Abstract: This study aimed to investigate the effects of different barnyardgrass varieties on yield formation of rice. A Japonica rice cultivar, Nanjing 9108, was used and co-cultured with three barnyardgrass varieties from transplanting to maturity under different nitrogen (N) levels of 0, 120, 240, and 360 kg N·hm^-2, taking baynyardgrass free as control. The three barnyardgrass varieties were Echinochloa crusgalli var. mitis, E. crusgali var. zelayensis and E. colonum. The results showed that, at the same N level, the plant height of the three barnyardgrass varieties was E. crusgali var. zelayensis > E. crusgalli var. mitis > E. colonum, and the growth duration was E. crusgalli var. mitis > E. crusgali var. zelayensis > E. colonum. The biomass of baynyardgrass increased with the increase of N application rates and reached a peak at the N rate of 240 kg N·hm^-2, and it decreased at 360 kg N·hm^-2. The biomass of either E. crusgalli var. mitis or E. crusgali var. zelayensis was significantly higher than that of E. colonum. At the 0 kg N· hm^-2 level, all barnyardgrass varieties showed no significant effect on rice yield. At the 120 kg N· hm^-2 level, rice yield was not significantly different among the three treatments of barnyardgrass free, co-cultured with E. crusgalli var. mitis, and co-cultured with E. colonum, but it was significantly decreased when co-cultured with E. crusgali var. zelayensis. At the 240 kg N·hm^-2 level, all treatments of co-cultured with barnyardgrass significantly decreased the rice yield. At the 360 kg N·hm^-2 level, rice yield was significantly decreased under the treatments of co-cultured with E. crusgali var. zelayensis or with E. crusgalli var. mitis, and showed no significant difference between barnyardgrass free and co-cultured with E. colonum. All these data indicated an interaction between barnyardgrass and N fertilizer, which mediated the formation of grain yield of rice. Furthermore, at the 120 kg N· hm^-2 level, the co-cultured E. crusgali var. zelayensis treatment significantly reduced leaf nitrate reductase activity and photosynthetic rate and root oxidation activity during the grain filling period, and decreased nitrogen accumulation and dry matter accumulation at the maturity stage, but other two treatments showed no significant effect when compared with barnyardgrass free treatment. These physiological indices of rice were significantly reduced by both E. crusgalli var. mitis and E. crusgali var. zelayensis treatments at either 240 or 360 kg N·hm^-2 level, and showed no significant difference among all treatments at the 0 kg N·hm^-2 level. Regression analysis showed that the order of effects of barnyardgrass phenotypes on rice grain yield was biomass, plant height, growth duration and tiller number. All these results suggested that the coexistence with large biomass of barnyardgrass inhibited the leaf photosynthetic rate, nitrate reductase activity, root oxidation activity, nitrogen accumulation and dry matter accumulation of rice, and consequently, reduced the rice grain yield.