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In high-yield irrigated rice systems, options for legume cover crops with the capacity for biological nitrogen (N) fixation in the period between harvest have been sought. However, options are limited due to excess water and low oxygen level in the soil. The persian clover (Trifolium resupinatum) is an exception as it is a promising species due to its adaptation to waterlogged soils. Thus, the objective of this work was to evaluate accumulated nitrogen in the legume, soil mineral and microbial N levels and the response of a rice hybrid established in succession to persian clover during four growing seasons in subtropical conditions in southern Brazil. The field experiment was carried out during four crop seasons in a 2 × 4 factorial, with two conditions of soil cover, fallow and legume (persian clover) under four levels of N (0, 60, 120 and 180 kg N ha−1). It was observed that the cultivation of Persian clover increased the availability of mineral and microbial N in the soil by 35% and 6.5%, respectively. There was a 14.1% reduction in the optimal nitrogen rate in the rice hybrid. The grain yield of rice grains using persian clover in winter is 7.8% higher in relation to fallow, with the greatest difference between clover and fallow at the lowest N rates (0 e 60 kg ha−1). Thus, persian clover is a promising legume to be used in the period between harvest in hydromorphic soils cultivated with irrigated rice in subtropical conditions. [Display omitted] • Persian clover increase 35% soil mineral nitrogen. • Use of legume (Persian clover) reduces nitrogen fertilization by 14.1%. • Persian clover legume increases rice grain yield by 7.8%. [ABSTRACT FROM AUTHOR]