Ultrastructure of the mature spermatozoon of the coconut crabBirgus latro (Coenobitidae: Paguroidea: Decapoda)

The spermatozoon ofBirgus latro (Linnaeus, 1767) is approx 14µm in length. It is composed of a large multi-layered oblong-ovoid acrosome which is capped by a conical operculum and lies anterior to a small ring of cytoplasm and an amorphous nucleus which is drawn out into a series of arms or extensions. Originating from the cytoplasmic area are three further long microtubular arms. The sperm ofB. latro is very similar to the sperm of the only other genus in the Coenobitidae,Coenobita, of whichC. clypeatus is a representative species. They share a suite of ultrastructural characters including: a long, cylindrical, capsule-bound acrosome containing an inner acrosome core, a large acrosome ray zone and a thin outer acrosome zone; an apical operculum anterior to a subopercular zone divided into two areas of differing density; an invaginated perforatorial zone with a bipartite granular matrix; microvillus-like extensions of the inner acrosome core projecting into the perforatorial invagination; a ring of cytoplasm, around the base of the acrosome, containing numerous mitochondria, extensive lamellar systems and the bases of three microtubular arms; granular nuclear material forming irregular arms; and at the posterior portion of the cell membrane a combination of nuclear and plasma membranes. Some ultrastructural characters which separate the two genera are: a domed operculum inC. clypeatus as opposed to a conical one inB. latro; inB. latro there is some residual cytoplasm external to the operculum and centrioles are absent from the mature spermatozoon; inC. clypeatus the inner acrosome core does not appear to invest the perforatorium and a series of dense rods are found subjacent and internal to the operculum. Similarities between the two coenobitids are greater than those shared with the paguridEupagurus bernhardus. In this latter species, the acrosome is more ovoid than cylindrical and the acrosome zones are less conspicuous; the operculum is absent from the mature sperm (although present during spermiohistogenesis); no subopercular zone is present; the perforatorium contains longitudinally arranged microtubules and extends the full length of the acrosome; and the nuclear material does not form separate nuclear arms. Sperm ultrastructure supports monophyly of the Paguroidea, while distinguishing coenobitids from pagurids within this superfamily.     

Spermiogenesis and sperm structure in the shrimp Parapenaeus longirostris (Crustacea: Dendrobranchiata): comparative aspects among decapods

Early spermatids of the dendrobranchiate shrimp Parapenaeus longirostris (Lucas, 1846) have a spherical nucleus with large patches of heterochromatin, surrounded by a cytoplasmic mass that contains the conspicuous proacrosomal vesicle. The highly polarized mid spermatid mainly consists of the nuclear region, displaying a discontinuous nuclear envelope, and a large proacrosomal vesicle located at the opposite side of the cell. The most recent spermiogenic transformations primarily concern elongation of the proacrosomal vesicle to form a tapering spike. This results in the typically tack-shaped sperm of natantian decapods. The initial steps of spermiogenesis in the two studied dendrobranchiates prove to be parallel to reptant spermiogenesis in some respects, namely rupture of the nuclear envelope, chromatin decondensation and differentiation of electron-dense regions within the proacrosomal vesicle content. Specifically, whereas the anteriormost condensation gives rise to the operculum in brachyurans, in dendrobranchiates it becomes the apical portion of the spike. Despite an unquestionable morphological similarity between the sperm of carideans and dendrobranchiates, spermiogenesis in both groups displays meaningful differences. Spermatids of caridean shrimps lack a distinct proacrosomal vesicle. In the course of spermiogenesis, the spike arises from aggregated cytosolic materials; hence it is not membrane-bound. Unlike in other decapods, caridean sperm do not undergo a conventional acrosome reaction, since exocytotic events are not involved in this process. The above arguments suggest that, in the Decapoda, separation into three sperm classes is more suitable than the two traditionally accepted classes. The dendrobranchiate and reptant sperm types share a number of spermiogenic and functional features, while the caridean sperm type appears to represent an independent evolutive line with regard to sperm development and function.     

Acrosome differentiation and the acrosome reaction in ascidian spermatozoa:<Emphasis Type="Italic"> Ascidiella aspersa</Emphasis> and<Emphasis Type="Italic"> Ascidia mentula</Emphasis> with some implications for tunicate phylogeny

The spermatozoa of both Ascidiella aspersa and Ascidia mentula have architectural features characteristic of ascidian spermatozoa that have previously been described. They have an elongated head (7 µm long for A. aspersa and 4 µm long for A. mentula), a single mitochondrion that is applied laterally to the nucleus and lacks a midpiece. The acrosome of A. aspersa spermatozoa is a flattened vesicle, about 200 nm×100 nm×40 nm (length, width and height). The acrosome of A. mentula spermatozoa consists of multiple vesicles; they are about 50 nm×50 nm×40 nm (length, width and height). During spermiogenesis in both species, several proacrosomal vesicles (50–70 nm in diameter) appear in a blister at the future apex of the spermatid. In A. aspersa, these vesicles fuse with each other to form a single acrosomal vesicle, while in A. mentula these vesicles do not fuse with each other, and form multiple acrosomal vesicles. In A. aspersa spermatozoa, calcium ionophore A23187 induces the acrosome reaction in which membrane fusion between the acrosomal apical membrane and the overlying sperm plasma membrane occurs along the peripheral margin of the acrosome, resulting in the release of a hybrid, membrane-bound, small vesicle. In A. mentula, multiple acrosomal vesicles disappear by releasing small vesicles after treatment with the calcium ionophore A23187; this also appears to be an acrosome reaction. This paper discusses the way in which acrosome structure and function may have changed during the evolution of the Tunicata.Communicated by T. Ikeda, Hakodate     

Ultrastructure of the spermatozoa of Hymenosomatidae (Crustacea: Brachyura) and the relationships of the family

The spermatozoa of the genus Odiomaris Ng and Richer de Forges, 1996 (=Amarinus Lucas, 1980) have the components typical of eubrachyuran (Heterotremata + Thoracotremata) sperm, but differ significantly from all other investigated eubrachyurans in at least ten characteristics: (1) presence of an epiopercular dome; (2) separation of all but the central region of the operculum from the remainder of the acrosome by an infra-opercular rim; (3) the fact that the acrosome is smaller in volume than the nucleus; (4) the acrosome is strongly emergent from the nucleus, being surrounded only basally by nuclear material; (5) the cytoplasmic sheath, ending anteriorly with the nucleus, is also basal; (6) division of the acrosome contents into an inner and outer acrosome zone is scarcely apparent in longitudinal section as the inner zone is narrow and of doubtful homology; (7) the thin, putative inner acrosome zone is anteriorly almost septate owing to several longitudinal corrugations; (8) basally there is a unique “fringe zone”; (9) the acrosome, including the epinuclear dome, is longer than wide; (10) the unique helical and posterolateral disposition of the nuclear arms. From a purely spermatological viewpoint, Odiomaris (as exemplified by O. pilosus and O. estuarius), and provisionally the Hymenosomatidae, are thus excluded from the Thora- cotremata, in which they were formerly placed, nor are they readily placeable in the Heterotremata. Received: 30 December 1996 / Accepted: 4 February 1997     

Spermatogenesis inPyrosoma atlanticum (Tunicata: Thaliacea: Pyrosomatida): Implications for tunicate phylogeny

Colonies ofPyrosoma atlanticum were collected by submersible in October 1988 in the Caribbean Sea, and testes were studied by electron microscopy. Spermatogonia, spermatocytes and early spermatids have two centrioles. The proximal centriole subsequently disappears, its remains apparently persisting in the spermatozoon as dense material adjacent to the distal centriole, which gives rise to the axoneme. At the tip of early spermatids are several 50 nm proacrosomal vesicles, which disappear leaving no trace in early elongating spermatids. The spermatozoon lacks an acrosome and has a head 35µm long. The head is differentiated into a bulbous posterior portion 5µm long × 1µm wide, a thinner anterior portion 25µm long tapering from a width of 0.7µm to a width of 0.4µm, and a very thin anterior extension 5µm long × 0.5µm wide. At the start of elongation, the anterior extension begins to form just lateral to the proacrosomal vesicles as a spiral projection comprising part of the nucleus, covered by a thin sheath of cytoplasm. This sheath of cytoplasm undergoes a complex differentiation. Ultimately, the nucleus in the anterior extension is overlain by two membrane-bound sheaths of cytoplasm connected by a spiral flange of cytoplasm. Between these two sheaths is a spiral space, open to the exterior through a subterminal pore near the sperm tip. In early spermatids the mitochondria fuse into a single mitochondrion, which remains lateral to the nucleus. The cristae become modified late in spermatogenesis. Throughout elongation of the spermatid there are patches of dense material between the nucleus and mitochondrion. A manchette of microtubules transiently encircles the thin anterior portion of the nucleus during the last phase of elongation. A manchette is not present during most of elongation. In the spermatozoon the mitochondrion, which has reticulate cristae, spirals a few times about the nucleus and extends from the junction between the bulbous portion and the thinner anterior portion of the nucleus to the junction between the thinner anterior portion and the nuclear extension. Spermatogenesis inP. atlanticum, compared to that in other tunicates, most closely resembles that in colonial ascidians, and supports the majority view that pyrosomes arose from aplousobranch ascidians that lost their attachment to the substratum. Pyrosome sperm are more highly derived than doliolid sperm, which have an acrosome that is probably capable of exocytosis. When salp and pyrosome sperm are compared, both are highly derived, but neither shares any apomorphies with the other that it does not share with at least one other tunicate order. Thus, sperm morphology does not support the majority view that pyrosomes gave rise to doliolids and neither confirms nor denies the idea that pyrosomes are intermediate between aplousobranch ascidians and salps. Therefore, it is likely that the class Thaliacea is polyphyletic, with doliolids arising very early from the ascidian lineage and with salps and pyrosomes arising somewhat later.     

Spermatogenesis,sperm storage and comparative sperm morphology in nine species of Capitella,Capitomastus and Capitellides (Polychaeta: Capitellidae)

Light microscopy and transmission electron microscopy (TEM) were used to describe spermatogenesis and the morphology of mature sperm and sperm storage organs in five sibling species of Capitella, three species in the related genus Capitomastus, and one species in the genus Capitellides. These capitellids lack a well-developed testis, but young males have a few specialized regions of the peritoneum in the eighth setiger, where germ cells proliferate and spermatogonia are released into the coelom, and spermiogenesis is completed. Mature sperm are stored in the central regions of paired genital ducts (coelomoducts), which lie between the seventh and eighth setigers. The cells forming the walls of the coelomostome and central region of the duct are ciliated and have large glycogen deposits. The lumenal borders have extensive microvilli and there is evidence that they secrete glycogen-containing materials into the duct. All species have modified primitive sperm with a conical acrosome, elongated nucleus, and long middle piece extending along the proximal portion of the flagellum. A single ring-shaped mitochondrion encircles the centriolar region of the middle piece and the cytoplasm is filled with glycogen. The sperm of all nine species differ significantly in the lengths of their middle pieces, acrosomes and especially in their nuclear lengths. The nuclear lengths have a twofold range among the sibling species of Capitella and Capitomastus. Subtle differences in the shape and volume of the acrosomal vesicle and acrosomal space characteristic of the Capitella sibling species seem to correlate with a basic division of these species into those with diploid chromosome numbers of 20 or 26. Spermiogenesis, the number of sperm produced, and the method of sperm storage are appropriate for efficient sperm utilization in fertilization. No evidence indicates that spermatophores are formed and transferred between individuals and the method of sperm transfer is not understood. The differences in the dimensions and acrosome morphology of mature sperm, and the previously demonstrated specializations in the egg envelopes in the Capitella sibling species, are characteristic features of the reproductive isolation that exists among these capitellid species.     

Podotreme affinities of Raninoides sp. and Lyreidus brevifrons: evidence from spermatozoal ultrastructure (Crustacea: Brachyura: Raninoidea)

Spermatozoal synapomorphies which singly or collectively distinguish Raninoidea are: (1) the presence of single (Ranina, Raninoides) or multiple (Lyreidus) keel-like projections of the acrosomal capsule; (2) a very large, weakly electron-opaque peripheral acrosomal zone (Ranina, Raninoides) and an homologous large outer zone in Lyreidus; (3) poor differentiation of the operculum from the capsule (autapomorphy); (4) a very well developed, perforate subopercular zone, of variable form; (5) presence of unique inward longitudinal projections (septa or corrugations) in the wall of the subacrosomal chamber (autapomorphy). Shared, presumably synapomorphic characters of Ranina and Raninoides but not of Lyreidus within the Raninidae, are: (1) branching of some of the subacrosomal septa (unbranched in Lyreidus); (2) the subspheroidal form of the acrosome in Ranina with a length: width ratio (L:W) of 0.76, and, although slightly more depressed, in Raninoides (L:W 0.73), considered apomorphic relative to the more depressed form in Lyreidus (L:W 0.52); (3) single or multiple coiled perforatorial filaments (Ranina, Raninoides) contrasting with a capitate perforatorium with “amoeboid” head in Lyreidus; (4) division of the capsule wall to give one posterior (Ranina) or multiple enclaves, plesiomorphically (?) absent in Lyreidus. Similarities of Lyreidus with other podotremes include the capitate perforatorium, questionably related to the radiate spiked-wheel structure of homolids in which acrosome proportions are similar or less closely to the bilateral capitate perforatorium of dromiids and dynomenids, and basal capsular projections as in the dromiid Stimdromia (=Petamolera) lateralis and in cyclodorippoids. No spermatozoal synapomorphies support a sister-group relationship of raninoids and heterotreme-thoracotreme crabs.     

Fine structure of spermatids and sperm of Dolioletta gegenbauri and Doliolum nationalis (Tunicata: Thaliacea): implications for tunicate phylogeny

In Dolioletta gegenbauri and Doliolum nationalis, collected in 1987 in the bay of Villefranche-sur-mer (French Mediterranean Sea), spermiogenesis is essentially the same. Early spermatids have a round head, a flagellum arising from a single centriole with short microtubules at 45° to its base, several mitochondria, and an acrosome 50 nm thick and 250 nm long with its long axis parallel to the plasma membrane. The acrosomal contents are dense, with a central denser plate. The nuclear envelope next to the acrosome is thickened and concave. In elongating nuclei, strands of chromatin become oriented parallel to the length of the nucleus and then twist helically. Although the mitochondria surround the nucleus, they remain relatively short and do not fuse into a single mitochondrion as in sperm of other tunicates. In very late doliolid spermatids, the acrosome undergoes exocytosis, and exposes fibrous material that stays associated with the tip of the sperm; no acrosomal tubule forms. Exocytosis at this stage may be triggered by fixation. If so, exocytosis probably occurs naturally at some time before fusion of sperm and egg. Sperm have elongate heads (1 m×10 m), with the anterior two-thirds of the nucleus surrounded by mitochondria. Spermiogenesis in doliolids, compared to that in other tunicates, is most like that in solitary members of the class Ascidiacea, except that in the latter the sperm mitochondria fuse and the acrosome appears incapable of exocytosis. In contrast, previous work has shown that salps (class Thaliacea) and colonial didemnid ascidians have an acrosomeless sperm with a spiral mitochondrion, while the class Appendicularia has a sperm with a midpiece, a compact head and an acrosome capable of exocytosis and acrosomal tubule formation. By outgroup comparison with echinoderms and acraniates, appendicularian sperm are plesiomorphic within the Tunicata. Thus, gamete morphology indicates that (1) solitary ascidians and doliolids had a common ancestor, (2) the popular idea that doliolids gave rise to appendicularians is incorrect, and (3) the Thaliacea are polyphyletic, doliolids having arisen very early from the ascidian lineage and salps having arisen later.     

Spermatozoon structure of some North American prosobranchs from the families Lottiidae (Patellogastropoda) and Fissurellidae (Archaeogastropoda)

The spermatozoa of four species of the patellogastropod family Lottiidae (Lottia pelta, L. digitalis, L. strigatella, Tectura scutum) and one species of the archaeogastropod family Fissurellidae (Diodora aspera) were examined in 1990 using transmission electron microscopy. All have primitive or ect-aquasperm, typical of invertebrates using external fertilization. Sperm of the lottiid limpets are characterized by a 5 to 9 m-long head composed of a conical acrosome which constitutes >50% of the head length, and a cylindrical nucleus. The acrosome of all species of lottiids is differentiated internally, and has a posterior invagination 0.9 to 1 m in depth, into which an elongate acrosomal lobe protrudes. Between the posterior acrosomal lobe and the nucleus, the subacrosomal material is aggregated as a fibrous column. The midpiece of the sperm has a ring of 4 to 5 spherical mitochondria of 0.6 m diam, posterior to which is a collar of cytoplasm 1 m long, which sheaths the anterior portion of the axoneme. The size and morphology of the acrosome and large cytoplasmic collar clearly distinguish the spermatozoa of the Lottiidae from other families of Patellogastropoda. The sperm of D. aspera (Fissurellidae) is typical of the family of archaeogastropod; the head has a length to breadth ratio of 4:1, and the cylindrical nucleus is capped by a small acrosome, <25% of the total head length, which is deeply invaginated.     

Variation in fertilisation abilities between hemiclonal hybrid and sexual parental males of sympatric water frogs (<Emphasis Type="Italic">Rana lessonae</Emphasis>,<Emphasis Type="Italic"> R. esculenta</Emphasis>,<Emphasis Type="Italic"> R. ridibunda</Emphasis>)

In many species, males and females mate with multiple partners, which gives rise to sperm competition and multiple paternity. The experiments on water frogs presented here demonstrate that such sperm competition can affect the structure and dynamics of mixed-species communities. The hybrid frog Rana esculenta (LR) mates with one of its parental species, usually R. lessonae (LL), although in some areas R. ridibunda (RR), to regain the premeiotically eliminated parental genome ("hybridogenesis"). Mixed LL/LR-populations are stable although hybrid numbers should continuously increase at the expense of parental animals, because of differences in female fecundity and other factors. This would finally lead to the extinction of the sexual host, followed by that of the sexual parasite, unless the reproductive superiority of R. esculenta is reduced by other factors, such as lower hybrid male fertility. Eggs from LL- and LR-females were fertilised in vitro by single- and multi-male sperm suspensions of LL-, LR- and RR-males. In all experiments, the proportion of offspring sired by R. esculenta sperm was significantly lower than that sired by R. lessonae or R. ridibunda sperm. Gonad mass, sperm morphology, sperm swimming velocity, and sperm survival did not explain these differences in fertilisation success; nor did gamete recognition and compatibility. Sperm density was the only trait that paralleled fertilisation success, but it offers no explanation either, because densities were equalised for the in-vitro fertilisations. In natural LL/LR populations, the significantly smaller amount, poorer competitive ability and lower long-term survival of R. esculenta compared to R. lessonae sperm will reduce the initial reproductive superiority of hybrids and contribute to the stabilisation of mixed water-frog populations. Differences in fertilisation ability are also likely to be relevant for the structure and dynamics of several other systems with encounters between eggs and sperm from different genotypes, ecotypes, ploidy levels and/or species.     

Juvenile length-at-age data reveal that spanner crabs (<Emphasis Type="Italic">Ranina ranina</Emphasis>) grow slowly

The spanner crab (Ranina ranina) is a widespread and abundant brachyuran in offshore sand substrata of the Indo-Pacific region. Little is known of this species biology, population dynamics and ecology, despite it being the target of commercial fishing operations in many areas. Previous studies of R. ranina growth using length-frequency analysis of samples collected with commercial fishing gear have derived widely divergent estimates of growth parameters. The estimated time taken to reach 100 mm rostral carapace length (minimum legally exploited size in Queensland, Australia) in those studies has ranged from 1.75 to 8.83 years for females and from 1.08 to 3.58 years for males. Our data show that the commercial fishing apparatus used in those studies is size selective and catches only adult crabs. The resulting size bias in samples collected using that apparatus precludes the application of length-frequency-based techniques to estimate growth parameters from those samples. We devised a new dredge to collect samples of juvenile R. ranina and to calculate juvenile growth rates from modal progression in those samples. We combined those data with estimated mean maximum lengths (L) of 121.7 mm for females and 155.9 mm for males from commercial catch data to model other von Bertalanffy growth parameters using bootstrap methods. Those modelled parameters (K=0.29, T₀=–0.24 for females; K=0.23, T₀=–0.25 for males) indicate that R. ranina grows more slowly than most previous estimates suggest, with females requiring an average of 6.35 years and males 4.31 years to reach 100 mm rostral carapace length. This slow growth is consistent with the slow metabolism of R. ranina, and indicates that this species would be likely to recover slowly from overexploitation.Communicated by G.F. Humphrey, Sydney     

Effects of water motion on propagule release from algae with complex life histories

Reproductive marine algae with complex life histories may respond differently to water motion depending upon whether the spore-producing or gamete-producing phase is considered. Two such species, the kelp Alaria esculenta (L.) Greville (Phaeophyceae) and Ulva lactuca L. (Chlorophyta), were examined experimentally in the laboratory. The kelp was collected in April–June 2000 and U. lactuca was collected in July–August 2000, from four intertidal habitats at Schoodic Point, Maine, USA. Orbital shakers were used to generate water motion. Sporophylls of A. esculenta released more zoospores under shaken versus calm conditions, whereas fewer antheridia on the microscopic male gametophytes released sperm under shaken versus calm conditions when male and female gametophytes were placed together. However, antheridial sperm release was equivalent when male gametophytes were exposed to undiluted media from dense cultures of mature female gametophytes under shaken versus calm conditions. These data suggest that water motion inhibited sperm release by diluting the sperm-releasing pheromone produced by ripe eggs below a threshold required to cause sperm release. Water motion stimulated both gamete and zoospore release from U. lactuca. This is the first report in an alga of stimulation of gamete release by increased water motion, but it is notable that parthenogenesis occurs in Ulva spp.; thus, gametes may develop into gametophytes (1 N) or parthenosporophytes (2 N). This study demonstrates that water motion has strong effects upon algal reproduction, but that the effects may vary between species, possibly due to their different life histories.Communicated by J.P. Grassle, New Brunswick     

Ultrastructure of sperm and spermathecae in Micromaldane spp. (Polychaeta: Capitellida: Maldanidae)

Assessing the possibility that external fertilization has re-evolved requires the study of monophyletic groups that exhibit various reproductive methods. Maldanid polychaetes show a range of reproductive mechanisms, though previous studies of reproduction have hitherto been restricted to larger species with external fertilization. Micromaldane pamelae Rouse and M. nutricula Rouse are small, gonochoristic maldanids that brood directly developing larvae. Both species have sperm with elongate nuclei and an acrosome extending down each side of the anterior end of the nucleus. A true midpiece is absent; two mitochondria extend along the posterior region of the nucleus. Spermatids develop synchronously in large clusters connected by a cytophore. In M. pamelae sperm are released into the water as spermatozeugmata. These are comprised of clusters of sperm with their tails oriented to the centre and the sperm heads facing outwards. Females of M. pamelae and M. nutricula bear pairs of spermathecae ventrally (M. pamelae three pairs, between setigers 10 and 11, 11 and 12 and 12 and 13 and M. nutricula two pairs, between setigers 10 and 11 and setigers 11 and 12). The blind sacs are epidermal invaginations bound closely together. The entrance to each spermatheca may only be 1 to 2 m across with each spermatheca holding several hundred sperm. This represents the first detailed study of spermathecae in the Capitellida. The occurrence and structure of spermathecae and spermatozeugmata in other groups are discussed and compared with Micromaldane spp. Comparisons are made with non-polychaetes with the purpose of discussing functional aspects of reproductive mechanisms in marine metazoans in general. Elongate sperm nuclei are associated with sperm storage and/or large egg size. The lack of an elongate sperm midpiece may be an indicator of having to swim in water but does not contraindicate sperm storage. Spermatozeugmata may serve as an indication of sperm storage and brooding of larvae. Speculations on the phylogenetic significance of these reproductive features are limited by the fact that supposedly modified (i.e., derived) states may reflect functional/structura, constraints of small body size.     

Sibling species in interstitial flatworms: a case study using<Emphasis Type="Italic"> Monocelis lineata</Emphasis> (Proseriata: Monocelididae)

The genetic relationships between morphologically indistinguishable marine and brackish-water populations of Monocelis lineata (O.F. Müller, 1774) (Proseriata: Monocelididae) were analysed by means of allozyme electrophoresis. Fifteen samples of M. lineata (13 from the Mediterranean and two from the Atlantic) from coastal marine and brackish-water habitats were examined for variation at 18 loci. Eleven loci were polymorphic in at least one population of M. lineata. Low levels of within-population genetic variability were found, with average observed and expected heterozygosity values ranging from H_o=0.015±0.015 to 0.113±0.044, and from H_e=0.028±0.028 to 0.138±0.054, respectively. The occurrence of a number of private alleles indicated a marked genetic divergence among populations of M. lineata, with Rogers genetic distances ranging from D_R=0.003 to 0.676 and a highly significant F_ST value (0.918±0.012, P<0.001). UPGMA (unweighted pair-group method using arithmetic average) cluster analysis and multidimensional scaling showed a clear genetic divergence between marine and brackish-water populations. Moreover, Atlantic and Mediterranean populations were sharply separated. Our results suggest that M. lineata is a complex of sibling species.Communicated by R. Cattaneo-Vietti, Genova     

Some polyphagous Homoptera gain protection from their natural enemies by feeding on the toxic plantsSpartium junceum andErythrina corallodendrum (Leguminosae)

Summary Interactions were studied among alkaloid-containing legumes (Erythrina corallodendrum andSpartium junceum) and non-toxic plants (Citrus sinensis, Cucurbita moschata andEuphorbia tirucalli), several polyphagous homopterans,Aphis craccivora (Aphididae),Icerya purchasi, I. aegyptiaca (Margarodidae),Lepidosaphes ulmi (Diaspididae) andPlanococcus citri (Pseudococcidae), and some major natural enemies of these homopterans. Significant reductions in survival due to negative effects of alkaloid containing as compared with non-alkaloidal plants were recorded for the predatorsRodolia cardinalis andChilocorus bipustulatus, but not forCryptolaemus montrouzieri (Coleoptera: Coccinellidae),Chrysoperla carnea (Neuroptera: Chrysopidae) andSympherobius sanctus (Neuroptera: Sympherobiidae). The development time of the larvae or pupae ofR. cardinalis, C. carnea andS. sanctus was longer on the toxic plants than on the non-toxic ones. The percentage of parasitism ofA. craccivora collected from the non-alkaloidal plantsVicia palaestina andMelilotus albus was much higher than that onS. junceum. The parasitoid complexes ofA. craccivora differed between both plant groups. The nutritive value of honeydew ofI. purchasi andA. craccivora, as expressed by the life span ofEncyrtus infelix (Hymenoptera: Encyrtidae) adults, was also investigated. Life spans were significantly longer when the wasps fed on honeydew produced on non-alkaloidal plants (C. sinensis andPittosporum tobira) than on alkaloid containing plants whenI. purchasi — but notA. craccivora — was the producer. It is suggested that the chemical defense ofE. corallodendrum andS. junceum is exploited by polyphagous phytophages to reduce predation. In nature, population growth and density of four of the investigated homopterans are conspicuously high when they developed on the alkaloid containing plant species, and very low on non-alkaloid plants. The efficiency of their natural enemies may be reduced by sequestration of alkaloids (or other toxic plant compounds) or their transfer into excreted honeydew. Therefore it is assumed that a generalist phytophagous homopteran may be protected from its natural enemies, although at different rates of efficiency, if it can safely sequester the host allelochemical when it develops on toxic species within its host range.     

Structure of the sperm of some South African archaeogastropods (Mollusca) from the superfamilies Haliotoidea,Fissurelloidea and Trochoidea

The structure of the spermatozoa of 11 species from five families of archaeogastropod (Haliotidae, Fissurellidae, Trochidae, Turbinidae, Phasianellidae) has been examined using transmission electron microscopy. All sperm are of the primitive, or ect-aquasperm type and each species has a sperm head (nucleus and acrosome) with unique morphology. Furthermore, the results from the present study together with those published in the literature reveal that, although there are a few exceptions, the size (length to breadth ratio) and spape of the nucleus and acrosome of sperm of species within each family are similar. It is therefore possible to identify members of a family and differentiate between families using sperm morphology. The intrafamilial similarities and interfamilial differences in sperm structure indicate that a spermiocladistic study of the suborder Vetigastropoda could provide interesting insights into the phylogenetic relationships of this group.     

Catch rate and emergence of male and female spanner crabs (Ranina ranina) in Australia

Catches of the spanner crab, Ranina ranina (Linnaeus, 1788), in southern Queensland, Australia in 1982 and 1983 showed a seasonal cycle, with peak catches from August to September. In most catches, males outnumbered females. Catches of both male and female spanner crabs declined in November and December, when a high proportion of females caught were ovigerous. Fish, echinoderms and polychaetes were the main food items. Closed-circuit time-lapse video recording was used to study the emergence behaviour of R. ranina in a 6 m-diameter tank. Spanner crabs in this tank were buried most of the time, emerging on average for only 1.7 h d^-1. They emerged mainly between 16.00 and 24.00 hrs and, less often, between 01.00 and 15.00 hrs. Males remained emerged significantly longer than females. The duration of emergence of the females peaked before the spawning season, becoming shorter during the period when they were ovigerous. Temperature and emergence were negatively correlated. It is concluded that reproductive behaviour has a strong influence on catch composition of spanner crabs.     

Crinoid phylogeny: a preliminary analysis (Echinodermata: Crinoidea)

We describe the first molecular and morphological analysis of extant crinoid high-level inter-relationships. Nuclear and mitochondrial gene sequences and a cladistically coded matrix of 30 morphological characters are presented, and analysed by phylogenetic methods. The molecular data were compiled from concatenated nuclear-encoded 18S rDNA, internal transcribed spacer 1, 5.8S rDNA, and internal transcribed spacer 2, together with part of mitochondrial 16S rDNA, and comprised 3,593 sites, of which 313 were parsimony-informative. The molecular and morphological analyses include data from the bourgueticrinid Bathycrinus; the antedonid comatulids Dorometra and Florometra; the cyrtocrinids Cyathidium, Gymnocrinus, and Holopus; the isocrinids Endoxocrinus, and two species of Metacrinus; as well as from Guillecrinus and Caledonicrinus, whose ordinal relationships are uncertain, together with morphological data from Proisocrinus. Because the molecular data include indel-rich regions, special attention was given to alignment procedure, and it was found that relatively low, gene-specific, gap penalties gave alignments from which congruent phylogenetic information was obtained from both well-aligned, indel-poor and potentially misaligned, indel-rich regions. The different sequence data partitions also gave essentially congruent results. The overall direction of evolution in the gene trees remains uncertain: an asteroid outgroup places the root on the branch adjacent to the slowly evolving isocrinids (consistent with palaeontological order of first appearances), but maximum likelihood analysis with a molecular clock places it elsewhere. Despite lineage-specific rate differences, the clock model was not excluded by a likelihood ratio test. Morphological analyses were unrooted. All analyses identified three clades, two of them generally well-supported. One well-supported clade (BCG) unites Bathycrinus and Guillecrinus with the representative (chimaeric) comatulid in a derived position, suggesting that comatulids originated from a sessile, stalked ancestor. In this connection it is noted that because the comatulid centrodorsal ossicle originates ontogenetically from the column, it is not strictly correct to describe comatulids as unstalked crinoids. A second, uniformly well-supported clade contains members of the Isocrinida, while the third clade contains Gymnocrinus, a well-established member of the Cyrtocrinida, together with the problematic taxon Caledonicrinus, currently classified as a bourgueticrinid. Another cyrtocrinid, Holopus, joins this clade with only weak molecular, but strong morphological support. In one morphological analysis Proisocrinus is weakly attached to the isocrinid clade. Only an unusual, divergent 18S rDNA sequence was obtained from the morphologically strange cyrtocrinid Cyathidium. Although not analysed in detail, features of this sequence suggested that it may be a PCR artefact, so that the apparently basal position of this taxon requires confirmation. If not an artefact, Cyathidium either diverged from the crinoid stem much earlier than has been recognised hitherto (i.e., it may be a Palaeozoic relic), or it has an atypically high rate of molecular evolution.Electronic Supplementary Material Supplementary material is available in the online version of this article at Communicated by J.P. Thorpe, Port Erin     

Reproduction of sympatric populations ofHeliocidaris erythrogramma andH. tuberculata (Echinoidea) in New South Wales

Reproduction ofHeliocidaris erythrogramma (Valenciennes) andH. tuberculata (Lamarck) was compared through examination of oogenesis, spermatogenesis and monthly measurement of the gonad index. These species occur sympatrically in the Sydney region. Their reproduction was examined at two sites near Botany Bay, New South Wales, Australia, from February 1989 through January 1990.H. erythrogramma produces buoyant, 450µm-diam eggs and the sperm have a head region 10µm in length. By contrast,H. tuberculata produces negatively-buoyant, 95µm-diam eggs and the sperm have a head region 4µm in length. Histochemical examination of the gonads revealed that periodic acid Schiff-positive (PAS +) material stored in the nutritive phagocytes appears to support vitellogenesis in both species. InH. tuberculata this material is utilized in the formation of PAS + yolk oligolecithal eggs, whereas inH. erythrogramma the PAS + material appears to be converted to lipid yolk in macrolecithal eggs.H. erythrogramma had a seasonal reproductive pattern with a 3 mo summer spawning period, whereas both populations ofH. tuberculata had a 9 mo breeding period characterized by the continual presence of nutrient reserves and vitellogenic oocytes which rapidly replaced spawned ova. Spawning ceased only for 3 mo over the summer. Due to the 9 mo spawning ofH. tuberculata it is not clear what factors serve to cue reproduction in this species.     

Comparative fertilization biology of gymnolaemate bryozoans

Locations and oogenic stages of eggs at the time of sperm-egg fusion were determined for nine gymnolaemate species exhibiting different reproductive strategies with respect to site of embryonic development (e.g., water column, introvert, embryo sac, or ovicell) larval type, and zooid sexuality. Ovarian, coelomic, and recently spawned oocytes obtained from freshly collected colonies between 1988 and 1995 were stained with aceto-orcein and examined for the presence of a sperm nucleus. All nine species are shown to have a similar fertilization biology in which sperm fuse with eggs in or near ovaries and egg activation is delayed until eggs are spawned. In species such as the etenostomeAlcyonidium sp. and the cheilostomeElectra pilosa (Linnaeus, 1767), which produce marierons oocytes that are spawned into the water column and have cyphonautes larvae, sperm fuse with late growth stage ovarian oocytes after germinal vesicle breakdown (GVBD) at or near ovulation. In species that produce few oocytes, brood embryos, and release coronate larvae, sperm fuse with late growth stage ovarian primary oocytes before GVBD, as in the ctenostomeBowerbankia gracilis Leidy, 1855, or with very early growth stage ovarian primary oocytes, as in the cheilostomesCribrilina corbicula (O'Donoghue, 1923),Dendrobeania lichenoides (Robertson, 1900),Hippodiplosia insculpta (Hincks, 1882),Schizoporella serialis (Heller, 1867),T ricellaria gracilis (Smitt, 1867), andWatersipora arcuata Banta, 1969. Fertilization success was high in all nine species, with 83 to 100% of oocytes at an appropriate oogenic stage containing a sperm nucleus or early male pronucleus. Gymnolaemate bryozoans may maintain high levels of fertilization success by: (1) concentrating spawned sperm from the water column using colony feeding currents; (2) localizing sperm-egg encounters in or near the ovary, (3) using sperm when available by permitting sperm to fuse with early growth stage oocytes; and (4) selfing.