Crude oil biodegradation by bacterial strains isolated from oil contaminated soil samples, Oman, were performed and its potential applications in crude oil waste management were analyzed. Accidental and occasional crude oil spills, treatment of produced water containing hydrocarbons and oil, and waste management are a major concern for petroleum industries. Various techniques such as, chemical, physical, biological and thermal treatments, are reported for treating spills and wastes on-site. We analyzed crude oil biodegradation by selected bacterial isolates from Oman, under reservoir conditions. Four potential bacterial isolates were selected, characterized by MALDI-Biotyper, and studied for crude oil biodegradation at 40 °C. The isolates were studied morphologically and by scanning electron microscope (SEM), and any changes in surface tension (biosurfactant production), during growth on crude oil as the only carbon source. Crude oil characteristics before and after biodegradation were analyzed by Gas chromatography-Mass specrtrometry (GC-MS). The bacterial strains were identified as Pseudomonas mendocina, Pseudomonas putida, and Brevibacillus agri. During the course of crude oil biodegradation, bacterial isolates showed growth, as analyzed by optical density measurement at 660 nm and cellular protein estimation; no changes were observed in surface tension values, and alteration in the cell morphology in presence of crude oil was observed. All four isolates showed oil clearing zones on agar plates coated with crude oil. Crude oil degradation was analyzed by GC-MS with respect to carbon numbers from C12 −C30. P. mendocina II, P. putida and B. agri showed reduction in all the compounds, but P. mendocina I showed very little degradation of hydrocarbons. Maximum crude oil biodegradation (~50%) was observed by P. mendocina II. It can be concluded that the present findings indicate the application potential of these bacterial isolates in the crude oil biodegradation. This could be the ideal solution to treat the contaminated soil and water, which can also be applied for the bioremediation of oil spills and water bodies as a cost effective and environmental friendly approach.