Abstract:
Cytochrome p450 1A (CYP1A) gene expression is a commonly used indicator of polycyclic aromatic hydrocarbons (PAHs -an organic compound in oil) exposure. We compared CYP1A gene expression levels in seaside sparrows (SESP) with the amount of oil present in the environment, as measured in sediment samples collected by Coastal Waters Consortium researchers (GRIIDC dataset R4.x264.000:0056). After initial data analysis indicated both CYP1A and hydrocarbons increased after a hurricane (Hurricane Isaac), we selected a subset of samples from R4.x264.000:0056 for oil-source fingerprinting in order to determine if the hydrocarbons in samples collected after the hurricane could be fingerprinted back to the Deepwater Horizon oil spill (fingerprinted samples can be referenced to R4.x264.000:0056 by using the ‘LSU Sample ID#’). The current dataset contains information regarding the field capture of seaside sparrows (including date, location of capture, age, and sex), dates of each major laboratory procedure and the RQ values for three genes: CYP1A5 (the gene of interest), PPIA and RPL4 (reference ‘housekeeping’ genes). It also contains data from the selected fingerprinted sediment samples mentioned above. Additional dataset authors: Christine Bergeon Burns, Stefan Woltmann, Philip Stouffer, Edward Overton, and Buffy Meyer.
Suggested Citation:
Perez-Umphrey, Anna, Taylor, Sabrina, Bergeon Burns, Chrisitne, Woltmann, Stefan, Stouffer, Phillip, Overton, Edward, Meyer, Buffy. 2018. CYP1A gene expression in seaside sparrows collected from oiled and unoiled plots in Plaquemines Parish, LA, 2011-2014. Distributed by: GRIIDC, Harte Research Institute, Texas A&M University–Corpus Christi. doi:10.7266/N7CZ35JV
Data Parameters and Units:
File: R4.x264.219-0002_SESP_FieldCollection-CYP1A_sub1.xlsx Date of field collection: DD-MMM-YY, Field Site Name: Descriptive site name, Center of plot latitude: The latitude of the center of the SESP sampling site in decimal degrees, Center of plot longitude: The longitude of the center of the SESP sampling site in decimal degrees, Treatment type: Site condition based on SCAT map criteria (in parentheses): oiled (heavily oiled), unoiled (very light or no oil observed), reference (East of the Mississippi), Corresponding site abbreviation: Closest regular SESP sampling site (O=oiled; U=unoiled; D=reference site, plus numeric identifier), Bird sex: M=male; F=female; U=unknown, New or recapture: N=new; R=recapture, Age: AHY=After Hatch Year; HY=Hatch Year, Band #: USGS band numerical ID; NA if no band, Sample ID: Collection identifier, Collector initials: Initials of collector, Region of liver subsampled: Section of liver used for RNA extraction (whole, right lobe, or top lobe), Date of RNA extraction: Date that RNA was extracted from liver sample (DD-MMM-YY), Date of cDNA synthesis: Date that extracted RNA was reverse transcribed into cDNA (DD-MMM-YY), cDNA sample #: cDNA sample numerical identifier, RQ CYP1A5: Relative Quantity value for CYP1A5, RQ PPIA3: Relative Quantity value for PPIA3, RQ RPL4: Relative Quantity value for RPL4 File: R4.x264.219-0002_SESP_FingerprintingData_sub1.xlsx Sample Description: User defined descriptor for sediment sample, assigned at time of field collection (also contained in R4.x264.000:0056), Date of field collection: DD-MMM-YY that sediment sample was collected from field site, Sample location latitude: Latitude where sediment sample was collected, in decimal degrees, Sample location longitude: Longitude where sediment sample was collected, in decimal degrees, Treatment type: Site condition based on SCAT map criteria (in parenthesis): oiled (heavily oiled), Corresponding site abbreviation: Closest SESP regular sampling site (O=oiled, plus numeric identifier) LSU sample ID #: Internal sample identifier assigned by the analytical lab (also contained in R4.x264.000:0056), Oil-source fingerprinting technique: Qualitative comparison analysis: Visual comparison of biomarker profiles of extracted sediment samples to MC252 Deepwater Horizon source oil. Qualitative match (Y=yes; N=no that a given sample matches MC252 Deepwater Horizon oil; N/A=oil detected but too weathered to fingerprint, inconclusive). Qualitative pattern: (Samples with an A, B, or AB pattern have MC252 oil in them; A=unweathered MC252 oil; B= weathered MC252 oil; AB=mixture of A and B; N/A=too weathered to fingerprint, inconclusive). Oil-source fingerprinting technique: Diagnostic biomarker ratio analysis: Triterpanes (m/z 191), steranes (m/z 217 and 218), and triaromatic steroids (m/z 231) GC/MS data are used for calculating diagnostic ratios based on peak heights for each sample and compared to the average MC252 source oil ratio using the critical difference statistical method. Percentages are then assigned to each sample based on degree of “matching”. Diagnostic Ratio Score: (87-100%=Match between the sample and MC252 oil; 79-86%=Probable match between the sample and MC252 oil; <79%= Inconclusive match between the sample and MC252 oil). Diagnostic Ratio Category: Descriptive result of the Diagnostic Ratio Score Oil-source fingerprinting technique: Chemometric analysis: Hierarchial cluster analysis of peak intensity data points for triterpanes, diasteranes, steranes, and triaromatic steroids in sediment samples and MC252 source oil, which uses a similarity matrix to assess relationships. Chemometrics, ALL: (Y=yes; N=no; N/A=inconclusive that there is a match between the sample and all four biomarker ion groups m/z 191, 217, 218, 231). Chemometrics, m/z 217 Only: (Y=yes; N=no; NA=inconclusive that there is a match between the sample and ion m/z 217). # of positive OSF techniques: Number of OSF analyses that matched a sample to MC252 oil # OSF techniques: Number of OSF techniques applied to the sample File: R4.x264.219-0002_SESP_DeltaCtValues_sub2.xlsx cDNA sample #: cDNA sample numerical identifier, Delta Ct CYP1A5: delta Ct values used to calculate Relative Quantity (RQ) for CYP1A5 genes (located in R4.x264.219-0002_SESP_FieldCollection-CYP1A_sub1.xlsx), Delta Ct PPIA3: delta Ct values used to calculate Relative Quantity (RQ) for PPIA3 genes (located in R4.x264.219-0002_SESP_FieldCollection-CYP1A_sub1.xlsx), Delta Ct RPL4: delta Ct values used to calculate Relative Quantity (RQ) for RPL4 genes (located in R4.x264.219-0002_SESP_FieldCollection-CYP1A_sub1.xlsx) File: R4.x264.219-0002_SESP_PeakHtDiagnosticRatios_sub2.xlsx Fingerprinting peak height data of triterpanes (m/z 191), steranes (m/z 217 and 218), and triaromatic steroids (m/z 231) for each sample and average MC252 oil source ratio data point listed in R4.x264.219-0002_SESP_FingerprintingData_sub1.xlsx. The MC252 source oil columns contain peak heights from one of the MC252 source oil samples analyzed in the sample batches and from each year the sample batches were analyzed (i.e., LSU ID#s: 2012270, 2013156, 2014255). File: R4.x264.219-0002_SESP_PeakIntensityData_sub2.xlsx Fingerprinting peak intensity data for hopanes, steranes, and triaromatic steroids in sediment samples and MC252 source oil contained in R4.x264.219-0002_SESP_FingerprintingData_sub1.xlsx. Worksheet Tabs “Peak Intensities_GT” and “Peak Intensities_MU” refer to the specific instrument samples were processed on. The analytical lab operates 2 GC/MS instruments, both with the same acquisition parameters and tuned to produce similar analytical results. Both worksheet tabs contain the same variables: Data File: Default Instrument export filename, LSU sample ID #: Internal identifier assigned to all samples by the analytical lab (contained in all oil data files as identifier). Note that MC252 source oil was analyzed with each sample batch. Time (minutes): Elapsed time of sample retention within the instrument. File: R4.x264.219-0002_SESP_KeyForChemicalConcentrationData_sub2.xlsx Filename key for locating concentration data referenced in this dataset. LSU sample ID #: Internal sample identifier assigned by the analytical lab (contained in all oil data files as identifier) Filename within R4.x264.000:0056 where sample data is located Worksheet in R4.x264.000:0056: Worksheet where sample data is located within the individual files listed Column: Column within the worksheet and individual files in R4.x264.000:0056 where sample data is located
Methods:
Seaside sparrows were collected from oiled and unoiled plots in Barataria Bay and Pointe-a-la-Hache, LA from 2011 to 2014. Birds were collected using a 0.22 shotgun (2011) and mist nests (2012-2014). Birds were euthanized by thoracic compression and livers and other tissues were collected and immediately frozen in liquid nitrogen. At Louisiana State University, total RNA was extracted from the livers and then reverse transcribed into cDNA. Primers were designed to amplify three genes in quantitative PCR reactions: CYP1A5 (the gene of interest), PPIA and RPL4 (reference ‘housekeeping’ genes). CYP1A5 gene expression levels were normalized against the abundance of reference genes to acquire the Relative Quantity (RQ) value of CYP1A5 in each liver sample. Sediment samples were selected for oil-source fingerprinting according to 1) their date of collection (sampled between the time of the hurricane and the end of 2012, i.e. August-December 2012), 2) proximity to the SESP collection sites (locations of the sediment sample collections had to be within 50m of the SESP collection sites, and 3) samples that were collected from 'oiled' sites (as determined by initial SCAT data).
Instruments:
Shotguns with 0.22 shotshells: Used to collect seaside sparrows 10m-long, 30mm mesh mist nest: Used to collect seaside sparrows Tissue Tearor (Biospec, Bartlesville, OK, USA): Used to homogenize liver tissues Qiagen RNeasy Plus Mini Kit: Used to extract total RNA from liver samples NanoDrop 2000 spectrophotometer (Thermo Fisher Scientific, Waltham, MA, USA): Used to measure RNA concentration in each sample Agilent 2011 Bioanalyzer (Santa Clara, CA, USA): Used to confirm RNA integrity Applied Biosystems 7900 Sequence Detection System instruments (Foster City, CA, USA): Used to run quantitative PCR reactions
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