Sculpins occasionally form interspecific hybrids in areas of mutual contact. These can be verified genetically and, potentially, morphologically. At the study site (Svratka River, Czech Republic), Cottus gobio are common whereas C. poecilopus represent a geographically isolated alien population. Potential hybrids between these two species were compared with the parent populations to establish simple identification criteria. Analysis of nuclear and mitochondrial DNA confirmed the existence of a new European hybridisation zone. Hybrids were the result of unidirectional crossing, with female C. gobio taking the maternal position. With the exception of the lateral line position, morphometric characteristics and the epidermis structure were similar in many traits, reflecting the contribution of both species, as well as in the number of sacciform cells where hybrids were similar to C. poecilopus. The vertical position of the lateral line allows recognition of hybrids directly in the field, a useful feature for future studies. The occurrence of these hybrids confirms expansion of non-native C. poecilopus at the expense of native C. gobio in rivers under anthropogenic pressure.