In this study, 97 microalgal strains purchased from algae bank and 50 microalgal strains isolated from local waters in Minnesota were screened for their adaptability growing on a 20-fold diluted digested swine manure wastewater (DSMW). A pool of candidate strains well adapted to the DSMW was established through a high-throughput screening process. Two top-performing facultative heterotrophic strains with high growth rate (0.536 day(-1) for UMN 271 and 0.433 day(-1) for UMN 231) and one strain with high omega-3 unsaturated fatty acid (EPA, 3.75 % of total fatty acids for UMN 231) were selected. Subsequently, a sequential two-stage mixo-photoautotrophic culture strategy was developed for biofuel and animal feed production as well as simultaneous swine wastewater treatment using above two strains. The maximal biomass concentration and lipid content at the first and second stages reached 2.03 g/L and 23.0 %, and 0.83 g/L and 19.0 % for UMN 271 and UMN 231, respectively. The maximal nutrient removals for total phosphorus and ammonia after second-stage cultivation were 100 and 89.46 %, respectively. The experiments showed that this sequential two-stage cultivation process has great potential for economically viable and environmentally friendly production of both renewable biofuel and high-value animal feed and at the same time for animal wastewater treatment.