On the morphology of antennular sensory and attachment organs in cypris larvae of the deep‐sea vent/seep barnacles,Ashinkailepas and Neoverruca

Barnacle cypris larvae show high morphological variation in the organs used in search of and attaching to a substratum. This variation may represent adaptation to the habitat of the species. Here, we studied SEM level morphologies of cypris antennular sensory and attachment organs in a deep‐sea vent endemic species (Neoverruca sp.) and a vent/seep inhabiting species (Ashinkailepas seepiophila). We compare them with three species from other environments. The antennular morphologies of Neoverruca sp. and A. seepiophila were similar, which is consistent with recent molecular studies showing a close relationship of the two species. The setation pattern of the antennules was very conservative among species from various environments. In contrast, striking differences were observed in the structure of the attachment organ (the third antennular segment). Neoverruca sp. and A. seepiophila had no velum or a skirt surrounding the attachment disc on the third segment, while other cirripede cyprids almost always have either of these structures. In addition, both cyprids of A. seepiophila and Neoverruca sp. had the attachment disc angled toward the substratum, whereas it faces distally in cyprids from hard bottom inhabiting barnacles. We suggest that both velum/skirt and the angle of the attachment disc play an important role, when the antennules are contacting the substratum during surface exploration. Differences in attachment organ structures may be highly adaptive, enabling cirripede species to enter new habitats during evolution. J. Morphol. 277:594–602, 2016. © 2016 Wiley Periodicals, Inc.     

How whale and dolphin barnacles attach to their hosts and the paradox of remarkably versatile attachment structures in cypris larvae

How larvae of whale and dolphin epibionts settle on their fast-swimming and migrating hosts is a puzzling question in zoology. We successfully reared the larvae of the whale and dolphin barnacle Xenobalanus globicipitis to the cyprid stage. We studied the larval developmental ecology and antennular morphology in an attempt to assess whether an epibiotic lifestyle on this extreme substratum entails any unique larval specializations. Morphological parameters were compared with five other barnacle species that also inhabit extreme substrata. We found no larval specializations to a lifestyle associated with marine mammals. The external morphology of the antennules in Xenobalanus cyprids is morphologically similar to species from strikingly different substrata. We found variation only in the structures that are in physical contact with the substratum, i.e., the third segments carrying the villi-covered attachment disc. The third segments of the Xenobalanus cyprid antennules are not spear-shaped as in the stony coral barnacles, which are here used to penetrate the live tissue of their hosts. The presence of a cyprid cement gland implies that Xenobalanus uses cement protein when attaching to its cetacean host. Naupliar instars developed outside of the mantle cavity, indicating dispersal is planktonic. Our results militate against the idea that the cyprids settle during ocean migrations of their hosts. We suggest cyprids settle during coastal aggregations of the cetacean hosts. We conclude that the ecological success of barnacles has ultimately depended on a larva that with little structural alteration possesses the ability to settle on an amazingly wide array of substrata, including cetaceans.

     

Cypris morphology in the barnacles Ibla and Paralepas (Crustacea: Cirripedia Thoracica) implications for cirripede evolution

We used scanning electron microscopy (SEM) to describe cypris morphology in species of the barnacles Ibla and Paralepas, both of which are pivotal in understanding cirripede evolution. In Ibla, we also studied late naupliar stages with video and SEM. Special emphasis was put on the lattice organs, the antennules and the thorax and telson. In Paralepas we had settled specimens only and could therefore only investigate the carapace with the lattice organs. Cyprids of Ibla quadrivalvis and Paralepas dannevigi have five sets of lattice organs, grouped as two anterior and three posterior pairs. The organs are of the pore‐field type and the terminal pore is situated anteriorly in the first pair, just as in the Rhizocephala and the Thoracica. In Ibla the armament of antennular sensilla resembles that found in the Thoracica but differs from the Rhizocephala. The absence of setules on the A and B setae sited terminally on the fourth antennular segment is a similarity with the Acrothoracica. The attachment disc is angled rather than facing distally and is encircled by a low cuticular velum. The thoracopods have two‐segmented endopods and exopods as in the Thoracica, but the number, shape, and position of thoracopodal setae differ somewhat from other species of that superorder. Both Ibla and Paralepas cyprids have a deeply cleaved telson, but no independent abdominal part. In cypris morphology, Ibla and Paralepas show several synapomorphies with the clade comprising Rhizocephala and Thoracica and there are no specific apomorphies with either the Acrothoracica, the Rhizocephala or any particular subgroup within the Thoracica. This is in agreement with recent molecular evidence that Ibla (Ibliformes) is the sister taxon to all other Thoracica and the ibliforms therefore become the outgroup of choice for studying character evolution within the superorder. Paralepas, and other pedunculated barnacles without shell plates, are apparently not primitive but are secondarily evolved and nested within the Thoracica. J. Morphol., 2009. © 2008 Wiley‐Liss, Inc.     

The cypris larvae of the rhizocephalan barnacle Heterosaccus lunatus with particular reference to antennular morphology

SEM investigation of laboratory-reared cyprids of the parasitic barnacle Heterosaccus lunatus has revealed morphological differences between the male and female larvae, particularly the attachment organ of their antennules. The detailed features of the attachment organ (segment III) and segment IV for each sex are described and compared with those of other rhizocephalan cyprids, particularly Sacculina carcini , a reportedly closely related species. The absence of the large posterior sac from segment III in the H. lunatus male highlights the need for careful larval investigations on other sacculinid species. Settlement of female H. lunatus cyprids occurred on the gill lamellae of the host crab with the subsequent kentrogon stage taking the sacculinid form. The inclusion of H. lunatus in the Sacculinidae was therefore confirmed, based on the morphology of the cypris antennules and kentrogon.     

Cypris Ultrastructure,Metamorphosis and Sex in Seven Families of Parasitic Barnacles (Crustacea: Cirripedia: Rhizocephala)

Cypris morphology in the Rhizocephala differs both between the sexes of individual species and between the species. The large aesthetasc on the third antennular segment is unique to male cyprids of the Lernaeodiscidae, Peltogasteridae and Sacculinidae (suborder Kentrogonida) and Mycetomorpha vancouverensis (suborder Akentrogonida). A fourth segmental aesthetasc, common to both sexes, is much longer in males than in females. A spinous process on the attachment disc is another male-specific character, but is absent altogether in the Lernaeodiscidae. It is argued that these aesthetascs help locate the settlement targets. Except in Mycetomorpha vancourverensis, aesthetascs have been secondarily lost in cyprids of the suborder Akentrogonida, and there are no other morphological means for separating the male and female settling stage. In cyprids of Sylon hippolytes (Sylonidae) and of Mycetomorpha vancouverensis and Thompsonia sp. (Akentrogonidae) the long and slender antennules are probably used in penetration without formation of a kentrogon, as already reported for Clistosaccus paguri (Clistosaccidae). Cyprids of Sylon hippolytes and Clistosaccus paguri are almost morphologically identical.     

Cypris larvae of acrothoracican barnacles (Thecostraca: Cirripedia: Acrothoracica)

We used SEM to investigate the morphology of the cypris larvae from a range of species of the Cirripedia Acrothoracica, representing all three families and including the first detailed account of cyprids in the highly specialized Cryptophialidae. Special attention was given to the head shield (carapace), the lattice organs, the antennules, the thoracopods, the telson and the furcal rami. The cypris larvae of the Acrothoracica fall into two morphological groups; those of the Trypetesidae and Lithoglyptidae have a well-developed carapace (head shield) that can completely enclose the body and sports fronto-lateral pores, numerous short setae and lattice organs perforated by numerous small, rounded pores and a single, conspicuous terminal pore. The fourth antennular segment has the setae arranged in subterminal and terminal groups. There is a developed thorax with natatory thoracopods and a distinct abdomen and telson. In comparison, the cyprids of the Cryptophialidae exhibit apomorphies in the morphology of the carapace, the antennules and the thorax, mostly in the form of simplifications and reductions. They have a much smaller head shield, leaving parts of the body directly exposed. The shield is conspicuously ornamented by deep pits and hexagonally arranged ridges and bears a few, very long setae but lacks fronto-lateral pores. The lattice organs have numerous elongated pores, but no large, terminal pore. The fourth antennular segment has all the setae clustered in one terminal group. The thorax and thoracopods are rudimentary and not suitable for swimming. These reductions and simplifications in morphology correlate with cryptophialid cyprids being unable to swim. They can only disperse by antennular walking resulting in small, but highly gregarious populations of adults. The variations in antennular morphology and telson structure were traced for the genera of the families Lithoglyptidae and Trypetesidae. The traditional non-cladistic taxonomy in the suborders Pygophora (Cryptophialidae+Lithoglyptidae) and Apygophora (Trypetesidae) was based largely on symplesiomorphies in adult morphology and cannot be upheld. The Lithoglyptidae and Trypetesidae may form a monophylum, but evidence remains scarce. We expect that the use of larval (cyprid) characters will in the future play an important part in more detailed phylogenetic analyses of the Acrothoracica and also shed new light on their reproductive ecology.     

Postembryonic development in Diaphanosoma brachyurum (Lievin, 1848) (Crustacea: Ctenopoda: Sididae)

Postembryonic females and males Diaphanosoma brachyurum from Lake Glubokoe (Moscow) have 3–4 and 3 juvenile instars, respectively. Females and males of the first three postembryonic instars can be identified by the different number of setae and setal rudiments on the proximal and distal segments of the exopodite of the swimming antennae: 3 + 7; (i + 3) + 7; 4 + (i + 7), respectively (i = rudiment of seta). The subsequent instars have 4 + 8 long plumose setae on these segments, but the fourth instar has the proximal lateral seta of the distal exopod segment slightly shorter and thinner than the others. The antennules and copulatory appendages of males are instar-specific. Diaphanosomas show small increments in body length during the postembryonic molts. The largest increments (about 115 m) occur during the first or second molts. The allometric equation of Huxley (1924) was used for a comparison of the relative growth rate of different body parts. In the middle of summer, the head and swimming antennae with the body and the antennal exopodite with the antennal basipodite grow in isometry. At the same time, the branches of the swimming antennae and their setae show allometric growth: the exopodite and distal setae grow faster than the endopodite and the lateral setae, respectively.     

Setal morphology and cirral setation of thoracican barnacle cirri: adaptations and implications for thoracican evolution

Thoracic cirripedes are sessile crustaceans that use six pairs of thoracic appendages (the cirri) to catch and handle food. We used scanning electron microscopy to examine the cirri, which include one to three pairs of maxillipeds in six species of thoracican barnacles, in search of correlations between cirral setation and feeding mode. The species studied comprise both pedunculate and sessile forms and represent a wide range of marine habitats as well as morphologies, viz., Ibla cumingi , Octolasmis warwickii , Capitulum mitella , Pollicipes polymerus , Tetraclita japonica japonica and Megabalanus volcano . Of the pedunculates, I. cumingi has the least complex setation pattern consisting of only serrulate types. This is consistent with its very simplified feeding mode and an apparent inability to discriminate between food items. Octolasmis warwickii is slightly more modified, while both P. polymerus and C. mitella have a more diversified setation. The balanomorphan species exhibit by far the most complex cirral setation. This is consistent with the several types of suspension feeding seen in these species, their ability to identify and sort captured food items and even to perform microfiltration in the mantle cavity using the setae on their three pairs of maxillipeds. Our results indicate that in thoracican barnacles, adaptations in feeding behaviour are associated with changes in the setation pattern of the cirri. In addition, the setal types and their distribution on the cirri are potential new characters in future morphology-based analyses of the phylogeny of the Cirripedia Thoracica.     

Probing the settlement signals of Amphibalanus amphitrite

To achieve their reproductive potential, barnacles combine tactile exploration of surface structural properties and integration of cellular signals originating from their antennular sensory setae within a developmentally defined, temporally narrow window of settlement opportunity. Behavioural assays with cyprids coupled with biometric analysis of scanning electron microscopy-acquired images in the presence of specific chemical compounds were used to investigate how settlement on a substratum is altered in response to the presence of these compounds. Impeding tactile exploration was shown which altered cellular signalling and/or induced malformation of anatomical features of the antennular sensory setae, which disrupted the settlement behaviour of the model barnacle species Amphibalanus amphitrite. It is concluded that surface exploration by the cyprids relies on mechanical and nociception-related and calcium-mediated signals while a protein kinase C signalling cascade controls the timely metamorphosis of the cyprids to sessile juveniles.     

Development of functional adaptation to clasping behaviour in harpacticoid copepods (Copepoda,Harpacticoida)

The copepodite antennule development of several harpacticoid families is studied and male antennule development of 5 species is schematically illustrated. The origin of newly formed segments can be determined by seta numbers of segments, relative segment length, furrows indicating previous articulations and by the position of the segment which bears the proximal aesthetasc. At least in some species sexual dimorphism of antennules is present from C II onwards. Females practically reach the adult antennular state at C V whereas males undergo drastic changes from C V to C VI with proximal addition of segments, and often with distal fusions and formation of a unique armature due to functional adaptations in clasping behaviour. No correlation has been found between clasping mode, male antennule type and taxonomic level. In closely related species of the same genus both chirocer and subchirocer antennules are found. The postnaupliar development of the chirocer type can proceed along different ways. Consequently it is suggested that at least chirocer antennules have originated several times independently during harpacticoid evolution.     

Hydaticus tuyuensis Trémouilles (Coleoptera: Dytiscidae): Larval Morphology and Phylogenetic Relationships within Dytiscinae

The three larval instars of Hydaticus (Guignotites) tuyuensis Trémouilles are described and illustrated for the first time, emphasizing the morphometry and chaetotaxy. Second- and third-instar larvae of the known species of the subgenus Guignotites Brinck are characterized by a trilobate median process of prementum. All larval instars of H. tuyuensis, however, have a bilobate process, similar to that present in the known species of the subgenus Hydaticus Leach. This implies that a trilobate process cannot be used as a diagnostic character for Guignotites. Alternatively, Guignotites as presently defined may not represent a natural group. A cladistic analysis of 57 larval characters suggests that the genus Hydaticus shares a common origin with the clade composed of the genera Eretes Laporte and Thermonectus Dejean, based on the following synapomorphies: (i) abdominal segment VII almost completely sclerotized ventrally; the absence of (ii) additional ventroapical pores on the third antennomere, (iii) setae FE4 and FE6, (iv) additional setae on the femur and (v) additional setae on the tibia; and the presence of (vi) setae on the median process of prementum and (vii) spinulae on the second labial palpomere. A bilobate or trilobate median process of the prementum and the submedial insertion of seta AN3 distinguish Hydaticus from the remaining genera of Dytiscinae studied.     

First report of antennular attachment organs in a barnacle nauplius larva

Larvae released from Newmaniverruca albatrossiana were cultured in the laboratory until the cypris stage. The brood size of individuals was low, about 60 larvae per brood. The exact number of instars was not determined. Early instars had the morphology normally seen in lecithotrophic nauplii of thoracican cirripedes. They had uniramous antennules with a few apical setae and biramous antennae and mandibles equipped with natatory setae only. Neither antennae nor mandibles carried any enditic spines or setae and the mouth cone was diminutive. The last nauplius stage obtained in our cultures was typical except in the structure of antennules. The head shield was enlarged but not flexed down, the antennae and mandibles were virtually unchanged from earlier instars, and the ventral thoracic process was well developed but without any external appendages. In contrast, the antennules had the complex shape and segmentation otherwise seen only in cypris larvae, where they are used for bipedal walking on the substratum in search of a settlement site. The similarity included the specialized shape of the first two antennular segments and the specialization of the third as an attachment organ. Nauplii just prior to this last instar had simple, straight antennules but completely lacked setae and instead terminated bluntly in what appears to be an incipient attachment organ. The presence of cypris-like antennules in late nauplii has not previously been recorded in cirripedes. We suggest that this will allow the larvae to attach on the substratum temporarily before they reach the cypris instar and this will increase the chance of settling successfully on their rare substratum (sea urchin spines). The specialization in late N. albatrossiana nauplii will therefore decrease mortality during the larval phase and thus counterbalance the very low breeding potential in this deep-sea species.     

A new species of Tigriopus (Copepoda, Harpacticoida, Harpacticidae) from Thailand with the description of its naupliar development

Both genders of Tigriopus thailandensis sp. nov. are described from a laboratory stock raised from individuals collected from the seaweed Enteromorpha clathrata in Thailand (Bangsaen Beach, Chonburi Province). Tigriopus thailandensis sp. nov. shares with its closest relative T. japonicus Mori, 1932 two setae on the third exopodal segment of leg 4 while other congeners bear 3 inner setae. However, allobasis and exopod of antenna in both genders are much more slender and elongate than in T. japonicus. All six naupliar stages of T. thailandensis are described from the offspring of isolated females. In comparison with nauplii of T. japonicus, T. thailandensis nauplii are characterized by the following: a smaller body size throughout the naupliar phase; first antennular segment without seta, second antennular segment with only one small seta plus two longer setae; third antennular segment with additional spinules from naupliar stage II onwards; antenna bears three small spinules on the terminal exopodal segment; one additional seta on the anterior surface of the antennary basis, tubular endopod of antenna with one tiny seta midlength at naupliar stage III that increases in size; mandibular basis with several spinules on anterior surface; mandibular coxa with one spinulose seta that is smooth in T. japonicus.     

Analysis of the Behaviours Mediating Barnacle Cyprid Reversible Adhesion

When exploring immersed surfaces the cypris larvae of barnacles employ a tenacious and rapidly reversible adhesion mechanism to facilitate their characteristic ‘walking’ behaviour. Although of direct relevance to the fields of marine biofouling and bio-inspired adhesive development, the mechanism of temporary adhesion in cyprids remains poorly understood. Cyprids secrete deposits of a proteinaceous substance during surface attachment and these are often visible as ‘footprints’ on previously explored surfaces. The attachment structures, the antennular discs, of cyprids also present a complex morphology reminiscent of both the hairy appendages used by some terrestrial invertebrates for temporary adhesion and a classic ‘suction cup’. Despite the numerous analytical approaches so-far employed, it has not been possible to resolve conclusively the respective contributions of viscoelastic adhesion via the proteinaceous ‘temporary adhesive’, ‘dry’ adhesion via the cuticular villi present on the disc and the behavioural contribution by the organism. In this study, high-speed photography was used for the first time to capture the behaviour of cyprids at the instant of temporary attachment and detachment. Attachment is facilitated by a constantly sticky disc surface – presumably due to the presence of the proteinaceous temporary adhesive. The tenacity of the resulting bond, however, is mediated behaviourally. For weak attachment the disc is constantly moved on the surface, whereas for a strong attachment the disc is spread out on the surface. Voluntary detachment is by force, requiring twisting or peeling of the bond – seemingly without any more subtle detachment behaviours. Micro-bubbles were observed at the adhesive interface as the cyprid detached, possibly an adaptation for energy dissipation. These observations will allow future work to focus more specifically on the cyprid temporary adhesive proteins, which appear to be fundamental to adhesion, inherently sticky and exquisitely adapted for reversible adhesion underwater.     

THE ACTION OF COPPER IN ANTIFOULING PAINTS

Experiments on the number of barnacle cyprids found on plane surfaces and on the surface of small pits coated with compositions containing different amounts of cuprous oxide show that the presence of copper does not deter the cyprids from attempting to settle. Copper-oxide paints do not inhibit settlement appreciably; their efficiency as antifouling agents is caused by the toxic action of copper on cyprids and young spat after initial attachment.
The cementing of the shell to the substratum in young barnacles which have survived metamorphosis appears to be hindered by the presence of copper paints so that they are easily dislodged.     

Metamorphosis in the Cirripedia Rhizocephala and the homology of the kentrogon and trichogon

The Rhizocephala are considered to be monophyletic due to several synapomorphies in the ontogeny of the cndoparasitic phase. The various types of metamorphosis described in the Rhizocephala are discussed and compared to metamorphosis in the Cirripedia Thoracica and Acrothoracica. In males and females of the suborder Kentrogonida. the cyprid settles and metamorphoses into a new instar, in males the trichogen and in females the infective kentrogon. The kentrogon goes through yet another. incomplete moult associated with the development of the stylet. Within the three kentrogonidan families. the Iernaeodiscid-peltogastrid type of kentrogon differs from the sacculinid type in the mode of attachment to the host. in the complexity of internal anatomy. in the position and penetration of the stylet, and in whether or not the cyprid carapace must be shed prior to penetration of the stylet. In the Akentrogonida metamorphosis never results in a new instar. Where observed (Clistosaccidae and Thompsoniidae). both male and female cyprids settle and penetrate into their substrate (female parasite or new host) with one of the antennules. Using the antennule as a syringe. male cyprids inject spermatogonia while female cyprids injects embryonic cells developing into an endoparasite. By comparison with metamorphosis in the Cirripedia Thoracica and Acrothoracica it is concluded that the presence of a metamorphic moult leading to a post-cyprid instar is plesiomorphic and that the trichogon and kentrogon are homologous with the first metamorphosed juvenile in these outgroups. The abbreviated ontogeny in the Akentrogonida without metamorphic moult and post-cyprid larval instars is considered apomorphic. This contradicts the long-held supposition that the Akentrogonida are the most‘primitive’Rhizocephala and dovetails with new information that this suborder contains many advanced traits. Within the Kentrogonida. the lernacodiseid-peltogastrid type of kentrogon is considered more plesiomorphic than the sacculinid type, which resembles the clistosaccidthompsoniid type in having the antennules involved in the penetration process. The homologization of the kentrogon with a juvenile barnacle indicates that presence of a kentrogon is plesiomorphic within the Rhizocephala and that the Kentrogonida is paraphyletic.     

Growth and molting in epizoic pedunculate barnacles genus Octolasmis (Crustacea: Thecostraca: Cirripedia: Thoracica)

Scanning electron microscopy, light microscopy, and histology were used to study growth in species of the pedunculate barnacle genus Octolasmis (O. angulata, O. cor, O. californiana, O. mülleri). These species are epizoic in the gill chamber of portunid crabs and have highly reduced capitular shell plates, with large areas of general cuticle in between. The external integument grows by means of a system of narrow growth zones, one encircling the peduncle and a Y-shaped system on either side of the capitulum. Growth is by a regular series of molts, but shedding of old cuticle and production of new layers is entirely restricted to the growth zones. Just prior to ecdysis, the new cuticle lies in a highly folded fashion beneath the old cuticle that is about to be shed. At ecdysis, the old cuticle breaks along the margins of the growth zones and the resulting scars remain as a system of "ecdysial lines" along either side of the zone. Once exposed after ecdysis, the new cuticle remains as a part of the permanent external integument. The growth zones divide the externa into five cuticular areas, two on the peduncle and three on the capitulum. The calcareous shell plates (carina, paired scuta, and, when present, paired terga) all lie within the capitular regions and the ecdysial lines pass across, not around, these mineralized areas. The number, relative spacing, and topology of the ecdysial lines form a record of the growth history of the specimen. These and other growth patterns demonstrate that size increase is due to the formation of new cuticle by molting in the growth zones, while expansion of the shell plates by mineralization follows only after production of the new cuticle. Thus, although specialized, growth in Octolasmis still complies with the general crustacean model, complicated only by the mineralization of parts of the capitular cuticle into shell plates. The results are compared with the very scarce information on molting in other barnacles. We argue that at least the circular peduncular growth zone is omnipresent in the Cirripedia Thoracica.