Peripheral isolation and demographic stability are reflected in the genetic diversity of the populations of the Helmeted Manakin: a bird endemic to the gallery forests.

The Cerrado encompasses a complex network of hydrographic basins, which is responsible for the formation and maintenance of the riparian and gallery forests. Alterations in the vegetation resulting from the paleoclimatic changes that occurred during the Pleistocene influenced the current distribution of these humid forests. To understand of the evolutionary dynamics of this landscape on the population structure of the associated organisms, we studied the population genetics of the Antilophia galeata (Pipridae), a bird endemic to the gallery forests of the Cerrado. For this, we evaluated the variability of the mitochondrial control region of 71 A. galeata specimens from 18 localities distributed throughout the Cerrado. The results indicated the existence of significant substructure among the populations located in the peripheral areas of the Cerrado, in comparison with the populations found in the central portion, reflecting historical changes in the environment. We also found high levels of diversity in all the populations, while the analysis of the demographic history revealed a scenario of stability. Overall, then, these findings indicate that the historical modifications of the gallery forest distribution may have been most accentuated in the peripheral zones, with a greater stability and connectivity persisting in the central portion of the biome.

Contrasting migratory journeys and changes in hippocampal astrocyte morphology in shorebirds.

Semipalmated sandpiper (Calidris pusilla) migration to the Southern Hemisphere includes a 5-day non-stop flight over the Atlantic Ocean, whereas semipalmated plover (Charadrius semipalmatus) migration, to the same area, is largely over land, with stopovers for feeding and rest. We compared the number and 3D morphology of hippocampal astrocytes of Ch. semipalmatus before and after autumnal migration with those of C. pusilla to test the hypothesis that the contrasting migratory flights of these species could differentially shape hippocampal astrocyte number and morphology. We captured individuals from both species in the Bay of Fundy (Canada) and in the coastal region of Bragança (Brazil) and processed their brains for selective GFAP immunolabeling of astrocytes. Hierarchical cluster analysis of astrocyte morphological features distinguished two families of morphological phenotypes, named type I and type II, which were differentially affected after migratory flights. Stereological counts of hippocampal astrocytes demonstrated that the number of astrocytes decreased significantly in C. pusilla, but did not change in Ch. semipalmatus. In addition, C. pusilla and Ch. semipalmatus hippocampal astrocyte morphological features were differentially affected after autumnal migration. We evaluated whether astrocyte morphometric variables were influenced by phylogenetic differences between C. pusilla and Ch. semipalmatus, using phylogenetically independent contrast approach, and phylogenetic trees generated by nuclear and mitochondrial markers. Our findings suggest that phylogenetic differences do not explain the results and that contrasting long-distance migratory flights shape plasticity of type I and type II astrocytes in different ways, which may imply distinct physiological roles for these cells.

Genetic diversity and structuring in the arapaima (Osteoglossiformes, Osteoglossidae) population reveal differences between the Amazon and the Tocantins-Araguaia basins.

Arapaima is a widely-distributed fish of enormous economic importance in the Amazon region. In the present study, a total of 232 specimens were sampled, 121 from five sites in the Amazon basin and 111 from five sites in the Tocantins-Araguaia basin. The analyses investigated fragments of the Cytochrome b, Control Region, Cytochrome Oxidase I, NADH dehydrogenase subunit 2 and seven loci microsatellites. The analyses revealed the existence of two mitochondrial lineages within the general area, with no haplotypes shared between basins, and genetic variability significantly higher in the Amazon than in the Tocantins-Araguaia basin. Two divergent, but sympatric mitochondrial lineages were found in the Amazon basin, but only a single lineage in the Tocantins-Araguaia basin. The existence of these two mitochondrial lineages indicates that past events, probably occurring during the Pleistocene, resulted in the separation of the populations of this species and molded its evolutionary history, which is reflected directly in its mitochondrial DNA. The analysis of the arapaima population structure identified distinct levels of diversity within the distribution of the species, indicating specific geographic regions that will require special attention for the development of conservation and management strategies.

Genetic diversity and structure in Arapaima gigas populations from Amazon and Araguaia-Tocantins river basins.

BACKGROUND: Arapaima gigas (Schinz, 1822) is the largest freshwater scaled fish in the world, and an emerging species for tropical aquaculture development. Conservation of the species, and the expansion of aquaculture requires the development of genetic tools to study polymorphism, differentiation, and stock structure. This study aimed to investigate genomic polymorphism through ddRAD sequencing, in order to identify a panel of single nucleotide polymorphisms (SNPs) and to simultaneously assess genetic diversity and structure in wild (from rivers Amazon, Solimões, Tocantins and Araguaia) and captive populations. RESULTS: Compared to many other teleosts, the degree of polymorphism in A. gigas was low with only 2.3% of identified RAD-tags (135 bases long) containing SNPs. A panel of 393 informative SNPs was identified and screened across the five populations. Higher genetic diversity indices (number of polymorphic loci and private alleles, Shannon's Index and HO) were found in populations from the Amazon and Solimões, intermediate levels in Tocantins and Captive, and very low levels in the Araguaia population. These results likely reflect larger population sizes from less urbanized environments in the Amazon basin compared to Araguaia. Populations were significantly differentiated with pairwise FST values ranging from 0.086 (Amazon × Solimões) to 0.556 (Amazon × Araguaia). Mean pairwise relatedness among individuals was significant in all populations (P < 0.01), reflecting a degree of inbreeding possibly due to severe depletion of natural stocks, the species sedentary behaviour and possible sampling biases. Although Mantel test was not significant (P = 0.104; R2 = 0.65), Bayesian analysis in STRUCTURE and discriminant analysis of principal components (DAPC) showed populations of Amazon and Solimões to be genetically differentiated from Araguaia, with Tocantins comprising individuals from both identified stocks. CONCLUSIONS: This relatively rapid genotyping by sequencing approach proved to be successful in delineating arapaima stocks. The approach and / or SNP panels identified should prove valuable for more detailed genetic studies of arapaima populations, including the elucidation of the genetic status of described discrete morphotypes and aid in delivery of conservation programs to maintain genetic diversity in reservoirs across the Amazon region.

Recent chapters of Neotropical history overlooked in phylogeography: Shallow divergence explains phenotype and genotype uncoupling in Antilophia manakins.

Establishing links between phenotypic and genotypic variation is a central goal of evolutionary biology, as they might provide important insights into evolutionary processes shaping genetic and species diversity in nature. One of the more intriguing possibilities is when no genetic divergence is found to be associated with conspicuous phenotypic divergence. In that case, speciation theory predicts that phenotypic divergence may still occur in the presence of significant gene flow-thereby resulting in little genomic divergence-when genetic loci underpinning phenotypes are under strong divergent selection. However, a finding of phenotypic distinctiveness with weak or no population genetic structure may simply result from low statistical power to detect shallow genetic divergences when small data sets are used. Here, we used a subgenomic data set of 2,386 ultraconserved elements to explore genomewide divergence between two species of Antilophia manakins, which are phenotypically distinct yet evidently lack strong genetic differentiation according to previous studies based on a limited number of loci. Our results revealed clear population structure that matches the two phenotypes, supporting the idea that smaller data sets lacked the power to detect this recent divergence event (likely <100 k ya). Indeed, we found little or no introgression between the species, as well as evidence of genomewide divergence. One implication of our study is that the Araripe plateau may be a hot spot of cryptic-diverging forest Cerrado populations. Besides their use in biogeography, subgenomic data sets may help redefine local conservation programmes by revealing cryptic population structure that may be key to population management.

Mitochondrial introgression obscures phylogenetic relationships among manakins of the genus Lepidothrix (Aves: Pipridae).

Lepidothrix is the most diverse genus of the family Pipridae, with eight recognized species. Although the genus' monophyly has been supported by both molecular and morphological characters, phylogenetic relationships and species limits within Lepidothrix remain uncertain. In the present study, we combined molecular sequences of mitochondrial (ND2 and COI) and nuclear (MYO, G3PDh and I5BF) markers in a multilocus analysis, to evaluate relationships and inter-specific limits among L. iris, L. nattereri, and L. vilasboasi, which are known to hybridize in eastern Amazonia. The results revealed a complex pattern, whereby events of secondary contact and gene flow after isolation and genetic and phenotypic differentiation prevented the recuperation of reciprocal monophyly among the studied taxa. The mitochondrial data indicate that L. nattereri is divided into two non-sister groups, one monophyletic, and the other, paraphyletic, with L. iris iris being more closely related to one of the two L. nattereri groups, while L. iris eucephala forms an undifferentiated clade with L. vilasboasi, probably resulting from an extensive process of mitochondrial introgression. In agreement with a previous study based on Single Nucleotide Polymorphism (SNP) data, mitochondrial haplotype networks also support that L. vilasboasi does not represent a recent "hybrid swarm" between L. iris and L. nattereri, but instead a genetically divergent lineage with a separate species status. Finally, the sister relationship recovered herein between L. iris iris and some western populations of L. nattereri currently in allopatry is also apparently explained by mitochondrial introgression, as also supported for nuclear genes by SNP data, indicating a complex scenario of past contact and gene flow between currently geographically distant Lepidothrix lineages.

Mutations in the melanocortin-1 receptor (MC1R) gene have no influence on the distinct patterns of melanic plumage found in the manakins of the genus Antilophia (Aves: Pipridae).

The melanocortin-1 receptor gene is the most widely-used marker for the investigation of the genetic determination of melanic plumage patterns. Studies of a number of wild bird species have shown an association between non-synonymous mutations of the MC1R gene and the presence of melanic variants. The genus Antilophia (Pipridae) includes only two manakin species (A. galeata and A. bokermanni), which are distinguished primarily by the differences in the pattern of melanic coloration of the plumage of the mantle in the adult males. In A. galeata, this plumage is black, while in A. bokermanni, it is predominantly white. This study investigates the possible association between mutations of the MC1R marker and the variation in plumage coloration observed in the two species. The MC1R sequences of the two species was analyzed, and the observed nucleotide variation was compared. Six polymorphic sites were identified, representing seven distinct genotypes. Five of these polymorphic mutations were non-synonymous, but were not related to the different phenotypes. Neutral evolution and the absence of any systematic association between the variants of the MC1R and plumage coloration in the Antilophia species indicate that alternative mechanisms regulate the expression of the coloration of the plumage in the adult males.

Molecular analyses reveal the occurrence of three new sympatric lineages of velvet worms (Onychophora: Peripatidae) in the eastern Amazon basin.

In the present study, we investigated the possible existence of new lineages of peripatids through comparisons between known Neotropical species and specimens obtained from two locations in Pará, a state in eastern Brazilian Amazonia using a molecular approach based on sequences of the mtDNA genes COI, 16Sr RNA, and 18S RNA. The analyses included also sequences of Asian and African taxa for a more systematic understanding of the phylogenetic relationships within the group. The analysis of the COI, 16S rRNA and 18S RNA sequences permitted the identification of three distinct lineages (A, B and C) based on two different phylogenetic approaches (Bayesian methods and ML). The three lineages presented here are completely distinct from all other peripatid taxa so far defined by molecular data. The presence of specimens of three independent onychophoran lineages occurring in sympatry in the Amazon basin was confirmed in all the analyses, providing consistent support for the phylogenies presented in this study. These findings reinforce the importance of the Amazon region in the diversification of Neotropical peripatids, and indicate that onychophoran diversity is much greater than previously thought, given that the number of taxa found at a single site was equivalent to the total number of allopatric species described for the entire region.

Molecular data indicate the presence of a novel species of Centropomus (Centropomidae - Perciformes) in the Western Atlantic.

Centropomus undecimalis is distributed in the coastal waters of the western Atlantic between North Carolina and São Paulo, although very little is known of the genetic structure of its populations. Here, 148 C. undecimalis samples were obtained from six sites in the southwestern Atlantic, representing the Brazilian distribution of this species. Segments of three mitochondrial (Cytb, COI and 16S) and one nuclear (IGF1) gene were sequenced. The results of all analyses indicated the presence of a previously undetected lineage of Centropomus in the northern extreme of Brazil (Amapá) in the region of the Oiapoque estuary. This taxon is genetically distinct from all 12 recognized species of Centropomus. The populations from the Brazilian states of Pará, Maranhão, Ceará, Rio Grande do Norte, Rio de Janeiro and São Paulo were genetically similar to C. undecimalis from coastal areas of the Caribbean and USA. Nucleotide divergence between C. undecimalis and the new Oiapoque taxon are greater than or similar to those found between a number of valid Centropomus species. The estimated time of divergence between C. undecimalis and the new taxon is approximately 2 millionyears. The findings of the present study emphasize the need for a thorough taxonomic revision of this genus.

Shorebirds' Longer Migratory Distances Are Associated With Larger ADCYAP1 Microsatellites and Greater Morphological Complexity of Hippocampal Astrocytes.

For the epic journey of autumn migration, long-distance migratory birds use innate and learned information and follow strict schedules imposed by genetic and epigenetic mechanisms, the details of which remain largely unknown. In addition, bird migration requires integrated action of different multisensory systems for learning and memory, and the hippocampus appears to be the integration center for this task. In previous studies we found that contrasting long-distance migratory flights differentially affected the morphological complexity of two types of hippocampus astrocytes. Recently, a significant association was found between the latitude of the reproductive site and the size of the ADCYAP1 allele in long distance migratory birds. We tested for correlations between astrocyte morphological complexity, migratory distances, and size of the ADCYAP1 allele in three long-distance migrant species of shorebird and one non-migrant. Significant differences among species were found in the number and morphological complexity of the astrocytes, as well as in the size of the microsatellites of the ADCYAP1 gene. We found significant associations between the size of the ADCYAP1 microsatellites, the migratory distances, and the degree of morphological complexity of the astrocytes. We suggest that associations between astrocyte number and morphological complexity, ADCYAP1 microsatellite size, and migratory behavior may be part of the adaptive response to the migratory process of shorebirds.

A new species of snook, Centropomus (Teleostei: Centropomidae), from northern South America, with notes on the geographic distribution of other species of the genus.

Centropomus irae sp. nov. (Centropomidae) is described from two localities in the state of Amapá, northern Brazil. The new species differs from all its congeners by the typically brown-colored lateral line, the relatively smaller mean interorbital width (4.1% of SL vs. 5.2-6.9%), mean snout length (8.9% of SL vs. 9.8-11.1%), mean diameter of the orbit (4.5% of SL vs. 5.4-7.3%), and mean mandible length (15.7% of SL vs.19.9-21.6%). The new species can also be distinguished from the similar Centropomus undecimalis by the number of scales around the caudal peduncle (18-23, usually 19-21, vs. 22-28, usually 24-27). With Centropomus irae sp. nov., seven species of the genus are now known to occur in the western Atlantic. The distribution of Centropomus in Brazilian waters is updated and a key to the species of this genus is presented. [Zoobank URL: urn:lsid:zoobank.org:act:3D497B38-48A5-4A6E-B377-6D57E82D4DE4[.

Low Genetic Diversity and Structuring of the Arapaima (Osteoglossiformes, Arapaimidae) Population of the Araguaia-Tocantins Basin.

The arapaima, Arapaima gigas, is a fish whose populations are threatened by both overfishing and the ongoing destruction of its natural habitats. In the Amazon basin, varying levels of population structure have been found in A. gigas, although no data are available on the genetic diversity or structure of the populations found in the Araguaia-Tocantins basin, which has a topographic profile, hydrological regime, and history of fishing quite distinct from those of the Amazon. In this context, microsatellite markers were used to assess the genetic diversity and connectivity of five wild A. gigas populations in the Araguaia-Tocantins basin. The results of the analysis indicated low levels of genetic diversity in comparison with other A. gigas populations, studied in the Amazon basin. The AMOVA revealed that the Arapaima populations of the Araguaia-Tocantins basin are structured significantly. No correlation was found between pairwise FST values and the geographical distance among populations. The low level of genetic variability and the evidence of restricted gene flow may both be accounted for by overfishing, as well as the other human impacts that these populations have been exposed to over the years. The genetic fragility of these populations demands attention, given that future environmental changes (natural or otherwise) may further reduce these indices and eventually endanger these populations. The results of this study emphasize the need to take the genetic differences among the study populations into account when planning management measures and conservation strategies for the arapaima stocks of the Araguaia-Tocantins basin.

Molecular confirmation of twinning in the West Indian Manatee (Trichechus manatus) / Confirmação molecular de gemelaridade em peixes-boi marinho (Trichechus manatus)

Abstract: Few twinning events have been recorded in the West Indian manatee (Trichechus manatus, Sirenia: Trichechidae) and no previous published study has provided confirmation of this phenomenon based in molecular tools. Here we investigate a possible case of twinning in an endangered Brazilian population of T. manatus using molecular tools. We analyzed two male neonates found stranded in Ceará State, on the northeastern coast of Brazil. The DNA of both individuals was isolated, and 10 microsatellite loci were amplified and genotyped. Following the identification of the alleles, the probabilities of identity by descent (∆7 and ∆8) and relatedness (rxy) were calculated using estimators that evaluate inbreeding. The two individuals shared most of the alleles, with differences in the genotypes being identified in only two loci. All the estimators identified a level of relatedness compatible with that found between siblings (selfed or outbred), indicating they were dizygotic twins. This is the first confirmed case of fraternal twins in free-ranging West Indian manatees in South America. The recognition of this type of twinning provides elements to improve actions for the rehabilitation of stranded animals and their subsequent release to the environment.

Resumo: Poucos eventos de gemelaridade foram registrados para o peixe-boi marinho (Trichechus manatus, Sirenia: Trichechidae) e nenhum estudo previamente publicado confirmou esse fenômeno com base em ferramentas moleculares. Aqui investigamos um possível caso de gemelaridade em uma ameaçada população brasileira de T. manatus utilizando ferramentas moleculares. Foram analisados dois neonatos machos encontrados encalhados no Ceará, costa nordeste do Brasil. O DNA dos indivíduos foi isolado e 10 loci microsatélites foram amplificados e genotipados. Após a identificação dos alelos, as probabilidades de identidade por descendência (∆7 e ∆8) e relação (rxy) foram calculadas usando estimadores que avaliam endogamia. Os dois indivíduos partilharam a maioria dos alelos, com diferenças nos genótipos sendo identificadas em apenas dois loci. Todos os estimadores identificaram um nível de parentesco compatível com o encontrado entre irmãos (com e sem endogamia), o que aponta para o fato de serem gêmeos dizigóticos. Este é o primeiro caso confirmado de gêmeos fraternais em peixes-boi marinho de vida livre na América do Sul. O reconhecimento deste tipo de gemelaridade fornece elementos para aprimorar ações que visem a reabilitação de animais encalhados e sua posterior soltura ao ambiente.

High levels of genetic connectivity among populations of yellowtail snapper, Ocyurus chrysurus (Lutjanidae-Perciformes), in the western South Atlantic revealed through multilocus analysis.

In the present study, five loci (mitochondrial and nuclear) were sequenced to determine the genetic diversity, population structure, and demographic history of populations of the yellowtail snapper, Ocyurus chrysurus, found along the coast of the western South Atlantic. O. chrysurus is a lutjanid species that is commonly associated with coral reefs and exhibits an ample geographic distribution, and it can therefore be considered a good model for the investigation of phylogeographic patterns and genetic connectivity in marine environments. The results reflected a marked congruence between the mitochondrial and nuclear markers as well as intense gene flow among the analyzed populations, which represent a single genetic stock along the entire coast of Brazil between the states of Pará and Espírito Santo. Our data also showed high levels of genetic diversity in the species (mainly mtDNA), as well a major historic population expansion, which most likely coincided with the sea level oscillations at the end of the Pleistocene. In addition, this species is intensively exploited by commercial fisheries, and data on the genetic structure of its populations will be essential for the development of effective conservation and management plans.

Dispersal capacity and genetic structure of Arapaima gigas on different geographic scales using microsatellite markers.

Despite the ecological and economic importance of the Arapaima gigas (Cuvier 1817), few data about its dispersal capacity are available. The present study was based on the analysis of microsatellite markers in order to estimate the dispersal capacity of the species on fine, meso, and large geographic scales. For this, 561 specimens obtained from stocks separated by distances of up to 25 km (fine scale), 100 km (meso scale), and 1300-2300 km (large scale) were analyzed. The fine scale analysis indicated a marked genetic similarity between lakes, with low genetic differentiation, and significant differences between only a few pairs of sites. Low to moderate genetic differentiation was observed between pairs of sites on a meso scale (100 km), which could be explained by the distances between sites. By contrast, major genetic differentiation was recorded in the large scale analysis, that is, between stocks separated by distances of over 1300 km, with the analysis indicating that differentiation was not related solely to distance. The genetic structuring analysis indicated the presence of two stocks, one represented by the arapaimas of the Mamirauá Reserve, and the other by those of Santarém and Tucuruí. The dispersal of arapaimas over short distances indicates a process of lateral migration within the várzea floodplains, which may be the principal factor determining the considerable homogeneity observed among the várzea lakes. The populations separated by distances of approximately 100 km were characterized by reduced genetic differentiation, which was associated with the geographic distances between sites. Populations separated by distances of over 1300 km were characterized by a high degree of genetic differentiation, which may be related primarily to historical bottlenecks in population size and the sedentary behavior of the species. Evidence was found of asymmetric gene flow, resulting in increasing genetic variability in the population of the Mamirauá Reserve.

Molecular analyses reveal the occurrence of three new sympatric lineages of velvet worms (Onychophora: Peripatidae) in the eastern Amazon basin

Abstract In the present study, we investigated the possible existence of new lineages of peripatids through comparisons between known Neotropical species and specimens obtained from two locations in Pará, a state in eastern Brazilian Amazonia using a molecular approach based on sequences of the mtDNA genes COI, 16Sr RNA, and 18S RNA. The analyses included also sequences of Asian and African taxa for a more systematic understanding of the phylogenetic relationships within the group. The analysis of the COI, 16S rRNA and 18S RNA sequences permitted the identification of three distinct lineages (A, B and C) based on two different phylogenetic approaches (Bayesian methods and ML). The three lineages presented here are completely distinct from all other peripatid taxa so far defined by molecular data. The presence of specimens of three independent onychophoran lineages occurring in sympatry in the Amazon basin was confirmed in all the analyses, providing consistent support for the phylogenies presented in this study. These findings reinforce the importance of the Amazon region in the diversification of Neotropical peripatids, and indicate that onychophoran diversity is much greater than previously thought, given that the number of taxa found at a single site was equivalent to the total number of allopatric species described for the entire region.

Early Pleistocene lineages of Bagre bagre (Linnaeus, 1766) (Siluriformes: Ariidae), from the Atlantic coast of South America, with insights into the demography and biogeography of the species

Coastal and marine environments are characterized by a lack of evident physical barriers or geographic isolation, and it may be difficult to understand how divergence can arise and be sustained in marine environments. The identification of 'soft' barriers is a crucial step towards the understanding of gene flow in marine environments. The marine catfishes of the family Ariidae are a demersal group with restricted migratory behavior, no pelagic larval stages, and mechanisms of larval retention, representing a potentially useful model for the understanding of historical processes of allopatric speciation in the marine environment. In the present study, two lineages of the Coco sea catfish, Bagre bagre , were recognized from their complete segregation at both mitochondrial and morphological levels. One lineage is distributed between Venezuela and the northern coast of Brazil, including the semiarid northeast coast, while the second lineage is found on the eastern coast of Brazil, including the humid northeast coast. Based on distribution area, habitats preference, and genetic variability, inferences are made in relation to biogeography and demography of lineages in Atlantic coast of South America.

Ambientes marinhos costeiros são caracterizados pela falta de barreiras físicas evidentes e ausência de isolamento geográfico, sendo difícil compreender como diferentes linhagens podem surgir e ser mantidas nestes ambientes. A identificação de barreiras permeáveis é um passo importante para a compreensão do fluxo gênico em ambientes marinhos. Os bagres marinhos da família Ariidae são um grupo demersal, com mecanismos de retenção larval e sem comportamento migratório ou estágio larval pelágico, representando um grupo potencialmente útil para a compreensão de processos históricos de especiação alopátrica no ambiente marinho. No presente estudo, duas linhagens do bandeirado ou bagre de fita, Bagre bagre , são reconhecidas com base na completa segregação mitocondrial e morfológica. Uma linhagem é encontrada da Venezuela à costa norte do Brasil, incluindo a costa nordeste semi-árida, enquanto a segunda linhagem é encontrada na costa leste e sudeste do Brasil, incluindo a costa nordeste úmida. Com base na área de distribuição, habitat preferencial e a variabilidade genética, inferências são feitas em relação à biogeografia e demografia das duas linhagens na costa do Atlântico da América do Sul.

Tocantins river as an effective barrier to gene flow in Saguinus niger populations

The Saguinus represent the basal genus of the Callitrichinae subfamily. Traditionally this genus is divided into three groups: Hairy, Mottled and Bare-face, however, molecular data failed to validate these groups as monophyletic units, as well as raised some subspecies to the species status. This is the case of the former subspecies Saguinus midas midas and S. midas niger, which are now considered as different species. In the present study, we sequenced a portion of the D-loop mtDNA region in populations from the East bank of the Xingu and from both banks of the Tocantins river, in order to test the effectiveness of large rivers as barriers to the gene flow in Saguinus. According to our results, the populations from the East and West banks of the Tocantins river are more divergent than true species like S. mystax and S. imperator. The Tocantins river may be acting as a barrier to gene flow, and consequently these very divergent populations may represent distinct taxonomic entities (species?).