Trophic Positions of Polyp and Medusa Stages of the Freshwater Jellyfish Craspedacusta sowerbii Based on Stable Isotope Analysis.

When species spread into new regions, competition with native species and predatory-prey relationships play a major role in whether the new species can successfully establish itself in the recipient food web and become invasive. In aquatic habitats, species with a metagenetic life cycle, such as the freshwater jellyfish Craspedacusta with benthic polyps and planktonic medusae, have to meet the requirements of two distinct life stages occurring in two habitats with different food webs. Here, we examined the trophic position of both life stages, known to be predatory, and compared their niches with those of putative native competitors using stable isotope analysis. We found that δ13C and δ15N signatures of medusae overlapped with those of co-occurring Chaoborus larvae and juvenile fish (Rutilus rutilus) in a well-studied lake, implying high competition with these native predators. The comparison of δ15N signatures of Hydra and Craspedacusta polyps in four additional lakes revealed their similar trophic position, matching their predatory lifestyle. However, their δ13C signatures differed not only across all four of the lakes studied but also within one lake over time, suggesting a preference for pelagic or benthic food sources. We conclude that invasive and native polyps differ in their niches due to different food spectra, which favors the invasion success of Craspedacusta.

Cytonuclear diversity and shared mitochondrial haplotypes among Daphnia galeata populations separated by seven thousand kilometres.

BACKGROUND: The zooplanktonic cladocerans Daphnia, present in a wide range of water bodies, are an important component of freshwater ecosystems. In contrast to their high dispersal capacity through diapausing eggs carried by waterfowl, Daphnia often exhibit strong population genetic differentiation. Here, to test for common patterns in the population genetic structure of a widespread Holarctic species, D. galeata, we genotyped two sets of populations collected from geographically distant areas: across 13 lakes in Eastern China and 14 lakes in Central Europe. The majority of these populations were genotyped at two types of markers: a mitochondrial gene (for 12S rRNA) and 15 nuclear microsatellite loci. RESULTS: Mitochondrial DNA demonstrated relatively shallow divergence within D. galeata, with distinct haplotype compositions in the two study regions but one widely distributed haplotype shared between several of the Chinese as well as European populations. At microsatellite markers, clear separation was observed at both large (between China and Europe) and small (within Europe) geographical scales, as demonstrated by Factorial Correspondence Analyses, Bayesian assignment and a clustering method based on genetic distances. Genetic diversity was comparable between the sets of Chinese and European D. galeata populations for both types of markers. Interestingly, we observed a significant association between genetic distance and geographical distance for D. galeata populations in China but not in Europe. CONCLUSIONS: Our results indicate relatively recent spread of D. galeata across wide expanses of the Palaearctic, with one mtDNA lineage of D. galeata successfully establishing over large distances. Despite a clear differentiation of Chinese and European D. galeata at a nuclear level, the pattern of genetic variation is nevertheless similar between both regions. Overall, our findings provide insights into the genetic population structure of a cladoceran species with extremely wide geographical range.

Daphnia parasite dynamics across multiple Caullerya epidemics indicate selection against common parasite genotypes.

Studies of parasite population dynamics in natural systems are crucial for our understanding of host-parasite coevolutionary processes. Some field studies have reported that host genotype frequencies in natural populations change over time according to parasite-driven negative frequency-dependent selection. However, the temporal patterns of parasite genotypes have rarely been investigated. Moreover, parasite-driven negative frequency-dependent selection is contingent on the existence of genetic specificity between hosts and parasites. In the present study, the population dynamics and host-genotype specificity of the ichthyosporean Caullerya mesnili, a common endoparasite of Daphnia water fleas, were analysed based on the observed sequence variation in the first internal transcribed spacer (ITS1) of the ribosomal DNA. The Daphnia population of lake Greifensee (Switzerland) was sampled and subjected to parasite screening and host genotyping during C. mesnili epidemics of four consecutive years. The ITS1 of wild-caught C. mesnili-infected Daphnia was sequenced using the 454 pyrosequencing platform. The relative frequencies of C. mesnili ITS1 sequences differed significantly among years: the most abundant C. mesnili ITS1 sequence decreased and rare sequences increased over the course of the study, a pattern consistent with negative frequency-dependent selection. However, only a weak signal of host-genotype specificity between C. mesnili and Daphnia genotypes was detected. Use of cutting edge genomic techniques will allow further investigation of the underlying micro-evolutionary relationships within the Daphnia-C. mesnili system.

Extreme Environments Facilitate Hybrid Superiority - The Story of a Successful Daphnia galeata × longispina Hybrid Clone.

Hybridization within the animal kingdom has long been underestimated. Hybrids have often been considered less fit than their parental species. In the present study, we observed that the Daphnia community of a small lake was dominated by a single D. galeata × D. longispina hybrid clone, during two consecutive years. Notably, in artificial community set-ups consisting of several clones representing parental species and other hybrids, this hybrid clone took over within about ten generations. Neither the fitness assay conducted under different temperatures, or under crowded and non-crowded environments, nor the carrying capacity test revealed any outstanding life history parameters of this hybrid clone. However, under simulated winter conditions (i.e. low temperature, food and light), the hybrid clone eventually showed a higher survival probability and higher fecundity compared to parental species. Hybrid superiority in cold-adapted traits leading to an advantage of overwintering as parthenogenetic lineages might consequently explain the establishment of successful hybrids in natural communities of the D. longispina complex. In extreme cases, like the one reported here, a superior hybrid genotype might be the only clone alive after cold winters. Overall, superiority traits, such as enhanced overwintering here, might explain hybrid dominance in nature, especially in extreme and rapidly changing environments. Although any favoured gene complex in cyclic parthenogens could be frozen in successful clones independent of hybridization, we did not find similarly successful clones among parental species. We conclude that the emergence of the observed trait is linked to the production of novel recombined hybrid genotypes.

Genetic structure of Daphnia galeata populations in Eastern China.

This study presents the first examination of the genetic structure of Daphnia longispina complex populations in Eastern China. Only one species, D. galeata, was present across the eight investigated lakes; as identified by taxon assignment using allelic variation at 15 microsatellite loci. Three genetically differentiated D. galeata subgroups emerged independent of the type of statistical analysis applied. Thus, Bayesian clustering, discriminant analysis based on results from factorial correspondence analysis, and UPGMA clustering consistently showed that populations from two neighbouring lakes were genetically separated from a mixture of genotypes found in other lakes, which formed another two subgroups. Clonal diversity was high in all D. galeata populations, and most samples showed no deviation from Hardy-Weinberg equilibrium, indicating that clonal selection had little effect on the genetic diversity. Overall, populations did not cluster by geographical origin. Further studies will show if the observed pattern can be explained by natural colonization processes or by recent anthropogenic impact on predominantly artificial lakes.

Population structure of a microparasite infecting Daphnia: spatio-temporal dynamics.

BACKGROUND: Detailed knowledge of spatial and temporal variation in the genetic population structure of hosts and parasites is required for understanding of host - parasite coevolution. As hot-spots of contemporary coevolution in natural systems are difficult to detect and long-term studies are restricted to few systems, additional population genetic data from various host - parasite systems may provide important insights into the topic. This is particularly true for parasites, as these players have been under-investigated so far due to the lower availability of suitable molecular markers. Here, we traced genetic variation (based on sequence variants in the internal transcribed spacer region, ITS) among seven geographically isolated populations of the ichthyosporean Caullerya mesnili, a common microparasite of the cladoceran Daphnia (here, the D. longispina hybrid complex). At three sites, we also studied parasite genetic variation over time (three to four sampling points) and tested for associations between parasite genotypes and host species. RESULTS: Parasite (and host) populations were significantly structured across space, indicating limited dispersal. Moreover, the frequency of parasite genotypes varied significantly over time, suggesting rapid evolutionary change in Caullerya. However, the distribution of parasite genotypes was similar across different host species, which might in turn have important consequences for parasite epidemiology. CONCLUSIONS: The approach proposed here can be applied to track spatial and temporal changes in the population structure of other microparasite species for which sequence variation in the ITS or other highly variable genome regions has been documented but other types of polymorphic markers are lacking. Screening of parasite sequence variants allows for reliable detection of cross-species infections and, using advanced sequencing techniques in the near future, for detailed studies of parasite evolution in natural host - parasite systems.

Hybridizing Daphnia communities from ten neighbouring lakes: spatio-temporal dynamics, local processes, gene flow and invasiveness.

BACKGROUND: In natural communities of cyclical parthenogens, rapid response to environmental change is enabled by switching between two reproduction modes. While long periods of asexual reproduction allow some clones to outcompete others, and may result in "clonal erosion", sexual reproduction restores genetic variation in such systems. Moreover, sexual reproduction may result in the formation of interspecific hybrids. These hybrids can then reach high abundances, through asexual clonal reproduction. In the present study, we explored genetic variation in water fleas of the genus Daphnia. The focus was on the short-term dynamics within several clonal assemblages from the hybridizing Daphnia longispina complex and the impact of gene flow at small spatial scales. RESULTS: Daphnia individuals belonged either to the parental species D. galeata and D. longispina, or to different hybrid classes, as identified by 15 microsatellite markers. The distribution and genotypic structure of parental species, but not hybrids, corresponded well with the geographical positions of the lakes. Within parental species, the genetic distance among populations of D. galeata was lower than among populations of D. longispina. Moreover, D. galeata dominance was associated with higher phosphorous load. Finally, there was no evidence for clonal erosion. CONCLUSIONS: Our results suggest that the contemporary structure of hybridizing Daphnia communities from ten nearby lakes is influenced by colonization events from neighbouring habitats as well as by environmental factors. Unlike the parental species, however, there was little evidence for successful dispersal of hybrids, which seem to be produced locally. Finally, in contrast to temporary Daphnia populations, in which a decrease in clonal diversity was sometimes detectable over a single growing season, the high clonal diversity and lack of clonal erosion observed here might result from repeated hatching of sexually produced offspring. Overall, our study provides insights into spatio-temporal dynamics in a hybridizing Daphnia species complex in a recently established lake system, and relates genetic similarities of populations to a scenario of secondary invasion enhanced by environmental factors.

Capturing the population structure of microparasites: using ITS-sequence data and a pooled DNA approach.

The internal transcribed spacer (ITS) region of nuclear ribosomal DNA is a common marker not only for the molecular identification of different taxa and strains, but also for the analysis of population structure of wild microparasite communities. Importantly, the multicopy nature of this region allows the amplification of low-quantity samples of the target DNA, a common problem in studies on unicellular, unculturable microparasites. We analysed ITS sequences from the protozoan parasite Caullerya mesnili (class Ichthyosporea) infecting waterflea (Daphnia) hosts, across several host population samples. We showed that analysing representative ITS-types [as identified by statistical parsimony network analysis (SPN)] is a suitable method to address relevant polymorphism. The spatial patterns were consistent regardless of whether parasite DNA was extracted from individual hosts or pooled host samples. Remarkably, the efficiency in detecting different sequence types was even higher after sample pooling. As shown by simulations, an easily manageable number of sequences from pooled DNA samples are sufficient to resolve the spatial population structure in this system. In summary, the ITS region analysed from pooled DNA samples can provide valuable insights into the spatial and temporal dynamics of microparasites. Moreover, the application of SPN analysis is a good alternative to the well-established neighbour-joining method (NJ) for the identification of representative ITS-types. SPN can even outperform NJ by joining most of the singleton sequences to representative sequence clusters.

Clonal diversity, clonal persistence and rapid taxon replacement in natural populations of species and hybrids of the Daphnia longispina complex.

Hybridization is common among cyclical parthenogens, especially in zooplankton species assemblages of the genus Daphnia. To explore hybridization dynamics and the extent of clonal diversity in the Daphnia longispina complex, we analysed population structure in eight permanent lakes. Based on 15 microsatellite loci, three major taxonomic units emerged: two species, D. galeata and D. longispina and their F1 hybrids, supported by factorial correspondence analysis and two Bayesian methods. At the same time, the detection of backcross classes differed between methods. Mean clonal diversity was lowest in the F1 hybrids, as expected from the high rate of asexual reproduction. Within taxa, replicated genotypes were of clonal origin, but clonal lineages persisted in subsequent years in only one of three resampled lakes. In another lake, the taxon composition changed from being dominated by hybrids to complete dominance by one parental taxon. Such a year-to-year taxon replacement has not been reported for the D. longispina complex before. Our data on this hybrid complex illustrate that high-resolution genotyping is essential for the understanding of ecological and evolutionary outcomes of hybridization in partially clonal taxa.

Molecular identification and hidden diversity of novel Daphnia parasites from European lakes.

Parasites play important roles in local population dynamics and genetic structure. However, due to insufficient diagnostic tools, detailed host-parasite interactions may remain concealed by hidden parasite diversity in natural systems. Microscopic examination of 19 European lake Daphnia populations revealed the presence of three groups of parasites: fungi, microsporidia, and oomycetes. For most of these parasites no genetic markers have been described so far. Based on sequence similarities of the nuclear small-subunit and internal transcribed spacer (ITS) rRNA gene regions, one fungus, four microsporidian, and nine oomycete taxa were discovered in 147 infected Daphnia (and/or three other zooplankton crustaceans). Additionally, cloning of rRNA gene regions revealed parasite sequence variation within host individuals. This was most pronounced in the ITS region of one microsporidian taxon, where the within-host sequence variation ranged from 1.7% to 5.3% polymorphic sites for parasite isolates from 14 different geographical locations. Interestingly, the parasite isolates from close locations grouped together based on sequence similarities, suggesting that there was parasite dispersal. Taken together, the data obtained in this study revealed hidden diversity of parasite communities in Daphnia lake populations. Moreover, a higher level of resolution for identifying parasite strains makes it possible to test new hypotheses with respect to parasite dispersal, transmission routes, and coinfection.

Dynamic reticulate evolution in a Daphnia multispecies complex.

Recent genomic data suggest that the role of hybridization in evolution might be more important than previously assumed. Here, we examine species-specific differentiation and signatures of reticulation in a multi-species complex of the crustacean genus Daphnia. We did a combined examination of mtDNA, allozymes and ITS1 (a part of the nuclear ribosomal DNA) in the Daphnia longispina group. We focused on the sequence variation of ITS1 in two unambiguous species (D. galeata, D. cucullata) and two ecotypes hyalina and rosea within the recently revised taxon D. longispina (O.F. Müller). We found two ITS1-types (S, L) and intra-individual and intra-specific polymorphisms. Another ITS1-type (XL) was restricted to the outgroup D. umbra. S was present in all taxa but occurred as only two variants. Surprisingly, D. galeata and D. cucullata, which were well differentiated by mtDNA and allozymes, were virtually indistinguishable with respect to S-ITS1 (S(cg)). The two ecotypes of D. longispina shared the second S-ITS1-variant (S(rh)) and were therefore almost indistinguishable for all types of molecular markers surveyed. The L-type differed between D. galeata and D. longispina samples, but L was absent in D. cucullata. Between hyalina and rosea ecotypes, we found some L-differentiation. Combined data suggest that reticulate evolution enabled the spread of one S-ITS1-variant (S(cg)) beyond species boundaries and that S-introgression was species-specific, despite contemporary hybridization between all species. Our data have implications for phylogenetic as well as phylogeographic surveys. Because of the dynamic impact of gene flow in multi-species complexes, misinterpretations of presumed species-specific data should be considered.