Wastewater treatment with microalgae is an ecologically sustainable process. In this study, the growth characteristics, nutrient removal, and spectral changes of dissolved organic matter (DOM) in microalgae bioreactors were investigated for treating low C/N ratio wastewater under different disturbance modes (agitation and aeration) and carbon sources (sucrose and humic acid). The results showed that the biomass and chlorophyll-a contents of Scenedesmus obliquus in the aeration condition (725.32–811.16 × 104 cells mL−1, 1.58–1.69 mg L−1) were higher than those in the agitation condition (426.06–465.14 × 104 cells mL−1, 1.48–1.61 mg L−1). The better removal of nutrients (TN, 29.62–36.39 mg L−1, TP, 1.84–2.30 mg L−1) by microalgae in sucrose-containing wastewater under agitation conditions occurred on the second day, with removal efficiencies of 21.33–30.67% and 44.84–58.51%, respectively; while it was on the fifth day both in sucrose and humic acid-containing wastewater under aeration conditions (TN, 19.56–31.20 mg L−1, TP, 0.26–0.30 mg L−1), with removal efficiencies of 13.92–46.75% and 88.36–90.50%, respectively. The wastewater DOM primarily consisted of humic-like substances under agitation and aeration conditions characterized by high levels of aromaticity, molecular weight and humification. Furthermore, the aromatization and humification properties of DOM in humic acid wastewater were higher than those in sucrose wastewater, which was corresponding with the lower removal and availability of pollutants by algae. Microalgae showed good biomass accumulation and nutrients removal at incubation time of 2 days (agitation condition) and 5 days (aeration condition), respectively. Consequently, a technical reference is provided for the microalgae coupled with other treatment processes.