Renewed sampling of inland aquatic habitats in southern Africa yields two novel freshwater crab species (Decapoda: Potamonautidae: Potamonautes)

A recent survey of the freshwater streams of the Mecula and Yao mountains in the Niassa province of Mozambique resulted in the discovery of a new freshwater crab species. This species is genetically and morphologically distinct from described species from Mozambique or its neighbouring countries, and is described as P otamonautes bellarussus sp. nov. In addition, a new semi‐terrestrial burrowing freshwater crab P otamonautes flavusjo sp. nov. from the Highveld of the Mpumalanga province in South Africa is described based on unique genetic and morphological characters. The phylogenetic affinities of the two new species in relation to the described eastern and southern African Potamonautes species is determined and the biogeographic implications are discussed. © 2014 The Linnean Society of London     

Phylogeographic patterning in a freshwater crab species (Decapoda: Potamonautidae: Potamonautes) reveals the signature of historical climatic oscillations

Aim The phylogeographic relationships among populations of the common Cape River crab, Potamonautes perlatus, are examined to investigate whether the contemporary population genetic structure is congruent with the hypothesized hydrographic evolution of drainage systems established during the Pliocene, or whether it reflects an older Miocene climatic amelioration. Location 139 samples of P. perlatus were collected from 31 populations distributed among the five major perennial drainage systems and a number of smaller catchments in the Western and Eastern Cape, South Africa. Methods Phylogeographic analysis using parsimony, maximum likelihood, minimum evolution and Bayesian inferences was employed for the 16S rRNA mtDNA gene region, while bootstrapping and posterior probabilities were used to assess the robustness of clades. In addition, nested clade analysis was performed in an attempt to disentangle the contemporary and historical factors that have sculpted genealogical relationships among conspecific populations of P. perlatus. Results Phylogenetic topologies were congruent irrespective of the evolutionary method employed. Two highly distinct reciprocally monophyletic clades characterized by marked levels of corrected sequence divergence were present, with no shared haplotypes between the two major phylogroups. Phylogroup one comprises the populations of the westward‐flowing drainages (mainly the Berg and Olifants drainages), and phylogroup two comprises all of the southward‐flowing drainages and can further be divided into two subclades – one containing the Breede River populations, and the other containing the Gamtoos and Gourits drainage systems. The nested clade analysis demonstrated restricted gene flow and long‐distance dispersal for a number of higher clade levels. The higher‐level groups and results for the total cladogram suggest either fragmentation or isolation by distance. Main conclusions Freshwater crabs are generally highly philopatric, and dispersal, although not common, has occurred historically. The westward‐flowing drainages (Berg, Olifants, Eerste, Liesbeeck and Tokai) are isolated from the southward‐flowing drainages by the Cape Fold Mountains, while the southward‐flowing drainages have a number of tributaries that extend into the low‐lying regions, allowing for gene flow between these three major drainages systems (Breede, Gamtoos and Gourits). Among the westward‐flowing drainages, a more intensive sampling regime is required to understand evolutionary relationships. Our molecular results suggest that the observed patterns pre‐date the formation of contemporary hydrographic patterns in the Cape. This suggests that an older Late Miocene event has severely impacted the contemporary population structure in this species, as recent Pliocene hydrographic boundaries do not correspond to the phylogeographic pattern observed. Conservation efforts for aquatic taxa should clearly be directed at the catchments, in an attempt to conserve biological diversity.     

Neglected refugia of biodiversity: mountainous regions in Mozambique and Malawi yield two novel freshwater crab species (Potamonautidae: Potamonautes)

Phylogenetic relationships amongst the southern African freshwater crab fauna are reinvestigated following the recent collection of morphologically distinct Potamonautes specimens from remote mountainous regions in Malawi and Mozambique. Specimens were subjected to DNA sequencing of three mtDNA loci, cytochrome c oxidase subunit I (COI), 12S rRNA, and 16S rRNA and compared to the 14 described species from the region. Phylogenetic analysis using maximum parsimony and Bayesian inference revealed the presence of two novel evolutionary lineages. The phylogeny demonstrates that Potamonautes obesus (A. Milne‐Edwards, 1868) is sister to a morphologically distinct novel species from Mount Namuli in Mozambique. Two sympatric and genetically distinct species from Mount Mulanje, in Malawi (forms A and B) were recognized. Form B is sister to the large‐bodied South African riverine freshwater crabs and represents a novel lineage whereas the remaining species (form A) from Mulanje, in Malawi was sister to samples from Mounts Inago and Mabu, and in Mozambique was identified as Potamonautes choloensis (Chace, 1953). The two novel evolutionary lineages were genetically distinct and morphologically different from the described species in each of the respective regions. Two new freshwater crab species Potamonautes namuliensis sp. nov. and Potamonautes mulanjeensis sp. nov. , are described in the present study. The samples from Mount Mulanje in Malawi, and Mounts Mabu and Inago in Mozambique represent new distribution records for Potamonautes choloensis. © 2012 The Linnean Society of London, Zoological Journal of the Linnean Society, 2012, 164 , 498–509.     

Phylogeography of Rhinichthys cataractae (Teleostei: Cyprinidae): pre‐glacial colonization across the Continental Divide and Pleistocene diversification within the Rio Grande drainage

The longnose dace, Rhinichthys cataractae, is a primary freshwater fish inhabiting riffle habitats in small headwater rivers and streams across the North American continent, including drainages east and west of the Continental Divide. The mitochondrially encoded cytochrome b gene (1140 bp) and 2298–2346 bp of the nuclear‐encoded genes S7 and RAG1 were obtained from 87 individuals of R. cataractae (collected from 17 sites throughout its range) and from several close relatives. Phylogenetic analyses recovered a monophyletic R. cataractae species‐group that contained Rhinichthys evermanni, Rhinichthys sp. ‘Millicoma dace’, and a non‐exclusive R. cataractae. Within the R. cataractae species‐group, two well‐supported lineages were identified, including a western lineage (containing R. evermanni, R. sp. ‘Millicoma dace’ and individuals of R. cataractae from Pacific slope drainages) and an eastern lineage (containing individuals of R. cataractae from Arctic, Atlantic, and Gulf slope drainages). Within the eastern lineage of R. cataractae, two well‐supported groups were recovered: a south‐eastern group, containing individuals from the Atlantic slope, southern tributaries to the Mississippi River, and the Rio Grande drainage; and a north‐eastern group, containing individuals from the Arctic slope and northern tributaries to the Mississippi River. Estimates of the timing of divergence within the R. cataractae species‐group, combined with ancestral area‐reconstruction methods, indicate a separation between the eastern and western lineages during the Pliocene to early‐Pleistocene, with a direction of colonization from the west of the Continental Divide eastward. Within the southern portion of its range, R. cataractae likely entered the Rio Grande drainage during the Pleistocene via stream capture events between the Arkansas River (Mississippi River drainage) and headwaters of the Rio Grande. A close relationship between populations of R. cataractae in the Rio Grande drainage and the adjacent Canadian River (Mississippi River drainage) is consistent with hypothesized stream capture events between the Pecos (Rio Grande drainage) and Canadian rivers during the late‐Pleistocene. The population of R. cataractae in the lower Rio Grande may have become separated from other populations in the Rio Grande drainage (upper Rio Grande and Pecos River) and Canadian River during the late‐Pleistocene, well before initiation of recent and significant anthropogenic disturbance within the Rio Grande drainage. © 2014 The Linnean Society of London, Biological Journal of the Linnean Society, 2014, 111 , 317–333.     

SOME ASPECTS OF THE ECOLOGY OF THE FRESHWATER CRAB (POTAMONAUTES PERLATUS MILNE EDWARDS) IN THE UPPER REACHES OF THE BUFFALO RIVER,EASTERN CAPE PROVINCE,SOUTH AFRICA.

Summary

Freshwater crabs (Potamonautes perlatus) are the largest naturally occurring invertebrates in southern African rivers. The ecology of these animals in South African riverine ecosystems is little understood. This study investigates some aspects of the population and feeding ecology of P. perlatus in the upper reaches of the Buffalo River. The densities of crabs (carapace width larger than 25 mm) at two sites in the river ranged between 1.72 and 5.25 crabs m², higher than previously recorded for freshwater crabs in rivers in southern Africa and southern Italy. Twenty-four hour observations revealed that P. perlatus has a nocturnal habit. The diet of P. perlatus was established by examination of the contents of the fore-gut. Crabs with a carapace width smaller than 40 mm fed predominantly on aquatic invertebrates while larger crabs fed on allochthanous material (leaf litter), detritus and algae. Through feeding, crabs reduced the surface area of leaves in the stream by 99.95%.     

Evolutionary insight into the Peripatopsis balfouri sensu lato species complex (Onychophora: Peripatopsidae) reveals novel lineages and zoogeographic patterning

Evolutionary relationships in the widely distributed velvet worm Peripatopsis balfouri sensu lato species complex were examined using DNA sequence data, gross and SEM morphology. Sequence data were generated for the COI mtDNA and the 18S rRNA loci and analysed using a Bayesian inference approach, maximum likelihood and maximum parsimony. Phylogenetic analyses of the combined DNA sequence data revealed that Peripatopsis clavigera specimens from the southern Cape (clade 1) was sister to P. balfouri sensu lato specimens from the Cederberg Mountains (clade 2). Within the main P. balfouri sensu lato species complex, three addition clades could be discerned (clades 3, 4 and 5). The obligatory troglobitic species Peripatopsis alba was equidistant between the Cape Peninsula and adjacent interior (clade 3) and the two Boland and Hottentots Holland Mountains (clades 4 and 5). On the Cape Peninsula, P. stelliporata specimens nested among the sympatric P. balfouri sensu lato specimens. The Cape Peninsula specimens were sister to specimens from Jonkershoek site 1, Kogelberg and Simonsberg. Two Boland clades were retrieved, comprising Du Toit's Kloof, Bain's Kloof and Mitchell's Pass and sister (in clade 4) to specimens from the Boland and adjacent Hottentots Holland Mountains in clade 5. These results revealed complex biogeographic patterning in the P. balfouri sensu lato species complex. The presence of sympatric, yet genetically discrete species pairs at six of the sample localities (Du Toit's Kloof, Simonsberg, Jonkershoek sites 1 and 2, Kogelberg and Landroskop) suggests that there is reproductive isolation between the lineages. Divergence time estimations suggest a Miocene/Pliocene/Pleistocene cladogenesis. A taxonomic revision of the P. balfouri sensu lato species complex was undertaken to stabilize the taxonomy. P. clavigera is monophyletic and retained for the southern Cape specimens, P. balfouri sensu stricto is now confined to the Cape Peninsula and adjacent interior, while P. stelliporata is regarded as a junior synonym of the latter taxon, P. alba is endemic to the Wynberg Cave systems on the Cape Peninsula. Three novel species (Peripatopsis cederbergiensis, sp. n., Peripatopsis bolandi sp. n. and Peripatopsis purpureus, sp. n.) are described.     

Ecological Assessment of Two Species of Potamonautid Freshwater Crabs from the Eastern Highlands of Zimbabwe,with Implications for Their Conservation

The spatial ecology of freshwater crabs and their conservation status is largely understudied in Africa. An ecological assessment was conducted at 104 localities in 51 rivers and/or streams in the Eastern Highlands of Zimbabwe whereby the distribution and abundances of freshwater crab species were mapped and the possible drivers of the observed trends in population structure explored. In addition, information on crab utilisation as a food resource by local communities was assessed via face to face interviews across the region. Finally, the conservation status of each species was assessed using the IUCN Red List criteria. Only two crab species Potamonautes mutareensis and Potamonautes unispinus were recorded within the region of study. Potamonautes mutareensis was largely restricted to less impacted environments in the high mountainous river system, whereas P. unispinus was found in low laying areas. In stretches of river where both species were found to co-occur, the species were never sampled from the same site, with P. mutareensis occurring in shallower, faster flowing environments and P. unispinus in deeper, slow flowing sites. Interview results revealed that the local communities, particularly in the southern part of the Eastern Highlands around the Chipinge area, had a considerable level of utilisation (55% of households) on the harvesting of crabs for household consumption during the non-agricultural season (May to September). Results from the IUCN Red List assessment indicate that both species should be considered as “Least Concern”. Threats to freshwater crabs in the Eastern Highlands, however, include widespread anthropogenic impacts such as habitat destruction associated with gold and diamond mining, inorganic and organic pollution and possibly exploitation for human consumption. The current study provides important information and insight towards the possible development of a freshwater crab conservation action plan within the region.     

Genetic variation of European grayling (Thymallus thymallus) populations in the Western Balkans

In order to elucidate genetic composition of European grayling (Thymallus thymallus) populations in the Western Balkans, the partial mitochondrial DNA (mtDNA) control region was sequenced and 12 microsatellite loci genotyped in 14 populations originating from tributaries of the Adriatic and Danube drainages. Eleven mtDNA haplotypes were found, one confined to the Adriatic clade, one to the Alpine group and the rest to the ‘Balkan’ grayling phylogenetic clade. Haplotypes from the Balkan clade were confined to the Danube drainage and constituted two groups: northern group with haplotypes found in the Slovenian part of the Danube drainage, and southern group, consisting from Bosnia–Herzegovina and Montenegro. Substantial genetic distance between northern and southern groups of haplotypes (0.75–1.8%) and well supported divisions within the northern group indicate very structured grayling population within the studied Danube basin that most probably did not evolve due to vicariance but rather as a consequence of multiple colonization waves that might have occurred during the Pleistocene. Furthermore, genetic distance of ~4% between Adriatic and Danube populations’ haplotypes, suggest that their separation occurred in mid-Pliocene. These findings imply a complex colonization pattern of the Western Balkans drainages. Microsatellite data also confirm high genetic diversity in Western Balkans populations of grayling (on average 7.5 alleles per microsatellite locus and H _exp 0.58). Limited stocking activities were detected based on microsatellites and mtDNA data. Regarding current knowledge of grayling phylogeography appropriate management strategies were proposed to preserve unique, autochthonous grayling populations in Western Balkan.     

Comparative phylogeography of livebearing fishes in the genera Poeciliopsis and Poecilia (Poeciliidae: Cyprinodontiformes) in central Mexico

Aim To test the hypothesis that the vicariant event responsible for north–south divergences in two clades of the fish genus Poeciliopsis Regan was also responsible for north–south divergences in the fish Poecilia butleri Jordan. Location Central Mexico. Methods Parsimony, distance, maximum likelihood and Bayesian phylogenetic analyses of a mitochondrial gene. Molecular clock test and Bayesian analyses of divergence time. Results We report concordant phylogeographical patterns between two clades in the genus Poeciliopsis (i.e. the other formed by P. latidens Garman and P. fasciata Meek, and the other formed by P. presidionis Jordan and P. turneri Miller) and the clade of Poecilia butleri, with northern and southern individuals within each clade grouping into separate lineages. There is also evidence for slower substitution rates in Poecilia compared with Poeciliopsis. After taking into account these substitution rate discrepancies with Bayesian relaxed molecular clock analyses, north–south divergences in Poecilia butleri were equivalent to those reported for Poeciliopsis latidens‐fasciata and P. presidionis‐turneri. Main conclusions The same Plio‐Pleistocene vicariant event associated with geological activity of the Trans‐Mexican Volcanic Belt (TMVB) appears to have caused divergence in three different freshwater fish lineages. This study is an example of how comparative phylogeography can strengthen inferences about vicariant events in regions of high biological diversity and complex geological history such as the TMVB.     

Population genetics of the freshwater mussel, <Emphasis Type="Italic">Amblema plicata</Emphasis> (Say 1817) (Bivalvia: Unionidae): Evidence of high dispersal and post-glacial colonization

Over 70% of North American freshwater mussel species (families Unionidae and Margaritiferidae) are listed as threatened or endangered. Knowledge of the genetic structure of target species is essential for the development of effective conservation plans. Because Ambelma plicata is a common species, its population genetic structure is likely to be relatively intact, making it a logical model species for investigations of freshwater mussel population genetics. Using mtDNA and allozymes, we determined the genotypes of 170+ individuals in each of three distinct drainages: Lake Erie, Ohio River, and the Lower Mississippi River. Overall, within-population variation increased significantly from north to south, with unique haplotypes and allele frequencies in the Kiamichi River (Lower Mississippi River drainage). Genetic diversity was relatively low in the Strawberry River (Lower Mississippi River drainage), and in the Lake Erie drainage. We calculated significant among-population structure using both molecular markers (A.p. Φ_st = 0.15, θ_st = 0.12). Using a hierarchical approach, we found low genetic structure among rivers and drainages separated by large geographic distances, indicating high effective population size and/or highly vagile fish hosts for this species. Genetic structure in the Lake Erie drainage was similar to that in the Ohio River, and indicates that northern populations were founded from at least two glacial refugia following the Pleistocene. Conservation of genetic diversity in Amblema plicata and other mussel species with similar genetic structure should focus on protection of a number of individual populations, especially those in southern rivers.     

Phylogeography and taxonomic status of trout and salmon from the Ponto‐Caspian drainages,with inferences on European Brown Trout evolution and taxonomy

Current taxonomy of western Eurasian trout leaves a number of questions open; it is not clear to what extent some species are distinct genetically and morphologically. The purpose of this paper was to explore phylogeography and species boundaries in freshwater and anadromous trout from the drainages of the Black and the Caspian Seas (Ponto‐Caspian). We studied morphology and mitochondrial phylogeny, combining samples from the western Caucasus within the potential range of five nominal species of trout that are thought to inhabit this region, and using the sequences available from GenBank. Our results suggest that the genetic diversity of trout in the Ponto‐Caspian region is best explained with the fragmentation of catchments. (1) All trout species from Ponto‐Caspian belong to the same mitochondrial clade, separated from the other trout since the Pleistocene; (2) the southeastern Black Sea area is the most likely place of diversification of this clade, which is closely related to the clades from Anatolia; (3) The species from the Black Sea and the Caspian Sea drainages are monophyletic; (4) except for the basal lineage of the Ponto‐Caspian clade, Salmo rizeensis, all the lineages produce anadromous forms; (5) genetic diversification within the Ponto‐Caspian clade is related to Pleistocene glacial waves; (6) the described morphological differences between the species are not fully diagnostic, and some earlier described differences depend on body size; the differences between freshwater and marine forms exceed those between the different lineages. We suggest a conservative taxonomic approach, using the names S. rizeensis and Salmo labrax for trout from the Black Sea basin and Salmo caspius and Salmo ciscaucasicus for the fish from the Caspian basin.     

Insights into historical drainage evolution based on the phylogeography of the chevron snakehead fish (Channa striata) in the Mekong Basin

1. The phylogeography of freshwater taxa is often integrally linked with landscape changes such as drainage re‐alignments that may present the only avenue for historical dispersal for these taxa. Classical models of gene flow do not account for landscape changes and so are of little use in predicting phylogeography in geologically young freshwater landscapes. When the history of drainage formation is unknown, phylogeographical predictions can be based on current freshwater landscape structure, proposed historical drainage geomorphology, or from phylogeographical patterns of co‐distributed taxa. 2. This study describes the population structure of a sedentary freshwater fish, the chevron snakehead (Channa striata), across two river drainages on the Indochinese Peninsula. The phylogeographical pattern recovered for C. striata was tested against seven hypotheses based on contemporary landscape structure, proposed history and phylogeographical patterns of co‐distributed taxa. 3. Consistent with the species ecology, analysis of mitochondrial and microsatellite loci revealed very high differentiation among all sampled sites. A strong signature of historical population subdivision was also revealed within the contemporary Mekong River Basin (MRB). Of the seven phylogeographical hypotheses tested, patterns of co‐distributed taxa proved to be the most adequate for describing the phylogeography of C. striata. 4. Results shed new light on SE Asian drainage evolution, indicating that the Middle MRB probably evolved via amalgamation of at least three historically independent drainage sections and in particular that the Mekong River section centred around the northern Khorat Plateau in NE Thailand was probably isolated from the greater Mekong for an extensive period of evolutionary time. In contrast, C. striata populations in the Lower MRB do not show a phylogeographical signature of evolution in historically isolated drainage lines, suggesting drainage amalgamation has been less important for river landscape formation in this region.     

Phylogeography of the Chinese endemic freshwater crab Sinopotamon acutum (Brachyura,Potamidae)

Phylogeographical inferences based on multiple molecular markers from different modes of inheritance (i.e. mtDNA and nucDNA) can help to explore drivers of current species divergence over different timescales and allow a deeper understanding of evolutionary processes. River systems and mountains, owing to their drainage networks and complex topography, are often a high priority when inferring external causes of phylogeographical patterns. Here, we selected the Chinese endemic freshwater crab Sinopotamon acutum inhabiting drainage networks across the Qinling–Daba Mountains, a watershed that divides northern from southern fauna in China, as a model species (1) to investigate whether river networks and mountain systems act as barriers in shaping the phylogeography of freshwater crabs and (2) to test the impact of historic environments and ecological habitats on crab distribution. Phylogenetic tree and network analyses based on mtDNA revealed divergence between eastern and western populations, but microsatellite DNA did not recover this. Frequent and male‐biased gene flow is evident. The adjacent tributaries, such as Weihe River 1 and Hanjiang River 1, although isolated by Qinling Mts, have low genetic variation in mtDNA. In addition, microsatellite DNA showed low variation between all rivers. Frequent gene flow, genetic variation and mitochondrial genetic structure in S. acutum indicated that the Qinling–Daba Mountains have had no effect on divergence in this species; instead, hydrological networks across these mountains serve as a continuous migration corridor for these crabs. Glacial refugia are probably responsible for the initial isolation of the eastern and western clades, and local adaptations after dispersal have further driven divergence. Cytonuclear discordance in phylogeographical patterns may be attributable to ongoing male‐biased gene flow.     

Granite boulders act as deep-time climate refugia: A Miocene divergent clade of rupicolous Cnemaspis Strauch, 1887 (Squamata: Gekkonidae) from the Mysore Plateau,India, with descriptions of three new species

We describe a divergent clade and three new rupicolous species of dwarf geckos of the genus Cnemaspis from the Mysore Plateau, southern India. Cnemaspis bangarasp. nov., C. graniticola sp. nov., and C.yelagiriensis sp. nov. differ from each other by 9%–18% uncorrected ND2 sequence divergence and other South Asian Cnemaspis by 17%–33% and are morphologically distinguishable from all regional congeners and each other. The new species are known from only granite boulders in montane habitats (>1,000 m asl.), just 60–120 km straight-line distance apart from each other. A fossil-calibrated timetree and ancestral area reconstructions indicate South Asian Cnemaspis originated in Western Ghats forests with initial divergence in the Paleocene-Eocene making it perhaps the oldest Indian squamate clade. Cnemaspis that occur in Peninsular India in the dry zone outside the Western Ghats form three independently derived clades that occur in significantly warmer and drier habitats than those in the Western Ghats. The discovery of a Miocene divergent clade from rocky hills on the southern edge of the Mysore Plateau reveals these habitats as novel, long-term climate refugia. This adds to the body of work revealing ancient origins of taxa in the Indian dry zone and supports the Mysore Plateau as an important and overlooked hot spot of lizard biodiversity.     

Phylogeography of the Wabash pigtoe, Fusconaia flava (Rafinesque, 1820) (Bivalvia: Unionidae)

Reconstructing the phylogeographic patterns of widely distributedand common freshwater mussel species (Bivalvia: Unionidae) mayprovide insight into unionid evolution and speciation. The Wabashpigtoe, Fusconaia flava, is currently recognized as a single,polytypic species that is widely distributed and common throughoutthe Mississippi River drainage and parts of the Canadian Interior,Great Lakes and Gulf Coast drainages. Sequence analysis of themitochondrial COI gene revealed two divergent (3.43%) clades.Clade A consisted of specimens located throughout the upperand lower Mississippi River drainage and in the Red River (Canada)and Lake Erie drainages and all F. cerina specimens. All haplotypeswithin clade A differed by three (0.55%) or fewer nucleotidesubstitutions from the most widely distributed and abundanthaplotype, F1. Clade B, consisting of specimens located in thefar western portion of the species' range, may comprise an undescribedspecies. There was no evidence of genetic differentiation amongF. flava inhabiting headwater and intermediate-sized river localitiesof the Muskingum River system and large river localities ofthe nearby Ohio River. The divergence among F. flava haplotypescomprising clade A (0.18–1.10%) was similar to the divergencebetween the F. cerina haplotypes and the F. flava haplotypescomprising clade A (0–1.10%). This study illustrates theimportance of accessing genetic diversity across the distributionof a polytypic species. Additional analyses based on a combinationof morphology and genetics are needed to determine the taxonomicstatus of clade B and to strengthen our understanding of therelationship between F. flava and F. cerina. (Received 28 May 2007; accepted 13 August 2007)     

Phylogenetic Relationships within GenusLeuciscus(Pisces, Cyprinidae) in Portuguese Fresh Waters, Based on Mitochondrial DNA CytochromebSequences

To investigate phylogenetic relationships amongLeuciscusspecies occurring in Portuguese inland waters, the cytochromebgene was sequenced from representatives of the main rivers. This study supports the recognition of the species level forL. pyrenaicus,including populations from the southern Portuguese drainages (Tejo, Sado, and Guadiana drainages), and forL. carolitertii,including populations from the northern Portuguese drainages. The existence of two new species occurring in the extreme southwestern drainages of Mira and Arade is also suggested. The present results support the monophyly of the Mira and the Arade populations, as well as an early divergence of these two lineages. The present-day distribution ofLeuciscusspecies is seen as a consequence of Pliocene and Pleistocene events, such as river disjunctions and posterior confluence in epicontinental seas and river captures. A mixture of haplotypes was observed in the Mondego and the Tejo drainages, which could be a consequence of ancient river captures, with a possible mitochondrial DNA introgression in the Tejo drainage and a recent introduction by man in the Mondego drainage. The pattern of differentiation among mtDNA haplotypes and their geographic distribution is discussed in terms of evolutionary aspects.     

Presence of itaconic acid in the hemolymph and tissues of the freshwater crab, Potamonautes warreni Calman

Itaconic acid was detected in hemolymph, hepatopancreas, gill and muscle tissue of the freshwater crab, Potamonautes warreni. This metabolite has previously only been found in unicellular organisms and fungi. The measured itaconic acid is either being manufactured by the crab itself or present due to infestation by unicellular organisms or fungi.     

Psilorhynchus nahlongthai,a new fish species (Teleostei: Psilorhynchidae) from the Brahmaputra drainage,northeast India

Psilorhynchus nahlongthai, a new psilorhynchid fish, is described from the Diyung River, a tributary to the Kopili River (itself a southern tributary of the Brahmaputra drainage) in Assam, northeast India. It is placed in the Psilorhynchus balitora species group and can be easily distinguished from all other members of this group by a combination of the following characters: dense and prominent tuberculation on the head region; thick and long pre- and post-epiphyseal fontanelles on the neurocranium; 9 + 8 caudal-fin rays; and 34 (24 + 10) vertebrae. Genetic divergence between P. nahlongthai and members of the P. balitora species group from the Brahmaputra and neighbouring drainages, with K2P distances ranging 3.7%–14.7% and 7.4%–20.7% in the mitochondrial COI and cyt b gene datasets respectively, support its report as a new species.     

Between an ocean and a high place: coastal drainage isolation generates endemic cryptic species in the Cape kurper Sandelia capensis (Anabantiformes: Anabantidae), Cape Region,South Africa

This study investigated the range-wide phylogenetics and biogeography of the Cape kurper Sandelia capensis, a primary freshwater fish endemic to and widespread within the Cape Floristic Region (CFR) of South Africa. Maximum likelihood, Bayesian phylogenetic and haplotype network analyses, based on two mitochondrial and two nuclear genes, revealed the existence of three reciprocally monophyletic, deeply divergent and allopatric clades that probably represent cryptic species. The West Coast Clade is largely confined to the Langvlei, Verlorenvlei, Berg and Diep Rivers, the Klein River Clade is endemic to the Klein River and the South Coast Clade is found everywhere else in the range of S. capensis sensu lato. It was hypothesised that divergences within S. capensis sensu lato probably occurred because of isolation of coastal drainages by persistent drainage divides or vicariance of current tributaries by the drowning of their confluences by high sea levels. The current distribution of lineages could be due to historical range expansion and gene flow via river capture or some other mode of transdivide dispersal or dispersal during periods of low sea level via palaeoriver confluences of currently isolated coastal rivers. Comparison of BEAST2 estimated divergence times with the timing of climatic, geological and geomorphological events supported long-term coastal drainage isolation, punctuated by rare transdivide dispersal events and limited palaeoriver dispersal, as the best explanation of current phylogeographic and divergence patterns in S. capensis. Hydrological barriers that block upstream passage in palaeotributaries could hypothetically explain why S. capensis failed to disperse through certain palaeoriver confluences. There were several sites where biogeographic patterns have likely been confounded by human translocation of S. capensis. Alien fish predators and water extraction may threaten the three cryptic species more severely than previously realised, due to their smaller population sizes and inhabitation of only a portion of the range previously ascribed to S. capensis sensu lato. The preponderance of cryptic diversity and endemism in the CFR suggests that additional undescribed cryptic species of obligate freshwater fishes may be found in short coastal river systems around the world, especially in regions with a history of geological stability and a narrow continental shelf.     

Pleistocene speciation of freshwater crabs (Crustacea: Potamidae: Geothelphusa) from northern Taiwan and southern Ryukyus, as revealed by phylogenetic relationships

The phylogenetic relationship among freshwater crab species of Geothelphusa from northern Taiwan and the Yaeyama Group of islands (including Iriomote and Ishigaki) in the southern Ryukyus was studied using the mitochondrial genes 16S rRNA and COI. Our results support the hypothesis that speciation of Geothelphusa among these islands was the result of cyclic glaciations and interglaciations during the Pleistocene. Two main clades, one the Taiwan Group (containing several clades, including most Taiwanese Geothelphusa species except Geothelphusa miyazakii but including Geothelphusa minei from Yaeyama), was estimated to be separated from its sister group, the southern Ryukyus-northern Taiwan (SRN) clade (including G. miyazakii, Geothelphusa shokitai, Geothelphusa fulva and G. marginata from northern Taiwan, the Pinnacle Islands [=Diaoyutai Islands or Senkaku Islands] and Yaeyama) at about 5.3 million years ago (mya). G. shokitai was separated from others within the SRN clade at 2.4 mya, but was probably derived from G. miyazakii in northern Taiwan. The ancestor of G. miyazakii is hypothesised to have dispersed from ancestors in Yaeyama and then isolated at 2.0 mya during the Pleistocene interglaciations. This is similar to the speciation of G. minei in Yaeyama at 1.5 mya, except that its ancestors originated from north-eastern Taiwan. Four clades of freshwater crabs are present in the Fushan Botanical Garden, located in the mountainous area of north-eastern Taiwan, which might be due to the historical rearrangements of the drainage and proximity of the various river origins.