Molecular phylogeny and species delimitation of the genus Schizodon (Characiformes, Anostomidae)

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Highlights

  • Schizodon is divided into two clades correlated with color pattern and biogeography.

  • Our analyses supported 13 consensus MOTUs distributed in 11 nominal species.

  • Six consensus MOTUs are in accordance with the current taxonomy.

  • Schizodon vittatus, S. nasutus and S. dissimilis were subdivided into two MOTUs.

  • S. platae and S. jacuiensis constituted a single MOTU, indicating a possible case of synonymy.

Abstract

The genus Schizodon is part of a group of headstanders and relatives (Family Anostomidae) that are widespread and ecologically important fishes in South American rivers. Schizodon includes 15 nominal species but their taxonomy has been challenging due to paucity of decisive characters to diagnose species. We present new molecular data to assess species boundaries or molecular operational taxonomic units (MOTUs), and to infer phylogenetic relationships among species. Evidence from two mitochondrial and three nuclear genes was used in these analyses. Mitochondrial DNA data for 112 specimens (from 11 nominal species) supported 13 consensus MOTUs, six of which matched valid nominal species (Schizodon borellii, S. fasciatus, S. intermedius, S. isognathus, S. knerii and S. scotorhabdotus). The nominal species Schizodon vittatus, S. nasutus, and S. dissimilis were subdivided into two MOTUs each, revealing either cryptic species or strong population structuring. In contrast, S. platae and S. jacuiensis constituted a single MOTU, indicating a possible case of synonymy. Our phylogenetic analysis subdivided the genus Schizodon into two large clades that are compatible with observed color patterns and biogeographic distribution. The first clade includes species with three to four conspicuous dark vertical bars on the flanks that originated in the Amazonas region (S. borellii, S. dissimilis, S. intermedius, S. fasciatus, S. scotorhabdotus, S. vittatus, and a cryptic species, Schizodon aff. vittatus). The second clade includes species with a conspicuous dark caudal blotch on the caudal peduncle, with vertical bars absent or inconspicuous, with a biogeographic origin in the La Plata drainage (S. isognathus, S. jacuiensis, S. knerii, S. nasutus and S. platae). Our results reinforce the importance of using molecular analyses to accelerate the study of diversity, particularly in groups with a wide distribution, few variable meristic characters, and high morphological plasticity, which may hide still unknown or underestimated diversity.

Introduction

The family Anostomidae includes a high diversity of freshwater fishes broadly distributed throughout South America. They are distinguished from other characiform fishes by the presence of a unique series of three to four teeth in each premaxillary or dentary section (Garavello and Britski, 2003). Anostomid species that attain large body sizes undertake characteristic spawning migrations along rivers and are exploited in commercial and subsistence fisheries (Garavello and Britski, 2003). The systematics and taxonomy of the Anostomidae have improved over the last decade (Ramirez et al., 2017b, Ramirez et al., 2016, Sidlauskas and Vari, 2008), with two new genera and more than 20 new species described. At the present time, Anostomidae consists of approximately 150 described species that are distributed over 15 genera, although hidden diversity at the species level is still suggested (Ramirez et al., 2017a, Ramirez et al., 2016, Ramirez and Galetti, 2015, Silva-Santos et al., 2018).

Schizodon Agassiz, 1829 is the second-most species-rich genus within the family, and it includes 15 nominal species. Schizodon is widespread in South America, occurring in almost all river basins draining to the east of the Andes, including the Amazonas, Orinoco, Paraná, Paraguay, Uruguay, Lagoa dos Patos, São Francisco, Tocantins, the northeastern Brazilian basins, and the Guianas coastal drainages, but it is absent from the isolated littoral basins of southeastern Brazil (Garavello and Britski, 2003, Sidlauskas and Vari, 2012). Garavello (1994) recognized four color patterns among the Schizodon species, which are considered as components of two hypothetical groups. The first is characterized by the presence of dark vertical brown bars on the flanks, sometimes combined with a longitudinal stripe or a macula at the base of the caudal peduncle. The second group is composed of species without vertical brown bars and with a conspicuous or inconspicuous stripe on the lateral line that masks the macula on the caudal peduncle.

The genus Schizodon has long been recognized by several diagnostic characters (Myers, 1950) and more recently by osteological synapomorphies (Sidlauskas and Vari, 2008), but delimitation of species within this genus has been somewhat problematic. Partially overlapping meristic counts and the relative stability of the color patterns among closely related species hampers identification of species within the genus (Garavello, 1994, Garavello and Britski, 1990). In view of this difficulty in identifying species, some of the most recently described species were delimited only after multivariate analyses of morphometric data (Garavello, 1994, Garavello and Britski, 1990). According to Reis et al. (Reis et al., 2016), the diversity of Neotropical ichthyofauna is underestimated primarily because of its phenotypic plasticity and cryptic species. In scenarios in which morphology-based identification is not sufficient, the use of molecular information can be helpful in delimitating species and indicating potential hidden diversity (e.g., Costa-Silva et al., 2015, Silva-Santos et al., 2018) as well as for constructing robust phylogenies (e.g., Oliveira et al., 2011, Ramirez et al., 2017b, Ramirez et al., 2016).

In the present study, we used a single-locus species delimitation approach with deep taxonomic sampling to identify the molecular operational taxonomic units (MOTUs) within Schizodon, and to provide new evidence to assess species richness in this genus. Additionally, two mitochondrial and three nuclear genes were used to reconstruct the evolutionary relationships within Schizodon and its phylogenetic position within Anostomidae. We also tested the two-groups hypothesis based on the presence or absence of vertical bars (Garavello 1994) by reconstructing the evolution of this trait on the molecular phylogeny. Finally, we performed an ancestral area reconstruction to study the biogeographic history of Schizodon.

Section snippets

Sampling

A total of 72 specimens from eleven of the fifteen valid Schizodon species from several hydrographic basins were sampled in this study (Table 1, Fig. 1, Fig. 2). Additionally, 40 sequences distributed over seven of these species that were obtained from the Barcode of Life Database platform (BOLD) were included (Table 1). The mean number of individuals represented per species was 10.2. Two species (S. jacuiensis and S. isognathus) were both represented by only one sample each, while S. nasutus

MOTU delimitation analyses

The final alignment of COI sequences resulted in 603 bp with 100 parsimony-informative sites. The shorter alignment length (compared with the amplified fragment) was due to the sequences included from the BOLD database. The PTP and bPTP analyses resulted in 13 MOTUs (Fig. 3), whereas the GMYC analysis resulted in 14 MOTUs, with a significant likelihood ratio test with P < 0.0001 (Fig. 3), and the BIN from the BOLD system clustered the sequences into 10 MOTUs. SPdel summarized the different

Discussion

The monophyly of Schizodon was strongly supported by our phylogenetic analyses based on two mitochondrial and three nuclear genes (Fig. 4). Morphologically, this genus is also supported on the basis of five synapomorphies, with special emphasis on the distinctive morphology of the teeth (Sidlauskas and Vari, 2008), with the primary characteristic being the presence of four multicuspid premaxillary and dentary teeth (Garavello, 1994).

The close relationship between Laemolyta, Schizodon and

Declaration of Competing Interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Acknowledgments

This work was supported by the Conselho Nacional de Desenvolvimento Cientifico e Tecnológico, CNPq (473474/2011-5 and 405309/2016-3 to P.M.G.J.), the SISBIOTA-Brazil Program (CNPq, 563299/2010-0; FAPESP, 10/52315-7), the Biota-Fapesp Program (FAPESP, 2011/50213-5 and 2018/04388-7), and the Consejo Nacional de Ciencia Tecnología e Innovación Tecnológica, CONCYTEC-FONDECYT (Proyecto Investigación Básica 2019-01, 431-2019-FONDECYT to J.L.R.). J.L.R had a fellowship from the Fundação de Amparo à

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