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The Scientific World Journal
Volume 2019, Article ID 6507954, 8 pages
Research Article

Morphological and Genetic Structure of Two Equivalent Astyanax Species (Characiformes: Characidae) in the Region of Paranaíba Arc

1Laboratory of Ecological and Evolutionary Genetics, Institute of Biological and Health Sciences, Federal University of Viçosa, Campus Rio Paranaíba, BR 354, Km 310, Campus Universitário, Post Code: 38810-000, Rio Paranaíba, MG, Brazil
2Graduate Program in Management and Conservation of Natural and Agricultural Ecosystems, Federal University of Viçosa, Campus Florestal, Rodovia LMG 818, Km 06 Post Code: 35690-000, Florestal, MG, Brazil

Correspondence should be addressed to Renan Rodrigues Rocha; rb.vfu@seugirdor.r.naner

Received 30 November 2018; Accepted 28 February 2019; Published 17 April 2019

Academic Editor: Lukas Kratochvil

Copyright © 2019 Renan Rodrigues Rocha et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.


The Astyanax scabripinnis complex is composed of a large number of almost morphological indistinguishable species, including Astyanax paranae and Astyanax rivularis, which exist in the Paraná and São Francisco Basins, respectively, and sometimes are considered subspecies of the A. scabripinnis group or even are cited just as A. scabripinnis. The two river basins are separated by the Upper Paranaíba Arc, likely the main cause of the isolation of these species. We used geometric morphometric tools and DNA analyses of populations of both species to identify the differences between them. Geometric morphometrics separated the two species into distinct groups, whose main difference was the body depth. This is generally related to the speed of the water flow in the river basins. The maximum likelihood phylogram based on mitochondrial DNA sequences formed two main clades: one composed of the population of A. rivularis and the other, of A. paranae. In the haplotype network, the species were similarly separated into two groups from the same ancestral haplotype, with A. rivularis dispersing into two lineages in the São Francisco River Basin. The distribution of A. paranae is a consequence of a secondary dispersion event in the Paraná River Basin. It forms two lineages from a haplotype derived from the ancestor. The vicariant effect of separate basins, through the elevation of the Upper Paranaíba Arc, led to the allopatric speciation of the populations originating the present species. The results of geometric morphometrics and molecular data of the fish show the importance of this geological event in the biogeography and evolutionary history of the ichthyofauna of the region and indicate that the isolation of these species seems to be effective.

1. Introduction

The genus Astyanax Baird and Girard, 1854, is composed of fishes popularly known as piabas or lambaris. The genus belongs to the Characidae family and has about 160 species distributed on the Neotropical region watershed [1].

The species complex Astyanax scabripinnis is an example of the morphological and genetic diversity of the genus. Previously considered as a single species, Moreira Filho and Bertollo [2], proposed that it was a species complex, based on variations found in cytogenetic and phenotypic characteristics. Fishes of the complex usually inhabit the headwater of rivers and small streams [3], which results in isolated populations that could be driven to allopatric speciation [2].

The fishes of the complex are widely distributed through large hydrographic basins, as Paraná River basin and São Francisco River basin [4]. These two hydrographic basins were separated by the uplift of the Upper Paranaíba Arc, which increased the degree of isolation of the existent populations [5]. As a representative species from São Francisco River basin, we have A. rivularis; meanwhile from Upper Paraná river basin A. paranae can be found, with low tendency to migration [6]. Astyanax rivularis and A. paranae are strongly related [7] and in spite of belonging to different basins these two species do have several ecological, morphological, and genetic similarities.

Delimiting related species or species within a complex is a hard task that demands studies on several areas to achieve a consensus on the procedure to separate them [8]. Despite the attempt to revise the Astyanax scabripinnis group, the taxonomists have difficulties to determine the nominal identification by just analyzing the samples. Molecular and morphological analyses could be some of the tools used for this delimitation [9].

Geometric morphometrics studies are efficient to demonstrate morphological differences between species within a species complex [10, 11], while phylogenetic and phylogeographic analyses based on mitochondrial DNA (mtDNA) allow elucidating evolutionary relationships and divergence of organisms [9].

Astyanax paranae and A. rivularis represent equivalent species formerly united under the A. scabripinnis complex that are distributed along an important watershed separating two major hydrographic basins. The present work aims to evaluate the morphological and/or genetic structuring among and within populations in adjacent region of the distribution of these two species.

2. Material and Methods

2.1. Specimens Sampling

The individuals of A. paranae were collected in Paranaíba and São João rivers and Água Grande and Lava Pés streams, belonging to Upper Paraná river basin. Astyanax rivularis samples were collected from Do Boi, Borrachudo, and Abaeté rivers, and Tiros and Vereda Grande streams, along the region of Upper São Francisco (Figure 1). All the specimens were deposited in the collection of Laboratory of Ecological and Evolutionary Genetics at the Federal University of Viçosa (UFV), campus Rio Paranaíba. The sampling of the specimens was carried out in accordance with SISBIO, Sistema de Autorização e Informação em Biodiversidade (license number 1938128), and SISGEN, Sistema Nacional de Gestão do Patrimônio Genético e do Conhecimento Tradicional Associado (license number A9FE946). The euthanizing was carried out according to the recommendations of the Conselho Nacional de Controle de Experimentação Animal of Brazil (CONCEA).

Figure 1: Sampling sites of specimens of this study. The locations numbered from 1 to 5 belong to the Paranaíba River basin; locations 6 to 10 belong to São Francisco River Basin. 1: São João river; 2: Paranaíba River; 3: Lava Pés stream; 4: Parque de Exposições stream; 5: Água Grande stream; 6: Abaeté river; 7: Tiros stream; 8; Borrachudo river; 9: Boi river; 10: Vereda Grande stream.
2.2. Geometric Morphometrics

Forty-four individuals of A. paranae and forty-eight of A. rivularis were used for morphometrics analysis. The specimens were photographed with the use of a Sony Cyber-Shot camera, 14.1-megapixel resolution and 4x zoom. The software TPSUtil 1.6 [12] was used to group and format the data at a suitable file. Fourteen anatomic landmarks were selected to represent the general body shape of the fishes (Figure 2).

Figure 2: Analyzed landmarks: 1: rostrum; 2: anterior insertion; 3: posterior insertion of dorsal fin; 4: posterior insertion of adipose fin; 5: superior insertion of the first ray of caudal fin; 6: inferior insertion of final ray of caudal ray; 7: posterior insertion; 8:anterior insertion of anal fin; 9: insertion of ventral fin; 10: insertion of pectoral fin; 11: inferior limit; 12: superior limit of operculum; 13: anterior limit; 14: posterior limit of ocular orbit.

The landmarks were digitized with the use of the TpsDig 2.26 software system [13]. The transformation of data from the matrix by procrustes superposition was conducted with the aid of the PAST v2.17 software system [14], aiming to delete errors of scale, orientation, and position. The differences observed resulted only from shape variation [15, 16]. The difference between body shapes over species was determined by analyzing the canonical variables associated with Multivariate Analysis of Variance/Canonical Variance Analysis (MANOVA/CVA) [17], with the inference of consensus shape for each species on software MorphoJ 1.18 [18].

2.3. Phylogenetic Analysis

Twenty A. paranae individuals from the three populations of Paranaíba River Basin (Paranaíba and São João rivers, and Água Grande stream) and twenty-four A. rivularis individuals from São Francisco River Basin (Boi, Borrachudo, and Abaeté rivers; Tiros and Vereda Grande streams) were used for molecular analysis.

DNA was isolated from liver and heart samples of each specimen following the commercial protocol of the PureLink Genomic DNA kit by Invitrogen. The samples were quantified by electrophoresis on 1% agarose gel and Low Mass DNA ladder by Invitrogen, with posterior dilution to the final concentration of 10 ng/μL of DNA. The amplification of Cytochrome b gene was carried out using primers H16460 - 5’CGAYCTTCGGATTACAAGAC3’ and GluDG.L - 5’TGACCTGAARAACCAYCGTT3’ [19]. The mitochondrial DNA from at least 3 individuals from each population was amplified and sequenced resulting in a fragment with 670 bp. The amplified sequences from Lava-Pés and Parque de Exposição streams were shorter and of poor quality than the others used in this work and therefore were excluded from further analysis. The PCR was conducted on a thermocycler with 25μL reaction tubes containing 2,5μL PCR 10X buffer, 1μL MgCl2 in 50mM, 1,5μL 10mM dNTP mix, 1μL of each primer, 0,2μL Taq polymerase, 5μL of DNA template, and 12,8μL of ultrapure water. The thermocycler program started at an initial step of 94°C for 4 minutes, followed by 35 cycles of 15 seconds at 94°C, 30 seconds at 56°C, and 2 minutes at 72°C, with a final extension of 10 minutes at 72°C, and was kept at 4°C [20].

The sequences obtained were aligned with ClustalW v1.6 algorithm [21] at the MEGA v6.06 [22]. The evolutionary model calculated by the software was HKY+G, with which the maximum likelihood phylogram was generated with 1000 replications. The p-distance between the sequences of the two species and the subsequently groupings was also calculated. The MEGA v6.06 software system [22], DnaSP 5.10 [23], and Network v4.6 were used to build the Cytochrome b haplotype network according to Median Joining Algorithm [24].

3. Results

3.1. Geometric Morphometrics

The multivariate analysis (MANOVA/CVA) for the species showed shape differences between them (Wilke’s lambda: 0.03236; df1 = 84; f = 4.69; p < 0.0001). The canonical axes CV1 explained 56.4% and CV2, 31% of the variation (Figure 3(a)). Alternatively, according to the molecular data, four groups were analyzed and showed morphological structuring (Wilke’s lambda: 0.01105; df1 = 112; f = 4.01; p < 0.0001) with canonical axes CV1 explaining 60.1% and CV2, 22.1% of the variation (Figure 3(b)). The consensus shapes for each species are presented in Figure 4. Astyanax paranae’s shape shows a higher body depth, while A. rivularis’ shape presented a lower body depth and longitudinal elongation, when compared to the first one.

Figure 3: (a) Score position of MANOVA/CVA for each species, at the space of the first and second canonical axes. Red dots: Astyanax paranae, blue dots: Astyanax rivularis. (b) Score position of MANOVA/CVA for each lineage, at the space of the first and second canonical axes. Green dots: lineage A1, blue dots: lineage A2, purple dots: lineage B1, and red dots: lineage B2.
Figure 4: Consensus shape for each species. (a) Astyanax rivularis; (b) Astyanax paranae. The dot represents the landmarks and the trace of the direction where the landmarks are moving in the deformation grid.
3.2. Phylogenetic Analysis

There were two main clades in the phylogram. One of them (clade A) is composed exclusively of A. rivularis individuals, while the other (clade B) is composed of A. paranae. Clades A and B presented two subclades each. Individuals from Borrachudo river were shared by the two A. rivularis subclades. Clade B1 was composed only of individuals from São João River, and clade B2 was composed of the remaining populations from Paranaíba basin (Figure 5).

Figure 5: Maximum likelihood phylogram built by MEGA v6.06. The values at each node are the bootstrap for 1000 replications. The numbers are equivalent to the sampling sites of Figure 1:1, São João river; 2, Paranaíba River; 5, Água Grande stream; 6, Abaeté river; 7, Tiros stream; 8, Borrachudo river; 9, Boi river; 10, Vereda Grande stream.

According to the results found on phylogenetic analysis and haplotype network, each species was separated on two groups from which the genetic distances between and within them were calculated. The groups match exactly with the clades observed on the phylogenetic analysis, so they were named as seen on there. The distances from the species as a whole were also calculated in 0.025. The results are shown in Table 1.

Table 1: Genetic distances between clades of Astyanax rivularis (A1, A2, and A3) and Astyanax paranae (B1 and B2). The values on the diagonal are genetic distances within each clade.
3.3. Haplotype Network

The haplotype network generated by the software system showed two extremities. Each one was from one species, connected at the center by the missing haplotype mv2 (Figure 6). Twelve haplotypes were identified, six from each species, with 22 variable sites of Cytochrome b gene. The A. rivularis’ extremity presented a bifurcation originated from haplotype mv2. Except for haplotype #1, represented by four individuals from Tiros stream, and haplotype #6, represented by only one individual from Do Boi River, the remaining haplotypes were represented by individuals from mixed populations.. In A. paranae, São João river specimens were represented by haplotypes #7 and #8, while haplotypes #9, #10, and #11 represent individuals from Paranaíba River and Água Grande stream. The haplotype #12 is present in one individual of Paranaíba River.

Figure 6: Haplotype network. Each line indicates a mutational event (step). The distance between each node is proportional to the number of mutational steps. The black circles are the missing haplotypes. Sampled populations: 1, Borrachudo River (yellow); 2, Tiros Stream (pink); 3, Vereda Grande stream (light green); 4, Do Boi River (dark blue); 5, Abaeté river (purple); 6, Água Grande stream (brown); 7, Paranaíba river (light blue); 8, São João river (red).

4. Discussion

The results showed that the species under study have visible morphological differentiation and genetic structuration. Astyanax paranae has a fusiform body, compared to A. rivularis, with a typical shape of higher water flow environments [25]. A. rivularis presents lower body depth in addition to a longitudinal elongation when compared to A. paranae. Such characteristics are usually associated to environments with lower speed water [25, 26]. According to Atlas das Águas [27], the average water speed at the sampling sites for the Upper Paranaíba populations is higher than that found in the São Francisco River Basin region, which matches with the body form variations found (Figure 4).

These data expand that one by Moreira-Filho and Bertollo [2], who proposed that species of A. scabripinnis complex would be morphologically adapted into a wide range of environments. Recent studies reinforce the great morphological plasticity of the genus and demonstrate that different species in the same environment could present distinct phenotypic adaptations [28, 29]. Thus, divergent evolutionary mechanisms at each basin could change the adaptive response, since environmental exploration and interaction with other species affect character selection, habitat colonization, and, hence, genes [30]. This could accelerate the evolutionary process that generates diversification of characters, even driving to allopatric speciation [31], since different genes tend to fix in different populations due to the particular selective pressures of each habitat.

The phylogenetic tree of the local tetras indicates a clear distinction between the two species, with the formation of a clade composed only by A. rivularis (clade A) and other composed only by A. paranae (clade B). The calculated 2,5% genetic distance corroborates the idea of different species, since April et al. [32] established 2% as the threshold of difference on DNA base composition from fish mitochondrial DNA to sustain the separation of distinct species.

In both clades, the relationships found are consistent with the geographic distribution of the rivers in which the individuals were collected. For A. rivularis (clade A), the diversification of populations seems to be more recent, due to lower structuration of the populations. Subclade A1 isrepresented by nearby and interconnected watercourses, which explain the clustering observed between them. However, between populations of subclade A2, there is no connection relationship between the rivers. Nevertheless, there is a geographical proximity at some points along the rivers, which would mean that the two initially isolated populations may have some contact, in case of floods, a common occurrence in the region [33], or even due to human intervention. The structuring of populations is more evident among the A. paranae (clade B), which demonstrates physical isolation between them. Subclade B1 is composed of an isolated river, which reflects on the structuration observed. Subclade B2 groups two distinct populations, which come from nearby and connected water bodies, since Água Grande stream is a tributary of the Paranaíba River. Despite the geographical proximity between these rivers, unlike the observed in São Francisco River Basin, fauna changes due to floods are improbable, since the hydromorphological characteristics of the basin do not support such event [34].

The haplotype network coincided with the results found in the phylogram of maximum likelihood, with two extremities, each one composed of only one species, which matches the geographic distribution of the basins. Besides the existence of three clades for A. rivularis on the phylogram, only two lineages were found for the species. The lineage B corresponds to clades A1 and A2, whose genetic distance is only 0.4%. That fact, in addition to shared mutational steps on same sites, as evidenced in Figure 6, may explain why there are only one lineage for these two different clades. Haplotype mv2 can be seen at the center of the network and could not be collected or may be extinct. According to Coalescent theory [35], mv2 would probably be the ancestral haplotype, i.e., the one that existed before the separation of the basins. Thus, we can infer that the process that drives the establishment and evolution of the ancestral haplotype was different in each basin.

The establishment of A. rivularis at the basin probably occurred with the dispersion of two lineages, A1 and A2, from the ancestral haplotype. The presence of individuals from Borrachudo River in both lineages could indicate the occurrence of homoplasy in the sequences, probably due to the geometry of the network [35] or maintenance of the ancestral haplotype [36]. However, in A. paranae, firstly there was the establishment of descendent haplotype mv1, derived from ancestor mv2. After this initial event, a secondary dispersion occurred along the basin, which also resulted in two lineages, B1 and B2, from the studied populations (Figure 6). Although lineages within each species appear to be divergent, most notably at A. rivularis case, the clustering found in the maximum likelihood phylogram and MANOVA/CVA and the low genetic distances between them indicate that there are only two species involved at the analysis.

The populations of both species, physically separated, may have passed through different evolutionary events, which led to the genetic and morphologic differentiation observed. Reduction or interruption of gene flow among them, due to isolation, leads to the accumulation of unique changes of the evolutionary history of each species. That fact, associated with the observed differentiation along with natural process of genetic drift and natural selection [37], leads us to believe in allopatric speciation as the responsible for the origin of species.

One of the main phenomena that affect this type of speciation is the vicariance [38], which occurred with the A. rivularis and A. paranae populations. The Upper Paranaíba Arc separates São Francisco River Basin from the Paraná River Basin, since a rock elevation emerged at Meso/Neocretaceous [5]. The rising of the elevation may have led to the isolation of species previously shared by both basins, acting as a vicariant event to them [39]. Different aspects of each basin, i.e., altitude, average speed of water, fauna, and flora composition, may have been agents to the diversification between the two species. Other works on species of the genus suggest the same board of speciation after the uplift of Upper Paranaíba Arc [40].

5. Conclusions

Although the identification of these species is often difficult due to the absence of diagnostic morphological characters, the genetic and morphological data shows lines of evidence that populations of Astyanax rivularis and Astyanax paranae are not intermixed, hence being different evolutionary units. The uplift of the Paranaíba Arc seems to be the main reason for this, reinforcing vicariance and allopatry roles in the evolution of the Astyanax genus and the strict relation between natural formation of hydrographic basins and their inhabitant ichthyofauna.

Data Availability

The DNA sequence data used to support the findings on this study have been deposited in the GenBank nucleotide repository under the accession numbers MK756216 to MK756259

Conflicts of Interest

The authors declare that there are no conflicts of interest regarding the publication of this paper.


The development of this work was supported by fellowships granted by Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq), Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) and Fundação de Amparo à Pesquisa de Minas Gerais (FAPEMIG, grant FORTIS-10254/2014), conceded to the Graduate Program in Management and Conservation of Natural and Agricultural Ecosystems (MCENA). The authors are grateful to the coworkers of the Laboratory of Ecological and Evolutionary Genetics for helping with the methodology and software analysis, especially Marcos Aurélio Silva.


  1. W. N. Eschmeyer and J. D. Fong, “Species by family/subfamily. Catalog of Fishes electronic version,”, [Accessed 02 August 2017].
  2. O. Moreira-Filho and L. A. Carlos Bertollo, “Astyanax scabripinnis (Pisces, Characidae): A species complex,” Revista Brasileira de Genética, vol. 14, no. 2, pp. 331–357, 1991. View at Google Scholar · View at Scopus
  3. A. H. Britski, “Peixes de água doce do estado de são paulo: sistemática,” in Poluição e Piscicultura, vol. 9, 1972,, [Accessed 9 August 2017]. View at Google Scholar
  4. V. A. Bertaco and C. A. Lucena, “Two new species of Astyanax (Ostariophysi: Characiformes: Characidae) from eastern Brazil, with a synopsis of the Astyanax scabripinnis species complex,” Neotropical Ichthyology, vol. 4, no. 1, pp. 53–60, 2006. View at Publisher · View at Google Scholar
  5. J. E. G. Campos and M. A. Dardenne, “Origem e evolução tectônica da bacia sanfranciscana,” Revista Brasileira de Geociências, vol. 27, no. 3, pp. 283–294, 1997. View at Publisher · View at Google Scholar
  6. H. I. Suzuki, A. E. A. M. Vazzoler, E. E. Marques, M. A. P. Lizama, and P. Inada, “Reproductive ecology of the fish assemblage,” in The Upper Paraná River and its Floodplain: Physical Aspects, Ecology and Conservation, S. M. Thomaz, A. A. Agostinho, and N. S. Hahn, Eds., pp. 271–292, Leiden: Backhuys Publishers, 2004. View at Google Scholar
  7. B. C. Rossini, C. A. Oliveira, F. A. Melo et al., “Highlighting Astyanax species diversity through DNA barcoding,” PLoS ONE, vol. 11, no. 12, p. e0167203, 2016. View at Publisher · View at Google Scholar
  8. K. De Queiroz, “Species concepts and species delimitation,” Systematic Biology, vol. 56, no. 6, pp. 879–886, 2007. View at Publisher · View at Google Scholar
  9. J. P. Castro, M. O. Moura, O. Moreira-Filho et al., “Diversity of the Astyanax scabripinnis species complex (Teleostei: Characidae) in the Atlantic Forest, Brazil: species limits and evolutionary inferences,” Reviews in Fish Biology and Fisheries, vol. 25, no. 1, pp. 231–244, 2015. View at Publisher · View at Google Scholar
  10. C. P. Klingenberg, M. Barluenga, and A. Meyer, “Body shape variation in cichlid fishes of the Amphilophus citrinellus species complex,” Biological Journal of the Linnean Society, 2003. View at Publisher · View at Google Scholar
  11. C. Clabaut, P. M. E. Bunje, W. Salzburger, and A. Meyer, “Geometric morphometric analyses provide evidence for the adaptive character of the Tanganyikan cichlid fish radiations,” Evolution, vol. 61, no. 3, pp. 560–578, 2007. View at Publisher · View at Google Scholar · View at Scopus
  12. F. J. Rohlf, tpsUtil version 1.6, Department of Ecology and Evolution, State University of New York at Stony Brook, 2013.
  13. F. J. Rohlf, tpsDig, version 2.26, Department of Ecology and Evolution, State University of New York, Stony Brook, 2015.
  14. Ø. Hammer, D. A. T. Harper, and P. D. Ryan, “PAST - Palaeontological statistics,” 2001,, [Accessed 02 August 2017].
  15. C. P. Klingenberg, “Morphometrics and the role of the phenotype in studies of the evolution of developmental mechanisms,” Gene, vol. 287, no. 1-2, pp. 3–10, 2002. View at Publisher · View at Google Scholar
  16. F. L. Bookstein, Morphometric Tools for Landmark Data: Geometry and Biology, Cambridge University Press, 1991. View at MathSciNet
  17. C. P. Klingenberg, M. Barluenga, A. Meyer, and P. Wainwright, “Shape analysis of symmetric structures: quantifying variation among individuals and asymmetry,” Evolution, 2002. View at Publisher · View at Google Scholar
  18. C. P. Klingenberg, “MorphoJ: an integrated software package for geometric morphometrics,” Molecular Ecology Resources, vol. 11, no. 2, pp. 353–357, 2011. View at Publisher · View at Google Scholar · View at Scopus
  19. A. Perdices, E. Bermingham, A. Montilla, and I. Doadrio, “Evolutionary history of the genus Rhamdia (Teleostei: Pimelodidae) in Central America,” Molecular Phylogenetics and Evolution, 2002. View at Publisher · View at Google Scholar
  20. S. M. Prioli, A. J. Prioli, H. F. Júlio Jr. et al., “Identification of Astyanax altiparanae (Teleostei, Characidae) in the Iguaçu River, Brazil, based on mitochondrial DNA and RAPD markers,” Genetics and Molecular Biology, vol. 25, no. 4, pp. 421–430, 2002. View at Publisher · View at Google Scholar
  21. J. D. Thompson, D. G. Higgins, and T. J. Gibson, “CLUSTAL W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice,” Nucleic Acids Research, vol. 22, no. 22, pp. 4673–4680, 1994. View at Publisher · View at Google Scholar · View at Scopus
  22. K. Tamura, G. Stecher, D. Peterson, A. Filipski, and S. Kumar, “MEGA6: molecular evolutionary genetics analysis version 6.0,” Molecular Biology and Evolution, vol. 30, no. 12, pp. 2725–2729, 2013. View at Publisher · View at Google Scholar
  23. P. Librado and J. Rozas, “DnaSP v5: a software for comprehensive analysis of DNA polymorphism data,” Bioinformatics, 2009. View at Publisher · View at Google Scholar
  24. H.-J. Bandelt, P. Forster, and A. Röhl, “Median-joining networks for inferring intraspecific phylogenies,” Molecular Biology and Evolution, vol. 16, no. 1, pp. 37–48, 1999. View at Publisher · View at Google Scholar · View at Scopus
  25. L. Breda, E. F. Oliveira, and E. Goulart, “Ecomorfologia de locomoção de peixes com enfoque para espécies neotropicais,” Acta Scientiarum. Biological Sciences, vol. 27, no. 4, 2005. View at Publisher · View at Google Scholar
  26. P. W. Webb, “Form and function in fish swimming,” Scientific American, vol. 251, no. 1, pp. 72–83, 1984. View at Google Scholar
  27. Atlas digital das águas de Minas, “Panorama Hídrico – Regiões hidrográficas – UPGRH,”, [Accessed 02 August, 2017].
  28. F. T. Mise, R. Fugi, J. P. A. Pagotto, and E. Goulart, “The coexistence of endemic species of Astyanax (Teleostei: Characidae) is propitiated by ecomorphological and trophic variations,” Biota Neotropica, vol. 13, no. 3, pp. 21–28, 2013. View at Google Scholar · View at Scopus
  29. M. A. Souza, D. C. Fagundes, C. G. Leal, and P. S. Pompeu, “Ecomorphology of Astyanax species in streams with different substrates,” Zoologia, 2014. View at Publisher · View at Google Scholar
  30. A. A. Agrawal, “Phenotypic plasticity in the interactions and evolution of species,” Science, vol. 294, no. 5541, pp. 321–326, 2001. View at Publisher · View at Google Scholar · View at Scopus
  31. M. J. West-Eberhard, “Phenotypic plasticity and the origins of diversity,” Annual Review of Ecology, Evolution, and Systematics, vol. 20, no. 1, pp. 249–278, 1989. View at Publisher · View at Google Scholar
  32. J. April, R. H. Hanner, A. Dion-Côté, and L. Bernatchez, “Glacial cycles as an allopatric speciation pump in north-eastern American freshwater fishes,” Molecular Ecology, vol. 22, no. 2, pp. 409–422, 2013. View at Publisher · View at Google Scholar
  33. Semed Secretaria de Estado de Meio Ambiente e Desenvolvimento Sustentável, “Atlas de vulnerabilidade às inundações de Minas Gerais,” Belo Horizonte, MG: Secretaria de Estado de Meio Ambiente e Desenvolvimento Sustentável, 2015.
  34. A. M. Veiga, C. C. P. dos Santos, M. R. D. Cardoso, and N. C. Lino, “Caracterização hidromorfológica da bacia do rio meia ponte,” Caminhos de Geografia, vol. 14, no. 46, 2013. View at Google Scholar
  35. D. Posada and K. A. Crandall, “Intraspecific gene genealogies: trees grafting into networks,” Trends in Ecology & Evolution, vol. 16, no. 1, pp. 37–45, 2001. View at Publisher · View at Google Scholar
  36. D. J. Funk and K. E. Omland, “Species-level paraphyly and polyphyly: frequency, causes, and consequences, with insights from animal mitochondrial DNA,” Annual Review of Ecology, Evolution and Systematics, vol. 34, no. 1, pp. 397–423, 2003. View at Publisher · View at Google Scholar
  37. C. J. Hoskin, M. Higgie, K. R. McDonald, and C. Moritz, “Reinforcement drives rapid allopatric speciation,” Nature, vol. 437, no. 7063, pp. 1353–1356, 2005. View at Publisher · View at Google Scholar
  38. D. E. Rosen, “Vicariant Patterns and Historical Explanation in Biogeography,” Systematic Zoology, vol. 27, no. 2, p. 159, 1978. View at Publisher · View at Google Scholar
  39. N. Hubert and J. Renno, “Historical biogeography of South American freshwater fishes,” Journal of Biogeography, 2006. View at Google Scholar
  40. R. Pazza, L. A. Cruvinel, and K. F. Kavalco, “Parallel evolution evidenced by molecular data in the banded-tetra (Astyanax fasciatus),” Biochemical Systematics and Ecology, vol. 70, pp. 141–146, 2017. View at Publisher · View at Google Scholar