Increased biodiversity can make valuable contributions to food production and security around the world. The role of plant species diversity for “ecological intensification” of agriculture has been widely recognised, but the potential contribution of multi-trophic-level production systems, such as rice-fish co-culture, has received less attention. A continuous 4-year experiment (2015–2018) was conducted comparing rice-fish (yellow finless eel and loach) co-culture, and mono-rice planting practices on the Chongming Eco-island of China. During the experiment, pests (insect herbivores and weeds), arthropods, pesticides, grain and marketable fish yield were sampled, soil quality (available nitrogen, phosphorus and potassium, total nitrogen, organic matter content and pH) and rice grain quality (protein content, chalkiness, gel consistency, amylose content) were evaluated, and an economic analysis were performed. Fish decreased herbivore insect abundance by 24.07%, reduced weeds abundance, richness and biomass by 67.62, 62.01 and 58.88% respectively, increased invertebrate predator abundance by 19.48%, and reduced the need for pesticide by 23.4%. Co-culture practice produced an average economic values 10.33% higher than in the mono-rice farming. In addition, rice-fish co-culture enhanced both soil and rice quality. Our results confirm that rice-fish co-culture can be an effective form of ecological intensification, incorporating and contributing ecosystem services in agricultural production and increasing sustainability.