Groundwater Amphipods Research Articles

Groundwater amphipods of the genus Niphargus Schiødte, 1834 in Boyer-Ahmad region (Iran) with description of two new species

Groundwater amphipods of the genus Niphargus Schiødte, 1834 in Boyer-Ahmad region (Iran) with description of two new species

Comparison of the gut microbiota in the groundwater amphipod Crangonyx islandicus Svavarsson & Kristjánsson, 2006 (Amphipoda: Crangonyctidae) to biofilms in its spring-source habitat

AbstractCrangonyx islandicus Svavarsson & Kristjánsson, 2006 is a groundwater amphipod endemic to Iceland and restricted to spring sources at edges of lava fields in the volcanically active zone. Genetic analysis indicate that the amphipod has survived in subglacial refugia in the groundwater in Iceland during past Ice Ages. We investigated the gut bacteria of C. islandicus and compared them with the bacterial diversity identified in biofilms from its groundwater habitat. The diversity of both uncultivated and cultivated bacteria was analyzed using partial 16S rRNA gene sequences. The results show that iron-oxidizing bacteria such as Sideroxydans and Gallionella and the methanotroph Methylobacter are common in the biofilm but absent from the gut samples. Thiomonas was common in both the biofilm and the gut samples. The cultivation of gut bacteria revealed that most of the taxa could be linked with terrestrial soil and/or freshwater habitat based on available database entries. This indicates that the food source for the amphipods is partly of terrestrial origin. The presence of primary producers in the subsurface habitat of C. islandicus indicate that microbial chemolithotrophic activity might fuel the system at least partly and thus explain the amphipod survival in the subsurface of Iceland during repeated glaciation for the past 5 mya.

Seasonal variation in the invertebrate community and diet of a top fish predator in a thermally stable spring

Many life-history events in aquatic invertebrates are triggered by seasonal changes in water temperature, but other ecological factors may be important as well. To rule out the confounding effects of changing water temperature, we studied the seasonal dynamics of an aquatic invertebrate community and their effect on a top fish predator in a thermally stable freshwater spring in South Iceland. We sampled benthic invertebrates five times over a year and conducted a mark-recapture study on the top predator in the system, small benthic Arctic charr, Salvelinus alpinus (L.). We assessed variation in diet composition and feeding preferences by calculating the electivity and individual specialisation of each fish at each sampling time. There was a clear separation of winter and summer communities for the benthic invertebrates. The variation in prey availability was also reflected in the fish diet, with higher feeding selectivity in summer than in winter for the highly abundant Chironomidae larvae. In contrast, individual specialisation as a measure of intrapopulation niche variation was higher in winter when prey availability was lower. We furthermore found that groundwater amphipods might play an important role in the winter diet of spring-dwelling Arctic charr. In conclusion, seasonal variation in the invertebrate community is an important factor to consider and has the potential to alter the phenotype (e.g. growth rates) and behaviour (e.g. feeding preferences) of higher trophic levels.

Demographic history, range size and habitat preferences of the groundwater amphipod Niphargus puteanus (C.L. Koch in Panzer, 1836)

Niphargus puteanus is the oldest described species of its genus and, in the past, was used as a taxonomic annotation for any subterranean amphipod record. For that reason, no clear knowledge exists about its actual range size and habitat preferences. We here applied a molecular taxonomic and phylogeographical approach based on mitochondrial and nuclear DNA to shed light on its distribution and to infer its demographic history. Furthermore, we analysed aquifer types and water flow regimes to provide a clearer picture of the species’ ecological requirements.Our results indicate that N. puteanus is widely distributed north of the Alps, having its core range in the geomorphological natural region of the ‘South German Scarplands’ (SGS). Additionally, isolated satellite populations exist in the Taunus and the Sauerland, and two single individuals were collected in Luxembourg and in Austria, respectively. The species’ maximal distribution range reaches 756 km between the two single-specimen records and 371 km within the SGS. A very high haplotype diversity was observed, revealing the presence of seven haplotype groups. All the haplogroups were present in the SGS and exhibited distinct spatial-genetic patterns. We thus inferred a high degree of population isolation, with the SGS being a potential long-term refugium for N. puteanus. Historical drift of specimens along major riverine networks (Rhine, Danube) may provide an evolutionary dispersal mechanism explaining the formation of satellite populations.From an ecological perspective, populations of N. puteanus were found to be closely associated with fast-flowing spring environments within fissured aquifers. This species should therefore be classified as both crenophile and stygophile, effectively exploiting resources of surface as well as subsurface spring waters.

Updates to the sporadic knowledge on microsporidian infections in groundwater amphipods (Crustacea, Amphipoda, Niphargidae)

A set of 69 specimens from 19 groundwater species of the genera Niphargus, Niphargellus, Microniphargus and Crangonyx was genetically screened for microsporidian infections. Samples mostly originated from groundwater-dependent spring environments (71%), natural caves (9%) and artificial caverns/tunnels (13%). Amphipod hosts were identified by morphology and/or molecular data, whereas microsporidian parasites were characterised by a genetic screening assay targeting a section of the small subunit rRNA gene. Five microsporidian species (Dictyocoela duebenum; Nosema sp.; Hyperspora aquatica and two undescribed Microsporidium spp.) were revealed from 13 host specimens (Niphargus schellenbergi; N. aquilex lineages B, F and G; Niphargellus arndti). In particular N. schellenbergi was frequently infected with D. duebenum as well as a new and potentially niphargid-specific Nosema sp. identified in Niphargellus arndti. Our results shed further light on the still largely unknown diversity and specificity of microsporidian parasites in groundwater amphipods and subterranean animals in general.

Range sizes of groundwater amphipods (Crustacea) are not smaller than range sizes of surface amphipods: a case study from Iran

The connectivity of groundwater aquifers is lower compared to surface waters. Consequently, groundwater species are expected to have smaller distributional ranges than their surface relatives. Molecular taxonomy, however, unveiled that many species comprise complexes of morphologically cryptic species, with geographically restricted distributional ranges in subterranean as well as in surface waters. Hence, the range sizes of surface and groundwater species might be more similar in size than hitherto thought. We tested this hypothesis by comparing the range size of surface amphipods of the genus Gammarus and subterranean amphipods of the genus Niphargus in Iran. We re-analyzed the taxonomic structure of both genera using two unilocus species delimitation methods applied to a fragment of the COI mitochondrial marker, to identify molecular operational taxonomic units (MOTUs), and assessed the maximum linear extent (MLE) of the ranges of MOTUs from both genera. Genus Gammarus comprised 44–58 MOTUs while genus Niphargus comprised 20–22 MOTUs. The MLEs of the two genera were not significantly different, regardless the delimitation method applied. The results remained unchanged also after exclusion of single site MOTUs. We tentatively conclude that in this case there is no evidence to consider that groundwater species are geographically more restricted than surface species.

Bacterial diversity in Icelandic cold spring sources and in relation to the groundwater amphipod Crangonyx islandicus.

Crangonyx islandicus is a groundwater amphipod endemic to Iceland, considered to have survived the Ice Ages in subglacial refugia. Currently the species is found in spring sources in lava fields along the tectonic plate boundary of the country. The discovery of a groundwater species in this inaccessible habitat indicates a hidden ecosystem possibly based on chemoautotrophic microorganisms as primary producers. To explore this spring ecosystem, we assessed its microbial diversity and analysed whether and how the diversity varied between the amphipods and the spring water, and if was dependent on environmental factors and geological settings. Isolated DNA from spring water and from amphipods was analysed using metabarcoding methods, targeting the 16S rRNA gene. Two genera of bacteria, Halomonas and Shewanella were dominating in the amphipod samples in terms of relative abundance, but not in the groundwater samples where Flavobacterium, Pseudomonas and Alkanindiges among others were dominating. The richness of the bacteria taxa in the microbial community of the groundwater spring sources was shaped by pH level and the beta diversity was shaped by geographic locations.

Two new species of groundwater amphipods of the genus Niphargus Schiödte, 1849 from northwestern Iran

This study was conducted to describe and illustrate two new species of groundwater amphipods from the northern parts of the Zagros Mountains in West Azerbaijan Province, Iran. Mitochondrial (COI) and nuclear (28S rDNA) fragments as well as several morphological traits were used to characterize Niphargus urmiensis sp. nov. and Niphargus fiseri sp. nov. The phylogenetic analyses showed that the nucleotide differences between the recently described species and their close allies are attributed to their distinctiveness. The molecular analysis also introduced that the new species are placed within the clade comprising Iranian species as a sister taxon. The genetic distances between N. urmiensis sp. nov. and N. fiseri sp. nov. are 7.6% and 1.6%, respectively based on the COI and 28S rDNA gene fragments.

Architects of the underworld: bioturbation by groundwater invertebrates influences aquifer hydraulic properties

The hydraulic properties of aquifers are critical to the storage and transmission of water to meet the needs of an increasingly groundwater-dependent global community. The hydraulic properties of aquifers can also influence the biota present, but inversely, invertebrate burrowing (bioturbation) may also influence the hydraulic properties and the flow of water through sediments. The aim of this study was to test whether groundwater invertebrates were capable of influencing the hydraulic properties of aquifer sediments in an experimental setting. Groundwater amphipods were added to sediment-filled laboratory columns, and the effective porosity (neff) and longitudinal dispersivity (αL) of the sediments were compared before and after 2 months of amphipod activity. The neff of columns without amphipods decreased significantly over time whereas in columns containing eight amphipods it remained relatively constant, and in columns with four amphipods it was highly variable. There was no difference in αL between columns with amphipod density or over time. These findings suggest that the amphipods were maintaining the amount of pore space that was actively contributing to transport but their activity was not influencing the distribution of flow paths, and amphipod density is critical to causing or maintaining changes in hydraulic properties.

Groundwater amphipods (Crustacea, Malacostraca) of India, with description of three new cavernicolous species.

Stygofaunistic surveys of peninsular India yielded, among others, three new cavernicolous amphipod taxa: Orientogidiella reducta gen. n., sp. n. in the Borra caves of Andhra Pradesh state, Bogidiella hindustanica sp. n. (Bogidiellidae) and Indoniphargus subterraneus sp. n. (Austroniphargidae) in the Kapiladevi caves of Telangana state. The new genus, Orientogidiella gen. n., is proposed to accommodate all the hitherto known species of the Bogidiella indica-group. This paper gives an illustrated description of the three new species together with their taxonomic affinities. A new record of O. indica comb. n., from the Guthikonda caves in Andhra Pradesh is provided as well. The structure of the Indoniphargus mandible is revisited with scanning electron microscopy (SEM) and revealed a poorly known flexible structure on the molar process which could be useful for phylogenetic purposes. The species diversity and geographic distribution of the Indian stygobiotic amphipods are briefly reviewed.

Is subterranean lifestyle reversible? Independent and recent large-scale dispersal into surface waters by two species of the groundwater amphipod genus Niphargus

Groundwater is an extreme environment due to its absence of light, resource scarcity and highly fragmentary nature. Successful groundwater colonizers underwent major evolutionary changes and exhibit eye and pigment loss (troglomorphies). Consequently, their chances of dispersal and survival in the well-connected surface waters are greatly decreased, resulting in significant endemism. The West Palaearctic subterranean amphipod genus Niphargus comprises hundreds of narrowly endemic and troglomorphic species. Nevertheless, a few are known to occur in surface waters, two of which, N. hrabei and N. valachicus, have extremely large ranges that even exceed those of many surface-water amphipods. We tested if this pattern results from a secondary colonization of the relatively well-connected epigean environment, and whether this ecological shift promoted the large-scale dispersal of these species. Results showed that despite their ecological and zoogeographic similarities, N. hrabei and N. valachicus are not closely related and independently colonized surface waters. Their phylogeographic patterns indicate Middle to Late Pleistocene dispersal episodes throughout the Danube lowlands, and relatively modest yet significant genetic differentiation among populations. Clustering based on morphology revealed that the two species are phenotypically closer to each other than they are to most other epigean congeners. We presume that the ecological shift to surface environments was facilitated by their ability to thrive in hypoxic waters where rheophilic competitors from the family Gammaridae cannot survive. In conclusion, our results indicate that adaptation to groundwater is not a one-way evolutionary path and that troglomorphic species can occasionally recolonize and widely disperse in surface waters.

Groundwater amphipods alter aquifer sediment structure

Video S1. Supporting information Please note: The publisher is not responsible for the content or functionality of any supporting information supplied by the authors. Any queries (other than missing content) should be directed to the corresponding author for the article.

Genetic analysis of ciliates living on the groundwater amphipod Crangonyx islandicus (Amphipoda: Crangonyctidae)

AbstractTwo endemic subterranean freshwater amphipod species have been discovered in groundwater of the volcanic active zone in Iceland, Crangonyx islandicus and Crymostygius thingvallensis. At least five different ciliate sequences were isolated from C. islandicus and the analysis of variation at the 18S ribosomal DNA gene suggests that they present previously unsequenced species, from the orders Apostomatida and Philasterida. Apostome ciliates are well known to be exuviotrophic epibionts on crustaceans. Analyses of ciliate epibionts from different groundwater amphipod species from North America and mainland Europe revealed distinct groups of ciliates and higher diversity on the epigean amphipod Crangonyx pseudogracilis. Analysis of geographic patterns of the most common ciliate within Iceland revealed population differentiation supporting limited current connectivity between the different groundwater systems. Our study reports an occurrence of previously undescribed ciliate species in a groundwater ecosystem characterized by low species diversity.

Completion of molecular taxonomy: description of four amphipod species (Crustacea: Amphipoda: Niphargidae) from Iran and release of database for morphological taxonomy

Niphargus is one of the major freshwater amphipod genera in the world, distributed in groundwater in the Western Palearctic. Iran presents the eastern-most boundary of its range. Recent surveys using molecular markers unveiled that many populations from Western Iran deserve species status. In this study, we morphologically analyzed, described and illustrated four new species: N. persicus sp. nov., N. hosseiniei sp. nov., N. ilamensis sp. nov. and N. sohrevardensis sp. nov. All species were found in the west and north-west of Iran in Elburz and Zagros Mountain ranges. Therefore, the number of Niphargus species from Iran has been raised to 15, and we expect more species will be discovered in the future. In order to facilitate and further stimulate the taxonomy of the genus in the region, we constructed a database in the taxonomic program DELTA. The database describes 31 the most important morphological characters for 29 Asian Niphargus species, and is freely distributed via the World Amphipoda Database. In order to stimulate research on groundwater amphipods in the Middle East, we present how further taxonomy can benefit from this web-available data on morphology.

Biogeography and conservation assessment of Bactrurus groundwater amphipods (Crangonyctidae) in the central and eastern United States

The subterranean amphipod genus Bactrurus (Amphipoda: Crangonyctidae) is comprised of eight species that occur in groundwater habitats in karst and glacial deposits of the central and eastern United States. We reexamine the distribution, biogeography, and conservation status of Bactrurus in light of new species distribution records and divergence time estimates in the genus from a recent molecular study. In particular, we discuss hypotheses regarding the distribution and dispersal of B. mucronatus and B. brachycaudus into previously glaciated regions of the Central Lowlands. We also conducted the first IUCN Red List conservation assessments and reassessed global NatureServe conservation ranks for each species. We identified 17 threats associated with increased extinction risk that vary in source, scope, and severity among species, with groundwater pollution being the most significant threat to all species. Our conservation assessments indicate that five of the eight species are at an elevated risk of extinction under IUCN Red List or NatureServe criteria, with one species (B. cellulanus) already extinct. However, none of the eight species are considered threatened or endangered by any state or federal agency. Significant knowledge gaps regarding the life history, ecology, and demography of each species exist. Given results of our conservation assessments and available information on threats to populations, we offer recommendations for conserva tion, management, and future research for each species.

Detection of catecholamines in single specimens of groundwater amphipods

Catecholamines play essential roles in several physiological processes in vertebrates as well as in invertebrates. While several studies have shown the presence of these substances in surface water invertebrates, their occurrence in groundwater fauna is unproven. In the present study, the presence of different catecholamines (i.e., noradrenaline, adrenaline, and dopamine) in individual specimens of groundwater amphipods of the genus Niphargus (mostly Niphargus inopinatus) was investigated via two independent analytical methods: HPLC/EcD and UPLC/TOF-MS. Mean values for catecholamine levels were 533 pg mg(-1) fresh weight for noradrenaline, 314 pg mg(-1) for adrenaline, and 16.4 ng mg(-1) for dopamine. The optimized protocol allowed the detection of CAs in single organisms of less than 1 mg fresh weight. Catecholamine concentration patterns in groundwater invertebrates are briefly discussed here with respect to their evolutionary adaptation to an environmentally stable, energy-poor habitat.

A new bioassay for the ecotoxicological testing of VOCs on groundwater invertebrates and the effects of toluene on Niphargus inopinatus

A protocol was developed for testing the ecotoxicological effects of volatile organic compounds (VOCs) on groundwater invertebrates. Test substance volatility was addressed in a “closed from start to analysis”-design. Since manifestation of toxic effects may be delayed in ‘slower metabolizing’ organisms such as groundwater fauna, a time-independent (TI-) approach was adopted. Toluene was used as a model substance and its toxicity to the groundwater amphipod Niphargus inopinatus was assessed as an example. The method evaluation process considered various methodological issues such as partitioning of the toxicant between the water and the gas phase (Henry equilibrium), the possible depletion of oxygen in closed test vials, as well as microbial biodegradation of the test substance. For N. inopinatus, an LC50,14 days of 46.6mgL−1 toluene was obtained. The ultimate LC50 value was estimated at 23.3mgL−1 toluene. No oxygen depletion occurred in the test vials and Henry equilibrium was found to be established after 6h. The new test system proposed now awaits broad practical application.

The ITS region of groundwater amphipods: length, secondary structure and phylogenetic information content in Crangonyctoids and Niphargids

The phylogenetic analysis of groundwater amphipods is challenging due to the lack of suitable morphological characters. However, molecular phylogenies based on the 18S and 28S nuclear genes of two Crangonyctoidea species endemic to Iceland, Crymostygius thingvallensis and Crangonyx islandicus, support the taxonomy of these species on the basis of morphological characters. Molecular analyses suggest that the genus Crangonyx is paraphyletic, with the species that is found in Eurasia being highly divergent genetically from the species present in North America and Iceland. Studies of the phylogenetic relationships within the genus Niphargus also warrant further work. The nuclear ITS2 region has recently been proposed as a barcoding marker for plants and animals. In addition, ITS2 has been used to build phylogenies at high taxonomic levels by including its secondary structure. In this study, we want to evaluate the applicability of the ITS region for this group of species and describe its characteristics. The taxonomy of C. thingvallensis, as well as the paraphyly of the genus Crangonyx, is supported herein by phylogenies based on the ITS2 variation. The secondary structure and the length of the ITS2 sequences of the Crangonyctoidea and the Niphargidae species studied are highly variable and are characterized by duplications. The ITS2 sequence of Niphargus plateaui is the longest metazoan sequence deposited in the ITS2 database so far. Although saturation was observed in the nucleotide variation of this marker, the addition of the secondary structure information for the reconstruction of the phylogeny did not add support to the phylogenetic trees. The ITS1 region, which is known to be more variable than ITS2 and bears a large duplication within C. islandicus, was found to be less useful for phylogenetic reconstruction.

Classification of Crangonyx islandicus (Amphipoda, Crangonyctidae) based on morphological characters and comparison with molecular phylogenies

Crangonyx islandicus is a groundwater amphipod species endemic to Iceland and a member of the family Crangonyctidaewhich is widespread throughout the Holarctic region. A previous study based on variation in nuclear genes showed thatC. islandicus closest relatives are Crangonyx species from North America, where most of the species diversity of the genusoccurs. In this study, we aim at describing the morphological affinities of C. islandicus with North American Crangonyxspecies and two Eurasian species, C. chlebnikovi and C. subterraneus. We used the morphological characters defined byZhang & Holsinger (2003) and scored them for different populations of C. islandicus as well as for C. chlebnikovi and C.subterraneus. The molecular phylogeny of the Crangonyctidae family and the morphological character states are com-pared in order to detect potential convergence in the morphological traits, common in subterranean species. C. islandicusappears to be close morphologically to C. subterraneus from Europe and C. longicarpus from North America. The morphological proximity of C. islandicus and C. subterraneus is incongruent with the pattern observed from molecular data.

Molecular taxonomy and phylogenetic affinities of two groundwater amphipods, Crangonyx islandicus and Crymostygius thingvallensis, endemic to Iceland

The amphipod superfamily Crangonyctoidea is distributed exclusively in freshwater habitats worldwide and is characteristic of subterranean habitats. Two members of the family, Crangonyx islandicus and Crymostygius thingvallensis, are endemic to Iceland and were recently discovered in groundwater underneath lava fields. Crangonyx islandicus belongs to a well-known genus with representatives both in North America and in Eurasia. Crymostygius thingvallensis defines a new family, Crymostygidae. Considering the incongruences observed recently between molecular and morphological taxonomy within subterranean species, we aim to assess the taxonomical status of the two species using molecular data. Additionally, the study contributes to the phylogenetic relationships among several crangonyctoidean species and specifically among species from four genera of the family Crangonyctidae. Given the available data we consider how the two Icelandic species could have colonized Iceland, by comparing geographical origin of the species with the phylogeny.Regions of two nuclear (18S and 28S rRNA) and two mitochondrial genes (16S rRNA and COI) for 20 different species of three families of the Crangonyctoidea were sequenced. Four different methods were used to align the RNA gene sequences and phylogenetic trees were constructed using bayesian and maximum likelihood analysis. The Crangonyctidae monophyly is supported. Crangonyx islandicus appeared more closely related to species from the Nearctic region. Crymostygius thingvallensis is clearly divergent from the other species of Crangonyctoidea. Crangonyx and Synurella genera are clearly polyphyletic and showed a geographical association, being split into a Nearctic and a Palearctic group.This research confirms that the studied species of Crangonyctidae share a common ancestor, which was probably widespread in the Northern hemisphere well before the break up of Laurasia. The Icelandic species are of particular interest since Iceland emerged after the separation of Eurasia and North America, is geographically isolated and has repeatedly been covered by glaciers during the Ice Age. The close relation between Crangonyx islandicus and North American species supports the hypothesis of the Trans-Atlantic land bridge between Greenland and Iceland which might have persisted until 6 million years ago. The status of the family Crymostygidae is supported, whereas Crangonyx islandicus might represent a new genus. As commonly observed in subterranean animals, molecular and morphological taxonomy led to different conclusions, probably due to convergent evolution of morphological traits. Our molecular analysis suggests that the family Crangonyctidae needs taxonomic revisions.