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Weeds are a concern to farmers mainly by reducing crop yield, especially in organic farming where herbicides are not an option. Additionally, reducing reliance on herbicides is among the goals to achieve sustainable crop production. By investigating alternatives for weed control, several studies have shown that grassland is more competitive with weeds than cash crops and adopting diversified crop rotations reduces overall weed infestation. However, in many of those studies the effect of the rotation is confounded with herbicide use. Moreover, there is a gap in understanding how diversified rotations influence the weed community and how changes in that weed community impact yield of the subsequent cash crop. Here, using a 4-year herbicide-free experiment in southern Brazil, we compared a grassland-cropping rotation (i.e., 3-year perennial tropical grassland before maize cultivation) vs. a crop-based rotation (i.e., sunflower-maize) to clearly characterize how a more diversified crop rotation changed weed patterns and weed interference in maize. We demonstrate the higher suppressive effect on weeds of a grassland-cropping rotation compared with a crop-based rotation, clearly evidenced in the early season of maize growth, in which the crop-based rotation had about twice as many number of weeds as the grassland-cropping rotation. However, there was no difference in seedbank size between both rotations. Weed community composition was not affected (i.e. rotations had 98% similarity in species composition), but weed assemblage was drastically changed with different rates of species-specific infestation in each rotation. The crop-based rotation selected weeds with rapid light acquisition ability (higher specific leaf area) that added to higher early season weed infestation and higher competitive pressure of weeds resulted in maize grain yield losses, whereas there was no significant weed interference on maize grain yield in the grassland-cropping rotation. The strategy of including a tropical grassland in rotation with cash crops as a herbicide-free weed management tool reduces overall weed infestation and prevents selection of weeds that cause a significant yield reduction in maize and represents an economically viable alternative. [ABSTRACT FROM AUTHOR]