Out of the ancient lake: Multiple riverine colonizations and diversification of the freshwater snails in the genus Semisulcospira around Lake Biwa.

Ancient lakes are a hotspot of biodiversity. Freshwater species often experience spectacular species radiation after colonizing lakes from riverine habitats. Therefore, the relationship between the fauna of the ancient lakes and the surrounding riverine system has a special significance in understanding their origin and evolutionary history. The study of ancient lake species often focused on the lake colonization of riverine species. In contrast, far less attention has been placed on the reverse direction: the riverine colonization of the lake species, despite its importance in disentangling their complex evolutionary history. The freshwater snails in the genus Semisulcospira involve endemic groups that radiated in the ancient Lake Biwa. Using genetics and fossil records, we inferred that the ancestors of these lake-endemic Semisulcospira snails historically colonized the riverine habitats at least three times during the Middle Pleistocene. Each colonization resulted in the formation of a new lineage that was genetically and morphologically distinct from other lineages. Further, one of these colonizations was followed by hybridization with a cosmopolitan riverine species, which potentially facilitated the population persistence of the colonizers in the new environment. Despite their complex histories, all these colonizers were currently grouped within a single species, Semisulcospira kurodai, suggesting cryptic diversity in this species. This study highlights the significance of the riverine colonizations of the lake species to fully understand the diversification history of freshwater fauna in and around the ancient lakes.

Deeply divergent freshwater fish species within a single river system in central Sulawesi.

Sulawesi is a biodiversity hotspot for ricefishes (Adrianichthyidae), with many species endemic to the central part of this island in single ancient lakes or lake systems. Frequent vicariance by lake fragmentation since the Pliocene may be largely responsible for diversification in this family. In this study, we demonstrate that not only lacustrine species but also riverine species in this area are also deeply divergent even within a single river system. A mitochondrial phylogeny revealed that a ricefish population newly discovered from Cerekang River is sister to Oryzias dopingdopingensis Mandagi, Mokodongan, Tanaka, & Yamahira, another riverine species endemic to Doping-doping River. However, the Cerekang Oryzias was genetically isolated from O. dopingdopingensis, despite that Cerekang River and Doping-doping River share a connection across estuarine waters. This separation was supported by phylogenomic trees and population genetic structure analyses based on genome-wide single nucleotide polymorphisms. Coalescent-based demographic inference demonstrated that the ancestral population of these two riverine ricefishes had experienced a substantial population decrease and subsequently diverged into two sub-populations. Because the Cerekang Oryzias was also morphologically distinguished from O. dopingdopingensis, we described it as a new species, O. landangiensis. We infer that O. landangiensis and O. dopingdopingensis are of lake-origin and are relic species which were left in these rivers after the lake disappeared, and that they have lost their dispersal ability when inhabiting the ancient lake. The lost dispersal ability possibly contributed to the formation of the biodiversity hotspot for this fish group on this island.

Resolving species-level diversity of Beringiana and Sinanodonta mussels (Bivalvia: Unionidae) in the Japanese archipelago using genome-wide data.

Accurate species identification is of primary importance in ecology and evolutionary biology. For a long time, the unionid mussels Beringiana and Sinanodonta have puzzled researchers trying to unravel their diversity because of their poorly discernible morphologies. A recent study conducted species delineation of unionid mussels based on mitochondrial DNA variation, opening up a new avenue to grasp species diversity of the mussels. However, mtDNA-based classification may not align with species boundaries because mtDNA is prone to introgression and incomplete lineage sorting that cause discordance between species affiliation and gene phylogeny. In this study, we evaluated the validity of the mtDNA-based classification of unionid mussels Beringiana and Sinanodonta in Japan using mitochondrial sequence data, double digest restriction site-associated DNA library (ddRAD) sequencing, and morphological data. We found significant inconsistencies in the mitochondrial and nuclear DNA phylogenies, casting doubt on the reliability of the mtDNA-based classification in this group. In addition, nuclear DNA phylogeny revealed that there are at least two unionid lineages hidden in the mtDNA phylogeny. Although molecular dating technique indicates that Beringiana and Sinanodonta diverged >35 million years ago, their shell morphologies are often indistinguishable. Specifically, morphological analyses exhibited the parallel appearance of nearly identical ball-like shell forms in the two genera in Lake Biwa, which further complicates species identification and the morphological evolution of unionid mussels. Our study adds to a growing body of literature that accurate species identification of unionid mussels is difficult when using morphological characters alone. Although mtDNA-based classification is a simple and convenient way to classify unionid mussels, considerable caution is warranted for its application in ecological and evolutionary studies.

Profiling metal contamination from ultramafic sediments to biota along the Albanian shoreline of Lake Ohrid (Albania/Macedonia).

Ultramafic sediments exhibit high levels of geogenically-derived and potentially toxic metals, with Ni, Cr and Co often exceeding benchmark values. As yet, a holistic understanding of the bioavailability, mobility, potential ecotoxicity and trophic transfer of trace elements in both benthic and pelagic food chains in aquatic ultramafic environments (UME) is lacking. We investigated potential environmental health issues due to metal contamination by jointly implementing chemical, ecological and toxicological tools, along the Albanian shoreline of Lake Ohrid. It is an aquatic system of worldwide importance, representative of temperate UME with a legacy of Ni and Cr contamination from mining activities. Levels of Ni, Cr, Cd, Cu, Co, Fe, Mn and Zn were determined in waters, sediments and native biota. The potential environmental mobility of sediment-bound elements was further assessed using CaCl2, EDTA and acetic acid extractions. Whole-sediment ecotoxicity tests were also carried out using ostracods and chironomids, according to standardized procedures. Despite Ni and Cr concentrations above the sediment quality guidelines for probable effect levels, we did not observe ecotoxic effects in laboratory tests. However, these elements were bioavailable to native organisms under field conditions, especially to benthic primary producers in direct contact with sediments (up to 139 mg Cr kg-1 and 785 mg Ni kg-1). Although biomagnification was not observed, further investigations of metal translocation, metabolism and elemental trophic transfer along benthic food webs appears to be a general research priority in the management of temperate UME. The present study shows that proper management of temperate UME requires not only the integration of data from different lines of evidence, but also laboratory vs. field approaches to understand the subtler, long-term effects of increased elemental body burdens in native organisms.

Effects of Long-Term Anthropogenic Disturbance on the Benthic Episammic Diatom Community of an Ancient, Tropical Lake.

Habitat homogenization, nutrient enrichment and loss of biodiversity are broadly recognized as the consequences of human activity in aquatic systems. Diatoms (Bacillariophyceae) are frequently used in aquatic environmental assessment and impact monitoring, but in unique habitats dominated by endemic taxa, traditional approaches may not be appropriate. We examined the impacts of long term anthropogenic impacts upon the littoral episammic diatom community around the town of Soroako, located on Lake Matano, an ancient tropical lake. Lake Matano is located on central Sulawesi Island, Indonesia, and socio-economic conditions are typical of developing nations. Although differences in nutrient concentrations were undetectable with field-based spectroscopy approaches, mean Shannon diversity was decreased in association with proximity the town-site. However, mean ß-diversity was maintained despite several decades of shoreline modification at Soroako. Elevated abundances of early-successional diatom taxa in the disturbed area drove differences between areas immediately offshore of Soroako and those farther away. These findings suggest that increased physical disturbance and TSS loads around Soroako, rather than increased nutrient loading, influenced shifts in the diatom community. These results suggest that microscopy-based biomonitoring approaches are sensitive indicators of environmental modification that could be useful in areas where access to cutting-edge analytical equipment is limited.

Genomic architecture of habitat-related divergence and signature of directional selection in the body shapes of Gnathopogon fishes.

Evolution of ecomorphologically relevant traits such as body shapes is important to colonize and persist in a novel environment. Habitat-related adaptive divergence of these traits is therefore common among animals. We studied the genomic architecture of habitat-related divergence in the body shape of Gnathopogon fishes, a novel example of lake-stream ecomorphological divergence, and tested for the action of directional selection on body shape differentiation. Compared to stream-dwelling Gnathopogon elongatus, the sister species Gnathopogon caerulescens, exclusively inhabiting a large ancient lake, had an elongated body, increased proportion of the caudal region and small head, which would be advantageous in the limnetic environment. Using an F2 interspecific cross between the two Gnathopogon species (195 individuals), quantitative trait locus (QTL) analysis with geometric morphometric quantification of body shape and restriction-site associated DNA sequencing-derived markers (1622 loci) identified 26 significant QTLs associated with the interspecific differences of body shape-related traits. These QTLs had small to moderate effects, supporting polygenic inheritance of the body shape-related traits. Each QTL was mostly located on different genomic regions, while colocalized QTLs were detected for some ecomorphologically relevant traits that are proxy of body and caudal peduncle depths, suggesting different degree of modularity among traits. The directions of the body shape QTLs were mostly consistent with the interspecific difference, and QTL sign test suggested a genetic signature of directional selection in the body shape divergence. Thus, we successfully elucidated the genomic architecture underlying the adaptive changes of the quantitative and complex morphological trait in a novel system.

A RAD-based phylogenetics for Orestias fishes from Lake Titicaca.

The fish genus Orestias is endemic to the Andes highlands, and Lake Titicaca is the centre of the species diversity of the genus. Previous phylogenetic studies based on a single locus of mitochondrial and nuclear DNA strongly support the monophyly of a group composed of many of species endemic to the Lake Titicaca basin (the Lake Titicaca radiation), but the relationships among the species in the radiation remain unclear. Recently, restriction site-associated DNA (RAD) sequencing, which can produce a vast number of short sequences from various loci of nuclear DNA, has emerged as a useful way to resolve complex phylogenetic problems. To propose a new phylogenetic hypothesis of Orestias fishes of the Lake Titicaca radiation, we conducted a cluster analysis based on morphological similarities among fish samples and a molecular phylogenetic analysis based on RAD sequencing. From a morphological cluster analysis, we recognised four species groups in the radiation, and three of the four groups were resolved as monophyletic groups in maximum-likelihood trees based on RAD sequencing data. The other morphology-based group was not resolved as a monophyletic group in molecular phylogenies, and some members of the group were diverged from its sister group close to the root of the Lake Titicaca radiation. The evolution of these fishes is discussed from the phylogenetic relationships.

Can fractal dimensions objectivize gastropod shell morphometrics? A case study from Lake Lugu (SW China).

Morphometrics are fundamental for the analysis of size and shape in fossils, particularly because soft parts or DNA are rarely preserved and hard parts such as shells are commonly the only source of information. Geometric morphometrics, that is, landmark analysis, is well established for the description of shape but it exhibits a couple of shortcomings resulting from subjective choices during landmarking (number and position of landmarks) and from difficulties in resolving shape at the level of micro-sculpture.With the aid of high-resolution 3D scanning technology and analyses of fractal dimensions, we test whether such shortcomings of linear and landmark morphometrics can be overcome. As a model group, we selected a clade of modern viviparid gastropods from Lake Lugu, with shells that show a high degree of sculptural variation. Linear and landmark analyses were applied to the same shells in order to establish the fractal dimensions. The genetic diversity of the gastropod clade was assessed.The genetic results suggest that the gastropod clade represents a single species. The results of all morphometric methods applied are in line with the genetic results, which is that no specific morphotype could be delimited. Apart from this overall agreement, landmark and fractal dimension analyses do not correspond to each other but represent data sets with different information. Generally, the fractal dimension values quantify the roughness of the shell surface, the resolution of the 3D scans determining the level. In our approach, we captured the micro-sculpture but not the first-order sculptural elements, which explains that fractal dimension and landmark data are not in phase.We can show that analyzing fractal dimensions of gastropod shells opens a window to more detailed information that can be considered in evolutionary and ecological contexts. We propose that using low-resolution 3D scans may successfully substitute landmark analyses because it overcomes the subjective landmarking. Analyses of 3D scans with higher resolution than used in this study will provide surface roughness information at the mineralogical level. We suggest that fractal dimension analyses of a combination of differently resolved 3D models will significantly improve the quality of shell morphometrics.

Inventory of the benthic eukaryotic diversity in the oldest European lake.

We have profound knowledge on biodiversity on Earth including plants and animals. In the recent decade, we have also increased our understanding on microorganisms in different hosts and the environment. However, biodiversity is not equally well studied among different biodiversity groups and Earth's systems with eukaryotes in freshwater sediments being among the least known. In this study, we used high-throughput sequencing of the 18S rRNA gene to investigate the entire diversity of benthic eukaryotes in three distinct habitats (littoral sediment and hard substrate, profundal sediment) of Lake Ohrid, the oldest European lake. Eukaryotic sequences were dominated by annelid and arthropod animals (54% of all eukaryotic reads) and protists (Ochrophyta and Ciliophora; together 40% of all reads). Eukaryotic diversity was 15% higher in the deep profundal than on either near-surface hard substrates or littoral sediments. The three habitats differed in their taxonomic and functional community composition. Specifically, heterotrophic organisms accounted for 92% of the reads in the profundal, whereas phototrophs accounted for 43% on the littoral hard substrate. The profundal community was the most homogeneous, and its network was the most complex, suggesting its highest stability among the sampled habitats.

Encyonopsis indonesica sp. nov. (Bacillariophyceae, Cymbellales), a new diatom from the ancient lake Matano (Sulawesi, Indonesia).

A new species, Encyonopsis indonesica, is described from the ancient lake Matano, Sulawesi island, Indonesia. The morphology of this species was studied by means of light and scanning electron microscopy. E. indonesica has a remarkable valve ultrastructure. The valve surface is ornamented with numerous longitudinal siliceous ribs and siliceous verrucae. Valve face delineated from the mantle by a thickened marginal ridge. Raised sterna border the raphe branches. Raphe is distinctly undulate with distal ends hooked strongly to the ventral side. The only similar species to E. indonesica is Amphora dissimilis described from New Caledonia. Comparison of both taxa is given and A. dissimilis is transferred to Encyonopsis. The taxonomic placement of both taxa is evaluated, and the phenomenon of external siliceous ornamentation is discussed.

Lake Poso's shrimp fauna revisited: the description of five new species of the genus Caridina (Crustacea, Decapoda, Atyidae) more than doubles the number of endemic lacustrine species.

Lake Poso, an ancient lake system on the Indonesian island Sulawesi, harbours an endemic species flock of six, four lacustrine and two riverine species of the freshwater shrimp genus Caridina. In this study, five new lacustrine species are described, bringing the total to eleven species altogether. The number of lacustrine species is more than doubled to nine species compared to the last taxonomic revision in 2009. One of them, Caridina mayamareenae Klotz, Wowor & von Rintelen, sp. nov., even represents the first case of an atyid shrimp associated with freshwater snails which is morphologically adapted to living in shells. An integrative approach was used by providing a combination of morphological, ecological, and molecular data. Based on standard morphological characters, distribution, substrate preferences, and colouration of living specimens in the field, five distinct undescribed species could be distinguished. To support our species-hypothesis based on the mitochondrial genes 16S and COI, a molecular phylogeny was used for all eleven species from Lake Poso. All species form a well-supported monophyletic group, but only four morphospecies consistently correspond to mtDNA clades - a possible reason could be introgressive hybridisation, incomplete lineage sorting, or not yet fixed species boundaries. These results are discussed further in the context of adaptive radiation, which turned out to be more diverse than previously described. Finally, yet importantly, subjecting all new species to similar threats and to the same IUCN category and criterion than the previously described species from the lake is recommended.

Ancient drainage networks mediated a large-scale genetic introgression in the East Asian freshwater snails.

Biogeography and genetic variation of freshwater organisms are influenced not only by current freshwater connections but also by past drainage networks. The Seto Inland Sea is a shallow enclosed sea in Japan, but geological evidence showed that a large freshwater drainage had intermittently appeared in this area between the late Pliocene and Pleistocene. Here, we demonstrated that this paleodrainage greatly affected the genetic variation of the East Asian freshwater snails, Semisulcospira spp. We found that the mtDNA haplotypes originated in the Lake Biwa endemic Semisulcospira species at the upstream side of the paleodrainage were frequently observed in the riverine Semisulcospira species at its downstream side. The genome-wide DNA and morphological analyses consistently showed that there was no clear evidence of nuclear introgression between the Lake Biwa endemics and riverine species. These results suggest that the large paleodrainage had facilitated mitochondrial introgression and had broadly spread the introgressed mtDNA haplotypes to its downstream region around the Seto Inland Sea. Our study highlights the role of paleodrainages in shaping the genetic variation of freshwater organisms.

New insights from museum specimens: a case of Viviparidae (Caenogastropoda: Mollusca) in Iwakawa's collection preserved in the National Museum of Nature and Science, Tokyo.

BACKGROUND: In this study, we clarify the classification of museum specimens of the family Viviparidae, which is composed of six species/subspecies in Japan, including three endangered species. We examined Viviparus sclateri specimens from the Tomotaro Iwakawa collection (1855-1933) in the National Museum of Nature and Science, Tokyo. The Iwakawa's collection was catalogued in 1919 and Viviparus sclateri, labelled with Naga-tanishi, the current Japanese name for H. longispira, which was, at the time, equivalent to Viviparus sclateri, was listed in this catalogue. The catalogue noted localities of Viviparus sclateri (Naga-tanishi) from outside Lake Biwa, including occurrences in Lake Kasumigaura and Lake Suwa. However, Heterogen longispira (Naga-tanishi) is currently considered to be endemic to Lake Biwa drainage. The actual status of Viviparus sclateri in Iwakawa (1919) has not been clarified until now. NEW INFORMATION: Our examination revealed that Viviparus sclateri from Iwakawa's catalogue included H. japonica, H. longispira and Sinotaia quadrata histrica, based on current taxonomy. Specimens assigned to H. longispira occurred only in Lake Biwa drainage. Heterogen japonica was confirmed to be present in all lots and some H. japonica from Lake Suwa had a distinctive morphology. Sinotaia quadrata histrica was only confirmed to occur in Lake Suwa. Furthermore, some specimens from southern Lake Biwa and the Seta River had intermediate characteristics between H. japonica and H. longispira and their populations are currently almost extinct.

Phylogeny and historical demography of endemic fishes in Lake Biwa: the ancient lake as a promoter of evolution and diversification of freshwater fishes in western Japan.

To elucidate the origins of the endemic fish of Lake Biwa, an ancient lake in Japan, and the role of the lake in the diversification of freshwater fish in western Japan, we established a molecular phylogenetic framework with an absolute time scale and inferred the historical demography of a large set of fish species in and around the lake. We used mtDNA sequences obtained from a total of 190 specimens, including 11 endemic species of Lake Biwa and their related species, for phylogenetic analyses with divergence time estimations and from a total of 2319 specimens of 42 species (including 14 endemics) occurring in the lake for population genetic analyses. Phylogenetic analysis suggested that some of the endemic species diverged from their closest relatives earlier (1.3-13.0 Ma) than the period in which the present environmental characteristics of the lake started to develop (ca. 0.4 Ma), whereas others diverged more recently (after 0.4 Ma). In contrast, historical demographic parameters suggested that almost all species, including endemic and nonendemic ones, expanded their populations after the development of the present lake environment. In phylogeographic analyses, common or very close haplotypes of some species were obtained from Lake Biwa and other regions of western Japan. The phylogenetic and historical demographic evidence suggests that there was a time lag between phylogenetic divergence and population establishment and that phenotypic adaptation of some endemic species to the limnetic environment occurred much later than the divergences of those endemic lineages. Population structure and phylogeographic patterns suggest that Lake Biwa has functioned not only as the center of adaptive evolution but also as a reservoir for fish diversity in western Japan.

A new freshwater snail genus (Hydrobiidae, Gastropoda) from Montenegro, with a discussion on gastropod diversity and endemism in Skadar Lake.

Karucia sublacustrina a new species of freshwater snails (Hydrobiidae, Gastropoda) is described based on material collected from Skadar Lake (Montenegro, Albania). The new species belongs to monotypic genus Karucia gen. n. The shell morphology and body shape of the new genus resembles Radomaniola Szarowska, 2006 and Grossuana Radoman, 1973, from which it differs in the larger shells with relatively slim and a slightly, but clearly shouldered body whorl. The number of gastropods from Skadar Lake basin tallies now 50 species. The adjusted rate of gastropod endemicity for Skadar Lake basin is estimated to be 38%. By compiling faunal and taxonomic data we also aim to provide information of relevance as to conservation efforts.

Integrative taxonomy of a new species of planarian from the Lake Ohrid basin, including an analysis of biogeographical patterns in freshwater triclads from the Ohrid region (Platyhelminthes, Tricladida, Dugesiidae).

A new species of the genus Dugesia is described from the Lake Ohrid region in the western part of the Balkan Peninsula, forming the first fully documented species description for this genus in the Ohrid area. The morphological species delimitation is supported by complementary molecular, karyological, and cytogenetic data available from the literature. Therefore, species delineation is based on a truly integrative approach. Further, a short account on the degree of freshwater planarian endemicity in the Ohrid region is provided.