afrotropical zoogeographic regions). Our results showed that all Amblyomma subgenera (Cernyomma, Anastosiella, Xiphiastor, Adenopleura, Aponomma and Dermiomma) are not monophyletic, except for Walkeriana and Amblyomma. Likewise, our best biogeographic scenario supports the origin of Amblyomma and its posterior diversification in the southern hemisphere at 47.8 and 36.8 Mya, respectively. This diversification could be associated with the end of the connection of Australasia and Neotropical ecoregions by the Antarctic land bridge. Also, the biogeographic analyses let us see the colonization patterns of some neotropical Amblyomma species to the Nearctic. CONCLUSIONS: We found strong evidence that the main theater of diversification of Amblyomma was the southern hemisphere, potentially driven by the Antarctic Bridge's intermittent connection in the late Eocene. In addition, the subgeneric classification of Amblyomma lacks evolutionary support. Future studies using denser taxonomic sampling may lead to new findings on the phylogenetic relationships and biogeographic history of Amblyomma genus.

Hepatozoon (Eucoccidiorida: Hepatozoidae) in wild mammals of the Americas: a systematic review.

BACKGROUND: The study of parasites provides insight into intricate ecological relationships in ecosystem dynamics, food web structures, and evolution on multiple scales. Hepatozoon Eucoccidiorida: Hepatozoidae) is a genus of protozoan hemoparasites with heteroxenous life cycles that switch infections between vertebrates and blood-feeding invertebrates. The most comprehensive review of the genus was published 26 years ago, and currently there are no harmonized data on the epizootiology, diagnostics, genotyping methods, evolutionary relationships, and genetic diversity of Hepatozoon in the Americas. METHODS: Here, we provide a comprehensive review based on the PRISMA method regarding Hepatozoon in wild mammals within the American continent, in order to generate a framework for future research. RESULTS: 11 out of the 35 countries of the Americas (31.4%) had data on Hepatozoon, with Carnivora and Rodentia orders having the most characterizations. Bats, ungulates, and shrews were the least affected groups. While Hepatozoon americanum, H. americanum-like, H. canis, H. didelphydis, H. felis, H. milleri, H. griseisciuri, and H. procyonis correspond to the identified species, a plethora of genospecies is pending for a formal description combining morphology and genetics. Most of the vectors of Hepatozoon in the Americas are unknown, but some flea, mite, and tick species have been confirmed. The detection of Hepatozoon has relied mostly on conventional polymerase chain reaction (PCR), and the implementation of specific real time PCR for the genus needs to be employed to improve its diagnosis in wild animals in the future. From a genetic perspective, the V4 region of the 18S rRNA gene has been widely sequenced for the identification of Hepatozoon in wild animals. However, mitochondrial and apicoplast markers should also be targeted to truly determine different species in the genus. A phylogenetic analysis of herein retrieved 18S ribosomal DNA (rDNA) sequences showed two main clades of Hepatozoon: Clade I associated with small mammals, birds, and herpetozoa, and Clade II associated with Carnivora. The topology of the tree is also reflected in the haplotype network. CONCLUSIONS: Finally, our review emphasizes Hepatozoon as a potential disease agent in threatened wild mammals and the role of wild canids as spreaders of Hepatozoon infections in the Americas.

New insights into the systematics of the afrotropical Amblyomma marmoreum complex (Acari: Ixodidae) and the genome of a novel Rickettsia africae strain using morphological and metagenomic approaches.

The Amblyomma marmoreum complex includes afrotropical species, such as Amblyomma sparsum, a three-host tick that parasitizes reptiles, birds, and mammals, and is a recognized vector of Ehrlichia ruminantium. However, the lack of morphological, genetic and ecological data on A. sparsum has caused considerable confusion in its identification. In this study, we used microscopy and metagenomic approaches to analyze A. sparsum ticks collected from a puff adder snake (Bitis arietans) in southwest Senegal (an endemic rickettsioses area) in order to supplement previous morphological descriptions, provide novel genomic data for the A. marmoreum complex, and describe the genome of a novel spotted fever group Rickettsia strain. Based on stereoscope and scanning electron microscopy (SEM) morphological evaluations, we provide high-quality images and new insights about punctation and enameling in the adult male of A. sparsum to facilitate identification for future studies. The metagenomic approach allowed us assembly the complete mitochondrial genome of A. sparsum, as well as the nearly entire chromosome and complete plasmid sequences of a novel Rickettsia africae strain. Phylogenomic analyses demonstrated a close relationship between A. sparsum and Amblyomma nuttalli for the first time and confirmed the position of A. sparsum within the A. marmoreum complex. Our results provide new insights into the systematics of A. sparsum and A. marmoreum complex, as well as the genetic diversity of R. africae in the Afrotropical region. Future studies should consider the possibility that A. sparsum may be a vector for R. africae.

Survey and Molecular Characterization of Sarcocystidae protozoa in Wild Cricetid Rodents from Central and Southern Chile.

In Chile, studies of parasites from the family Sarcocystidae (Apicomplexa) have mostly been related to domestic animals. We aimed to assess the presence of Sarcocystidae taxa in cricetid rodents from Central and Southern Chile. We studied 207 rodents, encompassing six species, from 13 localities. We isolated DNA from tissue samples, amplified the Sarcocystidae 18S rRNA gene with polymerase chain reaction, and performed phylogenetic analyses using maximum likelihood and Bayesian inferences. In addition, we examined blood smears and performed histological studies in organs from Sarcocystidae DNA-positive animals. Three specimens were DNA-positive and three genotypes were retrieved and named: Sarcocystis sp. P61, related to Sarcocystis strixi, was detected in two Abrothrix olivacea. Toxoplasmatinae gen. sp. P99 was retrieved from those same two specimens, and was related to Toxoplasma and other genera, although it branched independently. Besnoitia sp. R34 was detected in one Abrothrix hirta, and was clustered with congeneric species associated with rodents. No protozoa were found during microscopic studies; thus, it was not possible to confirm parasitic interactions rather than accidental encounters. However, the close relatedness of the retrieved genotypes to parasites of rodents supports the hypothesis of host-parasite associations. All three genotypes are suggested as potential new taxa, including a putative new genus.

Hidden species diversity and mito-nuclear discordance within the Mediterranean cone snail, Lautoconus ventricosus.

The Mediterranean cone snail, Lautoconus ventricosus, is currently considered a single species inhabiting the whole Mediterranean basin and the adjacent Atlantic coasts. Yet, no population genetic study has assessed its taxonomic status. Here, we collected 245 individuals from 75 localities throughout the Mediterranean Sea and used cox1 barcodes, complete mitochondrial genomes, and genome skims to test whether L. ventricosus represents a complex of cryptic species. The maximum likelihood phylogeny based on complete mitochondrial genomes recovered six main clades (hereby named blue, brown, green, orange, red, and violet) with sufficient sequence divergence to be considered putative species. On the other hand, phylogenomic analyses based on 437 nuclear genes only recovered four out of the six clades: blue and orange clades were thoroughly mixed and the brown one was not recovered. This mito-nuclear discordance revealed instances of incomplete lineage sorting and introgression, and may have caused important differences in the dating of main cladogenetic events. Species delimitation tests proposed the existence of at least three species: green, violet, and red + blue + orange (i.e., cyan). Green plus cyan (with sympatric distributions) and violet, had West and East Mediterranean distributions, respectively, mostly separated by the Siculo-Tunisian biogeographical barrier. Morphometric analyses of the shell using species hypotheses as factor and shell length as covariate showed that the discrimination power of the studied parameters was only 70.2%, reinforcing the cryptic nature of the uncovered species, and the importance of integrative taxonomic approaches considering morphology, ecology, biogeography, and mitochondrial and nuclear population genetic variation.

Phylogenetic relationships of the Amblyomma cajennense complex (Acari: Ixodidae) at mitogenomic resolution.

The genus Amblyomma is the third most diverse in the number of species within the Ixodidae, with practically half of its species distributed in the Americas, though there are also species occurring in Africa, Asia, and Australia. Within the genus, there are several species complexes with veterinary and public health importance. The Amblyomma cajennense complex, in the Americas, is represented by six species with a wide distribution, from Texas to northern Argentina. We combined two sequencing techniques to generate complete mitogenomes of species belonging to the Amblyomma cajennense complex: genome skimming and long-range PCRs sequencing methods. Thus, we generated seven new mitochondrial genomes for all species of the Amblyomma cajennense complex, except for Amblyomma interandinum. Genetic distances between the mitogenomes corroborate the clear differentiation between the five species of the Amblyomma cajennense complex. The phylogenetic relationships of these species had previously been evaluated by combining partial nuclear and mitochondrial genes and here these relationships are corroborated with a more robust framework of data, which demonstrates that the conjunction of mitochondrial and nuclear partial genes can resolve close relationships when entire genes or genomes are unavailable. The gene order, structure, composition, and length are stable across these mitogenomes, and they share the general characteristics of Metastriata. Future studies should increase the number of available mitogenomes for this genus, especially for those species from the Indo-Pacific region and Africa, by means of a better understanding of their relationships and evolutionary process.

Wild deer (Pudu puda) from Chile harbor a novel ecotype of Anaplasma phagocytophilum.

BACKGROUND: Deer species play an important role in the enzootic cycles of several Anaplasma species. While in the Northern Hemisphere ticks of genus Ixodes are well recognized vectors of these intracellular bacteria, less is known regarding the biological cycles of Anaplasma spp. in South America. METHODS: Using PCR protocols and Sanger sequencing, we assessed the presence of Anaplasma spp. in blood and ticks collected on a native deer species (Pudu puda) from southern Chile. RESULTS: Based on phylogenetic analyses of the 16S rRNA, gltA and groEL genes and calculation of average sequence divergence for groEL, our results bring to light a novel genovariant of Anaplasma phagocytophilum (named strain "Patagonia"). The strain represents a novel ecotype within the A. phagocytophilum species complex and was detected in both P. puda and their ticks. Using a larger matrix, denser taxon sampling and outgroup, our maximum-likelihood- and Bayesian-inferred phylogenies for groEL provide an accurate picture of the topology of A. phagocytophilum ecotypes and their evolutionary relationships. CONCLUSIONS: This is the first report of an ecotype of A. phagocytophilum in South America. Our results provide novel insight into the genetic diversity and ecology of this complex of bacterial lineages. Further studies should elucidate the enzootic cycle of A. phagocytophilum strain "Patagonia" and assess its pathogenic potential for pudues, domestic animals and humans in the region.

A Phylogenomic Backbone for Gastropod Molluscs.

Gastropods have survived several mass extinctions during their evolutionary history resulting in extraordinary diversity in morphology, ecology, and developmental modes, which complicate the reconstruction of a robust phylogeny. Currently, gastropods are divided into six subclasses: Caenogastropoda, Heterobranchia, Neomphaliones, Neritimorpha, Patellogastropoda, and Vetigastropoda. Phylogenetic relationships among these taxa historically lack consensus, despite numerous efforts using morphological and molecular information. We generated sequence data for transcriptomes derived from 12 taxa belonging to clades with little or no prior representation in previous studies in order to infer the deeper cladogenetic events within Gastropoda and, for the first time, infer the position of the deep-sea Neomphaliones using a phylogenomic approach. We explored the impact of missing data, homoplasy, and compositional heterogeneity on the inferred phylogenetic hypotheses. We recovered a highly supported backbone for gastropod relationships that is congruent with morphological and mitogenomic evidence, in which Patellogastropoda, true limpets, are the sister lineage to all other gastropods (Orthogastropoda) which are divided into two main clades 1) Vetigastropoda $s.l.$ (including Pleurotomariida $+$ Neomphaliones) and 2) Neritimorpha $+$ (Caenogastropoda $+$ Heterobranchia). As such, our results support the recognition of five subclasses (or infraclasses) in Gastropoda: Patellogastropoda, Vetigastropoda, Neritimorpha, Caenogastropoda, and Heterobranchia. [Compositional heterogeneity; fast-evolving; long-branch attraction; missing data; Mollusca; phylogenetics; systematic error.].

Extension of the distribution of Amblyomma triste Koch, 1844: Morphological and molecular confirmation of Morphotype I in Colombia.

Ticks of the Amblyomma maculatum (Acari: Ixodidae) group are represented by three species: Amblyomma maculatum, Amblyomma tigrinum, and Amblyomma triste. This group is widely distributed throughout the Nearctic and Neotropical regions, from the United States of America (USA) to Argentina. They parasitize several vertebrates, including domestic mammals and humans. Nonetheless, the identification and distribution of the species within the group have been controversial but can be solved with integrative studies on their taxonomy and ecology. In Colombia, a Neotropical country from which up to 58 tick species have been reported, only Amblyomma maculatum Koch, 1844 (sensu stricto - s.s.) - morphotype II - has been molecularly confirmed. Here, specimens of the A. maculatum group from three departments of Colombia were examined by morphological and molecular analyses. The results confirmed the wide distribution of A. maculatum s.s. in Colombia. Furthermore, the presence of A. triste s.s. - morphotype I was confirmed for the first time at the molecular level, which broadens the distribution of this species in the American continent. Considering that both taxa are vectors of pathogens such as Rickettsia parkeri s.s., additional taxonomic, reproductive compatibility and niche distribution studies are needed, as well as the application of new molecular markers due to the controversial taxonomy. This will allow us to explore cryptic diversity within the recently established putative groups.

The complete mitochondrial genome of Costapex baldwinae (Gastropoda: Neogastropoda: Turbinelloidea: Costellariidae) from the Caribbean Deep-Sea.

We report the complete mitochondrial genome sequence of Costapex baldwinae, a Caribbean representative of a predominantly Indo-Pacific genus of gastropods that occurs on sunken wood at bathyal depths. The mitogenome is 15,321 bp in length and has a base composition of 29.2% A, 41.8% T, 12.0% C and 17.0% G. It contains 13 protein-coding, two ribosomal RNA, and 22 tRNA genes with the same gene order and strand orientation as other non-toxoglossan neogastropods. Phylogenetic analyses indicate that the superfamily Turbinelloidea, represented by this species, diverged early within the Neogastropod radiation, forming the sister group to a clade that includes five of the seven presently recognized superfamilies.

Characterization of the complete mitochondrial genome of Amblyomma ovale, comparative analyses and phylogenetic considerations.

In this study, we sequenced two complete mitochondrial genomes of Amblyomma ovale, a tick of public health importance. Sequencing two distinct individuals, the resulting mitochondrial genomes were 14,756 and 14,760 bp in length and maintained the same gene order previously reported in Amblyomma. These were combined with RNA-seq derived mitochondrial sequences from three additional species, Amblyomma aureolatum, Amblyomma maculatum, and Amblyomma moreliae, to carry out mitogenome comparative and evolutionary analyses against all previously published tick mitochondrial genomes. We described a derivative genome rearrangement that isolates Ixodes from the remaining Ixodidae and consists of both a reverse translocation as well as an event of Tandem Duplication Random Loss. Genetic distance analyses indicated that cox2, nd1, nd5, and 16S are good candidates for future population studies in A. ovale. The phylogenetic analyses corroborated the utility of complete mitochondrial genomes as phylogenetic markers within the group. This study further supplements the genome information available for Amblyomma and facilitates future evolutionary and population genetic studies within the genus.

A mitogenomic phylogeny of chitons (Mollusca: Polyplacophora).

BACKGROUND: Polyplacophora, or chitons, have long fascinated malacologists for their distinct and rather conserved morphology and lifestyle compared to other mollusk classes. However, key aspects of their phylogeny and evolution remain unclear due to the few morphological, molecular, or combined phylogenetic analyses, particularly those addressing the relationships among the major chiton lineages. RESULTS: Here, we present a mitogenomic phylogeny of chitons based on 13 newly sequenced mitochondrial genomes along with eight available ones and RNAseq-derived mitochondrial sequences from four additional species. Reconstructed phylogenies largely agreed with the latest advances in chiton systematics and integrative taxonomy but we identified some conflicts that call for taxonomic revisions. Despite an overall conserved gene order in chiton mitogenomes, we described three new rearrangements that might have taxonomic utility and reconstructed the most likely scenario of gene order change in this group. Our phylogeny was time-calibrated using various fossils and relaxed molecular clocks, and the robustness of these analyses was assessed with several sensitivity analyses. The inferred ages largely agreed with previous molecular clock estimates and the fossil record, but we also noted that the ambiguities inherent to the chiton fossil record might confound molecular clock analyses. CONCLUSIONS: In light of the reconstructed time-calibrated framework, we discuss the evolution of key morphological features and call for a continued effort towards clarifying the phylogeny and evolution of chitons.

New patellogastropod mitogenomes help counteracting long-branch attraction in the deep phylogeny of gastropod mollusks.

Long-branch attraction (LBA) is a well-known artifact in phylogenetic reconstruction. Sparse taxon sampling and extreme heterogeneity of evolutionary rates among lineages generate propitious situations for LBA, even defying probabilistic methods of phylogenetic inference. A clear example illustrating LBA challenges is the difficulty of reconstructing the deep gastropod phylogeny, particularly using mitochondrial (mt) genomes. Previous studies consistently obtained unorthodox phylogenetic relationships due to the LBA between the mitogenomes of patellogastropods (true limpets, represented only by Lottia digitalis), heterobranchs, and outgroup taxa. Here, we use the reconstruction of the gastropod mitogenomic phylogeny as a case exercise to test the effect of key methodological approaches proposed to counteract LBA, including the selection of slow-evolving representatives, the use of different outgroups, the application of site-heterogeneous evolutionary models, and the removal of fast-evolving sites. In this regard, we sequenced three new patellogastropod mt genomes, which displayed shorter branches than the one of Lottia as well as gene organizations more similar to that of the hypothetical gastropod ancestor. Phylogenetic analyses incorporating the mt genomes of Patella ferruginea, Patella vulgata, and Cellana radiata allowed eliminating the artificial clustering of Patellogastropoda and Heterobranchia that had prevailed in previous studies. Furthermore, the use of site-heterogeneous models with certain combinations of lineages within the outgroup allowed eliminating also the LBA between Heterobranchia and the outgroup, and recovering Apogastropoda (i.e., Caenogastropoda + Heterobranchia). Hence, for the first time, we were able to obtain a mitogenomic phylogeny of gastropods that is congruent with both morphological and nuclear datasets.

Phylogenetic relationships of the conoidean snails (Gastropoda: Caenogastropoda) based on mitochondrial genomes.

With more than 5,000 species, Conoidea is one of the most diversified superfamilies of Gastropoda. Recently, the family-level classification of these venomous predator snails has undergone substantial changes, on the basis of a phylogenetic tree reconstructed combining partial mitochondrial and nuclear gene sequences, and up to 16 families are now recognized. However, phylogenetic relationships among these families remain largely unresolved. Here, we sequenced 20 complete or nearly complete mitochondrial (mt) genomes, which were combined with mt genomes available in GenBank to construct a dataset that included representatives of 80% of the known families, although for some we had only one species or genus as representative. Most of the sequenced conoidean mt genomes shared a constant genome organization, and observed rearrangements were limited exclusively to tRNA genes in a few lineages. Phylogenetic trees were reconstructed using probabilistic methods. Two main monophyletic groups, termed "Clade A" and "Clade B", were recovered with strong support within a monophyletic Conoidea. Clade A (including families Clavatulidae, Horaiclavidae, Turridae s.s., Terebridae, Drilliidae, Pseudomelatomidae, and Cochlespiridae) was composed of four main lineages, one of which was additionally supported by a rearrangement in the gene order. Clade B (including families Conidae, Borsoniidae, Clathurellidae, Mangeliidae, Raphitomidae, and Mitromorphidae) was composed of five main lineages. The reconstructed phylogeny rejected the monophyly of Clavatulidae, Horaiclavidae, Turridae, Pseudomelatomidae, and Conidae, indicating that several of the currently accepted families may be ill-defined. The reconstructed tree also revealed new phylogenetic positions for genera characterized as tentative (Gemmuloborsonia, Lucerapex, and Leucosyrinx), enigmatic (Marshallena) or challenging to place (Fusiturris), which will potentially impact the classification of the Conoidea.

Phylogenetic relationships of cone snails endemic to Cabo Verde based on mitochondrial genomes.

BACKGROUND: Due to their great species and ecological diversity as well as their capacity to produce hundreds of different toxins, cone snails are of interest to evolutionary biologists, pharmacologists and amateur naturalists alike. Taxonomic identification of cone snails still relies mostly on the shape, color, and banding patterns of the shell. However, these phenotypic traits are prone to homoplasy. Therefore, the consistent use of genetic data for species delimitation and phylogenetic inference in this apparently hyperdiverse group is largely wanting. Here, we reconstruct the phylogeny of the cones endemic to Cabo Verde archipelago, a well-known radiation of the group, using mitochondrial (mt) genomes. RESULTS: The reconstructed phylogeny grouped the analyzed species into two main clades, one including Kalloconus from West Africa sister to Trovaoconus from Cabo Verde and the other with a paraphyletic Lautoconus due to the sister group relationship of Africonus from Cabo Verde and Lautoconus ventricosus from Mediterranean Sea and neighboring Atlantic Ocean to the exclusion of Lautoconus endemic to Senegal (plus Lautoconus guanche from Mauritania, Morocco, and Canary Islands). Within Trovaoconus, up to three main lineages could be distinguished. The clade of Africonus included four main lineages (named I to IV), each further subdivided into two monophyletic groups. The reconstructed phylogeny allowed inferring the evolution of the radula in the studied lineages as well as biogeographic patterns. The number of cone species endemic to Cabo Verde was revised under the light of sequence divergence data and the inferred phylogenetic relationships. CONCLUSIONS: The sequence divergence between continental members of the genus Kalloconus and island endemics ascribed to the genus Trovaoconus is low, prompting for synonymization of the latter. The genus Lautoconus is paraphyletic. Lautoconus ventricosus is the closest living sister group of genus Africonus. Diversification of Africonus was in allopatry due to the direct development nature of their larvae and mainly triggered by eustatic sea level changes during the Miocene-Pliocene. Our study confirms the diversity of cone endemic to Cabo Verde but significantly reduces the number of valid species. Applying a sequence divergence threshold, the number of valid species within the sampled Africonus is reduced to half.

Beyond Conus: Phylogenetic relationships of Conidae based on complete mitochondrial genomes.

Understanding how the extraordinary taxonomic and ecological diversity of cone snails (Caenogastropoda: Conidae) evolved requires a statistically robust phylogenetic framework, which thus far is not available. While recent molecular phylogenies have been able to distinguish several deep lineages within the family Conidae, including the genera Profundiconus, Californiconus, Conasprella, and Conus (and within this one, several subgenera), phylogenetic relationships among these genera remain elusive. Moreover, the possibility that additional deep lineages may exist within the family is open. Here, we reconstructed with probabilistic methods a molecular phylogeny of Conidae using the newly sequenced complete or nearly complete mitochondrial (mt) genomes of the following nine species that represent all main Conidae lineages and potentially new ones: Profundiconus teramachii, Californiconus californicus, Conasprella wakayamaensis, Lilliconus sagei, Pseudolilliconus traillii, Conus (Kalloconus) venulatus, Conus (Lautoconus) ventricosus, Conus (Lautoconus) hybridus, and Conus (Eugeniconus) nobilis. To test the monophyly of the family, we also sequenced the nearly complete mt genomes of the following three species representing closely related conoidean families: Benthomangelia sp. (Mangeliidae), Tomopleura sp. (Borsoniidae), and Glyphostoma sp. (Clathurellidae). All newly sequenced conoidean mt genomes shared a relatively constant gene order with rearrangements limited to tRNA genes. The reconstructed phylogeny recovered with high statistical support the monophyly of Conidae and phylogenetic relationships within the family. The genus Profundiconus was placed as sister to the remaining genera. Within these, a clade including Californiconus and Lilliconus+Pseudolilliconus was the sister group of Conasprella to the exclusion of Conus. The phylogeny included a new lineage whose relative phylogenetic position was unknown (Lilliconus) and uncovered thus far hidden diversity within the family (Pseudolilliconus). Moreover, reconstructed phylogenetic relationships allowed inferring that the peculiar diet of Californiconus based on worms, mollusks, crustaceans and fish is derived, and reinforce the hypothesis that the ancestor of Conidae was a worm hunter. A chronogram was reconstructed under an uncorrelated relaxed molecular clock, which dated the origin of the family shortly after the Cretaceous-Tertiary boundary (about 59million years ago) and the divergence among main lineages during the Paleocene and the Eocene (56-30million years ago).

Phylogenetic relationships of Mediterranean and North-East Atlantic Cantharidinae and notes on Stomatellinae (Vetigastropoda: Trochidae).

The subfamily Cantharidinae Gray, 1857 (Trochoidea: Trochidae) includes 23 recognized genera and over 200 known living species. These marine top shell snails are microphagous grazers that generally live in shallow rocky shores and in macroalgae and seagrass beds of sub-tropical and temperate waters from the Central and Western Indo-Pacific biogeographic regions to the Mediterranean Sea and the Eastern Atlantic Ocean. Recent molecular phylogenetic studies revising the family Trochidae supported the monophyly of the subfamily Cantharidinae and its sister group relationship to the subfamily Stomatellinae. These studies and others has thus far mostly focused on Indo-Pacific members of the subfamily Cantharidinae whereas here, we investigated phylogenetic relationships among their counterparts from the Mediterranean Sea and the North-eastern (NE) Atlantic Ocean including 33 species of genera Gibbula, Jujubinus, Phorcus, Clelandella, and Callumbonella. The Mediterranean and NE Atlantic taxa were supplemented with 30 Indo-Pacific Cantharidinae species plus 19 members of the sister group subfamily Stomatellinae. Phylogenetic trees were constructed using Bayesian inference and maximum likelihood with two datasets comprised of partial sequences of four or six mitochondrial (cox1, rrnL, rrnS, and cob) and nuclear (28S rRNA and histone H3) genes. A clade comprised of all Mediterranean and NE Atlantic taxa was recovered with high support, but its sister group among the Indo-Pacific lineages could not be determined with confidence (although the assignment of "Trochus" kotschyi to Priotrochus could be rejected). Within the Mediterranean and NE Atlantic clade, genera Phorcus and Jujubinus were recovered as reciprocally monophyletic, and the deep-sea genera Clelandella and Callumbonella were placed with high support as sister to Jujubinus. However, the genus Gibbula as currently defined was not monophyletic and constituent species were divided into three major clades and two independent lineages. Phylogenetic relationships among Phorcus, Jujubinus (plus Clelandella and Callumbonella), and the different clades of Gibbula were not fully resolved but received higher support in the phylogenetic analyses based on six genes. A first approach to resolve phylogenetic relationships within Stomatellinae was conducted showing that the diversity of the subfamily is highly underestimated at present, and that Calliotrochus is possibly a member of this subfamily. A chronogram was reconstructed using an uncorrelated relaxed lognormal molecular clock and the origin of the Mediterranean and NE Atlantic clade was dated right after the Azolla phase in the Middle Eocene about 48 million years ago whereas diversification of major clades (genera) followed the eastern closure of the Tethys Ocean in the Middle Miocene about 14 million years ago.

Phylogenetic relationships among superfamilies of Neritimorpha (Mollusca: Gastropoda).

Despite the extraordinary morphological and ecological diversity of Neritimorpha, few studies have focused on the phylogenetic relationships of this lineage of gastropods, which includes four extant superfamilies: Neritopsoidea, Hydrocenoidea, Helicinoidea, and Neritoidea. Here, the nucleotide sequences of the complete mitochondrial genomes of Georissa bangueyensis (Hydrocenoidea), Neritina usnea (Neritoidea), and Pleuropoma jana (Helicinoidea) and the nearly complete mt genomes of Titiscania sp. (Neritopsoidea) and Theodoxus fluviatilis (Neritoidea) were determined. Phylogenetic reconstructions using probabilistic methods were based on mitochondrial (13 protein coding genes and two ribosomal rRNA genes), nuclear (partial 28S rRNA, 18S rRNA, actin, and histone H3 genes) and combined sequence data sets. All phylogenetic analyses except one converged on a single, highly supported tree in which Neritopsoidea was recovered as the sister group of a clade including Helicinoidea as the sister group of Hydrocenoidea and Neritoidea. This topology agrees with the fossil record and supports at least three independent invasions of land by neritimorph snails. The mitochondrial genomes of Titiscania sp., G. bangueyensis, N. usnea, and T. fluviatilis share the same gene organization previously described for Nerita mt genomes whereas that of P. jana has undergone major rearrangements. We sequenced about half of the mitochondrial genome of another species of Helicinoidea, Viana regina, and confirmed that this species shares the highly derived gene order of P. jana.

The complete mitogenome of the trilobite beetle, Platerodrilus sp. (Elateroidea: Lycidae).

The nucleotide sequence of the complete mitogenome of the trilobite beetle, Platerodrilus sp. was determined. This is the first complete mitogenome reported for the family Lycidae (Elateroidea: Coleoptera). The newly determined sequence is 16,394 bp long and shows a relatively high AT content (76.2%). The gene arrangement of the trilobite beetle mt genome is the same found in other related beetle mitogenomes. The reconstructed tree recovered Elateroidea as a strongly supported monophyletic group but could not resolve phylogenetic relationships among studied elateroid families, including Lycidae.