Project description:The spring bloom in the North Atlantic develops over a few weeks in response to the physical stabilization of the nutrient replete water column and is one of the biggest biological signals on earth. The composition of the phytoplankton assemblage during the spring bloom of 2008 was evaluated, using a microarray, on the basis of functional genes that encode key enzymes in nitrogen and carbon assimilation in eukaryotic and prokaryotic phytoplankton. Oligonucleotide archetype probes representing RuBisCO, nitrate reductase and nitrate transporter genes from major phytoplankton classes detected a diverse assemblage. For RuBisCO, the archetypes with strongest signals represented known phytoplankton groups, but for the nitrate related genes, the major signals were not closely related to any known phytoplankton sequences. Most of the assemblage's components exhibited consistent temporal/spatial patterns. Yet, the strongest archetype signals often showed quite different patterns, indicating different ecological responses by the main players. The most abundant phytoplankton genera identified previously by microscopy, however, were not well represented on the microarray. The lack of sequence data for well-studied species, and the inability to identify organisms associated with functional gene sequences in the environment, still limits our understanding of phytoplankton ecology even in this relatively well-studied system.
Project description:Background: Salmonid species have followed markedly divergent evolutionary trajectories in their interactions with sea lice. While sea lice parasitism poses significant economic, environmental, and animal welfare challenges for Atlantic salmon (Salmo salar) aquaculture, coho salmon (Oncorhynchus kisutch) exhibit near-complete resistance to sea lice, achieved through a potent epithelial hyperplasia response leading to rapid louse detachment. The molecular mechanisms underlying these divergent responses to sea lice are unknown. Results: We characterised the cellular and molecular responses of Atlantic salmon and coho salmon to sea lice using single-nuclei RNA sequencing. Juvenile fish were exposed to copepodid sea lice (Lepeophtheirus salmonis), and lice-attached pelvic fin and skin samples were collected 12h, 24h, 36h, 48h, and 60h after exposure, along with control samples. Comparative analysis of control and treatment samples revealed an immune and wound-healing response that was common to both species, but attenuated in Atlantic salmon, potentially reflecting greater sea louse immunomodulation. Our results revealed unique but complementary roles of three layers of keratinocytes in the epithelial hyperplasia response leading to rapid sea lice rejection in coho salmon. Our results suggest that basal keratinocytes direct the expansion and mobility of intermediate and, especially, superficial keratinocytes, which eventually encapsulate the parasite. Conclusions: Our results highlight the key role of keratinocytes in coho salmon’s sea lice resistance, and the diverged biological response of the two salmonid host species when interacting with this parasite. This study has identified key pathways and candidate genes that could be manipulated using various biotechnological solutions to improve Atlantic salmon sea lice resistance.
Project description:Time-series monitoring of potentially toxic diatoms of the genus Pseudo-nitzschia in Tokyo Bay, Japan with massively parallel sequencing technology
Project description:Mammalian feces can be collected non-invasively during field research and provides valuable information on the ecology and evolution of the host individuals. Undigested food objects, genome/metagenome, steroid hormones, and stable isotopes obtained from fecal samples provide evidence on diet, host/symbiont genetics, and physiological status of the individuals. However, proteins in mammalian feces have hardly been studied, which hampers the molecular investigations into the behavior and physiology of the host individuals. Here, we apply mass spectrometry-based proteomics to fecal samples (n = 10) that were collected from infant, juvenile, and adult captive Japanese macaques (Macaca fuscata) to describe the proteomes of the host, food, and intestinal microbes. The results show that fecal proteomics is a useful method to investigate dietary changes along with breastfeeding and weaning, to reveal the organ/tissue and taxonomy of dietary items, and to estimate physiological status inside intestinal tracts. These types of insights are difficult or impossible to obtain through other molecular approaches. Most mammalian species are facing extinction risk and there is an urgent need to obtain knowledge on their ecology and evolution for better conservation strategy. The fecal proteomics framework we present here is easily applicable to wild settings and other mammalian species, and provides direct evidence of their behavior and physiology.
Project description:Sequencing the metatranscriptome can provide information about the response of organisms to varying environmental conditions. We present a methodology for obtaining random whole-community mRNA from a complex microbial assemblage using Pyrosequencing. The metatranscriptome had, with minimum contamination by ribosomal RNA, significant coverage of abundant transcripts, and included significantly more potentially novel proteins than in the metagenome. Keywords: metatranscriptome, mesocosm, ocean acidification This experiment is part of a much larger experiment. We have produced 4 454 metatranscriptomic datasets and 6 454 metagenomic datasets. These were derived from 4 samples. The experiment is an ocean acidification mesocosm set up in a Norwegian Fjord in 2006. We suspended 6 bags containing 11,000 L of sea water in a Coastal Fjord and then we bubbled CO2 through three of these bags to simulate ocean acidification conditions in the year 2100. The other three bags were bubbled with air. We then induced a phytoplankton bloom in all six bags and took measurements and performed analyses of phytoplankton, bacterioplankton and physiochemical characteristics over a 22 day period. We took water samples from the peak of the phytoplankton bloom and following the decline of the phytoplankton bloom to analyses using 454 metagenomics and 454 metatranscriptomics. Day 1, High CO2 Bag and Day 1, Present Day Bag, refer to the metatranscriptomes from the peak of the bloom. Day 2, High CO2 Bag and Day 2, Present Day Bag, refer to the metatranscriptomes following the decline of the bloom. Obviously High CO2 refers to the ocean acidification mesocosm and Present Day refers to the control mesocosm. Raw data for both the metagenomic and metatranscriptomic components are available at NCBI's Short Read Archive at ftp://ftp.ncbi.nlm.nih.gov/sra/Studies/SRP000/SRP000101
Project description:<p>Phaeocystis pouchetii (Hariot) Lagerheim, 1893 regularly dominates phytoplankton blooms in the Arctic. Through zooplankton grazing and microbial activity, it is considered to be a key resource for the entire marine food web but the actual relevance of biomass transfer to higher trophic levels is still under discussion. Cell physiology and algal nutritional state are suggested to be major factors controlling the observed variability in zooplankton grazing. However, no data have so far yielded insights into the metabolic state of Phaeocystis populations that would allow testing this hypothesis. Therefore, endometabolic markers of different growth phases were determined in laboratory batch cultures using comparative metabolomics and quantified in different phytoplankton blooms in the field. Metabolites, produced during exponential, early and late stationary growth of P. pouchetii were profiled using gas chromatography-mass spectrometry. Then, metabolites were characterized that correlate with the growth phases using multivariate statistical analysis. Free amino acids characterized exponential growth, whereas the early stationary phase was correlated with sugar alcohols, mono- and disaccharides. In the late stationary phase free fatty acids, sterols and terpenes increased. These marker metabolites were then traced in Phaeocystis blooms during a cruise in the Barents Sea and North Norwegian fjords. About 50 endometabolites of P. pouchetii were detected in natural phytoplankton communities. Their relative abundances at Phaeocystis-dominated stations differed from diatom-dominated stations. Mannitol, scyllo- inositol, 24-methylcholesta-5,22-dien-3β-ol, and several free fatty acids were characteristic for Phaeocystis-dominated blooms. Distinct metabolic profiles were detected in the nutrient- depleted community in the inner Porsangerfjord (<0.5 μM NO3-, <0.1 μM PO4-), with high relative amounts of free mono- and disaccharides indicative for a limited culture. This study, therefore, shows how variable physiology of phytoplankton can alter the metabolic landscape of entire plankton communities.</p>
Project description:Poplars are known to be highly tolerant species to boron toxicity and accumulation. However, genes and molecular networks responsible in boron toxicity tolerance have not been investigated yet. Therefore, we performed a pot experiment with 20 black poplar clones collected from the vicinity of boron mines and polluted areas to investigate its potential role in phytoremediation and to select the most boron toxicity tolerant genotype. Trees were treated with irrigation water containing seven elevated boron concentrations from 0 to 160 ppm. Then a microarray based comparative transcriptome profiling was conducted to identify boron toxicity regulated genes responsible in defence responses of black poplar. The results of the study indicated that black poplar is quite suitable for phytoremediation of boron pollution. It could resist 15 ppm soil B content and < 1600 mg/kg boron accumulation in leaves which are highly toxic concentrations for almost all agricultural plants. Transcriptomics results of study revealed totally 1625 and 1419 altered probe sets under boron toxicity in leaf and root tissues, respectively. The highest induction were recorded for the probes sets annotated to tyrosine aminotransferase, ATP binding cassette transporters, glutathione S transferases and metallochaperone proteins. Strong up regulation of these genes attributed to internal excretion of boron into the cell vacuole and existence of detoxification processes in black poplar. Many candidate genes functional in signalling, gene regulation, antioxidation, boron uptake, transport and detoxification processes were also identified in the current study. This is the first transcriptomic study identifying boron toxicity regulated poplar genes and their potential role in boron toxicity tolerance. Total RNA used in microarray experiment was isolated from the leaves and roots of black poplar clone; N.92.237 which accumulated the highest amount of boron its tissues. Total RNA used in the microarray experiment was isolated from leaves and roots of three black poplar saplings grown in ~ 2 ppm (control) and ~ 15 ppm (toxic) soil B contents. RNA isolation was made according to Lithium chloride precipitation method described in Chang et al. (1993). These three isolated RNAs (biological replicates) for each tissue loaded onto three Affymetrix poplar Gene Chips (technical replicates). Totally, 12 GeneChips (2 tissues Ã? 2 different B treatment Ã? 3 biological replicates) were used for transcriptional analysis.
Project description:Staphylococcus aureus, a gram-positive bacterium, causes food poisoning and toxic shock syndrome through the production of superantigenic toxins known as Staphylococcal enterotoxins serotypes A-J (SEA, SEB, etc.) and toxic shock syndrome toxin-1 (TSST-1). A subset of these toxins have been classified as potential biothreat agents. The chronology of molecular events that could potentially characterize superantigenic toxicity and early pathogenesis is not well understood. The focus of this study was to determine the distinct and shared mechanisms of response to three toxins of the superantigenic family, namely SEA, SEB and TSST-1. Since skin functions as the front line of the host’s defense mechanism, melanocytes were selected for the study and treated with 25 µg/mL of one of these three toxins. Cells were collected after treatment for six different time periods, ranging from 0.5 h to 48 h. Total RNA was investigated using gene expression microarrays containing approximately 50,000 probes, and a subset of the results were validated using NanoString assays. Transcriptomic expression data indicated that each of these three toxins had a unique longitudinal trajectory. In particular, the gene expression profiles of SEB post-exposure was very distinct from those for SEA and TSST-1 superantigens. All three superantigens showed enriched biological networks related to necrosis, skin disorders, and inflammation. The three superantigens share some similarities in acting on the mechanisms underlying apoptosis, innate immunity, and other biological processes. Pathways related to innate immunity, such as the patterns of cytokine production and acute-phase response, showed toxin-specific regulation. The differentially regulated networks can be logical targets for early therapeutic intervention and can potentially serve as early diagnostic markers for superantigen-induced toxicity.