rbcL analysis supports the tribe Gymnothamnieae Kajimura (Ceramiaceae, Rhodophyta)

The tribe Gymnothamnieae Kajimura was proposed for the monotypic ceramiacean genus Gymnothamnion J. Agardh, previously placed either in the Ptiloteae Cramer or the Antithamnieae Hommersand. A bisporangial isolate of G. elegans (Schousboe ex C. Agardh) J. Agardh from Morocco formed only bisporangia in culture. Its smaller uninucleate cells and sporangia than those of tetrasporophytes suggested that bisporophytes may be haploid as in another member of the Ceramiaceae, Aglaothamnion diaphanum L'Hardy-Halos et Maggs. Phylogenetic analyses of the gene for the large subunit of rubisco (rbcL) from Gymnothamnion and representatives of eight other tribes of the Ceramiaceae confirmed that the removal of Gymnothamnion from the Ptiloteae and the Antithamnieae was warranted. Whereas all tribes with two or more representatives in our analyses were moderately or robustly resolved, Gymnothamnion did not form a strong clade with any other taxa. Analysis of rbcL sequences failed to resolve relationships between tribes, probably due to saturation at the high levels of sequence divergence found. In addition to reproductive features previously reported and interpreted as primitive, G. elegans shows a primitive vegetative feature and it is suggested that Gymnothamnion may be one of the most basal of the taxa presently included in the Ceramiaceae.

Phylogeographic structure of brown trout (Salmo trutta) in Britain and Ireland: glacial refugia, post-glacial colonisation, and origins of sympatric populations

The phylogeographical structure of brown trout Salmo trutta in Britain and Ireland was studied using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis of four mitochondrial DNA segments (16S/ND1, ND5/6, COXIII/ND5 and ND5/12S). Analysis of 3636 individuals from 83 sites-morphotypes revealed a total of 25 haplotypes. These haplotypes were nested in seven two-step clades. Although there was a clear geographical patterning to the occurrence of derived clades, admixture among ancestral clades was extensive throughout the studied area. A relevant feature of the data was that some populations contained mixtures of highly divergent clades. This type II phylogeographic pattern is uncommon in nature. Clade intermixing is likely to have taken place during earlier interglacials as well as since the Last Glacial Maximum. The anadromous life history of many S. trutta populations has probably also contributed to clade mixing. Based on the data presented here and published data, postglacial colonization of Britain and Ireland most likely involved S. trutta from at least five potential glacial refuges. Probable locations for such refugia were: south of England-western France, east of the Baltic Sea, western Ireland, Celtic Sea and North Sea. Ferox S. trutta, as defined by their longevity, late maturation and piscivory, exhibited a strong association with a particular clade indicating that they share a common ancestor. Current evidence indicates that the Lough Melvin gillaroo S. trutta and sonaghen S. trutta sympatric types diverged prior to colonization of Lough Melvin and, although limited gene flow has occurred since secondary contact, they have remained largely reproductively isolated due to inlet and outlet river spawning segregation. Gillaroo S. trutta may reflect descendents of a previously more widespread lineage that has declined due to habitat alterations particularly affecting outlet rivers. The mosaic-like distribution of mtDNA lineages means that conservation prioritization in Britain and Ireland should be based on the biological characteristics of local populations rather than solely on evolutionary lineages.

Crinoids for lunch? An unexpected biotic interaction from the Upper Ordovician of Scotland

Specimens of the polyplacophoran mollusk 'Helminthochiton' thraivensis Reed from the Upper Ordovician of southwest Scotland provide rare examples of complete valve series preserved in near life position, albeit as external molds. Application of high-resolution X-ray microtomography to one such specimen has revealed the exceptional preservation of its last meal, which included elements of a crinoid column, in its intestine. The interaction was either predatory or scavenging; extant chitons are not known to be crinoidivorous. This is the earliest direct record of predation or scavenging on crinoids in the fossil record. It is also the first indication that the broad axial canal of primitive crinoids may have contained nutritious tissues. The predatory or scavenging habit of H. thraivensis is consistent with its inferred phylogenetic position as a stem-group aplacophoran and provides new data suggesting an origin of carnivory early in the evolution of this clade.

Deep molluscan phylogeny: synthesis of palaeontological and neontological data

The position of the earliest-derived living molluscs, the Polyplacophora ( chitons) and shell-less vermiform Aplacophora, remains highly contentious despite many morphological, developmental and molecular studies of extant organisms. These two groups are thought to represent either a basal molluscan grade or a clade (Aculifera) sister to the 'higher' molluscs (Conchifera). These incompatible hypotheses result in very different predictions about the earliest molluscs. A new cladistic analysis incorporating both Palaeozoic and extant molluscs is presented here. Our results support the monophyly of Aculifera and suggest that extant aplacophorans and polyplacophorans both derive from a disparate group of multivalved molluscs in two major clades. Reanalysis of the critical Ordovician taxon 'Helminthochiton' thraivensis shows that this animal lacks a true foot despite bearing polyplacophoran-like valves. Its position within our phylogenetic reconstruction indicates that many fossil 'polyplacophorans' in the order Palaeoloricata are likely to represent footless stem-group aplacophorans. 'H.' thraivensis and similar forms such as Acaenoplax may be morphological stepping stones between chitons and the shell-less aplacophorans. Our results imply that crown-group molluscan synapomorphies include serial repetition, the presence of a foot, a mineralized scleritome and a creeping rather than worm-like mode of life.

Hibernation and non-shivering thermogenesis in the Hottentot golden mole (Amblysomus hottentottus longiceps)

Although heterothermy (hibernation and torpor) is a common feature among mammals, there is debate over whether it is a derived or ancestral trait relative to endothermic homeothermy. Determination of the physiological characteristics of primitive mammals is central to understanding the evolution of endothermy. Moreover, evaluation of physiological mechanisms responsible for endothermic heat production [e.g. non-shivering thermogenesis (NST)] is key to understanding how early mammals responded to historical climate changes and colonised different geographical regions. Here we investigated the capacity for NST and heterothermy in the Hottentot golden mole, a basal eutherian mammal. NST was measured as the metabolic response to injections of noradrenalin and heterothermy by recording body temperature in free-ranging animals. We found that hibernation and torpor occurred and that the seasonal phenotypic adjustment of NST capacity was similar to that found in other placental mammals. Using phylogenetically independent contrasts, we compared measured values of NST with those obtained from the literature. This showed that all variation in NST was accounted for by differences in phylogeny and not zoogeography. These findings lend support to the observation that NST and heterothermy occur in the Afrotheria, the basal placental mammalian clade. Furthermore, this work suggests that heterothermy, rather than homeothermy is a plesiomorphic trait in mammals and supports the notion that NST mechanisms are phylogenetically ancient.

Evolution in the deep sea: a combined analysis of the earliest diverging living chitons (Mollusca : Polyplacophora : Lepidopleurida)

Lepidopleurida is the earliest diverged group of living polyplacophoran molluscs. They are found predominantly in the deep sea, including sunken wood, cold seeps, other abyssal habitats, and a few species are found in shallow water. The group is morphologically identified by anatomical features of their gills, sensory aesthetes, and gametes. Their shell features closely resemble the oldest fossils that can be identified as modern polyplacophorans. We present the first molecular phylogenetic study of this group, and also the first combined phylogenetic analysis for any chiton, including three gene regions and 69 morphological characters. The results show that Lepidopleurida is unambiguously monophyletic, and the nine genera fall into five distinct clades, which partly support the current view of polyplacophoran taxonomy. The genus Hanleyella Sirenko, 1973 is included in the family Protochitonidae, and Ferreiraellidae constitutes another distinct clade. The large cosmopolitan genus Leptochiton Gray, 1847 is not monophyletic; Leptochiton and Leptochitonidae sensu stricto are restricted to North Atlantic and Mediterranean taxa. Leptochitonidae s. str. is sister to Protochitonidae. The results also suggest two separate clades independently inhabiting sunken wood substrates in the south-west Pacific. Antarctic and other chemosynthetic-dwelling species may be derived from wood-living species. Substantial taxonomic revision remains to be done to resolve lepidopleuran classification, but the phylogeny presented here is a dramatic step forward in clarifying the relationships within this interesting group.

Llama antibody fragments have good potential for application as HIV-1 topical microbicides

There is an urgent global need for preventative strategies against HIV-1 infections. Llama heavy-chain antibody fragments (VHH) are a class of molecules recently described as potent cross-clade HIV-1 entry inhibitors. We studied the potential of a VHH-based microbicide in an application-oriented fashion. We show that VHH can be inexpensively produced in high amounts in the GRAS organism S. cerevisiae, resulting in very pure, and endotoxin free product. VHH are very stable under conditions they might encounter during transport, storage or use by women. We developed active formulations of VHH in aqueous gel and compressed and lyophilized tablets for controlled release from an intra vaginal device. The release profile of the VHH from e.g. a vaginal ring suggests sufficient bioavailability and protective concentration of the molecule at the mucosal site at the moment of the infection. The ex vivo penetration kinetics through human tissues show that the VHH diffuse into the mucosal layer and open the possibility to create a second defense layer either by blocking the HIV receptor binding sites or by blocking the receptors of immune cells in the mucosa. In conclusion, our data show that VHH have

Functional and Structural Diversification of the Anguimorpha Lizard Venom System

Venom has only been recently discovered to be a basal trait of the Anguimorpha lizards. Consequently, very little is known about the timings of toxin recruitment events, venom protein molecular evolution, or even the relative physical diversifications of the venom system itself. A multidisciplinary approach was used to examine the evolution across the full taxonomical range of this similar to 130 million-year-old clade. Analysis of cDNA libraries revealed complex venom transcriptomes. Most notably, three new cardioactive peptide toxin types were discovered (celestoxin, cholecystokinin, and YY peptides). The latter two represent additional examples of convergent use of genes in toxic arsenals, both having previously been documented as components of frog skin defensive chemical secretions. Two other novel venom gland-overexpressed modified versions of other protein frameworks were also recovered from the libraries (epididymal secretory protein and ribonuclease). Lectin, hyaluronidase, and veficolin toxin types were sequenced for the first time from lizard venoms and shown to be homologous to the snake venom forms. In contrast, phylogenetic analyses demonstrated that the lizard natriuretic peptide toxins were recruited independently of the form in snake venoms. The de novo evolution of helokinestatin peptide toxin encoding do-mains within the lizard venom natriuretic gene was revealed to be exclusive to the helodermatid/anguid subclade. New isoforms were sequenced for cysteine-rich secretory protein, kallikrein, and phospholipase A 2 toxins. Venom gland morphological analysis revealed extensive evolutionary tinkering. Anguid glands are characterized by thin capsules and mixed glands, serous at the bottom of the lobule and mucous toward the apex. Twice, independently this arrangement was segregated into specialized serous protein-secreting glands with thick capsules with the mucous lobules now distinct (Heloderma and the Lanthanotus/Varanus clade). The results obtained highlight the importance of utilizing evolution-based search strategies for biodiscovery and emphasize the largely untapped drug design and development potential of lizard venoms. Molecular & Cellular Proteomics 9:2369-2390, 2010.

Taxonomy and phylogeny of Osmundea (Rhodomelaceae, Rhodophyta) in Atlantic Europe

Two species of Osmundea Stackhouse (Rhodomelaceae, Rhodophyta) that occur in Atlantic Europe have been confused under the names Osmundea ramosissima (Oeder) Athanasiadis and Osmundea truncata (Kutzing) Nam et Maggs, regarded until now as a synonym of O. ramosissima, An epitype from its type locality (Stavanger, Norway) is selected for Osmundea ramosissima Athanasiadis, recognized here as a valid name for Fucus ramosissimus Oeder, nom. illeg. Details of vegetative and reproductive morphology of O. ramosissima are reported, based on material from France, the British Isles, and Helgoland. Osmundea ramosissima resembles other species of Osmundea in its vegetative axial segments with two pericentral cells and one trichoblast, spermatangial development from apical and epidermal cells (filament type), the formation of five pericentral cells in the procarp-bearing segment of the female trichoblast, and tetrasporangial production from random epidermal cells. Among the species of Osmundea, O. ramosissima is most similar to O. truncata. Both species have discoid holdfasts, secondary pit connections between epidermal cells, and cup-shaped spermatangial pits. They differ in that: (a) O. ramosissima lacks lenticular wail thickenings and refractive needle-like inclusions in medullary cells, both of which are present in O. truncata; (b) O. ramosissima has branched spermatangial filaments that terminate in a cluster of several cells, whereas in O. truncata the unbranched spermatangial filaments have a single large terminal sterile cell; and (c) cystocarps of O. ramosissima lack protuberant ostioles but ostioles are remarkably protuberant in o. truncata. Phylogenetic analyses of rbcL sequences of Laurencia obtusa (Hudson) Lamouroux and all five Atlantic European species of Osmundea, including the type species, strongly support the generic status of Osmundea. Osmundea ramosissima and O. truncata are closely related (5.2% sequence divergence) and form a well-supported clade sister to a clade consisting of O. pinnatifida (Hudson) Stack-house, O. osmunda Stackhouse and O. hybrida (A. P. de Candolle) Nam. The formation of secondary pit connections between epidermal cells is a synapomorphy for the O. ramosissima + O. truncata clade. The close relationship between species with cup-shaped spermatangial pits (Osmundea hybrida) and urn-shaped pits (Osmundea pinnatifida and Osmundea osmunda) shows that spermatangial pit shape is not an important phylogenetic character. Parsimony analysis of a morphological data set also supports the genus Osmundea but conflicts with the molecular trees in infrageneric relationships, placing O. hybrida basal within the Osmundea clade and grouping O. osmunda and O. pinnatifida but not O. truncata and O. ramosissima. A key to Osmundea species is presented.

Small genetic differences between ericoid mycorrhizal fungi affect nitrogen uptake by Vaccinium

Ericoid mycorrhizal fungi have been shown to differ in their pattern of nitrogen (N) use in pure culture. Here, we investigate whether this functional variation is maintained in symbiosis using three ascomycetes from a clade not previously shown to include ericoid mycorrhizal taxa. Vaccinium macrocarpon and Vaccinium vitis-idaea were inoculated with three fungal strains known to form coils in Vaccinium roots, which differed in their patterns of N use in liquid culture. (15)N was used to trace the uptake of -N, -N and glutamine-N into shoots. (15)N transfer differed among the three fungal strains, including two that had identical internal transcribed spacer (ITS) sequences, and was quantitatively related to fungal growth in liquid culture at low carbon availability. These results demonstrate that functional differences among closely related ericoid mycorrhizal fungi are maintained in symbiosis with their hosts, and suggest that N transfer to plant shoots in ericoid mycorrhizas is under fungal control.

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(1-9). 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(3,6,8), proximate to other derived clades such as Cephalopoda(2,3,10), or as sister group to the Polyplacophora, forming the clade Aculifera(1,5,7,11,12). Resolution of this debate is required to allow the evolutionary origins of Mollusca to be reconstructed with confidence. Recent fossil finds(13-16) support the Aculifera hypothesis, demonstrating that the Palaeozoic-era palaeoloricate 'chitons' included taxa combining certain polyplacophoran and aplacophoran characteristics(5). 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 Lagerstatte(17,18) (about 425 million years BP), a Silurian deposit preserving a marine biota(18) in unusual three-dimensional detail. The specimen is reconstructed three-dimensionally through physical-optical tomography(19). Phylogenetic analysis indicates that this and many other palaeoloricate chitons are crown-group aplacophorans.

A tale of two chitons: Is habitat specialisation linked to distinct associated bacterial communities?

Although most chitons (Mollusca: Polyplacophora) are shallow-water molluscs, diverse species also occur in deep-sea habitats. We investigated the feeding strategies of two species, Leptochiton boucheti and Nierstraszella lineata, recovered on sunken wood sampled in the western Pacific, close to the Vanuatu Islands. The two species display distinctly different associations with bacterial partners. Leptochiton boucheti harbours Mollicutes in regions of its gut epithelium and has no abundant bacterium associated with its gill. Nierstraszella lineata displays no dense gut-associated bacteria, but harbours bacterial filaments attached to its gill epithelium, related to the Deltaproteobacteria symbionts found in gills of the wood-eating limpet Pectinodonta sp. Stable carbon and nitrogen isotope signatures and an absence of cellulolytic activity give evidence against a direct wood-feeding diet; both species are secondary consumers within the wood food web. We suggest that the distinct associations with bacterial partners are linked to niche specialisations of the two species. Nierstraszella lineata is in a taxonomic family restricted to sunken wood and is possibly adapted to more anoxic conditions thanks to its gill-associated bacteria. Leptochiton boucheti is phylogenetically more proximate to an ancestral form not specialised on wood and may itself be more of a generalist; this observation is congruent with its association with Mollicutes, a bacterial clade comprising gut-associated bacteria occurring in several metazoan phyla.

The C-Terminal Variable Region Specifies the Dynamic Properties of Arabidopsis Microtubule-Associated Protein MAP65 Isotypes

The microtubule-associated protein, MAP65, is a member of a family of divergent microtubule-associated proteins from different organisms generally involved in maintaining the integrity of the central spindle in mitosis. The dicotyledon Arabidopsis thaliana and the monocotyledon rice (Oryza sativa) genomes contain 9 and 11 MAP65 genes, respectively. In this work, we show that the majority of these proteins fall into five phylogenetic clades, with the greatest variation between clades being in the C-terminal random coil domain. At least one Arabidopsis and one rice isotype is within each clade, indicating a functional specification for the C terminus. In At MAP65-1, the C-terminal domain is a microtubule binding region (MTB2) harboring the phosphorylation sites that control its activity. The At MAP65 isotypes show differential localization to microtubule arrays and promote microtubule polymerization with variable efficiency in a MTB2-dependent manner. In vivo studies demonstrate that the dynamics of the association and dissociation of different MAP65 isotypes with microtubules can vary up to 10-fold and that this correlates with their ability to promote microtubule polymerization. Our data demonstrate that the C-terminal variable region, MTB2, determines the dynamic properties of individual isotypes and suggest that slower turnover is conditional for more efficient microtubule polymerization.

Pan-genome analysis of the emerging foodborne pathogen Cronobacter spp. suggests a species-level bidirectional divergence driven by niche adaptation

Background: Members of the genus Cronobacter are causes of rare but severe illness in neonates and preterm infants following the ingestion of contaminated infant formula. Seven species have been described and two of the species genomes were subsequently published. In this study, we performed comparative genomics on eight strains of Cronobacter, including six that we sequenced (representing six of the seven species) and two previously published, closed genomes.

Results: We identified and characterized the features associated with the core and pan genome of the genus Cronobacter in an attempt to understand the evolution of these bacteria and the genetic content of each species. We identified 84 genomic regions that are present in two or more Cronobacter genomes, along with 45 unique genomic regions. Many potentially horizontally transferred genes, such as lysogenic prophages, were also identified. Most notable among these were several type six secretion system gene clusters, transposons that carried tellurium, copper and/or silver resistance genes, and a novel integrative conjugative element.

Conclusions: Cronobacter have diverged into two clusters, one consisting of C. dublinensis and C. muytjensii (Cdub-Cmuy) and the other comprised of C. sakazakii, C. malonaticus, C. universalis, and C. turicensis, (Csak-Cmal-Cuni-Ctur) from the most recent common ancestral species. While several genetic determinants for plant-association and human virulence could be found in the core genome of Cronobacter, the four Cdub-Cmuy clade genomes contained several accessory genomic regions important for survival in a plant-associated environmental niche, while the Csak-Cmal-Cuni-Ctur clade genomes harbored numerous virulence-related genetic traits.

Neutral Genomic Microevolution of a Recently Emerged Pathogen, Salmonella enterica Serovar Agona

Salmonella enterica serovar Agona has caused multiple food-borne outbreaks of gastroenteritis since it was first isolated in 1952. We analyzed the genomes of 73 isolates from global sources, comparing five distinct outbreaks with sporadic infections as well as food contamination and the environment. Agona consists of three lineages with minimal mutational diversity: only 846 single nucleotide polymorphisms (SNPs) have accumulated in the non-repetitive, core genome since Agona evolved in 1932 and subsequently underwent a major population expansion in the 1960s. Homologous recombination with other serovars of S. enterica imported 42 recombinational tracts (360 kb) in 5/143 nodes within the genealogy, which resulted in 3,164 additional SNPs. In contrast to this paucity of genetic diversity, Agona is highly diverse according to pulsed-field gel electrophoresis (PFGE), which is used to assign isolates to outbreaks. PFGE diversity reflects a highly dynamic accessory genome associated with the gain or loss (indels) of 51 bacteriophages, 10 plasmids, and 6 integrative conjugational elements (ICE/IMEs), but did not correlate uniquely with outbreaks. Unlike the core genome, indels occurred repeatedly in independent nodes (homoplasies), resulting in inaccurate PFGE genealogies. The accessory genome contained only few cargo genes relevant to infection, other than antibiotic resistance. Thus, most of the genetic diversity within this recently emerged pathogen reflects changes in the accessory genome, or is due to recombination, but these changes seemed to reflect neutral processes rather than Darwinian selection. Each outbreak was caused by an independent clade, without universal, outbreak-associated genomic features, and none of the variable genes in the pan-genome seemed to be associated with an ability to cause outbreaks. © 2013 Achtman et al