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The European weather loach (Misgurnus fossilis) is classified as highly endangered in several countries of Central Europe. Populations of M. fossilis are predominantly found in ditches with low water levels and thick sludge layers and are thus hard to detect using conventional fishing methods. Therefore, environmental DNA (eDNA) monitoring appears particularly relevant for this species. In previous studies, M. fossilis was surveyed following eDNA water sampling protocols, which were not optimized for this species. Therefore, we created two full factorial study designs to test six different eDNA workflows for sediment samples and twelve different workflows for water samples. We used qPCR to compare the threshold cycle (Ct) values of the different workflows, which indicate the target DNA amount in the sample, and spectrophotometry to quantify and compare the total DNA amount inside the samples. We analyzed 96 water samples and 48 sediment samples from a pond with a known population of M. fossilis. We tested several method combinations for long‐term sample preservation, DNA capture, and DNA extraction. Additionally, we analyzed the DNA yield of samples from a ditch with a natural M. fossilis population monthly over one year to determine the optimal sampling period. Our results showed that the long‐term water preservation method commonly used for eDNA surveys of M. fossilis did not lead to optimal DNA yields, and we present a valid long‐term sample preservation alternative. A cost‐efficient high salt DNA extraction led to the highest target DNA yields and can be used for sediment and water samples. Furthermore, we were able to show that in a natural habitat of M. fossilis, total and target eDNA were higher between June and September, which implies that this period is favorable for eDNA sampling. Our results will help to improve the reliability of future eDNA surveys of M. fossilis. [ABSTRACT FROM AUTHOR]

Identification of the geographic extent of population boundaries, the distribution of genetic lineages, and the amount of genetic exchange among breeding groups is needed for effective conservation of vulnerable marine migratory species. This is particularly true of the flatback turtle (Natator depressus), which only breeds in Australia but has extensive migrations that can include international waters.This study investigated the phylogeography and genetic structure among 17 flatback turtle rookeries across their range by sequencing an 810 bp portion of the mitochondrial DNA in 889 samples and genotyping 10 microsatellite loci in 598 samples.There was low phylogenetic divergence among haplotypes and evidence of recent population expansion, likely in the late Pleistocene. A predominant haplotype was found across all rookeries, but other haplotype groups were regionally specific.In general, there was agreement in patterns of genetic differentiation in the mitochondrial DNA and microsatellite data, and in some pairwise comparisons a higher mutation rate of microsatellites provided stronger evidence of differentiation.These results suggest natal philopatry operates in the choice of breeding locations for males as well as females.Evidence of genetic connectivity among neighbouring rookeries led to the identification of seven genetic stocks. Geographic boundaries of rookeries used by genetic stocks varied widely (160–1,300 km), highlighting a need for field studies to better understand movement patterns.Hierarchical analysis of molecular variance identified significant genetic differentiation based upon genetic stock, nesting phenology (summer vs. winter nesters), and a west–east discontinuity across Torres Strait. A pattern of isolation by distance was identified, which was most strongly observed in the microsatellite data.In combination with tagging and telemetry studies, these results will allow better quantification of stock‐specific threats along migratory routes and in foraging habitats. Implications of climate change will be stock specific and may depend upon the extent of genetic connectivity between neighbouring stocks. [ABSTRACT FROM AUTHOR]

The Upper Rhine Valley, a "hotspot of biodiversity" in Germany, has been treated with the biocide Bacillus thuringiensis var. israelensis (Bti) for mosquito control for decades. Previous studies discovered Bti nontarget effects in terms of severe chironomid abundance reductions. In this study, we investigated the impact of Bti on species level and addressed the community composition of the nontarget family Chironomidae by use of community metabarcoding. Chironomid emergence data were collected in three mosquito‐control relevant wetland types in the Upper Rhine Valley. For all three sites the chironomid species composition, based on operational taxonomic units (OTUs), was different to varying degrees in the Bti‐treated samples versus control samples, ranging from a significant 63% OTU reduction to an OTU replacement. We assumed that predatory chironomids are less prone to Bti than filter feeders, as the latter feed on floating particles leading to direct ingestion of Bti. However, a comparable percentage of predators and filter feeders (63% and 65%, respectively) was reduced in the Bti samples, suggesting that the feeding strategy is not the main driver for Bti sensitivity in chironomids. Finally, our data was compared to a three‐year‐old data set, indicating possible chironomid community recovery due to species recolonization a few years after the last Bti application. Considering the currently discussed worldwide insect decline we recommend a rethinking of the usage of the biocide Bti, and to prevent its ongoing application especially in nature protection reserves to enhance ecological resilience and to prevent boosting the current biodiversity loss. [ABSTRACT FROM AUTHOR]

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 16 S r RNA 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. [ABSTRACT FROM AUTHOR]

Aim General models for understanding the climate-driven processes of post-glacial colonization in European arctic-alpine species are mainly derived from studies on temperate terrestrial taxa. However, cold-adapted freshwater species may tolerate or even thrive under colder climatic conditions as flowing water habitats are thermally buffered against freezing and extremely cold temperatures. Here, we investigate the European Pleistocene and Holocene history of the arctic-alpine stonefly Arcynopteryx dichroa. Location Europe. Methods We used two genetic data sets (mitochondrial sequence data and nuclear microsatellite data) to investigate the glacial survival and post-glacial recolonization routes of A. dichroa. We used species distribution models to critically evaluate our genetic data and phylogeographical interpretations. Results Among 344 sequenced individuals from eight European mountain ranges, 80 unique haplotypes were detected. Of these, 77 haplotypes were endemic to a single mountain range, indicating almost complete lineage sorting. Both sequence and microsatellite data suggested strong population differentiation between mountain ranges. The genetic hotspots were found in the Carpathians, the Balkans and the eastern Alps. The Black Forest and Fennoscandian populations exhibited shared and closely related haplotypes, indicating ancestral polymorphism in two populations that became disjunct due to vicariance or resulting from rare long-distance dispersal among disjunct northern and southern periglacial populations. Main conclusions Arcynopteryx dichroa is a glacial relict that survived glacial cycles through elevation shifts in isolated periglacial populations in the Pyrenees, the central European highlands, the Carpathians, the Balkans and the eastern Alps. The species probably recolonized the formerly glaciated Fennoscandian range from a refugium in the central European highlands, following the retreat of the ice sheet. This study suggests that aquatic organisms may have reacted differently to Pleistocene climate change compared with terrestrial species. [ABSTRACT FROM AUTHOR]

Summary 1. We investigated the Pleistocene and Holocene history of the rare mayfly Ameletus inopinatus EATON 1887 (Ephemeroptera: Siphlonuridae) in Europe. We used A. inopinatus as a model species to explore the phylogeography of montane, cold-tolerant aquatic insects with arctic-alpine distributions. 2. Using species distribution models, we developed hypotheses about the species demographic history in Central Europe and the recolonisation history of Fennoscandia. We tested these hypotheses using mitochondrial cytochrome oxidase I (mtCOI) sequence data and compared our genetic results with previously generated microsatellite data to explore genetic diversity distributions of A. inopinatus. 3. We observed old lineages, deep splits and almost complete lineage sorting of mtCOI sequences among mountain ranges. These results support a periglacial survival, i.e. persistence at the periphery of Pleistocene glaciers in Central Europe. 4. There was strong differentiation between the Fennoscandian and all other populations, indicating that Fennoscandia was recolonised from a refugium not accounted for in our sampling. High degrees of population genetic structure within the northern samples suggest that Fennoscandia was recolonised by more than one lineage. However, this structure was not apparent in previously published microsatellite data, consistent with secondary contact without sexual incompatibility or with sex-biased dispersal. 5. Our demographic analyses indicate that (i) the separation of northern and Central European lineages occurred during the early Pleistocene; (ii) Central European populations have persisted independently throughout the Pleistocene and (iii) the species extended its range about 150 000 years ago. [ABSTRACT FROM AUTHOR]

We describe the isolation of 11 polymorphic trinucleotide microsatellite loci from the stonefly Arcynopteryx compacta. Loci were highly variable with 3 to 14 alleles (mean = 6.45). Observed heterozygosity ranged from 0 to 0.867. Seven loci showed significant deviation from Hardy–Weinberg equilibrium across both populations. There was no evidence for null alleles, and thus, Hardy–Weinberg departures could have resulted from genetic structure between populations or subpopulations. No linkage between loci was found. The 11 loci should prove highly informative for population genetic studies. [ABSTRACT FROM AUTHOR]

We describe the isolation of ten polymorphic microsatellite loci from the mayfly Ameletus inopinatus. Loci had di- or trinucleotide repeat motifs and were highly variable with three to 17 alleles (mean = 7.15). Observed heterozygosity ranged from 0.143 to 0.905. One locus (Ami_202) showed significant deviation from Hardy–Weinberg equilibrium in one population, but no evidence for null alleles. One locus (Ami_73) was significantly linked with three other loci. The remaining nine loci should prove highly informative for population genetic studies. [ABSTRACT FROM AUTHOR]