## 4 Conclusions  
Germplasm diversity assessment helps in identifying genotypes useful in crop improvement programs whose effectiveness depends on the availability of various genetic resources. The results obtained indicate a high level of genetic diversity in wild populations of _Humulus lupulus_ L. and a low level of genetic differentiation among populations. Therefore, extensive sampling per population is clearly required to assess the genetic diversity of hop populations. Sampling strategies involving sampling across a large number of localities represented by only a few samples could lead to erroneous conclusions. In addition, there appears to be no isolation by distance between populations separated by up to 150 km. The results obtained indicate that there is a persistent gene flow between populations and that genetic material is frequently exchanged between populations, despite the fact that the sampling area is one of the most populated and infrastructurally developed regions in Croatia. The results show great potential of wild hop populations from Croatia for use in future breeding programs. Further studies will aim to sample the entire distribution range of wild hop in Croatia to gain better insight into genetic, morphological, and chemical variability in order to select the best performing genotypes for the development of new varieties.  
**Supplementary Materials:** The following are available online at [[https://www.mdpi.com/article/10](https://www.mdpi.com/article/10)]([https://www.mdpi.com/article/10](https://www.mdpi.com/article/10)) _3390/app11146484/s1, Table S1: Geographical and climatic data of eight wild hop sampling sites in northwestern Croatia, Table S2: Pairwise \(F_{ST}\) values between wild hop populations (lower diagonal) and their significance (upper diagonal). ns = non-significant value;* = significant at 0.01 \(<p<\) 0.05, Table S3: Microsatellite data of eight wild hop populations in northwestern Croatia, Figure S1: Log-likelihood values [\(ln\) Pr(X\(|\) K)] for each of the thirty independent runs of each K and ALK values for each K as inferred from microsatellite data of wild hop samples performed (A) without prior information on the populations to which the individuals belong and (B) with prior information on the populations (LOCPRIOR) using STRUCTURE, Figure S2: Genetic structure of wild hop populations based on STRUCTURE analysis of microsatellite data for K = 1 to 5 performed (A) without prior information on the populations to which the individuals belong and (B) with prior information on the populations (LOCPRIOR) option. Each vertical bar represents an individual genotype, with the probability of membership (Q) to genetic clusters differentiated by colour, Figure S3: Isolation-by-distance (IBD) plot showing the relationship between pairwise geographic [\(ln(\text{km})\)] and genetic (\(F_{ST}/1-F_{ST}\)) distances between eight wild hop populations in Croatia.  
**Author Contributions:** Conceptualization, S.S. and Z.S.; methodology, Z.S., S.S., J.J. and Z.L.; formal analysis, Z.S., Z.L., F.V.; investigation, S.S., Z.S., R.E.; writing--original draft preparation, M.G., Z.S., S.S., F.V.; writing--review and editing, M.G., Z.S., S.S. and J.J. All authors have read and agreed to the published version of the manuscript.  
This research received no external funding.  
**Institutional Review Board Statement:** Not applicable.  
**Informed Consent Statement:** Not applicable.  
**Data Availability Statement:** The data presented in this study are available in [Table S3].  
The authors declare no conflict of interest.