Visualizing the Hidden World: Utilizing CARD-FISH to Detect Microbes Exposed to Oil

Makeda Mills1,2, Amanda Achberger3 and Jason B Sylvan1, (1)Texas A&M University, Oceanography, College Station, TX, United States, (2)University of the Virgin Islands, St. Thomas, United States Virgin Islands, (3)Texas A&M University, Oceanography, College Station, United States
The Deepwater Horizon Oil Spill of 2010 released more than four million barrels of oil into the Gulf of Mexico, causing significant damage to marine life in the water column and surrounding shorelines. Several methods, including the application of chemical dispersants, were used to remediate the oil spill. Dispersants break oil into smaller droplets, enhancing degradation by abiotic processes as well as by marine microbes. This study takes a microscopic look at how two hydrocarbon-degrading genera of microbes, Alteromonas and Alcanivorax, aggregate in the presence of oil and oil plus dispersant. We used Catalyzed Reporter Deposition-Fluorescent in Situ Hybridization (CARD-FISH) to analyze microbial aggregation in three mesocosm treatments with natural surface seawater from coastal Texas: surface seawater (Control), water accommodated fraction of oil (WAF), and diluted chemically enhanced water accommodated fraction of oil (DCEWAF). We hypothesized that there will be higher amounts of micro-aggregates (<0.20 mm) compared to single cells in oil treatments vs untreated seawater, and that both genera would be more abundant in micro-aggregates than the free-living population. The utilized probe specific to Alcanivorax was designed and optimized for high intensity fluorescence, which we found was greatest at 55% formamide. Preliminary analysis of total community and CARD-FISH counts indicate that there is a higher number of micro-aggregates in WAF and DCEWAF compared to the control treatments, and that Alteromonas and Alcanivorax were proportionally more abundant in aggregates in oiled treatments. This study may lead to implications for further research in understanding the role micro-aggregates play in hydrocarbon degradation and the role known hydrocarbon degraders play in those micro-aggregates.