Ancestral morphology of crown-group molluscs revealed by a new Ordovician stem aculiferan.

Exceptionally preserved fossils provide crucial insights into extinct body plans and organismal evolution. Molluscs, one of the most disparate animal phyla, radiated rapidly during the early Cambrian period (approximately 535-520 million years ago (Ma)). The problematic fossil taxa Halkieria and Orthrozanclus (grouped in Sachitida) have been assigned variously to stem-group annelids, brachiopods, stem-group molluscs or stem-group aculiferans (Polyplacophora and Aplacophora), but their affinities have remained controversial owing to a lack of preserved diagnostic characters. Here we describe a new early sachitid, Calvapilosa kroegeri gen. et sp. nov. from the Fezouata biota of Morocco (Early Ordovician epoch, around 478 Ma). The new taxon is characterized by the presence of a single large anterior shell plate and polystichous radula bearing a median tooth and several lateral and uncinal teeth in more than 125 rows. Its flattened body is covered by hollow spinose sclerites, and a smooth, ventral girdle flanks an extensive mantle cavity. Phylogenetic analyses resolve C. kroegeri as a stem-group aculiferan together with other single-plated forms such as Maikhanella (Siphogonuchites) and Orthrozanclus; Halkieria is recovered closer to the aculiferan crown. These genera document the stepwise evolution of the aculiferan body plan from forms with a single, almost conchiferan-like shell through two-plated taxa such as Halkieria, to the eight-plated crown-group aculiferans. C. kroegeri therefore provides key evidence concerning the long debate about the crown molluscan affinities of sachitids. This new discovery strongly suggests that the possession of only a single calcareous shell plate and the presence of unmineralised sclerites are plesiomorphic (an ancestral trait) for the molluscan crown.

MolluscDB: an integrated functional and evolutionary genomics database for the hyper-diverse animal phylum Mollusca.

Mollusca represents the second largest animal phylum but remains poorly explored from a genomic perspective. While the recent increase in genomic resources holds great promise for a deep understanding of molluscan biology and evolution, access and utilization of these resources still pose a challenge. Here, we present the first comprehensive molluscan genomics database, MolluscDB (http://mgbase.qnlm.ac), which compiles and integrates current molluscan genomic/transcriptomic resources and provides convenient tools for multi-level integrative and comparative genomic analyses. MolluscDB enables a systematic view of genomic information from various aspects, such as genome assembly statistics, genome phylogenies, fossil records, gene information, expression profiles, gene families, transcription factors, transposable elements and mitogenome organization information. Moreover, MolluscDB offers valuable customized datasets or resources, such as gene coexpression networks across various developmental stages and adult tissues/organs, core gene repertoires inferred for major molluscan lineages, and macrosynteny analysis for chromosomal evolution. MolluscDB presents an integrative and comprehensive genomics platform that will allow the molluscan community to cope with ever-growing genomic resources and will expedite new scientific discoveries for understanding molluscan biology and evolution.

Biomineralization by particle attachment in early animals.

Crystallization by particle attachment (CPA) of amorphous precursors has been demonstrated in modern biomineralized skeletons across a broad phylogenetic range of animals. Precisely the same precursors, hydrated (ACC-H2O) and anhydrous calcium carbonate (ACC), have been observed spectromicroscopically in echinoderms, mollusks, and cnidarians, phyla drawn from the 3 major clades of eumetazoans. Scanning electron microscopy (SEM) here also shows evidence of CPA in tunicate chordates. This is surprising, as species in these clades have no common ancestor that formed a mineralized skeleton and appear to have evolved carbonate biomineralization independently millions of years after their late Neoproterozoic divergence. Here we correlate the occurrence of CPA from ACC precursor particles with nanoparticulate fabric and then use the latter to investigate the antiquity of the former. SEM images of early biominerals from Ediacaran and Cambrian shelly fossils show that these early calcifiers used attachment of ACC particles to form their biominerals. The convergent evolution of biomineral CPA may have been dictated by the same thermodynamics and kinetics as we observe today.

Marine diversity patterns in Australia are filtered through biogeography.

Latitudinal diversity gradients are among the most striking patterns in nature. Despite a large body of work investigating both geographic and environmental drivers, biogeographical provinces have not been included in statistical models of diversity patterns. Instead, spatial studies tend to focus on species-area and local-regional relationships. Here, we investigate correlates of a latitudinal diversity pattern in Australian coastal molluscs. We use an online database of greater than 300 000 specimens and quantify diversity using four methods to account for sampling variation. Additionally, we present a biogeographic scheme using factor analysis that allows for both gradients and sharp boundaries between clusters. The factors are defined on the basis of species composition and are independent of diversity. Regardless of the measure used, diversity is not directly explained by combinations of abiotic variables. Instead, transitions between regions better explain the observed patterns. Biogeographic gradients can in turn be explained by environmental variables, suggesting that environmental controls on diversity may be indirect. Faunas within provinces are homogeneous regardless of environmental variability. Thus, transitions between provinces explain most of the variation in diversity because small-scale factors are dampened. This explanation contrasts with the species-energy hypothesis. Future work should more carefully consider biogeographic gradients when investigating diversity patterns.

A mitogenomic phylogeny of chitons (Mollusca: Polyplacophora).

BACKGROUND: Polyplacophora, or chitons, have long fascinated malacologists for their distinct and rather conserved morphology and lifestyle compared to other mollusk classes. However, key aspects of their phylogeny and evolution remain unclear due to the few morphological, molecular, or combined phylogenetic analyses, particularly those addressing the relationships among the major chiton lineages. RESULTS: Here, we present a mitogenomic phylogeny of chitons based on 13 newly sequenced mitochondrial genomes along with eight available ones and RNAseq-derived mitochondrial sequences from four additional species. Reconstructed phylogenies largely agreed with the latest advances in chiton systematics and integrative taxonomy but we identified some conflicts that call for taxonomic revisions. Despite an overall conserved gene order in chiton mitogenomes, we described three new rearrangements that might have taxonomic utility and reconstructed the most likely scenario of gene order change in this group. Our phylogeny was time-calibrated using various fossils and relaxed molecular clocks, and the robustness of these analyses was assessed with several sensitivity analyses. The inferred ages largely agreed with previous molecular clock estimates and the fossil record, but we also noted that the ambiguities inherent to the chiton fossil record might confound molecular clock analyses. CONCLUSIONS: In light of the reconstructed time-calibrated framework, we discuss the evolution of key morphological features and call for a continued effort towards clarifying the phylogeny and evolution of chitons.

Early shell field morphogenesis of a patellogastropod mollusk predominantly relies on cell movement and F-actin dynamics.

BACKGROUND: The morphogenesis of the shell field is an essential step of molluscan shell formation, which exhibits both conserved features and interlineage variations. As one major gastropod lineage, the patellogastropods show different characters in its shell field morphogenesis compared to other gastropods (e.g., the pulmonate gastropod Lymnaea stagnalis), likely related to its epibolic gastrulation. The investigation on the shell field morphogenesis of patellogastropods would be useful to reveal the lineage-specific characters in the process and explore the deep conservation among different molluscan lineages. RESULTS: We investigated the early shell field morphogenesis in the patellogastropod Lottia goshimai using multiple techniques. Electron microscopy revealed distinct morphological characters for the central and peripheral cells of the characteristic rosette-like shell field. Gene expression analysis and F-actin staining suggested that the shell field morphogenesis in this species predominantly relied on cell movement and F-actin dynamics, while BrdU assay revealed that cell proliferation contributed little to the process. We found constant contacts between ectodermal and meso/endodermal tissues during the early stages of shell field morphogenesis, which did not support the induction of shell field by endodermal tissues in general, but a potential stage-specific induction was indicated. CONCLUSIONS: Our results emphasize the roles of cell movement and F-actin dynamics during the morphogenesis of the shell field in Lo. goshimai, and suggest potential regulators such as diffusible factors and F-actin modulators. These findings reflect the differences in shell field morphogenesis of different gastropods, and add to the knowledge of molluscan larval shell formation.

Less effective selection leads to larger genomes.

The evolutionary origin of the striking genome size variations found in eukaryotes remains enigmatic. The effective size of populations, by controlling selection efficacy, is expected to be a key parameter underlying genome size evolution. However, this hypothesis has proved difficult to investigate using empirical data sets. Here, we tested this hypothesis using 22 de novo transcriptomes and low-coverage genomes of asellid isopods, which represent 11 independent habitat shifts from surface water to resource-poor groundwater. We show that these habitat shifts are associated with higher transcriptome-wide [Formula: see text] After ruling out the role of positive selection and pseudogenization, we show that these transcriptome-wide [Formula: see text] increases are the consequence of a reduction in selection efficacy imposed by the smaller effective population size of subterranean species. This reduction is paralleled by an important increase in genome size (25% increase on average), an increase also confirmed in subterranean decapods and mollusks. We also control for an adaptive impact of genome size on life history traits but find no correlation between body size, or growth rate, and genome size. We show instead that the independent increases in genome size measured in subterranean isopods are the direct consequence of increasing invasion rates by repeat elements, which are less efficiently purged out by purifying selection. Contrary to selection efficacy, polymorphism is not correlated to genome size. We propose that recent demographic fluctuations and the difficulty of observing polymorphism variation in polymorphism-poor species can obfuscate the link between effective population size and genome size when polymorphism data are used alone.

Data, time and money: evaluating the best compromise for inferring molecular phylogenies of non-model animal taxa.

For over a decade now, High Throughput sequencing (HTS) approaches have revolutionized phylogenetics, both in terms of data production and methodology. While transcriptomes and (reduced) genomes are increasingly used, generating and analyzing HTS datasets remain expensive, time consuming and complex for most non-model taxa. Indeed, a literature survey revealed that 74% of the molecular phylogenetics trees published in 2018 are based on data obtained through Sanger sequencing. In this context, our goal was to identify the strategy that would represent the best compromise among costs, time and robustness of the resulting tree. We sequenced and assembled 32 transcriptomes of the marine mollusk family Turridae, considered as a typical non-model animal taxon. From these data, we extracted the loci most commonly used in gastropod phylogenies (cox1, 12S, 16S, 28S, h3 and 18S), full mitogenomes, and a reduced nuclear transcriptome representation. With each dataset, we reconstructed phylogenies and compared their robustness and accuracy. We discuss the impact of missing data and the use of statistical tests, tree metrics, and supertree and supermatrix methods to further improve phylogenetic data acquisition pipelines. We evaluated the overall costs (time and money) in order to identify the best compromise for phylogenetic data sampling in non-model animal taxa. Although sequencing full mitogenomes seems to constitute the best compromise both in terms of costs and node support, they are known to induce biases in phylogenetic reconstructions. Rather, we recommend to systematically include loci commonly used for phylogenetics and taxonomy (i.e. DNA barcodes, rRNA genes, full mitogenomes, etc.) among the other loci when designing baits for capture.

Osmoionic homeostasis in bivalve mollusks from different osmotic niches: Physiological patterns and evolutionary perspectives.

Physiological knowledge gained from questions focused on the challenges faced and strategies recruited by organisms in their habitats assumes fundamental importance about understanding the ability to survive when subjected to unfavorable situations. In the aquatic environment, salinity is particularly recognized as one of the main abiotic factors that affects the physiology of organisms. Although the physiological patterns and challenges imposed by each occupied environment are distinct, they tend to converge to osmotic oscillations. From a comparative perspective, we aimed to characterize the osmoregulatory patterns of the bivalve mollusks Corbicula largillierti (purple Asian cockle), Erodona mactroides (lagoon cockle), and Amarilladesma mactroides (white clam) - inhabitants of different osmotic niches - when submitted to hypo- and/or hyperosmotic salinity variations. We determined the hemolymph osmotic and ionic concentrations, tissue hydration, and the intracellular isosmotic regulation (IIR) from the use of osmolytes (organic and inorganic) after exposure to species-specific salinity intervals. Additionally, we incorporated phylogenetic perspectives to infer and even broaden the understanding about the patterns that comprise the osmoionic physiology of Bivalvia representatives. According to the variables analyzed in the hemolymph, the three species presented a pattern of osmoconformation. Furthermore, both ionic regulation and conformation patterns were observed in freshwater, estuarine, and marine species. The patterns verified experimentally show greater use of inorganic osmolytes compared to the participation of organic molecules, which varied according to the osmotic niche occupied in the IIR for the mantle, adductor muscle, and gills. This finding widens the classic vision about the preferential use of certain osmolytes by animals from distinct niches. Our phylogenetic perspective also indicates that environmental salinity drives physiological trait variations, including hemolymph osmolality and the ion composition of the extracellular fluid (sodium, chloride, magnesium, and calcium). We also highlight the important role played by the shared ancestry, which influences the interspecific variability of the hemolymph K+ in selected representatives of Bivalvia.

Massive expansion and diversity of nicotinic acetylcholine receptors in lophotrochozoans.

BACKGROUND: Nicotinic acetylcholine receptors (nAChRs) are among the oldest and most conserved transmembrane receptors involved in signal transduction. Despite the prevalence and significance of cholinergic signaling, the diversity and evolution of nAChRs are not fully understood. RESULT: By comparative genomic analysis, we found massive expansions of nAChR genes in molluscs and some other lophotrochozoans. The expansion is particularly pronounced in stationary bivalve molluscs with simple nervous systems, with the number of nAChR genes ranging from 99 to 217 in five bivalves, compared with 10 to 29 in five ecdysozoans and vertebrates. The expanded molluscan nAChR genes tend to be intronless and in tandem arrays due to retroposition followed by tandem duplication. Phylogenetic analysis revealed diverse nAChR families in the common ancestor of bilaterians, which subsequently experienced lineage-specific expansions or contractions. The expanded molluscan nAChR genes are highly diverse in sequence, domain structure, temporal and spatial expression profiles, implying diversified functions. Some molluscan nAChR genes are expressed in early development before the development of the nervous system, while others are involved in immune and stress responses. CONCLUSION: The massive expansion and diversification of nAChR genes in bivalve molluscs may be a compensation for reduced nervous systems as part of adaptation to stationary life under dynamic environments, while in vertebrates a subset of specialized nAChRs are retained to work with advanced nervous systems. The unprecedented diversity identified in molluscs broadens our view on the evolution and function of nAChRs that are critical to animal physiology and human health.

Putting keyhole limpets on the map: phylogeny and biogeography of the globally distributed marine family Fissurellidae (Vetigastropoda, Mollusca).

Fissurellidae are marine gastropods with a worldwide distribution and a rich fossil record. We integrate molecular, geographical and fossil data to reconstruct the fissurellid phylogeny, estimate divergence times and investigate historical routes of oceanic dispersal. With five molecular markers for 143 terminals representing 27 genera, we resolve deep nodes and find that many genera (e.g., Emarginula, Diodora, Fissurella) are not monophyletic and need systematic revision. Several genera classified as Emarginulinae are recovered in Zeidorinae. Future work should prioritize emarginuline genera to improve understanding of ancestral traits and the early evolution of fissurellids. Tree calibration with the fossilized birth-death model indicates that crown fissurellids originated around 175 Ma, and generally resulted in younger ages for the earliest nodes than the node dating approach. Model-based biogeographic reconstruction, supported by fossils, infers an Indo-West Pacific origin, with a westward colonization of new oceans via the Tethys Seaway upon the breakup of Pangea. Western Atlantic clades then served as source for dispersal towards other parts of the globe. As the sister group to all other fissurellids, Rimula is ranked in its own subfamily, Rimulinae stat. nov. New synonyms: Hemitominae syn. nov. of Zeidorinae stat. nov.; Cranopsissyn. nov. of Puncturella; Variegemarginulasyn. nov. of Montfortula.

An Antarctic flock under the Thorson's rule: Diversity and larval development of Antarctic Velutinidae (Mollusca: Gastropoda).

In most marine gastropods, the duration of the larval phase is a key feature, strongly influencing species distribution and persistence. Antarctic lineages, in agreement with Thorson's rule, generally show a short pelagic developmental phase (or lack it completely), with very few exceptions. Among them is the ascidian-feeding gastropod family Velutinidae, a quite understudied group. Based on a multilocus (COI, 16S, 28S and ITS2) dataset for 182 specimens collected in Antarctica and other regions worldwide, we investigated the actual Antarctic velutinid diversity, inferred their larval development, tested species genetic connectivity and produced a first phylogenetic framework of the family. We identified 15 Antarctic Molecular Operational Taxonomic Units (MOTUs), some of which represented undescribed species, which show two different types of larval shell, indicating different duration of the Pelagic Larval Phase (PLD). Antarctic velutinids stand as an independent lineage, sister to the rest of the family, with extensive hidden diversity likely produced by rapid radiation. Our phylogenetic framework indicates that this Antarctic flock underwent repeated events of pelagic phase shortening, in agreement with Thorson's rule, yielding species with restricted geographic ranges.

Molecular phylogeny of Caudofoveata (Mollusca) challenges traditional views.

The shell-less, worm-shaped Caudofoveata (=Chaetodermomorpha) is one of the least known groups of molluscs. The taxon consists of 141 recognized species found from intertidal environments to the deep-sea where they live burrowing in sediment. Evolutionary relationships of the group have been debated, but few studies based on morphological or molecular data have investigated the phylogeny of the group. Here we use molecular phylogenetics to resolve relationships among and within families of Caudofoveata. Phylogenetic analyses were performed using selected mitochondrial and nuclear genes from species from all recognized families of Caudofoveata. In resulting trees and contrary to traditional views, Prochaetodermatidae forms the sister clade to a clade containing the other two currently recognized families, Chaetodermatidae and Limifossoridae. The monophyly of Prochaetodermatidae is highly supported, but Limifossoridae and Chaetodermatidae are not recovered as monophyletic. Most of the caudofoveate genera are also not recovered as monophyletic in our analyses. Thus results from our molecular data suggest that the current classification of Caudofoveata is in need of revision, and indicate evolutionary scenarios that differ from previously proposed hypotheses based on morphology.

DNA metabarcoding from sample fixative as a quick and voucher-preserving biodiversity assessment method 1.

Metabarcoding is a powerful, increasingly popular tool for biodiversity assessment, but it still suffers from some drawbacks (specimen destruction, separation, and size sorting). In the present study, we tested a non-destructive protocol that excludes any sample sorting, where the ethanol used for sample preserving is filtered and DNA is extracted from the filter for subsequent DNA metabarcoding. When tested on macroinvertebrate mock communities, the method was widely successful but was unable to reliably detect mollusc taxa. Three different protocols (no treatment, shaking, and freezing) were successfully applied to increase DNA release to the fixative. The protocols resulted in similar success in taxa detection (6.8-7 taxa) but differences in read numbers assigned to taxa of interest (33.8%-93.7%). In comparison to conventional bulk sample metabarcoding of environmental samples, taxa with pronounced exoskeleton and small-bodied taxa were especially underrepresented in ethanol samples. For EPT (Ephemeroptera, Plecoptera, Trichoptera) taxa, which are important for determining stream ecological status, the methods detected 46 OTUs in common, with only 4 unique to the ethanol samples and 10 to the bulk samples. These results indicate that fixative-based metabarcoding is a non-destructive, time-saving alternative for biodiversity assessments focussing on taxa used for ecological status determination. However, for a comprehensive assessment on total invertebrate biodiversity, the method may not be sufficient, and conventional bulk sample metabarcoding should be applied.

Mollusc genomes reveal variability in patterns of LTR-retrotransposons dynamics.

BACKGROUND: The three superfamilies of Long Terminal Repeat (LTR) retrotransposons are a widespread kind of transposable element and a major factor in eukaryotic genome evolution. In metazoans, recent studies suggested that Copia LTR-retrotransposons display specific dynamic compared to the more abundant and diverse Gypsy elements. Indeed, Copia elements show a relative scarcity and the prevalence of only a few clades in specific hosts. Thus, BEL/Pao seems to be the second most abundant superfamily. However, the generality of these assumptions remains to be assessed. Therefore, we carried out the first large-scale comparative genomic analysis of LTR-retrotransposons in molluscs. The aim of this study was to analyse the diversity, copy numbers, genomic proportions and distribution of LTR-retrotransposons in a large host phylum. RESULTS: We compare nine genomes of molluscs and further added LTR-retrotransposons sequences detected in databases for 47 additional species. We identified 1709 families, which enabled us to define 31 clades. We show that clade richness was highly dependent on the considered superfamily. We found only three Copia clades, including GalEa and Hydra which appear to be widely distributed and highly dominant as they account for 96% of the characterised Copia elements. Among the seven BEL/Pao clades identified, Sparrow and Surcouf are characterised for the first time. We find no BEL or Pao elements, but the rare clades Dan and Flow are present in molluscs. Finally, we characterised 21 Gypsy clades, only five of which had been previously described, the C-clade being the most abundant one. Even if they are found in the same number of host species, Copia elements are clearly less abundant than BEL/Pao elements in copy number or genomic proportions, while Gypsy elements are always the most abundant ones whatever the parameter considered. CONCLUSIONS: Our analysis confirms the contrasting dynamics of Copia and Gypsy elements in metazoans and indicates that BEL/Pao represents the second most abundant superfamily, probably reflecting an intermediate dynamic. Altogether, the data obtained in several taxa highly suggest that these patterns can be generalised for most metazoans. Finally, we highlight the importance of using database information in complement of genome analyses when analyzing transposable element diversity.

Mitogenomics reveals phylogenetic relationships of caudofoveate aplacophoran molluscs.

The worm-shaped, shell-less aplacophoran molluscs Caudofoveata and Solenogastres have recently received attention as part of the clade Aculifera, but relationships within these two lineages are still largely unknown. Here, we use complete mitochondrial genomes to shed light on higher-level relationships within Caudofoveata. Mitochondrial genomes have been sequenced for many diverse molluscs, but only two mitochondrial genomes from aplacophoran molluscs (the caudofoveates Scutopus ventrolineatus and Chaetoderma nitidulum) are available to date. We sequenced and assembled complete or near complete mitochondrial genomes of five additional species of Caudofoveata (Falcidens acutargatus, Falcidens halanychi, Scutopus robustus, Psilodens balduri and Spathoderma clenchi) and one species of Solenogastres (Neomenia carinata) for comparison to available mitochondrial genomes of aculiferans. Comparison of mitochondrial gene order among different lineages revealed a highly conserved order of protein coding genes corresponding to the hypothesized ancestral gene order for Mollusca. Unique arrangements of tRNAs were found among lineages of aculiferan molluscs as well as among caudofoveate taxa. Phylogenetic analyses of amino acid sequences for the 13 protein-coding genes recovered a monophyletic Aplacophora. Within Caudofoveata, Chaetodermatidae, but not Limifossoridae, was recovered monophyletic. Taken together, our results suggest that mitochondrial genomes can serve as useful molecular markers for aculiferan phylogenetics, especially for more recent phylogenetic events.

Multiple reversals of strand asymmetry in molluscs mitochondrial genomes, and consequences for phylogenetic inferences.

Strand asymmetry in nucleotide composition is a remarkable feature of animal mitochondrial genomes. The strand-specific bias in the nucleotide composition of the mtDNA has been known to be highly problematic for phylogenetic analyses. Here, the strand asymmetry was compared across 140 mollusc species and analyzed for a mtDNA fragment including twelve protein-coding genes. The analyses show that almost all species in Gastropoda (except Heterobranchia) and all species in Bivalvia present reversals of strand bias. The skew values on individual genes for all codon positions (P123), third codon positions (P3), and fourfold redundant third codon positions (P4FD) indicated that CG skews are the best indicators of strand asymmetry. The differences in the patterns of strand asymmetry significantly influenced the amino acid composition of the encoded proteins. These biases are most striking for the amino acids Valine, Cysteine, Asparagine and Threonines, which appear to have evolved asymmetrical exchanges in response to shifts in nucleotide composition. Molluscs with strong variability of genome architectures (ARs) are usually characterized by a reversal of the usual strand bias. Phylogenetic analyses show that reversals of asymmetric mutational constraints have consequences on the phylogenetic inferences, as taxa characterized by reverse strand bias (Heterobranchia and Bivalvia) tend to group together due to long-branch attraction (LBA) artifacts. Neutral Transitions Excluded (NTE) model did not overcome the problem of heterogeneous biases present in molluscs mt genomes, suggested it may not be appropriate for molluscs mt genome data. Further refinement phylogenetic models may help us better understand internal relationships among these diverse organisms.

A Silurian armoured aplacophoran and implications for molluscan phylogeny.

The Mollusca is one of the most diverse, important and well-studied invertebrate phyla; however, relationships among major molluscan taxa have long been a subject of controversy. In particular, the position of the shell-less vermiform Aplacophora and its relationship to the better-known Polyplacophora (chitons) have been problematic: Aplacophora has been treated as a paraphyletic or monophyletic group at the base of the Mollusca, proximate to other derived clades such as Cephalopoda, or as sister group to the Polyplacophora, forming the clade Aculifera. Resolution of this debate is required to allow the evolutionary origins of Mollusca to be reconstructed with confidence. Recent fossil finds support the Aculifera hypothesis, demonstrating that the Palaeozoic-era palaeoloricate 'chitons' included taxa combining certain polyplacophoran and aplacophoran characteristics. However, fossils combining an unambiguously aplacophoran-like body with chiton-like valves have remained elusive. Here we describe such a fossil, Kulindroplax perissokomos gen. et sp. nov., from the Herefordshire Lagerstätte (about 425 million years bp), a Silurian deposit preserving a marine biota in unusual three-dimensional detail. The specimen is reconstructed three-dimensionally through physical-optical tomography. Phylogenetic analysis indicates that this and many other palaeoloricate chitons are crown-group aplacophorans.

Ammonite habitat revealed via isotopic composition and comparisons with co-occurring benthic and planktonic organisms.

Ammonites are among the best-known fossils of the Phanerozoic, yet their habitat is poorly understood. Three common ammonite families (Baculitidae, Scaphitidae, and Sphenodiscidae) co-occur with well-preserved planktonic and benthic organisms at the type locality of the upper Maastrichtian Owl Creek Formation, offering an excellent opportunity to constrain their depth habitats through isotopic comparisons among taxa. Based on sedimentary evidence and the micro- and macrofauna at this site, we infer that the 9-m-thick sequence was deposited at a paleodepth of 70-150 m. Taxa present throughout the sequence include a diverse assemblage of ammonites, bivalves, and gastropods, abundant benthic foraminifera, and rare planktonic foraminifera. No stratigraphic trends are observed in the isotopic data of any taxon, and thus all of the data from each taxon are considered as replicates. Oxygen isotope-based temperature estimates from the baculites and scaphites overlap with those of the benthos and are distinct from those of the plankton. In contrast, sphenodiscid temperature estimates span a range that includes estimates of the planktonic foraminifera and of the warmer half of the benthic values. These results suggest baculites and scaphites lived close to the seafloor, whereas sphenodiscids sometimes inhabited the upper water column and/or lived closer to shore. In fact, the rarity and poorer preservation of the sphenodiscids relative to the baculites and scaphites suggests that the sphenodiscid shells may have only reached the Owl Creek locality by drifting seaward after death.

Advances in DNA Barcoding of Toxic Marine Organisms.

There are more than 200,000 marine species worldwide. These include many important economic species, such as large yellow croaker, ribbonfish, tuna, and salmon, but also many potentially toxic species, such as blue-green algae, diatoms, cnidarians, ctenophores, Nassarius spp., and pufferfish. However, some edible and toxic species may look similar, and the correct identification of marine species is thus a major issue. The failure of traditional classification methods in certain species has promoted the use of DNA barcoding, which uses short, standard DNA fragments to assist with species identification. In this review, we summarize recent advances in DNA barcoding of toxic marine species such as jellyfish and pufferfish, using genes including cytochrome oxidase I gene (COI), cytochrome b gene (cytb), 16S rDNA, internal transcribed spacer (ITS), and Ribulose-1,5-bisphosphate carboxylase oxygenase gene (rbcL). We also discuss the application of this technique for improving the identification of marine species. The use of DNA barcoding can benefit the studies of biological diversity, biogeography, food safety, and the detection of both invasive and new species. However, the technique has limitations, particularly for the analysis of complex objects and the selection of standard DNA barcodes. The development of high-throughput methods may offer solutions to some of these issues.