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The reported persistence of the dioctyl sodium sulfosuccinate (DOSS) surfactant in Corexit 9500 in the oil plumes formed during the Deepwater Horizon oil spill has contributed to concerns regarding the biodegradability and bioavailability of dispersed oil and dispersants used as an oil spill countermeasure in the Gulf of Mexico. We studied the biodegradation of DOSS and dispersed South Louisiana crude oil (SLC) in laboratory microcosms. Two oil-degrading cultures from the Gulf of Mexico were isolated, one from the surface (meso) and one from close to the area of the Macondo well (cryo). Each was enriched on SLC, the former at 25 °C, the latter at 5 °C. Results indicated that the meso culture rapidly and completely degraded DOSS, alkanes, and aromatics. The cryo culture metabolized the same compounds but with a lag of 28 d and a remaining residual of iso-alkanes, n-C_30–35, and the 4-ring PAHs.