The spermatheca in the land snail, Arianta arbustorum (Pulmonata: Stylommatophora): muscle system and potential role in sexual selection

Abstract. Previous studies have shown variable patterns of paternity after multiple mating, and also variation in sperm storage among individuals of Arianta arbustorum , which suggests that the spermatheca may influence paternity in this promiscuous land snail. To identify possible morphological correlates of sperm manipulation, we investigated arrangement and ultrastructure of the muscles of the spermatheca. The musculature surrounding the 2–9 spermathecal tubules is arranged in a complex three dimensional network. In addition, each tubule has a thin sheath in which longitudinally oriented cells make up the innermost layer. Usually, the smooth muscle cells are enclosed by connective tissue. Only occasionally is direct muscle-muscle contact established through dense plaques. The short thick filaments, their small diameter, the relatively weak development of the tubular system and sarcoplasmic reticulum, and the low density of mitochondria indicate that the muscle cells contract relatively fast but with little strength, that they recover slowly, and have low endurance. A single muscle cell may be innervated by several axons and one axon may contact several muscle cells. Combining evidence of the present paper and a foregoing investigation on the spermathecal epithelium, we suggest that the main function of the spermathecal muscles is to expel sperm stored for fertilization, while the ciliation of the common duct is probably responsible for the distribution of sperm among the tubules.     

The pedal muscle of the land snail Megalobulimus oblongus (Gastropoda, Pulmonata): an ultrastructure approach

M. Cristina Faccioni-Heuser, Denise M. Zancan, Christiane Q. Lopes and Matilde Achaval. 1999. The pedal muscle of the land snail Megalobulimus oblongus (Gastropoda, Pulmonata): an ultrastructure approach. — Acta Zoologica (Stockholm) 80: 325–337
The ultrastructure of the pedal muscle of the Megalobulimus oblongus is described. This muscle consists of transverse, longitudinal and oblique bundles ensheathed in collagenous tissue. Each muscle cell is also ensheathed by collagen. The smooth muscle cells contain thin and thick filaments; the thin filaments are attached to dense bodies. These cells contain a simple system of sarcoplasmic reticulum, subsarcolemmal caveolae and mitochondria with dense granules in the matrix, and glycogen. Three types of muscle cells were identified. Type A cells exhibited densely packed myofilaments, abundant glycogen rosettes, numerous mitochondria and sarcoplasmic reticulum profiles. Type B cells exhibited scanty glycogen and mitochondria, few cisternae of sarcoplasmic reticulum and large intermyofibrillar spaces. Type C cells exhibited intermediate characteristics between type A and type B cells. Neither nexus nor desmosomes were observed between the muscle cell membranes. The muscle contains well developed connective tissue and blood vessels. These structures and the distribution of muscle cells are probably involved in the muscular-hydrostat system. The muscle is richly innervated, having neuromuscular junctions with clear and electron-dense synaptic vesicles. The clear vesicles probably contain acetylcholine because the axons to which they are connected arise from acetylcholinesterase positive neurones of the pedal ganglion. The other vesicles may secrete monoamines such as serotonin and/or neuropeptides such as substance P.     

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.     

Variation of distal genitalia in the simultaneously hermaphroditic land snail Arianta arbustorum (Pulmonata,Stylommatophora) caused by sexual selection?

The most likely explanation for genitalic extravagance may be sexual selection acting either before, during, or after copulation. In promiscuous species, the intensity of postulating sexual selection may be a function of population density. In this study we examined the variability of the distal genitalia involved in spcrmatophore production, reception and manipulation of 113 adult individuals of the simultaneously hermaphroditic land snail Arianta arbustorum (L.) from six natural populations in the Eastern Alps (Gesause, Austria). We investigated the hypothesis that these genitalia increase in relation to shell size with increasing population density (range: 0.9 to 39.8 individuals/m²) and expected a higher variance of the genitalia compared to shell dimensions due to sexual selection. Genitalic size was unexpectedly inversely related to population density, probably due to increased inhibitor)^- effects of snail mucus. Patterns of variation of female and male characters did not differ. Coefficients of variation of the genitalia were significantly higher than those of the shell dimensions as predicted. This was due to a higher dispersion around the regression lines rather than higher allometric values. However, the influence of sexual selection on genitalic size and variance cannot be unambiguously determined. We discuss different scenarios emphasizing the importance of sexual selection to differing degrees and conclude that sexual selection has probably played only a minor role.     

Dart shooting influences paternal reproductive success in the snail Helix aspersa (Pulmonata, Stylommatophora)

Although animal courtship behaviors are generally understoodwithin the context of sexual selection, the relevance of manysexual behaviors to sexual selection, and vice versa, remainsunexplained. For example, the adaptive function of the "lovedart" used in the precopulatory behavior of hermaphroditicland snails is only now becoming apparent. Contrary to previous assumptions, dart shooting is unlikely to function as a stimulusfor copulation. In searching for a more ultimate explanationof the dart's function, we tested whether variation in dartshooting influences reproductive fitness in Helix aspersa.Individual mother snails were mated sequentially to two potentialfathers. Dart shooting was closely observed and quantifiedfor all pairings, and percentages of offspring sired by each potential father were determined using allozymes. The resultsindicate that snails that shoot darts effectively have significantlygreater paternal reproductive success than snails that shootpoorly. In contrast, there was no significant effect of matingorder on either dart shooting or paternal reproductive success.     

Shell thickness in the land snail Theba pisana (Pulmonata: Helicidae)

Shell thickness in Theba pisana was measured in samples from Britain, northern France and the Mediterranean region. Mean adult thickness varied between about 0.08 mm and 0.20 mm. These values are much lower than previously published values. The considerable variation within samples was related to shell size, periods of growth, and resorption of shell material during breeding. Variation between samples was not great. This contrasts with previous work which linked shell thickness variation to large-scale climatic effects. Perhaps availability of calcium may be more important than climate in determining shell thickness in Theba Pisana .     

Systematics and distribution of the land snail Pene (Pulmonata: Enidae) in Israel

Formerly considered a subgenus of Buliminus, Pene is here, for reasons given, considered as a separate genus. Four species ( syriacus, sidoniensis, galilaea and auriculata ) inhabit Israel. A new subspecies, syriacus ramimensis , is described from north-east Galilee.
Northwards, towards the borderline with s. syriacus , the ratio mouth height/mouth diameter of s. ramimensis becomes first gradually larger, then reverses abruptly. The northernmost s. ramimensis sample is in a peculiar intermediate position suggesting that it represents a syriacus-ramimensis hybrid population which has reached stabilization.
Distribution borders are usually correlated with rain or substratum. The sidoniensis/galilaea borderline is unusual in that it crosses an area of nearly uniform climate, substratum and vegetation. Along it sidoneinsis and galilaea are found within centimetres of each other, suggesting that the borderline is here determined by interspecific competition rather than by physical factors.     

Phylogeny of the three brown banded land snail genus Humboldtiana (Pulmonata: Humboldtianidae)

The three brown banded land snail genus Humboldtiana (Pulmonata: Humboldtianidae) comprises 50 species of insular distribution from southern Texas to central México; its low vagility and dispersal potential are reflected in high levels of endemism and highly restricted distributions. A current scheme of classification recognizes six subgenera: Polyomphala, Oreades, Gymnopallax, Clydonacme, Aglotrochus, and Humboldtiana (the latter containing three species groups). In the present work, mitochondrial 16S rDNA and partial ITS region and ribosomal large subunit (5.8S+ITS-2+28S) sequences from 26 recognized species and 2 populations of unknown identity were studied to assess phylogenetic relationships of Humboldtiana species using parsimony and Bayesian analysis, and AU test was used to compare the different phylogenetic hypotheses. The same four major clades were recovered by both methods (buffoniana, queretaroana, montezumae and fortis), although none of them support the current scheme of classification, with the only exception being the subgenus Gymnopallax included in the clade buffoniana. The results suggest that speciation patterns observed in the genus Humboldtiana could be explained as a morphostatic radiation.     

Allozymic differentiation in the land snail Arianta arbustorum (Stylommatophora, Helicidae): historical inferences

The intraspecific genetic differentiation depends to a large part on historical factors. The mode of dispersal is an important determinant of the genetic diversity especially of newly founded populations. Fast long-distance colonization of formerly unsuitable habitats results in reduced genetic diversity and lineages originating from different glacial refugia form well-defined hybrid zones, where they meet. Slow dispersers, on the other hand, preserve their refugial genetic diversity. Paleontological findings and a previous analysis of mtDNA suggest that the land snail Arianta arbustorum, which today is widely spread across Europe, has survived the Pleistocene glaciations within the boundaries of permafrost. In the present paper, we asked, whether the differentiation of nuclear markers, i.e. allozymes, supports these results. We investigated the allozymic differentiation at 15 polymorphic loci in 14 Alpine and one English populations of the land snail A. arbustorum. Various tests of genetic population differentiation yielded ambiguous results. Allele frequencies differed significantly across populations and all but five pairwise F_ST values were significant. Genetic distances between populations were comparatively high. On the other hand, an exact test of population differentiation found no differences and the assignment of individuals to their populations of origin was poor. This indicates that allozymic differentiation is constrained. Whether these constraints are of natural or methodological nature remains open. Matrices of genetic and geographical distances were correlated only after removal of the English population from the analysis. Similarly, with one exception F_ST values increased as the number of regions included in the calculations increased. Thus, on a regional scale, we largely observed a pattern of isolation by distance, which does no longer hold on a larger scale. This is well in accordance with the previous findings based on mtDNA. Mitochondrial lineages were widely distributed across Europe and overlapped extensively. Considering the low potential for dispersal of a land snail, the patchy distribution of lineages, and the high genetic diversity also found in the present study suggests that A. arbustorum has survived the Pleistocene within the boundaries of permafrost.     

Self-fertilization and cross-fertilization in the land snail Arianta arbustorum (Mollusca, Pulmonata: Helicidae)

The simultaneously hermaphroditic land snail Arianta arbustorum (L.) (Pulmonata: Helicidae) was long believed to be self-incompatible. In the present study, however, uniparental reproduction was observed under laboratory conditions. Snails from two natural populations and from laboratory culture reared in isolation from the subadult stage produced fertile eggs. Over a period of three years after maturation, 38.5% of the unmated snails produced fertile eggs, mostly in the second and third years instead of the first year. Egg production and hatching success (the proportion of eggs that hatched) were lower in unmated snails than in mated snails. The number of hatchlings produced by unmated snails was 1–2% of that produced by mated snails. Hatchlings from unmated snails survived as well as those from mated snails under laboratory conditions. The result indicate that A. arbustorum can self-fertilize but with a great fitness reduction.     

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.     

Radiation of the land snail genus Sagda (Pulmonata: Sagdidae): comparative morphology, biogeography and ecology of the species of north-central Jamaica

The radiation of the endemic Jamaican land snail genus Sagda is examined through analysis of the comparative shell morphology, anatomy, ecology and biogeography of the four species inhabiting north-central Jamaica: S. centralis sp. nov. , S. spei, S. bondi and S. montegoensis. The species are all rather similar in their morphology and ecology. The shells show some differences in the presence or absence of internal lamellae, the position of the lamellae, the form of the depression in the centre of the base, the regularity of sculpture (low ribs) and the pattern of microsculpture. Slight differences in the form of the radular teeth are found between some species. The structure of the secondary ureter is peculiar: in all species it is open shortly behind the mantle collar, then branches into two closed tubes which open separately on the outside of the mantle collar. Despite its great complexity, the reproductive system is rather uniform among species; differences are seen in the number of pockets that form spines on the spermatophore. The complex, three-branched spermatophore is formed within the entire epiphallar branch of the penis, whereas the appendicular branch apparently assumes the copulatory function. When inactive, all species are found associated with rocks or, less commonly, among leaf litter. Activity occurs primarily at night and mostly on leaf litter rather than rocks. The snails eat dead plant material, including a variety of plant species. All species (with the possible exception of S. montegoensis) are ovoviviparous. They overlap broadly in the range of climates occupied. The geographic ranges of the four species are overlapping, although Sagda bondi has a parapatric distribution in relation to the western species S. grandis. Fossil and subfossil evidence shows that shifts in the ranges of the species have occurred in the late Quaternary; Pleistocene-Holocene environmental changes, habitat destruction by man, and possibly other environmental changes during historical times are probably involved. Radiations of other land snail groups on tropical islands have generally produced greater morphological and ecological differentiation than in Sagda. Allopatric or parapatric species distributions appear to be more common in other land snail radiations.     

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.     

Some aspects of the breeding biology of the equatorial land snail Limicolaria martensiana (Achatinidae: Pulmonata)

A study of the breeding season of the polymorphic equatorial land snail Limicolaria martensiana Smith (Achatinidae: Pulmonata) lasting six years and six months in populations in the Kampala District (lat. 0 20') and the Equator line shows that the snail breeds continuously throughout the year but has bimodal peaks in its breeding which occurs during the dry seasons, while during the rainy seasons breeding is low. It is the decrease in the amount of rainfall which induces peak breeding and as a result there is an inverse relationship between the amount of rainfall and the breeding intensity. The evolution of ovoviviparity with the ability to retain eggs and/or young in utero during aestivation is the phenomenon that enables L. martensiana to have peak breeding during the dry seasons. Moisture which enables young to feed and thus grow is an ultimate factor in determining breeding in L. martensiana . The gestation period is 10 to 14 days during the rainy seasons but the snail retains its eggs and/or young in utero during the dry seasons and only deposits them when rain returns. The eggs take 18.6±5.43 days ( N= 130) to hatch after they are laid. Hatching in utero and outside occurs by the young eating the egg shells. Oogenesis in the snails occurs at all times. L. martensiana stores viable allosperms which it can use for further reproduction.     

The ultrastructure of the epicardium epithelium of the slug, Arion ater (Linnaeus, 1758), (Pulmonata, Stylommatophora)

In the present paper we investigated the fine structure of terrestial slug Arion ater (L.) Interacisternal microtubules were formed in Arion ater, however they do not correspond to interacisternal polycylinders because they differ in size and they are not continuous with the membrane cistern. From our study it may be concluded that these intracisternal microtubules suggest an accumulation of protein.     

A histochemical study of the secretions of reproductive glands and of the egg envelopes of Achatina fulica (Pulmonata: Stylommatophora)

Histochemical investigations of the secretions of reproductive glands—albumen gland, apical uterus, basal uterus and prostate gland—indicate the presence of galactogen in the albumen gland, acid mucopolysaccharide in the apical uterus, and lipoprotein in the basal uterus and prostate gland of A. fulica. The proteinaceous secretions produced by the glands do differ in their terminal reactive sites. Intense alkaline phosphatase reaction is found in albumen gland and apical uterus; carbonic anhydrase activity could be detected mainly in the uterine glands. The cyclical secretory activity of the reproductive glands has been studied preparatory to egg-laying and in the spent phase. Histochemical characteristics of the egg envelopes—albumen, shell membrane and egg-shell—suggest a possible sequential deposition of glandular products during the descent of eggs through the repv. ductive tract. The factors contributing to the stability and resistant nature of the egg envelopes, and the possible role of nutritive materials contained in reproductive gland secretions in the development of the embryo (even while the eggs are inside the uterus), are discussed.     

Morphological modifications in stored heterospermatozoa of Oxyloma elegans (Pulmonata: Stylommatophora)

The authors describe a peculiar feature of the sperm of Oxyloma elegans (Pulmonata: Stylommatophora) which consists in a paracrystalline body encircling the apical portion of the nucleus. This body has been seen to vanish when spermatozoa are stored in the seminal receptacles after copulation. The meaning of this phenomenon and its possible relationship to capacitation are discussed.