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The stone crayfish Austropotamobius torrentium is globally endangered due to anthropogenic pressure on its habitats, climate change and the invasive crayfish species, particularly the signal crayfish Pacifastacus leniusculus. Aiming to guide A. torrentium conservation within the hotspot of its genetic diversity in the western Balkan Peninsula, we combined population genetics and species distribution models (SDMs) to reveal the impact of climate change and the invasive P. leniusculus on A. torrentium diversity. Population genetic analyses based on newly developed microsatellites revealed moderate within-population genetic diversity and high differentiation among populations, reflecting isolated populations with limited gene flow. Alongside strong genetic structuring, we discovered high level of inbreeding coefficient indicating homozygote excess within the majority of populations. The SDMs results predicted substantial reductions of suitable habitats for A. torrentium by 2070; 80 % of its currently suitable habitat is predicted to be lost under high‐warming climate change scenario. Obtained results indicated that 44 % of populations with high and/or unique genetic diversity, including three highly divergent and geographically restricted evolutionary lineages, are located in the areas predicted to become unsuitable in the future, highlighting their vulnerability to extinction. Further, SDMs revealed considerable decrease of future habitat suitability for P. leniusculus, suggesting that climate change represents greater threat to A. torrentium. Our study highlights the importance of conserving remnant A. torrentium populations, among other, through assisted migration and population mixing that could help populations overcome the risks of inbreeding and maladaptation, and thus enabling A. torrentium to withstand the ongoing climate change.

Cherax destructor, the yabby, is an iconic Australian freshwater crayfish species, which, similar to other major invertebrate groups, is grossly under-represented in genomic databases. The yabby is also the principal commercial freshwater crustacean species in Australia subject to explotation via inland fisheries and aquaculture. To address the genomics knowledge gap for this species and explore cost effective and efficient methods for genome assembly, we generated 106.8 gb of Nanopore reads and performed a long-read only assembly of the Cherax destructor genome. On a mini-server configured with an ultra-fast swap space, the de novo assembly took 131 h (∼5.5 days). Genome polishing with 126.3 gb of PCR-Free Illumina reads generated an assembled genome size of 3.3 gb (74.6% BUSCO completeness) with a contig N50 of 80,900 bp, making it the most contiguous for freshwater crayfish genome assemblies. We found an unusually large number of cellulase genes within the yabby genome which is relevant to understanding the nutritional biology, commercial feed development, and ecological role of this species and crayfish more generally. These resources will be useful for genomic research on freshwater crayfish and our methods for rapid and super-efficient genome assembly will have wide application.

Biosynthesis of glycosylphosphatidylinositol (GPI) is essential for the integrity of the fungal cell wall. Here, the authors show that the natural product jawsamycin inhibits GPI biosynthesis by targeting a subunit of the fungal UDP-glycosyltransferase, and displays pronounced activity against pathogenic fungi of the order Mucorales.

Background The noble crayfish (Astacus astacus) is a native European species in decline, with a contracting range and diminishing populations and abundance. Previous studies revealed this species significant genetic diversity in the south-eastern Europe, with populations from the western and the southern part of the Balkan Peninsula being the most divergent. However, sampling of populations from the western part of the Balkans was limited and insufficient for investigating genetic diversity and population divergence for the purpose of conservation planning and management. Thus, the major aim of this study was to fill in this knowledge gap by studying mitochondrial and microsatellite DNA diversity, using 413 noble crayfish from 18 populations from waterbodies in the western part of the Balkan Peninsula. Methods Phylogenetic analysis of studied populations and their mitochondrial diversity were studied using COI and 16S sequences and population genetic structure was described using 15 microsatellite loci. Results Phylogeographic analysis revealed new divergent mitochondrial haplotypes for the populations in the westernmost part of the Balkan Peninsula in the tributaries of the Sava and Drava rivers. Microsatellite data indicated that these populations harbour an important component of genetic diversity within A. astacus. The results suggest that the western part of the Balkans played an important role as microrefugia during the Pleistocene climate fluctuations, allowing the long term persistence of A. astacus populations in this region. These results will also be important to supporting conservation decision making and planning.

Abstract Background The recently published complete mitogenome of the European lobster (Homarus gammarus) that was generated using long-range PCR exhibits unusual gene composition (missing nad2) and gene rearrangements among decapod crustaceans with strong implications in crustacean phylogenetics. Such atypical mitochondrial features will benefit greatly from validation with emerging long read sequencing technologies such as Oxford Nanopore that can more accurately identify structural variation. Results We re-sequenced the H. gammarus mitogenome on an Oxford Nanopore Minion flowcell and performed a long-read only assembly, generating a complete mitogenome assembly for H. gammarus. In contrast to previous reporting, we found an intact mitochondrial nad2 gene in the H. gammarus mitogenome and showed that its gene organization is broadly similar to that of the American lobster (H. americanus) except for the presence of a large tandemly duplicated region with evidence of pseudogenization in one of each duplicated protein-coding genes. Conclusions Using the European lobster as an example, we demonstrate the value of Oxford Nanopore long read technology in resolving problematic mitogenome assemblies. The increasing accessibility of Oxford Nanopore technology will make it an attractive and useful tool for evolutionary biologists to verify new and existing unusual mitochondrial gene rearrangements recovered using first and second generation sequencing technologies, particularly those used to make phylogenetic inferences of evolutionary scenarios.

Pontastacus leptodactylus is a native European crayfish species found in both freshwater and brackish environments. It has commercial importance for fisheries and aquaculture industries. Up till now, most studies concerning P. leptodactylus have focused onto gaining knowledge about its phylogeny and population genetics. However, little is known about the chromosomal evolution and genome organization of this species. Therefore, we performed clustering analysis of a low coverage genomic dataset to identify and characterize repetitive DNA in the P. leptodactylus genome. In addition, the karyogram of P. leptodactylus (2n = 180) is presented here for the first time consisting of 75 metacentric, 14 submetacentric, and a submetacentric/metacentric heteromorphic chromosome pair. We determined the genome size to be at ~18.7 gigabase pairs. Repetitive DNA represents about 54.85% of the genome. Satellite DNA repeats are the most abundant type of repetitive DNA, making up to ~28% of the total amount of repetitive elements, followed by the Ty3/Gypsy retroelements (~15%). Our study established a surprisingly high diversity of satellite repeats in P. leptodactylus. The genome of P. leptodactylus is by far the most satellite-rich genome discovered to date with 258 satellite families described. Of the five mapped satellite DNA families on chromosomes, PlSAT3-411 co-localizes with the AT-rich DAPI positive probable (peri)centromeric heterochromatin on all chromosomes, while PlSAT14-79 co-localizes with the AT-rich DAPI positive (peri)centromeric heterochromatin on one chromosome and is also located subterminally and intercalary on some chromosomes. PlSAT1-21 is located intercalary in the vicinity of the (peri)centromeric heterochromatin on some chromosomes, while PlSAT6-70 and PlSAT7-134 are located intercalary on some P. leptodactylus chromosomes. The FISH results reveal amplification of interstitial telomeric repeats (ITRs) in P. leptodactylus. The prevalence of repetitive elements, especially the satellite DNA repeats, may have provided a driving force for the evolution of the P. leptodactylus genome.

The white-clawed crayfish (Austropotamobius pallipes) is an endangered species in Europe with limited genomic information. Despite its conservation status there is no transcriptomic data available for A. pallipes in public databases. The data here represents the first transcriptome profile of the white-clawed crayfish generated using Illumina stranded RNA sequencing. Pair-end reads were assembled de novo with three separate transcriptome assemblers (Trinity, RNABloom, and RNASpades) followed by transcript assembly reduction and gene reconstruction using the EvidentialGene pipeline. The transcriptome was functionally annotated using InterProScan and genes coding for carbohydrate-active enzymes were identified through the dbCAN2 server. Raw fastq reads and the final version of the transcriptome assembly have been deposited in the NCBI-SRA (SRR10549898) and NCBI-TSA (GICG01) databases. Keywords: Austropotamobius pallipes, Transcriptome, Muscle, Illumina, Crayfish

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Abstract The pace of biological invasions has increased in recent decades, leading to multiple invasions and the potential dominance of invasive species, destabilizing local ecological networks. This provides opportunities to study new ecological species interactions, including predation. Tropical freshwaters have been particularly concerned by aquatic invasions and we focused here on the Martinique island (Lesser Antilles). We examined the predator–prey relationships involving invasive Thiarid snails (Tarebia granifera and Melanoides tuberculata) and the native Neritina punctulata, both confronted with a newcomer predator, the redclaw crayfish (Cherax quadricarinatus). We conducted several mesocosm experiments to assess the impact of crayfish predation on snail survival and the passive and active antipredator responses of snails. A first experiment indicated snail survival rates between 50% and 80%, depending on crayfish size and sex. Notably, there was a negative correlation between snail survival and male crayfish size and the predation method (shell crushing vs. “body sucking”) varied with crayfish size. The second experiment suggested no refuge size for snails, with both very small (5 mm) unable to escape predation, regardless of crayfish size (from 77 to 138 mm) or sex. Finally, we investigated the escape behavior of Thiarids regarding three crayfish cues. Melanoides tuberculata tend to bury in the substrate and T. granifera to climb up aquarium walls, what was expected from their shell morphologies, and both responding to crayfish cues within minutes. Overall, C. quadricarinatus proves to be an efficient snail predator with limited escape options for snails, potentially contributing to the decline of certain snail populations in Martinique. This omnivorous predator might impact other native species across different groups, including shrimps and fish. Our study underscores the urgent need for monitoring efforts, solidifying the redclaw crayfish reputation as a dangerous invasive species for freshwater macrobenthic faunas worldwide.

The land cover data presented here is a reconstruction of the past landscape (1993) at Very High Resolution (VHR) for the city of Poitiers, France. This reconstruction is based on multiple sources of images and data. We combined the strengths of both mono-temporal and multi-temporal classifications. Orthophotos were created at a spatial resolution of 0.5 m using aerial raw images from the French National Geographic Institute (IGN), taken during two aerial missions in July and August 1993. These orthophotos were merged at a spatial resolution of 5 m to conduct a first object-based classification using Landsat-5 TM images. The goal was to identify croplands, grasslands, coniferous and deciduous forests, urban areas, water bodies, and shadows. This learning-based classification employed a dataset consisting of 1371 polygons and demonstrated strong classification performances, achieving an overall accuracy of 86.31% and a kappa index of 0.832. On the other hand, mono-temporal classifications at a 0.5 m spatial resolution were carried out on each orthophoto to extract trees and herbaceous vegetation, especially in urban contexts. As mono-temporal classifications contained less information, we used a larger number of polygons for the learning step: 3849 and 5173 polygons for the northern and southern classifications, respectively. The segmentation step performed better in urban areas compared to rural areas. Consequently, the performance of classifications was evaluated separately for both contexts. Urban areas exhibited excellent performances, achieving kappa indices of 0.897 and 0.881 for the northern and southern classifications, respectively, whereas only tree vegetation was accurately detected in rural areas. To compensate for the lack of information such as buildings, railways, or roads, we modified the BD TopoⓇ dataset from IGN. This land cover map provides highly detailed information, facilitating the understanding of urban sprawl and changes in urban and rural vegetation surrounding the city of Poitiers. Due to these reasons, this freely accessible map can be utilized by researchers, land managers, and private companies for addressing urban and ecological challenges.

Abstract The white‐clawed crayfish (Austropotamobius pallipes) is an emblematic taxon of European rivers, found mainly in oxygenated streams, known to be an excellent indicator of river quality. Since several decades, the population of A. pallipes declined in relation to anthropogenic pressure, habitat loss, and competition with pests (invasive crayfish, crayfish plague). This endangered species is now submitted to conservation strategies by freshwater managers in order to survey and protect the remaining populations. In France, traditional surveys in freshwater environments were performed by electric fishing, kick‐net fishing, or trapping, particularly disruptive for the environment and very time‐consuming. However, with the rise of molecular genetic technology, new methods based on the detection of environmental DNA (eDNA) have emerged. We present here the results of an optimized study for the detection of the endangered crayfish Austropotamobius pallipes in France, considering certain environmental co‐factors and comparing two PCR methods (qPCR and ddPCR). After improving laboratory procedures, we were able to detect the presence of the crayfish up to 2 km downstream from a known point of presence and unfortunately highlight the disappearance of a historical population, after sampling two consecutive years. Such a level of precision is interesting because it makes it possible to precise the presence of specimens in a relatively restricted area and to orient traditional prospecting, necessary for certain additional studies. During our study, we observed better probabilities of detection during the summer period, but in a growing context of climate change, we advise to adapt the sampling year by year. That said, this methodology is a very useful tool for the detection of rare and/or endemic species and we did not observe any difference between the two PCR methods used.

Introduction: Surgery is the cornerstone of ovarian cancer treatment. However, surgery and perioperative inflammation have been described as potentially pro-metastagenic. In various animal models and other human cancers, intraoperative administration of non-steroidal anti-inflammatory drugs (NSAIDs) appears to have a positive impact on patient outcomes. Materials and methods: In this unicentric retrospective study, we provide an exploratory analysis of the safety and potential benefit of intraoperative administration of ketorolac on the outcome of patients undergoing surgery for ovarian cancer. The study population included all patients who were given a diagnosis of ovarian, fallopian tube or peritoneal cancer by the multidisciplinary oncology committee (MOC) of the Cliniques universitaires Saint-Luc between 2015 and 2020. Results: We included 166 patients in our analyses, with a median follow-up of 21.8 months. Both progression-free survival and overall survival were superior in patients who received an intraoperative injection of ketorolac (34.4 months of progression-free survival in the ketorolac group versus 21.5 months in the non-ketorolac group (p = 0.002), and median overall survival was not reached in either group but there was significantly higher survival in the ketorolac group (p = 0.004)). We also performed subgroup analyses to minimise bias due to imbalance between groups on factors that could influence patient survival, and the group of patients receiving ketorolac systematically showed a better outcome. Uni- and multivariate analyses confirmed that administration of ketorolac intraoperatively was associated with better progression-free survival (HR = 0.47 on univariate analysis and 0.43 on multivariate analysis, p = 0.003 and 0.023, respectively). In terms of complications, there were no differences between the two groups, either intraoperatively or postoperatively. Conclusion: Our study has shown a favourable association between the use of ketorolac during surgery and the postoperative progression of ovarian cancer in a group of 166 patients, without any rise in intra- or postoperative complications. These encouraging results point to the need for a prospective study to confirm the benefit of intraoperative administration of ketorolac in ovarian cancer surgery.

Abstract Introductions of invasive non‐native species (and their subsequent impacts) are recognized as a major threat to native flora and fauna. This is especially true in island ecosystems such as the tropical island of Martinique. In 2018, one such aquatic invasive species, the suckermouth catfish Hypostomus robinii (Loricariidae), was reported for the first time in two of the islands rivers. H. robinii is a popular freshwater aquarium fish and native to tropical and sub‐tropical South America. Since its initial discovery, a growing number of populations have been found, suggesting a larger distribution of this species through Martinique's hydrographic network. Here, we developed a novel survey technique (utilizing environmental DNA) and conducted a widespread survey across the island to assess the distribution of this invasive species. We were able to detect H. robinii in 22% of sites surveyed (18 out of 83) via our eDNA‐based assay. The presence of these fish was confirmed using traditional trapping at 14 of these sites. Additionally, we used occupancy modeling to investigate the impact of different environmental covariates on the detection efficiency of the novel assay and the potential impacts of false positives and negatives. We highlight a decrease in the detection probability when water volume filtered increases. That said, the eDNA‐based method proves a useful tool for the detection of this invasive fish species and monitoring its spread for management purposes.

The newly described horned iguana Iguana insularis from the southern Lesser Antilles is separated in two easily recognized subspecies: I. insularis sanctaluciae from St. Lucia and I. insularis insularis from the Grenadines. Its former description is completed by the use of 38 new samples for genetic and morphological analysis. Seventeen microsatellites were used to estimate genetic diversity, population structure and the level of introgression with other Iguana species over nearly the whole range of the species. ND4 and PAC sequences were also used to better characterize hybridization and to complete the description of this lineage. The I. insularis population of St. Vincent shows a high level of introgression from I. iguana whereas in the Grenadines, most islands present pure insularis populations but several show evidence of introgressions. Of the two remaining populations of I. insularis sanctaluciae, only one is still purebred. The recent identification of this and other distinct insular species and subspecies in the eastern Caribbean, and evaluation of where hybridization has occurred, are timely and important because the native iguanas are in urgent need of conservation action. Among the greatest threats is the ongoing human-mediated spread of invasive iguanas from Central and South America, which are destroying the endemic insular lineages through multiple diachronic introgression events.

Urban landscapes are rapid changing ecosystems with diverse urban forms that impede the movement of organisms. Therefore, designing and modelling ecological networks to identify biodiversity reservoirs and their corridors are crucial aspects of land management in terms of population persistence and survival. However, the land cover/use maps used for landscape connectivity modelling can lack information in such a highly complex environment. In this context, remote sensing approaches are gaining interest for the development of accurate land cover/use maps. We tested the efficiency of an object-based classification using open-source projects and free images to identify vegetation strata at a very fine scale and evaluated its contribution to landscape connectivity modelling. We compared different spatial and thematic resolutions from existing databases and object-based image analyses in three French cities. Our results suggested that this remote sensing approach produced reliable land cover maps to differentiate artificial areas, tree vegetation and herbaceous vegetation. Land cover maps enhanced with the remote sensing products substantially changed the structural connectivity indices, showing an improvement up to four times the proportion of herbaceous and tree vegetation. In addition, functional connectivity indices evaluated for several forest species were mainly impacted for medium dispersers in quantitative (metrics) and qualitative (corridors) estimations. Thus, the combination of this reproductible remote sensing approach and landscape connectivity modelling at a very fine scale provides new insights into the characterisation of ecological networks for conservation planning.

The Lesser Antilles, in the Eastern Caribbean, is inhabited by three Iguana species: the Lesser Antillean iguana Iguana delicatissima, which is endemic to the northernmost islands of the Lesser Antilles, the introduced common iguana from South America, Iguana iguana iguana, represented also by the two newly described endemic subspecies Iguana iguana sanctaluciae from Saint Lucia and Iguana iguana insularis from Saint Vincent and the Grenadines, and Grenada, and the introduced Iguana rhinolopha from Central America. Drawing on both morphological and genetic data, this paper describes the Iguana populations from Saba and Montserrat as a new species, Iguana melanoderma. This species is recognized on the basis of the following combination of characteristics: private microsatellite alleles, unique mitochondrial ND4 haplotypes, a distinctive black spot between the eye and tympanum, a dorsal carpet pattern on juveniles and young adults, a darkening of body coloration with aging (except for the anterior part of the snout), a black dewlap, pink on the jowl, the high number of large tubercular nape scales, fewer than ten medium sized–triangular dewlap spikes, high dorsal spikes, and lack of horns on the snout. This new melanistic taxon is threatened by unsustainable harvesting (including for the pet trade) and both competition and hybridization from escaped or released invasive alien iguanas (I. iguana iguana and I. rhinolopha) from South and Central America, respectively. The authors call for action to conserve Iguana melanoderma in Saba and Montserrat and for further research to investigate its relationship to other melanistic iguanas from the Virgin Islands and coastal islands of Venezuela.

The lesser Antilles archipelago in the Caribbean is known as a biodiversity hotspot, hosting many endemic species. However, recent introduction of a highly invasive species, the Australian redclaw crayfish (Cherax quadricarinatus), has led to significant threats to this fragile ecosystem. Here we developed, validated, and optimized a species-specific eDNA-based detection protocol targeting the 16S region of the mitochondrial gene of C. quadricarinatus. Our aim was to assess the crayfish distribution across Martinique Island. Our developed assay was species-specific and showed high sensitivity in laboratory, mesocosm and field conditions. A significant and positive correlation was found between species biomass, detection probability and efficiency through mesocosm experiments. Moreover, we found eDNA persisted up to 23 days in tropical freshwaters. We investigated a total of 83 locations, spread over 53 rivers and two closed water basins using our novel eDNA assay and traditional trapping, the latter, undertaken to confirm the reliability of the molecular-based detection method. Overall, we detected C. quadricarinatus at 47 locations using eDNA and 28 using traditional trapping, all positive trapping sites were positive for eDNA. We found that eDNA-based monitoring was less time-consuming and less influenced by the crayfishes often patchy distributions, proving a more reliable tool for future large-scale surveys. The clear threat and worrying distribution of this invasive species is particularly alarming as the archipelago belongs to one of the 25 identified biodiversity hotspots on Earth.

Abstract Background Oomycetes are a group of filamentous eukaryotic microorganisms that have colonized all terrestrial and oceanic ecosystems, and they include prominent plant pathogens. The Aphanomyces genus is unique in its ability to infect both plant and animal species, and as such exemplifies oomycete versatility in adapting to different hosts and environments. Dissecting the underpinnings of oomycete diversity provides insights into their specificity and pathogenic mechanisms. Results By carrying out genomic analyses of the plant pathogen A. euteiches and the crustacean pathogen A. astaci, we show that host specialization is correlated with specialized secretomes that are adapted to the deconstruction of the plant cell wall in A. euteiches and protein degradation in A. astaci. The A. euteiches genome is characterized by a large repertoire of small secreted protein (SSP)-encoding genes that are highly induced during plant infection, and are not detected in other oomycetes. Functional analysis revealed an SSP from A. euteiches containing a predicted nuclear-localization signal which shuttles to the plant nucleus and increases plant susceptibility to infection. Conclusion Collectively, our results show that Aphanomyces host adaptation is associated with evolution of specialized secretomes and identify SSPs as a new class of putative oomycete effectors.

Wolbachia are endosymbiotic bacteria that commonly infect arthropods, inducing certain phenotypes in their hosts. So far, no endemic South American species of terrestrial isopods have been investigated for Wolbachia infection. In this work, populations from two species of Balloniscus (B. sellowii and B. glaber) were studied through a diagnostic PCR assay. Fifteen new Wolbachia 16S rDNA sequences were detected. Wolbachia found in both species were generally specific to one population, and five populations hosted two different Wolbachia 16S rDNA sequences. Prevalence was higher in B. glaber than in B. sellowii, but uninfected populations could be found in both species. Wolbachia strains from B. sellowii had a higher genetic variation than those isolated from B. glaber. AMOVA analyses showed that most of the genetic variance was distributed among populations of each species rather than between species, and the phylogenetic analysis suggested that Wolbachia strains from Balloniscus cluster within Supergroup B, but do not form a single monophyletic clade, suggesting multiple infections for this group. Our results highlight the importance of studying Wolbachia prevalence and genetic diversity in Neotropical species and suggest that South American arthropods may harbor a great number of diverse strains, providing an interesting model to investigate the evolution of Wolbachia and its hosts.

The white-clawed crayfish Austropotamobius pallipes complex is native to Europe, being present in 18 European countries, Italy included. However, the number and abundance of its populations are today restricted and it has been recently classified as “endangered” by IUCN (International Union for Conservation of Nature). Here, we report the first record of this freshwater crayfish in Sardinia Island (Italy). Using a fragment of the mitochondrial DNA 16S rRNA gene, we identified three haplotypes that correspond to the A. italicus meridionalis subclade. We provide information about the sampling area, population density and finally discuss hypotheses about the occurrence of this population in Sardinia, comparing it with other Mediterranean populations. Our results improve the existing knowledge about the phylogeography of the taxon across Italy, confirming its complex pattern of distribution. In addition to the non-native status of the Sardinian A. i. meridionalis crayfish, we showed that the most proximal Mediterranean population of white-clawed crayfish existing in Corsica belongs to A. pallipes from Southern France.

The worldwide increase of hybridization in different groups is thought to have become more important with the loss of isolating barriers and the introduction of invasive species. This phenomenon could result in the extinction of endemic species. This study aims at investigating the hybridization dynamics between the endemic and threatened Lesser Antillean iguana (Iguana delicatissima) and the invasive common green iguana (Iguana iguana) in the Lesser Antilles, as well as assessing the impact of interspecific hybridization on the decline of I. delicatissima. 59 I. delicatissima (5 localities), 47 I. iguana (12 localities) and 27 hybrids (5 localities), who were all identified based on morphological characters, have been genotyped at 15 microsatellites markers. We also sequenced hybrids using ND4 mitochondrial loci to further investigate mitochondrial introgression. The genetic clustering of species and hybrid genetic assignment were performed using a comparative approach, through the implementation of a Discriminant Analysis of Principal Component (DAPC) based on statistics, as well as genetic clustering approaches based on the genetic models of several populations (Structure, NewHybrids and HIest), in order to get full characterization of hybridization patterns and introgression dynamics across the islands. The iguanas identified as hybrids in the wild, thanks to morphological analysis, were all genetically F1, F2, or backcrosses. A high proportion of individuals were also the result of a longer-term admixture. The absence of reproductive barriers between species leads to hybridization when species are in contact. Yet morphological and behavioral differences between species could explain why males I. iguana may dominate I. delicatissima, thus resulting in short-term species displacement and extinction by hybridization and recurrent introgression from I. iguana toward I. delicatissima. As a consequence, I. delicatissima gets eliminated through introgression, as observed in recent population history over several islands. These results have profound implications for species management of the endangered I. delicatissima and practical conservation recommendations are being discussed in the light of these findings.

Aphanomyces astaci, the crayfish plague pathogen, first appeared in Europe in the mid-19(th) century and is still responsible for mass mortalities of native European crayfish. The spread of this parasite across the continent is especially facilitated by invasive North American crayfish species that serve as its reservoir. In France, multiple cases of native crayfish mortalities have been suggested to be connected with the presence of the signal crayfish Pacifastacus leniusculus, which is highly abundant in the country. It shares similar habitats as the native white-clawed crayfish Austropotamobius pallipes and, when infected, the signal crayfish might therefore easily transmit the pathogen to the native species. We investigated the prevalence of A. astaci in French signal crayfish populations to evaluate the danger they represent to local populations of native crayfish. Over 500 individuals of Pacifastacus leniusculus from 45 French populations were analysed, plus several additional individuals of other non-indigenous crayfish species Orconectes limosus, O. immunis and Procambarus clarkii. Altogether, 20% of analysed signal crayfish tested positive for Aphanomyces astaci, and the pathogen was detected in more than half of the studied populations. Local prevalence varied significantly, ranging from 0% up to 80%, but wide confidence intervals suggest that the number of populations infected by A. astaci may be even higher than our results show. Analysis of several individuals of other introduced species revealed infections among two of these, O. immunis and P. clarkii. Our results confirm that the widespread signal crayfish serves as a key reservoir of Aphanomyces astaci in France and therefore represents a serious danger to native crayfish species, especially the white-clawed crayfish. The prevalence in other non-indigenous crayfish should also be investigated as they likely contribute to pathogen transmission in the country.

Maternally inherited Wolbachia (α-Proteobacteria) are widespread parasitic reproductive manipulators. A growing number of studies have described the presence of different Wolbachia strains within a same host. To date, no naturally occurring multiple infections have been recorded in terrestrial isopods. This is true for Armadillidium vulgare which is known to harbor non simultaneously three Wolbachia strains. Traditionally, such Wolbachia are detected by PCR amplification of the wsp gene and strains are characterized by sequencing. The presence of nucleotide deletions or insertions within the wsp gene, among these three different strains, provides the opportunity to test a novel genotyping method. Herein, we designed a new primer pair able to amplify products whose lengths are specific to each Wolbachia strain so as to detect the presence of multi-infections in A. vulgare. Experimental injections of Wolbachia strains in Wolbachia-free females were used to validate the methodology. We re-investigated, using this novel method, the infection status of 40 females sampled in 2003 and previously described as mono-infected based on the classical sequencing method. Among these females, 29 were identified as bi-infected. It is the first time that naturally occurring multiple infections of Wolbachia are detected within an individual A. vulgare host. Additionally, we resampled 6 of these populations in 2010 to check the infection status of females.

Armadillidium vulgare is a terrestrial isopod (Crustacea, Oniscidea) which harbors Wolbachia bacterial endosymbionts. A. vulgare is the major model for the study of Wolbachia-mediated feminization of genetic males in crustaceans. As a consequence of their impact on host sex determination mechanisms, Wolbachia endosymbionts are thought to significantly influence A. vulgare evolution on various grounds, including population genetic structure, diversity and reproduction strategies. To provide molecular tools for examining these questions, we isolated microsatellite loci through 454 pyrosequencing of a repeat-enriched A. vulgare genomic library. We selected 14 markers and developed three polymorphic microsatellite multiplex kits. We tested the kits on two A. vulgare natural populations and found high genetic variation, thereby making it possible to investigate the impact of Wolbachia endosymbionts on A. vulgare nuclear variation at unprecedented resolution. In addition, we tested the transferability of these kits by cross-species amplification in five other terrestrial isopod species harboring Wolbachia endosymbionts. The microsatellite loci showed good transferability in particular in Armadillidium nasatum and Chaetophiloscia elongata, for which these markers represent promising tools for future genetic studies.