Karyological studies of five species of land snails (Helicoidea: Mollusca)

The karyotypes of five species of Helicoidea (Japanese land snails) were analyzed by the air-drying method, using embryonic cells. Somatic cells of Satsuma omphalodes and S. japonica had 2n=58. Karyotypes of Acusta despecta sieboldiana (2n=58), Euhadra dixoni and E. amaliae (2n=56) were also analyzed. Chromosomes of these five species are all metacentric or submetacentric. Two species of Euhadra, dixoni and amaliae, have identical karyotypes while karyotypes of S. omphalodes and S. japonica indicate some dissimilarity in medium-sized and small-sized chromosomes. On the basis of karyotypical characters, A. d. sieboldiana was recognized to be closely related to the Satsuma species, which belong to a different family rather than to Euhadra of the same family as Acusta.     

A phylogeny of the land snails (Gastropoda: Pulmonata)

We have undertaken the first large-scale molecular phylogenetic analysis of the Stylommatophora. Sequences of the ribosomal RNA gene-cluster were examined in 104 species of snails and slugs from 50 families, encompassing all the currently recognized major groups. It allows an independent test of the present classification based on morphology. At the level of families our molecular phylogeny closely supports the current taxonomy, but the deep branches within the tree do not. Surprisingly, a single assemblage including the families Achatinidae, Subulinidae and Streptaxidae lies near the base of the tree, forming a sister group to all remaining stylommatophorans. This primary division into 'achatinoid' and 'non-achatinoid' taxa is unexpected, and demands a radical reinterpretation of early stylommatophoran evolution. In particular, the Orthurethra appear to be relatively advanced within the 'non-achatinoid clade', and broadly equivalent to other super-familial clusters. This indicates that supposedly primitive features such as the orthurethran kidney are derived. The molecular tree also suggests that the origin of the Stylommatophora is much earlier than the main period of their diversification.     

Evolutionary relationships among the Pulmonate land snails and slugs (Pulmonata, Stylommatophora)

We have incorporated an additional 56 species of land snails and slugs in our ribosomal (r) RNA molecular phylogeny. The new taxa include representatives of several important groups. The molecular tree now includes 160 species of stylommatophoran land snails and slugs in 144 genera in 61 families. In the rDNA tree, the Stylommatophora are principally divided into an 'achatinoid' and a 'non-achatinoid' clade. Within these clades, several major land snail groups, including the Orthurethra, Elasmognatha, Limacoidea, and Helicoidea, are supported. Overall, the rDNA molecular phylogeny has remained stable following the incorporation of the additional taxa, with these additions having little impact on the major evolutionary patterns in the tree. Taxonomic coverage of the Orthurethra, Orthalicidae, Camaenidae, and Bradybaenidae is increased significantly. The camaenids and bradybaenids form a complex, and both appear to be paraphyletic. Several families of uncertain affinity, such as the Sagdidae and Thyrophorellidae, are included for the first time. The Sagdidae are shown to belong to the Helicoidea, and the Thyrophorellidae to the Achatinoidea.  © 2006 The Linnean Society of London, Biological Journal of the Linnean Society , 2006, 87 , 593–610.     

Molecular phylogeny and character evolution in the Western Palaearctic Helicidae s.l. (Gastropoda: Stylommatophora)

In this study, we present a molecular phylogeny for the west Palaearctic Helicidae sensu lato based on sequence data from two mitochondrial (COI, 16S rDNA) and two nuclear (ITS-1, 18S rDNA) genes. Maximum likelihood analysis and Bayesian inference revealed well supported monophyletic clades partly conflicting traditional classifications. Based on these results, we propose the following system. The Western Palaearctic Helicidae s.l. consist of two families, Helicidae and Hygromiidae. Within the Helicidae, three well supported subfamilies can be recognised: the Helicinae, Ariantinae, and Helicodontinae. The Hygromiidae consist of three clades: the Hygromiinae, the Helicellinae, and a yet unnamed clade comprising the genera Sphincterochila and Cochlicella. We then used the phylogeny to study the evolution of anatomical, and ecological characters traditionally used for systematic classification. In the Helicidae s.l., two independent evolutionary transitions to life in xeric environments occurred, which allowed the occupation of new niches with a subsequent radiation of the Helicellinae-Cochlicella/Sphincterochila clade and the Helicinae. Whereas, the multiplication of the Glandulae mucosae is a synapomorphy of the Hygromiidae, the lovedart sac apparatus is present in all groups and thus, the trait cannot provide a synapomorphy for either families or subfamilies. Additionally, we evaluated the use of structural molecular genetic characters for taxonomic assessment. The presence of an unique loop region of the 16S rDNA gene and a short tandem repeat in the ITS-1 region provide independent evidence for the monophyly of these major two groups, and can be used for preliminary classification.     

Biogeography of the Helicoidea (Mollusca: Gastropoda: Pulmonata): land snails with a Pangean distribution

The biogeography of helicoid land snails was investigated using cladistic methods. Parsimony analysis under Assumption 0 yielded twelve area cladograms (length=25, c.i.=0.76, r.i.=0.86). The pattern of vicariance for the Helicoidea indicated that families originated with the break up of eastern Gondwana and Laurasia between the late Mesozoic and mid-Tertiary, and possible vicariance events are identified. It is proposed that Asian terranes, located between India and Australia, maintained contact with northern Australia until the late Cretaceous, which is later than is suggested in current palaeogeographical hypotheses.     

A hairy business—Periostracal hair formation in two species of helicoid snails (Gastropoda,Stylommatophora, Helicoidea)

In molluscs, the calcareous shell is covered externally by a thin organic layer, the periostracum. The periostracum of some pulmonate species is of special taxonomic interest because it bears distinct microscale architectures. Where and how these structures are formed is as yet unknown. Using histological sections through their shells, gelatin cuts, and live observations I studied the pattern by which the periostracal hair‐like projections in two helicoid land snail species are secreted and evenly arranged on the shell. The results indicate a complex mechanism: a hair is formed in the periostracal groove independently of the periostracum, after which it is attached to the edge of the shell, drawn out of the tissue, and finally swivelled to the upper side of the periostracum. Upon further growth of the periostracum, the hairs are finally fixed upright on the shell. J. Morphol. 2011. © 2011 Wiley‐Liss, Inc.     

A molecular approach to the phylogenetic relationships of the western palaearctic Helicoidea (Gastropoda: Stylommatophora)

Molecular phylogenetic relationships among 45 members of the Helicoidea (Gastropoda: Stylommatophora) were examined using partial mitochondrial 16S rRNA sequences. Phylogenetic relationships were inferred using maximum parsimony, maximum likelihood and Bayesian methods. The reconstructed phylogenies showed a good degree of support for more recent branches, but gave little support to deeper nodes. Mitochondrial rDNA data further confirmed monophyletic status of helicids, recognized monachine hygromiid and bradybaenid clades and resolved a number of relationships in the helicelline hygromiids. With the respect to the latter assemblage, most of the anatomically based groups are confirmed, corroborating the diagnostic value of the dart-sac complex and a close affinity between Ichnusomunda sacchii and species of the genus Cernuella . Nevertheless, some well resolved branches challenge previous systematic arrangements, grouping species previously placed in different arrangements. In particular, support was not found for the monophyly of helicelline hygromiids with pedal penial innervation. Possible explanations for these incongruencies are suggested. 16S sequence data are appropriate for studies of relationships within the different species groups and less so for recovery of more ancient radiations in the Helicoidea. It will be valuable to combine the 16S data with other gene sequences to estimate basal relationships.  © 2005 The Linnean Society of London, Biological Journal of the Linnean Society , 2005, 85 , 501–512.     

Molecular phylogeny, taxonomy and evolution of the land snail genus Iberus (Pulmonata: Helicidae)

Partial DNA sequences of two mitochondrial genes [cytochrome oxidase subunit I (COI) and 16S rRNA] from 59 specimens of Iberus were used to test the validity of the described morphospecies of this genus, and examine genetic divergences within and between main phylogenetic groups. Both gene fragments showed phylogenetic concordance. The COI gene was found to be faster evolving than the 16S gene and was fully protein-coding with no insertions or deletions. 16S rRNA was more informative than COI for resolving basal nodes. Both individual and combined analyses of the two gene fragments revealed five main phylogroups. These five groups are genetically unique lineages that are allopatrically distributed and considered to have full species status. Further subdivisions were also considered. Shell morphology was suitable for delimiting species boundaries, but several incongruences between morphology and mtDNA phylogeny were observed. These incongruences were considered consequence of hybridization between Iberus cobosi and Iberus marmoratus , and the result of shell shape polymorphism in Iberus rositai . According to spatial patterns of sequence divergence, life habits and shell morphology may be concluded that the keeled-flat shelled snails independently originated several times within Iberus and they could represent cases of similar shell adaptation to a karstic arid environment.     

Polygyrid relations: a phylogenetic analysis of 17 subfamilies of land snails (Mollusca: Gastropoda: Stylommatophora)

The family of polygyrid land snails in North America is significant for its sympatric shell convergences, diversity of mating systems and complex zoogeography; its monophyly and its relation to other families has long been questionable. Cladistic analysis has been performed on one representative each of 17 subfamilies, including all three polygyrid subfamilies and one or more subfamilies each from all ten of the families that have been proposed as the polygyrid sister groups. Eighteen anatomical synapomorphies are used, of which eight are newly discovered, four are differently assessed from previous studies, and six are traditional. The resulting strict consensus tree of alternative maximum-parsimony cladograms is: (Acavidae (Ammonitellidae Corillidae ((Discidae Oreohelicidae) (Helminthoglyptidae Bradybaenidae Polygyridae (Thysanophoridae (Camaenidae Sagdidae)))))).
According to this working hypothesis, the Polygyridae are monophyletic, and their sister group remains unresolved, although the Acavidae, Ammonitellidae, Corillidae, Discidae and Oreohelicidae can be ruled out. Of the five classifications of stylommatophoran families that have been proposed in the past 12 years, the consensus tree is closest to that of Nordsieck. For future morphological work, three regions are recommended as potentially rich in unused phylogenetic information: the fertilization pouch-seminal receptacle complex, the ureter at the pneumostome and the ventral-chain ganglia. Simultaneous dissection, wkh side-by-side comparisons, is recommended over other methods for detecting homologies in land snails. Molecular characters should be exploited, because of the scarcity and the frequent homoplasy of morphological characters.     

Indications of diverse behavioural ecologies in the morphologically conservative Australian land snails Pommerhelix and Meridolum (Stylommatophora: Camaenidae)

The behavioural ecology of terrestrial molluscs is poorly known. We consider the behavioural ecology of three morphologically similar, parapatric camaenids of southeast Australia: Meridolum corneovirens, Meridolum middenense, and Pommerhelix duralensis. We critically review available literature on their behaviour and address knowledge gaps through field and laboratory studies, including spool-line tracking data for P. duralensis and M. corneovirens. These data are supplemented by in situ observation for each species and ex situ dietary preference studies for P. duralensis and M. corneovirens. Our results suggest a complex behavioural ecology for each species, with some inter-specific behavioural differences within the morphologically conservative group. Implications for invertebrate conservation planning are briefly discussed.     

Lessons from Timor: Shells are poor taxonomic indicators in Asperitas land snails (Stylommatophora,Dyakiidae)

The limacoid land snail Asperitas Gude, 1911 encompasses several vividly coloured, yet poorly known species that are distributed throughout the Lesser Sunda Islands, Sulawesi, and the Moluccas. We examined the variation in shell morphology, including the use of geometric morphometrics, and reproductive anatomy of snails from Timor and several adjacent islands. These studies revealed that none of the taxa described from Timor and considered to be endemic to this island could be reliably distinguished from any of the others. In addition to the systematic ambiguity of morphological characteristics, we uncovered rather low amounts of genetic differentiation in the mitochondrial marker cytochrome c oxidase subunit I that was not consistent with the current taxonomy. Based on these observations, we conclude that there is just a single Asperitas species on Timor that exhibits notable variability in shell characters and body colour. This variability is ascribed to the combined effects of polymorphic colour patterns with locally varying selection in different habitats and along altitudinal gradients. Individuals from the adjacent islands of Flores, Sumbawa, Solor and Romang, which are deemed to represent distinct species-level taxa, exhibit similar amounts of variation in shell and body colour. They exhibit a consistent reproductive anatomy and overall negligible amounts of mitochondrial differentiation from the populations on Timor. Patterns in shell variation do not lend support to previously held ideas that different Sunda Islands harbour distinct radiations of polytypic Asperitas species. By contrast, we suggest that the taxonomic diversity of Asperitas has been overestimated for the over-reliance on shell features and that many presently accepted taxa are likely mere synonyms of fewer and more widespread species.     

Molecular phylogeny and evolution of the AsianAconitum subgenusAconitum (Ranunculaceae)

Classification of the genusAconitum (Ranunculaceae) has long been considered quite difficult because its species show high levels of morphological and ecological variability. The molecular phylogeny of Asian aconites,Aconitum subgenusAconitum was, therefore, studied based on RFLP and sequences of the intergenic spacer between thetrnL (UAA) 3′ exon andtrnF (GAA), and of thetrnL intron, of the chloroplast DNA. UsingAconitum subgenusLycoctonum as an outgroup, we obtained a statistically reliable molecular tree composed of six clades branched radiatively at the very base. There are three clades of Japanese aconites, a single clade of the species of Yunnan and Himalayas, and two clades of Siberian plants. All the tetraploid taxa of Japan we studied did not show any difference based on the molecular characters analyzed, though they have been classified into many taxa. Evolution and phytogeography of the Asian aconites as well as the phylogeny are discussed.     

Molecular phylogeny and morphological evolution of the Acantharia (Radiolaria)

Acantharia are ubiquitous and abundant rhizarian protists in the world ocean. The skeleton made of strontium sulphate and the fact that certain harbour microalgal endosymbionts make them key planktonic players for the ecology of marine ecosystems. Based on morphological criteria, the current taxonomy of Acantharia was established by W.T. Schewiakoff in 1926, since when no major revision has been undertaken. Here, we established the first comprehensive molecular phylogeny from single morphologically-identified acantharian cells, isolated from various oceans. Our phylogenetic analyses based on 78 18S rDNA and 107 partial 28S rDNA revealed the existence of 6 main clades, sub-divided into 13 sub-clades. The polyphyletic nature of acantharian families and genera demonstrates the need for revision of the current taxonomy. This molecular phylogeny, which highlights the taxonomic relevance of specific morphological criteria, such as the presence of a shell and the organisation of the central junction, provides a robust phylogenetic framework for future taxonomic emendation. Finally, mapping all the existing environmental sequences available to date from different marine ecosystems onto our reference phylogeny unveiled another 3 clades and improved the understanding of the biogeography and ecology of Acantharia.     

Light and electron microscopy study of the salivary gland secretory cells of Helicoidea (Gastropoda, Stylommatophora)

We have studied the secretory goblet-cells of the salivary gland of six species of Helicoidea: Elona quimperiana, Trissexodon constrictus, Hygromia limbata, Cernuella aginnica, Cepaea nemoralis and Helix aspersa, using light microscopy and transmission electron microscopy. In each of the species studied, we have been able to demonstrate the presence of four cell types. Following a comparative study with the goblet-gland cells present in the six species in question, and the comparison of our data with previously published reports on the entire set of Stylommatophora, we have established the homologies corresponding to the cell types observed. Our hypotheses were based primarily on the morphology of the rough endoplasmic reticulum and of the secretory vesicles. Accordingly, we have defined five cell types for these six species. The first three cell types, A, B and C, appear in the six species studied and seem to be present in all the stylommatophores. In order to standardize the terminology used by the different authors, we propose that these cell types be called: 'swollen RER cisterns mucocyte', 'granular mucocyte' and 'alveolar cell', respectively. The D-cell type or 'basophilous cell' is present only in Hygromia limbata, Cernuella aginnica, Cepaea nemoralis and Helix aspersa. The E-cell type or 'vacuolated cell' appears only in Elona quimperiana and Trissexodon constrictus.     

Molecular phylogeny and plumage evolution in gulls (Larini)

We used DNA sequence data of the mitochondrial control region and cytochrome b gene to assess phylogenetic relationships among 32 gull species and two outgroup representatives. We tentatively estimated divergence times from transversional substitutions calibrated against DNA–DNA hybridization data. Several strongly supported species groups are identified, but the relationships between these species groups and the rooting of the gull tree remain unresolved. Geographical range extension appears as a factor of speciation, but several related, well‐differentiated species seem to have evolved within comparatively restricted areas. Some plumage characters used in the past for delimiting species groups appear inappropriate. The dark hooded species, for instance, do not constitute a natural assemblage. Molecular data also allowed the identification of several striking plumage convergences that had obscured the true relationships between gull species until now. For example, the dark tropical gulls analysed here each belong to totally different clades and are independent examples of convergent plumage evolution under common environmental constraints. The reverse situation also happened, with two arctic‐distributed species, the ivory gull (Pagophila eburnea) and the Sabine’s gull (Xema sabini), appearing as sister taxa despite completely different plumage features. Molecular data have thus significantly improved our understanding of gull evolution.     

Molecular phylogeny and character evolution of the chthamaloid barnacles (Cirripedia: Thoracica)

The Chthamaloidea (Balanomorpha) present the most plesiomorphic characters in shell plates and cirri, mouthparts, and oral cone within the acorn barnacles (Thoracica: Sessilia). Due to their importance in understanding both the origin and diversification of the Balanomorpha, the evolution of the Chthamaloidea has been debated since Darwin's seminal monographs. Theories of morphological and ontogenetic evolution suggest that the group could have evolved multiple times from pedunculated relatives and that shell plate number diminished gradually (8→6→4) from an ancestral state with eight wall plates surrounded by whorls of small imbricating plates; but this hypothesis has never been subjected to a rigorous phylogenetic test. Here we used multilocus sequence data and extensive taxon sampling to build a comprehensive phylogeny of the Chthamaloidea as a basis for understanding their morphological evolution. Our maximum likelihood and Bayesian analyses separate the Catophragmidae (eight shell plates and imbricating plates) from the Chthamalidae (8-4 shell plates and no imbricating plates), but do no support a gradual reduction in shell plates (8→6→4). This suggests that evolution at the base of the Balanomorpha involved a considerable amount of homoplasy.     

Molecular phylogeny of the scorpionflies Panorpidae (Insecta: Mecoptera) and chromosomal evolution

Panorpidae is the most species-rich family in Mecoptera with ca. 470 species in the Northern Hemisphere. However, the intergeneric phylogenetic relationships of Panorpidae remain unsatisfactorily resolved to date. Here, we used molecular and cytogenetic approaches to determine the phylogenetic relationships of Panorpidae in the evolutionary scenario of chromosomes, and estimated their divergence times using fossil-calibrated Bayesian analysis. In total, 89 species representing all seven genera of Panorpidae were used to reconstruct the phylogenetic trees using maximum parsimony, maximum likelihood and Bayesian inference based on the nuclear 28S rRNA and mitochondrial cox1 and cox2 genes. The results reveal that Panorpidae is a well-supported monophyletic group that can be categorized into two major clades. Major Clade I comprises Neopanorpa and Leptopanorpa, and Major Clade II consists of all the other genera (Cerapanorpa, Dicerapanorpa, Furcatopanorpa, Panorpa and Sinopanorpa). Neopanorpa and Cerapanorpa are regarded as paraphyletic groups for the first time. BEAST analysis indicates that Panorpidae originated in the Lower Cretaceous approximately 122.5 Ma (96.8–149.3 Ma), and that most diversification occurred from the Selandian (59.8 Ma) to the Middle Pleistocene (0.6 Ma) in the Cenozoic. Cytogenetic data plotted on the cladogram show that the lineage differentiation of Panorpidae is closely related to the chromosomal evolution, especially the reduction of chromosome number. Our study suggests that a taxonomic revision of Panorpidae is urgently needed at the generic level.