The barcode trap-Description of a new species of Microglanis, with a review of the status of Microglanis cibelae (Siluriformes: Pseudopimelodidae).

In a recent study based on the generalized mixed Yule coalescent method for delimiting species, a threshold of 2% genetic distance using cytochrome c oxidase subunit I sequences was used to delimit the species of Microglanis. That action resulted in assembling several populations of Microglanis from Atlantic coastal rivers between Rio Grande do Sul and São Paulo states as a single species, Microglanis cottoides, including Microglanis cibelae as a junior synonym. We reexamined these populations and found three species diagnosed by their morphology and that constitute separate mtDNA lineages, including a new species. The synonym of M. cibelae and M. cottoides is reviewed and refuted based on morphological and molecular evidence. M. cibelae and the new species are sympatric and occasionally syntopic in the Tramandaí, Mampituba, and Araranguá river basins. The new species is distinguished from M. cibelae and M. cottoides by the anterior margin of the posttemporosupracleitrum narrow articulated with the epioccipital, the short mental and maxillary barbels, and depressed head and body.

A new species of Geophagus (Teleostei: Cichlidae): Naming a cichlid species widely known in the aquarium hobby as 'Geophagus sp. Tapajos red head'.

A new species of Geophagus sensu stricto is described from the Tapajos River basin, Brazil, elevating the number of species of the genus to 21. The new species is of commercial importance and is known in the aquarist trade as Geophagus 'red head'. The new species is diagnosed using an integrative approach, based on mitochondrial DNA analysis along with morphological evidence. The new species is distinguished from all congeners by the absence of markings on the head, the bar pattern composed by nine vertical bars on the flanks and the presence of distinct longitudinal bands in the caudal fin. Additionally, it shows a genetic distance of at least 2.0% in cytochrome b gene sequences from its closest congeners. Molecular analysis including most genera of Cichlidae from South America corroborates that the new species belongs to the group of Geophagus sensu stricto.

The record and threats of the invasion of palometa Serrasalmus maculatus (Characiformes: Serrasalmidae) in the Patos lagoon drainage, southern Brazil.

We report the occurrence of an invasive alien species, palometa Serrasalmus maculatus, in the Patos Lagoon drainage. Primary occurrence data were based on three specimens captured and preserved as vouchers in scientific collections. Additionally, we searched for secondary records from unpublished scientific sources, public agencies reports and media news to find additional reports. We discussed the possible pathways of invasion, suggesting as the vector of introduction transpositions from the Uruguay River basin. Ecological implications for ichthyofauna, environmental impacts and risk of other events of invasion in the adjoining basins are discussed.

Hidden or unnoticed? Multiple lines of evidence support the recognition of a new species of Pseudocorynopoma (Characidae: Corynopomini).

Species delimitation is a permanent issue in systematics. The increasing recognition of geographically isolated populations as independent lineages allowed by new methods of analysis has inflated the species-populations dilemma, which involves deciding whether to consider separate lineages as different species or structured genetic populations. This is commonly observed between fishes of adjacent river basins, with some lineages being considered allopatric sister species and others considered isolated populations or variants of the same species. Pseudocorynopoma doriae is a characid diagnosed from its single congener by the number of anal-fin rays and sexually dimorphic characters of males, including distinct fin colouration. The authors found variation in the colour pattern between isolated populations previously identified as P. doriae but no variation in scale or fin-ray counts. They analysed molecular evidence at the population level and morphological differences related to life history (e.g., colour dimorphism related to inseminating behaviour). The results provide compelling evidence for the recognition of a new species of Pseudocorynopoma despite the lack of discrete differences in meristic data. The recognition of the new species is consistent with biogeographical evidence for the long-term isolation of the respective river drainages and with differences between the ichthyofaunal communities of these rivers.

Biogeography, habitat transitions and hybridization in a radiation of South American silverside fishes revealed by mitochondrial and genomic RAD data.

Rivers and lake systems in the southern cone of South America have been widely influenced by historical glaciations, carrying important implications for the evolution of aquatic organisms, including prompting transitions between marine and freshwater habitats and by triggering hybridization among incipient species via waterway connectivity and stream capture events. Silverside fishes (Odontesthes) in the region comprise a radiation of 19 marine and freshwater species that have been hypothesized on the basis of morphological or mitochondrial DNA data to have either transitioned repeatedly into continental waters from the sea or colonized marine habitats following freshwater diversification. New double digest restriction-site associated DNA data presented here provide a robust framework to investigate the biogeographical history of and habitat transitions in Odontesthes. We show that Odontesthes silversides originally diversified in the Pacific but independently colonized the Atlantic three times, producing three independent marine-to-freshwater transitions. Our results also indicate recent introgression of marine mitochondrial haplotypes into two freshwater clades, with more recurring instances of hybridization among Atlantic- versus Pacific-slope species. In Pacific freshwater drainages, hybridization with a marine species appears to be geographically isolated and may be related to glaciation events. Substantial structural differences of estuarine gradients between these two geographical areas may have influenced the frequency, intensity and evolutionary effects of hybridization events.

A new species of Odontostilbe cope (Characiformes: Cheirodontinae) from rio Madeira basin diagnosed based on morphological and molecular data.

A new species of Odontostilbe is described from the rio Jaciparaná, rio Madeira basin, Rondônia, Brazil. Odontostilbe pacaasnovos differs from all its congeners, except O. pequira, by the colour pattern. Additionally, it differs from its congeners by the terminal mouth, number of cusps in the teeth of the premaxilla (5-7), number of branched rays in the anal fin (19-22), by the shape of dentary teeth (5-7 cusps with central cusp larger and longer than laterals cusps) and by the number of lamellae of the olfactory rosette (17-18 in male and 14 in female). Morphological and molecular comparisons corroborate the distinctiveness between O. pacaasnovos and its congeners, justifying its recognition as a new species.

Phylogenetic relationships and historical biogeography of Oligosarcus (Teleostei: Characidae): Examining riverine landscape evolution in southeastern South America.

The pike-characin Oligosarcus is a group of Characidae composed of 22 species, which have mostly allopatric distributed species in southeastern South America and sympatric occurrence of few species. Oligosarcus shares a similar distribution pattern with other fish genera and therefore, can help us to understand biogeographic events that influenced freshwater fish distribution in the southeastern South America. Our paper presents the most extensive taxonomic coverage for molecular analysis of Oligosarcus and uses various methods to examine the evolutionary history of the genus. Phylogenetic relationships among species of Oligosarcus were examined using a multilocus dataset by Maximum Likelihood and Bayesian methods. A relaxed molecular clock was used to estimate lineage divergence times, which provide a framework to examine the biogeographic history of this clade across the drainage basins of southeastern South America. Oligosarcus was resolved as monophyletic with strong support, and related to lineages currently assigned to the genus Astyanax. Within Oligosarcus, two groups of approximately equal species richness were resolved as monophyletic, mainly restricted to continental and coastal drainages of southeastern South America. Oligosarcus radiation is estimated to the late Neogene, with its origin in the Pliocene and most speciation events occurring in the Pleistocene. Some apomorphic characteristics associated with piscivory (e.g. large caniniform teeth) in Oligosarcus likely have evolved once, and are convergent to similar phenotypes observed in a distantly related clade of Astyanax (formerly Bramocharax). In addition, the presence of morphological convergence within the genus Oligosarcus (e.g. trophic morphology) seems to explain the difference between the present molecular hypothesis and some previous morphological studies. Ancestral geographical range estimation using analytical methods (e.g. DIVALIKE and DEC) demonstrated the effects of different Landscape Evolution Models (LEMs) on diversification of Oligosarcus. The results suggest that the two main Oligosarcus clades evolved in allopatry in continental and coastal drainages, with subsequent range extension and vicariance events that established the modern distributions. LEM analyses indicate the importance of formation of riverine barriers across the watershed of the La Plata basin and the effects of sea-level changes during the Pleistocene for delineating lineage distributions of Oligosarcus.

Geographic distributions, phenotypes, and phylogenetic relationships of Phalloceros (Cyprinodontiformes: Poeciliidae): Insights about diversification among sympatric species pools.

With 22 described species, Phalloceros is the most species-rich genus of Poeciliidae in South America. Phalloceros diversity is characterized by high degrees of endemism and sympatry in coastal and inland drainages in southeastern South America. The taxa are also characterized by pronounced differentiation in sexual characters (i.e., female urogenital papilla and male gonopodium), which might have contributed to their diversification. Here we estimate phylogenetic relationships based on more than 18,000 loci in 93 individuals representing 19 described species and two putative undescribed species. Morphologically defined species correspond to monophyletic species lineages, with individuals within a species clustering together in phylogenetic estimates, with the main exception being P. harpagos, supporting undiscovered diversity in this morphospecies. Shifts in the female and male sexual traits (i.e., urogenital papilla and gonopodium) occurred in concert multiple times along the phylogeny highlighting the role of sexual selection in driving divergence in this genus. Out of 22 valid species, 14 species are found in sympatry with at least one other species of this genus. However, most co-occurrences are observed among non-sister species suggesting that diversification among closely related species involved mostly allopatric speciation, with only two instances of sympatric sister-species observed. A strong mismatch in sexual traits among sympatric taxa suggests that co-existence may be linked to divergent sexual traits that maintain species genetic distinctiveness through mechanical disruptions of interbreeding.

Two new species of Heptapterus (Siluriformes: Heptapteridae) from the Uruguay River basin, Brazil.

As part of an ongoing taxonomic revision of the genus Heptapterus from the Laguna dos Patos and Uruguay River drainages and Atlantic coastal streams of southern Brazil and Uruguay, two new species closely related to Heptapterus mustelinus were identified. Both species are endemic to small tributaries of the Uruguay River. The two new species are distinguished from each other and from other species of Heptapterus by arrangement of cephalic and trunk laterosensory systems, number of vertebrae and number of dorsal, pectoral and anal-fin rays. Phylogenetic analyses of mitochondrial DNA (coI and cytb) sequence data further supports distinctiveness of the two new species.

Genetics and telemetry indicate unexpected movements among structured populations for Brachyplatystoma platynemum in the Amazon.

The genetic analysis of Brachyplatystoma platynemum individuals sampled from the lower Madeira River reinforces the existence of two structured populations in the Amazon Basin (Madeira and Amazon populations). However, the recapture of an individual from the Amazon population in the Solimões River, which was telemetry-tagged in the Madeira River after the damming, indicates that fish from the Amazon population move between the two river systems. This has not yet been observed, however, in the Madeira River population, which is currently divided and isolated in the lower and upper Madeira River by the construction of two dams.

A new species of Diapoma (Characiformes, Characidae, Stevardiinae) from the Rio Paraná basin, with an identification key to the species of the genus.

Diapoma nandi is described from the Piray-Miní stream, a tributary of the Rio Paraná in Argentina. It is characterized among the Stevardiinae by having a terminal mouth, two unbranched and eight branched dorsal-fin rays, one unbranched and six branched pelvic-fin rays and the absence of a caudal-fin organ and is distinguished from all congeners by the following combination of characters: unmodified scales on the lower caudal-fin lobe, lack of enlarged opercle and subopercle, incomplete lateral line, hyaline adipose fin, anal-fin distal border straight or slightly convex in adult males, large tricuspid teeth on anterior region of the dentary, distal arrangements of the anal-fin bony hooks in adult males, middle caudal-fin rays lacking large round blotch and several morphometric variables associated with body shape. Additionally, we conducted a morphometric comparison focused on the congeners that co-occur in the Paraná basin.

Molecular phylogeny of the subfamily Stevardiinae Gill, 1858 (Characiformes: Characidae): classification and the evolution of reproductive traits.

BACKGROUND: The subfamily Stevardiinae is a diverse and widely distributed clade of freshwater fishes from South and Central America, commonly known as "tetras" (Characidae). The group was named "clade A" when first proposed as a monophyletic unit of Characidae and later designated as a subfamily. Stevardiinae includes 48 genera and around 310 valid species with many species presenting inseminating reproductive strategy. No global hypothesis of relationships is available for this group and currently many genera are listed as incertae sedis or are suspected to be non-monophyletic. RESULTS: We present a molecular phylogeny with the largest number of stevardiine species analyzed so far, including 355 samples representing 153 putative species distributed in 32 genera, to test the group's monophyly and internal relationships. The phylogeny was inferred using DNA sequence data from seven gene fragments (mtDNA: 12S, 16S and COI; nuclear: RAG1, RAG2, MYH6 and PTR). The results support the Stevardiinae as a monophyletic group and a detailed hypothesis of the internal relationships for this subfamily. CONCLUSIONS: A revised classification based on the molecular phylogeny is proposed that includes seven tribes and also defines monophyletic genera, including a resurrected genus Eretmobrycon, and new definitions for Diapoma, Hemibrycon, Bryconamericus sensu stricto, and Knodus sensu stricto, placing some small genera as junior synonyms. Inseminating species are distributed in several clades suggesting that reproductive strategy is evolutionarily labile in this group of fishes.

The phylogenetic placement of Hollandichthys Eigenmann 1909 (Teleostei: Characidae) and related genera.

The phylogenetic relationships among characids are complex with many genera remaining of uncertain systematic position inside the family. The genus Hollandichthys is one of these problematic genera. It has been considered as incertae sedis inside this family until two recently published phylogenies, one morphological and one molecular, arrived at alternative hypothesizes as to the relationships of Hollandichthys with Pseudochalceus or Rachoviscus, respectively. In this paper, we infer the phylogenetic relations of these taxa based on five genes (three mitochondrial - COI, ND2 and 16S; and two nuclear - Sia and Trop), totaling up to 2719 bp. The 41 analyzed species in the Characidae include four incertae sedis characid taxa once hypothesized as related to Hollandichthys, but never analyzed in a single phylogeny (Rachoviscus, Pseudochalceus, Nematocharax and Hyphessobrycon uruguayensis). Here we propose Rachoviscus as the sister-group of Hollandichthys, grouped in the large clade C previously defined, along with the remaining incertae sedis taxa studied here. In addition, we support the evidence that insemination evolved independently at least three times in the Characidae.

Relationships among major lineages of characid fishes (Teleostei: Ostariophysi: Characiformes), based on molecular sequence data.

The family Characidae is a group of freshwater bony fishes that exhibits high species-level diversity and whose members inhabit parts of Texas, Mexico, and Central and South America. Thus far, morphological data have been of limited use in discerning relationships among subfamilies and incertae sedis genera of the family Characidae. In this study, DNA sequence data from GenBank were combined with new sequences for analyses under Bayesian and parsimony schemes. Sequences fell into four gene partitions, with three genes in the mitochondrial subset (12S, 16S, COI genes) and one gene in the nuclear subset (RAG2 gene). Inferred Bayesian and parsimony-based phylogenies reject the monophyly of certain groups (e.g., Astyanax, Hyphessobrycon, and Bryconamericus), do not reject the monophyly of others (e.g., Cheirodontinae and "clade A" of previous authors), and present new sister-group hypotheses (e.g., Brittanichthys sister to Paracheirodon). Sister to clade A is a lineage referred to herein as "clade B" which includes Exodon and exemplars from Cheirodontinae (the most basal lineage within clade B), Aphyocharacinae, Tetragonopterinae, and Characinae (excluding Gnathocharax). "Clade C" is sister to A+B and contains representatives of large incertae sedis genera (e.g., Hyphessobrycon, Hemigrammus), as well as members of Stethaprioninae. Unless certain other subfamilial names are to be disregarded, the use of Tetragonopterinae should continue to be restricted to species of Tetragonopterus because other genera previously referred to this subfamily grouped in clades A or C, quite distant from Tetragonopterus.

Comparison and optimization for DNA extraction of archived fish specimens.

The DNA extracted from museum alcohol-fixed specimens can be a valuable source of information for solving taxonomic, phylogenetic, ecological and conservational questions. However, this type of DNA, also called ancient DNA, is routinely obtained in small portions and highly fragmented. We have tested two different extraction kits in museum type-specimens of the fish family Characidae. Aiming to increase the DNA yield, we made modifications on a Qiagen manufacturer protocol, in the elution step. Also, to overcome the issue of DNA fragmentation, we applied our efforts in Sanger sequencing, to find a highly variable and, in result, informative COI fragment. Based on our results, there is no correlation between amount of the DNA extracted and the age of the sample. The Sanger sequencing generated sequences which are useful in solving taxonomic puzzles. Here are presented the customization and guidelines that allowed us to recover DNA from the archived fish specimens. •DNA extraction from archived fish specimens is more effective when using silica columns.•Change of the elution times from minutes in room temperature to 24 h in freezer greatly improved the DNA yielded.•Short but highly variable sequences replace the need to sequence the entire gene to identify a species.

Sperm ultrastructure in the inseminating Macropsobrycon uruguayanae (Teleostei: Characidae: Cheirodontinae).

Macropsobrycon uruguayanae is a small, inseminating characid (tetra) of the tribe Compsurini. Although spermatozoa can be found within the ovarian cavity close to oocytes, the exact moment of fertilization has not yet been determined. Spermatozoa have moderately elongate nuclei with electron-dense chromatin. During spermiogenesis, nuclear rotation takes place. Elongate mitochondria with lamellar cristae are found posterior to the nucleus. Centrioles are parallel to one another with the proximal centriole slightly anterior to the longer distal one. The anterior tip of the proximal centriole is located within a shallow nuclear fossa. Electron-dense spurs are associated within the anterior and posterior ends of the distal centriole. Striated centriolar rootlets radiate both anteriorly and posteriorly from the distal centriole. Nine longitudinal accessory microtubules surround the axoneme in the proximal flagellum. The flagellum has a typical 9 + 2 axoneme with no intratubular differentiation. Atypical spermatozoa are also found in the testicular lumen. These cells resemble spermatozoa in most aspects, except that their nuclei are variable in shape, with the granular chromatin less electron-dense than that seen in spermatozoa. The origin and function of these cells could not be determined. The specializations seen in the spermatozoa are discussed as possible adaptations related to the habit of insemination.

Morphology and molecular data reveal the presence of two new species under Rhamdia quelen (Quoy Gaimard, 1824) (Siluriformes: Heptapteridae) species complex.

The eustatic movements triggered by glaciations during the Quaternary have shaped the landscape of Brazilian Atlantic Coast. Cyclic sea-level changes either isolated or connected freshwater fish populations, impacting their distribution and diversification. Rhamdia quelen has been widely recorded from the Brazilian Atlantic Coastal rivers, but it is also considered a species complex. A phylogeographic study carried out using three molecular markers of mtDNA and one of nDNA in the populations of Rhamdia from the hydrographic basins of Southern Atlantic Coast of Brazil recovered three evolutionarily distinct groups: one represented by the populations found in lowlands of all studied watersheds; the second group composed of populations found in the upper tributaries of the rio Araranguá, rio Mampituba and rio Tramandaí; and a third group found exclusively in the upper portions of rio Tubarão. The genetic divergences among these three lineages of Rhamdia is discussed according to sea level changes in the Quaternary. Two new species of Rhamdia are diagnosed and described based on morphological and molecular evidence.

A new species of Bryconops (Characiformes: Iguanodectidae) from the Rio Tapajós basin, Brazil.

A new species of Bryconops is described from upper rio Juruena drainage, rio Tapajós basin, Amazon basin, state of Mato Grosso, Brazil. The new species is hypothesized as belonging to the subgenus Creatochanes by presenting the posterior extension of maxilla reaching to the junction of second and third infraorbital bones, and the ventroposterior margin of second infraorbital forming a complete border with third infraorbital, resulting in the lack of a naked area between them. The new species is easily distinguished from other species of the subgenus Creatochanes by the color pattern of the caudal fin, which consists of the dorsal lobe conspicuously dark pigmented on its distal half and the ventral lobe dark gray pigmented along its ventral portion below the horizontal through the ventral margin of the caudal peduncle.