Five hundred million years to mobility: directed locomotion and its ecological function in a turtle barnacle.

Movement is a fundamental characteristic of life, yet some invertebrate taxa, such as barnacles, permanently affix to a substratum as adults. Adult barnacles became 'sessile' over 500 Ma; however, we confirm that the epizoic sea turtle barnacle, Chelonibia testudinaria, has evolved the capacity for self-directed locomotion as adults. We also assess how these movements are affected by water currents and the distance between conspecifics. Finally, we microscopically examine the barnacle cement. Chelonibia testudinaria moved distances up to 78.6 mm yr-1 on loggerhead and green sea turtle hosts. Movements on live hosts and on acrylic panels occasionally involved abrupt course alterations of up to 90°. Our findings showed that barnacles tended to move directly against water flow and independent of nearby conspecifics. This suggests that these movements are not passively driven by external forces and instead are behaviourally directed. In addition, it indicates that these movements function primarily to facilitate feeding, not reproduction. While the mechanism enabling movement remained elusive, we observed that trails of cement bore signs of multi-layered, episodic secretion. We speculate that proximal causes of movement involve one or a combination of rapid shell growth, cement secretion coordinated with basal membrane lifting, and directed contraction of basal perimeter muscles.

Population dynamics and development of the rhizocephalan Sacculina carcini, parasitic on the shore crab Carcinus maenas.

The ecologically important shore crab Carcinus maenas is commonly infected in its native range by the rhizocephalan Sacculina carcini. However, several aspects of this host-parasite interaction are poorly understood. Here, we analyse data from approximately 60000 Danish crabs to unravel factors governing infection patterns in time and space, and according to host sex and size. Female crabs were more frequently infected (12.6%) than males (7.9%). Sites with high salinity supported the highest infection prevalence. Infection prevalence peaked in summer (10 to 15%) and winter (20 to 35%) due in part to emergence of virginal externae in summer (main outbreak) and autumn (minor outbreak) preceded by peaks in crabs with lost externa (scars). Younger externae and scars dominated among males, whereas adult externae were most frequent among females. Infection prevalence increased with size in females but decreased in males, and modified (feminized) males showed lower scar frequency than unmodified ones. Modified males occurred frequently among the smaller size classes, whereas unmodified males dominated the larger size classes. Externa size was positively related to host size in both genders (same linear relationship). Molecular analyses suggested that hosts below 16 mm in carapace width do not become infected. Dissections of infected hosts revealed marked reduction of ovaries, whereas testes were unaffected by sacculinization. Our study demonstrates great spatio-temporal variation in infection prevalence mainly related to the parasite's life history. S. carcini appears capable of infecting all host sizes except the smallest. Owing to incomplete feminization of males, infections are rapidly lost from the larger and highly profitable male hosts.

The origins and evolution of dwarf males and habitat use in thoracican barnacles.

Barnacles are exceptional in having various sexual systems (androdioecy, hermaphroditism, dioecy) and with a high morphological diversity of males, though these are always minute (dwarf) compared to their female or hermaphrodite partners. For the first time, we use a multiple DNA marker-based phylogeny to elucidate the ancestral states and evolution of (1) dwarf males, (2) their morphology when present, (3) their attachment site on the partner, and (4) habitat use in thoracican barnacles. Our taxon sampling was especially rich in rare deep-sea Scalpelliformes and comprised species with diverse sexual systems and dwarf male morphologies. Within the thoracican barnacles dwarf male evolution is subject to extensive convergence, but always correlated to similar ecological conditions. Males evolved convergently at least four times from purely hermaphroditic ancestors, in each case correlated with the invasion into habitats with low mating group sizes. The independent evolution of dwarf males in these lineages dovetails with the males having different morphologies and occurring in several different locations on their sexual partner.

Molecular phylogeny, systematics and morphological evolution of the acorn barnacles (Thoracica: Sessilia: Balanomorpha).

The Balanomorpha are the largest group of barnacles and rank among the most diverse, commonly encountered and ecologically important marine crustaceans in the world. Paradoxically, despite their relevance and extensive study for over 150years, their evolutionary relationships are still unresolved. Classical morphological systematics was often based on non-cladistic approaches, while modern phylogenetic studies suffer from severe undersampling of taxa and characters (both molecular and morphological). Here we present a phylogenetic analysis of the familial relationships within the Balanomorpha. We estimate divergence times and examine morphological diversity based on five genes, 156 specimens, 10 fossil calibrations, and six key morphological characters. Two balanomorphan superfamilies, eight families and twelve genera were identified as polyphyletic. Chthamaloids, chionelasmatoid and pachylasmatoids split first from the pedunculated ancestors followed by a clade of tetraclitoids and coronuloids, and most of the balanoids. The Balanomorpha split from the Verrucidae (outgroup) in the Lower Cretaceous (139.6 Mya) with all the main lineages, except Pachylasmatoidea, having emerged by the Paleocene (60.9 Mya). Various degrees of convergence were observed in all the assessed morphological characters except the maxillipeds, which suggests that classical interpretations of balanomorphan morphological evolution need to be revised and reinterpreted.

Evolution of sex determination and sexually dimorphic larval sizes in parasitic barnacles.

The parasitic (rhizocephalan) barnacles include species of which larval sex is determined by the mother (genetic sex determination, GSD), male larvae are larger than female larvae, and a female accepts only two dwarf males who sire all the eggs laid by her. In contrast, other species of parasitic barnacles exhibit monomorphic larvae that choose to become male or female depending on the condition of the host they settle (environmental sex determination, or ESD), and a female accepts numerous dwarf males. Here, we ask why these set of traits are observed together, by examining the evolution of sex determination and the larval size. ESD has an advantage over GSD because each larva has a higher chance of encountering a suitable host. On the other hand, GSD has two advantages over ESD: the larval size can be chosen differently between sexes, and their larvae can avoid spending time for sex determination on the host. We conclude that, in species whose female accepts only two males, the male larvae engage in intense contest competition for reproductive opportunities, and male's success-size relation is very different from female's. Then, larvae with predetermined sex (GSD) with sexually dimorphic larvae is more advantageous than ESD. In contrast, in species whose females accept many dwarf males, the competition among males is less intense, and producing larvae with undetermined sex should evolve. We also discuss the condition for females to evolve receptacles to limit the number of males she accepts.

Morphometric and molecular identification of individual barnacle cyprids from wild plankton: an approach to detecting fouling and invasive barnacle species.

The present study used DNA barcodes to identify individual cyprids to species. This enables accurate quantification of larvae of potential fouling species in the plankton. In addition, it explains the settlement patterns of barnacles and serves as an early warning system of unwanted immigrant species. Sequences from a total of 540 individual cypris larvae from Taiwanese waters formed 36 monophyletic clades (species) in a phylogenetic tree. Of these clades, 26 were identified to species, but 10 unknown monophyletic clades represented non-native species. Cyprids of the invasive barnacle, Megabalanus cocopoma, were identified. Multivariate analysis of antennular morphometric characters revealed three significant clusters in a nMDS plot, viz. a bell-shaped attachment organ (most species), a shoe-shaped attachment organ (some species), and a spear-shaped attachment organ (coral barnacles only). These differences in attachment organ structure indicate that antennular structures interact directly with the diverse substrata involved in cirripede settlement.

Past, Present and Future of the Aral Sea -A Review of its Fauna and Flora before and during the Regression Crisis.

We review the past, present and possible future of the Aral Sea system in context of the human caused regression crisis that resulted in the drying out of the larger part of this original brackish water sea. The results are put into the context of other threatened saline lakes and the general water crisis in the world due to overexploitation of water resources and climate change. We cover the geographic history and hydrology from the origin of the sea 17,000 years ago to the present. The original biota including animals, higher plants and algae are covered in full detail, and tracked through the regression crisis. We put special emphasis on fish and fisheries because of their economic importance for the surrounding populations. We also review the side effects of the regression in terms of human health and changes to the terrestrial environment and local climate. We explain the dramatic improvements to the fauna in the northern Small Aral Sea following the construction of dams to retain its waters and discuss future options to further improve this restored water basin. We contrast this with the progressing hypersalinization of the remnants of the southern Large Aral Sea, which faces conditions that will eventually render a "Dead Sea" condition hostile to all metazoan life. We end by highlighting the partial restoration of the Small Aral Sea as an example of how much restoration can be achieved for relatively little financial expense and in a short period, when good ideas, kind hearts and hard work operate together for the benefit of the environment and our human society.

Adaptive evolution of sexual systems in pedunculate barnacles.

How and why diverse sexual systems evolve are fascinating evolutionary questions, but few empirical studies have dealt with these questions in animals. Pedunculate (gooseneck) barnacles show such diversity, including simultaneous hermaphroditism, coexistence of dwarf males and hermaphrodites (androdioecy), and coexistence of dwarf males and females (dioecy). Here, we report the first phylogenetically controlled test of the hypothesis that the ultimate cause of the diverse sexual systems and presence of dwarf males in this group is limited mating opportunities for non-dwarf individuals, owing to mating in small groups. Within the pedunculate barnacle phylogeny, dwarf males and females have evolved repeatedly. Females are more likely to evolve in androdioecious than hermaphroditic populations, suggesting that evolution of dwarf males has preceded that of females in pedunculates. Both dwarf males and females are associated with a higher proportion of solitary individuals in the population, corroborating the hypothesis that limited mating opportunities have favoured evolution of these diverse sexual systems, which have puzzled biologists since Darwin.

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.

Living on fire: Deactivating fire coral polyps for larval settlement and symbiosis in the fire coral-associated barnacle Wanella milleporae (Thoracicalcarea: Wanellinae).

Symbiosis is increasingly recognized as being an important component in marine systems, and many such relationships are initiated when free-swimming larvae of one partner settle and become sedentary on a host partner. Therefore, several crucial questions emerge such as the larva's mechanism of locating a host, selection of substratum and finally settlement on the surface of its future partner. Here, we investigated these mechanisms by studying how larvae of the fire coral-associated barnacle Wanella milleporae move, settle and establish symbiosis with their host, Millepora tenera. Cyprids of W. milleporae possess a pair of specialized antennules with bell-shaped attachment discs that enable them to explore and settle superficially on the hostile surface of the fire coral. Intriguingly, the stinging polyps of the fire coral remain in their respective pores when the cyprids explore the fire coral surface. Even when cyprids come into contact with the nematocysts on the extended stinging polyps during the exploratory phase, no immobilization effects against the cyprids were observed. The exploratory phase of Wanella cyprids can be divided into a sequence of wide searching (large step length and high walking speed), close searching (small step length and low speed) and inspection behavior, eventually resulting in permanent settlement and metamorphosis. After settlement, xenogeneic interactions occur between the fire coral and the newly metamorphosed juvenile barnacle. This involved tissue necrosis and regeneration in the fire coral host, leading to a callus ring structure around the juvenile barnacle, enhancing survival rate after settlement. The complex exploratory and settlement patterns and interactions documented here represent a breakthrough in coral reef symbiosis studies to show how invertebrates start symbiosis with fire corals.

A synthesis of the external morphology of cypridiform larvae of Facetotecta (crustacea: Thecostraca) and the limits of the genus Hansenocaris.

Although the naupliar and cypridiform stages of the enigmatic y-larvae of Facetotecta have been found in the marine plankton worldwide, they still represent the last significant group of crustaceans for which the adult forms are still unknown. From a number of y-cyprids representing different taxa from different locations, we employ scanning electron microscopy to describe fine morphological details of all external structures of this unique larval form. We document different segmentation patterns of the abdomen and presence/absence of the labrum and structural differences in the antennules, labrum, paraocular process, thoracopods, and telson lend support for the erection of several new genera as opposed to the single Hansenocaris. The data presented here emphasize the morphological limits of the genus Hansenocaris and the "bauplan" of cyprydiform larvae of Facetotecta. Although the optimum pathway is a joint analysis of both molecular and morphological characters, we use the morphological characters of y-cyprids to align them cladistically and determine the limits of the genus Hansenocaris s.s. and describe common characters for all y-cyprids including six pairs of the lattice organs instead five pairs considered as a ground pattern for all Thecostraca. We also determine plesiomorphic and apomorphic characters of all known y-cyprids and separate them from other thecostracan cypridiform larvae.

Independent and adaptive evolution of phenotypic novelties driven by coral symbiosis in barnacle larvae.

The invasion of novel habitats is recognized as a major promotor of adaptive trait evolution in animals. We tested whether similar ecological niches entail independent and adaptive evolution of key phenotypic structures related to larval host invasion in distantly related taxa. We use disparately related clades of coral barnacles as our model system (Acrothoracica: Berndtia and Thoracicalcarea: Pyrgomatidae). We analyze the larval antennular phenotypes and functional morphologies facilitating host invasion. Extensive video recordings show that coral host invasion is carried out exclusively by cypris larvae with spear-shaped antennules. These first exercise a series of complex probing behaviors followed by repeated antennular penetration of the soft host tissues, which subsequently facilitates permanent invasion. Phylogenetic mapping of larval form and function related to niche invasion in 99 species of barnacles (Thecostraca) compellingly shows that the spear phenotype is uniquely associated with corals and penetrative behaviors. These features evolved independently in the two coral barnacle clades and from ancestors with fundamentally different antennular phenotypes. The larval host invasion system in coral barnacles likely evolved adaptively across millions of years for overcoming challenges associated with invading and entering demanding coral hosts.

Video observation of surface exploration in cyprids of Balanus amphitrite: the movements of antennular sensory setae.

Video microscopy of cyprids of Balanus amphitrite was used to monitor the action of antennular setae during the exploratory behaviour prior to attachment. In addition, SEM was used to provide a revised description of all antennular setae for that species. The videos describe if a particular seta touches the substratum and the area it can cover during surface exploration. On the fourth segment, the plumose terminal setae A and B are never in contact with the substratum, lack a terminal pore and it is argued that they sense hydrodynamic forces. The aesthetasc-like terminal seta D is likewise held free in the water at all times and it is speculated that it senses dissolved substances, but, since it contains a scolopale rod, it must also have a mechano-receptive function. All remaining antennular setae on the second, third and fourth segments have a terminal pore and it is argued that these are bimodal receptors with both chemo- and mechano-receptive modalities. These setae are also at one time or another in contact with the substratum, except perhaps for the small preaxial seta 2 and terminal seta C. The first seta to contact the surface during a tentative step is radial seta 5, which is longer than all other radial setae. All other setae on the second and third segment are only in contact after a step is completed. When the attachment disc touches the surface (=a step completed) the long and curved postaxial seta 2 (on the second segment) and postaxial seta 3 on the third segment are both flexed to either side of the antennule. This lateral displacement ensures that these two setae can touch large surface areas to either side of the appendage. The four subterminal setae on the fourth segment contact the surface both immediately before and after a step has been completed, and the constant flicking of the segment significantly increases the surface area tested by both these chemoreceptors and by terminal seta E, which can sweep up to 60 µm laterally from the attachment disc. The flicking of the fourth segment may also serve to dilute the boundary layer of chemoreceptors on the fourth segment such as the aesthetasc-like terminal seta D and thus facilitate the detection of new stimuli.

Crustacean Fauna of the Aral Sea and its Relation to Ichthyofauna During the Modern Regression Crisis and Efforts at Restoration.

The regression and salinization of the Aral Sea, largely caused by water diversion for irrigation, is among the most severe ecological disasters of the 20th century, and has had severe health and economic consequences for the local population. Introductions of alien species to enhance commercial fisheries before the regression had already impacted the ecology of this system. Crustaceans made up about one-quarter of the original metazoan species and constituted the principal food for native and introduced fish. From 1960 on, crustaceans were recorded at numerous fixed sampling stations, including thanatocoenoses (dead animals from sediment cores). We use this previously unpublished information to document changes in species abundance and discuss their causes in the context of species interactions and changes to physical and chemical parameters. Competition from alien crustaceans led to declines in or even extinction of some native species, but eventually severe salinization became the main detriment, and resulted in the complete collapse of commercial fisheries. This seriously hurt a critical trade, which provided the principal protein source for the local population. We document how comparatively modest conservation efforts enabled the northern Small Aral Sea to partially recover and commercial fishing to resume.

'Flying barnacles': implications for the spread of non-indigenous species.

The presence of adult barnacles of Fistulobalanus pallidus (Darwin) and Fistulobalanus albicostatus (Pilsbry) attached to field-readable plastic leg rings on the Lesser Black-backed Gull Larus fuscus in Northern Europe is reported. L. fuscus is a long-distance palaearctic migrant, breeding in temperate areas spreading widely over inland and marine habitats outside the breeding season. The species is known to perform long-distance migration to Africa and the Middle East. Combining present knowledge on the birds' migratory pattern and the home range of the barnacle species, it is concluded that the cypris larvae of F. pallidus must have settled in African waters, whereas the area where F. albicostatus settled on the bird leg rings is less certain. The barnacles were of adult size and must thus have been attached for a period of no less than 2 months. More than 30 individual barnacles could occur together on a single field-readable plastic leg ring. The barnacles could therefore, if ported alive to a new area, reproduce successfully and thus either introduce the species or genetically affect other native populations. This may pose a new and wholly unexpected transportation pathway for barnacles as invasive species.

The monophyletic origin of a remarkable sexual system in akentrogonid rhizocephalan parasites: a molecular and larval structural study.

We use sequences from the nuclear ribosomal genes, 18S and 28S to analyze the phylogeny of the Rhizocephala Akentrogonida including two species, Clistosaccus paguri and Chthamalophilus delagei, that are critical for understanding rhizocephalan evolution but have not previously been part of a molecularly based study. In addition we use light and scanning electron microscopy to compare the cypris larvae of C. paguri, Sylon hippolytes and two species of the family Thompsoniidae, since this larval stage offers a suite of characters for analyzing the evolution of these otherwise highly reduced parasites. The Rhizocephala Akentrogonida form a monophyletic group nested within a paraphyletic "Kentrogonida". C. paguri and S. hippolytes are sistergroups confirming the monophyly of the Clistosaccidae that was originally based on similarities in the cypris larvae. We find numerous LM and SEM level similarities between the two species, many of which appear to be correlated with their specialized sexual system, where male cyprids use an antennule to implant cells into the virgin female parasite. Some of these traits are also found in cyprids of the thompsoniid species. We conclude that the special cypris morphology and the implantation of males by antennular penetration was present in the stem species to the Thompsoniidae and the Clistosaccidae and emphasize the power of larval characters in rhizocephalan systematics. C. delagei is a sister group to Boschmaella balani and the two are nested deep within the Akentrogonida. This confirms the monophyly of the Chthamalophilidae and falsifies the theory that C. delagei should represent the most primitive extant rhizocephalan. Instead, chthamalophilid rhizocephalans represent some of the most highly advanced members of the parasitic barnacles.

Remarkable convergent evolution in specialized parasitic Thecostraca (Crustacea).

BACKGROUND: The Thecostraca are arguably the most morphologically and biologically variable group within the Crustacea, including both suspension feeders (Cirripedia: Thoracica and Acrothoracica) and parasitic forms (Cirripedia: Rhizocephala, Ascothoracida and Facetotecta). Similarities between the metamorphosis found in the Facetotecta and Rhizocephala suggests a common evolutionary origin, but until now no comprehensive study has looked at the basic evolution of these thecostracan groups. RESULTS: To this end, we collected DNA sequences from three nuclear genes [18S rRNA (2,305), 28S rRNA (2,402), Histone H3 (328)] and 41 larval characters in seven facetotectans, five ascothoracidans, three acrothoracicans, 25 rhizocephalans and 39 thoracicans (ingroup) and 12 Malacostraca and 10 Copepoda (outgroup). Maximum parsimony, maximum likelihood and Bayesian analyses showed the Facetotecta, Ascothoracida and Cirripedia each as monophyletic. The better resolved and highly supported DNA maximum likelihood and morphological-DNA Bayesian analysis trees depicted the main phylogenetic relationships within the Thecostraca as (Facetotecta, (Ascothoracida, (Acrothoracica, (Rhizocephala, Thoracica)))). CONCLUSION: Our analyses indicate a convergent evolution of the very similar and highly reduced slug-shaped stages found during metamorphosis of both the Rhizocephala and the Facetotecta. This provides a remarkable case of convergent evolution and implies that the advanced endoparasitic mode of life known from the Rhizocephala and strongly indicated for the Facetotecta had no common origin. Future analyses are needed to determine whether the most recent common ancestor of the Thecostraca was free-living or some primitive form of ectoparasite.

Antennular sensory organs in cyprids of balanomorphan cirripedes: standardizing terminology using Megabalanus rosa.

Cirripedes are one of the major groups of fouling organism in the marine environment. The cyprid can, before a permanent attachment, actively explore and walk on the substratum using its antennules in a bipedal fashion without leaving the surface. Studying the structure of the cyprid antennule is therefore important for understanding the events that culminate in biofouling by barnacles. There are at present no complete, standardised accounts of the structure of the cyprid antennules in thoracican barnacles, and moreover, the existing accounts vary in their use of terminology. This article describes the cyprid antennule of the barnacle Megabalanus rosa. This barnacle species is common in E Asia, and the cyprids have previously been used in several biofouling studies. All externally visible setae on the antennules have been mapped; these comprise both chemosensors with a terminal pore, a putative aesthetasc-like seta and mechano-sensory setae. More setae were found on the attachment disc than in previous scanning electron microscope-based studies, but not all structures that can be seen with transmission electron microscopy were visible. The disc itself seems to have a variable surface area, which could assist in exploring rough surfaces. The various lengths of the antennular setae, coupled with the disposition of the segments, enable the cyprid to cover a wide swath of substratum during exploratory walking. A new terminology is proposed for cyprid antennular setae, which will form a basis for future comparative and functional studies of cirripede settlement.

Induced metamorphosis in crustacean y-larvae: towards a solution to a 100-year-old riddle.

BACKGROUND: The y-larva, a crustacean larval type first identified more than 100 years ago, has been found in marine plankton samples collected in the arctic, temperate and tropical regions of all oceans. The great species diversity found among y-larvae (we have identified more than 40 species at our study site alone) indicates that the adult organism may play a significant ecological role. However, despite intense efforts, the adult y-organism has never been identified, and nothing is therefore known about its biology. RESULTS: We have successfully and repeatedly induced metamorphosis of y-larvae into a novel, highly reduced juvenile stage by applying the crustacean molting hormone 20-HE. The new stage is slug-like, unsegmented and lacks both limbs and almost all other traits normally characterizing arthropods, but it is capable of vigorous peristaltic motions. CONCLUSION: From our observations on live and preserved material we conclude that adult Facetotecta are endoparasitic in still to be identified marine hosts and with a juvenile stage that represents a remarkable convergence to that seen in parasitic barnacles (Crustacea Cirripedia Rhizocephala). From the distribution and abundance of facetotectan y-larvae in the world's oceans we furthermore suggest that these parasites are widespread and could play an important role in the marine environment.

A new internal structure of nauplius larvae: A "ghostly" support sling for cypris y left within the exuviae of nauplius y after metamorphosis (Crustacea: Thecostraca: Facetotecta).

Facetotecta, or crustacean "y-larvae," occur in all the world's oceans although the adult forms remain completely unknown. At the metamorphic molt from the last naupliar instar to the terminal cypris larval stage a free carapace, six pairs of natatory thoracopods, and a segmented thorax and abdomen all develop anew. Unlike in earlier molts, the cephalic shield and the so-called "faciotruncal integument" usually remain together at this last naupliar molt, and the posterior "trunk" portion of the exuviae, while hollow, is not empty. In mounted preparations examined by phase contrast or differential interference contrast (DIC) microscopy, a ghost-like image of part of the cypris thorax, particularly the thoracopods and even their setae, is commonly visible inside the naupliar exuviae, and may be universally present in the Facetotecta. To investigate this "ghost," we used DIC and digital photographic stacking, and also scanning electron microscopy, on slide or stub-mounted final naupliar exuviae of an assortment of undescribed species of Facetotecta that had been reared from planktonic lecithotropic nauplii to the cypris stage at Sesoko Island, Okinawa, Japan, and at Keelung and Green Island, Taiwan. These techniques showed that the "ghost" is a delicate, three-dimensional, fibrous structure, essentially a sling-like mold or matrix with struts attached to the outer cuticle and pairs of deep pockets that previously held the thoracopods of the developing cypris y. Whether it is endoskeletal in nature, the (partial) exuvia of an additional instar, remnants of apoptosis, or something else is currently unknown. Nothing similar has been reported in other thecostracans, or in other crustaceans that undergo a similarly abrupt metamorphosis at the last naupliar molt.