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From offshore oil and gas platforms in North Atlantic Canada, crude oil, formation water, drilling mud, treated produced water and seawater samples were collected for screening potential biosurfactant producers. In total, 59 biosurfactant producers belong to 4 genera, namely, Bacillus, Rhodococcus, Halomonas, and Pseudomonas were identified and characterized. Phytogenetic trees based on 16S ribosomal deoxyribonucleic acid (16S rDNA) were constructed with isolated strains plus their closely related strains and isolated strains with biosurfactant producers in the literature, respectively. The distributions of the isolates were site and medium specific. The richness, diversity, and evenness of biosurfactant producer communities in oil and gas platform samples have been analyzed. Diverse isolates were found with featured properties such as effective reduction of surface tension, producing biosurfactants at high rate and stabilization of water-in-oil or oil-in-water emulsion. The producers and their corresponding biosurfactants had promising potential in applications such as offshore oil spill control, enhancing oil recovery and soil washing treatment of petroleum hydrocarbon-contaminated sites.