ALBERT

Description: Looking beyond stratification: a model-based analysis of the biological drivers of oxygen depletion in the North Sea Biogeosciences Discussions, 12, 12543-12610, 2015 Author(s): F. Große, N. Greenwood, M. Kreus, H. J. Lenhart, D. Machoczek, J. Pätsch, L. A. Salt, and H. Thomas The problem of low oxygen conditions, often referred to as hypoxia, occurs regularly in the North Sea, a temperate European shelf sea. Stratification represents a major process regulating the seasonal dynamics of bottom oxygen. However, lowest oxygen conditions in the North Sea do not occur in the regions of strongest stratification. This suggests that stratification is an important prerequisite for hypoxia, but that the complex interaction between hydrodynamics and the biological processes drives its development. In this study we use the ecosystem model HAMSOM-ECOHAM5 to provide a general characteristic of the different North Sea oxygen regimes, and to quantify the impact of the different physical and biological factors driving the oxygen dynamics below the thermocline and in the bottom layer. We show that the North Sea can be subdivided into three different regimes in terms of oxygen dynamics: (1) a highly productive, non-stratified coastal regime, (2) a productive, seasonally stratified regime with a small sub-thermocline volume, and (3) a productive, seasonally stratified regime with a large sub-thermocline volume, with regime 2 being highly susceptible to hypoxic conditions. Our analysis of the different processes driving the oxygen development reveals that inter-annual variations in the oxygen conditions are caused by variations in primary production, while spatial differences can be attributed to differences in stratification and water depth. In addition, we show that benthic bacteria represent the main oxygen consumers in the bottom layer, consistently accounting for more than 50 % of the overall consumption. By providing these valuable insights, we show that ecosystem models can be a useful tool for the interpretation of observations and the estimation of the impact of anthropogenic drivers on the North Sea oxygen conditions.

Description: Background: Spounavirinae viruses have received an increasing interest as tools for the control of harmful bacteria due to their relatively broad host range and strictly virulent phenotype. Results: In this study, we collected and analyzed the complete genome sequences of 61 published phages, either ICTV-classified or candidate members of the Spounavirinae subfamily of the Myoviridae. A set of comparative analyses identified a distinct, recently proposed Bastille-like phage group within the Spounavirinae. More importantly, type 1 thymidylate synthase (TS1) and dihydrofolate reductase (DHFR) genes were shown to be unique for the members of the proposed Bastille-like phage group, and are suitable as molecular markers. We also show that the members of this group encode beta-lactamase and/or sporulation-related SpoIIIE homologs, possibly questioning their suitability as biocontrol agents. Conclusions: We confirm the creation of a new genus—the “Bastille-like group”—in Spounavirinae, and propose that the presence of TS1- and DHFR-encoding genes could serve as signatures for the new Bastille-like group. In addition, the presence of metallo-beta-lactamase and/or SpoIIIE homologs in all members of Bastille-like group phages makes questionable their suitability for use in biocontrol.

Description: Background: The transcription factor p63 belongs to the p53/p63/p73 family and plays key functional roles during normal epithelial development and differentiation and in pathological states such as squamous cell carcinomas. The human TP63 gene, located on chromosome 3q28 is driven by two promoters that generate the full-length transactivating (TA) and N-terminal truncated (ΔN) isoforms. Furthermore alternative splicing at the C-terminus gives rise to additional α, β, γ and likely several other minor variants. Teasing out the expression and biological function of each p63 variant has been both the focus of, and a cause for contention in the p63 field. Results: Here we have taken advantage of a burgeoning RNA-Seq based genomic data-sets to examine the global expression profiles of p63 isoforms across commonly utilized human cell-lines and major tissues and organs. Consistent with earlier studies, we find ΔNp63 transcripts, primarily that of the ΔNp63α isoforms, to be expressed in most cells of epithelial origin such as those of skin and oral tissues, mammary glands and squamous cell carcinomas. In contrast, TAp63 is not expressed in the majority of normal cell-types and tissues; rather it is selectively expressed at moderate to high levels in a subset of Burkitt’s and diffuse large B-cell lymphoma cell lines. We verify this differential expression pattern of p63 isoforms by Western blot analysis, using newly developed ΔN and TA specific antibodies. Furthermore using unsupervised clustering of human cell lines, tissues and organs, we show that ΔNp63 and TAp63 driven transcriptional networks involve very distinct sets of molecular players, which may underlie their different biological functions. Conclusions: In this study we report comprehensive and global expression profiles of p63 isoforms and their relationship to p53/p73 and other potential transcriptional co-regulators. We curate publicly available data generated in part by consortiums such as ENCODE, FANTOM and Human Protein Atlas to delineate the vastly different transcriptomic landscapes of ΔNp63 and TAp63. Our studies help not only in dispelling prevailing myths and controversies on p63 expression in commonly used human cell lines but also augur new isoform- and cell type-specific activities of p63.

Description: Background: The genus Vulpes (true foxes) comprises numerous species that inhabit a wide range of habitats and climatic conditions, including one species, the Arctic fox (Vulpes lagopus) which is adapted to the arctic region. A close relative to the Arctic fox, the red fox (Vulpes vulpes), occurs in subarctic to subtropical habitats. To study the genetic basis of their adaptations to different environments, transcriptome sequences from two Arctic foxes and one red fox individual were generated and analyzed for signatures of positive selection. In addition, the data allowed for a phylogenetic analysis and divergence time estimate between the two fox species. Results: The de novo assembly of reads resulted in more than 160,000 contigs/transcripts per individual. Approximately 17,000 homologous genes were identified using human and the non-redundant databases. Positive selection analyses revealed several genes involved in various metabolic and molecular processes such as energy metabolism, cardiac gene regulation, apoptosis and blood coagulation to be under positive selection in foxes. Branch site tests identified four genes to be under positive selection in the Arctic fox transcriptome, two of which are fat metabolism genes. In the red fox transcriptome eight genes are under positive selection, including molecular process genes, notably genes involved in ATP metabolism. Analysis of the three transcriptomes and five Sanger re-sequenced genes in additional individuals identified a lower genetic variability within Arctic foxes compared to red foxes, which is consistent with distribution range differences and demographic responses to past climatic fluctuations. A phylogenomic analysis estimated that the Arctic and red fox lineages diverged about three million years ago. Conclusions: Transcriptome data are an economic way to generate genomic resources for evolutionary studies. Despite not representing an entire genome, this transcriptome analysis identified numerous genes that are relevant to arctic adaptation in foxes. Similar to polar bears, fat metabolism seems to play a central role in adaptation of Arctic foxes to the cold climate, as has been identified in the polar bear, another arctic specialist.

Description: Background: Insects belong to a class that accounts for the majority of animals on earth. With over one million identified species, insects display a huge diversity and occupy extreme environments. At present, there are dozens of fully sequenced insect genomes that cover a range of habitats, social behavior and morphologies. In view of such diverse collection of genomes, revealing evolutionary trends and charting functional relationships of proteins remain challenging. Results: We analyzed the relatedness of 17 complete proteomes representative of proteomes from insects including louse, bee, beetle, ants, flies and mosquitoes, as well as an out-group from the crustaceans. The analyzed proteomes mostly represented the orders of Hymenoptera and Diptera. The 287,405 protein sequences from the 18 proteomes were automatically clustered into 20,933 families, including 799 singletons. A comprehensive analysis based on statistical considerations identified the families that were significantly expanded or reduced in any of the studied organisms. Among all the tested species, ants are characterized by an exceptionally high rate of family gain and loss. By assigning annotations to hundreds of species-specific families, the functional diversity among species and between the major clades (Diptera and Hymenoptera) is revealed. We found that many species-specific families are associated with receptor signaling, stress-related functions and proteases. The highest variability among insects associates with the function of transposition and nucleic acids processes (collectively coined TNAP). Specifically, the wasp and ants have an order of magnitude more TNAP families and proteins relative to species that belong to Diptera (mosquitoes and flies). Conclusions: An unsupervised clustering methodology combined with a comparative functional analysis unveiled proteomic signatures in the major clades of winged insects. We propose that the expansion of TNAP families in Hymenoptera potentially contributes to the accelerated genome dynamics that characterize the wasp and ants.

Description: Optimizing the impact of temperature on bio-hydrogen production from food waste and its derivatives under no pH control using statistical modelling Biogeosciences Discussions, 12, 12823-12850, 2015 Author(s): A. Sattar, C. Arslan, C. Ji, S. Sattar, K. Yousaf, and S. Hashim The effect of temperature on bio-hydrogen production by co-digestion of sewerage sludge with food waste and its two derivatives, i.e. noodle waste and rice waste, was investigated by statistical modelling. Experimental results showed that increasing temperature from mesophilic (37 °C) to thermophilic (55 °C) was an effective mean for increasing bio-hydrogen production from food waste and noodle waste, but it caused a negative impact on bio-hydrogen production from rice waste. The maximum cumulative bio-hydrogen production of 650 mL was obtained from noodle waste under mesophilic temperature condition. Most of the production was observed during 48 h of incubation that continued till 72 h of incubation, and a decline in pH during this interval was 4.3 and 4.4 from a starting value of 7 under mesophilic and thermophilic conditions, respectively. Most of glucose consumption was also observed during 72 h of incubation and the maximum consumption was observed during the first 24 h, which was the same duration where the maximum pH drop occurred. The maximum hydrogen yields of 82.47 mL VS −1 , 131.38 mL COD −1 , and 44.90 mL glucose −1 were obtained from mesophilic food waste, thermophilic noodle waste and mesophilic rice waste respectively. The production of volatile fatty acids increased with an increase in time and temperature from food waste and noodle waste reactors whereas it decreased with temperature in rice waste reactors. The statistical modelling returned good results with high values of coefficient of determination ( R 2 ) for each waste type when it was opted for the study of cumulative hydrogen production, glucose consumption and volatile fatty acid production. The 3-D response surface plots developed by the statistical models helped a lot in developing better understanding of the impact of temperature and incubation time.

Description: Stable isotope study of a new chondrichthyan fauna (Kimmeridgian, Porrentruy, Swiss Jura): an unusual freshwater-influenced isotopic composition for the hybodont shark Asteracanthus Biogeosciences Discussions, 12, 12899-12921, 2015 Author(s): L. Leuzinger, L. Kocsis, J.-P. Billon-Bruyat, S. Spezzaferri, and T. Vennemann Chondrichthyan teeth (sharks, rays and chimaeras) are mineralised in isotopic equilibrium with the surrounding water, and parameters such as water temperature and salinity can be inferred from the oxygen isotopic composition (δ 18 O p ) of their bioapatite. We analysed a new chondrichthyan assemblage, as well as teeth from bony fish (Pycnodontiformes). All specimens are from Kimmeridgian coastal marine deposits of the Swiss Jura (vicinity of Porrentruy, Ajoie district, NW Switzerland). While the overall faunal composition and the isotopic composition of bony fish are consistent with marine conditions, unusually low δ 18 O p values were measured for the hybodont shark Asteracanthus . These values are also lower compared to previously published data from older European Jurassic localities. Additional analyses on material from Solothurn (Kimmeridgian, NW Switzerland) also have comparable, low- 18 O isotopic compositions for Asteracanthus . The data are hence interpreted to represent a so far unique, freshwater-influenced isotopic composition for this shark that is classically considered as a marine genus. While reproduction in freshwater or brackish realms is established for other hybodonts, a similar behaviour for Asteracanthus is proposed here. Regular excursions into lower salinity waters can be linked to the age of the deposits and correspond to an ecological adaptation, most likely driven by the Kimmeridgian transgression and by the competition of the primitive shark Asteracanthus with the rapidly diversifying neoselachians (modern sharks).

Description: Climate change impacts on net primary production (NPP) and export production (EP) regulated by increasing stratification and phytoplankton community structure in CMIP5 models Biogeosciences Discussions, 12, 12851-12897, 2015 Author(s): W. Fu, J. Randerson, and J. K. Moore We examine climate change impacts on net primary production (NPP) and export production (sinking particulate flux; EP) with simulations from nine Earth System Models (ESMs) performed in the framework of the fifth Coupled Model Inter-comparison Project (CMIP5). Global NPP and EP are reduced considerably by the end of the century for the intense warming scenario of Representative Concentration Pathway (RCP) 8.5. Relative to the 1990s, global NPP in the 2090s is reduced by 2.3–16 % and EP by 7–18 %. The models with the largest increases in stratification (and largest relative reductions in NPP and EP) also show the largest positive biases in stratification for the contemporary period, suggesting some potential overestimation of climate impacts on NPP and EP. All of the CMIP5 models show an increase in stratification in response to surface ocean warming and freshening that is accompanied by decreases in NPP, EP, and surface macronutrient concentrations. There is considerable variability across models in the absolute magnitude of these fluxes, surface nutrient concentrations, and their perturbations by climate change, indicating large model uncertainties. The negative response of NPP and EP to stratification increases reflects a bottom-up control, as nutrient flux to the euphotic zone declines. Models with dynamic phytoplankton community structure show larger declines in EP than in NPP. This is driven by phytoplankton community composition shifts, with a reduced percentage of NPP by large phytoplankton under RCP 8.5, as smaller phytoplankton are favored under the increasing nutrient stress. Thus, projections of the NPP response to climate change in the CMIP5 models are critically dependent on the simulated phytoplankton community structure, the efficiency of the biological pump, and the resulting (highly variable) levels of regenerated production. Community composition is represented relatively simply in the CMIP5 models, and should be expanded to better capture the spatial patterns and the changes in export efficiency that are necessary for predicting climate impacts on NPP.

Description: Technical Note: A simple calculation algorithm to separate high-resolution CH 4 flux measurements into ebullition and diffusion-derived components Biogeosciences Discussions, 12, 12923-12945, 2015 Author(s): M. Hoffmann, M. Schulz-Hanke, J. Garcia Alba, N. Jurisch, U. Hagemann, T. Sachs, M. Sommer, and J. Augustin Processes driving the production, transformation and transport of methane (CH 4 ) in wetland ecosystems are highly complex. Thus, serious challenges are constitutes in terms of the mechanistic process understanding, the identification of potential environmental drivers and the calculation of reliable CH 4 emission estimates. We present a simple calculation algorithm to separate open-water CH 4 fluxes measured with automatic chambers into diffusion- and ebullition-derived components, which helps facilitating the identification of underlying dynamics and potential environmental drivers. Flux separation is based on ebullition related sudden concentration changes during single measurements. A variable ebullition filter is applied, using the lower and upper quartile and the interquartile range (IQR). Automation of data processing is achieved by using an established R-script, adjusted for the purpose of CH 4 flux calculation. The algorithm was tested using flux measurement data (July to September 2013) from a former fen grassland site, converted into a shallow lake as a result of rewetting ebullition and diffusion contributed 46 and 55 %, respectively, to total CH 4 emissions, which is comparable to those previously reported by literature. Moreover, the separation algorithm revealed a concealed shift in the diurnal trend of diffusive fluxes throughout the measurement period.

Description: Background: Prokaryotic translation initiation involves the proper docking, anchoring, and accommodation of mRNA to the 30S ribosomal subunit. Three initiation factors (IF1, IF2, and IF3) and some ribosomal proteins mediate the assembly and activation of the translation initiation complex. Although the interaction between Shine-Dalgarno (SD) sequence and its complementary sequence in the 16S rRNA is important in initiation, some genes lacking an SD ribosome binding site (RBS) are still well expressed. The objective of this study is to examine the pattern of distribution and diversity of RBS in fully sequenced bacterial genomes. The following three hypotheses were tested: SD motifs are prevalent in bacterial genomes; all previously identified SD motifs are uniformly distributed across prokaryotes; and genes with specific cluster of orthologous gene (COG) functions differ in their use of SD motifs. Results: Data for 2,458 bacterial genomes, previously generated by Prodigal (PROkaryotic DYnamic programming Gene-finding ALgorithm) and currently available at the National Center for Biotechnology Information (NCBI), were analyzed. Of the total genes examined, ~77.0 % use an SD RBS, while ~23.0 % have no RBS. Majority of the genes with the most common SD motifs are distributed in a manner that is representative of their abundance for each COG functional category, while motifs 13 (5′-GGA-3′/5′-GAG-3′/5′-AGG-3′) and 27 (5′-AGGAGG-3′) appear to be predominantly used by genes for information storage and processing, and translation and ribosome biogenesis, respectively. Conclusion: These findings suggest that an SD sequence is not obligatory for translation initiation; instead, other signals, such as the RBS spacer, may have an overarching influence on translation of mRNAs. Subsequent analyses of the 5′ secondary structure of these mRNAs may provide further insight into the translation initiation mechanism.

Description: Background: Acidity is a major contributor to fruit quality. Several organic acids are present in apple fruit, but malic acid is predominant and determines fruit acidity. The trait is largely controlled by the Malic acid (Ma) locus, underpinning which Ma1 that putatively encodes a vacuolar aluminum-activated malate transporter1 (ALMT1)-like protein is a strong candidate gene. We hypothesize that fruit acidity is governed by a gene network in which Ma1 is key member. The goal of this study is to identify the gene network and the potential mechanisms through which the network operates. Results: Guided by Ma1, we analyzed the transcriptomes of mature fruit of contrasting acidity from six apple accessions of genotype Ma_ (MaMa or Mama) and four of mama using RNA-seq and identified 1301 fruit acidity associated genes, among which 18 were most significant acidity genes (MSAGs). Network inferring using weighted gene co-expression network analysis (WGCNA) revealed five co-expression gene network modules of significant (P 

Description: Nitrogen export from a boreal stream network following forest harvesting: seasonal nitrate removal and conservative export of organic forms Biogeosciences Discussions, 12, 12061-12089, 2015 Author(s): J. Schelker, R. Sponseller, E. Ring, L. Högbom, S. Löfgren, and H. Laudon Boreal streams are under pressure from large scale disturbance by forestry. Recent scenarios predict an increase in forest production in Scandinavia to meet market demands and to mitigate higher anthropogenic CO 2 emissions. Increased fertilization and shorter forest rotations are anticipated which will likely enhance the pressure on boreal streams in the near future. Among the major environmental impacts of forest harvesting is the increased mobilization of inorganic nitrogen (N), primarily as nitrate (NO 3 - ) into surface waters. But whereas NO 3 - inputs to first-order streams have been previously described, their downstream fate and impact is not well understood. We evaluated the downstream fate of N inputs in a boreal landscape that has been altered by forest harvests over a 10 year period to estimate the effects of multiple clear-cuts on aquatic N export in a boreal stream network. Small streams showed substantial leaching of NO 3 - in response to harvests with concentrations increasing by ~ 15 fold. NO 3 - concentrations at two sampling stations further downstream in the network were strongly seasonal and increased significantly in response to harvesting at the medium size, but not at the larger stream. Nitrate removal efficiency, E r , calculated as the percentage of "forestry derived" NO 3 - that was retained within the landscape using a mass balance model was highest during the snow melt season followed by the growing season, but declined continuously throughout the dormant season. In contrast, export of organic N from the landscape indicated little removal and was essentially conservative. Overall, net removal of NO 3 - between 2008 and 2011 accounted for ~ 70 % of the total NO 3 - mass exported from harvested patches distributed across the landscape. These results highlight the capacity and limitation of N-limited terrestrial and aquatic ecosystems to buffer inorganic N mobilization that arises from multiple clear-cuts within meso-scale boreal watersheds.

Description: No description available

Description: Background: Dickeya zeae is a bacterial species that infects monocotyledons and dicotyledons. Two antibiotic-like phytotoxins named zeamine and zeamine II were reported to play an important role in rice seed germination, and two genes associated with zeamines production, i.e., zmsA and zmsK, have been thoroughly characterized. However, other virulence factors and its molecular mechanisms of host specificity and pathogenesis are hardly known. Results: The complete genome of D. zeae strain EC1 isolated from diseased rice plants was sequenced, annotated, and compared with the genomes of other Dickeya spp.. The pathogen contains a chromosome of 4,532,364 bp with 4,154 predicted protein-coding genes. Comparative genomics analysis indicates that D. zeae EC1 is most co-linear with D. chrysanthemi Ech1591, most conserved with D. zeae Ech586 and least similar to D. paradisiaca Ech703. Substantial genomic rearrangement was revealed by comparing EC1 with Ech586 and Ech703. Most virulence genes were well-conserved in Dickeya strains except Ech703. Significantly, the zms gene cluster involved in biosynthesis of zeamines, which were shown previously as key virulence determinants, is present in D. zeae strains isolated from rice, and some D. solani strains, but absent in other Dickeya species and the D. zeae strains isolated from other plants or sources. In addition, a DNA fragment containing 9 genes associated with fatty acid biosynthesis was found inserted in the fli gene cluster encoding flagellar biosynthesis of strain EC1 and other two rice isolates but not in other strains. This gene cluster shares a high protein similarity to the fatty acid genes from Pantoea ananatis.ConlusionOur findings delineate the genetic background of D. zeae EC1, which infects both dicotyledons and monocotyledons, and suggest that D. zeae strains isolated from rice could be grouped into a distinct pathovar, i.e., D. zeae subsp. oryzae. In addition, the results of this study also unveiled that the zms gene cluster presented in the genomes of D. zeae rice isolates and D. solani strains, and the fatty acid genes inserted in the fli gene cluster of strain EC1 were likely derived from horizontal gene transfer during later stage of bacterial evolution.

Description: Background: To promote the clinical application of next-generation sequencing, it is important to obtain accurate and consistent variants of target genomic regions at low cost. Ion Proton, the latest updated semiconductor-based sequencing instrument from Life Technologies, is designed to provide investigators with an inexpensive platform for human whole exome sequencing that achieves a rapid turnaround time. However, few studies have comprehensively compared and evaluated the accuracy of variant calling between Ion Proton and Illumina sequencing platforms such as HiSeq 2000, which is the most popular sequencing platform for the human genome. The Ion Proton sequencer combined with the Ion TargetSeq™ Exome Enrichment Kit together make up TargetSeq-Proton, whereas SureSelect-Hiseq is based on the Agilent SureSelect Human All Exon v4 Kit and the HiSeq 2000 sequencer. Results: Here, we sequenced exonic DNA from four human blood samples using both TargetSeq-Proton and SureSelect-HiSeq. We then called variants in the exonic regions that overlapped between the two exome capture kits (33.6 Mb). The rates of shared variant loci called by two sequencing platforms were from 68.0 to 75.3 % in four samples, whereas the concordance of co-detected variant loci reached 99 %. Sanger sequencing validation revealed that the validated rate of concordant single nucleotide polymorphisms (SNPs) (91.5 %) was higher than the SNPs specific to TargetSeq-Proton (60.0 %) or specific to SureSelect-HiSeq (88.3 %). With regard to 1-bp small insertions and deletions (InDels), the Sanger sequencing validated rates of concordant variants (100.0 %) and SureSelect-HiSeq-specific (89.6 %) were higher than those of TargetSeq-Proton-specific (15.8 %). Conclusions: In the sequencing of exonic regions, a combination of using of two sequencing strategies (SureSelect-HiSeq and TargetSeq-Proton) increased the variant calling specificity for concordant variant loci and the sensitivity for variant loci called by any one platform. However, for the sequencing of platform-specific variants, the accuracy of variant calling by HiSeq 2000 was higher than that of Ion Proton, specifically for the InDel detection. Moreover, the variant calling software also influences the detection of SNPs and, specifically, InDels in Ion Proton exome sequencing.

Description: Hydroxy fatty acids in fresh snow samples from northern Japan: long-range atmospheric transport of Gram-negative bacteria by Asian winter monsoon Biogeosciences Discussions, 12, 13375-13397, 2015 Author(s): P. Tyagi, S. Yamamoto, and K. Kawamura Hydroxy fatty acids (FAs) in fresh snow from Sapporo, one of the heaviest snowfall regions in the world, have been studied to ascertain the airborne bacterial endotoxin concentrations and their biomass. The presence of β-hydroxy FAs (C 9 –C 28 ), constituents of Gram-negative bacteria (GNB), suggests long-range transport of soil microbes. Likewise, the occurrence of α- and ω-hydroxy FAs (C 9 –C 30 and C 9 –C 28 , respectively) in snow reveals their contribution from epicuticular waxes and soil microorganisms. Estimated endotoxin and GNB mass can aid in assessing their possible impacts on the diversity and functioning of aquatic and terrestrial ecosystems, as well as lethal effects on pedestrians through dispersal of microbes. Air mass back trajectories together with hydroxy FAs unveil their sources from Siberia, Russian Far East and North China by the Asian monsoon. This study highlights the role of fresh snow that reduces the human health risk of GNB and endotoxin by scavenging from the air.

Description: Background: Adaptive manipulation of animal behavior by parasites functions to increase parasite transmission through changes in host behavior. These changes can range from slight alterations in existing behaviors of the host to the establishment of wholly novel behaviors. The biting behavior observed in Carpenter ants infected by the specialized fungus Ophiocordyceps unilateralis s.l. is an example of the latter. Though parasitic manipulation of host behavior is generally assumed to be due to the parasite’s gene expression, few studies have set out to test this. Results: We experimentally infected Carpenter ants to collect tissue from both parasite and host during the time period when manipulated biting behavior is experienced. Upon observation of synchronized biting, samples were collected and subjected to mixed RNA-Seq analysis. We also sequenced and annotated the O. unilateralis s.l. genome as a reference for the fungal sequencing reads. Conclusions: Our mixed transcriptomics approach, together with a comparative genomics study, shows that the majority of the fungal genes that are up-regulated during manipulated biting behavior are unique to the O. unilateralis s.l. genome. This study furthermore reveals that the fungal parasite might be regulating immune- and neuronal stress responses in the host during manipulated biting, as well as impairing its chemosensory communication and causing apoptosis. Moreover, we found genes up-regulated during manipulation that putatively encode for proteins with reported effects on behavioral outputs, proteins involved in various neuropathologies and proteins involved in the biosynthesis of secondary metabolites such as alkaloids.

Description: Nitrogen cycling in the subsurface biosphere: nitrate isotopes in porewaters underlying the oligotrophic North Atlantic Biogeosciences Discussions, 12, 13545-13591, 2015 Author(s): S. D. Wankel, C. Buchwald, W. Ziebis, C. B. Wenk, and M. F. Lehmann Nitrogen (N) is a key component of fundamental biomolecules. Hence, the cycling and availability of N is a central factor governing the extent of ecosystems across the Earth. In the organic-lean sediment porewaters underlying the oligotrophic ocean, where low levels of microbial activity persist despite limited organic matter delivery from overlying water, the extent and modes of nitrogen transformations have not been widely investigated. Here we use the N and oxygen (O) isotopic composition of porewater nitrate (NO 3 − ) from a site in the oligotrophic North Atlantic (IODP) to determine the extent and magnitude of microbial nitrate production (via nitrification) and consumption (via denitrification). We find that NO 3 − accumulates far above bottom seawater concentrations (∼ 21 μM) throughout the sediment column (up to ∼ 50 μM) down to the oceanic basement as deep as 90 mbsf, reflecting the predominance of aerobic nitrification/remineralization within the deep marine sediments. Large changes in the δ 15 N and δ 18 O of nitrate, however, reveal variable influence of nitrate respiration across the three sites. We use an inverse porewater diffusion–reaction model, constrained by the N and O isotope systematics of nitrification and denitrification and the porewater NO 3 − isotopic composition, to estimate rates of nitrification and denitrification throughout the sediment column. Results indicate variability of reaction rates across and within the three boreholes that are generally consistent with the differential distribution of dissolved oxygen at this site, though not necessarily with the canonical view of how redox thresholds separate nitrate regeneration from dissimilative consumption spatially. That is, we provide isotope evidence for expanded zones of co-ocurring nitrification and denitrification. The isotope biogeochemical modeling also yielded estimates for the δ 15 N and δ 18 O of newly produced nitrate (δ 15 N NTR and δ 18 O NTR ), as well as the isotope effect for denitrification ( 15 ϵ DNF ), parameters with high relevance to global ocean models of N cycling. Estimated values of δ 15 N NTR were generally lower than previously reported δ 15 N values for sinking PON in this region. We suggest that these values can be related to sedimentary N-fixation and remineralization of the newly fixed organic N. Values of δ 18 O NTR generally ranged between −2.8 and 0.0 ‰, consistent with recent estimates based on lab cultures of nitrifying bacteria. Notably, some δ 18 O NTR values were elevated, suggesting incorporation of 18 O-enriched dissolved oxygen during nitrification, and possibly indicating a tight coupling of NH 4 + and NO 2 − oxidation in this metabolically sluggish environment. Our findings indicate that the production of organic matter by in situ autotrophy (e.g., nitrification, nitrogen fixation) supply a large fraction of the biomass and organic substrate for heterotrophy in these sediments, supplementing the small organic matter pool derived from the overlying euphotic zone. This work sheds new light on an active nitrogen cycle operating, despite exceedingly low carbon inputs, in the deep sedimentary biosphere.

Description: Spring bloom onset in the Nordic Seas Biogeosciences Discussions, 12, 13631-13673, 2015 Author(s): A. Mignot, R. Ferrari, and K. A. Mork The North Atlantic spring bloom is a massive annual growth event of marine phytoplankton, tiny free-floating algae that form the base of the ocean's food web and generates a large fraction of the global primary production of organic matter. The conditions that trigger the onset of the spring bloom in the Nordic Seas, at the northern edge of the North Atlantic, are studied using in-situ data from five bio-optical floats released above the Arctic Circle. It is often assumed that spring blooms start as soon as phytoplankton cells daily irradiance is sufficiently abundant that division rates exceed losses. The bio-optical float data instead suggest the tantalizing hypothesis that Nordic Seas blooms start when the photoperiod, the number of daily light hours experienced by phytoplankton, exceeds a critical value, independently of division rates. This bloom behavior may be explained by realizing that photosynthesis is impossible during polar nights and phytoplankton enters in a dormant stage in winter, only to be awaken by a photoperiodic trigger. While the first accumulation of biomass recorded by the bio-optical floats is consistent with the photoperiod hypothesis, it is possible that some biomass accumulation started before the critical photoperiod but at levels too low to be detected by the fluorometers. Thus more precise observations are needed to test the photoperiod hypothesis.

Description: Unusual biogenic calcite structures in two shallow lakes, James Ross Island, Antarctica Biogeosciences Discussions, 12, 13593-13629, 2015 Author(s): J. Elster, L. Nedbalová, R. Vodrážka, K. Láska, J. Haloda, and J. Komárek The floors of two shallow endorheic lakes, located on volcanic surfaces on James Ross Island, are covered with calcareous organosedimentary structures. Their biological and chemical composition, lake water characteristics, and seasonal variability of the thermal regime are introduced. The lakes are frozen down to the bottom eight-nine months per year and their water chemistry is characterized by low conductivity and neutral to slightly alkaline pH. The photosynthetic microbial mat is composed of filamentous cyanobacteria and microalgae that are considered to be Antarctic endemic species. The mucilaginous black biofilm is covered by green spots formed by a green microalga and the macroscopic structures are packed together with fine material. Thin sections consist of rock substrate, soft biofilm, calcite spicules and mineral grains originating from different sources. The morphology of the spicules is typical of calcium carbonate monocrystals having a layered structure and worn surface, which reflect growth and degradation processes. The spicules chemical composition and structure correspond to pure calcite. Lakes age, altitude, morphometry, geomorphological and hydrological stability, including low sedimentation rates, together with thermal regime predispose the existence of this community. We hypothesize that the precipitation of calcite is connected with the photosynthetic activity of the green microalgae that were not recorded in any other lake in the region. This study has shown that the unique community producing biogenic calcite spicules is quite different to any yet described.

Description: Background: In archaea and eukaryotes, ribonucleoprotein complexes containing small C/D box s(no)RNAs use base pair complementarity to target specific sites within ribosomal RNA for 2'-O-ribose methylation. These modifications aid in the folding and stabilization of nascent rRNA molecules and their assembly into ribosomal particles. The genomes of hyperthermophilic archaea encode large numbers of C/D box sRNA genes, suggesting an increased necessity for rRNA stabilization at extreme growth temperatures. Results: We have identified the complete sets of C/D box sRNAs from seven archaea using RNA-Seq methodology. In total, 489 C/D box sRNAs were identified, each containing two guide regions. A combination of computational and manual analyses predicts 719 guide interactions with 16S and 23S rRNA molecules. This first pan-archaeal description of guide sequences identifies (i) modified rRNA nucleotides that are frequently conserved between species and (ii) regions within rRNA that are hotspots for 2'-O-methylation. Gene duplication, rearrangement, mutational drift and convergent evolution of sRNA genes and guide sequences were observed. In addition, several C/D box sRNAs were identified that use their two guides to target locations distant in the rRNA sequence but close in the secondary and tertiary structure. We propose that they act as RNA chaperones and facilitate complex folding events between distant sequences. Conclusions: This pan-archaeal analysis of C/D box sRNA guide regions identified conserved patterns of rRNA 2'-O-methylation in archaea. The interaction between the sRNP complexes and the nascent rRNA facilitates proper folding and the methyl modifications stabilize higher order rRNA structure within the assembled ribosome.

Description: Background: The application of phages is a promising tool to reduce the number of Campylobacter along the food chain. Besides the efficacy against a broad range of strains, phages have to be safe in terms of their genomes. Thus far, no genes with pathogenic potential (e.g., genes encoding virulence factors) have been detected in Campylobacter phages. However, preliminary studies suggested that the genomes of group II phages may be diverse and prone to genomic rearrangements. Results: We determined and analysed the genomic sequence (182,761 bp) of group II phage CP21 that is closely related to the already characterized group II phages CP220 and CPt10. The genomes of these phages are comprised of four modules separated by very similar repeat regions, some of which harbouring open reading frames (ORFs). Though, the arrangement of the modules and the location of some ORFs on the genomes are different in CP21 and in CP220/CPt10. In this work, a PCR system was established to study the modular genome organization of other group II phages demonstrating that they belong to different subgroups of the CP220-like virus genus, the prototypes of which are CP21 and CP220. The subgroups revealed different restriction patterns and, interestingly enough, also distinct host specificities, tail fiber proteins and tRNA genes. We additionally analysed the genome of group II phage vB_CcoM-IBB_35 (IBB_35) for which to date only five individual contigs could be determined. We show that the contigs represent modules linked by long repeat regions enclosing some yet not identified ORFs (e.g., for a head completion protein). The data suggest that IBB_35 is a member of the CP220 subgroup. Conclusion: Campylobacter group II phages are diverse regarding their genome organization. Since all hitherto characterized group II phages contain numerous genes for transposases and homing endonucleases as well as similar repeat regions, it cannot be excluded that these phages are genetically unstable. To answer this question, further experiments and sequencing of more group II phages should be performed.

Description: Background: Mycoplasma pneumoniae is a common pathogen that causes upper and lower respiratory tract infections in people of all ages, responsible for up to 40 % of community-acquired pneumonias. It also causes a wide array of extrapulmonary infections and autoimmune phenomena. Phylogenetic studies of the organism have been generally restricted to specific genes or regions of the genome, because whole genome sequencing has been completed for only 4 strains. To better understand the physiology and pathogenicity of this important human pathogen, we performed comparative genomic analysis of 15 strains of M. pneumoniae that were isolated between the 1940s to 2009 from respiratory specimens and cerebrospinal fluid originating from the USA, China and England. Results: Illumina MiSeq whole genome sequencing was performed on the 15 strains and all genome sequences were completed. Results from the comparative genomic analysis indicate that although about 1500 SNP and indel variants exist between type1 and type 2 strains, there is an overall high degree of sequence similarity among the strains (〉99 % identical to each other). Within the two subtypes, conservation of most genes, including the CARDS toxin gene and arginine deiminase genes, was observed. The major variation occurs in the P1 and ORF6 genes associated with the adhesin complex. Multiple hsdS genes (encodes S subunit of type I restriction enzyme) with variable tandem repeat copy numbers were found in all 15 genomes. Conclusions: These data indicate that despite conclusions drawn from 16S rRNA sequences suggesting rapid evolution, the M. pneumoniae genome is extraordinarily stable over time and geographic distance across the globe with a striking lack of evidence of horizontal gene transfer.

Description: Background: Walnut (Juglans regia, 2n = 32, approximately 606 Mb per 1C genome) is an economically important tree crop. Resistance to anthracnose, caused by Colletotrichum gloeosporioides, is a major objective of walnut genetic improvement in China. The recently developed specific length amplified fragment sequencing (SLAF-seq) is an efficient strategy that can obtain large numbers of markers with sufficient sequence information to construct high-density genetic maps and permits detection of quantitative trait loci (QTLs) for molecular breeding. Results: SLAF-seq generated 161.64 M paired-end reads. 153,820 SLAF markers were obtained, of which 49,174 were polymorphic. 13,635 polymorphic markers were sorted into five segregation types and 2,577 markers of them were used to construct genetic linkage maps: 2,395 of these fell into 16 linkage groups (LGs) for the female map, 448 markers for the male map, and 2,577 markers for the integrated map. Taking into account the size of all LGs, the marker coverage was 2,664.36 cM for the female map, 1,305.58 cM for the male map, and 2,457.82 cM for the integrated map. The average intervals between two adjacent mapped markers were 1.11 cM, 2.91 cM and 0.95 cM for three maps, respectively. ‘SNP_only’ markers accounted for 89.25 % of the markers on the integrated map. Mapping markers contained 5,043 single nucleotide polymorphisms (SNPs) loci, which corresponded to two SNP loci per SLAF marker. According to the integrated map, we used interval mapping (Logarithm of odds, LOD 〉 3.0) to detect our quantitative trait. One QTL was detected for anthracnose resistance. The interval of this QTL ranged from 165.51 cM to 176.33 cM on LG14, and ten markers in this interval that were above the threshold value were considered to be linked markers to the anthracnose resistance trait. The phenotypic variance explained by each marker ranged from 16.2 to 19.9 %, and their LOD scores varied from 3.22 to 4.04. Conclusions: High-density genetic maps for walnut containing 16 LGs were constructed using the SLAF-seq method with an F1 population. One QTL for walnut anthracnose resistance was identified based on the map. The results will aid molecular marker-assisted breeding and walnut resistance genes identification.

Description: Background: In the male germline, neonatal prospermatogonia give rise to spermatogonia, which include stem cell population (undifferentiated spermatogonia) that supports continuous spermatogenesis in adults. Although the levels of DNA methyltransferases change dynamically in the neonatal and early postnatal male germ cells, detailed genome-wide DNA methylation profiles of these cells during the stem cell formation and differentiation have not been reported. Results: To understand the regulation of spermatogonial stem cell formation and differentiation, we examined the DNA methylation and gene expression dynamics of male mouse germ cells at the critical stages: neonatal prospermatogonia, and early postntal (day 7) undifferentiated and differentiating spermatogonia. We found large partially methylated domains similar to those found in cancer cells and placenta in all these germ cells, and high levels of non-CG methylation and 5-hydroxymethylcytosines in neonatal prospermatogonia. Although the global CG methylation levels were stable in early postnatal male germ cells, and despite the reported scarcity of differential methylation in the adult spermatogonial stem cells, we identified many regions showing stage-specific differential methylation in and around genes important for stem cell function and spermatogenesis. These regions contained binding sites for specific transcription factors including the SOX family members. Conclusions: Our findings show a distinctive and dynamic regulation of DNA methylation during spermatogonial stem cell formation and differentiation in the neonatal and early postnatal testes. Furthermore, we revealed a unique accumulation and distribution of non-CG methylation and 5hmC marks in neonatal prospermatogonia. These findings contrast with the reported scarcity of differential methylation in adult spermatogonial stem cell differentiation and represent a unique phase of male germ cell development.

Description: Background: The transposable element (TE) content of the genomes of plant species varies from near zero in the genome of Utricularia gibba to more than 80 % in many species. It is not well understood whether this variation in genome composition results from common mechanisms or stochastic variation. The major obstacles to investigating mechanisms of TE evolution have been a lack of comparative genomic data sets and efficient computational methods for measuring differences in TE composition between species. In this study, we describe patterns of TE evolution in 14 species in the flowering plant family Asteraceae and 1 outgroup species in the Calyceraceae to investigate phylogenetic patterns of TE dynamics in this important group of plants. Results: Our findings indicate that TE families in the Asteraceae exhibit distinct patterns of non-neutral evolution, and that there has been a directional increase in copy number of Gypsy retrotransposons since the origin of the Asteraceae. Specifically, there is marked increase in Gypsy abundance at the origin of the Asteraceae and at the base of the tribe Heliantheae. This latter shift in genome composition has had a significant impact on the diversity and abundance distribution of TEs in a lineage-specific manner. Conclusions: We show that the TE-driven expansion of plant genomes can be facilitated by just a few TE families, and is likely accompanied by the modification and/or replacement of the TE community. Importantly, large shifts in TE composition may be correlated with major of phylogenetic transitions.

Description: Background: Root rot caused by Aphanomyces euteiches is one of the most destructive pea diseases while a distantly related species P. pisi has been recently described as the agent of pea and faba bean root rot. These two oomycete pathogens with different pathogenicity factor repertories have both evolved specific mechanisms to infect pea. However, little is known about the genes and mechanisms of defence against these pathogens in pea. In the present study, the transcriptomic response of pea to these two pathogens was investigated at two time points during early phase of infection using a Medicago truncatula microarray. Results: Of the 37,976 genes analysed, 574 and 817 were differentially expressed in response to A. euteiches at 6 hpi and 20 hpi, respectively, while 544 and 611 genes were differentially regulated against P. pisi at 6 hpi and 20 hpi, respectively. Differentially expressed genes associated with plant immunity responses were involved in cell wall reinforcement, hormonal signalling and phenylpropanoid metabolism. Activation of cell wall modification, regulation of jasmonic acid biosynthesis and induction of ethylene signalling pathway were among the common transcriptional responses to both of these oomycetes. However, induction of chalcone synthesis and the auxin pathway were specific transcriptional changes against A. euteiches. Conclusions: Our results demonstrate a global view of differentially expressed pea genes during compatible interactions with P. pisi and A. euteiches at an early phase of infection. The results suggest that distinct signalling pathways are triggered in pea by these two pathogens that lead to common and specific immune mechanisms in response to these two oomycetes. The generated knowledge may eventually be used in breeding pea varieties with resistance against root rot disease.

Description: Background: In humans it is unknown if the composition of the gut microbiota alters the risk of Plasmodium falciparum infection or the risk of developing febrile malaria once P. falciparum infection is established. Here we collected stool samples from a cohort composed of 195 Malian children and adults just prior to an intense P. falciparum transmission season. We assayed these samples using massively parallel sequencing of the 16S ribosomal RNA gene to identify the composition of the gut bacterial communities in these individuals. During the ensuing 6-month P. falciparum transmission season we examined the relationship between the stool microbiota composition of individuals in this cohort and their prospective risk of both P. falciparum infection and febrile malaria. Results: Consistent with prior studies, stool microbial diversity in the present cohort increased with age, although the overall microbiota profile was distinct from cohorts in other regions of Africa, Asia and North America. Age-adjusted Cox regression analysis revealed a significant association between microbiota composition and the prospective risk of P. falciparum infection; however, no relationship was observed between microbiota composition and the risk of developing febrile malaria once P. falciparum infection was established. Conclusions: These findings underscore the diversity of gut microbiota across geographic regions, and suggest that strategic modulation of gut microbiota composition could decrease the risk of P. falciparum infection in malaria-endemic areas, potentially as an adjunct to partially effective malaria vaccines.

Description: Background: The hemibiotroph Colletotrichum lentis, causative agent of anthracnose on Lens culinaris (lentil) was recently described as a new species. During its interaction with the host plant, C. lentis likely secretes numerous effector proteins, including toxins to alter the plant’s innate immunity, thereby gaining access to the host tissues for nutrition and reproduction. Results: In silico analysis of 2000 ESTs generated from C. lentis-infected lentil leaf tissues identified 15 candidate effectors. In planta infection stage-specific gene expression waves among candidate effectors were revealed for the appressorial penetration phase, biotrophic phase and necrotrophic phase. No sign of positive selection pressure [ω (dN/dS) 

Description: Background: Improved understanding of bacterial-fungal interactions in the rhizosphere should assist in the successful application of bacteria as biological control agents against fungal pathogens of plants, providing alternatives to chemicals in sustainable agriculture. Rhizoctonia solani is an important soil-associated fungal pathogen and its chemical treatment is not feasible or economic. The genomes of the plant-associated bacteria Serratia proteamaculans S4 and Serratia plymuthica AS13 have been sequenced, revealing genetic traits that may explain their diverse plant growth promoting activities and antagonistic interactions with R. solani. To understand the functional response of this pathogen to different bacteria and to elucidate whether the molecular mechanisms that the fungus exploits involve general stress or more specific responses, we performed a global transcriptome profiling of R. solani Rhs1AP anastomosis group 3 (AG-3) during interaction with the S4 and AS13 species of Serratia using RNA-seq. Results: Approximately 104,504 million clean 75-100 bp paired-end reads were obtained from three libraries, each in triplicate (AG3-Control, AG3-S4 and AG3-AS13). Transcriptome analysis revealed that approximately 10 % of the fungal transcriptome was differentially expressed during challenge with Serratia. The numbers of S4- and AS13-specific differentially expressed genes (DEG) were 866 and 292 respectively, while there were 1035 common DEGs in the two treatment groups. Four hundred and sixty and 242 genes respectively had values of log 2 fold-change 〉 3 and for further analyses this cut-off value was used. Functional classification of DEGs based on Gene Ontology enrichment analysis and on KEGG pathway annotations revealed a general shift in fungal gene expression in which genes related to xenobiotic degradation, toxin and antioxidant production, energy, carbohydrate and lipid metabolism and hyphal rearrangements were subjected to transcriptional regulation. Conclusions: This RNA-seq profiling generated a novel dataset describing the functional response of the phytopathogen R. solani AG3 to the plant-associated Serratia bacteria S4 and AS13. Most genes were regulated in the same way in the presence of both bacterial isolates, but there were also some strain-specific responses. The findings in this study will be beneficial for further research on biological control and in depth exploration of bacterial-fungal interactions in the rhizosphere.

Description: Characterizing Leaf Area Index (LAI) and Vertical Foliage Profile (VFP) over the United States Biogeosciences Discussions, 12, 13675-13710, 2015 Author(s): H. Tang, S. Ganguly, G. Zhang, M. A. Hofton, R. F. Nelson, and R. Dubayah Leaf area index (LAI) and vertical foliage profile (VFP) are among the important canopy structural variables. Recent advances in lidar remote sensing technology have demonstrated the capability of accurately mapping LAI and VFP over large areas. The primary objective of this study was to derive and validate a LAI and VFP product over the contiguous United States using spaceborne waveform lidar data. This product was derived at the footprint level from the Geoscience Laser Altimeter System (GLAS) using a biophysical model. We validated GLAS derived LAI and VFP across major forest biomes using airborne waveform lidar. The comparison results showed that GLAS retrievals of total LAI were generally accurate with little bias ( r 2 = 0.67, bias = −0.13, RMSE = 0.75). The derivations of GLAS retrievals of VFP within layers was not as accurate overall ( r 2 = 0.36, bias = −0.04, RMSE = 0.26), and these varied as a function of height, increasing from understory to overstory −0 to 5 m layer: r 2 = 0.04, bias = 0.09, RMSE = 0.31; 10 to 15 m layer: r 2 = 0.53, bias = −0.08, RMSE = 0.22; and 15 to 20 m layer: r 2 = 0.66, bias =−0.05, RMSE = 0.20. Significant relationships were also found between GLAS LAI products and different environmental factors, in particular elevation and annual precipitation. In summary, our results provide a unique insight into vertical canopy structure distribution across North American ecosystems. This data set is a first step towards a baseline of canopy structure needed for evaluating climate and land use induced forest changes at continental scale in the future and should help deepen our understanding of the role of vertical canopy structure on terrestrial ecosystem processes across varying scales.

Description: Background: The radioactive isotope Strontium-90 ( 90 Sr) may be released as a component of fallout from nuclear accidents, or in the event of a radiological incident such as detonation of an improvised nuclear device, and if ingested poses a significant health risk to exposed individuals. In order to better understand the response to 90 Sr, using an easily attainable and standard biodosimetry sample fluid, we analyzed the global transcriptomic response of blood cells in an in vivo model system. Results: We injected C57BL/6 mice with a solution of 90SrCl2 and followed them over a 30-day period. At days 4, 7, 9, 25 and 30, we collected blood and isolated RNA for microarray analyses. These days corresponded to target doses in a range from 1–5 Gy. We investigated changes in mRNA levels using microarrays, and changes in specific microRNA (miRNA) predicted to be involved in the response using qRT-PCR. We identified 8082 differentially expressed genes in the blood of mice exposed to 90 Sr compared with controls. Common biological functions were affected throughout the study, including apoptosis of B and T lymphocytes, and atrophy of lymphoid organs. Cellular functions such as RNA degradation and lipid metabolism were also affected during the study. The broad down regulation of genes observed in our study suggested a potential role for miRNA in gene regulation. We tested candidate miRNAs, mmu-miR-16, mmu-miR-124, mmu-miR-125 and mmu-mir-21; and found that all were induced at the earliest time point, day 4. Conclusions: Our study is the first to report the transcriptomic response of blood cells to the internal emitter 90 Sr in mouse and a possible role for microRNA in gene regulation after 90 Sr exposure. The most dramatic effect was observed on gene expression related to B-cell development and RNA maintenance. These functions were affected by genes that were down regulated throughout the study, suggesting severely compromised antigen response, which may be a result of the deposition of the radioisotope proximal to the hematopoietic compartment in bone.

Description: Application of the 15 N-Gas Flux method for measuring in situ N 2 and N 2 O fluxes due to denitrification in natural and semi-natural terrestrial ecosystems and comparison with the acetylene inhibition technique Biogeosciences Discussions, 12, 12653-12689, 2015 Author(s): F. Sgouridis, S. Ullah, and A. Stott Soil denitrification is considered the most un-constrained process in the global N cycle due to uncertain in situ N 2 flux measurements, particularly in natural and semi-natural terrestrial ecosystems. 15 N tracer approaches can provide in situ measurements of both N 2 and N 2 O simultaneously, but their use has been limited to fertilised agro-ecosystems due to the need for large 15 N additions in order to detect 15 N 2 production against the high atmospheric N 2 . For 15 N-N 2 analyses, we have used an "in house" laboratory designed and manufactured N 2 preparation instrument which can be interfaced to any commercial continuous flow isotope ratio mass spectrometer (CF-IRMS). The N 2 prep unit has gas purification steps, a copper based reduction furnace, and allows the analysis of small gas injection volumes (4 μL) for 15 N-N 2 analysis. For the analysis of N 2 O, an automated Tracegas Pre-concentrator (Isoprime Ltd) coupled to an IRMS was used to measure the 15 N-N 2 O (4 mL gas injection volume). Consequently, the coefficient of variation for the determination of isotope ratios for N 2 in air and in standard N 2 O (0.5 ppm) was better than 0.5 %. The 15 N Gas-Flux method was adapted for application in natural and semi-natural land use types (peatlands, forests and grasslands) by lowering the 15 N tracer application rate to 0.04–0.5 kg 15 N ha −1 . For our chamber design (volume / surface = 8:1) and a 20 h incubation period, the minimum detectable flux rates were 4 μg N m −2 h −1 and 0.2 ng N m −2 h −1 for the N 2 and N 2 O fluxes respectively. The N 2 flux ranged between 2.4 and 416.6 μg N m −2 h −1 , and the grassland soils showed on average 3 and 14 times higher denitrification rates than the woodland and organic soils respectively. The N 2 O flux was on average 20 to 200 times lower than the N 2 flux, while the denitrification product ratio (N 2 O/N 2 + N 2 O) was low, ranging between 0.03 and 13 %. Total denitrification rates measured by the acetylene inhibition technique under the same field conditions correlated ( r = 0.58) with the denitrification rates measured under the 15 N Gas-Flux method but were underestimated by a factor of 4 and this was attributed to the incomplete inhibition of N 2 O reduction to N 2 under relatively high soil moisture content. The results show that the 15 N Gas-Flux method can be used for quantifying N 2 and N 2 O production rates in natural terrestrial ecosystems, thus significantly improving our ability to constrain ecosystem N budgets.

Description: Background: A complete genome sequence is an essential tool for the genetic improvement of wheat. Because the wheat genome is large, highly repetitive and complex due to its allohexaploid nature, the International Wheat Genome Sequencing Consortium (IWGSC) chose a strategy that involves constructing bacterial artificial chromosome (BAC)-based physical maps of individual chromosomes and performing BAC-by-BAC sequencing. Here, we report the construction of a physical map of chromosome 6B with the goal of revealing the structural features of the third largest chromosome in wheat. Results: We assembled 689 informative BAC contigs (hereafter reffered to as contigs) representing 91 % of the entire physical length of wheat chromosome 6B. The contigs were integrated into a radiation hybrid (RH) map of chromosome 6B, with one linkage group consisting of 448 loci with 653 markers. The order and direction of 480 contigs, corresponding to 87 % of the total length of 6B, were determined. We also characterized the contigs that contained a part of the nucleolus organizer region or centromere based on their positions on the RH map and the assembled BAC clone sequences. Analysis of the virtual gene order along 6B using the information collected for the integrated map revealed the presence of several chromosomal rearrangements, indicating evolutionary events that occurred on chromosome 6B. Conclusions: We constructed a reliable physical map of chromosome 6B, enabling us to analyze its genomic structure and evolutionary progression. More importantly, the physical map should provide a high-quality and map-based reference sequence that will serve as a resource for wheat chromosome 6B.

Description: Background: Base-resolution methylome data generated by whole-genome bisulfite sequencing (WGBS) is often used to segment the genome into domains with distinct methylation levels. However, most segmentation methods include many parameters to be carefully tuned and/or fail to exploit the unsurpassed resolution of the data. Furthermore, there is no simple method that displays the composition of the domains to grasp global trends in each methylome. Results: We propose to use changepoint detection for domain demarcation based on base-resolution methylome data. While the proposed method segments the methylome in a largely comparable manner to conventional approaches, it has only a single parameter to be tuned. Furthermore, it fully exploits the base-resolution of the data to enable simultaneous detection of methylation changes in even contrasting size ranges, such as focal hypermethylation and global hypomethylation in cancer methylomes. We also propose a simple plot termed methylated domain landscape (MDL) that globally displays the size, the methylation level and the number of the domains thus defined, thereby enabling one to intuitively grasp trends in each methylome. Since the pattern of MDL often reflects cell lineages and is largely unaffected by data size, it can serve as a novel signature of methylome. Conclusions: Changepoint detection in base-resolution methylome data followed by MDL plotting provides a novel method for methylome characterization and will facilitate global comparison among various WGBS data differing in size and even species origin.

Description: Background: Bi-parental mapping populations have been commonly utilized to identify and characterize quantitative trait loci (QTL) controlling resistance to soybean cyst nematode (SCN, Heterodera glycines Ichinohe). Although this approach successfully mapped a large number of SCN resistance QTL, it captures only limited allelic diversity that exists in parental lines, and it also has limitations for genomic resolution. In this study, a genome-wide association study (GWAS) was performed using a diverse set of 553 soybean plant introductions (PIs) belonging to maturity groups from III to V to detect QTL/genes associated with SCN resistance to HG Type 0. Results: Over 45,000 single nucleotide polymorphism (SNP) markers generated by the SoySNP50K iSelect BeadChip (http// www.soybase.org ) were utilized for analysis. GWAS identified 14 loci distributed over different chromosomes comprising 60 SNPs significantly associated with SCN resistance. Results also confirmed six QTL that were previously mapped using bi-parental populations, including the rhg1 and Rhg4 loci. GWAS identified eight novel QTL, including QTL on chromosome 10, which we have previously mapped by using a bi-parental population. In addition to the known loci for four simple traits, such as seed coat color, flower color, pubescence color, and stem growth habit, two traits, like lodging and pod shattering, having moderately complex inheritance have been confirmed with great precision by GWAS. Conclusions: The study showed that GWAS can be employed as an effective strategy for identifying complex traits in soybean and for narrowing GWAS-defined genomic regions, which facilitates positional cloning of the causal gene(s).

Description: Background: Unlike mammals, zebrafish have the ability to regenerate damaged parts of their central nervous system (CNS) and regain functionality of the affected area. A better understanding of the molecular mechanisms involved in zebrafish regeneration may therefore provide insight into how CNS repair might be induced in mammals. Although many studies have described differences in gene expression in zebrafish during CNS regeneration, the regulatory mechanisms underpinning the differential expression of these genes have not been examined. Results: We used microarrays to analyse and integrate the mRNA and microRNA (miRNA) expression profiles of zebrafish retina after optic nerve crush to identify potential regulatory mechanisms that underpin central nerve regeneration. Bioinformatic analysis identified 3 miRNAs and 657 mRNAs that were differentially expressed after injury. We then combined inverse correlations between our miRNA expression and mRNA expression, and integrated these findings with target predictions from TargetScan Fish to identify putative miRNA-gene target pairs. We focused on two over-expressed miRNAs (miR-29b and miR-223), and functionally validated seven of their predicted gene targets using RT-qPCR and luciferase assays to confirm miRNA-mRNA binding. Gene ontology analysis placed the miRNA-regulated genes (eva1a, layna, nefmb, ina, si:ch211-51a6.2, smoc1, sb:cb252) in key biological processes that included cell survival/apoptosis, ECM-cytoskeleton signaling, and heparan sulfate proteoglycan binding, Conclusion: Our results suggest a key role for miR-29b and miR-223 in zebrafish regeneration. The identification of miRNA regulation in a zebrafish injury model provides a framework for future studies in which to investigate not only the cellular processes required for CNS regeneration, but also how these mechanisms might be regulated to promote successful repair and return of function in the injured mammalian brain.

Description: Background: Ameloblast differentiation is the most critical stepwise process in amelogenesis, and it is controlled by precise molecular events. To better understand the mechanism controlling pre-ameloblasts (PABs) differentiation into secretory ameloblasts (SABs), a more precise identification of molecules and signaling networks will elucidate the mechanisms governing enamel formation and lay a foundation for enamel regeneration. Results: We analyzed transcriptional profiles of human PABs and SABs. From a total of 28,869 analyzed transcripts, we identified 923 differentially expressed genes (DEGs) with p   2. Among the DEGs, 647 genes showed elevated expression in PABs compared to SABs. Notably, 38 DEGs displayed greater than eight-fold changes. Comparative analysis revealed that highly expressed genes in PABs were involved in cell cycle control, DNA damage repair and apoptosis, while highly expressed genes in SABs were related to cell adhesion and extracellular matrix. Moreover, coexpression network analysis uncovered two highly conserved sub-networks contributing to differentiation, containing transcription regulators (RUNX2, ETV1 and ETV5), solute carrier family members (SLC15A1 and SLC7A11), enamel matrix protein (MMP20), and a polymodal excitatory ion channel (TRPA1). Conclusions: By combining comparative analysis and coexpression networks, this study provides novel biomarkers and research targets for ameloblast differentiation and the potential for their application in enamel regeneration.

Description: Background: Herbicide resistance in agrestal weeds is a global problem threatening food security. Non-target-site resistance (NTSR) endowed by mechanisms neutralising the herbicide or compensating for its action is considered the most agronomically noxious type of resistance. Contrary to target-site resistance, NTSR mechanisms are far from being fully elucidated. A part of weed response to herbicide stress, NTSR is considered to be largely driven by gene regulation. Our purpose was to establish a transcriptome resource allowing investigation of the transcriptomic bases of NTSR in the major grass weed Alopecurus myosuroides L. (Poaceae) for which almost no genomic or transcriptomic data was available. Results: RNA-Seq was performed from plants in one F2 population that were sensitive or expressing NTSR to herbicides inhibiting acetolactate-synthase. Cloned plants were sampled over seven time-points ranging from before until 73 h after herbicide application. Assembly of over 159M high-quality Illumina reads generated a transcriptomic resource (ALOMYbase) containing 65,558 potentially active contigs (N50 = 1240 nucleotides) predicted to encode 32,138 peptides with 74 % GO annotation, of which 2017 were assigned to protein families presumably involved in NTSR. Comparison with the fully sequenced grass genomes indicated good coverage and correct representation of A. myosuroides transcriptome in ALOMYbase. The part of the herbicide transcriptomic response common to the resistant and the sensitive plants was consistent with the expected effects of acetolactate-synthase inhibition, with striking similarities observed with published Arabidopsis thaliana data. A. myosuroides plants with NTSR were first affected by herbicide action like sensitive plants, but ultimately overcame it. Analysis of differences in transcriptomic herbicide response between resistant and sensitive plants did not allow identification of processes directly explaining NTSR. Five contigs associated to NTSR in the F2 population studied were tentatively identified. They were predicted to encode three cytochromes P450 (CYP71A, CYP71B and CYP81D), one peroxidase and one disease resistance protein. Conclusions: Our data confirmed that gene regulation is at the root of herbicide response and of NTSR. ALOMYbase proved to be a relevant resource to support NTSR transcriptomic studies, and constitutes a valuable tool for future research aiming at elucidating gene regulations involved in NTSR in A. myosuroides.

Description: Natural variability in the surface ocean carbonate ion concentration Biogeosciences Discussions, 12, 13123-13157, 2015 Author(s): N. S. Lovenduski, M. C. Long, and K. Lindsay We investigate variability in the surface ocean carbonate ion concentration ([CO 3 2− ]) on the basis of a long control simulation with a fully-coupled Earth System Model. The simulation is run with a prescribed, pre-industrial atmospheric CO 2 concentration for 1000 years, permitting investigation of natural [CO 3 2− ] variability on interannual to multi-decadal timescales. We find high interannual variability in surface [CO 3 2− ] in the tropical Pacific and at the boundaries between the subtropical and subpolar gyres in the Northern Hemisphere, and relatively low interannual variability in the centers of the subtropical gyres and in the Southern Ocean. Statistical analysis of modeled [CO 3 2− ] variance and autocorrelation suggests that significant anthropogenic trends in the saturation state of aragonite (Ω aragonite ) are already or nearly detectable at the sustained, open-ocean timeseries sites, whereas several decades of observations are required to detect anthropogenic trends in Ω aragonite in the tropical Pacific, North Pacific, and North Atlantic. The detection timescale for anthropogenic trends in pH is shorter than that for Ω aragonite , due to smaller noise-to-signal ratios and lower autocorrelation in pH. In the tropical Pacific, the leading mode of surface [CO 3 2− ] variability is primarily driven by variations in the vertical advection of dissolved inorganic carbon (DIC) in association with El Niño–Southern Oscillation. In the North Pacific, surface [CO 3 2− ] variability is caused by circulation-driven variations in surface DIC and strongly correlated with the Pacific Decadal Oscillation, with peak spectral power at 20–30 year periods. North Atlantic [CO 3 2− ] variability is also driven by variations in surface DIC, and exhibits weak correlations with both the North Atlantic Oscillation and the Atlantic Multidecadal Oscillation. As the scientific community seeks to detect the anthropogenic influence on ocean carbonate chemistry, these results will aid the interpretation of trends calculated from spatially- and temporally-sparse observations.

Description: Background: Species showing complex life cycles provide excellent opportunities to study the genetic associations between life cycle stages, as selective pressures may differ before and after metamorphosis. The European eel presents a complex life cycle with two metamorphoses, a first metamorphosis from larvae into glass eels (juvenile stage) and a second metamorphosis into silver eels (adult stage). We tested the hypothesis that different genes and gene pathways will be under selection at different life stages when comparing the genetic associations between glass eels and silver eels. Results: We used two sets of markers to test for selection: first, we genotyped individuals using a panel of 80 coding-gene single nucleotide polymorphisms (SNPs) developed in American eel; second, we investigated selection at the genome level using a total of 153,423 RAD-sequencing generated SNPs widely distributed across the genome. Using the RAD approach, outlier tests identified a total of 2413 (1.57 %) potentially selected SNPs. Functional annotation analysis identified signal transduction pathways as the most over-represented group of genes, including MAPK/Erk signalling, calcium signalling and GnRH (gonadotropin-releasing hormone) signalling. Many of the over-represented pathways were related to growth, while others could result from the different conditions that eels inhabit during their life cycle. Conclusions: The observation of different genes and gene pathways under selection when comparing glass eels vs. silver eels supports the adaptive decoupling hypothesis for the benefits of metamorphosis. Partitioning the life cycle into discrete morphological phases may be overall beneficial since it allows the different life stages to respond independently to their unique selection pressures. This might translate into a more effective use of food and niche resources and/or performance of phase-specific tasks (e.g. feeding in the case of glass eels, migrating and reproducing in the case of silver eels).

Description: Background: Avocado (Persea americana) is an economically important tropical fruit considered to be a good source of fatty acids. Despite its importance, the molecular and cellular characterization of biochemical and developmental processes in avocado is limited due to the lack of transcriptome and genomic information. Results: The transcriptomes of seeds, roots, stems, leaves, aerial buds and flowers were determined using different sequencing platforms. Additionally, the transcriptomes of three different stages of fruit ripening (pre-climacteric, climacteric and post-climacteric) were also analyzed. The analysis of the RNAseqatlas presented here reveals strong differences in gene expression patterns between different organs, especially between root and flower, but also reveals similarities among the gene expression patterns in other organs, such as stem, leaves and aerial buds (vegetative organs) or seed and fruit (storage organs). Important regulators, functional categories, and differentially expressed genes involved in avocado fruit ripening were identified. Additionally, to demonstrate the utility of the avocado gene expression atlas, we investigated the expression patterns of genes implicated in fatty acid metabolism and fruit ripening. Conclusions: A description of transcriptomic changes occurring during fruit ripening was obtained in Mexican avocado, contributing to a dynamic view of the expression patterns of genes involved in fatty acid biosynthesis and the fruit ripening process.

Description: Background: Deidentified newborn screening bloodspot samples (NBS) represent a valuable potential resource for genomic research if impediments to whole exome sequencing of NBS deoxyribonucleic acid (DNA), including the small amount of genomic DNA in NBS material, can be overcome. For instance, genomic analysis of NBS could be used to define allele frequencies of disease-associated variants in local populations, or to conduct prospective or retrospective studies relating genomic variation to disease emergence in pediatric populations over time. In this study, we compared the recovery of variant calls from exome sequences of amplified NBS genomic DNA to variant calls from exome sequencing of non-amplified NBS DNA from the same individuals. Results: Using a standard alignment-based Genome Analysis Toolkit (GATK), we find 62,000–76,000 additional variants in amplified samples. After application of a unique kmer enumeration and variant detection method (RUFUS), only 38,000–47,000 additional variants are observed in amplified gDNA. This result suggests that roughly half of the amplification-introduced variants identified using GATK may be the result of mapping errors and read misalignment. Conclusions: Our results show that it is possible to obtain informative, high-quality data from exome analysis of whole genome amplified NBS with the important caveat that different data generation and analysis methods can affect variant detection accuracy, and the concordance of variant calls in whole-genome amplified and non-amplified exomes.

Description: Background: Acanthus is a unique genus consisting of both true mangrove and terrestrial species; thus, it represents an ideal system for studying the origin and adaptive evolution of mangrove plants to intertidal environments. However, little is known regarding the two respects of mangrove species in Acanthus. In this study, we sequenced the transcriptomes of the pooled roots and leaves tissues for a mangrove species, Acanthus ilicifolius, and its terrestrial congener, A. leucostachyus, to illustrate the origin of the mangrove species in this genus and their adaptive evolution to harsh habitats. Results: We obtained 73,039 and 69,580 contigs with N50 values of 741 and 1557 bp for A. ilicifolius and A. leucostachyus, respectively. Phylogenetic analyses based on four nuclear segments and three chloroplast fragments revealed that mangroves and terrestrial species in Acanthus fell into different clades, indicating a single origin of the mangrove species in Acanthus. Based on 6634 orthologs, A. ilicifolius and A. leucostachyus were found to be highly divergent, with a peak of synonymous substitution rate (Ks) distribution of 0.145 and an estimated divergence time of approximately 16.8 million years ago (MYA). The transgression in the Early to Middle Miocene may be the major reason for the entry of the mangrove lineage of Acanthus into intertidal environments. Gene ontology (GO) classifications of the full transcriptomes did not show any apparent differences between A. ilicifolius and A. leucostachyus, suggesting the absence of gene components specific to the mangrove transcriptomes. A total of 99 genes in A. ilicifolius were identified with signals of positive selection. Twenty-three of the 99 positively selected genes (PSGs) were found to be involved in salt, heat and ultraviolet stress tolerance, seed germination and embryo development under periodic inundation. These stress-tolerance related PSGs may be crucial for the adaptation of the mangrove species in this genus to stressful marine environments and may contribute to speciation in Acanthus. Conclusions: We characterized the transcriptomes of one mangrove species of Acanthus, A. ilicifolius, and its terrestrial relative, A. leucostachyus, and provided insights into the origin of the mangrove Acanthus species and their adaptive evolution to abiotic stresses in intertidal environments.

Description: Spatial and temporal trends in summertime climate and water quality indicators in the coastal embayments of Buzzards Bay, Massachusetts Biogeosciences Discussions, 12, 13159-13192, 2015 Author(s): J. E. Rheuban, S. C. Williamson, J. E. Costa, D. M. Glover, R. W. Jakuba, D. C. McCorkle, C. Neill, T. Williams, and S. C. Doney Degradation of coastal ecosystems by eutrophication is largely defined by nitrogen loading from land via surface and groundwater flows. However, indicators of water quality are highly variable due to a myriad of other drivers, including temperature and precipitation. To evaluate these drivers, we examined spatial and temporal trends in a 22 year record of summer water quality data from 122 stations in 17 embayments within Buzzards Bay, MA (USA), collected through a citizen science monitoring program managed by Buzzards Bay Coalition. To identify spatial patterns across Buzzards Bay's embayments, we used a principle component and factor analysis and found that rotated factor loadings indicated little correlation between inorganic nutrients and organic matter and chlorophyll a (Chl a ) concentration. Factor scores showed that embayment geomorphology in addition to nutrient loading was a strong driver of water quality, where embayments with surface water inputs showed larger biological impacts than embayments dominated by groundwater influx. A linear regression analysis of annual summertime water quality indicators over time revealed that from 1992 to 2013, most embayments (15 of 17) exhibited an increase in temperature (mean rate of 0.082 ± 0.025 (SD) °C yr −1 ) and Chl a (mean rate of 0.0171 ± 0.0088 log 10 (Chl a ; mg m −3 ) yr −1 , equivalent to a 4.0 % increase per year). However, only 7 embayments exhibited an increase in total nitrogen (TN) concentration (mean rate 0.32 ± 0.47 (SD) μM yr −1 ). Average summertime log 10 (TN) and log 10 (Chl a ) were correlated with an indication that yield of Chl a per unit total nitrogen increased with time suggesting the estuarine response to TN may have changed because of other stressors such as warming, altered precipitation patterns, or changing light levels. These findings affirm that nitrogen loading and physical aspects of embayments are essential in explaining observed ecosystem response. However, climate-related stressors may also need to be considered by managers because increased temperature and precipitation may worsen water quality and partially offset benefits achieved by reducing nitrogen loading.

Description: Background: In higher eukaryotes, the genome is partitioned into large "Topologically Associating Domains" (TADs) in which the chromatin displays favoured long-range contacts. While a crumpled/fractal globule organization has received experimental supports at higher-order levels, the organization principles that govern chromatin dynamics within these TADs remain unclear. Using simple polymer models, we previously showed that, in mouse liver cells, gene-rich domains tend to adopt a statistical helix shape when no significant locus-specific interaction takes place. Results: Here, we use data from diverse 3C-derived methods to explore chromatin dynamics within mouse and Drosophila TADs. In mouse Embryonic Stem Cells (mESC), that possess large TADs (median size of 840 kb), we show that the statistical helix model, but not globule models, is relevant not only in gene-rich TADs, but also in gene-poor and gene-desert TADs. Interestingly, this statistical helix organization is considerably relaxed in mESC compared to liver cells, indicating that the impact of the constraints responsible for this organization is weaker in pluripotent cells. Finally, depletion of histone H1 in mESC alters local chromatin flexibility but not the statistical helix organization. In Drosophila, which possesses TADs of smaller sizes (median size of 70 kb), we show that, while chromatin compaction and flexibility are finely tuned according to the epigenetic landscape, chromatin dynamics within TADs is generally compatible with an unconstrained polymer configuration. Conclusions: Models issued from polymer physics can accurately describe the organization principles governing chromatin dynamics in both mouse and Drosophila TADs. However, constraints applied on this dynamics within mammalian TADs have a peculiar impact resulting in a statistical helix organization.

Description: Background: Field pea (Pisum sativum L.) is a cool-season grain legume that is cultivated world-wide for both human consumption and stock-feed purposes. Enhancement of genetic and genomic resources for field pea will permit improved understanding of the control of traits relevant to crop productivity and quality. Advances in second-generation sequencing and associated bioinformatics analysis now provide unprecedented opportunities for the development of such resources. The objective of this study was to perform transcriptome sequencing and characterisation from two genotypes of field pea that differ in terms of seed and plant morphological characteristics. Results: Transcriptome sequencing was performed with RNA templates from multiple tissues of the field pea genotypes Kaspa and Parafield. Tissue samples were collected at various growth stages, and a total of 23 cDNA libraries were sequenced using Illumina high-throughput sequencing platforms. A total of 407 and 352 million paired-end reads from the Kaspa and Parafield transcriptomes, respectively were assembled into 129,282 and 149,272 contigs, which were filtered on the basis of known gene annotations, presence of open reading frames (ORFs), reciprocal matches and degree of coverage. Totals of 126,335 contigs from Kaspa and 145,730 from Parafield were subsequently selected as the reference set. Reciprocal sequence analysis revealed that c. 87 % of contigs were expressed in both cultivars, while a small proportion were unique to each genotype. Reads from different libraries were aligned to the genotype-specific assemblies in order to identify and characterise expression of contigs on a tissue-specific basis, of which 87 % were expressed in more than one tissue, while others showed distinct expression patterns in specific tissues, providing unique transcriptome signatures. Conclusion: This study provided a comprehensive assembled and annotated transcriptome set for field pea that can be used for development of genetic markers, in order to assess genetic diversity, construct linkage maps, perform trait-dissection and implement whole-genome selection strategies in varietal improvement programs, as well to identify target genes for genetic modification approaches on the basis of annotation and expression analysis. In addition, the reference field pea transcriptome will prove highly valuable for comparative genomics studies and construction of a finalised genome sequence.

Description: Carbon dynamics in highly heterotrophic subarctic thaw ponds Biogeosciences Discussions, 12, 11707-11749, 2015 Author(s): T. Roiha, I. Laurion, and M. Rautio Global warming has accelerated the formation of permafrost thaw ponds in several subarctic and arctic regions. These ponds are net heterotrophic as evidenced by their greenhouse gas (GHG) supersaturation levels (CO 2 and CH 4 ), and generally receive large terrestrial carbon inputs from the thawing and eroding permafrost. We measured seasonal and vertical variations in the concentration and type of dissolved organic matter (DOM) in five subarctic thaw (thermokarst) ponds in northern Quebec, and explored how environmental gradients influenced heterotrophic and phototrophic biomass and productivity. Late winter DOM had low aromaticity indicating reduced inputs of terrestrial carbon, while the high concentration of dissolved organic carbon (DOC) suggests that some production of non-chromophoric dissolved compounds by the microbial food web took place under the ice cover. Summer DOM had a strong terrestrial signature, but was also characterized with significant inputs of algal-derived carbon, especially at the pond surface. During late winter, bacterial production was low (maximum of 0.8 mg C m −3 d −1 ) and was largely based on free-living bacterioplankton (58 %). Bacterial production in summer was high (up to 58 mg C m −3 d −1 ), dominated by particle-attached bacteria (67 %), and strongly correlated to the amount of terrestrial carbon. Primary production was restricted to summer surface waters due to strong light limitation deeper in the water column or in winter. The phototrophic biomass was equal to the heterotrophic biomass, but as the algae were mostly composed of mixotrophic species, most probably they used bacteria rather than solar energy in such shaded ponds. According to the δ 13 C analyses, non-algal carbon supported 51 % of winter and 37 % of summer biomass of the phantom midge larvae, Chaoborus sp., that are at the top of the trophic chain. Our results point to a strong heterotrophic energy pathway in these thaw pond ecosystems, where bacterioplankton dominates the production of new carbon in both summer and winter.

Description: Background: Formalin-fixed, paraffin-embedded (FFPE) tissues represent the most abundant resource of archived human specimens in pathology. Such tissue specimens are emerging as a highly valuable resource for translational proteomic studies. In quantitative proteomic analysis, reductive di-methylation of primary amines using stable isotopic formaldehyde variants is increasingly used due to its robustness and cost-effectiveness. Results: In the present study we show for the first time that isotopic amine dimethylation can be used in a straightforward manner for the quantitative proteomic analysis of FFPE specimens without interference from formalin employed in the FFPE process. Isotopic amine dimethylation of FFPE specimens showed equal labeling efficiency as for cryopreserved specimens. For both FFPE and cryopreserved specimens, differential labeling of identical samples yielded highly similar ratio distributions within the expected range for dimethyl labeling. In an initial application, we profiled proteome changes in clear cell renal cell carcinoma (ccRCC) FFPE tissue specimens compared to adjacent non–malignant renal tissue. Our findings highlight increased levels of glyocolytic enzymes, annexins as well as ribosomal and proteasomal proteins. Conclusion: Our study establishes isotopic amine dimethylation as a versatile tool for quantitative proteomic analysis of FFPE specimens and underlines proteome alterations in ccRCC.

Description: Phototrophic pigment diversity and picophytoplankton abundance in permafrost thaw lakes Biogeosciences Discussions, 12, 12121-12156, 2015 Author(s): A. Przytulska, J. Comte, S. Crevecoeur, C. Lovejoy, I. Laurion, and W. F. Vincent Permafrost thaw lakes (thermokarst lakes) are widely distributed across the northern landscape, and are known to be biogeochemically active sites that emit large amounts of carbon to the atmosphere as CH 4 and CO 2 . However, the abundance and composition of the photosynthetic communities that consume CO 2 have been little explored in this ecosystem type. In order to identify the major groups of phototrophic organisms and their controlling variables, we sampled 12 permafrost thaw lakes along a permafrost degradation gradient in northern Québec, Canada. Additional samples were taken from 5 rock-basin reference lakes in the region to determine if the thaw waters differed in limnological properties and phototrophs. Phytoplankton community structure was determined by high performance liquid chromatography analysis of their photoprotective and photosynthetic pigments, and autotrophic picoplankton concentrations were assessed by flow cytometry. One of the black colored lakes located in a andscape of rapidly degrading palsas (permafrost mounds) was selected for high-throughput 18S rRNA sequencing to help interpret the pigment and cytometry data. The results showed that the limnological properties of the thaw lakes differed significantly from the reference lakes, and were more highly stratified. However, both waterbody types contained similarly diverse phytoplankton groups, with dominance of the pigment assemblages by fucoxanthin-containing taxa, as well as chlorophytes, cryptophytes and cyanobacteria. Chlorophyll a concentrations (Chl a ) were correlated with total phosphorus (TP), and both were significantly higher in the thaw lakes (overall means of 3.3 μg Chl a L −1 and 34 μg TP L −1 ) relative to the reference lakes (2.0 μg Chl a L −1 and 8.2 μg TP L −1 ). Stepwise multiple regression of Chl a against the other algal pigments showed that it was largely a function of lutein, fucoxanthin and peridinin ( R 2 = 0.78). The bottom waters of two of the thaw lakes also contained high concentrations of bacteriochlorophyll d , showing the presence of green photosynthetic sulphur bacteria. The molecular analyses indicated a relatively minor contribution of diatoms, while chrysophytes, dinoflagellates and chlorophytes were well represented; the heterotrophic eukaryote fraction was dominated by numerous ciliate taxa, and also included Heliozoa, Rhizaria, chytrids and flagellates. Autotrophic picoplankton occurred in cell concentrations up to 8.8 × 10 5 mL −1 (picocyanobacteria) and 4.6 × 10 5 mL −1 (picoeukaryotes). Both groups of picophytoplankton were positively correlated with total phytoplankton abundance, as measured by Chl a ; picocyanobacteria were inversely correlated with dissolved organic carbon, while picoeukaryotes were correlated with conductivity. Despite their net heterotrophic character, subarctic thaw lakes are rich habitats for diverse phototrophic communities.

Description: Background: Lobosphaera incisa, formerly known as Myrmecia incisa and then Parietochloris incisa, is an oleaginous unicellular green alga belonging to the class Trebouxiophyceae (Chlorophyta). It is the richest known plant source of arachidonic acid, an ω-6 poly-unsaturated fatty acid valued by the pharmaceutical and baby-food industries. It is therefore an organism of high biotechnological interest, and we recently reported the sequence of its chloroplast genome. Results: We now report the complete sequence of the mitochondrial genome of L. incisa from high-throughput Illumina short-read sequencing. The circular chromosome of 69,997 bp is predicted to encode a total of 64 genes, some harboring specific self-splicing group I and group II introns. Overall, the gene content is highly similar to that of the mitochondrial genomes of other Trebouxiophyceae, with 34 protein-coding, 3 rRNA, and 27 tRNA genes. Genes are distributed in two clusters located on different DNA strands, a bipartite arrangement that suggests expression from two divergent promoters yielding polycistronic primary transcripts. The L. incisa mitochondrial genome contains families of intergenic dispersed DNA repeat sequences that are not shared with other known mitochondrial genomes of Trebouxiophyceae. The most peculiar feature of the genome is a repetitive palindromic repeat, the LIMP (L. Incisa Mitochondrial Palindrome), found 19 times in the genome. It is formed by repetitions of an AACCA pentanucleotide, followed by an invariant 7-nt loop and a complementary repeat of the TGGTT motif. Analysis of the genome sequencing reads indicates that the LIMP can be a substrate for large-scale genomic rearrangements. We speculate that LIMPs can act as origins of replication. Deep sequencing of the L. incisa transcriptome also suggests that the LIMPs with long stems are sites of transcript processing. The genome also contains five copies of a related palindromic repeat, the HyLIMP, with a 10-nt motif related to that of the LIMP. Conclusions: The mitochondrial genome of L. incisa encodes a unique type of repetitive palindromic repeat sequence, the LIMP, which can mediate genome rearrangements and play a role in mitochondrial gene expression. Experimental studies are needed to confirm and further characterize the functional role(s) of the LIMP.

Description: Background: The cereal rust fungi are destructive pathogens that affect grain production worldwide. Although the genomic and transcript sequences for three Puccinia species that attack wheat have been released, the functions of large repertories of genes from Puccinia still need to be addressed to understand the infection process of these obligate parasites. Host-induced gene silencing (HIGS) has emerged a useful tool to examine the importance of rust fungus genes while growing within host plants. In this study, HIGS was used to test genes from Puccinia with transcripts enriched in haustoria for their ability to interfere with full development of the rust fungi. Results: Approximately 1200 haustoria enriched genes from Puccinia graminis f. sp. tritici (Pgt) were identified by comparative RNA sequencing. Virus-induced gene silencing (VIGS) constructs with fragments of 86 Puccinia genes, were tested for their ability to interfere with full development of these rust fungi. Most of the genes tested had no noticeable effects, but 10 reduced Pgt development after co-inoculation with the gene VIGS constructs and Pgt. These included a predicted glycolytic enzyme, two other proteins that are probably secreted and involved in carbohydrate or sugar metabolism, a protein involved in thiazol biosynthesis, a protein involved in auxin biosynthesis, an amino acid permease, two hypothetical proteins with no conserved domains, a predicted small secreted protein and another protein predicted to be secreted with similarity to bacterial proteins involved in membrane transport. Transient silencing of four of these genes reduced development of P. striiformis (Pst), and three of also caused reduction of P. triticina (Pt) development. Conclusions: Partial suppression of transcripts involved in a large variety of biological processes in haustoria cells of Puccinia rusts can disrupt their development. Silencing of three genes resulted in suppression of all three rust diseases indicating that it may be possible to engineer durable resistance to multiple rust pathogens with a single gene in transgenic wheat plants for sustainable control of cereal rusts.

Description: Background: We present a genome-wide messenger RNA (mRNA) sequencing technique that converts small amounts of RNA from many samples into molecular phenotypes. It encompasses all steps from sample preparation to sequence analysis and is applicable to baseline profiling or perturbation measurements. Results: Multiplex sequencing of transcript 3′ ends identifies differential transcript abundance independent of gene annotation. We show that increasing biological replicate number while maintaining the total amount of sequencing identifies more differentially abundant transcripts. Conclusions: This method can be implemented on polyadenylated RNA from any organism with an annotated reference genome and in any laboratory with access to Illumina sequencing.

Description: Background: Verticillium wilt (VW) and Fusarium wilt (FW), caused by the soil-borne fungi Verticillium dahliae and Fusarium oxysporum f. sp. vasinfectum, respectively, are two most destructive diseases in cotton production worldwide. Root-knot nematodes (Meloidogyne incognita, RKN) and reniform nematodes (Rotylenchulus reniformis, RN) cause the highest yield loss in the U.S. Planting disease resistant cultivars is the most cost effective control method. Numerous studies have reported mapping of quantitative trait loci (QTLs) for disease resistance in cotton; however, very few reliable QTLs were identified for use in genomic research and breeding. Results: This study first performed a 4-year replicated test of a backcross inbred line (BIL) population for VW resistance, and 10 resistance QTLs were mapped based on a 2895 cM linkage map with 392 SSR markers. The 10 VW QTLs were then placed to a consensus linkage map with other 182 VW QTLs, 75 RKN QTLs, 27 FW QTLs, and 7 RN QTLs reported from 32 publications. A meta-analysis of QTLs identified 28 QTL clusters including 13, 8 and 3 QTL hotspots for resistance to VW, RKN and FW, respectively. The number of QTLs and QTL clusters on chromosomes especially in the A-subgenome was significantly correlated with the number of nucleotide-binding site (NBS) genes, and the distribution of QTLs between homeologous A- and D- subgenome chromosomes was also significantly correlated. Conclusions: Ten VW resistance QTL identified in a 4-year replicated study have added useful information to the understanding of the genetic basis of VW resistance in cotton. Twenty-eight disease resistance QTL clusters and 24 hotspots identified from a total of 306 QTLs and linked SSR markers provide important information for marker-assisted selection and high resolution mapping of resistance QTLs and genes. The non-overlapping of most resistance QTL hotspots for different diseases indicates that their resistances are controlled by different genes.

Description: Background: Genomic imprinting is an epigenetic mechanism that can lead to differential gene expression depending on the parent-of-origin of a received allele. While most studies on imprinting address its underlying molecular mechanisms or attempt at discovering genomic regions that might be subject to imprinting, few have focused on the amount of phenotypic variation contributed by such epigenetic process. In this report, we give a brief review of a one-locus imprinting model in a quantitative genetics framework, and provide a decomposition of the genetic variance according to this model. Analytical deductions from the proposed imprinting model indicated a non-negligible contribution of imprinting to genetic variation of complex traits. Also, we performed a whole-genome scan analysis on mouse body mass index (BMI) aiming at revealing potential consequences when existing imprinting effects are ignored in genetic analysis. Results: 10,021 SNP markers were used to perform a whole-genome single marker regression on mouse BMI using an additive and an imprinting model. Markers significant for imprinting indicated that BMI is subject to imprinting. Marked variance changed from 1.218 ×10 −4 to 1.842 ×10 −4 when imprinting was considered in the analysis, implying that one third of marked variance would be lost if existing imprinting effects were not accounted for. When both marker and pedigree information were used, estimated heritability increased from 0.176 to 0.195 when imprinting was considered. Conclusions: When a complex trait is subject to imprinting, using an additive model that ignores this phenomenon may result in an underestimate of additive variability, potentially leading to wrong inferences about the underlying genetic architecture of that trait. This could be a possible factor explaining part of the missing heritability commonly observed in genome-wide association studies (GWAS).

Description: Skeletal mineralogy of coral recruits under high temperature and p CO 2 Biogeosciences Discussions, 12, 12485-12500, 2015 Author(s): T. Foster and P. L. Clode Aragonite, which is the polymorph of CaCO 3 precipitated by modern corals during skeletal formation, has a higher solubility than the more stable polymorph calcite. This higher solubility leaves animals that produce aragonitic skeletons more vulnerable to anthropogenic ocean acidification. It is therefore, important to determine whether scleractinian corals have the plasticity to adapt and produce calcite in their skeletons in response to changing environmental conditions. Both high p CO 2 and lower Mg / Ca ratios in seawater are thought to have driven changes in the skeletal mineralogy of major marine calcifiers in the past ∼540 myr. Experimentally reduced Mg / Ca ratios in ambient seawater have been shown to induce some calcite precipitation in both adult and newly settled modern corals, however, the impact of high p CO 2 on the mineralogy of recruits is unknown. Here we determined the skeletal mineralogy of one-month old Acropora spicifera coral recruits grown under high temperature (+3 °C) and p CO 2 (∼900 μatm) conditions, using X-ray diffraction and Raman spectroscopy. We found that newly settled coral recruits produced entirely aragonitic skeletons regardless of the treatment. Our results show that elevated p CO 2 alone is unlikely to drive changes in the skeletal mineralogy of young corals. Not having an ability to switch from aragonite to calcite precipitation may leave corals and ultimately coral reef ecosystems more susceptible to predicted ocean acidification. An important area for prospective research would be to investigate the combined impact of high p CO 2 and reduced Mg / Ca ratio on coral skeletal mineralogy.

Description: The impact of sedimentary alkalinity release on the water column CO 2 system in the North Sea Biogeosciences Discussions, 12, 12395-12453, 2015 Author(s): H. Brenner, U. Braeckman, M. Le Guitton, and F. J. R. Meysman Recently, it has been proposed that alkalinity release from sediments can play an important role in the carbonate dynamics on continental shelves, lowering the p CO 2 of seawater and hence increasing the CO 2 uptake from the atmosphere. To test this hypothesis, sedimentary alkalinity generation was quantified within permeable and muddy sediments across the North Sea during two cruises in September 2011 (basin-wide) and June 2012 (Dutch coastal zone). Benthic fluxes of alkalinity ( A T ) and dissolved inorganic carbon (DIC) were determined using shipboard closed sediment incubations. These results show that sediments can be an important source for alkalinity, particularly in the shallow southern North Sea, where high A T and DIC fluxes were recorded in near shore sediments of the Belgian, Dutch and German coastal zone. In contrast, fluxes of A T and DIC are substantially lower in the deeper, seasonally stratified, northern part of the North Sea. Overall, our results show that sedimentary alkalinity generation should be considered an important factor in the CO 2 dynamics of shallow coastal systems.

Description: Contribution of Marine Group II Euryarchaeota to cyclopentyl tetraethers in the Pearl River estuary and coastal South China Sea: impact on the TEX 86 paleothermometer Biogeosciences Discussions, 12, 12455-12484, 2015 Author(s): J. X. Wang, C. L. Zhang, W. Xie, Y. G. Zhang, and P. Wang TEX 86 (TetraEther indeX of glycerol dialkyl glycerol tetraethers (GDGTs) with 86 carbon atoms) has been widely applied to reconstruct (paleo-) sea surface temperature (SST). While Marine Group I (MG I) Thaumarchaeota have been commonly believed to be the source for GDGTs, Marine Group II (MG II Euryarchaeota ) have recently been suggested to contribute significantly to the GDGT pool in the ocean. However, little is known how the MG II Euryarchaeota -derived GDGTs may influence TEX 86 in marine sediment record. In this study, we characterize MG II Euryarchaeota -produced GDGTs and assess the likely effect of these tetraether lipids on TEX 86 . Analyses of core lipid (CL-) and intact polar lipid (IPL-) based GDGTs, 454 sequencing and quantitative polymerase chain reaction (qPCR) targeting MG II Euryarchaeota were performed on suspended particulate matter (SPM) and surface sediments collected along a salinity gradient from the lower Pearl River (river water) and its estuary (mixing water) to the coastal South China Sea (seawater). The results showed that the community composition varied along the salinity gradient with MG II Euryarchaeota as the second dominant group in the mixing water and seawater. qPCR data indicated that the abundance of MG II Euryarchaeota in the mixing water was three to four orders of magnitude higher than the river water and seawater. Significant linear correlations were observed between the gene abundance ratio of MG II Euryarchaeota vs. total archaea and the relative abundance of GDGTs-1, -2, -3, or -4 as well as the ring index based on these compounds, which collectively suggest that MG II Euryarchaeota may actively produce GDGTs in the water column. These results also show strong evidence that MG II Euryarchaeota synthesizing GDGTs with 1–4 cyclopentane moieties may bias TEX 86 in the water column and sediments. This study highlights that valid interpretation of TEX 86 in sediment record, particularly in coastal oceans, needs to consider the contribution from MG II Euryarchaeota .

Description: Fate of peat-derived carbon and associated CO 2 and CO emissions from two Southeast Asian estuaries Biogeosciences Discussions, 12, 8299-8340, 2015 Author(s): D. Müller, T. Warneke, T. Rixen, M. Müller, A. Mujahid, H. W. Bange, and J. Notholt Coastal peatlands in Southeast Asia release large amounts of organic carbon to rivers, which transport it further to the adjacent estuaries. However, little is known about the fate of this terrestrial material in the coastal ocean. Although Southeast Asia is, by area, considered a hotspot of estuarine CO 2 emissions, studies in this region are very scarce. We measured dissolved and particulate organic carbon, carbon dioxide (CO 2 ) partial pressure and carbon monoxide (CO) concentrations in two tropical estuaries in Sarawak, Malaysia, whose coastal area is covered by peatlands. We surveyed the estuaries of the rivers Lupar and Saribas during the wet and dry season, respectively. The spatial distribution and the carbon-to-nitrogen ratios of dissolved organic matter (DOM) suggest that peat-draining rivers convey terrestrial organic carbon to the estuaries. We found evidence that a large fraction of this carbon is respired. The median p CO 2 in the estuaries ranged between 618 and 5064 μatm with little seasonal variation. CO 2 fluxes were determined with a floating chamber and estimated to amount to 14–272 mol m −2 yr −1 , which is high compared to other studies from tropical and subtropical sites. In contrast, CO concentrations and fluxes were relatively moderate (0.3–1.4 nmol L −1 and 0.8–1.9 mmol m −2 yr −1 ) if compared to published data for oceanic or upwelling systems. We attributed this to the large amounts of suspended matter (4–5004 mg L −1 ), limiting the light penetration depth. However, the diurnal variation of CO suggests that it is photochemically produced, implying that photodegradation might play a role for the removal of DOM from the estuary as well. We concluded that unlike smaller peat-draining tributaries, which tend to transport most carbon downstream, estuaries in this region function as an efficient filter for organic carbon and release large amounts of CO 2 to the atmosphere. The Lupar and Saribas mid-estuaries release 0.4 ± 0.2 Tg C yr −1 , which corresponds to approximately 80% of the emissions from the aquatic systems in these two catchments.

Description: Carbonate saturation state of surface waters in the Ross Sea and Southern Ocean: controls and implications for the onset of aragonite undersaturation Biogeosciences Discussions, 12, 8429-8465, 2015 Author(s): H. B. DeJong, R. B. Dunbar, D. A. Mucciarone, and D. A. Koweek Predicting when surface waters of the Ross Sea and Southern Ocean will become undersaturated with respect to biogenic carbonate minerals is challenging in part due to the lack of baseline high resolution carbon system data. Here we present ~ 1700 surface total alkalinity measurements from the Ross Sea and along a transect between the Ross Sea and southern Chile from the austral autumn (February–March 2013). We calculate the saturation state of aragonite (Ω Ar ) and calcite (Ω Ca ) using measured total alkalinity and p CO 2 . In the Ross Sea and south of the Polar Front, variability in carbonate saturation state (Ω) is mainly driven by algal photosynthesis. Freshwater dilution and calcification have minimal influence on Ω variability. We estimate an early spring surface water Ω Ar value of ~ 1.2 for the Ross Sea using a total alkalinity–salinity relationship and historical p CO 2 measurements. Our results suggest that the Ross Sea is not likely to become undersaturated with respect to aragonite until the year 2070.

Description: Biodegradability of dissolved organic carbon in permafrost soils and waterways: a meta-analysis Biogeosciences Discussions, 12, 8353-8393, 2015 Author(s): J. E. Vonk, S. E. Tank, P. J. Mann, R. G. M. Spencer, C. C. Treat, R. G. Striegl, B. W. Abbott, and K. P. Wickland As Arctic regions warm, the large organic carbon pool stored in permafrost becomes increasingly vulnerable to thaw and decomposition. The transfer of newly mobilized carbon to the atmosphere and its potential influence upon climate change will largely depend on the reactivity and subsequent fate of carbon delivered to aquatic ecosystems. Dissolved organic carbon (DOC) is a key regulator of aquatic metabolism and its biodegradability will determine the extent and rate of carbon release from aquatic ecosystems to the atmosphere. Knowledge of the mechanistic controls on DOC biodegradability is however currently poor due to a scarcity of long-term data sets, limited spatial coverage of available data, and methodological diversity. Here, we performed parallel biodegradable DOC (BDOC) experiments at six Arctic sites (16 experiments) using a standardized incubation protocol to examine the effect of methodological differences used as common practice in the literature. We further synthesized results from 14 aquatic and soil leachate BDOC studies from across the circum–arctic permafrost region to examine pan-Arctic trends in BDOC. An increasing extent of permafrost across the landscape resulted in higher BDOC losses in both soil and aquatic systems. We hypothesize that the unique composition of permafrost-derived DOC combined with limited prior microbial processing due to low soil temperature and relatively shorter flow path lengths and transport times, resulted in higher overall terrestrial and freshwater BDOC loss. Additionally, we found that the fraction of BDOC decreased moving down the fluvial network in continuous permafrost regions, i.e. from streams to large rivers, suggesting that highly biodegradable DOC is lost in headwater streams. We also observed a seasonal (January–December) decrease in BDOC losses in large streams and rivers, but no apparent change in smaller streams and soil leachates. We attribute this seasonal change to a combination of factors including shifts in carbon source, changing DOC residence time related to increasing thaw-depth, increasing water temperatures later in the summer, as well as decreasing hydrologic connectivity between soils and surface water as the seasons progress. Our results suggest that future, climate warming-induced shifts of continuous permafrost into discontinuous permafrost regions could affect the degradation potential of thaw-released DOC as well as its variability throughout the Arctic summer. We lastly present a recommended standardized BDOC protocol to facilitate the comparison of future work and improve our knowledge of processing and transport of DOC in a changing Arctic.

Description: Ideas and perspectives: use of tree-ring width as an indicator of tree growth Biogeosciences Discussions, 12, 8341-8352, 2015 Author(s): R. A. Hember, W. A. Kurz, and J. M. Metsaranta By taking core samples, dendroecological studies can reconstruct radial growth over the lifespan of a tree, providing a valuable way to estimate the sensitivity of tree productivity to environmental change. With increasing prevalence of such studies in global change science, it is worth cautioning that the incremental growth rate of a sub-dimension of a tree organ, such as annual ring width ( w ), does not respond to extrinsic perturbations with the same relative magnitude as the primary production of that organ. For example, if an extrinsic force causes a two-fold increase in the absolute growth rate of stemwood biomass (AGR), it should only theoretically translate into a 1.3-fold increase in w , or a 1.7-fold increase in basal area increment (BAI), when a 2:1 ratio in resource allocation to lateral and apical meristems is assumed. Expressing the magnitude of a response in relative terms does not, therefore, provide a valid means of comparing estimates of relative growth derived from measurement of different dimensional traits of the tree. From our perspective, enough conformity to facilitate comparison of environmental sensitivity across studies of tree growth is warranted so we emphasize the benefit of dimension analysis to transform measurements of w and BAI into the AGR. Although conversion to AGR introduces an error from the use of allometric equations, the approach is widely accepted in mainstream ecology and global change science at least partially because it avoids discrepancies in response magnitude owing to differences in dimension. Studies of organ elongation have historically provided invaluable information, yet it must be recognized that they systematically underestimate the response magnitude of primary production, and confound comparisons of growth sensitivity between many dendroecological studies that focus on w and studies of primary production.

Description: Background: Major secondary metabolites, including flavonoids, caffeine, and theanine, are important components of tea products and are closely related to the taste, flavor, and health benefits of tea. Secondary metabolite biosynthesis in Camellia sinensis is differentially regulated in different tissues during growth and development. Until now, little was known about the expression patterns of genes involved in secondary metabolic pathways or their regulatory mechanisms. This study aimed to generate expression profiles for C. sinensis tissues and to build a gene regulation model of the secondary metabolic pathways. Results: RNA sequencing was performed on 13 different tissue samples from various organs and developmental stages of tea plants, including buds and leaves of different ages, stems, flowers, seeds, and roots. A total of 43.7 Gbp of raw sequencing data were generated, from which 347,827 unigenes were assembled and annotated. There were 46,693, 8446, 3814, 10,206, and 4948 unigenes specifically expressed in the buds and leaves, stems, flowers, seeds, and roots, respectively. In total, 1719 unigenes were identified as being involved in the secondary metabolic pathways in C. sinensis, and the expression patterns of the genes involved in flavonoid, caffeine, and theanine biosynthesis were characterized, revealing the dynamic nature of their regulation during plant growth and development. The possible transcription factor regulation network for the biosynthesis of flavonoid, caffeine, and theanine was built, encompassing 339 transcription factors from 35 families, namely bHLH, MYB, and NAC, among others. Remarkably, not only did the data reveal the possible critical check points in the flavonoid, caffeine, and theanine biosynthesis pathways, but also implicated the key transcription factors and related mechanisms in the regulation of secondary metabolite biosynthesis. Conclusions: Our study generated gene expression profiles for different tissues at different developmental stages in tea plants. The gene network responsible for the regulation of the secondary metabolic pathways was analyzed. Our work elucidated the possible cross talk in gene regulation between the secondary metabolite biosynthetic pathways in C. sinensis. The results increase our understanding of how secondary metabolic pathways are regulated during plant development and growth cycles, and help pave the way for genetic selection and engineering for germplasm improvement.

Description: Background: Evidence is accumulating that perturbation of early life microbial colonization of the gut induces long-lasting adverse health effects in individuals. Understanding the mechanisms behind these effects will facilitate modulation of intestinal health. The objective of this study was to identify biological processes involved in these long lasting effects and the (molecular) factors that regulate them. We used an antibiotic and the same antibiotic in combination with stress on piglets as an early life perturbation. Then we used host gene expression data from the gut (jejunum) tissue and community-scale analysis of gut microbiota from the same location of the gut, at three different time-points to gauge the reaction to the perturbation. We analysed the data by a new combination of existing tools. First, we analysed the data in two dimensions, treatment and time, with quadratic regression analysis. Then we applied network-based data integration approaches to find correlations between host gene expression and the resident microbial species. Results: The use of a new combination of data analysis tools allowed us to identify significant long-lasting differences in jejunal gene expression patterns resulting from the early life perturbations. In addition, we were able to identify potential key gene regulators (hubs) for these long-lasting effects. Furthermore, data integration also showed that there are a handful of bacterial groups that were associated with temporal changes in gene expression. Conclusion: The applied systems-biology approach allowed us to take the first steps in unravelling biological processes involved in long lasting effects in the gut due to early life perturbations. The observed data are consistent with the hypothesis that these long lasting effects are due to differences in the programming of the gut immune system as induced by the temporary early life changes in the composition and/or diversity of microbiota in the gut.

Description: Background: Trifoliate orange (Poncirus trifoliata (L.) Raf.) is extremely cold hardy after a full acclimation; however the underlying molecular mechanisms underlying this economically valuable trait remain poorly understood. In this study, global transcriptome profiles of trifoliate orange under cold conditions (4 °C) over a time course were generated by high-throughput sequencing. Results: More than 68 million high-quality reads were produced and assembled into a non-redundant data of 77,292 unigenes with an average length of 1112 bp (N50 = 1778 bp). Of these, 23,846 had significant sequence similarity to known genes and these were assigned to 61 gene ontology (GO) categories and 25 clusters of orthologous groups (COG) involved in 128 KEGG pathways. Sequences derived from cold-treated and control plants were mapped to the assembled transcriptome, resulting in the identification of 5549 differentially expressed genes (DEGs). These comprised 600 (462 up-regulated, 138 down-regulated), 2346 (1631 up-regulated, 715 down-regulated), and 5177 (2702 up-regulated, 2475 down-regulated) genes from the cold-treated samples at 6, 24 and 72 h, respectively. The accuracy of the RNA-seq derived transcript expression data was validated by analyzing the expression patterns of 17 DEGs by qPCR. Plant hormone signal transduction, plant-pathogen interaction, and secondary metabolism were the most significantly enriched GO categories amongst in the DEGs. A total of 60 transcription factors were shown to be cold responsive. In addition, a number of genes involved in the catabolism and signaling of hormones, such as abscisic acid, ethylene and gibberellin, were affected by the cold stress. Meanwhile, levels of putrescine progressively increased under cold, which was consistent with up-regulation of an arginine decarboxylase gene. Conclusions: This dataset provides valuable information regarding the trifoliate orange transcriptome changes in response to cold stress and may help guide future identification and functional analysis of genes that are importnatn for enhancing cold hardiness.

Description: Spatial variability of diploptene δ 13 C values in thermokarst lakes: the potential to analyse the complexity of lacustrine methane cycling Biogeosciences Discussions, 12, 12157-12189, 2015 Author(s): K. L. Davies, R. D. Pancost, M. E. Edwards, K. M. Walter Anthony, P. G. Langdon, and L. Chaves Torres Cryospheric changes in northern high latitudes are linked to significant greenhouse gas flux to the atmosphere, including methane release that originates from organic matter decomposition in thermokarst lakes. The connections between methane production in sediments, transport pathways and oxidation are not well understood and this has implications for any attempts to reconstruct methane production from sedimentary archives. We assessed methane oxidation as represented by methane oxidising bacteria across the surface sediments of two interior Alaska thermokarst lakes in relation to methane emissions via ebullition (bubbling). The bacterial biomarker diploptene was present and had low δ 13 C values (lower than −38 ‰) in all sediments analysed, suggesting methane oxidation was widespread. The most δ 13 C-depleted diploptene was found in the area of highest methane ebullition emissions in Ace Lake (δ 13 C diplotene values between −68.2 and −50.1 ‰), suggesting a positive link between methane production, oxidation, and emission in this area. In contrast, significantly less depleted diploptene δ 13 C values (between −42.9 and −38.8 ‰) were found in the area of highest methane ebullition emissions in Smith Lake. Lower δ 13 C values of diploptene were found in the central area of Smith Lake (between −56.8 and −46.9 ‰), where methane ebullition rates are low but methane diffusion appears high. Using δ 13 C-diplotene as a proxy for methane oxidation activity, we suggest the observed differences in methane oxidation levels among sites within the two lakes could be linked to differences in source area of methane production (e.g. age and type of organic carbon) and bathymetry as it relates to varying oxycline depths and changing pressure gradients. As a result, methane oxidation is highly lake-dependent. The diploptene δ 13 C values also highlight strong within-lake variability, implying that single-value, down-core records of hopanoid isotopic signatures are not secure indicators of changing methane flux at the whole-lake scale.

Description: Metagenomic analyses of the late Pleistocene permafrost – additional tools for reconstruction of environmental conditions Biogeosciences Discussions, 12, 12091-12119, 2015 Author(s): E. Rivkina, L. Petrovskaya, T. Vishnivetskaya, K. Krivushin, L. Shmakova, M. Tutukina, A. Meyers, and F. Kondrashov A comparative analysis of the metagenomes from two 30 000 year-old permafrost samples, one of lake-alluvial origin and the other from late Pleistocene Ice Complex sediments, revealed significant differences within microbial communities. The late Pleistocene Ice Complex sediments (which have been characterized by the absence of methane with lower values of redox-potential and Fe 2+ content) showed both a low abundance of methanogenic archaea and enzymes from the carbon, nitrogen and sulfur cycles. The metagenomic and geochemical analyses described in the paper provide evidence that the formation of the late Pleistocene Ice Complex sediments likely took place under much more aerobic conditions than lake-alluvial sediments.

Description: Background: Chickens are susceptible to infection with a limited number of Influenza A viruses and are a potential source of a human influenza pandemic. In particular, H5 and H7 haemagglutinin subtypes can evolve from low to highly pathogenic strains in gallinaceous poultry. Ducks on the other hand are a natural reservoir for these viruses and are able to withstand most avian influenza strains. Results: Transcriptomic sequencing of lung and ileum tissue samples from birds infected with high (H5N1) and low (H5N2) pathogenic influenza viruses has allowed us to compare the early host response to these infections in both these species. Chickens (but not ducks) lack the intracellular receptor for viral ssRNA, RIG-I and the gene for an important RIG-I binding protein, RNF135. These differences in gene content partly explain the differences in host responses to low pathogenic and highly pathogenic avian influenza virus in chicken and ducks. We reveal very different patterns of expression of members of the interferon-induced transmembrane protein (IFITM) gene family in ducks and chickens. In ducks, IFITM1, 2 and 3 are strongly up regulated in response to highly pathogenic avian influenza, where little response is seen in chickens. Clustering of gene expression profiles suggests IFITM1 and 2 have an anti-viral response and IFITM3 may restrict avian influenza virus through cell membrane fusion. We also show, through molecular phylogenetic analyses, that avian IFITM1 and IFITM3 genes have been subject to both episodic and pervasive positive selection at specific codons. In particular, avian IFITM1 showed evidence of positive selection in the duck lineage at sites known to restrict influenza virus infection. Conclusions: Taken together these results support a model where the IFITM123 protein family and RIG-I all play a crucial role in the tolerance of ducks to highly pathogenic and low pathogenic strains of avian influenza viruses when compared to the chicken.

Description: Background: The co-chaperone Hop [ h eat shock protein (HSP) o rganizing p rotein] has been shown to act as an adaptor for protein folding and maturation, in concert with Hsp70 and Hsp90. The hop gene is of eukaryotic origin. Likewise, the chloroplast elongation factor G (cEF-G) catalyzes the translocation step in chloroplast protein synthesis. The chl-fus gene, which encodes the cEF-G protein, is of plastid origin. Both proteins, Hop and cEF-G, derived from domain duplications. It was demonstrated that the nuclear chl-fus gene locates in opposite orientation to a hop gene in Glycine max. We explored 53 available plant genomes from Chlorophyta to higher plants, to determine whether the chl-fus gene was transferred directly downstream of the primordial hop in the proto-eukaryote host cell. Since both genes came from exon/module duplication events, we wanted to explore the involvement of introns in the early origin and the ensuing evolutionary changes in gene structure. Results: We reconstructed the evolutionary history of the two convergent plant genes, on the basis of their gene structure, microsynteny and microcolinearity, from 53 plant nuclear genomes. Despite a high degree (72 %) of microcolinearity among vascular plants, our results demonstrate that their adjacency was a product of chromosomal rearrangements. Based on predicted exon − intron structures, we inferred the molecular events giving rise to the current form of genes. Therefore, we propose a simple model of exon/module shuffling by intronic recombinations in which phase-0 introns were essential for domain duplication, and a phase-1 intron for transit peptide recruiting. Finally, we demonstrate a natural susceptibility of the intergenic region to recombine or delete, seriously threatening the integrity of the chl-fus gene for the future. Conclusions: Our results are consistent with the interpretation that the chl-fus gene was transferred from the chloroplast to a chromosome different from that of hop, in the primitive photosynthetic eukaryote, and much later before the appearance of angiosperms, it was recombined downstream of hop. Exon/module shuffling mediated by symmetric intron phases (i.e., phase-0 introns) was essential for gene evolution. The intergenic region is prone to recombine, risking the integrity of both genes.

Description: Background: Soil salinity is one of the most significant abiotic stresses affecting plant shoots and roots growth. The adjustment of root architecture to spatio-temporal heterogeneity in salinity is particularly critical for plant growth and survival. Bermudagrass (Cynodon dactylon) is a widely used turf and forage perennial grass with a high degree of salinity tolerance. Salinity appears to stimulate the growth of roots and decrease their mortality in tolerant bermudagrass. To estimate a broad spectrum of genes related to root elongation affected by salt stress and the molecular mechanisms that control the positive response of root architecture to salinity, we analyzed the transcriptome of bermudagrass root tips in response to salinity. Results: RNA-sequencing was performed in root tips of two bermudagrass genotypes contrasting in salt tolerance. A total of 237,850,130 high quality clean reads were generated and 250,359 transcripts were assembled with an average length of 1115 bp. Totally, 103,324 unigenes obtained with 53,765 unigenes (52 %) successfully annotated in databases. Bioinformatics analysis indicated that major transcription factor (TF) families linked to stress responses and growth regulation (MYB, bHLH, WRKY) were differentially expressed in root tips of bermudagrass under salinity. In addition, genes related to cell wall loosening and stiffening (xyloglucan endotransglucosylase/hydrolases, peroxidases) were identified. Conclusions: RNA-seq analysis identified candidate genes encoding TFs involved in the regulation of lignin synthesis, reactive oxygen species (ROS) homeostasis controlled by peroxidases, and the regulation of phytohormone signaling that promote cell wall loosening and therefore root growth under salinity.

Description: Nonlinear thermal and moisture dynamics of high Arctic wetland polygons following permafrost disturbance Biogeosciences Discussions, 12, 11797-11831, 2015 Author(s): E. Godin, D. Fortier, and E. Lévesque Low-centre polygonal terrain developing within gentle sloping surfaces and lowlands in the high Arctic have a potential to retain snowmelt water in their bowl-shaped centre and as such are considered high latitude wetlands. Such wetlands in the continuous permafrost regions have an important ecological role in an otherwise generally arid region. In the valley of the glacier C-79 on Bylot Island (Nunavut, Canada), thermal erosion gullies are rapidly eroding the permafrost along ice wedges affecting the integrity of the polygons by breaching and collapsing the surrounding rims. While intact polygons were characterized by a relative homogeneity (topography, snow cover, maximum active layer thaw depth, ground moisture content, vegetation cover), eroded polygons had a non-linear response for the same elements following their perturbation. The heterogeneous nature of disturbed terrains impacts active layer thickness, ground ice aggradation in the upper portion of permafrost, soil moisture and vegetation dynamics, carbon storage and terrestrial green-house gas emissions.

Description: Controls on microalgal community structures in cryoconite holes upon high Arctic glaciers, Svalbard Biogeosciences Discussions, 12, 11751-11795, 2015 Author(s): T. R. Vonnahme, M. Devetter, J. D. Žárský, M. Šabacká, and J. Elster Glaciers are known to harbor surprisingly complex ecosystems. On their surface, distinct cylindrical holes filled with meltwater and sediments are considered as hot spots for microbial life. The present paper addresses possible biological interactions within the community of prokaryotic cyanobacteria and eukaryotic microalgae (microalgae) and relations to their potential grazers, additional to their environmental controls. Svalbard glaciers with substantial allochthonous input of material from local sources reveal high microalgal densities. Small valley glaciers with high sediment coverages and high impact of birds show high biomasses and support a high biological diversity. Invertebrate grazer densities do not show any significant negative correlation with microalgal abundances, but a positive correlation with eukaryotic microalgae. Most microalgae found in this study form large colonies ( 〈 10 cells, or 〉 25 μm), which may protect them against invertebrate grazing. This finding rather indicates grazing as a positive control on eukaryotic microalgae by nutrient recycling. Density differences between the eukaryotic microalgae and prokaryotic cyanobacteria and their high distinction in RDA and PCA analyses indicate that these two groups are in strong contrast. Eukaryotic microalgae occurred mainly in unstable cryoconite holes with high sediment loads, high N : P ratios, and a high impact of bird guano, as a proxy for nutrients. In these environments autochthonous nitrogen fixation appears to be negligible. Selective wind transport of Oscillatoriales via soil and dust particles is proposed to explain their dominance in cryoconites further away from the glacier margins. We propose that, for the studied glaciers, nutrient levels related to recycling of limiting nutrients is the main factor driving variation in the community structure of microalgae and grazers.

Description: Background: The filamentous cyanobacterium Nostoc sp. strain PCC 7120 can fix N 2 when combined nitrogen is not available. Furthermore, it has to cope with reactive oxygen species generated as byproducts of photosynthesis and respiration. We have previously demonstrated the synthesis of Ser/Thr kinase Pkn22 as an important survival response of Nostoc to oxidative damage. In this study we wished to investigate the possible involvement of this kinase in signalling peroxide stress and nitrogen deprivation. Results: Quantitative RT-PCR experiments revealed that the pkn22 gene is induced in response to peroxide stress and to combined nitrogen starvation. Electrophoretic motility assays indicated that the pkn22 promoter is recognized by the global transcriptional regulators FurA and NtcA. Transcriptomic analysis comparing a pkn22-insertion mutant and the wild type strain indicated that this kinase regulates genes involved in important cellular functions such as photosynthesis, carbon metabolism and iron acquisition. Since metabolic changes may lead to oxidative stress, we investigated whether this is the case with nitrogen starvation. Our results rather invalidate this hypothesis thereby suggesting that the function of Pkn22 under nitrogen starvation is independent of its role in response to peroxide stress. Conclusions: Our analyses have permitted a more complete functional description of Ser/Thr kinase in Nostoc. We have decrypted the transcriptional regulation of the pkn22 gene, and analysed the whole set of genes under the control of this kinase in response to the two environmental changes often encountered by cyanobacteria in their natural habitat: oxidative stress and nitrogen deprivation.

Description: Background: Despite a number of recent reports of insect resistance to transgenic crops expressing insecticidal toxins from Bacillus thuringiensis (Bt), little is known about the mechanism of resistance to these toxins. The purpose of this study is to identify genes associated with the mechanism of Cry1F toxin resistance in European corn borer (Ostrinia nubilalis Hübner). For this, we compared the global transcriptomic response of laboratory selected resistant and susceptible O. nubilalis strain to Cry1F toxin. We further identified constitutive transcriptional differences between the two strains. Results: An O. nubilalis midgut transcriptome of 36,125 transcripts was assembled de novo from 106 million Illumina HiSeq and Roche 454 reads and used as a reference for estimation of differential gene expression analysis. Evaluation of gene expression profiles of midgut tissues from the Cry1F susceptible and resistant strains after toxin exposure identified a suite of genes that responded to the toxin in the susceptible strain (n = 1,654), but almost 20-fold fewer in the resistant strain (n = 84). A total of 5,455 midgut transcripts showed significant constitutive expression differences between Cry1F susceptible and resistant strains. Transcripts coding for previously identified Cry toxin receptors, cadherin and alkaline phosphatase and proteases were also differentially expressed in the midgut of the susceptible and resistant strains. Conclusions: Our current study provides a valuable resource for further molecular characterization of Bt resistance and insect response to Cry1F toxin in O. nubilalis and other pest species.

Description: Thermo-erosion gullies boost the transition from wet to mesic vegetation Biogeosciences Discussions, 12, 12191-12228, 2015 Author(s): N. Perreault, E. Lévesque, D. Fortier, and L. J. Lamarque Continuous permafrost zones with well-developed polygonal ice-wedge networks are particularly vulnerable to climate change. Thermo-mechanical erosion can initiate the development of gullies that lead to substantial drainage of adjacent wet habitats. How vegetation responds to this particular disturbance is currently unknown but has the potential to strongly disrupt function and structure of Arctic ecosystems. Focusing on three major gullies of Bylot Island, Nunavut, we aimed at estimating the effects of thermo-erosion processes in shaping plant community changes. Over two years, we explored the influence of environmental factors on plant species richness, abundance and biomass studying 197 polygons that covered the whole transition from intact wet to disturbed and mesic habitats. While gullying decreased soil moisture by 40 % and thaw front depth by 10 cm in breached polygons, we observed a gradual vegetation shift within five to ten years with mesic habitat plant species such as Arctagrostis latifolia and Salix arctica replacing wet habitat dominant Carex aquatilis and Dupontia fisheri . This transition was accompanied by a five time decrease in graminoid above-ground biomass in mesic sites. Our results illustrate that wetlands are highly vulnerable to thermo-erosion processes that may rapidly promote the decrease of food availability for herbivores and reduce methane emissions of Arctic ecosystems.

Description: Effects of dust additions on phytoplankton growth and DMS production in high CO 2 northeast Pacific HNLC waters Biogeosciences Discussions, 12, 12281-12319, 2015 Author(s): J. Mélançon, M. Levasseur, M. Lizotte, M. Scarratt, J.-É. Tremblay, P. Tortell, G.-P. Yang, G.-Y. Shi, H.-W. Gao, D. M. Semeniuk, M. Robert, M. Arychuk, K. Johnson, N. Sutherland, M. Davelaar, N. Nemcek, A. Peña, and W. Richardson Ocean acidification (OA) is likely to have an effect on the fertilizing potential of desert dust in high-nutrient, low-chlorophyll oceanic regions, either by modifying Fe speciation and bioavailability, or by altering phytoplankton Fe requirements and acquisition. To address this issue, short incubations (4 days) of northeast subarctic Pacific waters enriched with either FeSO 4 or dust, and set at pH 8.0 (in situ) and 7.8 were conducted in August 2010. We assessed the impact of a decrease in pH on dissolved Fe concentration, phytoplankton biomass, taxonomy and productivity, and the production of dimethylsulfide (DMS) and its algal precursor dimethylsulfoniopropionate (DMSP). Chlorophyll a (chl a ) remained unchanged in the controls and doubled in both the FeSO 4 -enriched and dust-enriched incubations, confirming the Fe-limited status of the plankton assemblage during the experiment. In the acidified treatments, a significant reduction (by 16–38 %) of the final concentration of chl a was measured compared to their non-acidified counterparts, and a 15 % reduction in particulate organic carbon (POC) concentration was measured in the dust-enriched acidified treatment compared to the dust-enriched non-acidified treatment. FeSO 4 and dust additions had a fertilizing effect mainly on diatoms and cyanobacteria. Lowering the pH affected mostly the haptophytes, but pelagophyte concentrations were also reduced in some acidified treatments. Acidification did not significantly alter DMSP and DMS concentrations. These results show that dust deposition events in a low-pH iron-limited Northeast subarctic Pacific are likely to stimulate phytoplankton growth to a lesser extent than in today's ocean during the few days following fertilization and point to a low initial sensitivity of the DMSP and DMS dynamics to OA.

Description: Projected climate change impacts on North Sea and Baltic Sea: CMIP3 and CMIP5 model based scenarios Biogeosciences Discussions, 12, 12229-12279, 2015 Author(s): D. Pushpadas, C. Schrum, and U. Daewel Climate change impacts on the marine biogeochemistry and lower trophic level dynamics in the North Sea and Baltic Sea have been assessed using regional downscaling in a number of recent studies. However, most of these where only forced by physical conditions from Global Climate Models (GCMs) and regional downscaling considering the climate change impact on oceanic nutrient conditions from Global Earth System Models (ESMs) are rare and so far solely based on CMIP3-generation climate models. The few studies published show a large range in projected future primary production and hydrodynamic condition. With the addition of CMIP5 models and scenarios, the demand to explore the uncertainty in regional climate change projections increased. Moreover, the question arises how projections based on CMIP5-generation models compare to earlier projections and multi-model ensembles comprising both AR4 and AR5 generation forcing models. Here, we investigated the potential future climate change impacts to the North Sea and the Baltic Sea ecosystem using a coherent regional downscaling strategy based on the regional coupled bio-physical model ECOSMO. ECOSMO was forced by output from different ESMs from both CMIP3 and CMIP5 models. Multi-model ensembles using CMIP3/A1B and CMIP5/RCP4.5 scenarios are examined, where the selected CMIP5 models are the successors of the chosen CMIP3 models. Comparing projected changes with the present day reference condition, all these simulations predicted an increase in Sea Surface Temperature (SST) in both North Sea and Baltic Sea, reduction in sea ice in the Baltic, decrease in primary production in the North Sea and an increase in primary production in the Baltic Sea. Despite these largely consistent results on the direction of the projected changes, our results revealed a broad range in the amplitude of projected climate change impacts. Our study strengthens the claim that the choice of the ESM is a major factor for regional climate projections. The change in oceanic nutrient input appeared to be the major driver for the projected changes in North Sea primary production. Assessing the spread in ensemble groups, we found that there is for the North Sea a significant reduction in the spread of projected changes among CMIP5 forced model simulations compared to those forced by CMIP3 ESMs, except for salinity. The latter was due to an unexpected salinification observed in one of the CMIP5 model while all other models exhibit freshening in the future. However, for the Baltic Sea substantial differences in inter-model variability in projected climate change impact to primary production is lacking.

Description: Background: Trichoderma reesei is used for industry-scale production of plant cell wall-degrading enzymes, in particular cellulases, but also xylanases. The expression of the encoding genes was so far primarily investigated on the level of transcriptional regulation by regulatory proteins. Otherwise, the impact of chromatin remodelling on gene expression received hardly any attention. In this study we aimed to learn if the chromatin status changes in context to the applied conditions (repressing/inducing), and if the presence or absence of the essential transactivator, the Xylanase regulator 1 (Xyr1), influences the chromatin packaging. Results: Comparing the results of chromatin accessibility real-time PCR analyses and gene expression studies of the two prominent cellulase-encoding genes, cbh1 and cbh2, we found that the chromatin opens during sophorose-mediated induction compared to D-glucose-conferred repression. In the strain bearing a xyr1 deletion the sophorose mediated induction of gene expression is lost and the chromatin opening is strongly reduced. In all conditions the chromatin got denser when Xyr1 is absent. In the case of the xylanase-encoding genes, xyn1 and xyn2, the result was similar concerning the condition-specific response of the chromatin compaction. However, the difference in chromatin status provoked by the absence of Xyr1 is less pronounced. A more detailed investigation of the DNA accessibility in the cbh1 promoter showed that the deletion of xyr1 changed the in vivo footprinting pattern. In particular, we detected increased hypersensitivity on Xyr1-sites and stronger protection of Cre1-sites. Looking for the players directly causing the observed chromatin remodelling, a whole transcriptome shotgun sequencing revealed that 15 genes encoding putative chromatin remodelers are differentially expressed in response to the applied condition and two amongst them are differentially expressed in the absence of Xyr1. Conclusions: The regulation of xylanase and cellulase expression in T. reesei is not only restricted to the action of transcription factors but is clearly related to changes in the chromatin packaging. Both the applied condition and the presence of Xyr1 influence chromatin status.

Description: Background: A systems toxicology investigation comparing and integrating transcriptomic and proteomic results was conducted to develop holistic effects characterizations for the wildlife bird model, Northern bobwhite (Colinus virginianus) dosed with the explosives degradation product 2-amino-4,6-dinitrotoluene (2A-DNT). A subchronic 60d toxicology bioassay was leveraged where both sexes were dosed via daily gavage with 0, 3, 14, or 30 mg/kg-d 2A-DNT. Effects on global transcript expression were investigated in liver and kidney tissue using custom microarrays for C. virginianus in both sexes at all doses, while effects on proteome expression were investigated in liver for both sexes and kidney in males, at 30 mg/kg-d. Results: As expected, transcript expression was not directly indicative of protein expression in response to 2A-DNT. However, a high degree of correspondence was observed among gene and protein expression when investigating higher-order functional responses including statistically enriched gene networks and canonical pathways, especially when connected to toxicological outcomes of 2A-DNT exposure. Analysis of networks statistically enriched for both transcripts and proteins demonstrated common responses including inhibition of programmed cell death and arrest of cell cycle in liver tissues at 2A-DNT doses that caused liver necrosis and death in females. Additionally, both transcript and protein expression in liver tissue was indicative of induced phase I and II xenobiotic metabolism potentially as a mechanism to detoxify and excrete 2A-DNT. Nuclear signaling assays, transcript expression and protein expression each implicated peroxisome proliferator-activated receptor (PPAR) nuclear signaling as a primary molecular target in the 2A-DNT exposure with significant downstream enrichment of PPAR-regulated pathways including lipid metabolic pathways and gluconeogenesis suggesting impaired bioenergetic potential. Conclusion: Although the differential expression of transcripts and proteins was largely unique, the consensus of functional pathways and gene networks enriched among transcriptomic and proteomic datasets provided the identification of many critical metabolic functions underlying 2A-DNT toxicity as well as impaired PPAR signaling, a key molecular initiating event known to be affected in di- and trinitrotoluene exposures.

Description: Dissolved organic carbon lability and stable isotope shifts during microbial decomposition in a tropical river system Biogeosciences Discussions, 12, 12761-12782, 2015 Author(s): N. Geeraert, F. O. Omengo, G. Govers, and S. Bouillon A significant amount of carbon is transported to the ocean as dissolved organic carbon (DOC) in rivers. During transport, it can be transformed through microbial consumption and photochemical oxidation. In dark incubation experiments with water from the Tana River, Kenya, we examined the consumption of DOC through microbial decomposition and the associated change in its carbon stable isotope composition (δ 13 C). In 15 of the 18 incubations, DOC concentrations decreased significantly by 10 to 60 %, with most of the decomposition taking place within the first 24–48 h. After 8 days, the remaining DOC was up to 3 ‰ more depleted in 13 C compared with the initial pool, and the change in δ 13 C correlated strongly with the fraction of DOC remaining. We propose that the shift in δ 13 C is consistent with greater microbial lability of DOC originating from herbaceous C 4 vegetation than DOC derived from woody C 3 vegetation in the semi-arid lower Tana. The findings complement earlier data that riverine C sources do not necessarily reflect their proportion in the catchment: besides spatial distribution, also processing within the river can further influence the riverine δ 13 C.

Description: Ocean dynamic processes causing spatially heterogeneous distribution of sedimentary caesium-137 massively released from the Fukushima Dai-ichi Nuclear Power Plant Biogeosciences Discussions, 12, 12713-12759, 2015 Author(s): H. Higashi, Y. Morino, N. Furuichi, and T. Ohara Massive amounts of anthropogenic radiocaesium 137 Cs that was released into the environment by the Fukushima Dai-ichi Nuclear Power Plant accident on March 2011 are widely known to have extensively migrated to Pacific oceanic sediment off of east Japan. Several recent reports have stated that the sedimentary 137 Cs is now stable with a remarkably heterogeneous distribution. The present study elucidates ocean dynamic processes causing this heterogeneous sedimentary 137 Cs distribution in and around the shelf off Fukushima and adjacent prefectures. We performed a numerical simulation of oceanic 137 Cs behaviour for about 10 months after the accident, using a comprehensive dynamic model involving advection–diffusion transport in seawater, adsorption and desorption to and from particulate matter, sedimentation and suspension on and from the bottom, and vertical diffusion transport in the sediment. A notable simulated result was that the sedimentary 137 Cs significantly accumulated in a swath just offshore of the shelf break (along the 50–100 m isobath) as in recent observations, although the seabed in the entire simulation domain was assumed to have ideal properties such as identical bulk density, uniform porosity, and aggregation of particles with a single grain diameter. This result indicated that the heterogeneous sedimentary 137 Cs distribution was not necessarily a result of the spatial distribution of 137 Cs sediment adsorptivity. The present simulation suggests that the shape of the swath is mainly associated with spatiotemporal variation between bottom shear stress in the shallow shelf ( 〈 50 m depths) and that offshore of the shelf break. In a large part of the shallow shelf, the simulation indicated that strong bottom friction suspending particulate matter from the seabed frequently occurred via a periodic spring tide about every 2 weeks and via occasional strong wind. The sedimentary 137 Cs thereby could hardly stay on the surface of the seabed with the result that the simulated sediment-surface 137 Cs activity tended to decrease steadily for a long term after the initial 137 Cs migration. By contrast, in the offshore region, neither the spring tide nor the strong wind caused bottom disturbance. Hence, the particulate matter incorporated with 137 Cs, which was horizontally transported from the adjacent shallow shelf, readily settled and remained on the surface of the sediment just offshore of the shelf break. The present simulation also clearly demonstrated that the bottom disturbance influenced the sedimentary 137 Cs distributions not only horizontally but also vertically. In particular, within a part of the near-shore off the nuclear power plant, the simulation indicated that large amounts of the sedimentary 137 Cs were present in both upper and deeper sediments. As a result, total sedimentary 137 Cs in the entire simulation domain (1.4 x 10 5 km 2 ) at the end of 2011 was 3.2 x 10 15 Bq, more than 10 times that in previous estimates using samples of upper sediments.

Description: Soil N 2 O and NO emissions from land use and land-use change in the tropics and subtropics: a meta-analysis Biogeosciences Discussions, 12, 12783-12821, 2015 Author(s): J. van Lent, K. Hergoualc'h, and L. V. Verchot Deforestation and forest degradation in the tropics may substantially alter soil N-oxide emissions. It is particularly relevant to accurately quantify those changes to properly account for them in a REDD+ climate change mitigation scheme that provides financial incentives to reduce the emissions. With this study we provide updated land use (LU)-based emission rates (103 studies, 387 N 2 O and 111 NO case studies), determine the trend and magnitude of flux changes with land-use change (LUC) using a meta-analysis approach (43 studies, 132 N 2 O and 37 NO cases) and evaluate biophysical drivers of N 2 O and NO emissions and emission changes for the tropics. The average N 2 O and NO emissions in intact upland tropical forest amounted to 2.0 ± 0.2 ( n = 88) and 1.7 ± 0.5 ( n = 36) kg N ha −1 yr −1 , respectively. In agricultural soils annual N 2 O emissions were exponentially related to N fertilization rates and average water-filled pore space (WFPS) whereas in non-agricultural sites a Gaussian response to WFPS fit better the observed NO and N 2 O emissions. The sum of soil N 2 O and NO fluxes and the ratio of N 2 O to NO increased exponentially and significantly with increasing nitrogen availability (expressed as NO 3 − /[NO 3 − +NH 4 + ]) and WFPS, respectively; following the conceptual Hole-In-the-Pipe model. Nitrous and nitric oxide fluxes did not overall increase significantly as a result of LUC (Hedges's d of 0.11 ± 0.11 and 0.16 ± 0.19, respectively), however individual LUC trajectories or practices did. Nitrous oxide fluxes increased significantly after intact upland forest conversion to croplands (Hedges's d = 0.78 ± 0.24) and NO increased significantly following the conversion of low forest cover (secondary forest younger than 30 years, woodlands, shrublands) (Hedges's d of 0.44 ± 0.13). Forest conversion to fertilized systems significantly and highly raised both N 2 O and NO emission rates (Hedges's d of 1.03 ± 0.23 and 0.52 ± 0.09, respectively). Changes in nitrogen availability and WFPS were the main factors explaining changes in N 2 O emissions following LUC, therefore it is important that experimental designs monitor their spatio-temporal variation. Gaps in the literature on N oxide fluxes included geographical gaps (Africa, Oceania) and LU gaps (degraded forest, wetland (notably peat) forest, oil palm plantation and soy cultivation).

Description: Phytoplankton calcification as an effective mechanism to prevent cellular calcium poisoning Biogeosciences Discussions, 12, 12691-12712, 2015 Author(s): M. N. Müller, J. Barcelos e Ramos, K. G. Schulz, U. Riebesell, J. Kaźmierczak, F. Gallo, L. Mackinder, Y. Li, P. N. Nesterenko, T. W. Trull, and G. M. Hallegraeff Marine phytoplankton has developed the remarkable ability to tightly regulate the concentration of free calcium ions in the intracellular cytosol at a level of ~ 0.1 μmol L −1 in the presence of seawater Ca 2+ concentrations of 10 mmol L −1 . The low cytosolic calcium ion concentration is of utmost importance for proper cell signalling function. While the regulatory mechanisms responsible for the tight control of intracellular Ca 2+ concentration are not completely understood, phytoplankton taxonomic groups appear to have evolved different strategies, which may affect their ability to cope with changes in seawater Ca 2+ concentrations in their environment on geological time scales. For example, the Cretaceous (145 to 66 Ma ago), an era known for the high abundance of coccolithophores and the production of enormous calcium carbonate deposits, exhibited seawater calcium concentrations up to four times present-day levels. We show that calcifying coccolithophore species ( Emiliania huxleyi , Gephyrocapsa oceanica and Coccolithus braarudii ) are able to maintain their relative fitness (in terms of growth rate and photosynthesis) at simulated Cretaceous seawater calcium concentrations, whereas these rates are severely reduced under these conditions in some non-calcareous phytoplankton species ( Chaetoceros sp., Ceratoneis closterium and Heterosigma akashiwo ). Most notably, this also applies to a non-calcifying strain of E. huxleyi which displays a calcium-sensitivity similar to the non-calcareous species. We hypothesize that the process of calcification in coccolithophores provides an efficient mechanism to prevent cellular calcium poisoning and thereby offered a potential key evolutionary advantage, responsible for the proliferation of coccolithophores during times of high seawater calcium concentrations.

Description: EUROSPEC: at the interface between remote sensing and ecosystem CO 2 flux measurements in Europe Biogeosciences Discussions, 12, 13069-13121, 2015 Author(s): A. Porcar-Castell, A. Mac Arthur, M. Rossini, L. Eklundh, J. Pacheco-Labrador, K. Anderson, M. Balzarolo, M. P. Martín, H. Jin, E. Tomelleri, S. Cerasoli, K. Sakowska, A. Hueni, T. Julitta, C. J. Nichol, and L. Vescovo Resolving the spatial and temporal dynamics of gross primary productivity (GPP) of terrestrial ecosystems across different scales remains a challenge. Remote sensing is regarded as the solution to upscale point observations conducted at the ecosystem level, using the eddy covariance (EC) technique, to the landscape and global levels. In addition to traditional vegetation indices, the photochemical reflectance index (PRI) and the emission of solar-induced chlorophyll fluorescence (SIF), now measurable from space, provide a new range of opportunities to monitor the global carbon cycle using remote sensing. However, the scale mismatch between EC observations and the much coarser satellite-derived data complicates the integration of the two sources of data. The solution is to establish a network of in situ spectral measurements that can act as bridge between EC measurements and remote sensing data. In situ spectral measurements have been already conducted for many years at EC sites, but using variable instrumentation, setups, and measurement standards. In Europe in particular, in situ spectral measurements remain highly heterogeneous. The goal of EUROSPEC Cost Action ES0930 was to promote the development of common measuring protocols and new instruments towards establishing best practices and standardization of in situ spectral measurements. In this review we describe the background and main tradeoffs of in situ spectral measurements, review the main results of EUROSPEC Cost Action, and discuss the future challenges and opportunities of in situ spectral measurements for improved estimation of local and global carbon cycle.

Description: Improved end-member characterization of modern organic matter pools in the Ohrid Basin (Albania, Macedonia) and evaluation of new palaeoenvironmental proxies Biogeosciences Discussions, 12, 12975-13039, 2015 Author(s): J. Holtvoeth, D. Rushworth, A. Imeri, M. Cara, H. Vogel, T. Wagner, and G. A. Wolff We present elemental, lipid biomarker and compound-specific isotope (δ 13 C, δ 2 H) data for soils and leaf litter collected in the catchment of Lake Ohrid (Albania, Macedonia), as well as macrophytes, particulate organic matter and sediments from the lake itself. Lake Ohrid provides an outstanding archive of continental environmental change of at least 1.2 M years and the purpose of our study is to ground truth organic geochemical proxies that we developed in order to study past changes in the terrestrial biome. We show that soils dominate the lipid signal of the lake sediments rather than the vegetation or aquatic biomass, while compound-specific isotopes (δ 13 C, δ 2 H) determined for n-alkanoic acids confirm a dominant terrestrial source of organic matter to the lake. There is a strong imprint of suberin monomers on the composition of total lipid extracts and chain-length distributions of n-alkanoic acids, n-alcohols, ω-hydroxy acids and α,ω-dicarboxylic acids. Our end-member survey identifies that ratios of mid-chain length suberin-derived to long-chain length cuticular-derived alkyl compounds as well as their average chain length distributions can be used as new molecular proxies of organic matter sources to the lake. We tested these for the 8.2 ka event, a pronounced and widespread Holocene climate fluctuation. In SE Europe climate became drier and cooler in response to the event, as is clearly recognizable in the carbonate and organic carbon records of Lake Ohrid sediments. Our new proxies indicate biome modification in response to hydrological changes, identifying two phases of increased soil OM supply, first from topsoils and then from mineral soils. Our study demonstrates that geochemical fingerprinting of terrestrial OM should focus on the main lipid sources, rather than the living biomass. Both can exhibit climate-controlled variability, but are generally not identical.

Description: The root economics spectrum: divergence of absorptive root strategies with root diameter Biogeosciences Discussions, 12, 13041-13067, 2015 Author(s): D. Kong, J. Wang, P. Kardol, H. Wu, H. Zeng, X. Deng, and Y. Deng Plant roots usually vary along a dominant ecological axis, the root economics spectrum (RES), depicting a tradeoff between resource acquisition and conservation. For absorptive roots, which are mainly responsible for resource acquisition, we hypothesized that root strategies as predicted from the RES shift with increasing root diameter. To test this hypothesis, we used seven contrasting plant species for which we separated absorptive roots into two categories: thin roots ( 〈 247 μm diameter) and thick roots. For each category, we analyzed a~range of root traits closely related to resource acquisition and conservation, including root tissue density, carbon (C) and nitrogen (N) fractions as well as root anatomical traits. The results showed that trait relationships for thin absorptive roots followed the expectations from the RES while no clear trait relationships were found in support of the RES for thick absorptive roots. Our results suggest divergence of absorptive root strategies in relation to root diameter, which runs against a single economics spectrum for absorptive roots.

Description: Millennial changes in North Atlantic oxygen concentrations Biogeosciences Discussions, 12, 12947-12973, 2015 Author(s): B. A. A. Hoogakker, D. J. R. Thornalley, and S. Barker Glacial–interglacial changes in bottom water oxygen concentrations [O 2 ] in the deep Northeast Atlantic have been linked to decreased ventilation relating to changes in ocean circulation and the biological pump (Hoogakker et al., 2015). In this paper we discuss seawater [O 2 ] changes in relation to millennial climate oscillations in the North Atlantic ocean over the last glacial cycle, using bottom water [O 2 ] reconstructions from 2 cores: (1) MD95-2042 from the deep northeast Atlantic (Hoogakker et al., 2015), and (2) ODP 1055 from the intermediate northwest Atlantic. Deep northeast Atlantic core MD95-2042 shows decreased bottom water [O 2 ] during millennial scale cool events, with lowest bottom water [O 2 ] of 170, 144, and 166 ± 17 μmol kg −1 during Heinrich ice rafting events H6, H4 and H1. Importantly, at intermediate core ODP 1055 bottom water [O 2 ] was lower during parts of Marine Isotope Stage 4 and millennial cool events, with lowest values of 179 and 194 μmol kg −1 recorded during millennial cool events C21 and a cool event following Dansgaard–Oeschger event 19. Our reconstructions agree with previous model simulations suggesting that glacial cold events may be associated with lower seawater [O 2 ] across the North Atlantic below ~1 km (Schmittner et al., 2007), although in our reconstructions the changes are less dramatic. The decreases in bottom water [O 2 ] during North Atlantic Heinrich events and earlier cold events at the deep site can be linked to water mass changes in relation to ocean circulation changes, and possibly productivity changes. At the intermediate depth site a strong North Atlantic Intermediate Water cell precludes water mass changes as a cause for decreased bottom water [O 2 ]. Instead we propose that the lower bottom [O 2 ] there can be linked to productivity changes through increased export of organic material from the surface ocean.

Description: Background: Insertions and deletions (indels) are the most abundant form of structural variation in all genomes. Indels have been increasingly recognized as an important source of molecular markers due to high-density occurrence, cost-effectiveness, and ease of genotyping. Coupled with developments in bioinformatics, next-generation sequencing (NGS) platforms enable the discovery of millions of indel polymorphisms by comparing the whole genome sequences of individuals within a species. Results: A total of 1,973,746 unique indels were identified in 345 maize genomes, with an overall density of 958.79 indels/Mbp, and an average allele number of 2.76, ranging from 2 to 107. There were 264,214 indels with polymorphism information content (PIC) values greater than or equal to 0.5, accounting for 13.39 % of overall indels. Of these highly polymorphic indels, we designed primer pairs for 83,481 and 29,403 indels with major allele differences (i.e. the size difference between the most and second most frequent alleles) greater than or equal to 3 and 8 bp, respectively, based on the differing resolution capabilities of gel electrophoresis. The accuracy of our indel markers was experimentally validated, and among 100 indel markers, average accuracy was approximately 90 %. In addition, we also validated the polymorphism of the indel markers. Of 100 highly polymorphic indel markers, all had polymorphisms with average PIC values of 0.54. Conclusions: The maize genome is rich in indel polymorphisms. Intriguingly, the level of polymorphism in genic regions of the maize genome was higher than that in intergenic regions. The polymorphic indel markers developed from this study may enhance the efficiency of genetic research and marker-assisted breeding in maize.

Description: Background: Soybean cyst nematode (SCN, Heterodera glycines Ichinohe) is one of the most fatal pests of soybean (Glycine max (L.) Merr.) worldwide and causes huge loss of soybean yield each year. Multiple sources of resistance are urgently needed for effective management of SCN via the development of resistant cultivars. The aim of the present study was to investigate the genetic architecture of resistance to SCN HG Type 0 (race 3) and HG Type 1.2.3.5.7 (race 4) in landraces and released elite soybean cultivars mostly from China. Results: A total of 440 diverse soybean landraces and elite cultivars were screened for resistance to SCN HG Type 0 and HG Type 1.2.3.5.7. Exactly 131 new sources of SCN resistance were identified. Lines were genotyped by SNP markers detected by the Specific Locus Amplified Fragment Sequencing (SLAF-seq) approach. A total of 36,976 SNPs were identified with minor allele frequencies (MAF) 〉 4 % that were present in 97 % of all the genotypes. Genome-wide association mapping showed that a total of 19 association signals were significantly related to the resistance for the two HG Types. Of the 19 association signals, eight signals overlapped with reported QTL including Rhg1 and Rhg4 genes. Another eight were located in the linked regions encompassing known QTL. Three QTL were found that were not previously reported. The average value of female index (FI) of soybean accessions with resistant alleles was significantly lower than those with susceptible alleles for each peak SNP. Disease resistance proteins with leucine rich regions, cytochrome P450s, protein kinases, zinc finger domain proteins, RING domain proteins, MYB and WRKY transcription activation families were identified. Such proteins may participate in the resistant reaction to SCN and were frequently found in the tightly linked genomic regions of the peak SNPs. Conclusions: GWAS extended understanding of the genetic architecture of SCN resistance in multiple genetic backgrounds. Nineteen association signals were obtained for the resistance to the two Hg Types of SCN. The multiple beneficial alleles from resistant germplasm sources will be useful for the breeding of cultivars with improved resistance to SCN. Analysis of genes near association signals may facilitate the recognition of the causal gene(s) underlying SCN resistances.

Description: Background: Haloquadratum walsbyi represents up to 80 % of cells in NaCl-saturated brines worldwide, but is notoriously difficult to maintain under laboratory conditions. In order to establish the extent of genetic diversity in a natural population of this microbe, we screened a H. walsbyi enriched metagenomic fosmid library and recovered seven novel version of its cell-wall associated genomic island. The fosmid inserts were sequenced and analysed. Results: The novel cell-wall associated islands delineated two major clades within H. walsbyi. The islands predominantly contained genes putatively involved in biosynthesis of surface layer, genes encoding cell surface glycoproteins and genes involved in envelope formation. We further found that these genes are maintained in the population and that the diversity of this region arises through homologous recombination but also through the action of mobile genetic elements, including viruses. Conclusions: The population of H. walsbyi in the studied saltern brine is composed of numerous clonal lineages that differ in surface structures including the cell wall. This type of variation probably reflects a number of mechanisms that minimize the infection rate of predating viruses.

Description: Background: The maturation of the brain involves the coordinated expression of thousands of genes, proteins and regulatory elements over time. In sensory pathways, gene expression profiles are modified by age and sensory experience in a manner that differs between brain regions and cell types. In the auditory system of altricial animals, neuronal activity increases markedly after the opening of the ear canals, initiating events that culminate in the maturation of auditory circuitry in the brain. This window provides a unique opportunity to study how gene expression patterns are modified by the onset of sensory experience through maturity. As a tool for capturing these features, next-generation sequencing of total RNA (RNAseq) has tremendous utility, because the entire transcriptome can be screened to index expression of any gene. To date, whole transcriptome profiles have not been generated for any central auditory structure in any species at any age. In the present study, RNAseq was used to profile two regions of the mouse auditory forebrain (A1, primary auditory cortex; MG, medial geniculate) at key stages of postnatal development (P7, P14, P21, adult) before and after the onset of hearing (~P12). Hierarchical clustering, differential expression, and functional geneset enrichment analyses (GSEA) were used to profile the expression patterns of all genes. Selected genesets related to neurotransmission, developmental plasticity, critical periods and brain structure were highlighted. An accessible repository of the entire dataset was also constructed that permits extraction and screening of all data from the global through single-gene levels. To our knowledge, this is the first whole transcriptome sequencing study of the forebrain of any mammalian sensory system. Although the data are most relevant for the auditory system, they are generally applicable to forebrain structures in the visual and somatosensory systems, as well. Results: The main findings were: (1) Global gene expression patterns were tightly clustered by postnatal age and brain region; (2) comparing A1 and MG, the total numbers of differentially expressed genes were comparable from P7 to P21, then dropped to nearly half by adulthood; (3) comparing successive age groups, the greatest numbers of differentially expressed genes were found between P7 and P14 in both regions, followed by a steady decline in numbers with age; (4) maturational trajectories in expression levels varied at the single gene level (increasing, decreasing, static, other); (5) between regions, the profiles of single genes were often asymmetric; (6) GSEA revealed that genesets related to neural activity and plasticity were typically upregulated from P7 to adult, while those related to structure tended to be downregulated; (7) GSEA and pathways analysis of selected functional networks were not predictive of expression patterns in the auditory forebrain for all genes, reflecting regional specificity at the single gene level. Conclusions: Gene expression in the auditory forebrain during postnatal development is in constant flux and becomes increasingly stable with age. Maturational changes are evident at the global through single gene levels. Transcriptome profiles in A1 and MG are distinct at all ages, and differ from other brain regions. The database generated by this study provides a rich foundation for the identification of novel developmental biomarkers, functional gene pathways, and targeted studies of postnatal maturation in the auditory forebrain.

Description: Background: Root-colonizing fluorescent pseudomonads are known for their excellent abilities to protect plants against soil-borne fungal pathogens. Some of these bacteria produce an insecticidal toxin (Fit) suggesting that they may exploit insect hosts as a secondary niche. However, the ecological relevance of insect toxicity and the mechanisms driving the evolution of toxin production remain puzzling. Results: Screening a large collection of plant-associated pseudomonads for insecticidal activity and presence of the Fit toxin revealed that Fit is highly indicative of insecticidal activity and predicts that Pseudomonas protegens and P. chlororaphis are exclusive Fit producers. A comparative evolutionary analysis of Fit toxin-producing Pseudomonas including the insect-pathogenic bacteria Photorhabdus and Xenorhadus, which produce the Fit related Mcf toxin, showed that fit genes are part of a dynamic genomic region with substantial presence/absence polymorphism and local variation in GC base composition. The patchy distribution and phylogenetic incongruence of fit genes indicate that the Fit cluster evolved via horizontal transfer, followed by functional integration of vertically transmitted genes, generating a unique Pseudomonas-specific insect toxin cluster. Conclusions: Our findings suggest that multiple independent evolutionary events led to formation of at least three versions of the Mcf/Fit toxin highlighting the dynamic nature of insect toxin evolution.

Description: Background: In complex Metazoans a given gene frequently codes for multiple protein isoforms, through processes such as alternative splicing. Large scale functional annotation of these isoforms is a key challenge for functional genomics. This annotation gap is increasing with the large numbers of multi transcript genes being identified by technologies such as RNASeq. Furthermore attempts to characterise the functions of splicing in an organism are complicated by the difficulty in distinguishing functional isoforms from those produced by splicing errors or transcription noise. Tools to help prioritise candidate isoforms for testing are largely absent. Results: In this study we implement a Time-course Switch (TS) score for ranking isoforms by their likelihood of producing additional functions based on their developmental expression profiles, as reported by modENCODE. The TS score allows us to better investigate functional roles of different isoforms expressed in multi transcript genes. From this analysis, we find that isoforms with high TS scores have sequence feature changes consistent with more deterministic splicing and functional changes and tend to gain domains or whole exons which could carry additional functions. Furthermore these functions appear to be particularly important for essential regulatory roles, establishing functional isoform switching as key for regulatory processes. Based on the TS score we develop a Transcript Annotations Pipeline for Alternative Splicing (TAPAS) that identifies functional neighbourhoods of potentially interesting isoforms. Conclusions: We have identified a subset of protein isoforms which appear to have high functional significance, particularly in regulation. This has been made possible through the development of novel methods that make use of transcript expression profiles.The methods and analyses we present here represent important first steps in the development of tools to address the near complete lack of isoform specific function annotation. In turn the tools allow us to better characterise the regulatory functions of alternative splicing in more detail.

Description: The carbon cycle in Mexico: past, present and future of C stocks and fluxes Biogeosciences Discussions, 12, 12501-12541, 2015 Author(s): G. Murray-Tortarolo, P. Friedlingstein, S. Sitch, V. J. Jaramillo, F. Murguía-Flores, A. Anav, Y. Liu, A. Arneth, A. Arvanitis, A. Harper, A. Jain, E. Kato, C. Koven, B. Poulter, B. D. Stocker, A. Wiltshire, S. Zaehle, and N. Zeng We modelled the carbon (C) cycle in Mexico with a process-based approach. We used different available products (satellite data, field measurements, models and flux towers) to estimate C stocks and fluxes in the country at three different time frames: present (defined as the period 2000–2005), the past century (1901–2000) and the remainder of this century (2010–2100). Our estimate of the gross primary productivity (GPP) for the country was 2137 ± 1023 Tg C yr −1 and a total C stock of 34 506 ± 7483 Tg C, with 20 347 ± 4622 Pg C in vegetation and 14 159 ± 3861 in the soil. Contrary to other current estimates for recent decades, our results showed that Mexico was a C sink over the period 1990–2009 (+31 Tg C yr −1 ) and that C accumulation over the last century amounted to 1210 ± 1040 Tg C. We attributed this sink to the CO 2 fertilization effect on GPP, which led to an increase of 3408 ± 1060 Tg C, while both climate and land use reduced the country C stocks by −458 ± 1001 and −1740 ± 878 Tg C, respectively. Under different future scenarios the C sink will likely continue over 21st century, with decreasing C uptake as the climate forcing becomes more extreme. Our work provides valuable insights on relevant driving processes of the C-cycle such as the role of drought in marginal lands (e.g. grasslands and shrublands) and the impact of climate change on the mean residence time of C in tropical ecosystems.

Description: Influence of timing of sea ice retreat on phytoplankton size during marginal ice zone bloom period in the Chukchi and Bering shelves Biogeosciences Discussions, 12, 12611-12651, 2015 Author(s): A. Fujiwara, T. Hirawake, K. Suzuki, L. Eisner, I. Imai, S. Nishino, T. Kikuchi, and S. I. Saitoh Timing of sea ice retreat (TSR) as well as cell size of primary producers (i.e., phytoplankton) plays crucial roles in seasonally ice-covered marine ecosystem. Thus, it is important to monitor the temporal and spatial distribution of phytoplankton community size structure. Prior to this study, an ocean color algorithm has been developed to derive phytoplankton size index F L , which is defined as the ratio of chlorophyll a derived from the cells larger than 5 μm to the total chl a using satellite remote sensing for the Chukchi and Bering shelves. Using this method, we analyzed pixel-by-pixel relationships between F L during marginal ice zone (MIZ) bloom period and TSR over a period of 1998–2013. The influence of TSR on sea surface temperature (SST) and changes in ocean heat content (ΔOHC) during the MIZ bloom period were also investigated. A significant negative relationship between F L and TSR was widely found in the shelf region during MIZ bloom season. On the other hand, we found a significant positive (negative) relationship between SST (ΔOHC) and TSR. That is, earlier sea-ice retreat was associated with a dominance of larger phytoplankton during a colder and weakly stratified MIZ bloom season, suggesting that duration of nitrate supply, which is important for large-sized phytoplankton growth in this region (i.e., diatoms), can change according to TSR. In addition, under-ice phytoplankton blooms are likely to occur in years with late ice retreat, because sufficient light for phytoplankton growth can pass through the ice and penetrate into the water columns due to an increase in solar radiation toward the summer solstice. Moreover, we found not only the length of ice-free season but also annual median of F L positively correlated with annual net primary production (APP). Thus, both phytoplankton community composition and growing season are important for APP in the study area. Our findings showed quantitative relationship between the inter-annual variability of F L , TSR and APP suggesting satellite remote sensing of phytoplankton community size structure is suitable to document the impact of recent rapid sea ice loss on ecosystem of the study region.

Description: Background: PCR amplicon sequencing has been widely used as a targeted approach for both DNA and RNA sequence analysis. High multiplex PCR has further enabled the enrichment of hundreds of amplicons in one simple reaction. At the same time, the performance of PCR amplicon sequencing can be negatively affected by issues such as high duplicate reads, polymerase artifacts and PCR amplification bias. Recently researchers have made some good progress in addressing these shortcomings by incorporating molecular barcodes into PCR primer design. So far, most work has been demonstrated using one to a few pairs of primers, which limits the size of the region one can analyze. Results: We developed a simple protocol, which enables the use of molecular barcodes in high multiplex PCR with hundreds of amplicons. Using this protocol and reference materials, we demonstrated the applications in accurate variant calling at very low fraction over a large region and in targeted RNA quantification. We also evaluated the protocol’s utility in profiling FFPE samples. Conclusions: We demonstrated the successful implementation of molecular barcodes in high multiplex PCR, with multiplex scale many times higher than earlier work. We showed that the new protocol combines the benefits of both high multiplex PCR and molecular barcodes, i.e. the analysis of a very large region, low DNA input requirement, very good reproducibility and the ability to detect as low as 1 % mutations with minimal false positives (FP).

Description: Background: Whole genome sequencing provides the most comprehensive collection of an organism’s genetic information. The availability of complete genome sequences is expected to dramatically deliver a high impact on biology. However, to achieve this impact in the area of crop improvement, significant efforts are still required on functional genomics, including the areas of gene annotation, cloning, expression profiling, and functional validation. Results: Here we report our efforts in generating the first transcription factor (TF) open reading frame (ORF)eome resource associated with drought resistance in soybean (Glycine max), a major oil/protein crop grown worldwide. This study provides a highly annotated soybean TF-ORFeome associated with drought resistance. It contains information from experimentally verified protein-coding sequences (CDS), expression profiling under several abiotic stresses (drought, salinity, dehydration and ABA), and computationally predicted protein subcellular localization and cis-regulatory elements (CREs) analysis. All the information is available to plant researchers through a freely accessible and user-friendly database, Soybean Knowledge Base (SoyKB). Conclusions: The soybean TF-ORFeome provides a valuable public resource for functional genomics studies, especially in the area of plant abiotic stresses. It will accelerate findings in the areas of abiotic stresses and lead to the generation of crops with enhanced resistance to multiple stresses.

Description: Background: Identifying the transcription start sites (TSS) of genes is essential for characterizing promoter regions. Several protocols have been developed to capture the 5′ end of transcripts via Cap Analysis of Gene Expression (CAGE) or linker-ligation strategies such as Paired-End Analysis of Transcription Start Sites (PEAT), but often require large amounts of tissue. More recently, nanoCAGE was developed for sequencing on the Illumina GAIIx to overcome these difficulties. Results: Here we present the first publicly available adaptation of nanoCAGE for sequencing on recent ultra-high throughput platforms such as Illumina HiSeq-2000, and CapFilter, a computational pipeline that greatly increases confidence in TSS identification. We report excellent gene coverage, reproducibility, and precision in transcription start site discovery for samples from Arabidopsis thaliana roots. Conclusion: nanoCAGE-XL together with CapFilter allows for genome wide identification of high confidence transcription start sites in large eukaryotic genomes.

Description: Southwestern Tropical Atlantic coral growth response to atmospheric circulation changes induced by ozone depletion in Antarctica Biogeosciences Discussions, 12, 13193-13213, 2015 Author(s): H. Evangelista, I. Wainer, A. Sifeddine, T. Corrège, R. C. Cordeiro, S. Lamounier, D. Godiva, C.-C. Shen, F. Le Cornec, B. Turcq, C. E. Lazareth, and C.-Y. Hu Climate changes induced by stratospheric ozone depletion over Antarctica have been recognized as an important consequence of the recently observed Southern Hemisphere atmospheric circulation. Here we present evidences that the Brazilian coast (Southwestern Atlantic) may have been impacted from both winds and sea surface temperature changes derived from this process. Skeleton analysis of massive coral species living in shallow waters off Brazil are very sensitive to air–sea interactions, and seem to record this impact. Growth rates of Brazilian corals show a trend reversal that fits the ozone depletion evolution, confirming that ozone impacts are far reaching and potentially affect coastal ecosystems in tropical environments.

Description: Contrasting pH buffering patterns in neutral-alkaline soils along a 3600 km transect in northern China Biogeosciences Discussions, 12, 13215-13240, 2015 Author(s): W. Luo, P. N. Nelson, M.-H. Li, J. Cai, Y. Zhang, Y. Zhang, Y. Shan, R. Wang, X. Han, and Y. Jiang Soil pH buffering capacity (pHBC) plays a crucial role in predicting acidification rates, yet its large-scale patterns and controls are poorly understood, especially for neutral-alkaline soils. Here, we evaluated the spatial patterns and drivers of pHBC along a 3600 km long transect (1900 km sub-transect with carbonate containing soils and 1700 km sub-transect with non-carbonate containing soils) across northern China. Soil pHBC was greater in the carbonate containing soils than in the non-carbonate containing soils. Acid addition decreased soil pH in the non-carbonate containing soils more markedly than in the carbonate containing soils. Within the carbonate soil sub-transect, soil pHBC was positively correlated with cation exchange capacity (CEC), carbonate content and exchangeable sodium (Na) concentration, but negatively correlated with initial pH and clay content, and not correlated with soil organic carbon (SOC) content. Within the non-carbonate sub-transect, soil pHBC was positively related to initial pH, clay content, CEC and exchangeable Na concentration, but not related to SOC content. Carbonate content was the primary determinant of pHBC in the carbonate containing soils and CEC was the main determinant of buffering capacity in the non-carbonate containing soils. Soil pHBC was positively related to aridity index and carbonate content across the carbonate containing soil sub-transect. Our results indicated that mechanisms controlling pHBC differ among neutral-alkaline soils of northern China, especially between carbonate and non-carbonate containing soils, leading to different rates, risks, and impacts of acidification. This understanding should be incorporated into the acidification risk assessment and landscape management in a changing world.