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.

Phylogenetic relationships and description of two new species of Diapoma (Characidae: Stevardiinae) from the La Plata River basin

Herein we describe two new species of Diapoma, one from the Negro River, a tributary of the Uruguay River in Brazil and Uruguay, and one from the Iguaçu River, in Brazil and Argentina. The new species from the Negro River basin is distinguished from its congeners by the following combination of characters: a black narrow and conspicuous line restricted to the body horizontal septum, incomplete lateral line, tricuspid teeth in the inner series of the premaxilla, and a lower body depth at vertical through the dorsal-fin origin (29.3-32.8% SL in males and 27.7-33.3% SL in females). The new species from the Iguaçu River basin is distinguished from its congeners by the following combination of characters: a discontinuous lateral line, adipose fin hyaline, longer anal-fin base (26.5-32.4% SL), and a longitudinal black stripe along the median region of caudal-fin rays. Additionally, we updated the molecular phylogeny of the genus, including new sequences from these two new species and Diapoma thauma. An identification key for species of Diapoma is presented, modified from previous study.(AU)

Neste trabalho descrevemos duas espécies novas de Diapoma, uma do rio Negro, afluente do rio Uruguai, Brasil e Uruguai, e a segunda do rio Iguaçu, Brasil e Argentina. A espécie nova do rio Negro é diagnosticada de suas congêneres pela combinação das seguintes características: uma linha preta, estreita e conspícua restrita ao septo horizontal do corpo, linha lateral incompleta, dentes da série interna da pré-maxila tricuspidados, e baixa altura do corpo na vertical que passa pela origem da nadadeira dorsal (29,3-32,8% SL em machos e 27,7-33,3% SL em fêmeas). A espécie nova do rio Iguaçu é diagnosticada de suas congêneres pela combinação das seguintes características: linha lateral descontínua, nadadeira adiposa não pigmentada de preto, base da nadadeira anal longa (26,5-32,4% SL) e raios médios da nadadeira caudal com uma linha preta longitudinal. Adicionalmente, atualizamos a filogenia molecular do gênero, incluindo novas sequências destas duas espécies novas e de Diapoma thauma. Uma chave de identificação para as espécies de Diapoma é apresentada, modificada de estudo anterior.(AU)

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.

The genital and anal papillae of Compsura heterura (Characidae: Cheirodontinae): morphological structure and possible role in insemination

The function of the genital and anal papillae for insemination in Compsura heterura is discussed based on the description of their morphologies at different stages of the life cycle and during copulation and spawning. In males and females both the genital and anal papillae are involved in copulation in C. heterura. A pre-anal organ, anterior to the anus and stretched posteriorly, is present in adult males. The openings of the urinary channel and oviduct are separate in females. During copulation, there is an approximation between the female genital and anal papillae which become temporarily juxtaposed, forming a chamber enclosing the opening of the oviduct, which may also function in sperm capture. During spawning, the lateral edges of the female genital papilla are projected anteriorly, acquiring a tubular shape for oviposition.(AU)

A função das papilas genital e anal na inseminação em Compsura heterura é discutida com base na descrição de suas morfologias em diferentes estágios do ciclo de vida e por ocasião da cópula e desova. Em machos e fêmeas, as papilas genital e anal estão envolvidas na cópula. Um órgão pré-anal, anterior ao ânus e alongado posteriormente, está presente em machos adultos. Fêmeas possuem aberturas urinária e genital separadas. Durante a cópula, ocorre uma aproximação entre as papilas genital e anal femininas, que se justapõem temporariamente, formando uma câmara fechada onde se abre o oviduto, e que serve possivelmente à captura de esperma. Durante a desova, as bordas laterais da papila genital feminina são projetadas anteriormente, adquirindo um formato tubular para ovoposição.(AU)

Hydrography rather than lip morphology better explains the evolutionary relationship between Gymnogeophagus labiatus and G. lacustris in Southern Brazil (Cichlidae: Geophagini)

Gymnogeophagus labiatus and G. lacustris have been long recognized as sister species exhibiting different ecological requirements. Gymnogeophagus labiatus occurs in rock bottom rivers in the hydrographic basins of Patos Lagoon (HBP) and Tramandaí River (HBT), while G. lacustris is exclusive from sand bottom coastal lagoons of the HBT. In this study, we used molecular markers, morphological measurements and data from nuptial male coloration to investigate the evolutionary relationship between these species in each hydrographic basin. We found, for all data sets, a closer relationship between G. labiatus and G. lacustris from the HBT than between G. labiatus populations from HBT and HBP. In particular, lip area had a large intraspecific plasticity, being uninformative to diagnose G. lacustris from G. labiatus. Molecular clock-based estimates suggest a recent divergence between species in the HBT (17,000 years ago), but not between G. labiatus from HBP and HBT (3.6 millions of years ago). Finally, we also found a divergent G. labiatus genetic lineage from the Camaquã River, in the HBP. These results show that the current taxonomy of G. labiatus and G. lacustris does not properly represent evolutionary lineages in these species.(AU)

Gymnogeophagus labiatus e G. lacustris vêm sendo consideradas espécies irmãs que possuem diferentes exigências ecológicas. Gymnogeophagus labiatus ocorre em rios de fundo de pedra nas bacias hidrográficas da Laguna dos Patos (HBP) e do rio Tramandaí (HBT), enquanto G. lacustris é exclusivo da HBT, ocorrendo em lagoas costeiras de fundo de arenoso. Nesse estudo, foram usados marcadores moleculares, medidas morfológicas e dados sobre a coloração nupcial em machos para investigar a relação evolutiva entre estas espécies em cada bacia hidrográfica. Para todos os conjuntos de dados foi observada uma relação mais próxima entre G. labiatus e G. lacustris da HBT do que entre as populações de G. labiatus da HBP e HBT. Em particular, a área do lábio teve uma grande plasticidade intraespecífica, não sendo informativa para diagnosticar G. lacustris de G. labiatus. Estimativas baseadas no relógio molecular sugeriram uma divergência recente entre as espécies da HBT (17.000 anos atrás), mas não entre as populações de G. labiatus da HBP e HBT (3,6 milhões de anos atrás). Finalmente, também foi encontrada uma linhagem genética de G. labiatus divergente no rio Camaquã, na HBP. Esses resultados mostram que a taxonomia atual de G. labiatus e G. lacustris não representa adequadamente as linhagens evolutivas nessas espécies.(AU)

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.

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.

Systematic revision of the Neotropical catfish genus Scleronema (Siluriformes: Trichomycteridae), with descriptions of six new species from Pampa grasslands

The Neotropical genus Scleronema is revised based on the re-examination of the type specimens and 1,713 newly collected specimens. Scleronema is diagnosed from other trichomycterids by the following unambiguous derived characters: fleshy flap at the base of the maxillary barbell; skin flap in the posterior margin of the opercle; articulation between the autopalatine and the vomer ventrally located, with the medial margins of the autopalatines very close to each other; and autopalatine with an interrupted or not interrupted ossified arch-shaped process on its dorsal surface forming a canal. Scleronema minutum and S. operculatum are redescribed, S. angustirostre is considered a junior synonym of S. minutum, and six new species are described. A lectotype is designated for Trichomycterus minutus. The type localities of S. angustirostre, S. minutum, and S. operculatum are reviewed in order to correct erroneous information cited in articles and catalogs subsequent to the original descriptions. Species of Scleronema are geographically distributed in the La Plata basin and Atlantic coastal drainages from Southern Brazil, Southern Paraguay, Northeastern Argentina and Uruguay. They inhabit rivers or streams with sand- or gravel-bottoms across the Pampa grasslands. We provide evidences to recognize two putative monophyletic units within the genus, namely the S. minutum species group and the S. operculatum species group, and discuss the distribution patterns of their species.(AU)

O gênero Neotropical Scleronema é revisado com base na análise do material-tipo e outros 1.713 espécimes recentemente coletados. Scleronema é diagnosticado de outros tricomicterídeos pelos seguintes caracteres derivados não ambíguos: aba de pele na base do barbilhão maxilar; aba de pele na margem posterior do opérculo; articulação entre o autopalatino e o vômer posicionada ventralmente, deixando as margens mediais dos autopalatinos muito próximas entre si; e o autopalatino com um processo com formato de arco na sua superfície dorsal. Scleronema minutum e S. operculatum são redescritas, S. angustirostre é considerada um sinônimo júnior de S. minutum e seis novas espécies são descritas. Um lectótipo é designado para S. minutum. As localidades-tipo de S. angustirostre, S. minutum e S. operculatum são revisadas com o intuito de corrigir informações errôneas citadas em artigos e catálogos após suas descrições. As espécies de Scleronema distribuem-se na bacia do rio da Plata e drenagens costeiras atlânticas no sul do Brasil, sul do Paraguai, nordeste da Argentina e Uruguai, habitando rios e riachos ao longo do Pampa com fundos de areia ou cascalho. Dois grupos de espécies supostamente monofiléticos, o grupo S. minutum e o grupo S. operculatum, são reconhecidos no gênero e os padrões de distribuição de suas espécies são discutidos.(AU)

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.

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.

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.

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.

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.

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.

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.

Reappraisal of the systematics of Microglanis cottoides (Siluriformes, Pseudopimelodidae), a catfish from southern Brazil.

The southern region of Brazil is characterized by high species diversity and endemism of freshwater fishes distributed across geographically isolated river basins. Microglanis cottoides has a widespread range across these river basins and occurs in sympatry with other endemic species of the genus (e.g. M. cibelae, M. eurystoma, and M. malabarbai). Herein we tested the monophyly of M. cottoides and presented for the first time information about the molecular phylogeny of species in the genus. The results suggest that M. cottoides currently forms a non-monophyletic group which includes populations endemic to the Uruguay River basin that are more closely related to M. malabarbai, and excludes M. cibelae, found to be nested within M. cottoides. Based on an integrative approach using morphological and molecular data, we propose M. cibelae as a junior synonym of M. cottoides, and the populations of the Uruguay River basin previously assigned to M. cottoides in fact belong to M. malabarbai. Our molecular phylogeny shows that M. cottoides is sister to M. parahybae, which is also a coastal species, and M. malabarbai is sister of M. garavelloi, both endemic to inland river basins. The time-calibrated phylogeny indicates that the separation between inland and the coastal clades occurred in the Tertiary period, and that the species within the coastal basins diverged in the Pliocene, which overlaps with the diversification times estimated for the two inland species as well. This pattern of diversification corroborates some previous studies with other fishes from the same region.

Revision of Banded Knifefishes of the Gymnotus carapo and G. tigre clades (Gymnotidae Gymnotiformes) from the Southern Neotropics.

Banded Knifefishes (Gymnotus, Gymnotidae) comprise the most species-rich, ecologically tolerant (eurytopic), and geographically widespread genus of Neotropical electric fishes (Gymnotiformes), with 40 valid species occupying most habitats and regions throughout the humid Neotropics. Despite substantial alpha-taxonomic work in recent years, parts of the genus remain characterized by taxonomic confusion. Here we describe and delimit species of the G. carapo and G. tigre clades from the southern Neotropics, using body proportions (caliper-based morphometrics), fin-ray, scale and laterosensory-pore counts (meristics), quantitative shape differences (geometric morphometrics), osteology, color patterns and electric organ discharges. We report these data from 174 Gymnotus specimens collected from 100 localities throughout the southern Neotropics, and delimit species boundaries in a multivariate statistical framework. We find six species of the G. carapo clade (G. carapo australis, G. cuia n. sp., G. chimarrao, G. omarorum, G. pantanal, and G. sylvius), and two species of the G. tigre clade (G. inaequilabiatus and G. paraguensis) in the southern Neotropics. The new species G. cuia is readily distinguished from the morphologically similar and broadly sympatric G. c. australis by a shorter head and deeper head and body, and from the morphologically similar and sympatric G. omarorum by fewer lateral-line ventral rami and fewer pored lateral-line scales anterior to the first ventral ramus. We also review the geographic distributions of all eight species of the G. carapo and G. tigre clades in the southern Neotropics, showing that G. cuia is the most widespread species in the region. These results affirm the importance of understanding the structure of variation within and between species, both geographic and ontogenetic, in delimiting species boundaries.