There has been chronic contamination of the surrounding aquatic and terrestrial environments in the Niger Delta region of Nigeria due to the enormous activities of crude oil exploration. This study aimed to characterize and assess the crude oil tolerance potential of indigenous microbes from crude oil spills in Santa Barbara River, Bayelsa State, Nigeria, that could serve as potential consortia for bioremediation of the crude oil spills. Total crude oil hydrocarbon-utilizing bacteria count (THUBC) and total crude oil hydrocarbon-utilizing fungi count (THUFC) in the samples were determined with a culture-dependent spread plate technique. Bacterial and fungal isolates were characterised using standard phenotypic and 16S/Internal Transcribed Spacer region rRNA gene sequencing techniques, respectively. The tolerance of autochthonous bacterial isolates to different concentrations of crude oil was subsequently determined. THUBC and THUFC in crude oil-polluted water samples ranged from 1.88 log10 CFU/ml to 2.74 log10 CFU/ml and from 0.00 log10 CFU/ml to 1.70 log10 CFU/ml, respectively. Representative strains of Proteus mirabilis, Bacillus subtilis, Pseudomonas aeruginosa, Micrococcus luteus and Aspergillus flavus isolates obtained from crude oil-polluted water samples were deposited in the GenBank (NCBI) under accession numbers OQ969924, OQ969951, OQ969987, OQ970009 and OQ975908. Proteus mirabilis, Pseudomonas aeruginosa and Aspergillus flavus demonstrated the most significant tolerance to crude oil pollutants (minimum crude oil inhibitory concentrations (MIC) = 80%) followed by Bacillus subtilis and Micrococcus luteus (MIC= 40%). The findings from this study are pointers to the potential role of the microbial isolates as bioremediation consortia to remediate the polluted Santa Barbara River.