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Whole genome sequencing is instrumental for the study of genome variation in natural populations, delivering important knowledge on genomic modifications and potential targets of natural selection at the population level. Large dormant eggbanks of aquatic invertebrates such as the keystone herbivore Daphnia, a microcrustacean widespread in freshwater ecosystems, provide detailed sedimentary archives to study genomic processes over centuries. To overcome the problem of limited DNA amounts in single Daphnia dormant eggs, we developed an optimized workflow for whole genome amplification (WGA), yielding sufficient amounts of DNA for downstream whole genome sequencing of individual historical eggs, including polyploid lineages. We compare two WGA kits, applied to recently produced Daphnia magna dormant eggs from laboratory cultures, and to historical dormant eggs of Daphnia pulicaria collected from Arctic lake sediment between 10 and 300 years old. Resulting genome coverage breadth in most samples was ~70%, including those from >100-year-old isolates. Sequence read distribution was highly correlated among samples amplified with the same kit, but less correlated between kits. Despite this, a high percentage of genomic positions with single nucleotide polymorphisms in one or more samples (maximum of 74% between kits, and 97% within kits) were recovered at a depth required for genotyping. As a by-product of sequencing we obtained 100% coverage of the mitochondrial genomes even from the oldest isolates (~300 years). The mitochondrial DNA provides an additional source for evolutionary studies of these populations. We provide an optimized workflow for WGA followed by whole genome sequencing including steps to minimize exogenous DNA.

DNA can be preserved in marine and freshwater sediments both in bulk sediment and in intact, viable resting stages. Here, we assess the potential for combined use of ancient, environmental, DNA and timeseries of resurrected long-term dormant organisms, to reconstruct trophic interactions and evolutionary adaptation to changing environments. These new methods, coupled with independent evidence of biotic and abiotic forcing factors, can provide a holistic view of past ecosystems beyond that offered by standard palaeoecology, help us assess implications of ecological and molecular change for contemporary ecosystem functioning and services, and improve our ability to predict adaptation to environmental stress., Ellegaard et al. discuss the potential for using ancient environmental DNA (eDNA), combined with resurrection ecology, to analyse trophic interactions and evolutionary adaptation to changing environments. Their Review suggests that these techniques will improve our ability to predict genetic and phenotypic adaptation to environmental stress.

Whole genome sequencing is instrumental for the study of genome variation in natural populations, delivering important knowledge on genomic modifications and potential targets of natural selection at the population level. Large dormant eggbanks of aquatic invertebrates such as the keystone herbivore Daphnia, a microcrustacean widespread in freshwater ecosystems, provide detailed sedimentary archives to study genomic processes over centuries. To overcome the problem of limited DNA amounts in single Daphnia dormant eggs, we developed an optimised workflow for whole genome amplification (WGA), yielding sufficient amounts of DNA for downstream whole genome sequencing of individual historical eggs, including polyploid lineages. We compare two WGA kits, applied to recently produced Daphnia magna dormant eggs from laboratory cultures, and to historical dormant eggs of Daphnia pulicaria collected from Arctic lake sediment between 10y and 300y old. Resulting genome coverage breadth in most samples was ~70%, including those from >100y old isolates. Sequence read distribution was highly correlated among samples amplified with the same kit, but less correlated between kits. Despite this, a high percentage of genomic positions with SNPs in one or more samples (maximum of 74% between kits, and 97% within kits) were recovered at a depth required for genotyping. As a by-product of sequencing we obtained 100% coverage of the mitochondrial genomes even from the oldest isolates (~300y). The mtDNA provides an additional source for evolutionary studies of these populations. We provide an optimised workflow for WGA followed by whole genome sequencing including steps to minimise exogenous DNA.

The American brine shrimp Artemia franciscana is important in aquaculture and has become invasive in other continents, aided by dispersal via waterbirds. However, little is known about processes underlying its genetic diversity and population structure in its natural habitat in North America. These processes, including dispersal and local adaptation, are pivotal drivers of species distribution and community structure, and therefore central to aquatic biodiversity. We studied 15 populations in natural saline lakes of Saskatchewan, Canada to determine the influence of variation in geological history, water chemistry, lake size, and location. We aimed to determine the relative importance of isolation by distance and isolation by environment using the cytochrome c oxidase subunit 1 gene (CO1) as a mitochondrial marker and five nuclear microsatellite markers. Geographic patterns for CO1 and microsatellites differed, with lakes clustering in different groups based on genetic distances according to the marker used. CO1 better indicated historical colonisation processes, suggesting potential routes of initial colonisation when lakes were formed after deglaciation 11,000–15,000 years ago. Differentiation between lakes based on nuclear markers was strongly related to variation in hydrochemistry, suggested by distance-based redundancy analysis, but there was no indication of isolation by distance. The ratio between alkalinity and the sum of Ca and Mg concentrations was particularly important, although a lake with a high Cl concentration caused by potash mining also had a unique Artemia population. Geochemistry is important in the adaptive radiation of anostracan crustaceans. Our study suggests that it also underlies intraspecific genetic variation between populations, promoting isolation by environment, and making dispersal ineffective when cysts are moved by birds between lakes with different hydrochemistry

Paleolimnologists have utilized lake sediment records to understand historical lake and landscape development, timing and magnitude of environmental change at lake, watershed, regional and global scales, and as historical datasets to target watershed and lake management. Resurrection ecologists have long recognized lake sediments as sources of viable propagules (“seed or egg banks”) with which to explore questions of community ecology, ecological response, and evolutionary ecology. Most researchers consider Daphnia as the primary model organism in these efforts, but many other aquatic biota, from viruses to macrophytes, similarly produce viable propagules that are incorporated in the sediment record but have been underutilized in resurrection ecology. The common goals shared by these two disciplines have led to mutualistic and synergistic collaborations—a development that must be encouraged to expand. We give an overview of the achievements of paleolimnology and the reconstruction of environmental history of lakes, review the untapped diversity of aquatic organisms that produce dormant propagules, compare Daphnia as a model of resurrection ecology with other organisms amenable to resurrection studies, especially diatoms, and consider new research directions that represent the nexus of these two fields.

This perspective provides an overview to the Special Issue on Resurrection Ecology (RE). It summarizes the contributions to this Special Issue, and provides background information and future prospects for the use of RE in both basic and applied evolutionary studies.

Knowledge of the molecular basis of phenotypic responses to environmental cues is key to understanding the process of adaptation. Insights to adaptation at an evolutionary time scale can be gained by observing organismal responses before and after a shift in environmental conditions, but such observations can rarely be made. Using the ecological and genomic model Daphnia, we linked transcriptomic responses and phosphorus (P)-related phenotypic traits under high and low P availability. We mapped weighted gene coexpression networks to traits previously assessed in resurrected ancient (600 years old) and modern Daphnia pulicaria from a lake with a historic shift in P-enrichment. Subsequently, we assessed evolutionary conservation or divergence in transcriptional networks of the same isolates. We discovered highly preserved gene networks shared between ancient genotypes and their modern descendants, but also detected clear evidence of transcriptional divergence between these evolutionarily separated genotypes. Our study highlights that phenotypic evolution is a result of molecular fine-tuning on different layers ranging from basic cellular responses to higher order phenotypes. In a broader context, these findings advance our understanding how populations are able to persist throughout major environmental shifts.

Cultural eutrophication is one of the largest threats to aquatic ecosystems with devastating ecological and economic consequences. AlthoughDaphnia, a keystone grazer crustacean, can mitigate some negative effects of eutrophication in freshwater habitats, it is itself affected by changes in nutrient composition. Previous studies have shown evolutionary adaptation ofDaphniato environmental change, including high phosphorus levels. While phosphorus is an essential macronutrient, it negatively affectsDaphnialife history when present in excessive concentrations. Here, we map weighted gene co-expression networks to phosphorus-related phenotypic traits in centuries-old ancestralDaphniaand their modern descendants, contrasting pre- and post-eutrophication environments. We find that evolutionary fine-tuning of transcriptional responses is manifested at a basic (cellular) phenotypic level, and is strongly correlated to an evolved plasticity of gene expression. At a higher phenotypic level, this contributes to the maintenance of physiological homeostasis, and a robust preservation of somatic phosphorus concentration.

1. The high Arctic, including the Svalbard archipelago in the North Atlantic, has been exposed to direct and indirect drivers of climatic change such as rising temperatures and associated changes in hydrology and nutrient fluxes. In addition, the number of migrating birds, particularly geese, increased remarkably in the Svalbard archipelago during the second half of the last century. The higher number of breeding birds potentially affects water quality and the biota in ponds and lakes. 2. We aimed to investigate the potential influence of increasing goose abundance on trophic state, taxon richness, and species composition of freshwater communities in the high Arctic. We hypothesised that higher goose abundance affects the trophic state of shallow lakes and ponds and their taxon richness and species composition. We conducted a survey of selected ponds at Svalbard along a goose abundance gradient. We used the number of area-specific goose droppings (range of 0–94 droppings m2) as a proxy of goose presence and measured proxies for productivity as well as taxon richness and composition of phytoplankton and invertebrate communities. 3. Presence and abundance of geese were associated with higher productivity of ponds. Invertebrate and phytoplankton taxon richness correlated (positively) with goose abundance. Both phytoplankton and invertebrate taxon richness increased with increasing nitrogen (N) concentrations. Goose abundance significantly affected phytoplankton species composition, while concentrations of total-N and total phosphorus (P) did not. Species composition of aquatic invertebrates was most strongly affected by goose abundance, but the effect of total-N concentration was also significant. 4. Increased goose abundance was associated with bird driven nutrient enrichment, increased phytoplankton and invertebrate taxon richness and changes of these biological communities. Thus, in addition to climate change, the higher abundances of large migratory water fowl in many polar areas may pose a major additional stress to arctic lakes and ponds. In fact, climate change and bird impact may interact, accelerating ongoing environmental change of arctic freshwater ecosystems. climate change, geese, invertebrates, phytoplankton, Svalbard

Little is known about the role of transcriptomic changes in driving phenotypic evolution in natural populations, particularly in response to anthropogenic environmental change. Previous analyses of Daphnia genotypes separated by centuries of evolution in a lake using methods in resurrection ecology revealed striking genetic and phenotypic shifts that were highly correlated with anthropogenic environmental change, specifically phosphorus (P)-driven nutrient enrichment (i.e. eutrophication). Here, we compared the transcriptomes of two ancient (~700-year-old) and two modern (~10-year-old) genotypes in historic (low P) and contemporary (high P) environmental conditions using microarrays. We found considerable transcriptomic variation between 'ancient' and 'modern' genotypes in both treatments, with stressful (low P) conditions eliciting differential expression (DE) of a larger number of genes. Further, more genes were DE between 'ancient' and 'modern' genotypes than within these groups. Expression patterns of individual genes differed greatly among genotypes, suggesting that different transcriptomic responses can result in similar phenotypes. While this confounded patterns between 'ancient' and 'modern' genotypes at the gene level, patterns were discernible at the functional level: annotation of DE genes revealed particular enrichment of genes involved in metabolic pathways in response to P-treatments. Analyses of gene families suggested significant DE in pathways already known to be important in dealing with P-limitation in Daphnia as well as in other organisms. Such observations on genotypes of a single natural population, separated by hundreds of years of evolution in contrasting environmental conditions before and during anthropogenic environmental changes, highlight the important role of transcriptional mechanisms in the evolutionary responses of populations.

A laboratory microcosm experiment was conducted to examine the roles of food quantity and food quality on the competitive abilities of clones of the keystone aquatic zooplankter, Daphnia pulicaria. Using methods of resurrection ecology, clones were established by hatching dormant eggs from sediment layers of a lake (South Center, MN, U.S.A) that were separated by centuries of environmental change (nutrient enrichment). Two sets of paired clones (“modern” vs. “ancient”) were competed against each other in a 2 × 2 factorial design of high/low food quantity and high/low food quality. Experimental carbon:phosphorus (C:P) ratios were designed to mimic food quantity and food quality parameters respective to each of the two time periods from which the clones were extracted to test whether clones are competitively superior under conditions representative of the time period in which they existed. Contrary to predictions, we found that “modern” clones were able to outcompete “ancient” clones under all food treatments. We discuss potential mechanisms (i.e., mutational input, viability of long-dormant eggs, differences in phenotypic plasticity) that may have differentially impacted the performance of experimental clones. Our results should aid in deciphering microevolutionary dynamics observed in resurrection ecology studies, a powerful tool that can illuminate adaptive dynamics of organisms to environmental changes on long-term (centuries-long) temporal scales.

Population genetic surveys approximately 25 years apart examined the distribution and abundance of asexual clones of the freshwater zooplankter Daphnia pulex complex in rock pools near Churchill, Manitoba, Canada. In 1984–1985, melanic members of this species complex were present in 131 rock pools at this site, but were only detected in 90 of these pools in 2007–2008. Allozymic surveys conducted during these two time periods revealed that 59 per cent of these populations showed unchanged clonal composition. Total clonal replacement occurred in 8 per cent of the populations, while the others (33%) included a mixture of ‘resident’ clones and new ‘colonists’. We discuss these changes in light of shifts in biotic and abiotic factors. We also discuss the use of rock pool habitats as ‘sentinel’ systems for examining long-term environmental changes in the ecological genetics of aquatic organisms in the Arctic.

Zooplankton in temporary ponds is often collected with gear originally designed for lakes, and mostly unsuitable for sampling shallow habitats. We describe a new simple and inexpensive device for sampling zooplankton in very shallow, vegetated temporary ponds. We tested the sampling efficiency by comparing species composition and density of cyclopoid copepods, an important component of the zooplankton, by sampling with both the new bag sampler and a plastic beaker frequently employed for collections of zooplankton in small waterbodies. With the bag sampler we collected a larger number of species and higher densities of copepods due to its higher efficiency in vegetated areas and near the sediment. The beaker appeared to sample almost only the water surface. The samples collected with the bag sampler revealed a distinct distribution of copepod life cycle stages in a shallow pond, which differed between depths and microhabitats. Additional advantages of the bag sampler are its small size and weight, and the possibility of fast exchange of sample bags between sample locations, thus preventing accidental faunal exchange between sample locations. We conclude that the bag sampler is a device especially useful for sampling zooplankton of shallow ponds and wetlands rich in vegetation, for diversity studies as well as for quantitative sampling. (© 2005 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim)

We examined the impact of lateral and longitudinal hydrological connectiv- ity on copepod diversity (local species richness, diapause strategies and population genetics) in ponds of a North American floodplain. Results of a General Linear Model (GLM) which included three environmental pond variables showed that species rich- ness in these ponds was significantly influenced only by pond type (permanent ponds, temporary ponds with connections to other waterbodies during floods, isolated tempo- rary ponds). Local species richness in connected temporary ponds had significantly higher scores of local richness compared to isolated temporary ponds (means 7.4 and 2.7, respectively). Such enhancement of species diversity in connected ponds appears to be facilitated by increased dispersal during floods and also pertains to the diversity of diapause strategies in the ponds studied. Species with and without diapause were present in connected temporary ponds, but absent from isolated temporary ponds, which contained only species capable of diapausing during the dry season. To explore the significance of hydrological connectivity of longitudinal dispersal between up- stream and downstream sites, we used RAPD markers in populations of a calanoid co- pepod. The increase of average heterozygosity H (0.301 to 0.477) in an upstream- downstream direction and generally low genetic distances (Nei's D < 0.05) suggested maintenance of gene flow by flood-mediated dispersal between connected ponds. Lower heterozygosity (0.289) of the population in a downstream pond protected by le- vees suggested the interruption of gene flow when dispersal by flooding is blocked.

SUMMARY 1. We explored the quantitative contribution of two alternative recolonisation strategies by a combination of experimental and field data from the Little Tallahatchie floodplain (Mississippi, U.S.A.). In this area, several floods of short duration occur in winter and spring. During floods, lentic and lotic waterbodies of different size are connected for a limited period. 2. Shortly before the onset of the flood season we collected sediment samples from six temporary ponds. The emergence of seven cyclopoid species was recorded from the experimentally flooded sediments and was quantified over a 4-week period. Water samples were taken in the same ponds after flooding, from which eight further species were recorded and quantified. 3. The contribution of flood-dispersed individuals was variable among flooded ponds (between 38 and 94%), but numbers of flood-dispersed colonists did not differ significantly from those of hatching individuals in flooded ponds. Our results demonstrate the equal importance of both hydrological connections and hatching from dormant stages as pathways for recolonisation by copepods if ponds are hydrologically connected. 4. The significance of recolonisation by hatching individuals in disconnected waterbodies was apparent in an isolated temporary pond that was exclusively recolonised by cyclopoid copepods emerging from the sediment.

1. The survival of cyclopoid copepods was investigated in a floodplain for 2 years where flooding occurred during the cold season. The cyclopoid community was studied in three waterbodies with distinct hydroperiods: a permanent pond connected to the flooded area during inundation, a temporary pool that is part of the flooded area and an isolated temporary pool. 2. Field studies, including data obtained from samples of water, sediment and soil, showed the overall predominance of species with a summer diapause over those with a winter diapause or without diapause. Emergence of cyclopoid copepods at the onset of flooding, examined using emergence traps and data from recently filled or still isolated temporary pools, showed that some species can survive several months of drying. 3. The ability of the diapausing fourth copepodid stages of Cyclops strenuus and C. insignis, the two cyclopoids most abundant during winter and spring flooding, to survive terrestrial conditions was tested in laboratory experiments. Both species survived for several months, but rates differed among the species. A higher percentage of C. strenuus survived for a longer period, possibly explaining why this species was relatively more abundant in more temporary habitats. 4. Both dormancy and dispersal facilitated survival of cyclopoid copepods in transient habitats connected to each other during flood periods. Dormancy was the most important survival strategy, whereas dispersal could be more important following prolonged periods without flooding.

For an accurate assessment of the anthropogenic impacts on evolutionary change in natural populations, we need long-term environmental, genetic and phenotypic data that predate human disturbances. Analysis of c. 1600 years of history chronicled in the sediments of South Center Lake, Minnesota, USA, revealed major environmental changes beginning c. 120 years ago coinciding with the initiation of industrialised agriculture in the catchment area. Population genetic structure, analysed using DNA from dormant eggs of the keystone aquatic herbivore, Daphnia pulicaria, suggested no change for c. 1500 years prior to striking shifts associated with anthropogenic environmental alterations. Furthermore, phenotypic assays on the oldest resurrected metazoan genotypes (potentially as old as c. 700 years) indicate significant shifts in phosphorus utilisation rates compared to younger genotypes. Younger genotypes show steeper reaction norms with high growth under high phosphorus (P), and low growth under low P, while ‘ancient’ genotypes show flat reaction norms, yet higher growth efficiency under low P. Using this resurrection ecology approach, environmental, genetic and phenotypic data spanning pre- and post-industrialised agricultural eras clearly reveal the evolutionary consequences of anthropogenic environmental change.

Daphnia lumholtzi is a planktonic crustacean native to subtropical regions in Africa, Asia and Australia. Since its invasion to the southern USA in ~1990 it has spread across North America as far north as the Laurentian Great Lakes. We assessed invasion history using microsatellite makers and to explore the influence of mean annual temperature on the genetic structure along a latitudinal gradient in North America. Genotypic data were obtained from 9 microsatellite markers for 178 individuals from 13 populations (eight populations introduced to North America and five populations in the native range). Pairwise Fst values as well as Bayesian clustering showed a strong subdivision between native and introduced populations. Bayesian clustering identified multiple genetic clusters in recently invaded locations, suggestive of multiple invasions from various sources, including Asia and Africa. Using variation partitioning, we determined the amount of variation for genetic clusters of populations in the invaded range due to mean annual air temperature and the year of first detection. The results point to a primary introduction into the southern range of North America, with a subsequent northward expansion, and multiple introductions possibly from both the native range and by secondary spread from previously-invaded locations. Separate analysis of genetic clusters within the invaded range suggests additional effects of temperature conditions on geographic genetic structure, possibly as a consequence of D. lumholtzi's tropical origin. © 2012 Springer Science+Business Media B.V.

Background Copepods outnumber every other multicellular animal group. They are critical components of the world's freshwater and marine ecosystems, sensitive indicators of local and global climate change, key ecosystem service providers, parasites and predators of economically important aquatic animals and potential vectors of waterborne disease. Copepods sustain the world fisheries that nourish and support human populations. Although genomic tools have transformed many areas of biological and biomedical research, their power to elucidate aspects of the biology, behavior and ecology of copepods has only recently begun to be exploited. Discussion The extraordinary biological and ecological diversity of the subclass Copepoda provides both unique advantages for addressing key problems in aquatic systems and formidable challenges for developing a focused genomics strategy. This article provides an overview of genomic studies of copepods and discusses strategies for using genomics tools to address key questions at levels extending from individuals to ecosystems. Genomics can, for instance, help to decipher patterns of genome evolution such as those that occur during transitions from free living to symbiotic and parasitic lifestyles and can assist in the identification of genetic mechanisms and accompanying physiological changes associated with adaptation to new or physiologically challenging environments. The adaptive significance of the diversity in genome size and unique mechanisms of genome reorganization during development could similarly be explored. Genome-wide and EST studies of parasitic copepods of salmon and large EST studies of selected free-living copepods have demonstrated the potential utility of modern genomics approaches for the study of copepods and have generated resources such as EST libraries, shotgun genome sequences, BAC libraries, genome maps and inbred lines that will be invaluable in assisting further efforts to provide genomics tools for copepods. Summary Genomics research on copepods is needed to extend our exploration and characterization of their fundamental biological traits, so that we can better understand how copepods function and interact in diverse environments. Availability of large scale genomics resources will also open doors to a wide range of systems biology type studies that view the organism as the fundamental system in which to address key questions in ecology and evolution.

We investigated the phylogeography of the main lineages in the tadpole shrimp Triops mauritanicus Ghigi in the south-western Iberian Peninsula, using mito- chondrial 12S and 16S rDNA sequences. Our results indicate that a fourth, hitherto unknown main phylogenetic lineage occurs in Iberia, so that in total, the species is divided into six distinct clades, comprising T. m. mauritanicus, T. m. simplex Ghigi, and four as yet unnamed lineages that appear to be endemic to Iberia. Percentages of sequence divergence among the main clades in T. mauritanicus reach the range reported for recognized species in other notostracan lineages. A thorough morphological investigation also revealed that the differentiation among these lineages is higher than previously thought, and that populations of three of the main clades within T. mauritanicus can be reliably separated from each other and from the remaining lineages based on the morphology of adult males. The remaining clades also show a significant level of morphological differentiation, but include a certain proportion of populations for which the additional application of molecular methods is needed for a reliable determination. The geographic distribu- tions of 12S haplotypes are indicative of frequent dispersal events and gene flow among populations belonging to the same main lineage, but give no evidence of recent migration events among different main lineages, suggesting that there is no gene flow among the latter. Our data thus suggest that the six main lineages within T. mauritanicus represent distinct species. We therefore describe the Iberian lineages as T. baeticus Korn n. sp., T. emeritensis Korn & Pérez-Bote n. sp., T. gadensis Korn & García-de-Lomas n. sp., and T. vicentinus Korn, Machado, Cristo & Cancela da Fonseca n. sp., and reinstate T. simplex Ghigi to full species status. Our data confirm the general, previously recognized pattern of a lower dispersal probability in gonochoric Triops taxa. However, we found evidence that passive dispersal in Triops may be further complicated by a strong habitat dependence of dispersal probability, mediated by prevailing dispersal vectors

Over 30 years after drainage for agricul- ture, a 2700 ha temporary marshland was recently restored in Don˜ ana National Park. We describe the recovery of zooplankton communities (copepods, cladocerans and rotifers) in 47 new temporary ponds excavated as part of the restoration project during the first two hydroperiods (April 2006 and 2007), and compare them to those of eight reference sites in the surrounding marshland. Major changes in the species composition and abundance occurred in new ponds between years. While rotifers and cyclopoid cope- pods dominated in terms of number of individuals in 2006, calanoid copepods and cladocerans were the most abundant groups in 2007. Rotifer species richness was significantly lower in 2007, but there was an increase in Simpson and b-diversity in 2007 owing largely to a dramatic decline in the abundance of Hexarthra cf. fennica (rare in reference sites) from 93% of all rotifer individuals in new ponds in 2006 to only 32% in 2007. In contrast, species richness of copepods and cladocerans was significantly higher in new ponds in 2007, but there were no changes in Simpson diversity. b-Diversity of cladocerans was also significantly higher in 2007. In 2006, the species richness of cladocera and copepods was significantly lower in new ponds than in reference sites, but by 2007 there were no differences in richness or Simpson diversity. Overall, 7 copepod, 13 cladoceran and 26 rotifer taxa were recorded in new ponds, including 80% of taxa recorded in reference sites. These results indicate that zooplankton communities can be rapidly restored in Mediterranean temporary wetlands, at least when large source populations in the surrounding area reduce dispersal limitation. They also illustrate the importance of comparing different metrics of richness and diversity in studies of zooplankton restoration.

The viable propagule banks of a tempo- rary stream were studied from sections with different agricultural history. Hatching of zooplankton (cope- pods, rotifers and cladocerans) was recorded in the laboratory under controlled temperature and light conditions from an agriculturally modified area with average hydroperiods of about a week per year and two semi-natural reference areas with average hyd- roperiods of more than 3 weeks per year. We found significant differences in both taxon richness and abundance of zooplankton hatching between areas, which were lower in the agriculturally modified section, compared to the reference sections. Another factor likely to have influenced hatching in our experiment was conductivity, which differed between the two reference sections and might have affected hatching at high conductivities. For restora- tion purposes, hydrological reconnection of stream segments is important to facilitate dispersal from the high diversity upstream segments to the depleted sites downstream

The sequence in which new colonists reach an empty habitat can be crucial for future development and species structure of communities. It is therefore important to assess species composition and abundance in the initial stages of habitat existence. In the present study we focussed on colonization of newly constructed temporary ponds in Doñana, Southwest Spain, created by removing 30 or 60 cm of top soil. To confirm that no egg-bank was present, we conducted hatching experiments using sediment cores from six new ponds and two reference sites. Hatching was not recorded in the sediment of the new ponds with the exception of two rotifer individuals. In contrast, in the reference sites a maximum of 103 individuals per sample hatched, including cyclopoids, cladocerans, ostracods and rotifers. In the field, water samples were collected from seven ponds after 19 days of their first hydroperiod. Cyclopoid copepods, mostly Metacyclops minutus, had arrived and dominated all sampled ponds. Other taxa were the monogonont rotifer Brachionus plicatilis in four ponds, and the cladoceran Moina brachiata in one pond. The abundance of zooplankton was negatively correlated with conductivity, suggesting that ponds with higher conductivity undergo delayed colonization. We suggest that fast dispersal and dominance of certain cyclopoid copepods during early colonization is related to their ability to store sperm and fast individual development. Given that cyclopoid copepods can survive drought periods in the sediment of temporary ponds, first and early colonization by cyclopoids is likely to have a profound effect on the propagule bank and future plankton communities when the ponds refill.

Speculation about the role of waterbirds in the dispersal of aquatic invertebrates pre-dates Dar- win. However, there is a critical shortage of field studies quantifying such dispersal. We quantified the viability of aquatic invertebrates in the faeces of different waterfowl species collected in the field at different times during winter. Faeces were collected from four duck species (Northern Pintail Anas acuta, Mallard A. platyrhynchos, Shoveler A. clypeata, Eurasian Teal A. crecca) and Eurasian Coot Fulica atra in November 2004 and January 2005. We also collected soil samples from resting sites as an indicator of what may be transported on birds< feet and plumage. Faecal and soil samples were incubated using two treatments (0.4 and 4.0 mS cm–1) to quantify the potential for dispersal between aquatic habitats of different salinities. We found that viable Nematoda, Rotifera, Copepoda, Ostracoda, Insecta (Tipulidae), and Daphnia and Moina cladocerans were trans- ported internally by birds in the wild. We also found evidence that nematodes, rotifers, ostracods, cope- pods, tipulids, chironomids and hemipterans can be dispersed on birds< feet and feathers. The overall incidence of hatching from all samples was higher in January (59.4 %) than in November (11.5 %). With the exception of bdelloid rotifers, we found no evidence that the potential for dispersal between two habitats would be impeded by salinity in the range tested. Our data suggest that the taxonomic range of dispersed invertebrates and the frequency of their dispersal via waterfowl has previously been underestimated

1. During a study of five artificial brackish ponds situated in Donana Natural Park and the Guadalquivir delta between July 2001 and June 2002, a total of six copepod species (two calanoids, three cyclopoids and one harpacticoid) were recorded. All of them are typical of brackish or estuarine habitats. 2. The estuarine calanoid copepod Acartia tonsa (originally from North America) was present in all the ponds studied. This exotic species has been found previously in European estuaries, but has not previously been recorded from artificial wetlands. 3. Both the relative and absolute abundance of the species varied significantly among months and ponds. A. tonsa was most abundant during autumn and spring, while Calanipeda aquae-dulcis was most abundant in summer. This seasonal pattern of the copepod community composition was different from that reported in studies from other European estuaries or wetlands. 4. The invasion by and dominance of A. tonsa in the area of Donana Natural Park has important implications for the conservation of the diverse native zooplankton fauna in the natural marshes of the adjacent Donana National Park, which includes endemic species

Twenty-five sites located in five wetland zones within Donana, south-west Spain were studied for copepod and cladoceran species richness and composition in relation to habitat characteristics from January to March 2004. The c-diversity of copepods and cladocerans combined varied between wetland zones, which differed significantly in conductivity, surface area and abundance of vegetation. However, there were no significant differences between zones in local species richness of either copepods or cladocerans or the two combined. Species richness was significantly higher in sites with intermediate hydroperiods (duration between 3 and 5 months) than in sites with shorter or longer hydroperiods. CCA analyses performed separately for copepods and cladocerans extracted conductivity, surface area, and vegetation cover as the main factors structuring species composition of both groups. Ours is one of the first studies of zooplankton in the natural marshes of the Donana World Heritage Site, and we recorded five species new to the Donana area

Life cycles of the freshwater cyclopoids Cyclops strenuus Fischer and Cyclops insignis Claus are described from populations of one permanent and two temporary waters in a floodplain area. The abundance of these two species was studied in detail, with special regard to instars surviving summer drought. Both species coexist in two locations during the flood period in the cold season. Cyclops insignis is strictly univoltine, whereas Cyclops strenuus may produce two generations. Differences in timing of egg production, mortality of juvenile instars at four different temperatures, and survival of resting stages in terrestrial conditions are discussed as possible characteristics facilitating the coexistence of the two species.