In the past, the freshwater snail Theodoxus fluviatilis was abundant in the river Rhine, but was considered to be extinct in the Upper and HighRhine in 2004. We found individuals of T. fluviatilis for the first time at sites in the southern part of the Upper Rhine and High Rhine in 2020,where this species went extinct 16 years ago. There are several distinct haplotypes of this species; some of them occurring in geographicallyseparated regions of Europe. We used mitochondrial DNA (cytochrome c oxidase subunit I, COI) to trace the origin of the newly arrivedT. fluviatilis. We examined 269 individuals collected at 29 sites and compared their sequence data with known haplotypes of T. fluviatilisand other species of Theodoxus deposited in GenBank. We also analysed a historical sample from 1995 (collected before the species wentextinct), which revealed that the haplotype of the native T. fluviatilis in the High Rhine at that time was F28. In the recent samples, however,we found T. fluviatilis with the haplotypes F31 and “euxinus”, which are reported in the Ponto-Caspian region. Both haplotypes are crypticinvaders, colonizing the Rhine via the Rhine–Main–Danube canal and adversely affecting the remaining native populations of T. fluviatilisin Western Europe. Our findings are important for this species management by nature conservation authorities: lineages of endangeredspecies should be preserved and supported, while the spreading of invasive, non-native lineages (or species) should be prevented.