Abstract:

Mesocosm experiments were carried out to test the effect of oil and Corexit on microbial community composition and structure over time in coastal seawater. Filtered water samples were collected in triplicate every 12 hours in the Control, WAF, CEWAF, dilute CEWAF, and the WAF silicon tube treatment. DNA was extracted from filtered water samples and sequenced using barcoded primers specific for the V4 hypervariable region of the 16S rRNA gene.

Suggested Citation:

Doyle, Shawn; Sylvan, Jason. 2018. GOM coastal waters (GOMCOAST): Illumina MiSeq sequenced 16S library of Microbial Community Response to Oil and Corexit. Distributed by: GRIIDC, Harte Research Institute, Texas A&M University–Corpus Christi. doi:10.7266/N77D2SPZ

Publications:

Kamalanathan, M., Xu, C., Schwehr, K., Bretherton, L., Beaver, M., Doyle, S. M., … Quigg, A. (2018). Extracellular Enzyme Activity Profile in a Chemically Enhanced Water Accommodated Fraction of Surrogate Oil: Toward Understanding Microbial Activities After the Deepwater Horizon Oil Spill. Frontiers in Microbiology, 9. doi:10.3389/fmicb.2018.00798

Bera, G., Doyle, S., Passow, U., Kamalanathan, M., Wade, T. L., Sylvan, J. B., … Knap, A. H. (2019). Biological response to dissolved versus dispersed oil. Marine Pollution Bulletin, 110713. doi:10.1016/j.marpolbul.2019.110713

Doyle, S. M., Lin, G., Morales-McDevitt, M., Wade, T. L., Quigg, A., & Sylvan, J. B. (2020). Niche Partitioning between Coastal and Offshore Shelf Waters Results in Differential Expression of Alkane and Polycyclic Aromatic Hydrocarbon Catabolic Pathways. mSystems, 5(4). doi:10.1128/msystems.00668-20

Purpose:

To characterize the microbial community composition and structure in the GOMCOAST Mesocosms (Meso4 and MesoSi).

Data Parameters and Units:

Illumina MiSeq reads in FastQ format Mesocosm 4 - GOMCOAST Treatments: Control: seawater only WAF: seawater containing Water Accommodated oil Fraction CEWAF: Chemically-enhanced WAF (WAF prepared with Corexit) DCEWAF: Diluted chemically-enhanced WAF (a 1:10 dilution of CEWAF) In addition to the above treatments, two additional treatments were prepared for Mesocosm Si: Si-Control: seawater only containing empty silicon tubing Si-WAF: seawater containing a passively dosed WAF, delivered via silicon tubing Timepoints: T0: 0 hours T1: 12 hours T2: 24 hours T3: 36 hours T4: 48 hours T5: 60 hours T6: 72 hours Triplicate replicates are labeled (A), (B), and (C)

Methods:

WAF generated via silicone tube was prepared following Redman, A.D., Butler, J.D., Letinski, D.J., Parkerton, T.F., 2017. Investigating the role of dissolved and droplet oil in aquatic toxicity using dispersed and passive dosing systems. Environmental Toxicology and Chemistry Vol 36 No. 4 pp 1020-1028. In summary, a medical grade silicone tube (6 inch-1.5 ft) from A-M System Inc. WA with 0.058-inch X 0.077-inch X 0.0095-inch dimension was used. The predetermined amount of oil (e.g. 0.84 ml/2L for oil loading of 360 mg/L) was introduced into the silicone tube by gas tight Hamilton syringe and both ends were closed tightly by knotting. The silicone tube was then put into the 2L aspirator attached to the stir bar. Analogous to the CROSERF method, the loaded tubing was stirred for 72 h at 300 rpm before collection of samples. Twelve 100L mesocosm tanks were filled with Gulf of Mexico seawater collected from the Texas coastline, near TABS buoy R (29° 38.1000'N, 93° 38.5020'W) which is located ~100 miles away from Galveston (TX). Four treatments were prepared in triplicate. Control tanks were filled with seawater. Water accommodated fraction (WAF) of oil was prepared by mixing 25 mL (5 ml ~ every 30 min for 2.5 hrs) of Macondo surrogate oil into 130 L of seawater. Mixing ended 24 hrs. after the initial oil addition (Knap et al. 1983; Wade et al. 2017). The WAF was then introduced into the WAF mesocosm tanks and filled to 87 L and mixed. From these WAF tanks 6 L was removed for other experiments and analyses (2 L dark/light, 4 L hydrocarbon analyses). In order to make chemically enhanced water accommodated fraction (CEWAF), Corexit was mixed with oil in a ratio of 1:20 and 25 mL of this mixture (5 ml every 30 min for 2.5 hrs) of surrogate oil plus Corexit was added to 130 L of seawater. Mixing ended 24 hrs after the initial oil addition. The CEWAF was then introduced into the CEWAF mesocosm tanks and filled to 96 L and mixed. From these CEWAF tanks 13 L was removed for other experiments and analyses (7 L for the DCEWAF mesocosms, 2 L dark/light, 4 L hydrocarbon analyses). Diluted CEWAF (DCEWAF) was prepared by mixing 9 L of CEWAF with 78 L of the original seawater for a total volume of 87 L. From these DCEWAF tanks 6 L was removed for other experiments and analyses (2 L dark/light, 4 L hydrocarbon analyses). To the water in the 12 mesocosms, nutrients were added (final concentration f/20) and the tanks stirred. Banks of lights were placed behind each of the glass mesocosm tanks and a 12:12 light/dark cycle employed. Sampling commenced and defined as time zero. The estimated oil equivalents (EOE) were determined using Macondo surrogate oil as the calibration standard (Wade et al. 2011) for the fluorescence analyses (Horiba Scientific Aqualog Fluorometer). The EOE mean concentration of the three mesocosms for the control, WAF, DCEWAF and CEWAF at the start of the experiments were 0 mg/L, 0.26 mg/L, 2.74 mg/L and 41.5 mg/L, respectively. The EOE mean concentration of the three mesocosms for the in the control, WAF, DCEWAF and CEWAF after 72 hours were 0 mg/L , 0.06 mg/L, 1.03. and 17.3 mg/L, respectively. Raw sequence reads in FASTQ format for each replicate of each treatment (i.e. Control, WAF, CE-WAF, Diluted CE-WAF, Si-Control, and Si-WAF). The hyper-variable V4 region of the 16S rRNA gene was PCR amplified from the DNA extracts with GoTaq Flexi DNA Polymerase (Promega) using a methodology similar to that described by Caporaso et al. 2011. Each sample was amplified in triplicate 25μL reactions with the following cycling parameters: 95°C for 3 min, 30 cycles of 95°C for 45 s, 50°C for 60 s, and 72°C for 90 s, and a final elongation step at 72°C for 0 min. All reactions were performed using the 515F-806R primer pair (10µM each) which was modified to include Golay barcodes and adapters for Illumina MiSeq sequencing (Caporaso et al. 2012). The primer pair was also modified to include recently published revisions that reduce bias against the Crenarchaeota/Thaumarchaeota archaeal lineages and the SAR11 bacterial clade (Parada et al. 2016; Apprill et al. 2015). Following amplification, triplicate reactions were combined together and run on a 1.5% agarose gel to assess amplification and relative band intensity. Amplicons were then quantified with the QuantiFluor dsDNA System (Promega), pooled at equimolar concentrations, and purified with an UltraClean PCR Clean-Up Kit (MoBio Laboratories). The purified library, along with aliquots of the three sequencing primers, were sent to the Georgia Genomics Facility (Athens, GA, USA) for MiSeq sequencing (v2 chemistry, 2 x 250bp). Illumina MiSeq Sequencing: V2-Chemistry; PE-250.

Provenance and Historical References:

Apprill, A., McNall, S., Parsons, R., and L. Weber (2015). Minor revision to V4 region SSU rRNA 806R gene primer greatly increases detection of SAR11 bacterioplankton. Aquat. Microb. Ecol. 75: 129-137, doi: 10.3354/ame01753 Caporaso, J.G., Lauber, C.L., Walters, W.A., Berg-Lyons, D., Lozupone, C.A., Turnbaugh, P.J., Fierer, N., and R. Knight (2011). Global patterns of 16S rRNA diversity at a depth of millions of sequences per sample. PNAS, 108(Supplement 1): 4516-4522, doi: 10.1073/pnas.1000080107 Caporaso, J.G., Lauber, C.L., Walters, W.A., Berg-Lyons, D., Huntley, J., Fierer, N., Owens, S.M., Betley, J., Fraser, L., Bauer, M., Gormley, N., Gilbert, J.A., Smith, G. and R. Knight (2012). Ultra-high throughput microbial community analysis on the Illumina HiSeq and MiSeq platforms. ISME J. 6: 1621-1624, doi: 10.1038/ismej.2012.8 Knap, A.H., Sleeter, T.D., Dodge, R.E., Wyers, S.C., Frith, H.R., and S.R. Smith. (1983). The effects of oil spills and dispersant use on corals: A review and multidisciplinary experimental approach. Oil and Petrochemical Pollution, 1(3): 157-169. doi: 10.1016/S0143-7127(83)90134-5 Parada, A.E., Needham, D.M., and J.A. Fuhrman (2016). Every base matters: assessing small subunit rRNA primers for marine microbiomes with mock communities, time series and global field samples. Env. Microbiol. 18(5): 1403-1414, doi: 10.1111/1462-2920.13023 Wade, T. L., S. T. Sweet, J. N. Walpert, J. L. Sericano, J. J. Singer, and N. L. Guinasso Jr. (2011). Evaluation of possible inputs of oil from the Deepwater Horizon spill to the Loop Current and associated eddies in the Gulf of Mexico, in Monitoring and Modeling the Deepwater Horizon Oil Spill: A Record-Breaking Enterprise. Geophys. Monogr. Ser., doi:10.1029/2011GM 001095. Wade, T.L., Morales-McDevitt, M., Bera, G., Shi, D., Sweet, S., Want, B., Gold-Bouchot, G., Quigg, A., and A.H. Knap (2017). A method for the production of large volumes of WAF and CEWAF for dosing mesocosms to understand marine oil snow formation. Heliyon, 3(10): e00419, doi: 10.1016/j.heliyon.2017.e00419

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