Aim of study: The goal of the study described in this manuscript was to evaluate qualitative and quantitative changes in weed flora infes - ting winter wheat stands grown in crop rotation and wheat monoculture. Area of study: South-eastern Poland, Europe (2018-2020). Materials and methods: The experiment was established in the system of randomized blocks (25 m × 6 m) in three replications. Winter wheat was sown in (1) crop rotation (CR): potato – winter wheat – peas – winter barley; and (2) in monoculture (MON). Weed infestation was evaluated in two terms: (1) at the tillering stage and (2) at the waxy maturity stage of winter wheat. Main results: The number and air-dry weight of weeds evaluated in both terms were higher in MON than in CR. Before wheat harvest, its plots in MON were massively infested by Apera spica-venti and significantly populated by Avena fatua and Anthemis arvensis. In this evaluation term, the weeds of the upper and middle levels accounted for 88.4% of the whole weed community in CR and for 97.7% in MON. In both terms of evaluation, greater biodiversity of the weed community was observed in CR than in MON. Research highlights: Indicators used to assess the weed flora in crop rotation and monoculture, i.e. weed species composition, number and air-dry weight of weeds, weed distribution in wheat crop levels, and Shannon-Wiener's diversity index.