Due to its beneficial effects on river ecosystems, black alder (Alnus glutinosa) is one of the tree species selected for planting on riverbanks in the cross-border area encompassing Wallonia in Belgium, Lorraine in France, and Luxembourg. The preservation of this species, however, is threatened by an invasive pathogen that particularly targets and kills young alder individuals. The objectives of this study were to characterize the genetic diversity and the genetic structure of A. glutinosa at this local level with the aim of assisting the conservation and replanting strategies and to determine if a germplasm collection comprising individuals from the same cross-border area captures the diversity present in the region. Nuclear simple sequence repeat (SSR) and chloroplastic DNA (cpDNA) markers were used to analyze four local wild populations and the germplasm collection which is representative of two river catchments and six legal provenance regions. Three populations distant from the studied area were also included. A panel of 14 nuclear SSR loci revealed high allelic diversity and very low differentiation among wild populations (mean F-ST = 0.014). The germplasm collection displayed a range of alleles that were representative of the different populations, and no significant differentiation between the germplasm collection and the local wild populations was observed, making this collection, as far as allelic diversity is concerned, suitable for providing trees for riverbank replanting programs. Using SSR markers, various statistical approaches consistently indicated the lack of a significant geographical structure at the level of the river catchments or provenance regions. In contrast, two cpDNA haplotypes were detected and displayed a cross-border geographically structured distribution that could be taken into account in defining new cross-border provenance regions.