Amaranthus tuberculatus, commonly known as the rough-fruited water-hemp, is a dioecious annual weed species originating from North America. It has been introduced into Europe and Asia, where it is mostly considered a naturalized or casual alien occupying riparian areas, agricultural fields and disturbed habitats, especially in commercial port areas. Here we report the first occurrence of the species in Croatia, the second in Southeastern Europe. Flowering male and female individuals were found in 19 localities in the Pannonian part of Croatia. Female individuals from all localities had flowers with only one tepal and a dehiscent fruit and were therefore identified as A. tuberculatus var. rudis. In the studied area, the species was most frequently recorded along the banks of the Sava River and eutrophicated drainage channels passing through arable land. Here, it was present within vegetation of the alliance Nanocyperion Koch 1926 (class Isoëto-Nanojuncetea Br.-Bl. et Tx. in Br.- Bl. et al. 1952) and the alliance Bidention tripartitae (class Bidentetea Tx. et al. ex von Rochow 1951), as well as on the edges of alluvial willow and poplar forests (Salicion albae Soó 1951, class Salicetea purpureae Moor 1951). Furthermore, it was recorded on river sandbars, concrete embankments and other disturbed habitats (along the railroad, a road, on a construction site). Although the species is a well known agricultural weed and the majority of our study area is composed of arable land, A. tuberculatus was not detected in this habitat type. However, we believe that it is highly probable that it will spread into new areas and suitable habitats, and strongly recommend the occupied area to be monitored.