Unveiling the Ongoing Expansion of Bullheads of the Genus Ameiurus (Actinopteri: Siluriformes) in the Dnipro River Basin, Ukraine: New Distributional Records and Genetic Diversity

ABSTRACT Invasive bullheads of the genus Ameiurus are actively expanding across Eastern Europe, yet accurate species identification remains problematic owing to phenotypic plasticity and high intraspecific morphological variability. This study aims to resolve the taxonomic identity, spatial distribution, and invasion dynamics of Ameiurus populations within the Dnipro River catchment, Ukraine, where mapping the current frontiers of these non‐native species is critical for freshwater conservation. We employed an integrated framework combining laboratory molecular analysis (mtDNA COI barcoding) with field‐based morphological scoring via Discriminant Analysis of Principal Components (DAPC). Sampling was conducted across multiple riverine networks within the catchment to evaluate whether range expansion is driven by natural downstream dispersal or human‐mediated pathways. A total of 66 specimens from six sampling localities were analysed, yielding 66 COI sequences. Phylogenetic reconstruction consistently recovered two discrete, high‐support clades corresponding to A. nebulosus and A. melas , a topology consistently supported across multiple distance‐based and phylogeny‐based delimitation methods. These results document the continued range expansion of both species in the Dnipro River basin, including the first molecularly confirmed occurrence record of A. melas in this basin, previously known only from the Southern Buh and Danube basins. Genetic analysis revealed five COI haplotypes per species, most of which are new to Ukraine, a pattern consistent with multiple independent introduction events or the retention of standing genetic variation from diverse source populations. DAPC profiling identified body colouration, pectoral spine serration, anal fin membrane pigmentation, and gill raker count as the most robust diagnostic traits for reliable field delimitation. The observed lack of isolation‐by‐distance and the allopatric establishment of both species in adjacent water bodies support a ‘jumpy’ colonisation pattern driven by unauthorised human‐mediated translocations rather than natural dispersal. This study provides a scalable, integrated framework to resolve taxonomic ambiguities in cryptic or plastic invasive fish populations. Our findings establish an ecological baseline for range forecasting and risk assessment of Ameiurus in European inland waters, underscoring the urgent need for coordinated molecular monitoring at the leading edges of freshwater biological invasions.