Abstract:

A mesocosm scale experiment was conducted to monitor the temporal changes in microbial cell abundance as well as the concentration of microaggregates that form in response to the presence of oil and chemically dispersed oil. Water samples were collected and viewed under the microscope to quantify cell and microaggregate concentrations over time.

Suggested Citation:

Achberger, Amanda and Jason Sylvan. 2021. Mesocosm: Characterization of cell concentrations and microaggregate abundance, MICROX, to study the microbial oxidation and degradation of oil. Distributed by: GRIIDC, Harte Research Institute, Texas A&M University–Corpus Christi. doi:10.7266/n7-p1yv-k581

Purpose:

To monitor microbial cell and microaggregate population dynamics over time after exposure to oil.

Data Parameters and Units:

The headers are Treatment (Control, WAF, DCEWAF), Timepoint (days), Cells per mL, and Aggregates per mL

Methods:

The water accommodated fraction (WAF) of oil was prepared by mixing 25 mL (5 ml ~ 5 min over 25 min) of Macondo surrogate oil into six baffled recirculating tanks (Wade et al., 2017) containing ~130 L of seawater were mixed vigorously for 4 hours to create a “high energy” WAF (Knap et al. 1983). The WAF was then introduced into the WAF mesocosm tanks, filled to 104L, and mixed. From these tanks, 3.5 L WAF was removed for hydrocarbon analyses. Corexit was mixed with oil in a ratio of 1:20 to make diluted chemically enhanced water accommodated fraction (DCEWAF). Then 25 mL of this mixture (5 ml every 5 min for 25 min) of surrogate oil plus Corexit was added to a baffled recirculating tank containing ~130 L of seawater which was not vigorously mixed for 4 hours prior to being loaded into the mesocosm tanks. From these CEWAF tanks 5L was removed for other analyses (3.5 L hydrocarbon analyses). Diluted CEWAF (DCEWAF) was prepared by mixing 10.4 L of CEWAF with 93.6 L of the original seawater for a total volume of 104 L. This concentrated plankton mass was introduced to each of the tanks (1.5 L to each tank which contained 104L of treated seawater, total volume of seawater 105.5L per tank) immediately prior to starting the experiments. Banks of lights were placed behind each of the glass mesocosm tanks and a 12:12 light/dark cycle employed. Direct cell counts were performed with an epifluorescence microscope (Zeiss Axio Imager.M2) after staining the 2% formalin fixed samples with DAPI (45uM final concentration) for 5 minutes in the dark and filtering them onto 25mm, 0.2um black polycarbonate filters. Each filter was mounted on a glass microscope slide with coverslip using two drops of AF1 antifade. A 100um×100um ocular counting grid was used at 1000× magnification for cell counts. Due to their much larger size, aggregates were instead counted for each field of view at 400× magnification.

Instruments:

Zeiss Axio Imager.M2

Provenance and Historical References:

Knap, Anthony H., Thomas D. Sleeter, Richard E. Dodge, Sheila C. Wyers, Harold R. Frith, and S. R. Smith. 1983. The effects of oil spills and dispersants 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 Wade, Terry L., Maya Morales-McDevitt, Gopal Bera, Dawai Shi, Stephen Sweet, Binbin Wang, Gerado Gold-Bouchot, Antonietta Quigg, and Anthony 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 3e00419 DOI: 10.1016/j.heliyon.2017.e00419

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