Widespread infection, diversification and old host associations of Nosema Microsporidia in European freshwater gammarids (Amphipoda).

The microsporidian genus Nosema is primarily known to infect insects of economic importance stimulating high research interest, while other hosts remain understudied. Nosema granulosis is one of the formally described Nosema species infecting amphipod crustaceans, being known to infect only two host species. Our first aim was to characterize Nosema spp. infections in different amphipod species from various European localities using the small subunit ribosomal DNA (SSU) marker. Second, we aimed to assess the phylogenetic diversity, host specificity and to explore the evolutionary history that may explain the diversity of gammarid-infecting Nosema lineages by performing a phylogenetic reconstruction based on RNA polymerase II subunit B1 (RPB1) gene sequences. For the host species Gammarus balcanicus, we also analyzed whether parasites were in excess in females to test for sex ratio distortion in relation with Nosema infection. We identified Nosema spp. in 316 individuals from nine amphipod species being widespread in Europe. The RPB1-based phylogenetic reconstruction using newly reported sequences and available data from other invertebrates identified 39 haplogroups being associated with amphipods. These haplogroups clustered into five clades (A-E) that did not form a single amphipod-infecting monophyletic group. Closely related sister clades C and D correspond to Nosema granulosis. Clades A, B and E might represent unknown Nosema species infecting amphipods. Host specificity seemed to be variable with some clades being restricted to single hosts, and some that could be found in several host species. We show that Nosema parasite richness in gammarid hosts is much higher than expected, illustrating the advantage of the use of RPB1 marker over SSU. Finally, we found no hint of sex ratio distortion in Nosema clade A infecting G. balcanicus. This study shows that Nosema spp. are abundant, widespread and diverse in European gammarids. Thus, Nosema is as diverse in aquatic as in terrestrial hosts.

Histopathological screening of Pontogammarus robustoides (Amphipoda), an invader on route to the United Kingdom.

Biological invasions may act as conduits for pathogen introduction. To determine which invasive non-native species pose the biggest threat, we must first determine the symbionts (pathogens, parasites, commensals, mutualists) they carry, via pathological surveys that can be conducted in multiple ways (i.e., molecular, pathological, and histological). Whole animal histopathology allows for the observation of pathogenic agents (virus to Metazoa), based on their pathological effect upon host tissue. Where the technique cannot accurately predict pathogen taxonomy, it does highlight pathogen groups of importance. This study provides a histopathological survey of Pontogammarus robustoides (invasive amphipod in Europe) as a baseline for symbiont groups that may translocate to other areas/hosts in future invasions. Pontogammarus robustoides (n = 1,141) collected throughout Poland (seven sites), were noted to include a total of 13 symbiotic groups: a putative gut epithelia virus (overall prevalence = 0.6%), a putative hepatopancreatic cytoplasmic virus (1.4%), a hepatopancreatic bacilliform virus (15.7%), systemic bacteria (0.7%), fouling ciliates (62.0%), gut gregarines (39.5%), hepatopancreatic gregarines (0.4%), haplosporidians (0.4%), muscle infecting microsporidians (6.4%), digeneans (3.5%), external rotifers (3.0%), an endoparasitic arthropod (putatively: Isopoda) (0.1%), and Gregarines with putative microsporidian infections (1.4%). Parasite assemblages partially differed across collection sites. Co-infection patterns revealed strong positive and negative associations between five parasites. Microsporidians were common across sites and could easily spread to other areas following the invasion of P. robustoides. By providing this initial histopathological survey, we hope to provide a concise list of symbiont groups for risk-assessment in the case of a novel invasion by this highly invasive amphipod.

Histopathological survey for parasite groups in Gammarus varsoviensis (Amphipoda).

Invasive non-native amphipods (Crustacea) are becoming a model system in which to explore the impact and diversity of invasive parasites-parasites that are carried along an invasion route with their hosts. Gammarus varsoviensis is a freshwater amphipod species that has a recently explored invasion history. We provide a histopathological survey for a putatively invasive non-native population of this amphipod, identifying 8 symbiotic groups: Acanthocephala, Rotifera, Digenea, ciliated protozoa, Haplosporidia, Microsporidia, 'Candidatus Aquirickettsiella', and a putative nudivirus, at various prevalence. Our survey indicates that the parasites have no sex bias and that each has the potential to be carried in either sex along an invasion route. We discuss the pathology and prevalence of the above symbiotic groups and whether those that are parasitic may pose a risk if G. varsoviensis were to carry them to novel locations.

Dictyocoela microsporidia diversity and co-diversification with their host, a gammarid species complex (Crustacea, Amphipoda) with an old history of divergence and high endemic diversity.

BACKGROUND: Although the processes of co-evolution between parasites and their hosts are well known, evidence of co-speciation remains scarce. Microsporidian intracellular parasites, due to intimate relationships with their hosts and mixed mode of transmission (horizontal but also vertical, from mother to offspring), may represent an interesting biological model for investigating co-speciation. Amphipod crustaceans, especially gammarids, are regular hosts of microsporidian parasites, in particular the Dictyocoela spp., which have so far been found limited to these amphipods and are known to use a vertical mode of transmission. The amphipod genus Gammarus has a diversification history spanning the last 50-60 Mya and an extensive cryptic diversity in most of the nominal species. Here, we investigated the degree of co-diversification between Dictyocoela and Gammarus balcanicus, an amphipod with high degrees of ancient cryptic diversification and lineage endemism, by examining the genetic diversity of these parasites over the entire geographic range of the host. We hypothesised that the strong host diversification and vertical transmission of Dictyocoela would promote co-diversification. RESULTS: Using the parasite SSU rDNA as a molecular marker, analyzing 2225 host specimens from 88 sites covering whole host range, we found 31 haplogroups of Dictyocoela, 30 of which were novel, belonging to four Dictyocoela species already known to infect other Gammarus spp. The relationships between Dictyocoela and gammarids is therefore ancient, with the speciation in parasites preceding those of the hosts. Each novel haplogroup was nevertheless specific to G. balcanicus, leaving the possibility for subsequent co-diversification process during host diversification. A Procrustean Approach to Co-phylogeny (PACo) analysis revealed that diversification of Dictyocoela was not random with respect to that of the host. We found high degrees of congruence between the diversification of G. balcanicus and that of Dictyocoela roeselum and D. muelleri. However, we also found some incongruences between host and Dictyocoela phylogenies, e.g. in D. duebenum, probably due to host shifts between different G. balcanicus cryptic lineages. CONCLUSION: The evolutionary history of Dictyocoela and Gammarus balcanicus represents an example of an overall host-parasite co-diversification, including cases of host shifts.

'Candidatus Aquirickettsiella gammari' (Gammaproteobacteria: Legionellales: Coxiellaceae): A bacterial pathogen of the freshwater crustacean Gammarus fossarum (Malacostraca: Amphipoda).

Invasive and non-native species can pose risks to vulnerable ecosystems by co-introducing bacterial pathogens. Alternatively, co-introduced bacterial pathogens may regulate invasive population size and invasive traits. We describe a novel candidate genus and species of bacteria ('Candidatus Aquirickettsiella gammari') found to infect Gammarus fossarum, from its native range in Poland. The bacterium develops intracellularly within the haemocytes and cells of the musculature, hepatopancreas, connective tissues, nervous system and gonad of the host. The developmental cycle of 'Candidatus Aquirickettsiella gammari' includes an elementary body (496.73 nm ±â€¯37.56 nm in length, and 176.89 nm ±â€¯36.29 nm in width), an elliptical, condensed spherical stage (737.61 nm ±â€¯44.51 nm in length and 300.07 nm ±â€¯44.02 nm in width), a divisional stage, and a spherical initial body (1397.59 nm ±â€¯21.26 nm in diameter). We provide a partial genome for 'Candidatus Aquirickettsiella gammari', which clades phylogenetically alongside environmental 16S rRNA sequences from aquatic habitats, and bacterial symbionts from aquatic isopods (Asellus aquaticus), grouping separately from the Rickettsiella, a genus that includes bacterial pathogens of terrestrial insects and isopods. Increased understanding of the diversity of symbionts carried by G. fossarum identifies those that might regulate host population size, or those that could pose a risk to native species in the invasive range. Identification of 'Candidatus Aquirickettsiella gammari' and its potential for adaptation as a biological control agent is explored.

Isolation and characterization of 8 microsatellite loci for the "killer shrimp'', an invasive Ponto-Caspian amphipod Dikerogammarus villosus (Crustacea: Amphipoda).

Dikerogammarus villosus is a freshwater amphipod of the Ponto-Caspian origin recognized as one of the 100 worst alien species in Europe, having negative impact on biodiversity and functioning of the invaded aquatic ecosystems. The species has a wide ecophysiological tolerance and during the last 20 years it has rapidly spread throughout European inland waters. In consequence, it presents a major conservation management problem. We describe eight polymorphic microsatellite loci developed for D. villosus by combining a biotin-enrichment protocol and new generation 454GS-FLX Titanium pyrosequencing technology. When genotyped in 64 individuals from two locations, the loci exhibited a mean diversity of 4.87 alleles per locus (2-13). The mean observed and expected heterozygosities were, respectively, 0.439 (0.091-0.844) and 0.468 (0.089-0.843). Gametic disequilibrium was not detected for any pair of loci. The microsatellite markers will be a valuable tool in assessing the demographic processes associated with invasion of the killer shrimp from a genetic point of view.

Taming the terminological tempest in invasion science.

Standardised terminology in science is important for clarity of interpretation and communication. In invasion science - a dynamic and rapidly evolving discipline - the proliferation of technical terminology has lacked a standardised framework for its development. The result is a convoluted and inconsistent usage of terminology, with various discrepancies in descriptions of damage and interventions. A standardised framework is therefore needed for a clear, universally applicable, and consistent terminology to promote more effective communication across researchers, stakeholders, and policymakers. Inconsistencies in terminology stem from the exponential increase in scientific publications on the patterns and processes of biological invasions authored by experts from various disciplines and countries since the 1990s, as well as publications by legislators and policymakers focusing on practical applications, regulations, and management of resources. Aligning and standardising terminology across stakeholders remains a challenge in invasion science. Here, we review and evaluate the multiple terms used in invasion science (e.g. 'non-native', 'alien', 'invasive' or 'invader', 'exotic', 'non-indigenous', 'naturalised', 'pest') to propose a more simplified and standardised terminology. The streamlined framework we propose and translate into 28 other languages is based on the terms (i) 'non-native', denoting species transported beyond their natural biogeographic range, (ii) 'established non-native', i.e. those non-native species that have established self-sustaining populations in their new location(s) in the wild, and (iii) 'invasive non-native' - populations of established non-native species that have recently spread or are spreading rapidly in their invaded range actively or passively with or without human mediation. We also highlight the importance of conceptualising 'spread' for classifying invasiveness and 'impact' for management. Finally, we propose a protocol for classifying populations based on (i) dispersal mechanism, (ii) species origin, (iii) population status, and (iv) impact. Collectively and without introducing new terminology, the framework that we present aims to facilitate effective communication and collaboration in invasion science and management of non-native species.

Wide geographic distribution of overlooked parasites: Rare Microsporidia in Gammarus balcanicus, a species complex with a high rate of endemism.

Parasites and other symbionts deeply influence host organisms, and no living organism can be considered to have evolved independent of its symbionts. The first step towards understanding symbiotic influences upon host organisms is a strong supporting knowledge of parasite/symbiont diversity. Parasites of freshwater amphipods are diverse, with Microsporidia being a major group. These intracellular parasites impact gammarid fitness in different ways, ranging from reduced fitness to increased fecundity. Many Microsporidia have been recorded using molecular data, with multiple taxa pending formal taxonomic description. While some parasites are common, others are known only through sporadic records of single infections. In this study, we focus on rare/sporadic microsporidian infections within Gammarus balcanicus, a host species complex with a high level of endemism. In addition to enriching our knowledge on Microsporidia parasite diversity in amphipod hosts, we test whether these symbionts are specific to G. balcanicus or if they are the same taxa infecting other gammarid species. Of 2231 hosts from 87 sites, we catalogued 29 sequences of "rare" Microsporidia clustering into 19 haplogroups. These haplogroups cluster into 11 lineages: four pre-described taxa (Cucumispora roeselum, C. ornata, C. dikerogammari and Enterocytospora artemiae) and seven 'Molecular Operational Taxonomic Units', which are known from previously published studies to infect other European amphipod species. Our study significantly widens the geographic range of these Microsporidia and expands the known spectrum of hosts infected. Our results suggest that these parasites are ancient infections of European gammarids. For some host-parasite systems, we hypothesize that the common parasite ancestors that infected the hosts' common ancestors, diversified alongside host diversification. For others, we observe Microsporidia taxa with wide host ranges that do not follow host phylogeny.

Successful post-glacial colonization of Europe by single lineage of freshwater amphipod from its Pannonian Plio-Pleistocene diversification hotspot.

Gammarus roeselii Gervais, 1835 is a morphospecies with a wide distribution range in Europe. The Balkan Peninsula is known as an area of pre-Pleistocene cryptic diversification within this taxon, resulting in at least 13 Molecular Operational Taxonomic Units (MOTUs). The morphospecies diversified there during Neogene and has probably invaded other parts of the continent very recently, in postglacial or even historical times. Thus, the detailed goals of our study were to (1) identify which lineage(s) colonized Central-Western Europe (CWE), (2) determine their possible geographical origin, (3) verify, whether the colonisation was associated with demographic changes. In total, 663 individuals were sequenced for the cytochrome oxidase I (COI) barcoding fragment and 137 individuals for the internal transcribed spacer II (ITS2). We identified two MOTUs in the study area with contrasting Barcode Index Number and haplotype diversities. The Pannonian Basin (PB) appeared to be a potential ice age refugium for the species, while CWE was colonised by a single lineage (also present in PB), displaying low genetic diversity. Our results suggest that G. roeselii is a relatively recent coloniser in CWE, starting demographic expansion around 10 kya.

Microsporidian infections in the species complex Gammarus roeselii (Amphipoda) over its geographical range: evidence for both host-parasite co-diversification and recent host shifts.

BACKGROUND: Microsporidians are obligate endoparasites infecting taxonomically diverse hosts. Both vertical (from mother to eggs) and horizontal (between conspecifics or between species) transmission routes are known. While the former may promote co-speciation and host-specificity, the latter may promote shifts between host species. Among aquatic arthropods, freshwater amphipod crustaceans are hosts for many microsporidian species. However, despite numerous studies, no general pattern emerged about host specificity and co-diversification. In south-eastern Europe, the gammarid Gammarus roeselii is composed of 13 cryptic lineages of Miocene to Pleistocene age but few genotypes of one lineage have spread postglacially throughout north-western Europe. Based on nearly 100 sampling sites covering its entire range, we aim to: (i) explore the microsporidian diversity present in G. roeselii and their phylogenetic relationships, especially in relation to the parasites infecting other Gammaridae; (ii) test if the host phylogeographical history might have impacted host-parasite association (e.g. co-diversifications or recent host shifts from local fauna). METHODS: We used part of the small subunit rRNA gene as source of sequences to identify and determine the phylogenetic position of the microsporidian taxa infecting G. roeselii. RESULTS: Microsporidian diversity was high in G. roeselii with 24 detected haplogroups, clustered into 18 species-level taxa. Ten microsporidian species were rare, infecting a few individual hosts in a few populations, and were mostly phylogenetically related to parasites from other amphipods or various crustaceans. Other microsporidians were represented by widespread genera with high prevalence: Nosema, Cucumispora and Dictyocoela. Two contrasting host association patterns could be observed. First, two vertically transmitted microsporidian species, Nosema granulosis and Dictyocoela roeselum, share the pattern of infecting G. roeselii over most of its range and are specific to this host suggesting the co-diversification scenario. This pattern contrasted with that of Dictyocoela muelleri, the three species of Cucumispora, and the rare parasites, present only in the recently colonised region by the host. These patterns suggest recent acquisitions from local host species, after the recent spread of G. roeselii. CONCLUSIONS: Microsporidians infecting G. roeselii revealed two scenarios of host-parasite associations: (i) ancient associations with vertically transmitted parasites that probably co-diversified with their hosts, and (ii) host shifts from local host species, after the postglacial spread of G. roeselii.

Some like it hot: factors impacting thermal preferences of two Ponto-Caspian amphipods Dikerogammarus villosus (Sovinsky, 1894) and Dikerogammarus haemobaphes (Eichwald, 1841).

Temperature is a crucial factor determining biology and ecology of poikilothermic animals. It often constitutes an important barrier for invasive species originating from different climate zones but, on the other hand, may facilitate the invasion process of animals with wide thermal preferences and high resistance to extreme temperatures. In our experimental study, we investigated the thermal behaviour of two Ponto-Caspian amphipod crustaceans-Dikerogammarus villosus and Dikerogammarus haemobaphes. Both species are known to live under a wide range of thermal conditions which may promote their invasion. Moreover, both these amphipods are hosts for microsporidian parasites which co-evolved with them within the Ponto-Caspian region and spread in European waters. As the presence of a parasite may influence the thermal preferences of its host, we expected to observe behavioural changes in infected individuals of the studied amphipods leading to (1) behavioural fever (selecting a warmer habitat) or (2) anapyrexia (selecting a colder habitat). The experiment (N = 20) was carried out for 30 min in a 100 cm. 20 cm from boths sides were not avaliable for amphipods long thermal gradient (0-40 °C), using 30 randomly selected adult amphipod individuals of one species. At the end of each trial, we checked the position of amphipods along the gradient and determined their sex and infection status (uninfected or infected by one of microsporidium species). D. villosus was infected with Cucumispora dikerogammari whereas D. haemobaphes was a host for C. dikerogammari, Dictyocoela muelleri or D. berillonum. Thermal preferences of amphipods depended on their species and sex. Females of D. villosus preferred warmer microhabitats (often much above 30 °C) than conspecific males and females of D. haemobaphes, whereas no significant differences were found among males of both species and both sexes of D. haemobaphes. Moreover, infected males of D. villosus stayed in warmer water more often than uninfected males of this species, selecting temperatures higher than 30 °C, which may be explained either as a behavioural fever constituting a defence mechanism of a host against the infection, or as a parasite manipulation of the host behaviour increasing the parasite fitness. On the other hand, none of the parasite species affected the thermal preferences of D. haemobaphes, including also C. dikerogammari, changing the behaviour of D. villosus. Our research presents the complexity of the thermal behaviour of studied amphipods and the evidence that microsporidia may trigger a change in temperature preferendum of their host species and those observations may be the result of different host-parasite coevolution time which may vary for the two host species (Poulin, 2010).

Europe-wide reassessment of Dictyocoela (Microsporidia) infecting native and invasive amphipods (Crustacea): molecular versus ultrastructural traits.

Microsporidia are common parasites infecting animals and protists. They are specifically common pathogens of amphipods (Crustacea, Malacostraca), with Dictyocoela spp. being particularly frequent and highly prevalent, exhibiting a range of phenotypic and ecological effects. Until now, seven species of Dictyocoela were defined, predominantly based on the genetic distance. However, neither the taxonomic status of this provisionally erected genus (based on eight novel sequences and one micrograph of the spore), nor its internal phylogenetic relationships have been clearly revealed. The formal description of the genus and of most of the putative species are still lacking. Here we aimed to fill this gap and performed both ultrastructural and molecular studies (based on SSU, ITS and partial LSU) using different species delimitation methods. As a consensus of these results and following conservative data interpretation, we propose to distinguish five species infecting gammarid hosts, and to keep the names introduced by the authors of the type sequences: Dictyocoela duebenum, D. muelleri, D. berillonum and D. roeselum. We provide full descriptions of these species. Moreover, thanks to our extensive sampling, we extend the known host and geographic range of these Microsporidia.

Neogene paleogeography provides context for understanding the origin and spatial distribution of cryptic diversity in a widespread Balkan freshwater amphipod.

BACKGROUND: The Balkans are a major worldwide biodiversity and endemism hotspot. Among the freshwater biota, amphipods are known for their high cryptic diversity. However, little is known about the temporal and paleogeographic aspects of their evolutionary history. We used paleogeography as a framework for understanding the onset of diversification in Gammarus roeselii: (1) we hypothesised that, given the high number of isolated waterbodies in the Balkans, the species is characterised by high level of cryptic diversity, even on a local scale; (2) the long geological history of the region might promote pre-Pleistocene divergence between lineages; (3) given that G. roeselii thrives both in lakes and rivers, its evolutionary history could be linked to the Balkan Neogene paleolake system; (4) we inspected whether the Pleistocene decline of hydrological networks could have any impact on the diversification of G. roeselii. MATERIAL AND METHODS: DNA was extracted from 177 individuals collected from 26 sites all over Balkans. All individuals were amplified for ca. 650 bp long fragment of the mtDNA cytochrome oxidase subunit I (COI). After defining molecular operational taxonomic units (MOTU) based on COI, 50 individuals were amplified for ca. 900 bp long fragment of the nuclear 28S rDNA. Molecular diversity, divergence, differentiation and historical demography based on COI sequences were estimated for each MOTU. The relative frequency, geographic distribution and molecular divergence between COI haplotypes were presented as a median-joining network. COI was used also to reconstruct time-calibrated phylogeny with Bayesian inference. Probabilities of ancestors' occurrence in riverine or lacustrine habitats, as well their possible geographic locations, were estimated with the Bayesian method. A Neighbour Joining tree was constructed to illustrate the phylogenetic relationships between 28S rDNA haplotypes. RESULTS: We revealed that G. roeselii includes at least 13 cryptic species or molecular operational taxonomic units (MOTUs), mostly of Miocene origin. A substantial Pleistocene diversification within-MOTUs was observed in several cases. We evidenced secondary contacts between very divergent MOTUs and introgression of nDNA. The Miocene ancestors could live in either lacustrine or riverine habitats yet their presumed geographic localisations overlapped with those of the Neogene lakes. Several extant riverine populations had Pleistocene lacustrine ancestors. DISCUSSION: Neogene divergence of lineages resulting in substantial cryptic diversity may be a common phenomenon in extant freshwater benthic crustaceans occupying areas that were not glaciated during the Pleistocene. Evolution of G. roeselii could be associated with gradual deterioration of the paleolakes. The within-MOTU diversification might be driven by fragmentation of river systems during the Pleistocene. Extant ancient lakes could serve as local microrefugia during that time.

Parasites, pathogens and commensals in the "low-impact" non-native amphipod host Gammarus roeselii.

BACKGROUND: Whilst vastly understudied, pathogens of non-native species (NNS) are increasingly recognised as important threats to native wildlife. This study builds upon recent recommendations for improved screening for pathogens in NNS by focusing on populations of Gammarus roeselii in Chojna, north-western Poland. At this location, and in other parts of continental Europe, G. roeselii is considered a well-established and relatively 'low-impact' invader, with little understanding about its underlying pathogen profile and even less on potential spill-over of these pathogens to native species. RESULTS: Using a combination of histological, ultrastructural and phylogenetic approaches, we define a pathogen profile for non-native populations of G. roeselii in Poland. This profile comprised acanthocephalans (Polymorphus minutus Goese, 1782 and Pomphorhynchus sp.), digenean trematodes, commensal rotifers, commensal and parasitic ciliated protists, gregarines, microsporidia, a putative rickettsia-like organism, filamentous bacteria and two viral pathogens, the majority of which are previously unknown to science. To demonstrate potential for such pathogenic risks to be characterised from a taxonomic perspective, one of the pathogens, a novel microsporidian, is described based upon its pathology, developmental cycle and SSU rRNA gene phylogeny. The novel microsporidian Cucumispora roeselii n. sp. displayed closest morphological and phylogenetic similarity to two previously described taxa, Cucumispora dikerogammari (Ovcharenko & Kurandina, 1987), and Cucumispora ornata Bojko, Dunn, Stebbing, Ross, Kerr & Stentiford, 2015. CONCLUSIONS: In addition to our discovery extending the host range for the genus Cucumispora Ovcharenko, Bacela, Wilkinson, Ironside, Rigaud & Wattier, 2010 outside of the amphipod host genus Dikerogammarus Stebbing, we reveal significant potential for the co-transfer of (previously unknown) pathogens alongside this host when invading novel locations. This study highlights the importance of pre-invasion screening of low-impact NNS and, provides a means to document and potentially mitigate the additional risks posed by previously unknown pathogens.

Zebra mussel beds: an effective feeding ground for Ponto-Caspian gobies or suitable shelter for their prey?

Aggregations of the Ponto-Caspian invasive zebra mussel (Dreissena polymorpha) constitute a suitable habitat for macroinvertebrates, considerably increasing their abundance and providing effective antipredator protection. Thus, the overall effect of a mussel bed on particular predator species may vary from positive to negative, depending on both prey density increase and predator ability to prey in a structurally complex habitat. Alien Ponto-Caspian goby fish are likely to be facilitated when introduced into new areas by zebra mussels, provided that they are capable of utilizing mussel beds as habitat and feeding grounds. We ran laboratory experiments to find which prey (chironomid larvae) densities (from ca. 500 to 2,000 individuals m-2) in a mussel bed make it a more beneficial feeding ground for the racer goby Babka gymnotrachelus (RG) and western tubenose goby Proterorhinus semilunaris (WTG) compared to sandy and stone substrata (containing the basic prey density of 500 ind. m-2). Moreover, we checked how food availability affects habitat selection by fish. Mussel beds became more suitable for fish than alternative mineral substrata when food abundance was at least two times higher (1,000 vs. 500 ind. m-2), regardless of fish size and species. WTG was associated with mussel beds regardless of its size and prey density, whereas RG switched to this habitat when it became a better feeding ground than alternative substrata. Larger RG exhibited a stronger affinity for mussels than small individuals. WTG fed more efficiently from a mussel bed at high food abundances than RG. A literature review has shown that increasing chironomid density, which in our study was sufficient to make a mussel habitat an attractive feeding ground for the gobies, is commonly observed in mussel beds in the field. Therefore, we conclude that zebra mussels may positively affect the alien goby species and are likely to facilitate their establishment in novel areas, contributing to an invasional meltdown in the Ponto-Caspian invasive community.

Out of the Black Sea: phylogeography of the invasive killer shrimp Dikerogammarus villosus across Europe.

The amphipod Dikerogammarus villosus has colonized most of the European main inland water bodies in less than 20 years, having deteriorating effect on the local benthic communities. Our aim was to reveal the species phylogeography in the native Black Sea area, to define the source populations for the colonization routes in continental Europe and for the newly established UK populations. We tested for the loss of genetic diversity between source and invasive populations as well as along invasion route. We tested also for isolation by distance. Thirty three native and invasive populations were genotyped for mtDNA (COI, 16S) and seven polymorphic nuclear microsatellites to assess cryptic diversity (presence of deeply divergent lineages), historical demography, level of diversity within lineage (e.g., number of alleles), and population structure. A wide range of methods was used, including minimum spanning network, molecular clock, Bayesian clustering and Mantel test. Our results identified that sea level and salinity changes during Pleistocene impacted the species phylogeography in the Black Sea native region with four differentiated populations inhabiting, respectively, the Dnieper, Dniester, Danube deltas and Durungol liman. The invasion of continental Europe is associated with two sources, i.e., the Danube and Dnieper deltas, which gave origin to two independent invasion routes (Western and Eastern) for which no loss of diversity and no isolation by distance were observed. The UK population has originated in the Western Route and, despite very recent colonization, no drastic loss of diversity was observed. The results show that the invasion of the killer shrimp is not associated with the costs of loosing genetic diversity, which may contribute to the success of this invader in the newly colonized areas. Additionally, while it has not yet occurred, it might be expected that future interbreeding between the genetically diversified populations from two independent invasion routes will potentially even enhance this success.