Attachment of Alcanivorax borkumensis to Hexadecane-In-Artificial Sea Water Emulsion Droplets

Alcanivorax borkumensis (AB) is a marine bacterium that dominates bacterial communities around many oil spills because it enzymatically degrades the oil while using it as a nutrient source. Several dispersants have been used to produce oil-in-water emulsions following a spill. Compared to surface slicks, the additional oil water surface area produced by emulsification provides greater access to the oil and accelerates its degradation. We deliberately cultured AB cells using hexadecane as the only nutrient source. We then examined the first critical step of the biodegradation process, the attachment of these AB cells to hexadecane-water interfaces, using fluorescence microscopy and cryogenic scanning electron microscopy. The hexadecane-in-artificial sea water (ASW) emulsions were produced by gentle shaking and were stabilized either by AB alone, by Corexit 9500, by Tween 20, or by carbon black particles. When no dispersants were used, AB stabilizes the emulsion, and bacterial cells attach to the hexadecane droplets within the first 3 days. When Corexit 9500 was used as the dispersant, AB did not attach to the hexadecane droplets over 3 days, and many AB cells in the aqueous phase appeared dead. Only limited attachment was observed after 7 days. No AB attachment was observed over 3 days when Tween 20 was used as the dispersant. However, the bacteria used Tween 20 in the ASW as a nutrient. Large amounts of AB attached to carbon black stabilized hexadecane droplets within 3 days. An analysis that accounts for van der Waals and electrostatic interactions is unable to predict all of these observations, indicating that the attachment of AB to the hexadecane is a complex phenomenon that goes beyond simple physiochemical effects. While these experiments do not mimic conditions in the open ocean where the large amount of water dilutes any emulsion stabilizer, they provide important insights on bacteria adhesion to oil, a critical step in the oil degradation process following a marine spill.