BACKGROUND:The garden dormouse (Eliomys quercinus) is one of the fastest-declining mammals in Europe, and action is needed to prevent further population losses. The primary causes of declines are not well-understood, as the species experiences variable conditions and threats across its range, but likely include habitat fragmentation and loss. Previous genetic studies have provided evidence of highly structured garden dormouse populations in Western Europe, despite this region having been defined as a single clade with mitochondrial DNA analysis. Within Western Europe, the magnitude of declines has been recognized to be greater on the eastern edge of the species' range, which could explain differentiation within the clade as resulting from diversity loss and genetic drift for regions under greater risk of extirpation. Here, we focus on fine-scale genomic differentiation across the Western European clade to explore the consequences of genomic erosion on the eastern region and to help identify mechanisms driving genetic differentiation within this species. RESULTS:We found genetic differentiation both between and within major geographic regions. Populations located in the eastern edge of the species' range showed stronger signs of population isolation, including structure between spatially distant populations, lower genetic diversity, and greater rates of inbreeding. However, all populations exhibited signals of recent rapid population decline. Outlier analyses indicated that differentiation between regions was primarily due to genetic drift resulting from isolation-by-distance rather than adaptive differentiation. We also found genetic structuring between populations within the Rhine Valley, despite apparent lack of physical barriers preventing dispersal among groups within this region. CONCLUSIONS:Our findings indicate that population isolation following habitat loss and fragmentation has likely been a major contributor to garden dormouse declines. Dispersal among disparate garden dormouse sampling regions is restricted-even across local spatial scales-leading to loss of genetic diversity and potential erosion of evolutionary potential. With 21st century declines expected to continue across the species' range, even relatively common and well-connected populations are likely to follow the trajectory of the eastern populations, with increasing loss of diversity as populations contract and become more isolated.

While viral diseases of domestic cats (Felis catus) can threaten the recovery of the European wildcat (Felis s. silvestris), their epidemiology in wildcat populations remains poorly understood. Here, we analyzed 428 road-killed wildcats from Western and Central Germany for the presence of antibodies/antigens of six feline viruses. The presence of at least one viral antigen or antiviral antibody was detected in 53.3% of the animals. Antibodies against feline parvovirus (FPV) showed the highest seroprevalence (29.2%), while feline leukemia virus (FeLV) antigens were detected in 22.2% of the animals. Antibodies to feline coronavirus (FCoV), feline herpesvirus (FHV) and feline calicivirus (FCV) were detected in 10% or fewer of the wildcats. No antibodies to feline immunodeficiency virus (FIV) were detected. FeLV antigens clustered spatially, with prevalence declining from southwest to northeast, consistent with the geographic expansion of the virus antigens. Seroprevalence of FPV and prevalence of FeLV increased with age, suggesting cumulative exposure, while juvenile males were particularly unlikely to be seropositive for FPV. Proximity to built-up areas did not predict seroprevalence for any virus. FeLV and FPV in particular warrant further investigation as potential threats to wildcat recovery in Germany and highlight the need for longitudinal health monitoring alongside existing conservation efforts.

Sex chromosomes differ in their inheritance properties from autosomes and hence may encode complementary information about past demographic events. We compiled and analyzed a range-wide resequencing data set of the red deer (Cervus elaphus), one of the few Eurasian herbivores of the Late Pleistocene megafauna still found throughout much of its historic range. Our analyses of 144 whole genomes reveal striking discrepancies between the population clusters suggested by autosomal and X-chromosomal data. We postulate that the genetic legacy of Late Glacial population structure is better captured and preserved by the X chromosome than by autosomes, for two reasons. First, X chromosomes have a lower Ne and hence lose genetic variation faster during isolation in glacial refugia, causing increased population differentiation. Second, following postglacial recolonization and secondary contact, immigrant males pass on their X chromosomes to female offspring only, which effectively halves the migration rate when gene flow is male mediated. Our study illustrates how a comparison between autosomal and sex chromosomal phylogeographic signals unravels past demographic processes that otherwise would remain hidden.

Abstract Insect pollinators face numerous threats, including habitat loss and population fragmentation. The effects of human‐altered landscapes on population connectivity need to be better understood to inform effective mitigation measures. We examined the literature on the effects of landscape heterogeneity on population genetic connectivity in two key pollinator groups: bees (Hymenoptera: Anthophila) and hoverflies (Diptera: Syrphidae). We identified 113 studies from 59 countries, covering 96 bee and 21 hoverfly species. However, biased taxonomic and geographical coverage limited broad conclusions regarding species susceptibility to genetic isolation. Notably, hoverflies remain significantly understudied, hampering comprehensive assessments of genetic connectivity patterns. While some species demonstrated genetic differentiation across tens of kilometres, others maintained connectivity across entire continents. Various landscape features, including water bodies and mountain ranges, often acted as barriers to gene flow, while the impacts of deforestation, agriculture, and urbanisation on genetic connectivity were mixed. Biological traits like body size, resource specialisation, and effective population sizes were found to influence susceptibility to population isolation. However, contrasting results and limited species coverage precluded conclusive findings. Future research should include evaluations of genetic time lags and statistical power to determine the appropriateness of selected genetic tools for testing hypotheses of recent genetic effects of human‐altered landscapes on pollinators. Owing to increased interest in pollinator habitat corridors, population genetic research encompassing a wider range of species, habitats, and improved study designs, is needed to provide an evidence‐based framework for conservation measures.

AbstractHoverflies (Syrphidae) are essential pollinators, and their severe decline jeopardizes their invaluable contribution to plant diversity and agricultural production. However, we know little about the dispersal abilities of hoverflies in urbanized landscapes, limiting our understanding of the spatiotemporal dynamics of plant–pollinator systems and reducing our ability to preserve biodiversity in the context of global change. Previous work has not addressed how urbanization affects the functional connectivity of hoverflies and whether dispersal is a limiting factor in their population dynamics. In this study, we investigated the spatial genetic structure of two nonmigratory species of hoverflies in two urban areas. We collected more than a thousand specimens of each Syritta pipiens and Myathropa florea, each, by hand netting in two western European urbanized study areas of 490 and 460 km2 in 2021, and we genotyped them at 14 and 24 microsatellite loci, respectively. Based on spatial and nonspatial Bayesian clustering methods, we failed to reject the null hypothesis of panmixia, suggesting that both species exhibited high genetic connectivity despite urbanization. The distribution of allele frequencies was not correlated with geographic distance, implying that isolation‐by‐distance was negligible at the investigated spatial scale in both species. Although anthropogenic land cover changes generally have dramatic consequences on biodiversity, these hoverfly species retain high connectivity, which suggests that dispersal is not a strong limiting factor in their metapopulational dynamics. However, high effective population size and its confounding effect on signals of genetic drift may limit our ability to conclude confidently about landscape effects on gene flow in those two species. Provided we maintain or restore habitat, recolonization may be prompt even in urban areas.

Background Batrachochytrium dendrobatidis ( Bd ) and Batrachochytrium salamandrivorans ( Bsal ) are two pathogenic fungi that are a significant threat to amphibian communities worldwide. European populations are strongly impacted and the monitoring of the presence and spread of these pathogens is crucial for efficient decision-making in conservation management. Results Here we proposed an environmental DNA (eDNA) monitoring of these two pathogenic agents through droplet digital PCR (ddPCR) based on water samples from 24 ponds in Luxembourg. In addition, amphibians were swabbed in eight of the targeted ponds in order to compare the two approaches at site-level detection. This study allowed the development of a new method taking below-Limit of Detection (LOD) results into account thanks to the statistical comparison of the frequencies of false positives in no template controls (NTC) and below-LOD results in technical replicates. In the eDNA-based approach, the use of this method led to an increase in Bd and Bsal detection of 28 and 50% respectively. In swabbing, this resulted in 8% more positive results for Bd . In some samples, the use of technical replicates allowed to recover above-LOD signals and increase Bd detection by 35 and 33% respectively for eDNA and swabbing, and Bsal detection by 25% for eDNA. Conclusions These results confirmed the usefulness of technical replicates to overcome high levels of stochasticity in very low concentration samples even for a highly sensitive technique such as ddPCR. In addition, it showed that below-LOD signals could be consistently recovered and the corresponding amplification events assigned either to positive or negative detection via the method developed here. This methodology might be particularly worth pursuing in pathogenic agents’ detection as false negatives could have important adverse consequences. In total, 15 ponds were found positive for Bd and four for Bsal . This study reports the first record of Bsal in Luxembourg.

Human infection by Baylisascaris procyonis can result in larva migrans syndromes, which can cause severe neurological sequelae and fatal cases. The raccoon serves as the definitive host of the nematode, harboring adult worms in its intestine and excreting millions of eggs into the environment via its feces. Transmission to paratenic hosts (such as rodents, birds and rabbits) or to humans occurs by accidental ingestion of eggs. The occurrence of B. procyonis in wild raccoons has been reported in several Western European countries. In France, raccoons have currently established three separate and expanding populations as a result of at least three independent introductions. Until now the presence of B. procyonis in these French raccoon populations has not been investigated. Between 2011 and 2021, 300 raccoons were collected from both the south-western and north-eastern populations. The core parts of the south-western and north-eastern French raccoon populations were free of B. procyonis. However, three worms (molecularly confirmed) were detected in a young raccoon found at the edge of the north-eastern French raccoon population, close to the Belgian and Luxemburg borders. Population genetic structure analysis, genetic exclusion tests and factorial correspondence analysis all confirmed that the infected raccoon originated from the local genetic population, while the same three approaches showed that the worms were genetically distinct from the two nearest known populations in Germany and the Netherlands. The detection of an infected raccoon sampled east of the northeastern population raises strong questions about the routes of introduction of the roundworms. Further studies are required to test wild raccoons for the presence of B. procyonis in the area of the index case and further east towards the border with Germany.

1. Population genetics is a valuable tool for assessing the impact of human -altered landscapes on genetic connectivity in various species. However, when applied to insects, challenges arise due to potentially large effective population sizes (Ne), high dispersal capacities and the recency of anthropogenic impacts. 2. This study assessed the population genetic structure of two pollinators across a human -altered landscape in Luxembourg. Samples from the ashy mining bee (Andrena cineraria, N = 201) and the greater bee fly (Bombylius major, N = 637) were genotyped at 25 microsatellite loci, including a genotyping-by-ampliconsequencing approach for A. cineraria. 3. Despite high statistical power of FST > 0.002 in B. major and FST > 0.0025 in A. cineraria, no deviations from genetic homogeneity were detected. For both species, there was no evidence for isolation -by -distance or genetic clustering. Genetic homogeneity was most likely the result of high levels of gene flow that compensate for the effects of genetic drift. 4. Estimates of Ne ranged between several thousand to tens of thousands, although precision was low. Simulations highlighted that genetic lag times can substantially affect our ability to detect recent (<50 generations) population differentiation when Ne is very large. Lag times were shorter with data from 25,000 simulated di -allelic loci, but only when sample sizes remained high. 5. Insect genetic studies should consider lag times due to large Ne and ensure sample size and markers offer adequate power to reject the null hypothesis of no landscape effect on genetic connectivity.

Biological invasions are a major threat to biodiversity and have particularly devastating impacts on island ecosystems. The New Caledonia archipelago is considered a biodiversity hotspot due to its diverse native flora. Javan rusa deer ( Rusa timorensis ) were introduced to New Caledonia in 1870 and the population consists of several hundred thousand individuals today. They directly threaten rare endemic species and affect the composition and structure of the vegetation. While a rusa deer management plan has identified ten priority areas for deer control operations, removing deer could be offset by the dispersal of animals back into the control areas. Here, we genotyped 628 rusa deer using 16 microsatellite markers to analyse the genetic structure of the animals in New Caledonia. We aimed to assess fine-scale genetic structure, to identify natural barriers to deer movement and to assess functional connectivity by optimising individual-based landscape resistance models. Our results suggested that rusa deer formed a single genetic population on the main New Caledonian island. The isolation-by-distance pattern suggested that female dispersal was limited, whereas males had larger dispersal distances. We assessed functional connectivity using different genetic distance metrics and all models performed poorly (m R 2 ≤ 0.0043). Landscape features thus hardly affected deer movement. The characteristics of our results suggested that they were not an artefact of the colonisation history of the species. Achieving an effective reduction of deer population sizes in specific management areas will be difficult because of the deer’s high dispersal capabilities and impossible without very substantial financial investment.

Estimates of gene flow resulting from landscape resistance inferences frequently inform conservation management decision-making processes. Therefore, results must be robust across approaches and reflect real-world gene flow instead of methodological artefacts. Here, we tested the impact of 32 individual-based genetic distance metrics on the robustness and accuracy of landscape resistance modelling results. We analysed three empirical microsatellite datasets and 36 simulated datasets that varied in landscape resistance and genetic spatial autocorrelation. We used ResistanceGA to generate optimised multi-feature resistance surfaces for each of these datasets using 32 different genetic distance metrics. Results of the empirical dataset demonstrated that the choice of genetic distance metric can have strong impacts on inferred optimised resistance surfaces. Simulations showed accurate parametrisation of resistance surfaces across most genetic distance metrics only when a small number of environmental features was impacting gene flow. Landscape scenarios with many features impacting gene flow led to a generally poor recovery of true resistance surfaces. Simulation results also emphasise that choosing a genetic distance metric should not be based on marginal R2 -based model fit. Until more robust methods are available, resistance surfaces can be optimised with different genetic distance metrics and the convergence of results needs to be assessed via pairwise matrix correlations. Based on the results presented here, high correlation coefficients across different genetic distance categories likely indicate accurate inference of true landscape resistance. Most importantly, empirical results should be interpreted with great caution, especially when they appear counter-intuitive in light of the ecology of a species.

The raccoon roundworm (Baylisascaris procyonis), a gastrointestinal nematode parasite of the raccoon (Procyon lotor), is the causative agent of a zoonotic disease that can cause severe neurological sequelae and even fatal cases. The parasite is now established in Europe and, while not present in all raccoon populations, it is currently expanding its range. Understanding the geographic distribution of the roundworm is a necessity for public health. Here, we report the first detections of the raccoon roundworm from Luxembourg. We used genetic techniques to test whether the roundworm originated from a distinct introduction event or entered the country by natural dispersal. We detected two roundworms in a raccoon that was road-killed in Echternach, northeastern Luxembourg. Furthermore, in a wildlife rescue centre, roundworms were detected in the latrine of an enclosure containing seven raccoons. Using different genetic techniques, we showed the Echternach roundworms to originate from the nearest German population, while the rescue centre roundworms were genetically distinct. By genotyping the host from a roundworm DNA extract, we identified a raccoon originating from southwest Luxembourg as the host of at least one roundworm. Roundworms are thus colonising Luxembourg both by natural dispersal and from an introduced population. The (potential) hosts all originated from the local population, suggesting that imported/introduced roundworms have already spread to the local raccoon population. Authorities should act as though the parasite is already widespread in the country, raising awareness among the public and individuals with occupational contact with raccoons.

Abstract The intensification of agriculture and urbanisation has precipitated the decline of numerous species, including the northern crested newt (Triturus cristatus). To mitigate the impacts of habitat degradation and loss, restoration initiatives depend on evidence‐based decision‐making to identify priority sites for conservation efforts. Here, we employed a multidisciplinary approach, drawing on population genetics, species distribution modelling (SDMs) and landscape genetics to develop a spatial prioritisation framework for the creation and/or restoration of crested newt breeding ponds in Luxembourg. For the genetic analyses, more than 1000 crested newts from 98 ponds across Luxembourg were genotyped at 15 microsatellite loci to assess genetic diversity and spatial clustering. The relationship between newt presence and environmental variables was evaluated using SDMs. Priority restoration areas were identified with PRIORITZR, integrating genetic diversity, habitat suitability and connectivity. The population genetic structure was characterised by spatial clustering, which followed an isolation‐by‐distance pattern. Five sites were identified as significantly differentiated from the remaining populations, warranting targeted conservation efforts. Standing water bodies and proximity to the nearest occupied pond were the most important predictors of crested newt presence. The highest gain in habitat suitability through the creation of new ponds was predicted to occur in southwest Luxembourg. Based on the country‐wide predicted occurrence of crested newts and gains in habitat suitability, areas of highest restoration potential were identified in close proximity to existing crested newt populations. The produced maps of priority restoration areas should inform future conservation efforts of the crested newt in Luxembourg. While the importance of preserving genetic diversity is widely recognised, the theory‐practice gap frequently precludes the incorporation of genetic aspects into in situ conservation measures or management policies. This case study demonstrates the integration of multidisciplinary analyses within an evidence‐based framework to guide species conservation efforts.

The raccoon roundworm ( Baylisascaris procyonis ), a gastrointestinal nematode of the raccoon ( Procyon lotor ), may cause a severe form of larva migrans in humans, which can lead to death or permanent neurological damage. Although roundworms were inadvertently introduced to Europe alongside their raccoon hosts, the parasite is not present in every raccoon population. It is important to understand the geographic distribution of B. procyonis , as early and rapid treatment can prevent severe pathologies in humans. We present evidence for the roundworm spreading into a naive raccoon population through natural dispersal of infected raccoons. We sampled 181 raccoons from Saxony-Anhalt, a German federal state containing contact zones of different raccoon populations, two of which were previously free of the parasite. We screened the raccoons for roundworms and used microsatellite-based assignment tests to determine the genetic origin of the raccoons and their parasites. We detected roundworms in 16 of 45 raccoons sampled in a previously roundworm-free area in the northern part of the state. The largest proportion of the genetic ancestry (≥ 0.5) of the 16 raccoon hosts was assigned to the previously naive raccoon population. Conversely, the genetic ancestry of almost all the roundworms was assigned to the nearest roundworm population in the southern part of the state. Infected raccoons have, therefore, spread to the north of the state, where they interbred with and infected local raccoons. It seems likely that the roundworms will continue to spread. Health authorities should consider continuous surveillance programmes of naive populations and raise public awareness.

AimThe biodiversity of the Malay Archipelago is the product of the region's rich biogeographical history with periods of island connectivity and isolation during the Pleistocene glacial cycles. Here, the case of two endemic suid species, the Javan (Sus verrucosus) and Bawean (S. blouchi) warty pigs, was used to illustrate how biogeographic processes and recent anthropogenic pressures can shape demographic histories with significant implications for species conservation. LocationMalay Archipelago, with focus on Bawean and Java. MethodsWe employed genome-wide single nucleotide polymorphisms from the Porcine SNP60 v2 BeadChip to assess interspecific genetic differentiation, to estimate divergence times and to perform demographic model selection. ResultsIn contrast to the hypothesis of recent divergence during the last glacial maximum, S. blouchi was found to have diverged from S. verrucosus at least 166 k years ago following a founder event. The contemporary S. blouchi population was characterised by a recent bottleneck that reduced the effective population size to less than 20. The genomic assessment supports the single species status of S. blouchi, as was previously proposed based on morphometrics. The demographic history of S. verrucosus showed evidence of secondary contact with the sympatric banded pig (S. scrofa vittatus) that colonised Java 70 k years ago. Main ConclusionsWhile the Javan and Bawean warty pigs have persisted throughout the Pleistocene climatic oscillations, contemporary pressures from human activities threaten their survival and immediate action should be taken to grant legal protection to both S. verrucosus and S. blouchi. This study highlighted the use of demographic history modelling using genomic data to identify evolutionary significant units and inform conservation.

BACKGROUND:Important information on movement pathways and introduction routes of invasive parasites can be obtained by comparing the genetic makeup of an invader with its spatial genetic structure in other distribution areas. Sometimes, the population genetic structure of the host might be more informative than that of the parasite itself, and it is important to collect tissue samples of both host and parasite. However, host tissue samples are frequently not available for analysis. We aimed to test whether it is possible to generate reliable microsatellite profiles of host individuals by amplifying DNA extracted from a nematode parasite, using the raccoon (Procyon lotor) and the raccoon roundworm (Baylisascaris procyonis) as a test case.METHODS:Between 2020 and 2021, we collected tissue as well as a single roundworm each from 12 raccoons from central Germany. Both the raccoon and the roundworm DNA extracts were genotyped using 17 raccoon-specific microsatellite loci. For each roundworm DNA extract, we performed at least eight amplification reactions per microsatellite locus.RESULTS:We extracted amplifiable raccoon DNA from all 12 roundworms. We obtained at least two amplification products for 186 of the 204 possible genotypes. Altogether 1077 of the 1106 genotypes (97.4%) matched the host-DNA derived reference genotypes and thus did not contain genotyping errors. Nine of the 12 roundworm-derived genetic profiles matched the reference profiles from the raccoon hosts, with one additional genetic profile containing genotyping errors at a single locus. The remaining two genetic profiles were deemed unsuitable for downstream analysis because of genotyping errors and/or a high proportion of missing data.CONCLUSIONS:We showed that reliable microsatellite-based genetic profiles of host individuals can be obtained by amplifying DNA extracted from a parasitic nematode. Specifically, the approach can be applied to reconstruct invasion pathways of roundworms when samples of the raccoon hosts are lacking. Further research should assess whether this method can be replicated in smaller species of parasitic nematodes and other phyla of parasites more generally.

Landscape genetics provides powerful tools to quantify the effects of landscape features on population connectivity, but robust results are imperative to inform conservation planning. The robustness of landscape genetic inferences was assessed using the case of the northern crested newt (Triturus cristatus) in Luxembourg. Specifically, the effect of different study designs and genetic distance metrics was tested in terms of model convergence and misspecification rates (Type I error). The optimisation of resistance surfaces was performed in ResistanceGA, using individual- and population-based sampling designs and 16 genetic distance metrics inferred from 897 multilocus genotypes from 85 locations. Empirical results were complemented with simulations to assess Type I error rates and correlation between ‘true’ and optimised resistance surfaces. Individual-based optimisations seemed prone to overfitting, with little convergence among empirical resistance surfaces from different sets of individuals. Simulations showed significant differences in performance among population genetic distance metrics. Linear topographical features exhibited higher Type I error rates (83.3 F_ST , and PCA axes 1–45 were the top three genetic distance metrics for recovering true resistance features. Topographic roughness consistently drove spatial genetic clustering of T. cristatus, but variability existed among conductivity maps derived from optimised resistance surfaces. These findings underscore the importance of carefully selecting genetic distance metrics and addressing potential sources of uncertainty in resistance surface optimisation. By doing so, we can enhance the effectiveness of conservation planning efforts for T. cristatus and species with similar ecological considerations.

The major histocompatibility complex (MHC) has been intensively studied for the relative effects of different evolutionary forces in recent decades. Pathogen-mediated balancing selection is generally thought to explain the high polymorphism observed in MHC genes, but it is still unclear to what extent MHC diversity is shaped by selection relative to neutral drift. In this study, we genotyped MHC class II DRB genes and 15 neutral microsatellite loci across 26 geographic populations of European badgers (Meles meles) covering most of their geographic range. By comparing variation of microsatellite and diversity of MHC at different levels, we demonstrate that both balancing selection and drift have shaped the evolution of MHC genes. When only MHC allelic identity was investigated, the spatial pattern of MHC variation was similar to that of microsatellites. By contrast, when functional aspects of the MHC diversity (e.g., immunological supertypes) were considered, balancing selection appears to decrease genetic structuring across populations. Our comprehensive sampling and analytical approach enable us to conclude that the likely mechanisms of selection are heterozygote advantage and/or rare-allele advantage. This study is a clear demonstration of how both balancing selection and genetic drift simultaneously affect the evolution of MHC genes in a widely distributed wild mammal.

Abstract Genetic identification methods have become increasingly important for species that are difficult to identify in the field. A case in point is Pelophylax water frogs. While their morphological determination is highly complex, they include species protected under EU law and some that are classified as invasive. Additionally, genetic data can provide insights into their complex breeding systems, which may or may not involve the reproductive dependency of one species on another. Here, we generate baseline data for water frog monitoring in Luxembourg. We applied a countrywide sampling approach and used SNPs generated by ddRAD sequencing to identify individuals and infer the breeding systems present in the country. We found Pelophylax lessonae and P. kl. esculentus throughout Luxembourg, mostly living in syntopy. In general, a reproductive dependency of P. kl. esculentus on P. lessonae (L‐E system) was revealed. Besides this general system, we detected triploid P. kl. esculentus in six ponds. This indicates a modified L‐E system with reproductive dependency of the triploids on the diploid P. kl. esculentus. The invasive P. cf. bedriagae was detected in three ponds in southern Luxembourg, with evidence for hybridization with native water frogs. In addition to the ddRAD data, we tested a simple genetic method for future monitoring based on the MND1 marker. It showed in almost all cases, an identical species identification as the ddRAD data and was successfully applied to DNA extracts from mouth swabs. Combining this method with our baseline data will enable informed choices for the protection of native water frog species in Luxembourg.

The urinary bladder and lower urinary tract of domestic and wild carnivores can be parasitised by filamentous nematodes from the genus Pearsonema (syn. Capillaria). Infestations are often asymptomatic, but severe courses in dogs and cats have been described. Hosts are infested through the ingestion of earthworms (Lumbricidae) which act as intermediate hosts. Epidemiological studies of Pearsonema in raccoons (Procyon lotor) in North America are scarce and previous studies of urinary bladder parasites of European raccoons did not provide evidence of infestation. We examined urine sediment or rinse water from urinary bladders of 499 wild raccoons from Luxembourg, Poland and five study sites in Germany. Pearsonema eggs were found in the urine sediment of 31 (6.2%) raccoons. Infested animals were found in all study areas with prevalence values ranging from 3.7% to 8.7%. No significant difference in prevalence was found either between animals in urban and rural areas or between sexes and age classes. Based on their morphology, the eggs were likely to be P. plica. Considering their increasing density in Central Europe, raccoons may play a previously overlooked role in environmental contamination with Personema eggs.

Niphargus amphipods were collected from 2007 to 2018 at 98 sites comprising artificial caverns, springs and interstitial waters in the Grand Duchy of Luxembourg. Opportunistic sampling was combined with passive trapping. Specimen identification was achieved using morphological keys and molecular data. Initial morphological determination and literature data suggested five species, whereas sequencing of fragments of the mitochondrial cytochrome c oxidase subunit 1 gene and nuclear 28S rDNA marker supported the presence of seven species: Niphargus schellenbergi, Niphargus puteanus, Niphargus fontanus, one species of the Niphargus kochianus complex, and three species of the Niphargus aquilex complex. Niphargus schellenbergi was by far the most abundant and widespread species. Limited overlap was observed between literature-based records, our initial morphological determinations based on classical taxonomic characters, and genetic sequence data. In general, the combination of phenotypically variable taxa, such as N. schellenbergi, and cryptic or near-cryptic species, as in the N. aquilex complex, renders morphological identification of niphargids from Luxembourg a challenging or even impossible task. DNA taxonomy will therefore have to be used in future studies of the fauna of this region.