Analyses of mitochondrial (mt) DNA and microsatellite variation were carried out to examine the relationships between 10 freshwater populations of three-spined sticklebacks Gasterosteus aculeatus along the eastern coast of the Adriatic Sea. Partial sequences of the mtDNA control region and cytochrome b gene, in addition to 15 microsatellite loci, were used to analyse populations from four isolated river catchments. Results uncovered an Adriatic lineage that was clearly divergent from the European lineage, and confirmed that the most divergent and ancient populations are located within the Adriatic lineage as compared with other European populations. Two northern Adriatic populations formed independent clades within the European mitochondrial lineage, suggesting different colonization histories of the different Adriatic populations. Nuclear marker analyses also indicated deep divergence between Adriatic and European populations, albeit with some discordance between the mtDNA phylogeny of the northern Adriatic populations, further highlighting the strong differentiation among the Adriatic populations. The southern populations within the Adriatic lineage were further organized into distinct clades corresponding to respective river catchments and sub-clades corresponding to river tributaries, reflecting a high degree of population structuring within a small geographic region, concurrent with suggestions of existence of several microrefugia within the Balkan Peninsula. The highly divergent clades and haplotypes unique to the southern Adriatic populations further suggest, in accordance with an earlier, more limited survey, that southern Adriatic populations represent an important reservoir for ancient genetic diversity of G. aculeatus.

Fish introductions, particularly in areas of high biological diversity and endemism, represent a major threat for biodiversity. In the Balkan Peninsula, 60 fish species have been introduced to date, of which 36 have become naturalized in inland waters. Since the Balkans are one of the world's 35 biodiversity hot spots, this large presence of alien fish species poses a serious threat for the stability of freshwater ecosystems and the survival of the native ichthyofauna and of aquatic biodiversity in general. The motivation for the introductions, and the historical timeline, varies among the Balkan states. Despite recent attempts to implement and align legislation aimed at preventing the introduction of potentially invasive species, and the implementation of rigorous controls of introductions and increased protection of open waters, the majority of current introductions remain intentional, primarily via aquaculture. This review article provides a historical overview of freshwater fish introductions, the motivation behind them and the current distribution of alien freshwater fishes in the Balkans. The ecological implications and future perspectives concerning alien fish species in the region are also discussed.

The reproductive biology and gonadogenesis cycle of the Vrgorac goby, Knipowitschia croatica is described [Conservation of Endangered Freshwater Fish in Europe, Birkhauser Verlag, Basel]. The species displays sexual dimorphism during the spawning period. Sexual maturity is achieved at an early age, with 50% of males and females sexually mature at total lengths of 40-45 mm. Fecundity of gravid females ranged from 188 to 593 eggs, with an egg diameter of 0.22-1.11 mm. Though the extended spawning period lasts from March to November, the highest intensity is observed from April to September. A comparison is made of the reproductive biology of this species with other sand goby species of the genera Knipowitschia and Pomatoschistus.

and degree to which populations are expected to respond to natural selection is thought to depend significantly on demographic and genetic characteristics of populations and the traits under selection (Lynch & Walsh, 1998). For instance, small populations are less likely to carry potentially adaptive alleles and are more likely to be subject to strong genetic drift depleting genetic variation and reducing the effect of selection than large populations, and thus are probably subject to reduced adaptive potential (Lanfear et al., 2014;Stern & Orgogozo, 2009;Willi et al., 2006). Constraints on local adaptation may also be imposed by the genetic architecture of the traits under selection (Arnold, 1992;Kemppainen et al., 2021), or strong gene

Morphometric differentiation among freshwater fish populations is a commonplace occurrence, although the underlying causes for this divergence often remain elusive. We analysed the degree and patterns of morphological differentiation among nine freshwater three-spined stickleback (Gasterosteus aculeatus) populations inhabiting isolated karst rivers of the Adriatic Sea basin, to characterise the phenotypic diversity and differentiation in these populations. The analyses revealed marked and significant morphometric differentiation -especially in traits related to predator defence amongst most populations -even among those located within close geographic proximity in the same catchment system. Accordingly, the degree of morphometric and neutral genetic differentiation, as assessed from variability in 15 microsatellite loci from a parallel study, were uncorrelated across the populations. However, PST/FST comparisons revealed that the degree of phenotypic differentiation (PST) among populations exceeded that to be expected from genetic drift alone, suggesting a possible adaptive basis for the observed differentiation. In fact, avian predation pressure and several physiochemical environmental variables were identified as the main putative drivers of the observed differentiation, particularly in the dorsal spines, ascending process and lateral plates. Hence, the high degree of morphometric differentiation among Adriatic three-spined stickleback populations appears to reflect adaptation to local ecological conditions.