A close phylogenetic relationship between Sipuncula and Annelida evidenced from the complete mitochondrial genome sequence of Phascolosoma esculenta

Background: There are many advantages to the application of complete mitochondrial (mt) genomes in the accurate reconstruction of phylogenetic relationships in Metazoa. Although over one thousand metazoan genomes have been sequenced, the taxonomic sampling is highly biased, left with many phyla without a single representative of complete mitochondrial genome. Sipuncula (peanut worms or star worms) is a small taxon of worm-like marine organisms with an uncertain phylogenetic position. In this report, we present the mitochondrial genome sequence of Phascolosoma esculenta, the first complete mitochondrial genome of the phylum. Results: The mitochondrial genome of P. esculenta is 15,494 bp in length. The coding strand consists of 32.1% A, 21.5% C, 13.0% G, and 33.4% T bases (AT = 65.5%; AT skew = -0.019; GC skew = -0.248). It contains thirteen protein-coding genes (PCGs) with 3,709 codons in total, twenty-two transfer RNA genes, two ribosomal RNA genes and a non-coding AT-rich region (AT = 74.2%). All of the 37 identified genes are transcribed from the same DNA strand. Compared with the typical set of metazoan mt genomes, sipunculid lacks trnR but has an additional trnM. Maximum Likelihood and Bayesian analyses of the protein sequences show that Myzostomida, Sipuncula and Annelida (including echiurans and pogonophorans) form a monophyletic group, which supports a closer relationship between Sipuncula and Annelida than with Mollusca, Brachiopoda, and some other lophotrochozoan groups. Conclusion: This is the first report of a complete mitochondrial genome as a representative within the phylum Sipuncula. It shares many more similar features with the four known annelid and one echiuran mtDNAs. Firstly, sipunculans and annelids share quite similar gene order in the mitochondrial genome, with all 37 genes located on the same strand; secondly, phylogenetic analyses based on the concatenated protein sequences also strongly support the sipunculan + annelid clade (including echiurans and pogonophorans). Hence annelid "key-characters" including segmentation may be more labile than previously assumed.

鸡胚吗啡用药对学习记忆和成瘾易感性的影响及其受体机制

Prenatal morphine exposure affects neural development of fetus by impairing learning and memory, and increasing susceptibility to morphine abuse. Because nervous systems have different developmental characteristics during different developmental stages, administration of morphine at different stages also has different effects on learning, memory, and susceptibility to morphine. Due to the precise developmental processes of neurotransmitter systems in chick embryo’s brain, and unique superiority of chick embryo model, the purpose of the present studies was to explore critical periods correlated to the memory impairment and the increasing susceptibility to morphine, via one-trial passive avoidance and conditioned place preference as behavior models. Then the possible roles of mu and delta opioid receptors as the possible mechanism were analyzed. Experiment 1 showed that injecting low dose of morphine (1 mg/kg) during the period embryonic 5 to 8 significantly impaired the function of learning and memory, worse than any other periods of the same treatment. Experiment 2 showed that injecting low dose of morphine during the period embryonic 17 to 20 significantly increased the susceptibility to morphine in the new-born chicks. The affected chicks acquired the morphine conditioned place preference more quickly, and maintained it much longer. Experiment 3 showed that during E5-8, injecting delta receptor antagonist naltrindole reversed the learning and memory impairment caused by morphine while delta receptor agonist DPDPE impaired learning and partial memory function. On the other hand, mu opioid receptors had little effect. As for E17-20, given naloxonazine can reverse the increases of susceptibility to morphine, and the mu receptor agonist DAGO cause the increases of susceptibility to morphine. Delta receptors have no effect. The above results demonstrated that prenatal morphine expousure at different developmental periods of chick embryo caused different influences on memory and susceptibility to morphine. That is, E5-8 is the critical period correlate to memory impairment; and E17-20 is the critical period correlate to susceptibility to morphine. Delta receptors were critical in learning and memory impairment while mu receptors in susceptibility.

La trayectoria deportiva hacia el rendimiento en los deportes colectivos

¿Cómo es la trayectoria seguida por un jugador de fútbol desde que empieza a dar sus primeros pasos con el balón hasta que alcanza el rendimiento que le permita competir en la liga profesional de fútbol?, ¿cómo ocurre en el baloncesto o en el balonmano? Son muchos los factores que influirán sin duda alguna en dicho proceso. Entre dichos factores, en los últimos años, se ha considerado de forma detenida la influencia de la “practica deliberada” en el desarrollo del deportista. Sin embargo, son varios los autores y estudios que explican que no solo influye dicha practica, sino que también es muy importante la influencia del “juego deliberado”, bien en el mismo deporte, bien en otras especialidades deportivas, y que ambos tipos de practica son compatibles formando un continuum en el tiempo. Este artículo tiene como objetivo presentar el estado de la arte en torno a este debate, en el ámbito de los deportes colectivos, analizando si en los deportes colectivos los deportistas se especializan al principio en un solo deporte o bien si practican varias disciplinas deportivas para finalmente dedicarse exclusivamente a un deporte. Los resultados apuntan a que no existe un único camino en el desarrollo del deportista, y que razones de carácter social y cultural son las que realmente condicionan dicho proceso

Radioactive in situ hybridization for detecting diverse gene expression patterns in tissue.

Knowing the timing, level, cellular localization, and cell type that a gene is expressed in contributes to our understanding of the function of the gene. Each of these features can be accomplished with in situ hybridization to mRNAs within cells. Here we present a radioactive in situ hybridization method modified from Clayton et al. (1988)(1) that has been working successfully in our lab for many years, especially for adult vertebrate brains(2-5). The long complementary RNA (cRNA) probes to the target sequence allows for detection of low abundance transcripts(6,7). Incorporation of radioactive nucleotides into the cRNA probes allows for further detection sensitivity of low abundance transcripts and quantitative analyses, either by light sensitive x-ray film or emulsion coated over the tissue. These detection methods provide a long-term record of target gene expression. Compared with non-radioactive probe methods, such as DIG-labeling, the radioactive probe hybridization method does not require multiple amplification steps using HRP-antibodies and/or TSA kit to detect low abundance transcripts. Therefore, this method provides a linear relation between signal intensity and targeted mRNA amounts for quantitative analysis. It allows processing 100-200 slides simultaneously. It works well for different developmental stages of embryos. Most developmental studies of gene expression use whole embryos and non-radioactive approaches(8,9), in part because embryonic tissue is more fragile than adult tissue, with less cohesion between cells, making it difficult to see boundaries between cell populations with tissue sections. In contrast, our radioactive approach, due to the larger range of sensitivity, is able to obtain higher contrast in resolution of gene expression between tissue regions, making it easier to see boundaries between populations. Using this method, researchers could reveal the possible significance of a newly identified gene, and further predict the function of the gene of interest.

Interspecies avian brain chimeras reveal that large brain size differences are influenced by cell-interdependent processes.

Like humans, birds that exhibit vocal learning have relatively delayed telencephalon maturation, resulting in a disproportionately smaller brain prenatally but enlarged telencephalon in adulthood relative to vocal non-learning birds. To determine if this size difference results from evolutionary changes in cell-autonomous or cell-interdependent developmental processes, we transplanted telencephala from zebra finch donors (a vocal-learning species) into Japanese quail hosts (a vocal non-learning species) during the early neural tube stage (day 2 of incubation), and harvested the chimeras at later embryonic stages (between 9-12 days of incubation). The donor and host tissues fused well with each other, with known major fiber pathways connecting the zebra finch and quail parts of the brain. However, the overall sizes of chimeric finch telencephala were larger than non-transplanted finch telencephala at the same developmental stages, even though the proportional sizes of telencephalic subregions and fiber tracts were similar to normal finches. There were no significant changes in the size of chimeric quail host midbrains, even though they were innervated by the physically smaller zebra finch brain, including the smaller retinae of the finch eyes. Chimeric zebra finch telencephala had a decreased cell density relative to normal finches. However, cell nucleus size differences between each species were maintained as in normal birds. These results suggest that telencephalic size development is partially cell-interdependent, and that the mechanisms controlling the size of different brain regions may be functionally independent.

Sea Urchin Body Plan Development and Evolution: An Integrative Transcriptomic Approach

My dissertation work integrates comparative transcriptomics and functional analyses to investigate gene expression changes underlying two significant aspects of sea urchin evolution and development: the dramatic developmental changes associated with an ecologically significant shift in life history strategy and the development of the unusual radial body plan of adult sea urchins.

In Chapter 2, I investigate evolutionary changes in gene expression underlying the switch from feeding (planktotrophic) to nonfeeding (lecithotrophic) development in sea urchins. In order to identify these changes, I used Illumina RNA-seq to measure expression dynamics across 7 developmental stages in three sea urchin species: the lecithotroph Heliocidaris erythrogramma, the closely related planktotroph Heliocidaris tuberculata, and an outgroup planktotroph Lytechinus variegatus. My analyses draw on a well-characterized developmental gene regulatory network (GRN) in sea urchins to understand how the ancestral planktotrophic developmental program was altered during the evolution of lecithotrophic development. My results suggest that changes in gene expression profiles occurred more frequently across the transcriptome during the evolution of lecithotrophy than during the persistence of planktotrophy. These changes were even more pronounced within the GRN than across the transcriptome as a whole, and occurred in each network territory (skeletogenic, endomesoderm and ectoderm). I found evidence for both conservation and divergence of regulatory interactions in the network, as well as significant changes in the expression of genes with known roles in larval skeletogenesis, which is dramatically altered in lecithotrophs. I further explored network dynamics between species using coexpression analyses, which allowed me to identify novel players likely involved in sea urchin neurogenesis and endoderm patterning.

In Chapter 3, I investigate developmental changes in gene expression underlying radial body plan development and metamorphosis in H. erythrogramma. Using Illumina RNA-seq, I measured gene expression profiles across larval, metamorphic, and post-metamorphic life cycle phases. My results present a high-resolution view of gene expression dynamics during the complex transition from pre- to post-metamorphic development and suggest that distinct sets of regulatory and effector proteins are used during different life history phases.

Collectively, my investigations provide an important foundation for future, empirical studies to investigate the functional role of gene expression change in the evolution of developmental differences between species and also for the generation of the unusual radial body plan of sea urchins.

Feeding of anchovy Engraulis encrasicolus larvae in the northwestern Adriatic Sea in response to changing hydrobiological conditions

Results from depth integrated and vertically stratified plankton sampling in the northwestern Adriatic Sea were used for comparison of gut contents of larvae of European anchovy Engraulis encrasicolus with composition and concentration of potential prey in the plankton. Sampling was carried out over a grid of stations both before and after a period of increased wind mixing to investigate changes in food availability and larval feeding success. All larvae had empty guts soon after dusk, indicating daytime feeding and rapid gut clearance. With increasing larval length there was a greater percentage of specimens with empty guts, despite suitable food being available in the plankton for these larger larvae; this suggests differential gut evacuation during sampling-possibly related to the degree of gut development. Larval diet was principally the various developmental stages of copepods, especially calanoid and cyclopoid nauplii, which were preferentially selected by larvae, whereas selection was against harpacticoid nauplii. Lamellibranch larvae and Peridinium were generally abundant in the plankton, but were only present in the gut contents in any number when the preferred dietary organisms were present in the plankton at low concentrations. The number of food organisms in the gut contents increased with concentration of the preferred food organisms in the plankton up to a limit of similar to 50 organisms/l. Within the upper 18 m of the water column, there was a reduction in the proportion of larvae with food in their guts with increasing depth, irrespective of the vertical profile of food concentration. Following a period of wind mixing the composition of the plankton changed. This was reflected in the diet of anchovy larvae, which altered in parallel. There was also an overall 41% decrease in concentration of the preferred food particles of larvae in the plankton following the period of wind mixing, but larvae were still able to maintain their food intake. These results show that anchovy larvae can successfully adapt their diet to a changing prey field and suggest that in the conditions observed in the northern Adriatic, quite radical changes in the feeding environment were probably insufficient to affect overall larval mortality.

Variability In Abundance Vertical-Distribution And Ontogenetic Migrations Of Thysanoessa-Longicaudata (Crustacea Euphausiacea) In The Northeastern Atlantic-Ocean

The vertical distribution, seasonal and ontogenetic migrations and seasonal variability in abundance of Thysanoessa longicaudata (Krøyer) were investigated using the Longhurst-Hardy Plankton Recorder for a 4 yr period (March, 1971 to May, 1975) at Ocean Weather Station “I” (59°00′N; 19°00′W) in the north-eastern Atlantic Ocean. Of 8 species of euphausiids identified at this position, the vast majority were T. longicaudata (for example, 99.5% of the total euphausiids in 1972 belonged to this species). From March to October the majority of calyptopes, furciliae and adults of T. longicaudata were found in the upper 100 m. The major spawning occurred in spring at a water temperature of 9° to 10°C and calyptopes and furciliae appeared in late April, reaching their maximum abundance in May. There was no evidence of large-scale diurnal migrations, although an extensive ontogenetic migration of young developmental stages was observed. The eggs were found from 100 m down to 800 m, the maximum depth of sampling, and the vertical distribution of the three naupliar stages showed a “developmental ascent” as they matured. During the main reproductive period in May, over 70% of all nauplii were below 500 m while more than 94% of Calyptopis Stage I were above 500 m with their maximum abundance in the euphotic zone (0 to 50 m). Calyptopis Stage I is the first feeding stage and it is this stage which shows the largest ontogenetic migration. Brief descriptions of the egg and nauplii are given.

Respiration Of The Meiobenthic Harpacticoid Copepod Tachidius-Discipes Giesbrecht From An Estuarine Mudflat

The relationships between respiration (R) and body volume (V) for all developmental stages of the harpacticoid copepod Tachidius discipes Giesbrecht have been investigated. The relationships for laboratory-reared animals and animals from the field are significantly different. They are: logR = −0.07 + 1.10 logV for laboratory-reared animals and log R = −0.10 + 0.82 logV for field animals. The effect of temperature on the respiration rate of adult males, over the temperature range 5–20°C, was described by a Q10 of 2.09 ± 0.24. The respiration rate of an adult T. discipes is very similar to that of a similar sized nematode from the same field site and is compared with published data for other harpacticoids.

Spatial demography of Calanus finmarchicus in the Irminger Sea

Continuous Plankton Recorder data suggest that the Irminger Sea supports a major proportion of the surface-living population of the copepod Calanus finmarchicus in the northern North Atlantic, but there have been few studies of its population dynamics in the region. In this paper, we document the seasonal changes in the demographic structure of C finmarchicus in the Irminger Sea from a field programme during 2001/2002, and the associations between its developmental stages and various apparent bio-physical zones. Overwintering stages were found widely at depth (>500 m) across the Irminger Sea, and surviving females were widely distributed in the surface waters the following spring. However, recruitment of the subsequent generation was concentrated around the fringes of the Irminger Sea basin, along the edges of the Irminger and East Greenland Currents, and not in the central basin. In late summer animals were found descending back to overwintering depths in the Central Irminger Sea. The key factors dictating this pattern of recruitment appear to be (a) the general circulation regime, (b) predation on eggs in the spring, possibly by the surviving GO stock, and (c) mortality of first feeding naupliar stages in the central basin where food concentrations appear to be low throughout the year. We compared the demographic patterns in 2001/2002 with observations from the only previous major survey in 1963 and with data from the Continuous Plankton Recorder (CPR) surveys. In both previous data sets, the basic structure of GO ascent from the central basin and G1 recruitment around the fringes was a robust feature, suggesting that it is a recurrent phenomenon. The Irminger Sea is a complex mixing zone between polar and Atlantic water masses, and it has also been identified as a site of sporadic deep convection. The physical oceanographic characteristics of the region are therefore potentially sensitive to climate fluctuations. Despite this, the abundance of C finmarchicus in the region, as measured by the CPR surveys, appears not to have responded to climate factors linked to the North Atlantic Oscillation Index, in contrast with the stocks in eastern Atlantic areas. We speculate that this may because biological factors (production and mortality), rather than transport processes are the key factors affecting the population dynamics in the Irminger Sea. (c) 2007 Elsevier Ltd. All rights reserved.

Transmission electron microscopy of marine crustacean eggs

Ultrastructural investigations of eggs can be important in helping to understand embryonic development. There are few transmission electron microscope studies of marine arthropod eggs, however, as they have proved difficult to fix and infiltrate with resin. Here, we describe a modification of a standard method that allows the preparation of the quite different eggs of the marine copepod, Acartia tonsa and the lobster, Homarus gammarus, for transmission electron microscopy. By using double fixation and an extended resin infiltration time we obtained good preparations for electron microscopy. We anticipate that these modifications to the standard protocol will be widely applicable and useful for the study of the eggs and early developmental stages of many marine arthropod taxa. Les recherches sur l'ultrastructure des oeufs peuvent être importantes en aidant à comprendre le développement embryonnaire. Il existe cependant peu d'études en microscopie électronique à transmission sur les oeufs d'arthropodes marins, car il est difficile de les fixer et d'y infiltrer de la résine. Dans ce travail, nous décrivons une modification de la méthode standard, qui permet la préparation pour la microscopie électronique à transmission d'oeufs aussi différents que ceux du copépode marin Acartia tonsa et du homard Homarus gammarus. En utilisant une double fixation et un temps plus long d'infiltration de la résine, nous avons obtenu de bonnes préparations pour la microscopie électronique. Nous prévoyons que ces modifications du protocole standard seront largement applicables et utiles pour l'étude des oeufs et des premiers stades de développement de nombreux taxons d'arthropodes marins.

High dispersal potential has maintained long-term population stability in the North Atlantic copepod Calanus finmarchicus

The cool-water copepod Calanus finmarchicus is a key species in North Atlantic marine ecosystems since it represents an important food resource for the developmental stages of several fish of major economic value. Over the last 40 years, however, data from the Continuous Plankton Recorder survey have highlighted a 70 per cent reduction in C. finmarchicus biomass, coupled with a gradual northward shift in the species's distribution, which have both been linked with climate change. To determine the potential for C. finmarchicus to track changes in habitat availability and maintain stable effective population sizes, we have assessed levels of gene flow and dispersal in current populations, as well as using a coalescent approach together with palaeodistribution modelling to elucidate the historical population demography of the species over previous changes in Earth's climate. Our findings indicate high levels of dispersal and a constant effective population size over the period 359 000–566 000 BP and suggest that C. finmarchicus possesses the capacity to track changes in available habitat, a feature that may be of crucial importance to the species's ability to cope with the current period of global climate change.

Have we been underestimating the effects of ocean acidification in zooplankton?

Understanding how copepods may respond to ocean acidification (OA) is critical for risk assessments of ocean ecology and biogeochemistry. The perception that copepods are insensitive to OA is largely based on experiments with adult females. Their apparent resilience to increased carbon dioxide (pCO2 ) concentrations has supported the view that copepods are 'winners' under OA. Here, we show that this conclusion is not robust, that sensitivity across different life stages is significantly misrepresented by studies solely using adult females. Stage-specific responses to pCO2 (385-6000 μatm) were studied across different life stages of a calanoid copepod, monitoring for lethal and sublethal responses. Mortality rates varied significantly across the different life stages, with nauplii showing the highest lethal effects; nauplii mortality rates increased threefold when pCO2 concentrations reached 1000 μatm (year 2100 scenario) with LC50 at 1084 μatm pCO2 . In comparison, eggs, early copepodite stages, and adult males and females were not affected lethally until pCO2 concentrations ≥3000 μatm. Adverse effects on reproduction were found, with >35% decline in nauplii recruitment at 1000 μatm pCO2 . This suppression of reproductive scope, coupled with the decreased survival of early stage progeny at this pCO2 concentration, has clear potential to damage population growth dynamics in this species. The disparity in responses seen across the different developmental stages emphasizes the need for a holistic life-cycle approach to make species-level projections to climate change. Significant misrepresentation and error propagation can develop from studies which attempt to project outcomes to future OA conditions solely based on single life history stage exposures.

A molecular method for the identification of resting eggs of acartiid copepods in the Thau lagoon, France

Acartia and Paracartia species, often known to co-occur, can exhibit complex life cycles, including the production of resting eggs. Studying and understanding their population dynamics is hindered by the inability to identify eggs and early developmental stages using morphological techniques. We have developed a simple molecular technique to distinguish between the three species of the Acartiidae family (Acartia clausi, A. discaudata and Paracartia grani) that co-occur in the Thau lagoon (43�250N; 03�400E) in southern France. Direct amplification of a partial region of the mitochondrial cytochrome oxidase I gene by polymerase chain reaction and subsequent restriction fragment length polymorphism results in a unique restriction profile for each species. The technique is capable of determining the identity of individual eggs, including resting eggs retrieved from sediment samples, illustrating its application in facilitating population dynamic studies of this ubiquitous and important member of the zooplankton community.

Specific and sensitive detection of Nosema bombi (Microsporidia : Nosematidae) in bumble bees (Bombus spp.; Hymenoptera : Apidae) by PCR of partial rRNA gene sequences

A polymerase chain reaction (PCR) based method was developed for the specific and sensitive diagnosis of the microsporidian parasite Nosema bombi in bumble bees (Bombus spp.). Four primer pairs, amplifying ribosomal RNA (rRNA) gene fragments, were tested on N. bombi and the related microsporidia Nosema apis and Nosema ceranae, both of which infect honey bees. Only primer pair Nbombi-SSU-Jf1/Jr1 could distinguish N. bombi (323 bp amplicon) from these other bee parasites. Primer pairs Nbombi-SSU-Jf1/Jr1 and ITS-f2/r2 were then tested for their sensitivity with N. bombi spore concentrations from 107 down to 10 spores diluted in 100 mu l of either (i) water or (ii) host bumble bee homogenate to simulate natural N. bombi infection (equivalent to the DNA from 10(6) spores down to 1 spore per PCR). Though the N. bombi-specific primer pair Nbombi-SSU-Jf1/Jr1 was relatively insensitive, as few as 10 spores per extract (equivalent to 1 spore per PCR) were detectable using the N. bombi-non-specific primer pair ITS-f2/r2, which amplifies a short fragment of similar to 120 bp. Testing 99 bumble bees for N. bombi infection by light microscopy versus PCR diagnosis with the highly sensitive primer pair ITS-f2/r2 showed the latter to b more accurate. PCR diagnosis of N. bombi using a combination of two primer pairs (Nbombi-SSU-Jf1/Jr1 and ITS-f2/r2) provides increased specificity, sensitivity, and detection of all developmental stages compared with light microscopy. (c) 2005 Elsevier Inc. All rights reserved.