Project description:This study investigates the baseline or inducible differences in between populations of Atlantic salmon lice Lepeophtheirus salmonis with differing levels of resistance to the parasiticidal drug emamectin benzoate (EMB), as well as the induced effects of EMB exposure to Pacific salmon lice. F1 generation lice were exposed in bioassays to a dilution series of emamectin benzoate. Two separate experiments were conducted, one for Atlantic and one for Pacific salmon lice (to be analyzed separately). Atlantic pre-adult salmon lice, separated into male and female, and sensitive or resistant to EMB populations, and exposed to a dilution series: 0 (control), 0.1, 25, 300, and 1000 parts per billion EMB. For each combination four biological replicates were included, except male resistant 25 (n = 3) and female resistant 300 (n = 2). Pacific pre-adult lice of both sexes were exposed to a dilution series: 0 (control), 25, 50 parts per billion EMB.
Project description:Global metaproteomic analyses of microbial biomass from the upper water column of the Central Pacific Ocean. This dataset was used as a discovery dataset to identify peptide biomarkers for cyanobacterial populations for use in targeted metaproteomic calibrated multiple reaction monitoring (MRM) analyses published in in Saito et al., 2014 and 2015. Saito, M. A., McIlvin, M. R., Moran, D. M., Goepfert, T. J., DiTullio, G. R., Post, A. F., and Lamborg, C. H.: Multiple nutrient stresses at intersecting Pacific Ocean biomes detected by protein biomarkers, Science, 345, 1173-1177, 2014. Saito, M. A., Dorsk, A., Post, A. F., McIlvin, M., Rappé, M. S., DiTullio, G., and Moran, D.: Needles in the Blue Sea: Sub‐Species Specificity in Targeted Protein Biomarker Analyses Within the Vast Oceanic Microbial Metaproteome, PROTEOMICS, 15, 3521-3531, 2015.
Project description:This study investigates sex-biased gene expression between populations of Atlantic and Pacific salmon lice, Lepeophtheirus salmonis. Two Atlantic L. salmonis populations were previously used for an array study (GSE56024) while a third dataset using Pacific L. salmonis was novel. Using all three populations, a consensus-based, meta-analysis approach was used to identify sex-biased and sex-specific genes. Two separate experiments were conducted, one for Atlantic and one for Pacific salmon lice. As the Atlantic data has been previously published for other comparisons (GSE56024), only the Pacific data is uploaded here. Lice from three populations (2 in the Atlantic and 1 in the Pacific) were collected for in vitro bioassay analysis using emamectin benzoate. After 24hrs, lice were collected as per treatment protocol below. Males and females from all populations were compared separately before forming a consensus probe list of sex-biased genes concordantly expressed across all three populations. Please note that each raw data file contains three or four 'block' data and each block data correspond to individual sample raw data. Therefore, each raw data file contains raw data for 3-4 samples (as indicated in the description field).
Project description:Metaproteomics is a powerful analytical approach that can assess the functional capabilities deployed by microbial communi-ties in both environmental and biomedical microbiome settings. Yet the mass spectra resulting from these mixed biological communities are challenging due to the high number of low intensity peak features. The use of multiple dimensions of chroma-tographic separation prior to mass spectrometry analyses has been applied to proteomics previously but can require increased sampling handling and instrument time. Here we demonstrate an automated online comprehensive active-modulation two-dimensional liquid chromatography method for metaproteome sample analysis. A high pH PLRP-S column was used in the first dimension followed by low pH separation in the second dimension using dual modulating C18 traps and a C18 column. This method increased the number of unique peptides found in ocean metaproteome samples by more than 50% when com-pared to a one dimension separation while using the same amount of sample and instrument time.
Project description:In this study the impact of protein fractionation techniques prior to LC/MS analysis was investigated on activated sludge samples derived at winter and summer condition from a full-scale wastewater treatment plant (WWTP). For reduction of the sample complexity, different fractionation techniques including RP-LC (1D-approach), SDS-PAGE and RP-LC (2D-approach) as well as RP-LC, SDS-PAGE and liquid IEF (3D-approach) were carried out before subsequent ITMS analysis. The derived spectra were identified by MASCOT search using a combination of the public UniProtKB/Swiss-Prot protein database and metagenome data from a WWTP (data are available via ProteomeXchange with identifier PXD001547). The results showed a significant increase of identified spectra, enabled by applying IEF and SDS-PAGE to the proteomic workflow. Based on meta-proteins, a core metaproteome and the corresponding taxonomic profile of the wastewater activated sludge was revealed and functional aspects using the KEGG (Kyoto Encyclopedia of Genes and Genomes) pathway library were analyzed. Hereby, the KEGG Orthology identifiers (KO numbers) of protein hits were plotted into pathway maps of the central carbon (map01200) and nitrogen metabolism (map00910). Using the 3D-approach, most proteins involved in glycolysis and citrate cycle and nearly all proteins of the nitrogen removal were identified, qualifying this approach as the most promising for future studies.
Project description:Environmental meta-omics is rapidly expanding as sequencing capabilities improve, computing technologies become more accessible, and associated costs are reduced. The in situ snapshots of marine microbial life afforded by these data provide a growing knowledge of the functional roles of communities in ecosystem processes. Metaproteomics allows for the characterization of the dynamic proteome of a complex microbial community. It has the potential to reveal impacts of microbial metabolism on biogeochemical transport, storage and cycling (for example, Hawley et al., 2014), while additionally clarifying which taxonomic groups perform these roles. Previous work illuminated many of the important functions and interactions within marine microbial communities (for example, Morris et al., 2010), but a review of ocean metaproteomics literature revealed little standardization in bioinformatics pipelines for detecting peptides and inferring and annotating proteins. As prevalence of these data sets grows, there is a critical need to develop standardized approaches for mass spectrometry (MS) proteomic spectrum identification and annotation to maximize the scientific value of the data obtained. Here, we demonstrate that bioinformatics decisions made throughout the peptide identification process are as important for data interpretation as choices of sampling protocol and bacterial community manipulation experimental design. Our analysis offers a best practices guide for environmental metaproteomics.
Project description:In this study we characterize microbial community features on the surface of Indian Ocean. 11 samples were collected from Indian Ocean and subjected for quantitative metaproteomics analysis for taxonomic and functional analysis. Our results suggested that metabolic tuning at metaproteomics levels enabled microbial community to sustain stable when subjected to environmental perturbations in the oligotrophic ocean.
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:This study investigates transcriptomic responses of Atlantic salmon lice, Lepeophtheirus salmonis exposed to cypermethrin, a commonly used antiparasitic agent used in aquaculture. Copepodid L. salmonis were exposed to cypermethrin (Betamax®) at a concentration of 1.0ppb An in vitro bioassay experiment was conducted using cypermethrin exposures on copepodid (larvae) sea lice (F1 generation) collected from BMA-2a New Brunswick, Canada in 2014. The bioassay exposed copepodids to 1.0 μg/L cypermethrin or a sea water control for 24 hours in glass beakers (VWR) at 12 ± 1oC. Each condition had a total volume of 500mL with six replicates and approximately 500 lice per beaker. After 24 hours, pools of ~500 copepodids were flash frozen for RNA extractions. Post 24-hours, lice were assessed for survival similarly to the technique used for staging and enumeratio first
Project description:Hornyhead turbot (Pleuronichthys verticalis) captured near sewage outfalls are used as sentinel fish for monitoring exposure to industrial and agricultural chemicals of ~20 million people living in coastal southern California. Although analyses of hormones in blood and organ morphology and histology in fish are useful for assessing exposure, there is a need for quantitative and sensitive molecular measurements, as many contaminants produce subtle effects. A novel multispecies microarray and qRT-PCR were used to investigate endocrine disruption in turbot captured near sewage outfalls in San Diego, Orange County and Los Angeles California. Analysis of expression of genes involved in hormone [e.g. estrogen, androgen, thyroid] responses and xenobiotic metabolism in turbot livers was correlated with phenotypic end points.