Mastacembelid eels support Lake Tanganyika as an evolutionary hotspot of diversification

Background. Lake Tanganyika (LT) is the oldest of the African Rift Lakes and is one of the richest freshwater ecosystems on Earth, with high levels of faunal diversity and endemism. The endemic species flocks that occur in this lake, such as cichlid fishes, gastropods, catfish and crabs, provide unique comparative systems for the study of patterns and processes of speciation. Mastacembelid eels (Teleostei: Mastacembelidae) are a predominately riverine family of freshwater fish, occurring across Africa and Asia, but which also form a small species flock in LT. Methods. Including 25 species across Africa, plus Asian representatives as outgroups, we present the first molecular phylogenetic analysis for the group, focusing particularly on the evolutionary history and biodiversity of LT mastacembelid eels. A combined matrix of nuclear and mitochondrial genes based on 3118 bp are analysed implementing different phylogenetic methods, including Bayesian inference and maximum likelihood. Results. LT Mastacembelus are recovered as monophyletic, and analyses reveal the rapid diversification of five main LT lineages. Relaxed molecular clock dates provide age estimates for the LT flock at ∼7-8 Myr, indicating intralacustrine diversification, with further speciation events coinciding with periods of lower lake level. Our analyses also reveal as yet undescribed diversity of lacustrine and riverine species. A Southern-Eastern African clade, that is younger than the LT flock, is also recovered, while West African taxa are basal members of the African mastacembelid clade. Conclusions. That the LT species flock of mastacembelid eels appears to have colonised and immediately diversified soon after the formation of the lake, supports the view of LT as an evolutionary hotspot of diversification. We find evidence for biogeographic clades mirroring a similar pattern to other ichthyological faunas. In addition, our analyses also highlight a split of African and Asian mastacembelid eels at ∼19 Myr that is considerably younger than the split between their associated continents, suggesting a dispersal scenario for their current distribution. © 2010 Brown et al; licensee BioMed Central Ltd

Phylogenetic and biogeographic history of brook lampreys (Lampetra: Petromyzontidae) in the river basins of the Adriatic Sea based on DNA barcode data.

The Adriatic brook lamprey, Lampetra zanandreai Vladykov 1955, was described from northeastern Italy. Its distribution is thought to include left tributaries of the River Po and the river basins of the Adriatic Sea from the River Po to the River Isonzo/Soča in Italy, Switzerland and Slovenia. It also shows a geographically isolated distribution in the Potenza River on the Adriatic slope in Central Italy. Lampetra from the Neretva River system in Croatia and Bosnia and Herzegovina and the Morača River system in Montenegro that were previously identified as L. zanandreai were recently described as a new species Lampetra soljani Tutman, Freyhof, Dulčić, Glamuzina & Geiger 2017 based on morphological data and a genetic distance between the two species of roughly 2.5% in the DNA barcoding gene cytochrome oxidase I (COI). Since DNA barcodes for L. zanandreai are only available for one population from the upper Po River in northwestern Italy, we generated additional COI nucleotide sequence data of this species from Switzerland, northeastern and central Italy comprising near topotypic material and obtained GenBank sequences of the species from Slovenia to better assess the evolutionary history of the two brook lamprey species in the river basins of the Adriatic Sea. Our data show a low sequence divergence of <1% between L. zanandreai from Switzerland, northeastern and central Italy and Slovenia and the Balkan species L. soljani. However, members of the population previously identified as 'L. zanandreai' from northwest Italy are genetically highly divergent from those of L. zanandreai and likely belong to an undescribed species, L. sp. 'upper Po'. The presence of a unique and highly divergent brook lamprey lineage in the upper Po River suggests that L. zanandreai and Lampetra sp. 'upper Po' may have evolved in separate paleo drainages during the formation of the modern Po Valley subsequent to marine inundations in the Pliocene

Molecular phylogeny and phylogeography of ricefishes (Teleostei: Adrianichthyidae: Oryzias ) in Sri Lanka

Ricefishes of the genus Oryzias occur commonly in the fresh and brackish waters in coastal lowlands ranging from India across Southeast Asia and on to Japan. Among the three species of Oryzias recorded from peninsular India, two widespread species, O. carnaticus and O. dancena, have previously been reported from Sri Lanka based on museum specimens derived from a few scattered localities. However, members of the genus are widespread in the coastal lowlands of Sri Lanka, a continental island separated from India by the shallow Palk Strait. Although recent molecular phylogenies of Adrianichthyidae represent near-complete taxon representation, they lack samples from Sri Lanka. Here, based on sampling at 13 locations representative of the entire geographic and climatic regions of the island's coastal lowlands, we investigate for the first time the molecular phylogenetic relationships and phylogeography of Sri Lankan Oryzias based on one nuclear and two mitochondrial markers. Sri Lankan Oryzias comprise two distinct non-sister lineages within the javanicus species group. One of these is represented by samples exclusively from the northern parts of the island; it is recognized as O. dancena. This lineage is recovered as the sister group to the remaining species in the javanicus group. The second lineage represents a species that is widespread across the island's coastal lowlands. It is recovered as the sister group of O. javanicus and is identified as O. cf. carnaticus. Ancestral-range estimates suggest two independent colonizations of Indian subcontinent and Sri Lanka by widespread ancestral species of Oryzias during two discrete temporal windows: late Miocene and Plio-Pleistocene. No phylogeographic structure is apparent in Sri Lankan Oryzias, suggesting that there are no strong barriers to gene flow and dispersal along the coastal floodplains, as is the case also for other generalist freshwater fishes in the island

General principles for assignments of communities from eDNA : Open versus closed taxonomic databases

Metabarcoding of environmental DNA (eDNA) is a powerful tool for describing biodiversity, such as finding keystone species or detecting invasive species in environmental samples. Continuous improvements in the method and the advances in sequencing platforms over the last decade have meant this approach is now widely used in biodiversity sciences and biomonitoring. For its general use, the method hinges on a correct identification of taxa. However, past studies have shown how this crucially depends on important decisions during sampling, sample processing, and subsequent handling of sequencing data. With no clear consensus as to the best practice, particularly the latter has led to varied bioinformatic approaches and recommendations for data preparation and taxonomic identification. In this study, using a large freshwater fish eDNA sequence dataset, we compared the frequently used zero-radius Operational Taxonomic Unit (zOTU) approach of our raw reads and assigned it taxonomically (i) in combination with publicly available reference sequences (open databases) or (ii) with an OSU (Operational Sequence Units) database approach, using a curated database of reference sequences generated from specimen barcoding (closed database). We show both approaches gave comparable results for common species. However, the commonalities between the approaches decreased with read abundance and were thus less reliable and not comparable for rare species. The success of the zOTU approach depended on the suitability, rather than the size, of a reference database. Contrastingly, the OSU approach used reliable DNA sequences and thus often enabled species-level identifications, yet this resolution decreased with the recent phylogenetic age of the species. We show the need to include target group coverage, outgroups and full taxonomic annotation in reference databases to avoid misleading annotations that can occur when using short amplicon sizes as commonly used in eDNA metabarcoding studies. Finally, we make general suggestions to improve the construction and use of reference databases for metabarcoding studies in the future

General principles for assignments of communities from <scp>eDNA</scp> : Open versus closed taxonomic databases

Metabarcoding of environmental DNA (eDNA) is a powerful tool for describing biodiversity, such as finding keystone species or detecting invasive species in environmental samples. Continuous improvements in the method and the advances in sequencing platforms over the last decade have meant this approach is now widely used in biodiversity sciences and biomonitoring. For its general use, the method hinges on a correct identification of taxa. However, past studies have shown how this crucially depends on important decisions during sampling, sample processing, and subsequent handling of sequencing data. With no clear consensus as to the best practice, particularly the latter has led to varied bioinformatic approaches and recommendations for data preparation and taxonomic identification. In this study, using a large freshwater fish eDNA sequence dataset, we compared the frequently used zero-radius Operational Taxonomic Unit (zOTU) approach of our raw reads and assigned it taxonomically (i) in combination with publicly available reference sequences (open databases) or (ii) with an OSU (Operational Sequence Units) database approach, using a curated database of reference sequences generated from specimen barcoding (closed database). We show both approaches gave comparable results for common species. However, the commonalities between the approaches decreased with read abundance and were thus less reliable and not comparable for rare species. The success of the zOTU approach depended on the suitability, rather than the size, of a reference database. Contrastingly, the OSU approach used reliable DNA sequences and thus often enabled species-level identifications, yet this resolution decreased with the recent phylogenetic age of the species. We show the need to include target group coverage, outgroups and full taxonomic annotation in reference databases to avoid misleading annotations that can occur when using short amplicon sizes as commonly used in eDNA metabarcoding studies. Finally, we make general suggestions to improve the construction and use of reference databases for metabarcoding studies in the future

Multiple independent colonizations into the Congo Basin during the continental radiation of African Mastacembelus spiny eels

AIM: There has been recent interest in the origin and assembly of continental biotas based on densely sampled species-level clades, however, studies from African freshwaters are few so that the commonality of macroevolutionary patterns and processes among continental clades remain to be tested. Within the Afrotropics, the Congo Basin contains the highest diversity of riverine fishes, yet it is unclear how this fauna was assembled. To address this, and the diversification dynamics of a continental radiation, we focus on African Mastacembelus spiny eels. LOCATION: Afrotropical freshwaters. METHODS: The most complete molecular phylogeny to date was reconstructed for African spiny eels. Divergence times were estimated applying a Bayesian relaxed clock comparing fossil and geological calibrations across nuclear and mitochondrial trees. Biogeographic reconstructions, applying a dispersal–extinction–cladogenesis model and lineage diversification dynamics were examined. RESULTS: Spiny eels originated in Asia and colonized Africa c. 15.4 Ma (95% HPD: 23.9–8.8 Ma) from which their subsequent radiation across the Afrotropics was best fitted by a constant rate model. Ancestral state estimation identified multiple colonization events into the Congo Basin, whereas all other regions were likely to have been colonized once indicating considerable geographic constraints. Application of the fossil calibration gave similar age estimates across datasets, whereas a geological calibration estimated considerably older nuclear divergences. MAIN CONCLUSIONS: Despite profound environmental events during the evolutionary history of the group, there is no evidence for rapid lineage diversification. This finding supports several recent studies on tropical continental radiations that contrast to the common pattern of density-dependent diversification. We further show that dispersal has occurred into, as well as out of the Congo Basin, indicating the importance of this region in the generation of biodiversity

Paleo-Drainage Basin Connectivity Predicts Evolutionary Relationships across Three Southeast Asian Biodiversity Hotspots

Understanding factors driving diversity across biodiversity hotspots is critical for formulating conservation priorities in the face of ongoing and escalating environmental deterioration. While biodiversity hotspots encompass a small fraction of Earth's land surface, more than half the world's plants and two-thirds of terrestrial vertebrate species are endemic to these hotspots. Tropical Southeast (SE) Asia displays extraordinary species richness, encompassing four biodiversity hotspots, though disentangling multiple potential drivers of species richness is confounded by the region's dynamic geological and climatic history. Here, we use multilocus molecular genetic data from dense multispecies sampling of freshwater fishes across three biodiversity hotspots, to test the effect of Quaternary climate change and resulting drainage rearrangements on aquatic faunal diversification. While Cenozoic geological processes have clearly shaped evolutionary history in SE Asian halfbeak fishes, we show that paleo-drainage re-arrangements resulting from Quaternary climate change played a significant role in the spatiotemporal evolution of lowland aquatic taxa, and provide priorities for conservation efforts. [Freshwater; geology; halfbeak; island radiation; Miocene; Pleistocene; river; Southeast Asia.

Climate, immigration and speciation shape terrestrial and aquatic biodiversity in the European Alps

Quaternary climate fluctuations can affect speciation in regional biodiversity assembly in two non-mutually exclusive ways: a glacial species pump, where isolation in glacial refugia accelerates allopatric speciation, and adaptive radiation in underused adaptive zones during ice-free periods. We detected biogeographic and genetic signatures associated with both mechanisms in the assembly of the biota of the European Alps. Age distributions of endemic and widespread species within aquatic and terrestrial taxa (amphipods, fishes, amphibians, butterflies and flowering plants) revealed that endemic fish evolved only in lakes, are highly sympatric, and mainly of Holocene age, consistent with adaptive radiation. Endemic amphipods are ancient, suggesting preglacial radiation with limited range expansion and local Pleistocene survival, perhaps facilitated by a groundwater-dwelling lifestyle. Terrestrial endemics are mostly of Pleistocene age and are thus more consistent with the glacial species pump. The lack of evidence for Holocene adaptive radiation in the terrestrial biome is consistent with faster recolonization through range expansion of these taxa after glacial retreats. More stable and less seasonal ecological conditions in lakes during the Holocene may also have contributed to Holocene speciation in lakes. The high proportion of young, endemic species makes the Alpine biota vulnerable to climate change, but the mechanisms and consequences of species loss will likely differ between biomes because of their distinct evolutionary histories

Climate, immigration and speciation shape terrestrial and aquatic biodiversity in the European Alps.

Quaternary climate fluctuations can affect speciation in regional biodiversity assembly in two non-mutually exclusive ways: a glacial species pump, where isolation in glacial refugia accelerates allopatric speciation, and adaptive radiation in underused adaptive zones during ice-free periods. We detected biogeographic and genetic signatures associated with both mechanisms in the assembly of the biota of the European Alps. Age distributions of endemic and widespread species within aquatic and terrestrial taxa (amphipods, fishes, amphibians, butterflies and flowering plants) revealed that endemic fish evolved only in lakes, are highly sympatric, and mainly of Holocene age, consistent with adaptive radiation. Endemic amphipods are ancient, suggesting preglacial radiation with limited range expansion and local Pleistocene survival, perhaps facilitated by a groundwater-dwelling lifestyle. Terrestrial endemics are mostly of Pleistocene age and are thus more consistent with the glacial species pump. The lack of evidence for Holocene adaptive radiation in the terrestrial biome is consistent with faster recolonization through range expansion of these taxa after glacial retreats. More stable and less seasonal ecological conditions in lakes during the Holocene may also have contributed to Holocene speciation in lakes. The high proportion of young, endemic species makes the Alpine biota vulnerable to climate change, but the mechanisms and consequences of species loss will likely differ between biomes because of their distinct evolutionary histories

Disentangling the taxonomy of the subfamily Rasborinae (Cypriniformes, Danionidae) in Sundaland using DNA barcodes

Sundaland constitutes one of the largest and most threatened biodiversity hotspots; however, our understanding of its biodiversity is afflicted by knowledge gaps in taxonomy and distribution patterns. The subfamily Rasborinae is the most diversified group of freshwater fishes in Sundaland. Uncertainties in their taxonomy and systematics have constrained its use as a model in evolutionary studies. Here, we established a DNA barcode reference library of the Rasborinae in Sundaland to examine species boundaries and range distributions through DNA-based species delimitation methods. A checklist of the Rasborinae of Sundaland was compiled based on online catalogs and used to estimate the taxonomic coverage of the present study. We generated a total of 991 DNA barcodes from 189 sampling sites in Sundaland. Together with 106 previously published sequences, we subsequently assembled a reference library of 1097 sequences that covers 65 taxa, including 61 of the 79 known Rasborinae species of Sundaland. Our library indicates that Rasborinae species are defined by distinct molecular lineages that are captured by species delimitation methods. A large overlap between intraspecific and interspecific genetic distance is observed that can be explained by the large amounts of cryptic diversity as evidenced by the 166 Operational Taxonomic Units detected. Implications for the evolutionary dynamics of species diversification are discussed