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  1. Swaraj Basu
  2. Roger Butlin
  3. Henrik Christiansen
  4. Friederike Eimer
  5. Johan Hollander
  6. Kerstin Johannesson
  7. Anja Kamp (poster)
  8. Jeanine Olsen
  9. Mattias Ormestad
  10. Mats Töpel (poster)
  11. Marina Rafajlović
  12. Mark Ravinet (poster)
  13. Kirsikka Sillanpää (poster)
  14. Helen Sköld

Computational approaches in comparative genomics with a focus towards the non-coding genome.

Swaraj Basu
Department of Medical Biochemistry and Cell Biology, Institute of Biomedicine, The Sahlgrenska Academy, Sweden

A pertinent question in biology is to identify relevant molecularmechanisms which shape the evolution of species. Advances in high throughput technologies have led to a drastic rise in the number of species with available genomes and transcriptomes. Developing novel computational strategies which utilize this huge repository of genomic and transcriptomic data to identify and answer relevant biological problems remains a continuous challenge. Specially the recent focus on the non-coding part of the genome has led to a rising interest in tools and techniques which aid in elucidating the functional role of micro RNAs,
long non-coding RNAs (lncRNA) and conserved non-coding elements (CNEs) with respect to disease and development in several species (Dimitrieva and Bucher, 2013; Necsulea et al., 2014; Nepal et al., 2015).

In this context a standardized protocol for genome and transcriptome annotation is of utmost importance for all downstream analyses. Hence we developed sequence annotation platform which can identify both coding and non-coding genes along with additional plugins which perform gene ontology enrichment and differential expression analyses. The pipeline was used to annotate the transcriptome of the spotted green pufferfish and the genome of a marine diatom. The annotation was followed by downstream analyses which identified, lncRNAs transcribed during early development in pufferfish and CNEs associated with regulation of transcription in the marine diatom (Pseudo-nitzschia multistriata). Further the results also suggest that it is imperative to remain highly stringent while analyzing the non-coding genome to avoid encountering transcriptional artefacts which might be mis-annotated as functional ncRNAs. Finally a comparative approach taking into account several species adds to the relevance of the findings and highlights potential candidates optimal for experimental validation.

Dimitrieva, S., and Bucher, P. (2013). UCNEbase--a database of ultraconserved non-coding elements and genomic regulatory blocks. Nucleic Acids Res. 41, D101-109.

Necsulea, A., Soumillon, M., Warnefors, M., Liechti, A., Daish, T., Zeller, U.,
Baker, J.C., Grützner, F., and Kaessmann, H. (2014). The evolution of lncRNA repertoires and expression patterns in tetrapods. Nature 505, 635–640.

Nepal, C., Coolen, M., Hadzhiev, Y., Cussigh, D., Mydel, P., Steen, V.M., Carninci, P., Andersen, J.B., Bally-Cuif, L., Müller, F., et al. (2015). Transcriptional, posttranscriptional and chromatin-associated regulation of pri-miRNAs, pre-miRNAs and moRNAs. Nucleic Acids Res. gkv1354.

The origin of barriers to gene exchange

Roger Butlin
Department of Animal and Plant Sciences, University of Sheffield and Dept Marine Sciences, University of Gothenburg

I will outline plans for the next five years with ERC funding, using Littorina contact zones to understand how the processes of coupling and reinforcement might contribute to speciation.


Application of ddRAD sequencing for population genomics of Antarctic fish 

Henrik Christiansen1, Anton P. Van de Putte1,2, Filip A.M. Volckaert1
1 Laboratory of Biodiversity and Evolutionary Genomics (LBEG), KU Leuven, Ch. Deberiotstraat 32, B-3000 Leuven, Belgium
2 Royal Belgian Institute of Natural Sciences, Vautiersstraat 29, B-1000 Brussels, Belgium

Contact email: henrik.christiansen@bio.kuleuven.be

The teleost fauna of the Southern Ocean is dominated by largely endemic fishes of the suborder Notothenioidei. As a rare example of a marine adaptive radiation and characterized by specific adaptations such as anti-freeze glycoproteins, they have attracted interest of evolutionary and developmental biologists alike. The Antarctic continent and its surrounding Southern Ocean are differently affected by global change with the Antarctic Peninsula being one of the fastest warming regions on Earth. Additionally, several notothenioid species are targeted by commercial fisheries. Rapid environmental change and high fishing pressure may threaten the survival of Antarctic fish and/or reduce their intraspecific genetic diversity. Developing genomic resources, such as single nucleotide polymorphisms (SNPs), can help identifying sub-populations and their evolutionary potential, thus enabling modeling forecasts and the determination of prime targets for conservation measures.

A variety of restriction-site associated DNA (RAD) sequencing approaches has recently been used widely to employ the power of next generation sequencing for non-model organisms. Here, we apply a double-digest RAD (ddRAD) sequencing protocol to Notothenia coriiceps, the first notothenioid with an assembled draft genome. We assess the accuracy of this approach by comparing empirical data to in silico predictions of the number of fragments and SNPs, which should be obtained. Different bioinformatical pipelines and the effect of various assembly and SNP calling parameters are evaluated. Finally, loci that passed strict quality filtering are used to assess genetic population structure of N. coriiceps on different spatial scales. Results are expected to facilitate further ddRAD studies of Antarctic fish that can provide insight into ecological, evolutionary, and conservation-related questions.

Utilising Seaweed-Microbial Interactions For The Optimisation Of Seaweed Aquaculture

Friederike Eimer, PhD Student, Dept Marine Sciences, University of Gothenburg
Supervisor: Gunilla Toth, Henrik Pavia

In order to cultivate any crop successfully, knowledge about optimum growing and reproductive conditions and a detailed understanding of its life history is essential. Red seaweeds of the genus Porphyra sp. (also kown as nori) are economically the most valuable seaweed in the world with annual revenue of 1,5 billion USD. Similarly, the green seaweed of the genus Ulva sp. is also harvested for human consumption but has also shown potential as a bioremediator. While both species have been cultivated extensively in Asia for many centuries, the rest of the world is just beginning to develop sustainable methodologies and the demand for seaweed products globally is expanding rapidly. Studies that look at optimal growing conditions and induction of reproduction are extensive, however, the majority of studies have focused on the effects of abiotic factors such as temperature, photoperiod, light intensity and wave action only. Studies of biotic interactions between seaweeds and their epiphytic microbes are rare and focus mainly on the pathogenic traits of microbes since disease outbreaks can be economically and ecologically devastating. In recent years, the focus has shifted towards the role microbes play in the completion of the hosts life cycle and results indicate that microbes associated with seaweeds produce growth promoting substances and morphogenesis inducing compounds. Additionally, Quorum-Sensing signaling compounds (AHLs), which are used by bacteria in order to communicate with each either, have gained special attention in algae research as spore release and spore settlement have been shown to be controlled by (AHLs). This PhD thesis is aimed at exploring the potential of introducing seaweed-bacterial interactions as part of cultivation optimization with a focus in spore release, growth and net settlement.

Publication bias in Climate-Change science?

Johan Hollander
Department of Biologi - Aquatic Ecology, Lund University, Sweden

Non-significant results are less likely to be reported by authors and, when submitted for per review, less likely to be published by journal editors. This phenomenon, known collectively as publication bias, is found in a variety of scientific disciplines and can erode public trust in the scientific method. Public trust in science is especially important for fields like climate change science, where scientific consensus can influence state policies on a global scale, including strategies for industrial and agricultural management and development. Here we used meta-analysis to test for biases in the statistical results of climate change articles, including 1,154 experimental results from 120 articles. Funnel plots revealed no evidence of non-significant results being underreported, even at low sample sizes. However, we discovered three other types of reporting bias relating to writing style and relative prestige of journals: First, the magnitude of statistical effects was significantly larger in the abstract than the main body of articles. Second, the difference in effect sizes in abstracts versus main body of articles was especially pronounced in journals with high impact factors. Finally, the number of published articles about climate change, and the magnitude of effect sizes therein, both increased within two years of the seminal report by the Intergovernmental Panel on Climate Change 2007. Although we found evidence of stylistic biases within many of these reports, results of the meta-analysis confirmed that our sample of 120 articles about climate change gave credence to both significant and non-significant results and was characteristic of unbiased publishing practices.

What explains rare and conspicuous colours in a snail? - A test of time-series data against models of drift, migration and selection

Kerstin Johannesson
Dept Marine Sciences, University of Gothenburg

It is intriguing that conspicuous colour morphs of a prey species may be maintained at low frequencies alongside cryptic morphs. Negative frequency dependent selection by predators using search images ("apostatic selection") is often suggested without rejecting alternative explanations. Using a maximum likelihood approach we fitted predictions from models of genetic drift, migration, constant selection, heterozygote advantage and negative frequency-dependent selection to time-series data of colour frequencies in isolated populations of a marine snail (Littorina saxatilis), re-established with perturbed colour morph frequencies and followed for >20 generations. Snails of conspicuous colours (white, red, banded) are naturally rare in the study area (usually less than 10%) but frequencies were manipulated to levels of ~50% (one colour per population) in 8 populations at the start of the experiment in 1992. In 2013 frequencies had declined to ~15-45%. Drift alone could not explain these changes. Migration could not be rejected in any population, but required rates much higher than those recorded. Directional selection was rejected in three populations in favour of balancing selection. Heterozygote advantage and negative frequency dependent selection could not be distinguished statistically, although overall the results favoured the latter. Populations varied idiosyncratically as mild or variable colour selection (3-11%) interacted with demographic stochasticity, and the overall conclusion was that multiple mechanisms may contribute to maintaining the polymorphisms.

Impact of Skeletonema marinoi on anaerobic nitrate metabolism in marine snow

by Anja Kamp*, Ronnie N. Glud, Laura A. Bristow, Bo Thamdrup, and Peter Stief

*AIAS, Aarhus Institute of Advanced Studies, Aarhus University, Denmark

Diatom-bacteria aggregates can form in masses in oceanic surface layers and sink out of the photic zone toward the sea floor. Most research on sinking diatom aggregates and “marine snow” in general focuses on the vertical transport of organic carbon. Here we address the use and transport of intracellular nitrate within sinking diatom-bacteria aggregates. Only recently, it was discovered that diatoms are able to use stored nitrate for dissimilation in anoxic habitats, more specifically for Dissimilatory Nitrate Reduction to Ammonium (DNRA). Hypoxic or anoxic microsites within diatom-bacteria aggregates might therefore be important “hotspots” for dissimilatory nitrate reduction, especially in aggregates sinking through oxygen-depleted water.

Here, we studied the aggregate-forming, nitrate-storing diatom Skeletonema marinoi with 15N-labeling experiments, both in axenic cultures and in laboratory-produced diatom-bacteria aggregates. Our results show that (a) axenic S. marinoi is able to perform DNRA under dark and anoxic conditions, (b) nitrate is stored intracellularly by aggregate-associated S. marinoi in concentrations > 50 mM, (c) intracellular nitrate is used up within 2-3 days after shifting diatom-bacteria aggregates to dark and anoxic conditions, and (d) part of the intracellular nitrate of S. marinoi becomes available to the bacterial community of diatom-bacteria aggregates (e.g. by cell leakage) as indicated by the conversion of intracellular nitrate not solely to ammonium, as found in axenic S. marinoi, but also to nitrite and dinitrogen. Thus, intracellular nitrate may fuel complex nitrogen transformations including loss of reactive nitrogen when diatom aggregates sink into oxygen-depleted waters.

Return to the sea: the genome of the seagrass Zostera marina

Jeanine L. Olsen* (University of Groningen, NL), Yves van de Peer (Ghent University, BE) and the 35 members of the Zostera marina consortium. j.l.olsen@rug.nl.

The genome of Z. marina, the first marine angiosperm to be fully sequenced (online Nature 27 Jan 16, hard copy 18 Feb 16) reveals unique insights into the genomic losses and gains involved in achieving the structural and physiological adaptations required for its fully marine lifestyle, arguably the most extreme habitat shift ever accomplished by flowering plants. Seagrasses returned to the sea some 100 mya and Zostera underwent a whole genome duplication near the Cretaceous-Tertiary boundary thus opening new ecological opportunities.
In returning to the sea, seagrasses found a vast new habitat free of terrestrial competitors, insect pests and drought stress; but they also had to adapt to high salinity, develop flexible leaves able to take up nutrients, and perform photosynthesis in the spectrally reduced light of the submarine environment.

Here we present a number of highlights including modified cell walls that are more algal-like than plant-like, loss of all stomata and all of the genes, reduced plant defense genes and volatiles, and modifications for underwater pollination involving exineless pollen.

The genome resource will advance a wide range of functional ecological studies from adaptation of marine ecosystems under climate warming, to unraveling the mechanisms of osmoregulation under high salinities that may further inform our understanding of the evolution of salt tolerance.

The Skeletonema marinoi genome project

Mats Töpel1,2, Sylvie VM Tesson2, Magnus Alm Rosenblad2, Tomas Larsson2, Susanna Gross2, Alvar Almstedt2, Kerstin Johannesson2, Anders Blomberg2, Anna Godhe2

1. Bioinformatics Infrastructure for Life Sciences, BILS (http://bils.se/)
2. Department of Marine Sciences, University of Gothenburg.

We have sequenced the genome and transcriptome of the centric diatom Skeletonema marinoi GUMACC st54, as part of the Linnaeus Centre for Marine Evolutionary Biology (CeMEB, http://cemeb.science.gu.se/). This work is part of the Infrastructure for Marine Genetic Model Organisms (IMAGO), aimed at developing new marine model systems and provide genomic tools to study costal marine ecosystems. Skeletonema marinoi is an abundant primary producer during spring blooms in the North Atlantic and a valuable food source for zooplankton. Its generation time is 24 hours, which makes it ideal for studies of phenotypic response to environmental cues. Benthic cells act as resting stages, with up to 50 000 cells per gram of sediment, and can survive for at least hundred years and thereby provide short-term evolutionary genomic archives in the sediment.
In this project we have used Illumina paired-end libraries of insert sizes 180, 300 and 650 bp, mate pair libraries of insert sizes 3Kb and 6Kb, together with SMRT technology long reads from Pacific Biosciences to assemble a first version draft genome, to be used for population genomics studies.


A universal mechanism generating clusters of differentiated loci during divergence-with-migration

Marina Rafajlović1,2

In collaboration with:
Anna Emanuelsson1, Kerstin Johannesson3,2, Roger K. Butlin4,2, Bernhard

1 Department of Physics, University of Gothenburg, Gothenburg, Sweden
2 The Linnaeus Centre for Marine Evolutionary Biology, University of Gothenburg, Gothenburg, Sweden
3 Department of Marine Sciences, University of Gothenburg, Tjärnö, Strömstad, Sweden
Department of Animal and Plant Sciences, University of Sheffield, Sheffield, UK

Genome-wide patterns of genetic divergence help to reveal mechanisms of adaptation under gene flow. Empirical data show that divergence is mostly concentrated in narrow genomic regions [1]. This pattern may arise because differentiated loci protect nearby mutations from gene flow [2], but recent theory suggests this mechanism is insufficient to explain the emergence of concentrated differentiation during biologically realistic timescales [3]. Critically, earlier theory neglects an inevitable consequence of genetic drift: stochastic loss of local genomic divergence. Here we demonstrate that the rate of stochastic loss of weak local differentiation increases with recombination distance to a strongly diverged locus and, above a critical recombination distance, local loss is faster than local ‘gain’ of new differentiation [4]. Under high migration and weak selection this critical recombination distance is much smaller than the total recombination distance of the genomic region under selection [4]. Consequently, divergence between populations increases by net gain of new differentiation within the critical recombination distance, resulting in tightly-linked clusters of divergence [4]. The mechanism responsible is the balance between stochastic loss and gain of weak local differentiation, a mechanism acting universally throughout the genome [4]. Our results will help to explain empirical observations and lead to novel predictions regarding changes in genomic architectures during adaptive divergence.

[1] O. Seehausen et al. Genomics and the origin of species. Nature Reviews Genetics 15: 176-192, 2014.
[2] J. L. Feder, R. Gejii, S. Yeaman and P. Nosil. Establishment of new mutations under divergence and genome hitchhiking. Phil. Trans. R. Soc. B., 367: 461-474, 2012.
[3] S. Yeaman. Genomic rearrangements and the evolution of clusters of locally adaptive loci. PNAS, 110: E1743-E1751, 2013.
[4] M. Rafajlović, A. Emanuelsson, K. Johannesson, R. K. Butlin and B. Mehlig. A universal mechanism generating clusters of differentiated loci during divergence-with-migration. Under review.

Ancestral polymorphism or recent gene flow? Insight from comparative genome scans in allopatry and sympatry

Mark Ravinet1,2, Kohta Yoshida1 & Jun Kitano1

1Centre for Ecological and Evolutionary Synthesis, University of Oslo, Norway
2National Institute of Genetics, Mishima, Japan

How informative are genome scans for understanding speciation? Peaks and troughs of differentiation metrics have become a central focus in speciation studies yet relatively few attempts have been made to address the difficulties of interpreting these patterns. For example, does low divergence reflect gene flow or shared ancestral polymorphism?

One way to deal with this is to investigate systems where it is possible to compare populations in sympatry and allopatry. If low divergence is due to shared ancestral polymorphism, we would expect no difference in differentiation between comparisons. We therefore focused on the recent speciation between two highly divergent three-spined stickleback species in the Japan Sea (Gasterosteus nipponicus) and the Pacific Ocean (Gasterosteus aculeatus).

We used 26 resequenced whole genomes from sympatric and allopatric populations across the distributions of both species. Our spatially distributed sampling design allows us to identify introgression (lower differentiation in sympatry relative to allopatry) as well as candidate loci underlying reproductive isolation during divergence (divergence common to allopatry) and on secondary contact (divergence found only in sympatry). We suggest that careful consideration of study design can improve the interpretation of genome scan approaches.

Calcium transport in the mantle tissue of Pacific oyster, Crassostrea gigas

Sillanpää, J.K., Salman, Z., Sundell, K.

Fish Endocrinology Laboratory, Department of Biology and Environmental Sciences, University of Gothenburg, PO Box 463, S-405 30, Gothenburg, Sweden
E-mail: kirsikka.sillanpaa@bioenv.gu.se

Calcium for shell building in marine bivalves needs to be taken up from the environment and transported to the site of calcification via the hemolymph. It further needs to be transported through the mantle tissue, which surrounds the body of the animal and connects the soft tissues to the shell. The last passage of the calcium to reach the shell occurs across the outer mantle epithelium (OME). This epithelial cell layer is separating the circulatory system from the extrapallial space, i.e. the space between the mantle and the shell. The suggested calcium transport mechanisms through the OME include both passive and active transports, where the calcium is transported either in the ionic form or as a ligand bound to inorganic or organic compounds. Calcium can also be transported via the cells e.g. as vesicle-mediated transport. Active transport of the ionized calcium across the OME is probably maintained through multiple Ca2+ transport systems, such as Ca2+ ATPases (PMCA), Na+/Ca2+ exchangers and/or Ca channels.

The OME of Crassostrea gigas was prepared by peeling off the inner mantle epithelium and the connective tissues. The OME was thereafter mounted into Ussing chamber and analysed using a 4 chamber Ussing chamber set-up. Radiolabeled calcium was used to trace the transport of calcium through the OME. Commonly used, but less selective ATPase inhibitor, vanadate, and a newly developed, specific PMCA inhibitor, caloxin, were used to test the presence of PMCA like transporters in the OME. A decrease in Ca2+ transport was induced by vanadate, whereas caloxin instead increased the Ca2+ transport. These results indicate that although PMCA-like proteins might be present, their structure, and potential function, differs from mammalian models. Additionally, other transport mechanisms are likely working alongside PMCA-like proteins. We will continue this work to elucidate the transporters present and responsible for active Ca-transport across the OME of C. gigas by using combinations of multiple inhibitors together with concentration gradients of calcium.

Page Manager: Eva Marie Rödström|Last update: 4/22/2016

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