Membrane biofouling is the primary cause of inefficiency in seawater reverse osmosis desalination. The identification and subsequent removal of causative microorganisms would therefore be beneficial. To achieve this aim, the assembly of microorganisms onto the reverse osmosis membranes was first modeled to reveal a niche-selective process. Specifically, bacterial genera Hyphomonas, Muricauda, Bacillus and Pseudoruegeria were detected in occurrence frequency higher than predicted, and likely play a role in biofouling due to production of exopolymers. Subsequently, four different pretreatment systems, namely ultrafiltration (UF) membranes, intake wells, dual media filtration and cartridge filters (CF), were evaluated for their log removal efficiencies of these four genera. UF outperformed the others in removing the potential biofouling-associated genera, but intake wells achieved a higher log removal of cell densities. Microbial regrowth, as denoted by an increase in cell numbers, was consistently observed within the CF. Using well intakes provides the highest degree of pretreatment in removing total cells in a chemical-free manner, while UF is the next best process to remove bacteria and organic carbon compounds most responsible for membrane biofouling.