Comparative spermatozoal ultrastructure and molecular analysis in dromiid crabs and their phylogenetic implications for Dromiidae and Podotremata (Decapoda: Brachyura)

We described the spermatozoal ultrastructure and conducted a molecular analysis of Dromiidae Hypoconcha parasitica, Hypoconcha arcuata, Moreiradromia antillensis and Dromia erythropus. To elucidate the relationship between the different species of this brachyuran group, we also compared the spermatozoal morphologies and phylogenetic positioning among species of Dromiidae, Dromioidea and Podotremata. Specimens were collected from the northern coast of São Paulo, Brazil and were fixed and processed followed by transmission electron microscopy and molecular analysis routines. The Dromiidae spermatozoa studied are characterized by a discoidal acrosome, with three or four concentric zones, which are centrally separated by a bilaterally capitate perforatorial chamber, with a “mushroom”-shaped apex in the Hypoconchinae and a “T-shape” in Dromiinae. Above the perforatorial chamber, there is an apical protuberance, continuous with the subopercular region and the operculum, which forms a low, centrally perforated dome. Under differential interference contrast microscopy, the spermatozoa show 3 to 4 radial arms. The spermatozoal characters in Hypoconchinae and Dromiinae do not separate these subfamilies from the Dromiidae and Dromioidea. Ultrastructural differentiation was only found between representative Dromioidea and other Podotremata. Thus, the spermiotaxonomy of these Hypoconcha, Moreiradromia and Dromia species corroborated previous morphological and molecular studies, supporting the monophyly of Dromiidae and Dynomenidae in relation to Homolidae and Latreilliidae.     

Not only for males: the pleon‐holding mechanism in female calappids (Crustacea: Decapoda: Brachyura)

Adult female Calappidae were examined with light microscopy and microcomputed tomography for the presence of a pleon‐holding mechanism. A typical press‐button consisting of a protrusion on the fifth thoracic sternite and a socket on the sixth pleon segment was found in species of the genera Calappa, Calappula, Cycloes, Mursia, and Paracyclois. Within the genus Calappa, 15 of 17 species showed a complete press‐button mechanism, whereas in two species only a sternal button or none of the holding structures could be observed. The functionality of the mechanism was assessed by manually flexing and opening the pleon. This revealed a lack of function in species with fully developed holding structures, while others exhibited full function. Additionally, signs of abrasive wear could be identified with scanning electron microscopy on the surface of the holding structures of Calappa hepatica. The documentation of pleon‐holding structures of varying degrees of functionality in adult female eubrachyurans provides new insight into a morphological characteristic known exclusively from Brachyura.     

Evolutionary morphology of the hemolymph vascular system in hermit and king crabs (Crustacea: Decapoda: Anomala)

The morphological transformation of hermit crabs into crab‐like king crabs in the evolution of decapod crustaceans represents a remarkable case of carcinization or evolutionary shaping into a crab‐like form. In this study, we focus on internal organs such as the hemolymph vascular system and adjacent anatomical structures of several Recent hermit crab (Paguridae) and king crab (Lithodidae) species. There are various correspondences in the morphology of the arterial systems in the dorsal cephalothorax of the two taxa, especially with regard to the anterior aorta, anterior lateral arteries, and hepatic arteries. In the pleon, the posterior aorta in both taxa displays a proximal bifurcation and follows an asymmetrical course. The ventral vessel system, on the other hand, which mainly supplies the limbs, differs significantly between the taxa, with pagurids displaying the plesiomorphic condition. The pattern of the ventral vessel system in Lithodidae is influenced by morphological transformations of integumental structures during carcinization. One of these transformations was the broadening of the sternites, which resulted in a widening of the space between the endosternites. In addition, changes in the morphology of the endophragmal skeleton in Lithodidae led to an increase in the potential for intraspecific variability and interspecific variation in the arterial branching pattern. J. Morphol., 2013. © 2013 Wiley Periodicals, Inc.     

Female reproductive morphology of coral‐inhabiting gall crabs (Crustacea: Decapoda: Brachyura: Cryptochiridae)

Gall crabs are obligate associates of stony corals in which they induce skeletal modifications. In some cryptochirid species, females live in open depressions accessible to males; while in others, females are rather isolated in semiclosed galls, which necessitates elaborate sperm storage capabilities by the female. In this study we investigate the female gross morphology and reproductive systems of Fungicola syzygia lodged in semiclosed flattened pits, Opecarcinus cathyae with semiopen pits and Pseudocryptochirus viridis from shallow open depressions using line drawings and histological methods. The general morphology of the cryptochirids' reproductive systems is uniform and conforms to other thoracotreme brachyurans: paired muscular vaginae of the concave pattern lead from the sternal gonopores into paired seminal receptacles where sperm is stored. The seminal receptacle is internally lined by distinct types of epithelia: a cuticle underlined by a columnar epithelium ventrally, a monolayered secretory epithelium dorsally and a multilayered transfer tissue where the oviducts enter the seminal receptacle. In all studied specimens, the seminal receptacle contained free spermatozoa; however, in specimens of Pseudocryptochirus viridis it also contained spermatophores, indicating a recent insemination. In contrast to most other brachyurans ovaries of the investigated cryptochirids extend into the pleon. The specific degree of ovary extension differs between the studied species and is closely related to female body shape.     

Phylogenetic analysis of the Mexican freshwater crabs of the tribe Pseudothelphusini (Decapoda: Brachyura: Pseudothelphusidae)

The Mexican pseudothelphusid crabs are classified in one subfamily, three tribes, and 13 genera. Up to now, 56 species have been recognized, distributed in a strictly Neotropical pattern, with some of them reaching the state of Sonora on the western slope of Mexico. The tribe Pseudothelphusini is the most diverse, with five genera and 35 species, all of them endemic to Mexico: the two most species‐rich genera are Pseudothelphusa, with 23 species, and Tehuana, with eight species; Epithelphusa includes two species, whereas Disparithelphusa and Smalleyus are monotypic. The Pseudothelphusini lack an updated systematic revision, which could serve as a framework to analyse the monophyletic origin of the group, to clarify the relationships among genera and species, as well as to resolve the taxonomic status of various species complexes. In the present study, an exhaustive morphological revision was conducted using somatic and sexual characters. A phylogenetic analysis was performed using 77 characters and 183 character states, taken from 41 species. Ten trees of the same length were obtained using PAUP 4.0 through a heuristic search. The results show that the tribe as it is actually known constitutes a paraphyletic group, in which the species of Epithelphusa and Pseudothelphusa puntarenas are excluded from the internal group. According to the obtained results, the tribe Pseudothelphusini s.s. includes five genera: Smalleyus, Pseudothelphusa, Tehuana, and two new ones to accommodate Pseudothelphusa galloi and Pseudothelphusa sulcifrons, respectively. This new arrangement considers the provisional suppression of the genus Disparithelphusa, which remained as another species of Pseudothelphusa throughout the cladistic analysis. The phylogenetic results show a strong congruence with the distribution of the species, in several cases grouping species that form morphological clines along a geographical gradient. The previously proposed southern origin of the tribe Pseudothelphusini gains support with the results obtained. © 2010 The Linnean Society of London, Zoological Journal of the Linnean Society, 2010, 160 , 457–481     

Global diversity of crabs (Crustacea: Decapoda: Brachyura) in freshwater

An assessment of the global freshwater crab diversity is presented. A total of 1,476 species in 14 families are currently known from all zoogeographical regions (except Antarctica), including 1,306 species in eight exclusively freshwater families (Pseudothelphusidae, Trichodactylidae, Potamonautidae, Deckeniidae, Platythelphusidae, Potamidae, Gecarcinucidae and Parathelphusidae). Estimates of true freshwater crab diversity including likely numbers of undescribed taxa suggest that the field remains largely in a “discovery” phase. Main ideas on the origins, diversification, and phylogeny of true freshwater crabs are briefly discussed. The economic importance of freshwater crabs is also highlighted. Guest editors: E. V. Balian, C. Lévêque, H. Segers & K. Martens Freshwater Animal Diversity Assessment     

Ultrastructure of the annual cycle of female sperm storage in spermathecae of the torrent salamander, Rhyacotriton variegatus (Amphibia: Rhyacotritonidae)

This study is the first report on the ultrastructure of the sperm storage glands (spermathecae) in the salamander Rhyacotriton variegatus. The population studied is associated with cold-water, rocky streams of the redwood (Sequoia) zone in northern California. Males possess sperm in their vasa deferentia and undergo spermiation throughout the year, but mating is seasonal. Most females with large, vitellogenic follicles (2.0-3.9 mm mean dia.) collected from February-June contain sperm in their spermathecae, although some females with large follicles lack sperm. Other mature-size females collected during this period have small ovarian follicles (0.9-1.2 mm mean dia.) and lack stored sperm. All females collected from September-November have small follicles (0.6-1.6 mm mean dia.) and lack sperm, except in one instance in which a female collected in November had a small amount of degraded sperm, apparently retained from the previous breeding season. The spermathecae consist of simple tubulo-alveolar glands in which the neck tubules produce a mucoid secretory product, and the distal bulbs, where sperm are stored, contain secretory vacuoles of uniform density that stain positively for glycosaminoglycans. In specimens containing sperm, some bulbs have abundant sperm and others lack sperm, but the ultrastructure is similar in both conditions. The acini contain columnar epithelial cells with wide intercellular canaliculi, and a merocrine process releases the secretion. Spermiophagy occurs. In specimens from spring and summer with small ovarian follicles, the neck tubules are similar to those of breeding females, but the distal bulbs are reduced to cords of cells lacking a discernible lumen. Secretory activity in the distal bulbs is initiated in the fall. Spermathecae of R. variegatus are most similar to those of a stream-dwelling plethodontid, Eurycea cirrigera.     

Morphology and function of the female reproductive system of Ebalia tumefacta (Decapoda,Brachyura, Leucosiidae)

In this article, the morphology and function of the female reproductive organs of Ebalia tumefacta were investigated using histological methods. While the vagina conforms to the concave type, the study reveals a new orientation of seminal receptacle compartments. The seminal receptacle consists of two chambers, which are oriented in anterior‐posterior direction. This is in contrast to the dorso‐ventral orientation of seminal receptacle chambers in all other known brachyuran crabs. The anterior chamber is lined by cuticle, whereas the posterior chamber is covered with a holocrine glandular epithelium. The oviduct connection is located ventrally, close to the opening of the vagina. The oviduct orifice is characterized by a transition of the epithelium lining of the oviduct to the seminal receptacle holocrine glandular epithelium. Special features are muscle fibers, which are attached to the oviduct orifice and to the sternal cuticle as well. The muscle fibers can be found exactly at that point where the oviduct opens into the seminal receptacle and are secondly attached to the sternum beneath. This musculature is newly described for Eubrachyuran crabs. This musculature can be interpreted as an important feature in the fertilization and egg‐laying process in relation to supporting and controling the inflow of eggs into the seminal receptacle lumen. These new discoveries were compared to the known pattern of an Eubrachyuran seminal receptacle. J. Morphol. 276:517–525, 2015. © 2014 Wiley Periodicals, Inc.     

Evolution of sperm morphology in potamid freshwater crabs (Crustacea: Brachyura: Potamoidea)

We investigated sperm cells and spermatophores of four species of Old World freshwater crabs belonging to three different genera of the subfamily Potaminae (family Potamidae). Characters previously believed to be apomorphic for the potamid subfamily Potamiscinae were also found to occur in the Potaminae. To infer the morphological ancestral character state combination of the Potamidae, ancestral character state analysis of four different sperm traits was performed, based on a 16S rDNA phylogeny of the investigated species. Comparing molecular phylogeny and character state distribution, several cases of convergent evolution could be identified. The densely packed, coenospermic spermatophores and the occurrence of a ‘tongue‐and‐groove’ connection between operculum and acrosomal zones are probably apomorphies for the whole Potamidae. The spermatozoa of Socotrapotamon socotrense show several unique characters. We also analysed the evolution of acrosome size. The sperm cells of the Potamidae and their sister‐group Gecarcinucidae only slightly overlap in acrosome size. Within the investigated species, the ‘East Asia’ subclade (subfamily Potamiscinae) developed significantly larger acrosomes than the subfamily Potaminae. Our results suggest that the use of brachyuran acrosome morphology for phylogenetic inference at the family level is strongly affected by small sample size, and by convergent character evolution. © 2010 The Linnean Society of London, Zoological Journal of the Linnean Society, 2010.     

Prevalence of bacteria in the spermathecae of female snow crab, Chionoecetes opilio (Brachyura: Majidae)

The spermathecal contents of primiparous, multiparous and barren female snow crabs were observed by a scanning electron microscope. Bacteria colonies were observed with a significantly higher frequency in the spermathecae of primiparous and old barren females. Bacteria infect the spermathecae and destroy the spermatophores and spermatozoa inside. These observations suggest that bacteria in the spermathecae do not exclude opportunistic microbes by modifying pH of the medium as suggested in the literature. The prevalence of bacteria in primiparous and old barren females suggests that they infect individuals with a weak anti-microbial protection. The absence of bacteria in the highly acidic seminal fluid derived from males upon copulation suggests that it may provide anti-microbial protection. Bacteria do not seem to be able to survive in an anaerobic environment for a long period.     

Variation in safety factors of claws within and among six species of Cancer crabs (Decapoda: Brachyura)

To better understand how safety factors of biological structures evolve, we examined the frequency of claw failure, and the intra‐ and interspecific patterns of variation in maximum biting force and breaking strength in the claws of six species of Cancer (Linnaeus) crabs that live in sympatrv along the coast of the northeastern Pacific: C. antennarius, C. branneri, C. gracilis, C. maguter, C. oregonensis and C. productus. Although the breakage frequencies in natural populations were similar among species (6%), they were higher than predicted based on failure probabilities calculated from laboratory measurements of biting force and breaking strength for healthy pristine claws. The incidence of claw damage was correlated with the degree of wear, suggesting that claws later in the intermolt interval were more likely to fail. Within species, safety factors increased from 3.1 to 4.6 with increasing instar number due primarily to a decline in muscle stress (force per unit area of apodeme). Surprisingly, the lower maximum muscle stress generated by later instars appeared to be due to behavioral restraint, since it was not accompanied by relatively lower muscle mass. In addition, among individuals of the same claw size, lower breaking forces were correlated with lower maximum biting force, and both were correlated with lighter cuticle and closer muscle mass, suggesting a coupling that maintains a more stable safety factor over the moult cycle. In some species, size‐adjusted maximum biting forces were higher for males than females, but this paralleled differences in breaking strength, so safety factors did not differ between the sexes. Among the six Cancer species, one exhibited an unusually high safety factor (C. oregonensis, 7.4) and another an unusually low one (C. maguter, 2.6). The remaining four species were similar to each other and exhibited an intermediate safety factor (3.6). From a phylogenetic perspective, the species with more extreme safety factors appeared to be derived from a common ancestor with an intermediate safety factor. From an ecological perspective, species more closely associated with rocky substrata, and presumably a higher incidence of hard‐shelled prey, exhibited higher safety factors. But safety factors were also correlated with relative claw size, and sexual dimorphism in claw size. Although we cannot say whether habitat, diet or sexual selection are primarily responsible for the differences in safety factors observed among species, the cost of producing a relatively larger claw seems an unlikely explanation because safety‐factors did not differ between males and females in any of the sexually dimorphic species.     

A light and electron microscopical study of the spermathecae and ventral receptacle of Anastrepha suspensa (Diptera: Tephritidae) and implications in female influence of sperm storage

Female insects with multiple sperm storage organs may potentially influence patterns of paternity by differential storage of sperm from competing males. The Caribbean Fruit Fly, Anastrepha suspensa, stores sperm differentially with respect to its three spermathecae. To understand the mechanisms and processes responsible for patterns of sperm storage and use in A. suspensa, details of the fine structure of female sperm storage organs were resolved by UV-light microscopy, confocal microscopy, tissue sectioning, and scanning and transmission electron microscopy. Structures not previously described for this species include a ventral receptacle for sperm storage and osmoregulation, a conical-shaped valve at the junction between the spermathecal capsules and their ducts, laminar and granular secretions, secretions from the signum, hemocytes surrounding the spermathecae, and spermathecae with sclerotized, hollow projections that terminate in single glandular cells. The independent organization of sperm storage organs, spermathecal ducts, associated musculature, gland cells, and innervation offer possible mechanisms by which sperm movement may be influenced by females. The implications of these structures for insemination and fertilization events are discussed.     

The ecology and biology of Southeast Asian false spider crabs (Crustacea: Decapoda: Brachyura: Hymenosomatidae)

The false spider crabs of the family Hymenosomatidae are one of the most poorly known group of brachyuran crabs in Southeast Asia. This is largely attributed to their small size and cryptic behaviour. Despite the many studies of decapods in Southeast Asia, only eleven species of hymenosomatids are known. Of these, two genera and six species were only described in the last two years. From Singapore the largest number of species (five) has been reported probably because it has been the best explored, while from Thailand three species and Peninsular Malaysia two species are known. There are no published records of hymenosomatids from Borneo or Java. The hymenosomatid fauna is perhaps best known in Australia and New Zealand. It is anticipated that when proper collections are made and studies implemented, the Southeast Asian hymenosomatid fauna will prove to be more diverse than what has been reported. The ecology of the Southeast Asian species is reviewed, especially in the view that two species are completely freshwater, one of which is a troglobite. Most species are littoral or sublittoral in habit and very sensitive to human activities (e.g. pollution).     

New species and new records of pinnotherid crabs (Crustacea: Decapoda: Brachyura) from the Yellow Sea

Five species belonging to two genera of pinnotherid crabs are reported from the Yellow Sea. Two are new to science, namely: Sakaina glabra sp. nov. and Sakaina granulata sp. nov.; while three species are recorded for the first time from China: Pinnaxodes majorOrtmann, 1894, Pinnaxodes mutuensisSakai, 1939, and Sakaina japonicaSerène, 1964. Sakaina glabra can be easily distinguished from congeners by lacking a thick rim of pubescence along the anterolateral margin of the carapace. Sakaina granulata closely resembles S. asiatica (Sakai, 1933), but differs mainly in having a row of granule-like denticles along the anterolateral margin of its carapace.     

Ultrastructure of spermatozoa of orsolobidae (Haplogynae,Araneae) with implications on the evolution of sperm transfer forms in Dysderoidea

Haplogynae are highly diverse with respect to the primary male genital system and sperm characteristics. Additionally, all sperm transfer forms (STF) known for spiders are present. Besides individually transferred sperm (cleistospermia), sperm are transferred as conjugates, both primary (synspermia) and secondary sperm conjugates (coenospermia, rouleaux) occur. Nevertheless, the ultrastructure of spermatozoa and STF are described for few Haplogynae and often only one representative species was studied, resulting in a superficial insight in the evolution of these traits. To elucidate the evolution of STF within Haplogynae we investigated representatives of four genera of the dysderoid family Orsolobidae. Our data show the presence of synspermia (Orsolobus, Osornolobus, Hickmanolobus, and Tasmanoonops) and also cleistospermia (Osornolobus). The occurrence of different STF within one family or even genus has not been described for any other spider taxon so far. Moreover, the synspermia of species of Tasmanoonops and Hickmanolobus were not covered by a secretion sheath suggesting a previously unknown strategy of transferring sperm that is possibly related to sperm residency time or female triggered processes after copulation. Based on serial ultrathin sectioning and subsequent 3D‐reconstruction, we obtained detailed measurements revealing remarkable size differences of STF. To evaluate the previously suggested correlation with the most distal region of the spermophor inside the embolus (intromittent part of the copulatory organ) we measured the diameter of the spermophor using micro‐computed X‐ray tomography data to obtain corresponding morphometric parameters. Based on these data only two species show similarity in STF and spermophor diameter. J. Morphol. 275:1238–1257, 2014. © 2014 Wiley Periodicals, Inc.     

Morphology and evolutionary implications of the annual cycle of secretion and sperm storage in spermathecae of the salamander Ambystoma opacum (Amphibia: Ambystomatidae)

Females of the marbled salamander, Ambystoma opacum, store sperm in exocrine glands called spermathecae in the roof of the cloaca. Eggs are fertilized by sperm released from the spermathecae during oviposition. Some sperm remain in the spermathecae following oviposition, but these sperm degenerate within a month and none persists more than 6 mo after oviposition. Thus, sperm storage between successive breeding seasons does not occur. Apical secretory vaculoes are abundant during the fall mating season and contain a substance that is alcian blue+ at pH 2.5. Production of secretory vacuoles decreases markedly after oviposition, and the glands are inactive by the summer months. Ambystoma opacum is a terrestrial breeder, and some mating occurs prior to arrival at pond basins where oviposition occurs. Mating prior to arrival at the ovipository site may prolong the breeding season, leading to fitness implications for both males and females. Females have opportunities for more matings, and the possibilities for sperm competition in the spermathecae are enhanced. © 1995 Wiley-Liss, Inc.     

Three‐dimensional reconstruction of the odontophoral cartilages of Caenogastropoda (Mollusca: Gastropoda) using micro‐CT: Morphology and phylogenetic significance

Odontophoral cartilages are located in the molluscan buccal mass and support the movement of the radula during feeding. The structural diversity of odontophoral cartilages is currently known only from limited taxa, but this information is important for interpreting phylogeny and for understanding the biomechanical operation of the buccal mass. Caenogastropods exhibit a wide variety of feeding strategies, but there is little comparative information on cartilage morphology within this group. The morphology of caenogastropod odontophoral cartilages is currently known only from dissection and histology, although preliminary results suggest that they may be structurally diverse. A comparative morphological survey of 18 caenogastropods and three noncaenogastropods has been conducted, sampling most major caenogastropod superfamilies. Three‐dimensional models of the odontophoral cartilages were generated using X‐ray microscopy (micro‐CT) and reconstruction by image segmentation. Considerable morphological diversity of the odontophoral cartilages was found within Caenogastropoda, including the presence of thin cartilaginous appendages, asymmetrically overlapping cartilages, and reflexed cartilage margins. Many basal caenogastropod taxa possess previously unidentified cartilaginous support structures below the radula (subradular cartilages), which may be homologous to the dorsal cartilages of other gastropods. As subradular cartilages were absent in carnivorous caenogastropods, adaptation to trophic specialization is likely. However, incongruence with specific feeding strategies or body size suggests that the morphology of odontophoral cartilages is constrained by phylogeny, representing a new source of morphological characters to improve the phylogenetic resolution of this group. J. Morphol. 2009. © 2008 Wiley‐Liss, Inc.     

Morphology of the female reproductive system of three dorippid crabs (Crustacea: Decapoda: Brachyura: Dorippidae) and the role of accessory cuticle structures associated with seminal receptacles

The reproductive systems of crabs reveal characters of considerable importance for the understanding of brachyuran phylogeny and evolution. The Dorippoidea show several plesiomorphic characters within Eubrachyura and similarities to podotreme crabs. Hence, they are often considered as an early diverging lineage, sometimes even as the sister group to all remaining eubrachyurans. Due to their role as prime candidates for putative plesiomorphic characters of the reproductive system of the Eubrachyura, we compared the morphology of the vaginae, seminal receptacles, and ovaries of three dorippid species using histological methods, micro‐computed tomography, and 3‐D reconstructions. Despite the putative phylogenetic position of dorippids, the female reproductive system shows features that are regarded as derived characters in eubrachyurans, including a concave vagina and a ventral‐type seminal receptacle. In contrast to other eubrachyurans, the oviduct does not enter the seminal receptacle directly but through specific cuticular valves. The female reproductive systems of Dorippe sinica and Dorippe quadridens are remarkable in further aspects. The seminal receptacles of both species are completely cuticle‐lined and have accessory sperm storage structures, the bursae. Our findings on the morphology of the female reproductive system of dorippids with its unique combination of basal, derived, and new characters challenges the prevailing hypothesis on the evolution of sperm storage organs in Eubrachyura.     

Isolation and characterization of microsatellites in Pachygrapsus marmoratus (Grapsidae; Decapoda; Brachyura)

Seven polymorphic microsatellite loci were isolated and characterized from the grapsid crab Pachygrapsus marmoratus using FIASCO (fast isolation by AFLP of sequences containing repeats) protocol. Twenty‐seven primer pairs were designed, 14 of which worked well in polymerase chain reaction (PCR), amplifying a fragment of the expected size. Variability was tested in 18 specimens collected along the coast of Tuscany (Italy). Five loci were discarded due to stutter products during their amplification, and two resulted to be monomorphic. The remaining seven loci, showed a number of alleles ranging from two to 14 and an observed heterozygosity ranging from 0.12 to 0.67.