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Unravelling the evolutionary history of taxa requires solid delimitation of the traits characterising these. This can be challenging especially in groups with a highly complex taxonomy. The squat lobster family Munididae contains more than 450 species distributed among 21 genera, Munida being the most speciose (~300 species). Previous phylogenetic studies, based on a small part of the diversity of the group, have suggested polyphyletic origins for Munida and the paraphyly of Munididae. Here, we use an integrative approach based on multi-locus phylogenies (two mitochondrial and three nuclear markers) paired with 120 morphological characters, to resolve taxonomic and evolutionary relationships within Munididae. Our study covers ~60% of the family’s known diversity (over 800 specimens of 291 species belonging to 19 of the 21 genera collected from the Atlantic, Indian and Pacific oceans). Using this information, we confirm the validity of most genera, proposing new ones in cases where the genetic analyses are compatible with morphological characters. Four well-defined munidid clades were recovered, suggesting that new genera should be erected in the currently recognised Munididae (three for the genus Agononida and eleven in Munida), and the genus Grimothea is resurrected. A key to all genera of the family is presented. Molecular clock estimates and ancestral biogeographic area reconstructions complement the taxonomic profiles and suggest some explosive diversification within Munididae during the Cretaceous and the Palaeogene. Further anagenetic events and narrow sympatry accounting for changes in distribution indicate a more limited dispersal capacity than previously considered. Our study unravels how diversification may occur in deep waters and further highlights the importance of the integrative approach in accurately delineating species in understanding the history of a family and the factors driving the evolution. ZooBank LSID: urn:lsid:zoobank.org:pub:16A61C4A-8D96-4372-820F-8EBDF179B43C

This study reports on new squat lobsters of the genus Munida collected during recent surveys of Australia’s continental margins. We report on 33 species of Munida including seven new species and 14 new range extensions for Australia. More than 500 specimens were collected, mostly from the western continental margin of Australia, but also including a new species from deep water (>2000 m) off Tasmania. We provide new data on the colour patterns of some species and include molecular data from two mitochondrial markers (16S rRNA and COI) to support the taxonomic status of the new species.

Marine invertebrates harbour a complex suite of bacterial and archaeal symbionts, a subset of which are probably linked to host health and homeostasis. Within a complex microbiome it can be difficult to tease apart beneficial or parasitic symbionts from nonessential commensal or transient microorganisms; however, one approach is to detect strong cophylogenetic patterns between microbial lineages and their respective hosts. We employed the Procrustean approach to cophylogeny (PACo) on 16S rRNA gene derived microbial community profiles paired with COI, 18S rRNA and ITS1 host phylogenies. Second, we undertook a network analysis to identify groups of microbes that were co-occurring within our host species. Across 12 coral, 10 octocoral and five sponge species, each host group and their core microbiota (50% prevalence within host species replicates) had a significant fit to the cophylogenetic model. Independent assessment of each microbial genus and family found that bacteria and archaea affiliated to Endozoicomonadaceae, Spirochaetaceae and Nitrosopumilaceae have the strongest cophylogenetic signals. Further, local Moran's I measure of spatial autocorrelation identified 14 ASVs, including Endozoicomonadaceae and Spirochaetaceae, whose distributions were significantly clustered by host phylogeny. Four co-occurring subnetworks were identified, each of which was dominant in a different host group. Endozoicomonadaceae and Spirochaetaceae ASVs were abundant among the subnetworks, particularly one subnetwork that was exclusively comprised of these two bacterial families and dominated the octocoral microbiota. Our results disentangle key microbial interactions that occur within complex microbiomes and reveal long-standing, essential microbial symbioses in coral reef invertebrates.

Aim To develop a comparative phylogeographic framework to understand the origins, evolution, taxonomic richness, and distribution of Australian demersal fish endemics in the context of the Indo‐West Pacific (IWP). Location Southern Australia and the IWP. Taxon Platycephalidae (flathead fishes). Methods Nuclear and mitochondrial phylogenies of flathead fishes were inferred from 46 of 85 nominal species, and 14 additional cryptic species‐level lineages, representing 17 of the 18 genera. Molecular clocks and habitat trait reconstructions were used to infer the palaeoclimatic and geological events responsible for shaping the evolution and diversification of the group. Results The family Platycephalidae comprises two sister subfamilies; Platycephalinae and Onigociinae, which diverged in the Eocene into predominantly temperate and tropical assemblages respectively. The basal platycephalin taxa are confined to southern Australia with the most derived groups in the tropics, following a high‐ to low‐latitudinal evolutionary trajectory. In contrast, the onigociins are predominantly associated with the tropics, and have diversified across the region since the early Miocene with very few introductions into temperate Australia. Main conclusions Platycephalinae and Onigociinae show contrasting evolutionary scenarios. Platycephalins have a temperate to tropical evolutionary trajectory consistent with their arrival into the region via tectonic rafting and subsequent dispersal. This dispersal was likely facilitated by formation of shallow‐water environments along the Sunda Arc margin following collision of the Australian and Eurasian Plates. In contrast, the Onigociinae has likely maintained a tropical presence across the IWP since the Eocene and has experienced higher diversification rates leading to circa three times the species diversity found in the Platycephalinae. Rounds of dispersal and allopatric speciation have subsequently played out across both low and high latitudes with both subfamilies harbouring cryptic species‐level lineages. This work provides an explicit working hypothesis for exploring origins and diversification in other demersal fishes endemic to the Australian continent.

The mitochondrial genetic diversity, distribution and invasive potential of multiple cryptic operational taxonomic units (OTUs) of the red invasive seaweed Asparagopsis were assessed by studying introduced Mediterranean and Hawaiian populations. Invasive behavior of each Asparagopsis OTU was inferred from phylogeographic reconstructions, past historical demographic dynamics, recent range expansion assessments and future distributional predictions obtained from demographic models. Genealogical networks resolved Asparagopsis gametophytes and tetrasporophytes into four A. taxiformis and one A. armata cryptic OTUs. Falkenbergia isolates of A. taxiformis L3 were recovered for the first time in the western Mediterranean Sea and represent a new introduction for this area. Neutrality statistics supported past range expansion for A. taxiformis L1 and L2 in Hawaii. On the other hand, extreme geographic expansion and an increase in effective population size were found only for A. taxiformis L2 in the western Mediterranean Sea. Distribution models predicted shifts of the climatically suitable areas and population expansion for A. armata L1 and A. taxiformis L1 and L2. Our integrated study confirms a high invasive risk for A. taxiformis L1 and L2 in temperate and tropical areas. Despite the differences in predictions among modelling approaches, a number of regions were identified as zones with high invasion risk for A. taxiformis L2. Since range shifts are likely climate-driven phenomena, future invasive behavior cannot be excluded for the rest of the lineages.

The red algal genus Asparagopsis (Bonnemaisoniaceae) is a significant resource for bioactive natural products. However, prior to domestication for commercial production, we need to understand the potential variation in growth and concentration of natural products between isolates of Asparagopsis and, beyond that, how these traits are affected by environmental conditions. Ten isolates of Asparagopsis taxiformis were collected from tropical and warm-temperate regions in Queensland, Australia, and identified by molecular barcoding of the mitochondrial intergenic spacer (cox2–3 spacer). The isolates were cultured at three temperatures ranging from the minimum of the warm-temperate region to the maximum of the tropical region. Growth rates and the concentration of natural products varied between the region of origin, between isolates within region and between temperatures. Growth differed by up to 50% between isolates, whereas the concentration of natural products differed more than tenfold. Growth rates were highest at the minimum temperature of 20.2°C, irrespective of region of origin, and were lowest at the maximum temperature of 28.1°C. Natural products were threefold higher in tropical isolates, and this variation was not correlated to growth. Consequently, targeting isolates with high concentrations of natural products should be the primary strategy for the domestication of Asparagopsis for biotechnology applications.

Vibrios of the Harveyi clade are responsible for severe economic losses in the aquaculture industry worldwide, but so far no rapid method has been available to discriminate between the closely related species Vibrio campbellii, Vibrio harveyi, Vibrio owensii and Vibrio rotiferianus. Here, we describe a multiplex PCR assay capable of fast and reliable detection and identification of these species by amplification of three conserved protein-coding genes (topA, ftsZ and mreB). The developed species-specific primer sets produce PCR amplicons of unequal length and can be used separately or in combination. When combined with a 3-tube most-probable-number (MPN) based protocol, the multiplex PCR allows highly sensitive detection and enumeration in complex, aquaculture-relevant samples such as water, Artemia and spiny lobster (Panulirus ornatus) larvae. The described multiplex PCR represents a significant advance for aquaculture by providing an easy, fast and low-cost method for identification of several closely related pathogens that are difficult to discriminate by traditional approaches.

Understanding the factors that cause population divergence has long been ofinterest to marine biologists in their attempts to interpret the effect of human-mediated vectors. Broadcast-spawning species with limited dispersal capabilityare excellent candidates to measure the present-day patterns of genetic diversity.The tunicate Ciona intestinalis (Ascidiacea) is comprised of a complex ofmorphologically cryptic species that form vigorous aggregates in eutrophic habi-tats (harbors, gulfs and lagoons) where they can compete with the epibenthiccommunity and cause biofouling problems. This study investigated biogeograph-ic variability and migration patterns of C. intestinalis sp. A along NortheastAtlantic and Mediterranean coasts using microsatellite markers. Data presentedhere on 371 specimens collected from 17 populations reveal high genetic poly-morphism, but with a deficit of heterozygote deficiency. Absence of evidence forisolation by distance suggests that the genetic patterns do not reflect the geo-graphic distribution of sampled populations. Substantial gene flow and artificialpotential for dispersal boost high levels of within-population genetic variabilityand prevent genetic differentiation within and between seas. A predominant east-ward migration pattern was revealed by the data set, with very limited opportu-nity for C. intestinalis sp. A to travel westward. This directional movementindicates that other properties (e.g. habitat quality, genetic traits, mating system,life cycle) may cause adaptive divergence at a large biogeographic scale.IntroductionDispersal plays an important role in the biogeographicdistribution of marine organisms, influencing the coloni-zation of new habitats, the connectivity of populationsand ultimately species persistence (Bilton et al. 2001;Grantham et al. 2003). At sea, physical barriers are oftenabsent and water currents may accelerate larval dispersal,determining population homogeneity over large geo-graphic areas (Knutsen et al. 2003; Cowen & Sponaugle2009). In benthic invertebrates, large larval output,increased larval duration and long-distance dispersal mayallow high genetic connectivity thereby mitigating theeffect of genetic drift and inbreeding (Hedgecock 1986),and eventually leading to panmixia. Hydrologic (e.g. pat-terns of marine currents, existence of different watermasses) and ecological barriers (e.g. habitat discontinuity,larval behavior) may hinder long-distance movements(Palumbi 1994; Riginos & Nachman 2001; Bierne et al.2003). Once barriers have been established, internal (e.g.life strategy and larval survival) and external (e.g. envi-ronmental parameters) factors may favor, together with

Invasive species display remarkable levels of ecophysiological plasticity, which supports colonization, population establishment and fitness across their introduction range. The red seaweed genus Asparagopsis comprises genetically homogeneous invasive species (A. armata) and cryptic species complexes (A. taxiformis sensu lato) consisting of invasive mitochondrial lineages introduced worldwide. The photosynthetic plasticity of Australian, Mediterranean and Hawaiian Falkenbergia stages (i.e. the tetrasporophytic stage) of A. taxiformis lineages 2, 3 and 4 and Mediterranean isolates of A. armata was assessed by challenging their photosynthetic performance at five different temperatures (12–26 °C). Our aim is to portray the photosynthetic profiles in relation to temperature for each of the aforementioned Asparagopsis OTUs. We additionally test the physiological response of A. taxiformis lineage 2 sampled within its invasive (Mediterranean Sea) and native range (Australia) to identify physiological features associated with invasive strains. Based on photosynthesis optima, Asparagopsis isolates were recovered into a tropical (NL2 and L4) and a temperate (AA, Il2 and L3) group that presented no differences in most photosynthetic parameters at the experimental temperatures, thus indicating a greater physiological plasticity. On the other hand, low Ic values together with an apparent lack of sensitivity in the photosynthetic response to changing temperatures were revealed for the Mediterranean lineage 2, indicative of adaptive benefits that likely support its invasive success compared to the rest of the genus. Our results represent a valuable resource to predict distributional shifts in some of the lineages and to anticipate control programs for lineage 3, potentially invasive.

Sponge taxonomy can be challenging as many groups exhibit extreme morphological plasticity induced by local environmental conditions. Foliose keratose sponges of the sub-family Phyllospongiinae (Dictyoceratida, Thorectidae: Strepsichordaia, Phyllospongia and Carteriospongia) are commonly found in intertidal and subtidal habitats of the Indo-Pacific. Lacking spicules, these sponges can be difficult to differentiate due to the lack of reliable morphological characters for species delineation. We use molecular phylogenies inferred from the nuclear Internal Transcribed Spacer 2 region (ITS2) and morphometrics (19 characters; 52 character states) to identify evolutionarily significant units (ESUs; sensu Moritz) within foliose Phyllosponginiids collected from seven geographic locations across tropical eastern and Western Australia. The ITS2 topology was congruent with the tree derived from Bayesian inference of discrete morphological characters supporting expected taxonomic relationships at the genus level and the identification of five ESUs. However, phylogenies inferred from the ITS2 marker revealed multiple sequence clusters, some of which were characterised by distinct morphological features and specific geographic ranges. Our results are discussed in light of taxonomic incongruences within this study, hidden sponge diversity and the role of vicariant events in influencing present day distribution patterns.

Rapid identification of introduced seaweeds is crucial to support management and conservation decisions, especially when multiple cryptic lineages of high-profile invasive taxa occur sympatrically. The red seaweed genus Asparagopsis (Bonnemaisoniales, Rhodophyta) comprises two recognised morpho-species characterised by heteromorphic life cycles and presumably morphologically identical 'Falkenbergia' tetrasporophyte stages: A. armata and A. taxiformis. Populations of the former were easily identified by the presence of distinctive harpoon-like braches on the gametophyte thalli. Four morphologically cryptic yet genetically distinct mitochondrial lineages of invasive nature were recognised within A. taxiformis. We reported a morphological delineation of tetrasporophytes and gametophytes of Asparagopsis, including cryptic lineages collected from the Mediterranean Sea and the Hawaiian Islands, where multiple Asparagopsis lineages were present. Vegetative anatomical characters of the tetrasporophytes were useful in differentiating A. armata from those of A. taxiformis as well as among tetrasporophyte isolates belonging to the four A. taxiformis lineages. In addition, these characters distinguished lineage 2 native range specimens (Hawaii) from the invasive specimens (Mediterranean Sea), which suggested high levels of morphological plasticity in the invasive taxon. We propose that the taxonomic status of the lineages within A. taxiformis needs to be revised.

Molecular studies have been instrumental for refining species boundaries in the coral genus Pocillopora and revealing hidden species diversity within the extensively studied global species Pocillopora damicornis . Here we formally revise the taxonomic status of species closely related to and within the P. damicornis species complex, taking into account both genetic evidence and new data on morphometrics, including fine-scale corallite and coenosteum structure. We found that mitochondrial molecular phylogenies are congruent with groups based on gross-morphology, therefore reflecting species-level differentiation. However, high levels of gross morphological plasticity and shared morphological characteristics mask clear separation for some groups. Fine-scale morphological variation, particularly the shape and type of columella, was useful for differentiating between clades and provides an excellent signature of the evolutionary relationships among genetic lineages. As introgressive hybridization and incomplete lineage sorting complicate the delineation of species within the genus on the basis of a single species concept, the Unified Species Concept may represent a suitable approach in revising Pocillopora taxonomy. Eight species are herein described ( P. damicornis , P. acuta , P. aliciae , P. verrucosa , P. meandrina , P. eydouxi , P. cf. brevicornis ), including a novel taxon – P ocillopora bairdi sp. nov. (Schmidt-Roach, this study). Citation synonyms and type materials are presented.

Lack of morphological features of diagnostic value and high levels of environmental phenotypic plasticity obscure species boundaries for most taxa in the genus Pocillopora Lamarck, 1816 and complicate the definition of taxonomically distinct units. Species of the genus are colonial, generally ramose, rarely massive or encrusting and mostly hermatypic; corallite arrangement is plocoid, septa are generally poorly developed and usually arranged in two cycles; the columella is mostly poorly developed (Veron & Pichon 1976). Verrucae are common, although reduced in some species. Currently 17 species are formally acknowledged within the genus (Veron 2000).

Sponges are abundant, diverse and functionally important components of aquatic biotopes with crucial associations for many reef fish and invertebrates. Sponges have strict temperature optima, and mass mortality events have occurred after unusually high temperatures. To assess how sponges may adapt to thermal stress associated with a changing climate, we applied gene expression profiling to both stages of their bipartite life cycles. Adult Rhopaloeides odorabile are highly sensitive to thermal stress (32 °C), yet their larvae can withstand temperatures up to 36 °C. Here, we reveal the molecular mechanisms that underpin these contrasting thermal tolerances, which may provide sponges with a means to successfully disperse into cooler waters. Heat shock protein 70 was induced by increasing temperature in adult sponges, and genes involved in important biological functions including cytoskeleton rearrangement, signal transduction, protein synthesis/degradation, oxidative stress and detoxification were all negatively correlated with temperature. Conversely, gene expression in larvae was not significantly affected until 36 °C when a stress response involving extremely rapid activation of heat shock proteins occurred. This study provides the first transcriptomic assessment of thermal stress on both life history stages of a marine invertebrate facilitating better predictions of the long-term consequences of climate change for sponge population dynamics.

Six species of Paramunida are reported from the continental margin of north-western Australia. Three species are new to science: Paramunida christinae sp. nov., P. ioannis sp. nov., and P. spiniantennata sp. nov. Two species are reported for the first time from Australian waters, P. evexa Macpherson, 1996 and P. tricarinata (Alcock, 1894). These species were confirmed by molecular evidence from the mitochondrial markers ND1 and 16S. We also examine phylogenetic relationships within the genus, and provide an identification key for all known Paramunida species.

International audience; SOUL homologs constitute a heme-binding protein superfamily putatively involved in heme and tetrapyrrole metabolisms associated with a number of physiological processes. Despite their omnipresence across the tree of life and the biochemical characterization of many SOUL members, their functional role and the evolutionary events leading to such remarkable protein repertoire still remain cryptic. To explore SOUL evolution, we apply a computational phylogenetic approach, including a relevant number of SOUL homologs, to identify paralog forms and reconstruct their genealogy across the tree of life and within species. In animal lineages, multiple gene duplication or loss events and paralog functional specializations underlie SOUL evolution from the dawn of ancestral echinoderm and mollusc SOUL forms. In photosynthetic organisms, SOUL evolution is linked to the endosymbiosis events leading to plastid acquisition in eukaryotes. Derivative features, such as the F2L peptide and BH3 domain, evolved in vertebrates and provided innovative functionality to support immune response and apoptosis. The evolution of elements such as the N-terminal protein domain DUF2358, the His42 residue, or the tetrapyrrole heme-binding site is modern, and their functional implications still unresolved. This study represents the first in-depth analysis of SOUL protein evolution and provides novel insights in the understanding of their obscure physiological role.

The incredible range of morphological plasticity present in scleractinian corals has confused the taxonomy of the group, prompting the introduction of “ecomorphs” to explain the observed correlation between local environmental conditions and phenotypic variation. Pocillopora damicornis (Linnaeus, 1758) represents one of the best known examples of eco-phenotypic variation in scleractinian corals with a variety of forms and reproductive strategies reported across its global distribution range. Here, we reconstruct genealogical relationships of P. damicornis colonies collected from thirteen locations along the East Australian coast to examine the relationship between genetic and phenotypic diversity in this species. Haplotype networks computed from two mitochondrial DNA regions (CR, ORF) indicate that the range of morphotypes observed within this taxon fall into at least five genetically distinct mitochondrial lineages. Nuclear (HSP70, ITS2) haplowebs on the other hand recover sharp genetic discontinuities among three of the morphological groups. We conclude that P. damicornis from Eastern Australia constitutes a cryptic species complex. The misinterpretation of taxonomical units within P. damicornis may well explain its perceived variation in the ecology, biology and life history across its range.

Morphological delineation of sponge species is hindered by the narrow range of fixed diagnostic characters and our limited knowledge of how much phenotypic plasticity the sponge body plan assumes in response to environmental conditions. Here, we make use of the partial mitochondrial cytochrome c oxidase subunit I (COI) gene and the second internal transcribed spacer (ITS2) region to assess the taxonomic validity of colour morphotypes observed in the elephant ear sponge Ianthella basta (Pallas, 1776) across its distribution range in northern Australia, and explore levels and patterns of genetic diversity among populations of the species collected from both sides of the Torres Strait. Molecular phylogenetic analyses revealed congruent topologies consistent with three evolutionarily significant units (ESUs) that were independent of the morphology of the sponge. ESU I includes previously morphologically and genetically delineated western Pacific specimens of I. basta (Guam), and probably corresponds to the type specimen originally described from Indonesia. ESU I occurs in almost all sampling sites across northern Australia, suggesting considerable levels of connectivity among reefs throughout the Torres Strait. ESUs II and III are each exclusively associated with a geographic region of high sponge species richness separated by Torres Strait, and probably represent the result of historical population fragmentation.

Identification of taxonomical units underpins most biological endeavours ranging from accurate biodiversity estimates to the effective management of sustainably harvested, protected or endangered species. Successful species identification is now frequently based on a combination of approaches including morphometrics and DNA markers. Sequencing of the mitochondrial COI gene is an established methodology with an international campaign directed at barcoding all fishes. We employed COI sequencing alongside traditional taxonomic identification methods and uncovered instances of deep intraspecific genetic divergences among flathead species. Sixty-five operational taxonomic units (OTUs) were observed across the Indo-West Pacific from just 48 currently recognized species. The most comprehensively sampled taxon, Platycephalus indicus, exhibited the highest levels of genetic diversity with eight lineages separated by up to 16.37% genetic distance. Our results clearly indicate a thorough reappraisal of the current taxonomy of P. indicus (and its three junior synonyms) is warranted in conjunction with detailed taxonomic work on the other additional Platycephalidae OTUs detected by DNA barcoding.

The Munidopsidae, one of three squat lobster families in the Galatheoidea, contains the deepest dwelling squat lobsters, with some occurring at abyssal depths. Munidopsids were formerly divided into two subfamilies: Shinkaiinae, for the unusual hydrothermal vent genus Shinkaia; and Munidopsinae for remaining taxa. Four munidopsid genera are currently recognised (Shinkaia, Leiogalathea, Galacantha and Munidopsis) but the largest genus, Munidopsis, is highly diverse morphologically, with multiple genera or subgenera currently in its synonymy. Phylogenetic studies of galatheoids focussed on high level relationships indicate that Leiogalathea is sister to other munidopsids, but the position of Shinkaia with respect to Munidopsis and Galacantha is unclear, as is the reciprocal monophyly of the latter two genera. Phylogenetic analyses of the Munidopsidae based on mitochondrial 16S and COI sequences, sampling all current genera (including the majority of the formerly recognised subgenera), indicate that the generic and former subfamily classifications do not reflect the phylogeny. Shinkaia and Galacantha clades are nested within Munidopsis rendering the genus paraphyletic and the bi-subfamily classification phylogenetically invalid. Many of the Munidopsis clades recovered, however, correspond well to formerly recognised genera or subgenera, indicating good prospects for a natural subdivision of Munidopsis. Crown Copyright © 2011 Published by Elsevier GmbH. All rights reserved.

Natural populations of marine invertebrates often exhibit measureable morphologic variation resulting in taxonomic confusion. This potentially has severe consequences for experimental design and data management. Species of the sponge genus Ianthella embody a number of different morphologies and a diverse range of secondary metabolites. Among them, Ianthella basta (Pallas, 1776), a common sponge in Papua New Guinea and the Great Barrier Reef (GBR), exhibits two dominant colour morphotypes: yellow and purple. Specimens collected from Orpheus Island on the GBR were investigated using phylogenetic (CO1, ITS-2 sequence analysis), chemical (mass spectrometry) and microbial (DGGE and 16S rRNA clone library) techniques in an effort to fully characterise the two colour morphs. Phylogenetic analyses indicated sharp genetic discontinuities within I. basta sensu lato independent of colour variation. The two morphotypes did, however, correspond to distinct DGGE profiles largely due to the presence of additional bands in the purple morpho-group. Further comparison of the microbial communities by 16S rRNA gene sequencing revealed that whilst both colour morphs were dominated by only two bacterial symbionts (residing within the Gamma and Alphaproteobacteria), the purple morph also contained minor representatives of the Cyanobacteria, Chloroflexi and Verrucomicrobia. Untargeted metabolic profiling by Fourier transform ion cyclotron resonance mass spectrometry (FTICR-MS) indicated two distinct clusters corresponding to the different sponge colours. A clear association was found between the araplysillin class of compounds and the purple morphotype of I. basta, indicating the utility of a metabolomic approach to assess differences between colour morphs. These results have important implications for ecological investigations in sponges and other invertebrate taxa whose morphology is fundamentally dynamic, stressing the need for precise taxonomic, chemical and microbial descriptions.

We report an accurate multiplex reverse transcription quantitative polymerase chain reaction (RT–qPCR) assay, capable of reproducing gene expression profiles from 16 target genes [12 genes of interest (GOIs) and four reference genes (RGs)] in Acropora millepora, a common reef-building model coral species. The 12 GOIs have known or putative roles in the coral bleaching response, yet the method is not restricted to this particular assay and gene set. The procedure is based on the Beckman Coulter (Fullerton, CA, USA) GenomeLab™ GeXP Genetic Analysis System and bridges the gap between quantitative real-time PCR (qPCR) expression analysis of a single or a small number of genes and microarray gene expression surveys of thousands of genes. Despite large variation among biological replicates, the majority of GOIs were up-regulated (up to 4000%) in most colonies during a laboratory-based thermal stress experiment. Two genes, Nf-kβ2 and MnSod, were consistently up-regulated in all colonies tested, and we therefore propose these as candidate markers useful for population-level evaluations of thermal stress. Our assay provides an important new tool for coral bleaching studies; because of the lower cost, labour and amount of cDNA required compared with singleplex qPCR, population-level studies with large biological replication are feasible.

Nitric oxide (NO) is essential to many physiological functions and operates in several signaling pathways. It is not understood how and when the different isoforms of nitric oxide synthase (NOS), the enzyme responsible for NO production, evolved in metazoans. This study investigates the number and structure of metazoan NOS enzymes by genome data mining and direct cloning of Nos genes from the lamprey. In total, 181 NOS proteins are analyzed from 33 invertebrate and 63 vertebrate species. Comparisons among protein and gene structures, combined with phylogenetic and syntenic studies, provide novel insights into how NOS isoforms arose and diverged. Protein domains and gene organization--that is, intron positions and phases--of animal NOS are remarkably conserved across all lineages, even in fast-evolving species. Phylogenetic and syntenic analyses support the view that a proto-NOS isoform was recurrently duplicated in different lineages, acquiring new structural configurations through gains and losses of protein motifs. We propose that in vertebrates a first duplication took place after the agnathan-gnathostome split followed by a paralog loss. A second duplication occurred during early tetrapod evolution, giving rise to the three isoforms--I, II, and III--in current mammals. Overall, NOS family evolution was the result of multiple gene and genome duplication events together with changes in protein architecture.

The red alga, Asparagopsis taxiformis, has recently expanded its distribution range into the Western Mediterranean Sea, and populations have now even been found on the Portuguese South coast. All Western Mediterranean populations belong to a single mitochondrial cryptic lineage (referred to as lineage 2 in earlier studies) and probably result from a recent invasion of Indo-Pacific origin. Here we investigate fine-scale population genetic diversity and structure within and among Mediterranean populations of lineage 2 using eight microsatellite loci and compare the obtained patterns with those observed in a Californian population of the same lineage. To generate an appropriate analytical method suitable to the polyploid status of this species, we score microsatellite loci as in a dominant marker system. Thereafter, we produce robust descriptors inferred from frequencies of both microsatellite alleles and multilocus genotypes. Populations from California and the Mediterranean Sea differ considerably in their levels of genetic diversity. In the Mediterranean, populations reproduce predominantly sexually and exhibit high levels of genotypic variation, suggestive either of multiple introductions or of a single introduction by a genetically diverse and large group of individuals. Bayesian clustering revealed one or possibly two weakly supported panmictic subpopulations, indicative of extensive admixture. The expansion of this lineage is rapid, possibly due to the absence of eco-physiological barriers to gene flow throughout its invasive trajectory.

Molecular phylogeography has for decades been a frequently used approach to delineate novel evolutionarily significant units (ESUs) and to study the dynamics of invasive species. Next-generation sequencing technology (NGS) and the use of environmental DNA (eDNA) have the potential to revolutionize our way of understanding biodiversity and to establish rapid protocols for early-stage detection of invasive species. In seaweeds, however, several years of research on iconic invasive taxa of ambiguous taxonomic status (e.g. Caulerpa, Codium, Asparagopsis) have suggested that an integrative approach, namely the combination of multiple lines of evidence (e.g. phylogeographic, ecological, physiological and predictive modelling), is necessary to accurately resolve the taxonomy and their invasive potential. At present, integrative approaches in these fields are often weak because of incongruences among species delineation, newly discovered ESUs which remain undescribed taxonomically, and because databases containing vouchers of barcoded specimens are incomplete. As relocations of marine biota accelerate and climatic changes offer new potential niches for invasive seaweeds, new, transferable and internationally adopted protocols are necessary for exploring, monitoring and managing marine biodiversity. This is particularly urgent in areas of intense maritime traffic, such as the Mediterranean Sea and the Hawaiian archipelago, in order to achieve sustainable socio-economic development without compromising the local marine resources.

Eight polymorphic nuclear microsatellite loci were identified from the invasive Indo-Pacific Mediterranean strain of Asparagopsis taxiformis. Microsatellite markers were tested against a panel of specimens collected along the Italian (Elba, Naples) and Californian (Catalina Island) coasts, all belonging to the same mitochondrial lineage. In addition, we used Hawaiian specimens, belonging to a closely related mitochondrial lineage. The markers amplified in all of the specimens but failed consistently in thalli of two more distantly related mitochondrial lineages of A. taxiformis as well as in specimens belonging to the sister species Asparagopsis armata. Since haploid female individuals among the Mediterranean specimens contained cystocarps, genotyping was performed on supposedly haploid female specimens and supposedly diploid cystocarps separately. As expected, external allelic contribution was detected in the cystocarps. However, even after removal of these reproductive structures, gametophyte thalli exhibited patterns consisting of up to three alleles in all of the tested populations indicating polyploidy. An elevated number of distinct genotypes (up to 85%) were found per population, suggesting high intra-population variation. Results showed high genetic similarity between the two Mediterranean populations screened and lower similarity between these two and the Californian one within the same mitochondrial lineage. Lowest similarity was found between these three and the Hawaiian population belonging to the other related mitochondrial lineage 1.

The rhodophyte seaweed Asparagopsis armata Harvey is distributed in the northern and southern temperate zones, and its congener Asparagopsis taxiformis (Delile) Trevisan abounds throughout the tropics and subtropics. Here, we determine intraspecific phylogeographic patterns to compare potential causes of the disjunctions in the distributions of both species. We obtained specimens throughout their ranges and inferred phylogenies from the hypervariable domains D1-D3 of the nuclear rDNA LSU, the plastid spacer between the large and small subunits of RuBisCo and the mitochondrial cox 2-3 intergenic spacer. The cox spacer acquired base changes the fastest and the RuBisCo spacer the slowest. Median-joining networks inferred from the sequences revealed the absence of phylogeographic structure in the introduced range of A. armata, corroborating the species' reported recent introduction. A. taxiformis consisted of three nuclear, three plastid and four mitochondrial genetically distinct, lineages (1-4). Mitochondrial lineage 3 is found in the western Atlantic, the Canary Islands and the eastern Mediterranean. Mitochondrial lineages 1, 2, and 4 occur in the Indo-Pacific, but one of them (lineage 2) is also found in the central Mediterranean and southern Portugal. Phylogeographic results suggest separation of Atlantic and Indo-Pacific lineages resulted from the emergence of the Isthmus of Panama, as well as from dispersal events postdating the closure event, such as the invasion of the Mediterranean Sea by mitochondrial lineages 2 and 3. Molecular clock estimates using the Panama closure event as a calibration for the split of lineages 3 and 4 suggest that A. taxiformis diverged into two main cryptic species (1 + 2 and 3 + 4) about 3.2-5.5 million years ago (Ma), and that the separation of the mitochondrial lineages 1 and 2 occurred 1-2.3 Ma.

A method for isolating high-quality DNA is presented for the green algae Caulerpa sp. (C. racemosa, C. prolifera, and C. taxifolia) and the brown alga Sargassum muticum. These are introduced, and invasive species in Europe, except for the native C. prolifera. Previous methods of extraction, using cetyl trimethyl ammonium bromide or various commercial kits, were used to isolate genomic DNA but either no DNA or DNA of very low quality was obtained. Genomic libraries were attempted with Caulerpa sp. on three occasions and either the restriction enzyme, the Taq polymerase, or the T4 ligase was inhibited, probably by the large amount of polysaccharides in these algae. The method presented here consists of the rapid isolation of stable nuclei, followed by DNA extraction. Yields of 6-10 μg genomic DNA from 1 g fresh blades were obtained. After genomic DNA was isolated from fresh material, the quality was checked by agarose gel. Quantification of DNA concentration was performed using UV spectrophotometric measurement of the A 260 /A 280 ratio. The DNA was suitable for PCR, cloning, and hybridization. The DNA isolated using this method allowed successful construction of microsatellite libraries for Caulerpa species and S. muticum. The technique is inexpensive and appropriate for the isolation of multiple samples of DNA from a small amount of fresh material.

The tropical-subtropical red seaweed Asparagopsis Montagne (Bonnemaisoniales) constitutes the haploid, gametophytic phase in a heteromorphic diplo-haplontic life cycle. The diploid tetrasporophyte is known as the ‘Falkenbergia’ stage. The genus contains two species, A. armata and A. taxiformis, both present in the Mediterranean Sea where they are regarded as introduced. A. armata is morphologically distinct from A. taxiformis in that it possesses long stolons bearing harpoon-like hooks. The seemingly morphologically identical ‘Falkenbergia’ stages of the two Asparagopsis species and phenotypic variation within these species have caused taxonomic confusion. We defined species boundaries in the Mediterranean Sea by inferring phylogenies from sequence data from a variable region in the nuclear LSU rDNA gene, the plastid RuBisCo spacer, and the mitochondrial cox2 – 3 spacer of specimens from the Mediterranean, western Europe and the Canary Islands. Results indicate that A. armata and its ‘Falkenbergia’ tetras...

Twenty-five landraces of the cherry-like tomato named Corbarino, a typical niche product grown in the Sarno valley of the Campania region, have been characterized. The landraces used have been compared to eight cultivars widely spread in the same area of cultivation. The genetic diversity within and between landraces was evaluated through the comparison of DNA fingerprints obtained with (GATA)(4) probe hybridized to TaqI digested genomic DNA. Twenty-two of the Corbarino landraces were homogeneous and were unequivocally characterized by their DNA fingerprints. The others are probably "population varieties" in that within each of them polymorphic DNA fragments were identified. According to the characterization of fruit shapes, four groups were identified. One landrace from each group, harvested at the same ripening degree, was further characterized for yield, fruit quality components, antioxidant activities, and carotenoid contents and compared to Faino F(1) and Tomito F(1) hybrids. The Corbarino landraces were all characterized by high yield, a high level of carbohydrates, and high levels of soluble and total solids. Interestingly, the biotype named ISCI 05 was the most suitable for canning. Landraces ISCI 07 and ISCI 05 gave the highest value of antioxidant activities and carotenoid content. These data, together with previously published results, suggested that ISCI 05 should be proposed for the institution of a Label of Origin.

Three different species of Caulerpa (Caulerpaceae, Chlorophyta) co-occur in the Mediterranean Sea: two of them are found at the central-eastern basin and are typically considered non aggressive components of the Lessepsian flora [(C. chemnitzia(Esper) J.V. Lamouroux and C. racemosa var. lamourouxii (Turner) Weber-van Bosse f. requienii(Montagne) Weber van Bosse)]; a third taxon, C. cylindracea Sonder has aggressively expanded its range since its first observation in 1990, and it is nowadays reported from nearly all the Mediterranean countries. We report a population of C. cylindracea from Almería (Andalusia, Southern Iberian Peninsula) at −30 m depth as to be the westernmost record of the invasive variety on the Mediterranean European coast. Therefore, we made use of morphological description and molecular phylogenetics to provide a complete identification of this invasive seaweed in Southern Spain. Our findings are discussed in light of the composition of the receptor communities, such as maërl bed, edges of Posidonia oceanica (Linnaeus) Delile, and their ecology. Our results confirmed the suggested directionality of the invasive pathway to be westward to the Strait of Gibraltar, mainly supported by sea currents and vectors of anthropogenic origin.En el Mar Mediterráneo se encuentran tres especies diferentes de Caulerpa (Caulerpaceae, Chlorophyta): dos de ellas se localizan en la cuenca centro oriental y se consideran componentes no agresivos de la flora Lessepsiana (C. chemnitzia(Esper) J.V. Lamouroux and C. racemosa var. lamourouxii (Turner) Weber-van Bosse f. requienii(Montagne) Weber van Bosse); un tercer taxon, C. cylindracea Sonder ha expandido de manera agresiva su rango de distribución desde su primera observación en 1990, y actualmente se registra en casi todos los países de la cuenca mediterránea. En este trabajo se informa sobre una población de C. cylindracea en Almería (Andalucía, Sur de España), a −30 m de profundidad, que representaría la cita más occidental de la variedad invasora en las costas mediterráneas europeas. Se realiza una completa identificación de esta alga invasora en el Sur de España mediante una descripción morfológica y filogenia molecular. Los datos obtenidos se discuten en relación a la composición de las comunidades receptoras, tales como fondos de maërl y bordes de Posidonia oceanica (Linnaeus) Delile, y su ecología. Los resultados confirman la direccionalidad de la ruta invasora de la especie hacia el oeste en las proximidades del Estrecho de Gibraltar, principalmente debido a corrientes marinas y vectores de origen antropogénico.

Squat lobsters from Madagascar, Vanuatu, Papua New Guinea, Fiji, eastern Australia and French Polynesia belonging to the Agononida incerta (Henderson, 1888) species complex are described as four new species: A. madagascerta, A. polycerta, A. tasmancerta and A. vanuacerta. This brings to ten the number of species in this complex. All species are morphologically distinguishable only on the basis of the shape of the anterolateral margin of the telson and setation of the dactyli of pereopods 2-4. The morphological delineation of nine of the species and their taxonomic status are robustly supported by phylogenetic analysis of the partial 16S rDNA gene and the partial mitochondrial cytochrome oxidase subunit 1 genes, and in some cases by colour. A phylogenetic analysis of the nine species for which molecular data are available grouped the species in two clades, one of four species with facial spines on the upper surface of pereopod 4 and the other of five species lacking facial spines.

Squat lobsters from Madagascar, Vanuatu, Papua New Guinea, Fiji, eastern Australia and French Polynesia belonging to the Agononida incerta (Henderson, 1888) species complex are described as four new species: A. madagascerta, A. polycerta, A. tasmancerta and A. vanuacerta. This brings to ten the number of species in this complex. All species are morphologically distinguishable only on the basis of the shape of the anterolateral margin of the telson and setation of the dactyli of pereopods 2-4. The morphological delineation of nine of the species and their taxonomic status are robustly supported by phylogenetic analysis of the partial 16S rDNA gene and the partial mitochondrial cytochrome oxidase subunit 1 genes, and in some cases by colour. A phylogenetic analysis of the nine species for which molecular data are available grouped the species in two clades, one of four species with facial spines on the upper surface of pereopod 4 and the other of five species lacking facial spines.

Eight nuclear polymorphic microsatellite markers were characterized from the ascidian Ciona intestinalis whole genome sequence. The behaviour of these loci was investigatedagainst two geographically distinct populations: one from Plymouth, UK the other fromthe Fusaro Lagoon, Italy, both belonging to the type A Ciona cryptic species. The markersexhibited six to 29 alleles and average observed heterozygosity ranging from 0.06 to 0.73.These new microsatellite loci demonstrated to be valuable tools for both population geneticanalysis at different scales and genetic identification of mutant phenotypes frequentlyencountered in Mediterranean populations of C. intestinalis . Keywords : Ciona intestinalis , cryptic species, Mediterranean populations, microsatellites, mutantphenotypes Received 18 September 2006; revision accepted 6 November 2006 The ascidian Ciona intestinalis Linnaeus (Enterogona,Cionidae), with its recently sequenced genome ( Dehal et al .2002), has become an important model system in evolutionaryand developmental investigations (reviewed in Satoh &Levine 2005). In all studies, typically

Surprisingly little is known of the toxic arsenal of cnidarian nematocysts compared to other venomous animals. Here we investigate the toxins of nematocysts isolated from the jellyfish Olindias sambaquiensis. A total of 29 unique ms/ms events were annotated as potential toxins homologous to the toxic proteins from diverse animal phyla, including cone-snails, snakes, spiders, scorpions, wasp, bee, parasitic worm and other Cnidaria. Biological activities of these potential toxins include cytolysins, neurotoxins, phospholipases and toxic peptidases. The presence of several toxic enzymes is intriguing, such as sphingomyelin phosphodiesterase B (SMase B) that has only been described in certain spider venoms, and a prepro-haystatin P-IIId snake venom metalloproteinase (SVMP) that activates coagulation factor X, which is very rare even in snake venoms. Our annotation reveals sequence orthologs to many representatives of the most important superfamilies of peptide venoms suggesting that their origins in higher organisms arise from deep eumetazoan innovations. Accordingly, cnidarian venoms may possess unique biological properties that might generate new leads in the discovery of novel pharmacologically active drugs.

Squat lobsters from Australia, east Africa, Taiwan, Philippines and the Norfolk Ridge (southwestern Pacific) previouslyidentified as Agononida incerta (Henderson, 1888) are redescribed as four new species in addition to the original: A.africerta, A. auscerta, A. indocerta and A. norfocerta. A. rubrizonata Macpherson & Baba, 2009, also earlier confusedwith this species, is redescribed. All six species are morphologically distinguishable only on the basis of the shape of theanterolateral lobe of the telson and the shape and setation of the dactyli of pereopods 2–4. The morphological delineationof these species and their taxonomic status are robustly supported by phylogenetic analysis of the partial mitochondrialCOI marker. Taken together, subtle morphological differences, geographical distribution patterns and genetic discontinuities have important implications for understanding diversity, systematics and evolution of squat lobsters.

Maskrays of the genus Neotrygon (Dasyatidae) have dispersed widely in the Indo-West Pacific being represented largely by an assemblage of narrow-ranging coastal endemics. Phylogenetic reconstruction methods reproduced nearly identical and statistically robust topologies supporting the monophyly of the genus Neotrygon within the family Dasyatidae, the genus Taeniura being consistently basal to Neotrygon, and Dasyatis being polyphyletic to the genera Taeniurops and Pteroplatytrygon. The Neotrygon kuhlii complex, once considered to be an assemblage of color variants of the same biological species, is the most derived and widely dispersed subgroup of the genus. Mitochondrial (COI, 16S) and nuclear (RAG1) phylogenies used in synergy with molecular dating identified paleoclimatic fluctuations responsible for periods of vicariance and dispersal promoting population fragmentation and speciation in Neotrygon. Signatures of population differentiation exist in N. ningalooensis and N. annotata, yet a large-scale geological event, such as the collision between the Australian and Eurasian Plates, coupled with subsequent sea-level falls, appears to have separated a once homogeneous population of the ancestral form of N. kuhlii into southern Indian Ocean and northern Pacific taxa some 4–16 million years ago. Repeated climatic oscillations, and the subsequent establishment of land and shallow sea connections within and between Australia and parts of the Indo-Malay Archipelago, have both promoted speciation and established zones of secondary contact within the Indian and Pacific Ocean basins.

Identification of taxonomical units underpins most biological endeavours ranging from accurate biodiversity estimates to the effective management of sustainably harvested, protected or endangered species. Successful species identification is now frequently based on a combination of approaches including morphometrics and DNA markers. Sequencing of the mitochondrial COI gene is an established methodology with an international campaign directed at barcoding all fishes. We employed COI sequencing alongside traditional taxonomic identification methods and uncovered instances of deep intraspecific genetic divergences among flathead species. Sixty-five operational taxonomic units (OTUs) were observed across the Indo-West Pacific from just 48 currently recognized species. The most comprehensively sampled taxon, Platycephalus indicus, exhibited the highest levels of genetic diversity with eight lineages separated by up to 16.37% genetic distance. Our results clearly indicate a thorough reappraisal of the current taxonomy of P. indicus (and its three junior synonyms) is warranted in conjunction with detailed taxonomic work on the other additional Platycephalidae OTUs detected by DNA barcoding.

Seaweeds were among the first harvested human food supplies in several parts of the world and are today valuable natural resources. They are, however, part of one of the most pressing conservation issues of our time: biological invasions. Global patterns of biodiversity are changing by relocations of organisms at the species and subspecies levels, with the latter often remaining cryptic. The increasing occurrence of marine invasions is mainly due to intensifying maritime traffic and global environmental changes. Following introduction, if suitable conditions for survival occur in the “recipient” environment, seaweeds will establish and spread. Many high-profile invasive seaweeds are commercially used in their native range and have biological traits similar to high-yield terrestrial crops, e.g., high growth rates. The incentives to introduce potentially invasive taxa for commercial use are significant. However, the associated environmental risks are high and robust strategies to prevent and control intentional and accidental seaweed introductions remain essential. For industrial and commercial use, preference should be given to the harvesting and culture of native seaweeds.

Biofilms of the bacterium Pseudoalteromonas induce metamorphosis of acroporid coral larvae. The bacterial metabolite tetrabromopyrrole (TBP), isolated from an extract of Pseudoalteromonas sp. associated with the crustose coralline alga (CCA) Neogoniolithon fosliei, induced coral larval metamorphosis (100%) with little or no attachment (0-2%). To better understand the molecular events and mechanisms underpinning the induction of Acropora millepora larval metamorphosis, including cell proliferation, apoptosis, differentiation, migration, adhesion and biomineralisation, two novel coral gene expression assays were implemented. These involved the use of reverse-transcriptase quantitative PCR (RT-qPCR) and employed 47 genes of interest (GOI), selected based on putative roles in the processes of settlement and metamorphosis. Substantial differences in transcriptomic responses of GOI were detected following incubation of A. millepora larvae with a threshold concentration and 10-fold elevated concentration of TBP-containing extracts of Pseudoalteromonas sp. The notable and relatively abrupt changes of the larval body structure during metamorphosis correlated, at the molecular level, with significant differences (p

The red seaweed genus Asparagopsis Montagne (Bonnemaisoniales) contains two widely introduced species that are considered notorious seaweed invaders worldwide, Asparagopsis armata and A. taxiformis, both characterised by heteromorphic, diplo-haplontic life histories. To uncover cryptic introductions of Asparagopsis along the South African coastline and identify ‘Falkenbergia’ isolates (i.e. tetrasporophytic life-history phase morphologically identical between species), the mitochondrial cox2–3 spacer was sequenced from gametophytes of Asparagopsis taxiformis from Scottburgh, KwaZulu-Natal, on the East Coast, Knysna Lagoon on the South Coast and from tetrasporophytes, otherwise unidentifiable to species level, collected from False Bay near Cape Town on the South-West Coast and Tsitsikamma on the South Coast. Only tetrasporophytes of the temperate Asparagopsis armata were encountered from the Cape Peninsula (Cape Town) probably as far east as to Port St Johns, Eastern Cape province. This is considered an introduced species, and was first collected at Kommetjie (Cape Peninsula) in 1935. Gametophytes of the warm-temperate to tropical A. taxiformis were first collected at Reunion Rocks near Durban in 1984; the KwaZulu-Natal material studied here belongs to an Atlantic Mediterranean cryptic lineage. This taxon is an ecological dominant in some intertidal and shallow subtidal areas in northern KwaZulu-Natal, and is thus considered ‘introduced’ and ‘invasive’. In contrast, A. taxiformis gametophytes, collected in Knysna Lagoon in 2008, clustered with individuals of Indo-Pacific lineage 2. The latter is considered a major invasive lineage in the western Mediterranean, but at present is categorised as introduced in South Africa. This study provides molecular evidence of three independent, cryptic introductions in South Africa, one of them probably very recent, and this is discussed with respect to potential vectors responsible for transport.Keywords: Bonnemaisoniaceae, cryptogenic, introduced, invasive, seaweedAfrican Journal of Marine Science 2011, 33(2): 263–271