A hotspot of groundwater amphipod diversity on a crossroad of evolutionary radiations

Aim Groundwater harbours an exceptional fauna and provides invaluable ecosystem services, yet is among the least explored and consequently least protected ecosystems. Successful protection of its biodiversity depends on complete species inventories, knowledge of species spatial distribution, and quantification of biodiversity patterns, as well as disentanglement of the processes that shaped biodiversity patterns. We studied the hyper-speciose amphipod genus Niphargus as a model system within a global subterranean biodiversity hotspot. We linked the biodiversity patterns with possible underlying processes and discuss the needs to include information on different origins of biodiversity into conservation approaches. Location Europe, Western Balkans. Methods We analysed biodiversity patterns of Niphargus using two biodiversity metrics, species richness and phylogenetic diversity, on a grid-based approach. To account for high cryptic diversity, we replaced nominal species with taxonomic units identified in unilocus delimitations (MOTUs). We built a time-calibrated multilocus phylogeny of 512 Niphargus MOTUs from within and outside the study area, and calculated Faith's phylogenetic diversity, standardized effect sizes of phylogenetic diversity, and residual of phylogenetic diversity regressed onto species richness. Results Within the study area, we recognized 245 MOTUs, belonging to different Niphargus clades. Species richness is highest in a north-western hotspot, although some species-rich cells were detected also in the south-east. High phylogenetic diversity coincides with high species richness in the north-west, while in the south-east it is lower than expected. Main conclusions We have shown that species richness does not predictably correlate with phylogenetic diversity. This difference suggests that different processes have led to the formation of species-rich areas in the Western Balkans: through a combination of dispersal and speciation in the north-west, and local radiation in the south-east, respectively. This calls for caution in conservation strategies relying solely on number of species and may change the view on conservation priorities within this region

Fifty Years of the Hypotelminorheic: What Have We Learned?

Originally described by Meštrov in 1962, hypotelminorheic habitats are superficial subterranean drainages, typically less than a meter or so in depth, that emerge at small seepage springs. These are persistent wet spots, typically with blackened leaves in small depressions. There may be no flow during dry periods, but the underlying clay retains water above. They share the landscape with other small bodies of water (močila in Slovenian), not necessarily connected with groundwater. Hypotelminorheic habitats (mezišča in Slovenian) usually harbor a fauna dominated by species adapted to subterranean life, characteristically without eyes or pigment. The basic chemistry and hydrology of the habitat is described as are the basic faunal elements. The habitat is placed in a more general context by reviewing how species invade the habitat, their morphology, and their possible connection to deeper subterranean habitats

Description of a widely distributed but overlooked amphipod species in the European Alps

Amphipods are keystone species in many freshwater ecosystems. Understanding their distribution and diversity is crucial to ensure and preserve freshwater ecosystem functioning, particularly in the northern hemisphere. For the European Alps information on amphipods has been relatively limited until recently. We describe a new, widely distributed amphipod species, Gammarus alpinus sp. nov., found across the Alps and analyse its distribution, biogeography as well as genetic and morphological differentiation. Until now, this species has been reported as Gammarus lacustris. Based on genetic and morphometric measurements, we show that G. alpinus is highly divergent from G. lacustris. The latter has a circumpolar distribution, but may be absent from the Alps. The observed occurrence pattern of G. alpinus might be explained by a Pliocene range expansion and vicariance due to climate warming following the Quaternary glaciation. Historical drainage divides suggest a single recolonization route from a distinct southern refugium. While G. lacustris is widely distributed and not endangered at a global scale, G. alpinus is endemic to the Alps and its habitat is negatively affected by eutrophication, non-native species and possibly climate change

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

Abstract 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

Phylogenetic structure and molecular species delimitation hint a complex evolutionary history in an Alpine endemic Niphargus clade (Crustacea, Amphipoda)

Subterranean fauna is an important contributor to the global fauna, but it is still understudied and a large part of its taxonomy is not yet resolved. One species complex with unresolved taxonomy is the groundwater amphipod Niphargus ruffoi, endemic to the Alpine chain. Here, we used new samples from across the Alpine arc to review the taxonomic status of the entire clade, including the species N. ruffoi and Niphargus arolaensis. We sequenced four genetic markers from the collected specimens, assessed the phylogenetic position of N. ruffoi within the genus, and studied the structure of this species complex using four molecular species delimitation methods. We tested for recombination using the alignments of the concatenated nuclear rDNA genes. The phylogenetic analyses revealed high support for the monophyly of the studied species complex, defining two lineages (i.e., N. arolaensis and N. ruffoi) within the clade. Molecular species delimitation methods suggested that N. arolaensis is a single species, while N. ruffoi should be considered as a species complex of three (using ITS) to eight (using COI) putative species. Moreover, we found a discrepancy between the different nuclear ribosomal DNA markers, indicating a possible recombination with fragments of 28S DNA of N. ruffoi s. lat. present in the genome of N. arolaensis. For the above-mentioned reasons, the internal phylogenetic structure of N. ruffoi s. lat. could not be fully resolved. Moreover, no clear morphological evidence supported the molecular species delimitation. Consequently, no taxonomic changes were proposed. We postulate that this complex scenario was influenced by Pleistocene climate oscillations with subsequent fragmentation events and secondary contacts, making this an interesting study system to investigate the evolution and biogeography of Alpine clades

Translating Niphargus barcodes from Switzerland into taxonomy with a description of two new species (Amphipoda, Niphargidae)

The amphipod genus Niphargus (Amphipoda: Niphargidae Bousfield, 1977) is the most species-rich genus of freshwater amphipods in the World. Species of this genus, which live almost exclusively in subterranean water, offer an interesting model system for basic and applied biodiversity science. Their use, however, is often limited due to the hitherto unresolved taxonomy within the whole genus. As a comprehensive taxonomic revision of the currently >425 Niphargus species is too demanding, it has been suggested that the taxonomy of the genus could be advanced in smaller steps, by reviewing regional faunas, that would eventually integrate into a global revision. In this study, we provide such a revision of Niphargus in Switzerland. First, we molecularly delimited, morphologically diagnosed, and formally described two new species, namely Niphargus luchoffmanni sp. n. and Niphargus tonywhitteni sp. n. Second, we updated and revised a checklist of Niphargus in Switzerland with new findings, and prepared a list of reference sequences for routine molecular identification, available at BOLD and GenBank. All available specimens of 22 known species from the area were morphologically examined, and their morphological variation was compiled in a data file of DEscription Language for TAxonomy, which can be used for automated generation of dichotomous or interactive keys. The data file is freely available at the World Amphipoda Database. Together, the checklist, the library of reference sequences, the DELTA file, but also a list of hitherto unresolved aspects are an important step towards a complete revision of the genus within a well-defined and biogeographically interesting area in Central Europe

Low intraspecific genetic divergence and weak niche differentiation despite wide ranges and extensive sympatry in two epigean Niphargus species (Crustacea: Amphipoda)

The amphipod genus Niphargus comprises hundreds of eyeless and depigmented species with narrow ranges in Western Palaearctic subterranean freshwaters. Niphargus hrabei and N. valachicus, two morphologically and ecologically similar species, are atypical due to their epigean lifestyle and large ranges. Given their wide and largely sympatric distributions, we explored their potential ecological niche overlap by comparing morphological functional traits, patterns of co-occurrence and habitat selection, and tested for cryptic diversity by examining variation of mitochondrial and nuclear markers in selected populations. Mitochondrial sequences and the presence of shared nuclear alleles among individuals collected along the study area did not support the existence of cryptic lineages; two other Niphargus species described in the past from Hungary seem to be their younger synonyms. Except for body size, both species were morphologically similar and seem only weakly differentiated ecologically. Though smaller in size, N. hrabei is ecologically more tolerant, while N. valachicus might be a superior competitor due to its larger size. Despite large-scale sympatry, co-occurrence of both species was rare. Their present distributions may result from a dynamic relationship between dispersal ability and competitive strength. These widespread and euryoecious Niphargus species demonstrate that the genus may be successful not only in the subterranean realm but also in various surface waters

Groundwater is a hidden global keystone ecosystem

Groundwater is a vital ecosystem of the global water cycle, hosting unique biodiversity and providing essential services to societies. Despite being the largest unfrozen freshwater resource, in a period of depletion by extraction and pollution, groundwater environments have been repeatedly overlooked in global biodiversity conservation agendas. Disregarding the importance of groundwater as an ecosystem ignores its critical role in preserving surface biomes. To foster timely global conservation of groundwater, we propose elevating the concept of keystone species into the realm of ecosystems, claiming groundwater as a keystone ecosystem that influences the integrity of many dependent ecosystems. Our global analysis shows that over half of land surface areas (52.6%) has a medium‐to‐high interaction with groundwater, reaching up to 74.9% when deserts and high mountains are excluded. We postulate that the intrinsic transboundary features of groundwater are critical for shifting perspectives towards more holistic approaches in aquatic ecology and beyond. Furthermore, we propose eight key themes to develop a science‐policy integrated groundwater conservation agenda. Given ecosystems above and below the ground intersect at many levels, considering groundwater as an essential component of planetary health is pivotal to reduce biodiversity loss and buffer against climate change