Chondrocladia (Symmetrocladia) lyra subgen. nov., sp. nov., is described from northeast Pacific sites at Escanaba Ridge and Monterey Canyon at depths of 3316–3399 m. Two retrieved specimens are described in detail, while variations are described in ten photographed or videotaped specimens. The basic structure, termed a vane, is harp- or lyre-shaped. From 1 to 6 vanes extend by radial growth from the organism's center. The orientation among the vanes is approximately equiangular, such that together they display pentaradiate, tetraradiate, triradiate, or biradiate symmetries. Each vane is formed by a horizontal stolon supporting a series of upright, equidistantly spaced branches each of which terminates at its apex in a swollen ball in all observed specimens except the paratype. Swellings occur midway along the branches in the holotype, but not in the paratype. A linear row of filaments project from the sides, front, and back of each branch, and also from the tops of each stolon. The terminal balls are the sites of spermatophore production and release; mid-branch swellings are sites of oocyte maturation. The two megasclere spicule types have specific distributions; styles support rhizoids, stolons, and branches, while subtylostyles support filaments and terminal balls. Anchorate isochelae cover all surfaces. Enclosed crustacean prey on branches and stolons provide direct evidence of carnivory. The structure of the vanes maximizes surface area for passive suspension feeding. Increased surface area could also maximize spermatophore capture, with the sigmas projecting from the spermatophore surface being caught by projecting isochelae on filaments. Swellings on filaments are snared spermatophores, firmly fused to recipient tissues and undergoing destruction. Spermatophores on filaments are present in branch swellings containing early and mature oocytes. Oogenesis and maturation occur only in proximity to branch swellings, suggesting that development is induced by spermatophore reception. Symmetrical development of uniserial branched stolons (the vanes) characterized members of the new subgenus Symmetrocladia.

In freshwater systems, parasitological studies have mainly been carried out on vertebrates and molluscs, but little is known about parasites of aquatic insects. We describe the trematodes and nematodes parasitizing the benthic insects of an Andean Patagonian stream and the presence of parasites in the terrestrial adult stages. Members of 3 of 20 insect taxa were found to be parasitized by larval nematodes, and members of six taxa harbored metacercariae of digeneans. In benthic samples, chironomids, simuliids (Order Diptera), and baetids (Order Ephemeroptera) harbored mermithid larvae (Nematoda). The stonefly Antarctoperla michaelseni (Order Plecoptera), the caddisfly Smicridea annulicornis (Order Trichoptera), a watersnipe fly (Order Diptera: Athericidae), and three species of leptophlebiid mayflies (Order Ephemeroptera) were parasitized by encysted plagiorchiid metacercariae (Order Plagiorchiida). Most metacercariae were found in the three species of mayflies with prevalences ranging 15–63% and mean intensities ranging 1.2–4.9. Prevalence declined from summer to early winter, probably because of the emergence of infected nymphs and the recruitment of uninfected new cohorts. The imagos had live metacercariae with higher prevalences and intensities of infection than nymphs. We suggest that these plagiorchiids have an allogenic life cycle, involving a terrestrial definitive host.

The ecology of “Gordian knots” – aggregates of entwined reproductive individuals of nematomorphs, or hair worms – remains poorly understood. We studied the ecology of Gordian knots in a wild population of the hair worm Paragordius tricuspidatus sampled from a stream near Montpellier, France, in 2005 and 2006. At our study site, knots were composed entirely of males. The number of adults per knot varied throughout the summer, most likely due to the seasonal decrease in stream water level and the appearance of females later in the season. The average length of individual worms in each knot did not change significantly over time, but it became significantly more variable. We also showed that knots within these streams were more likely to be attached to floating rather than immobile substrates. Because the water levels in our sampling stream decrease as the season progresses, males in knots attached to floating substrates could be in a better position to cope with decreasing water levels, as they always remain immersed. An important avenue of future study would be to explore the effects of varying water level and the presence of females on the size and maintenance of knots in laboratory conditions.

The presence of sibling species within the marine gastropod genus Crepipatella has complicated the taxonomy of members of the group. Since the establishment of the genus, 15 species have been described, but recent studies have indicated that there are only five valid species, two of which inhabit the coasts of Chile, namely C. dilatata and C. fecunda. The two species are morphologically indistinguishable as adults, but can be differentiated on the basis of their encapsulated developmental stages. The primary aim of this study was to reconstruct phylogeny within the genus, and to establish species limits of C. dilatata and C. fecunda, using mitochondrial DNA data. To this end, we used maximum parsimony, maximum likelihood, and Bayesian inference to reconstruct phylogenies using 589 bp of the cytochrome oxidase I (COI) gene. The mtDNA phylogenies were then used as input in a general mixed Yule-coalescent (GMYC) analysis to estimate species boundaries. In addition, quarter likelihood mapping was used to test a posteriori the confidence of inner branch patterns in the phylogenetic tree. Both DNA tree-based and GMYC methods provide support for five isolated lineages within this species complex. Our data also suggest that Late Pleistocene and Holocene fragmentation and subsequent range expansion events may have shaped contemporary genetic patterns of Crepipatella in South America.

Vitellogenin (VTG), the precursor of yolk protein, plays an important role during vitellogenesis and oogenesis. However, little is known about the vitellogenin gene in marine bivalves. We cloned the full-length cDNA of Chlamys farreri vitellogenin (Cf-vtg) using a homologous cloning strategy and rapid amplification of cDNA ends. The full-length cDNA consisted of 7604 nucleotides with an open reading frame encoding 2296 amino acid residues. The deduced amino acid sequence shared high similarity with the VTG of other species, particularly in the N-terminal region. We analyzed the spatiotemporal expression of Cf-vtg transcripts by semiquantitative and quantitative real-time PCR. Cf-vtg was primarily expressed in the ovary, and levels were 1.917±0.035 (mean±S.E.) and 2.570±0.252 times higher in growing and mature stages, respectively, than in the proliferative stage. There was negligible expression at the resting stage. Interestingly, we observed weak expression in the female hepatopancreas. The level was 4.50±0.90% (mean±S.E.) that in the ovary during the proliferative stage. We did not detect any expression in testis or in other tissues. Using in situ hybridization, Cf-vtg transcripts were localized to the follicle cells surrounding the oocytes in the ovary. Moreover, synthesis of Cf-vtg transcripts in the ovary increased when estradiol-17β was injected in vivo into the gonads of early growing stage. We conclude that VTG synthesis is heterosynthetic in C. farreri. Also, we hypothesize that Cf-VTG synthesis occurs primarily in the follicle cells and is mediated by estradiol-17β.

Serine proteases, ubiquitous enzymes known to function in digestion and immune protection in both vertebrates and invertebrates and implicated in regeneration in some species, were investigated in the California blackworm, Lumbriculus variegatus. Several serine proteases, rather than a single enzyme with broad specificity, were present in tissue extracts from the worms. Extracts were treated with a fluorescein-labeled peptide chloromethyl ketone that specifically binds to trypsin/thrombin-like proteases. Denaturing gel electrophoresis of labeled extracts showed several serine proteases with their molecular weight ranging 28,000–38,000 daltons. The trypsin/thrombin-like activity was localized, using the fluorescein-conjugated reagent, to the pharynx and digestive tract of L. variegatus. Movement of cells labeled by the reagent into regenerating tissues suggests that some differentiated endodermal tissues were used for reformation of digestive structures during regeneration in L. variegatus. The types of serine proteases in the extracts were further characterized by inhibitor studies. Presence of plasmin-like activity was indicated by degradation of fibrin by tissue homogenates from the worms and the inhibitory effect of aprotinin on enzymes in these extracts. The ability of L. variegatus extracts to generate clots when incubated with rabbit plasma and partial inhibition of extract activity by phenylmethylsulfonyl fluoride and hirudin indicated presence of thrombin-like activity. Consistent with the detection of trypsin, chymotrypsin, and plasmin-like enzymes in the extracts was partial inhibition of L. variegatus serine protease activity by aminoethyl benzenesulfonyl fluoride and soybean trypsin inhibitor. Selective inhibition of chymotrypsin-like activity by N-tosyl-L-phenylalanine chloromethyl ketone and chymostatin as well as trypsin-like activity by N-tosyl-L-lysine chloromethyl ketone was observed. A potential role during regeneration for serine proteases is suggested by blockage of formation of head and tail structures by aminoethyl benzenesulfonyl fluoride, an inhibitor of these proteases.

Xerothermic species are rare and threatened in central and eastern Europe. In light of the continuing loss of steppe-like habitats due to anthropogenic fragmentation and degradation, the evaluation of genetic variation in populations inhabiting them is of immediate importance if appropriate conservation measures are to be undertaken. Here we report on the genetic diversity of the rare leaf beetle Crioceris quatuordecimpunctata, whose populations in central and eastern Europe inhabit highly geographically isolated areas. All of the studied populations (in Poland, Ukraine, and Slovakia) were differentiated at the mitochondrial marker COI. However, with respect to the nuclear marker ITS1, Polish populations were monomorphic, but distinct from all other populations. The distinctiveness of the studied populations was confirmed by Wolbachia screening, which showed that all populations carried different strains (one or two), which were probably transferred independently from other insects. On the other hand, no diversity was found in any marker within particular populations, which could be caused (at least for mtDNA) by a Wolbachia selective sweep. Crioceris quatuordecimpunctata probably consists of isolated populations, which went through narrow bottlenecks leading to a drastic reduction in their genetic diversity. As these populations are reciprocally monophyletic for mtDNA haplotypes and show a significant divergence of allele frequencies at nuclear loci, they could be classified as evolutionarily significant units (ESUs). In addition, DNA barcodes were used to identify Asparagus officinalis as the host plant for members of all studied populations. These data should be valuable in efforts to conserve populations of C. quatuordecimpunctata (e.g., for guiding reintroductions).

The morphologically convergent larvae of the echinoderm classes Ophiuroidea and Echinoidea have been suggested to be functionally dissimilar when it comes to their capacities to feed, but little is known about whether these larvae are similar in terms of energetics. Here, we compare the energetics of early development of a tropical ophiuroid, Ophiocoma alexandri, and a temperate to tropical echinoid, Arbacia punctulata, two species with similarly sized eggs. Measurements of respiration and constituent analyses were performed on eggs and unfed larvae of both species. Members of both species showed an increase in oxygen consumption during morphogenesis followed by a lower, static rate once morphogenesis was complete (3 d for O. alexandri and 1.3 d for A. punctulata). Compared to the echinoid larvae, the ophiuroid larvae developed more slowly and had peak respiration rates that were 3.1× lower. Eggs of O. alexandri contained significantly more protein and significantly less triacylglycerol than eggs of A. punctulata. Energy utilization, as calculated via respiration measurements, closely matched decreases in energy content from the eggs to larvae as measured with biochemical constituent assays. Larvae of A. punctulata used 1.4× more energy to reach the pluteus stage than larvae of O. alexandri, and used 4× more energy during the first 9 d of larval life. These data suggest that echinoid larvae require more energy to develop to the feeding stage than ophiuroid larvae, and likewise have higher requirements for maintenance metabolism. Ophiuroid larvae may be more tolerant of low food levels due to their very low metabolic rates, but this advantage may be offset by their slower rate of development.

The shallow water comatulid crinoid Tropiometra carinata is native to both the Atlantic and Indian Oceans, a distribution anomalous among shallow water crinoids and many other broadcast spawning species. Given this species' short pelagic larval duration, the findings of previous work that suggest that the Benguela upwelling is a significant barrier to gene flow in broadcast spawning species, and T. carinata's unexpected geographic distribution, we predicted that the crinoids presently recognized as T. carinata consisted of a species complex. To test this prediction, we sequenced a portion of the mitochondrial cytochrome oxidase 1 gene from 30 individuals of T. carinata collected from Brazil, the Mozambique Channel, Madagascar, and Reunion Island. We found that nucleotide divergence ranged 0.02–3.10% among haplotypes. Moreover, while a Bayesian phylogenetic tree indicated that there were two substantially divergent genetic lineages, there was no evidence to support that T. carinata is comprised of a species complex due to isolation-by-distance. Surprisingly, both lineages were found in sympatry in both the Atlantic and Indian Oceans. Likewise, a 95% parsimony haplotype network revealed that identical haplotypes are found in both oceans, suggesting that a species complex may indeed exist, just not one caused by geographic isolation. We discuss possible explanations for this unexpected genetic structure, such as natural dispersal or human-mediated movement, and how the genetic structure found here is relevant to other marine organisms and to cryptic speciation.