Summary Reconstructing the phylogeographic history of a species can aid in defining areas of conservation priority. For freshwater species, historical river structure plays a significant role in explaining genetic differentiation and population structure. However, human‐induced translocations can erase the natural genetic structure, especially for species of commercial interest such as the noble crayfish (Astacus astacus). Our aim was to reconstruct the current genetic structure of the endangered noble crayfish in central Europe to identify refugial areas that are hotspots of genetic diversity. We analysed a fragment of the mitochondrial cytochrome oxidase subunit I, and the 16S rRNA from 540 noble crayfish specimens from 156 sampling sites distributed around five European sea basins. Additionally, we conducted a microsatellite analysis of 289 individuals from 22 sites. Both mitochondrial and nuclear markers revealed genetically relatively homogenous populations in central Europe that had been influenced by anthropogenic translocations. However, some areas (Eider catchment in northern Germany and Rhineland‐Palatinate in south‐western Germany) show a distinct genetic structure with endemic haplotypes and private alleles indicating (i) that these areas were refugia for A. astacus in central Europe and (ii) that these populations have not been subject to anthropogenic translocations. Further, we found the highest genetic diversity in the Black Sea basin and particularly high differentiation between populations from the western Balkans and the remaining Black Sea populations. The split between Western Balkan and the remaining European populations is estimated to have occurred approximately 700 k years before present, whereas remaining differentiations occurred within the last 450 k years. Using migration modelling, we detected that the North Sea basin and the Baltic Sea basin were colonised independently via different colonisation paths from the eastern Black Sea basin, while the western Balkans did not contribute to this colonisation. Our results suggest the existence of at least two refugial areas in south‐eastern Europe. To conserve maximum genetic diversity, conservation priorities for noble crayfish should focus on the south‐eastern European genetic hotspots and on populations in central Europe that hold an autochthonous genetic structure (e.g. Langsee in the Eider catchment area). We further propose that each river catchment should form a separate management unit to reduce anthropogenic genetic homogenisation.

The parthenogenetic all-female marbled crayfish is a novel research model and potent invader of freshwater ecosystems. It is a triploid descendant of the sexually reproducing slough crayfish, Procambarus fallax, but its taxonomic status has remained unsettled. By cross-breeding experiments and parentage analysis we show here that marbled crayfish and P. fallax are reproductively separated. Both crayfish copulate readily, suggesting that the reproductive barrier is set at the cytogenetic rather than the behavioural level. Analysis of complete mitochondrial genomes of marbled crayfish from laboratory lineages and wild populations demonstrates genetic identity and indicates a single origin. Flow cytometric comparison of DNA contents of haemocytes and analysis of nuclear microsatellite loci confirm triploidy and suggest autopolyploidisation as its cause. Global DNA methylation is significantly reduced in marbled crayfish implying the involvement of molecular epigenetic mechanisms in its origination. Morphologically, both crayfish are very similar but growth and fecundity are considerably larger in marbled crayfish, making it a different animal with superior fitness. These data and the high probability of a divergent future evolution of the marbled crayfish and P. fallax clusters suggest that marbled crayfish should be considered as an independent asexual species. Our findings also establish the P. fallax–marbled crayfish pair as a novel paradigm for rare chromosomal speciation by autopolyploidy and parthenogenesis in animals and for saltational evolution in general.

The oomycete Aphanomyces astaci, the causative agent of crayfish plague, is listed as one of the 100 worst invasive species in the world, destroying the native crayfish populations throughout Eurasia. The aim of this study was to examine the potential of selected mitochondrial (mt) genes to track the diversity of the crayfish plague pathogen A. astaci. Two sets of primers were developed to amplify the mtDNA of ribosomal rnnS and rnnL subunits. We confirmed two main lineages, with four different haplogroups and five haplotypes among 27 studied A. astaci strains. The haplogroups detected were (1) the A-haplogroup with the a-haplotype strains originating from Orconectes sp., Pacifastacus leniusculus and Astacus astacus; (2) the B-haplogroup with the b-haplotype strains originating from the P. leniusculus; (3) the D-haplogroup with the d1 and d2-haplotypes strains originating from Procambarus clarkii; and (4) the E-haplogroup with the e-haplotype strains originating from the Orconectes limosus. The described markers are stable and reliable and the results are easily repeatable in different laboratories. The present method has high applicability as it allows the detection and characterization of the A. astaci haplotype in acute disease outbreaks in the wild, directly from the infected crayfish tissue samples.

1. Declines in populations of stone crayfish (Austropotamobius torrentium) have recently been reported throughout many parts of their range, including southern Germany. To assess the rate of decline and the probable causes, a crayfish survey was conducted in the River Argen catchment, where the species was known to be widely distributed and abundant.2. Using mixed-effects regression analysis, the presence or absence of the species was subsequently correlated with riparian land-use and habitat characteristics. To elucidate the potential role of crayfish plague in stone crayfish decline, the crayfish plague-carrier status of newly discovered alien signal crayfish (Pacifastacus leniusculus) was evaluated using real-time polymerase chain reaction (PCR).3. Twenty-five of the 98 surveyed sites were colonized by stone crayfish, including only 12 of 25 sites with a previous record. The probability of stone crayfish presence was positively correlated with overhanging bank structures and the proportion of broad-leafed woods in riparian buffers. In contrast, high proportions of intensive grassland had a distinct negative effect on the probability of stone crayfish presence, and cropland also tended to decrease the probability of stone crayfish presence. Both land-use forms have significantly increased in the study area within the last decade, signifying an intensification of riparian land-use.4. Plague-carrying signal crayfish were present at two headwater sites. Crayfish plague transmission from highly resistant signal crayfish to highly susceptible stone crayfish has most likely contributed to the decline in stone crayfish, and crayfish plague outbreaks may account for up to 67% of the observed stone crayfish disappearances.5. The present study confirms a marked decline in stone crayfish populations, which is probably driven by the persistent spread of invasive alien species and intensified riparian land-use. Therefore, conservation efforts for stone crayfish should incorporate strategies to prevent and control the spread of invasive, plague-carrying crayfish, and to mitigate pressures from land use.