& Key message High levels of genetic diversity, pronounced genetic structure and limitations to gene flow in Serbian spruce, a rare and endangered tree species from the refugial Balkan region, point towards a "one population-one unit" strategy for assembling a network of Genetic Conservation Units (GCUs) for its dynamic conservation. On the other hand, genetic information also permits to prioritize populations for conservation based on their contribution to genetic diversity and differentiation. & Context Serbian spruce, Picea omorika (Panč.) Purk., is a rare, IUCN red-listed European conifer endemic to the Balkan region. Its current rigid conservation (without any intervention allowed in~30 remnant populations) and the extant network of Genetic Conservation Units (four natural populations and three planted stands from the western part of the species range, in the Republic of Srpska, Bosnia and Herzegovina, RS-BH) might be ineffective in preserving the species' genetic diversity. & Aims To facilitate implementation of dynamic conservation of Serbian spruce by reassessing the number and size of remnant populations in RS-BH and updating genetic knowledge on these understudied western Serbian spruce populations. & Methods Comprehensive field survey in RS-BH, genotyping 689 individuals from 14 western populations with ten highly informative nuclear EST-SSRs and analytical methods for prioritizing populations for conservation based on their contribution to the geographical structuring of genetic diversity. & Results The genetic diversity of western Serbian spruce populations (Ae = 2.524, H E = 0.451) is comparable with what was found for eastern ones; they are highly genetically differentiated (Hedrick's G' ST = 0.186; Jost's D = 0.097) and comprise ten distinct gene pools. Effective population size is often ≥ 15. As much as 14% of alleles is not preserved in the extant GCUs established in natural populations. Eight populations positively contribute to within-population genetic diversity, four to genetic differentiation, and two are globally important in terms of diversity and differentiation. Although wildfires may contribute to admixture of different gene pools, re-establishment from seeds from extirpated populations has likely prevailed in studied populations. & Conclusions A larger network of GCUs is required for the dynamic conservation of western Serbian spruce populations. A "one population-one unit" strategy, with 14 GCUs, would represent the safest approach to conserve species extant genetic variation in this part of the species range. Nonetheless, a strategy to prioritize populations for conservation based on their contribution to allelic diversity has been put forward. Given the rapid global warming and peculiarities of Serbian spruce distribution, habitat and life history traits, conservation measures based on a rigorously designed GCU network are urgent for its rescue and survival.