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Overuse of chemical fertilizers and pesticides has led to severe ecological and food safety problems. The application of seaweed fertilizer meets the needs of green and sustainable agriculture. In the present study, we aimed to evaluate whether seaweed fertilizer was suitable for long-term application as a green fertilizer. Seaweed fertilizer derived from brown algae residues was applied to the okra planting field for 2 years, and the soil’s physical and chemical properties, enzyme activity indicators, and bacterial flora structure were tested. We found that seaweed fertilizer could effectively improve the soil pH, water content, organic carbon, alkali nitrogen, and the activities of various enzymes, such as catalase, sucrase, and cellulase. In addition, compared with the short-term use of seaweed fertilizer previously studied, our study showed that long-term fertilization could more significantly improve soil bacterial diversity. The abundance of some bacteria involved in polysaccharide degradation and nitrogen fixation was significantly increased, such as Acidobacteria and Cyanobacteria at the phylum level, and Methylobacterium, norank Acidobacteriales, and Micromonosporaat the genus level. This study showed that the long-term application of seaweed fertilizer was one of the effective ways to develop sustainable green agriculture.