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Background: The application of organic fertilizer is a sustainable approach to maintain soil fertility in agricultural crop production. In contrast to other organic fertilizers, the digestate from by-products of anaerobic digestion has not been well characterized in terms of its agronomic properties. In this study, different fertilization treatments were investigated to evaluate their impacts on growth, yield and fruit quality of tomatoes and on soil properties under greenhouse and field conditions. The experiments comprised a control (unfertilized) and three treatments with the same nitrogen dose: chemical fertilizer, digestate from by-product (organic fertilizer) and digestate combined with chemical fertilizer. Results: The results showed that the application of digestate significantly increased the growth and fruit quality of tomato including height, stem diameter, leaf chlorophyll content index, and photosynthetic rate of tomato plant and sugar–acid ratio, protein content, and ascorbic acid content of the fruit. The nitrate contents in tomato fruit were lower in the digestate treatment and digestate combined with chemical fertilizer treatment than in the chemical fertilizer. The digestate combined with chemical fertilization resulted in the greatest increase in tomato yield, up to 26.29% and 10.78% higher than that in the chemical fertilizer treatment under field and greenhouse conditions, respectively. Moreover, fertilization with digestate treatment and digestate combined with chemical fertilizer treatment increased soil fertility, including soil nitrogen and carbon contents, and enhanced soil enzyme activities under both field and greenhouse conditions. In addition, the growth, yield, and fruit quality of tomato were significantly correlated with soil chemical characteristics and soil enzyme activities. Conclusions: The effects of digestate treatments to maintain a stable tomato yield and improve fruit quality may be due to the enhanced soil enzymatic activities and chemical properties. These results suggest that the use of digestate as a full or partial replacement for chemical fertilizer could improve the growth and fruit quality of tomato, maintain the yield, and reduce the use of inorganic fertilizers in tomato production. [ABSTRACT FROM AUTHOR]