Dataset Information

HoloFish - Untargeted Metabolomics

ABSTRACT: Sample Collection Samples used for this study were obtained as part of the HoloFish project (Norwegian Seafood Research Fund, project no. 901436). This cohort has been described previously 19. Briefly, we sampled 460 ready-to-harvest Atlantic salmon from a commercial production site close to Bergen, Norway, owned by Leroy Seafood Group in April 2018. Samples were obtained from two groups reared in separate sea pens and fed two different standard commercial diets. These diets have been anonymised but were manufactured respectively by BioMar and EWOS in 2018. Six biological samples were taken from each fish, including muscle tissue for fatty acid profiling, gill tissue for host genomics, gut epithelia for host transcriptomics, gut epithelial cell scrapes for 16S metabarcoding and two gut content samples for metagenomics and metabolomics. Approximate 100 mg distal gut content for each individual was sampled for metabolomics. Gut content for metabolomics was preserved at -80 degrees Celsius. All the sampling tools and equipment used for each sample were sterile. Extraction Gut content samples were cryo-homogenised in 25% water, 25% methanol and 50% dichloromethane in a 1:15 sample: solvent ratio (w:v). Homogenisation was carried out using an OMNI Bead Ruptor 24, using liquid nitrogen to keep homogenised samples below 0 degrees Celsius to minimise degradation of metabolites during extraction. Homogenates were centrifuged at 20,000 g (0 degrees Celsius) and the polar phase from all samples was concentrated using SpeedVac (ThermoFisher Scientific) and resuspended in 200 microL 5% methanol. Four procedural blanks were included in homogenisation. A volume of 100 microL of all samples was collected into a Quality Control sample used for normalisation to enhance the detection of metabolites. Chromatography Samples were measured on a nano-flow ultra-high pressure liquid chromatography tandem high-resolution mass spectrometry analysis. Mass spectrometry Metabolites were detected using a Q Exactive HF Hybrid Quadrupole-Orbitrap Mass Spectrometer (ThermoFisher Scientific) operated in positive ion data-dependent acquisition mode. Data Transformation ThermoFisher Scientific UHPLC-Orbitrap-MS/MS RAW files were converted into mzML files using Proteo Wizard. For an increased deciphering of molecular spectres, MZmine2 was applied for mass detection of MS1 and MS2 spectres, followed by chromatogram detection and deconvolution. Subsequently, detected isotopes and features were grouped according to a tolerance of mass-charges (5 ppm for m/z) and retention time (6 sec.) and the features were further aligned according to retention time and m/z. Lastly, only features with an MS2 spectrum were kept for further substructural analysis and in silico analysis.

INSTRUMENT(S): Q Exactive HF

ORGANISM(S): Salmo Salar (ncbitaxon:8030)

SUBMITTER: Jacob Rasmussen

PROVIDER: MSV000093872 | MassIVE | Tue Jan 16 03:50:00 GMT 2024

REPOSITORIES: MassIVE

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Project description:Sample Collection Samples used for this study were obtained as part of the HoloFish project (Norwegian Seafood Research Fund, project no. 901436). This cohort has been described previously 19. Briefly, we sampled 460 ready-to-harvest Atlantic salmon from a commercial production site close to Bergen, Norway, owned by Leroy Seafood Group in April 2018. Samples were obtained from two groups reared in separate sea pens and fed two different standard commercial diets. These diets have been anonymised but were manufactured respectively by BioMar and EWOS in 2018. Six biological samples were taken from each fish, including muscle tissue for fatty acid profiling, gill tissue for host genomics, gut epithelia for host transcriptomics, gut epithelial cell scrapes for 16S metabarcoding and two gut content samples for metagenomics and metabolomics. Approximate 100 mg distal gut content for each individual was sampled for metabolomics. Gut content for metabolomics was preserved at -80 degrees Celsius. All the sampling tools and equipment used for each sample were sterile. Extraction Gut content samples were cryo-homogenised in 25% water, 25% methanol and 50% dichloromethane in a 1:15 sample: solvent ratio (w:v). Homogenisation was carried out using an OMNI Bead Ruptor 24, using liquid nitrogen to keep homogenised samples below 0 degrees Celsius to minimise degradation of metabolites during extraction. Homogenates were centrifuged at 20,000 g (0 degrees Celsius) and the polar phase from all samples was concentrated using SpeedVac (ThermoFisher Scientific) and resuspended in 200 microL 5% methanol. Four procedural blanks were included in homogenisation. A volume of 100 microL of all samples was collected into a Quality Control sample used for normalisation to enhance the detection of metabolites. Chromatography Samples were measured on a nano-flow ultra-high pressure liquid chromatography tandem high-resolution mass spectrometry analysis. Mass spectrometry Metabolites were detected using a Q Exactive HF Hybrid Quadrupole-Orbitrap Mass Spectrometer (ThermoFisher Scientific) operated in positive ion data-dependent acquisition mode. Data Transformation ThermoFisher Scientific UHPLC-Orbitrap-MS/MS RAW files were converted into mzML files using Proteo Wizard. For an increased deciphering of molecular spectres, MZmine2 was applied for mass detection of MS1 and MS2 spectres, followed by chromatogram detection and deconvolution. Subsequently, detected isotopes and features were grouped according to a tolerance of mass-charges (5 ppm for m/z) and retention time (6 sec.) and the features were further aligned according to retention time and m/z. Lastly, only features with an MS2 spectrum were kept for further substructural analysis and in silico analysis.

Project description:This study measured the differential expression of genes from a single clonal Daphnia pulex isolate obtained from an eutrophic flooded opencast mine near Gräfenhain (Saxony, Germany) and has been kept in the laboratory since 2002. The daphniids (50 adult animals per batch) were raised in 3-L glass beakers under a 16 h:8 h L:D photoperiod. Three-quarter of the culture medium (M4) was renewed once a week, and the animals were equally fed ad libitum with algae (Desmodesmus subspicatus; SAG 53.80, Göttingen, Germany). Long-term-acclimated animals were kept at three different temperatures (i.e., 10 ± 0.5°C, 20 ± 0.3°C, and 24 ± 0.3°C) under normoxia (Po2 ≥ 20 kPa) for at least twelve weeks. Time-resolved experiments (acute heat stress) were carried out with long-term 20°C acclimated animals, which were exposed for three different time intervals (2, 4, and 8 h) to either 30 ± 0.2°C (test conditions) or 20 ± 0.3°C (control conditions). For this, 50 animals each were rapidly transferred with a sieve (mesh size: 1.2 mm) from 20°C-medium to a set of beakers containing 30°C-medium. As control, 50 animals each were transferred from 20°C-medium to another set of beakers with 20°C-medium. Only adult female animals with a body length of 2−2.5 mm (between the base of the apical spine and the anterior part of the head) and carrying parthenogenetic eggs and embryos were used for experiments. Animals were not fed during short-term exposures. During experiments, all animals were in good physical condition. After thermal acclimation or heat exposure, animals were transferred into 1.5-ml microcentrifuge tubes and shock-frozen in liquid nitrogen after removing adhering water with a tissue paper. Samples were short-term stored at -80°C. For all experimental conditions, four independent replicates (50 animals each) were analyzed. This GEO record is for the contrasts between animals conditioned to 10 degrees Celsius (condition 1) versus 20 degrees Celsius (condition 2), 10 degrees Celsius (condition 1) versus 24 degrees Celsius (condition 3), and 20 degrees Celsius (condition 2) versus 24 degrees Celsius (condition 3)

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HoloFish - Untargeted Metabolomics

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