A new genus of Anostomidae (Ostariophysi: Characiformes): Diversity, phylogeny and biogeography based on cytogenetic, molecular and morphological data

Highlights

• Megaleporinus is described to include ten valid extant species.

• Megaleporinus is supported by morphological, cytogenetical and molecular data.

• At least four undescribed, cryptic species, were identified.

• Synapomorphies include a ZW sex chromosome system.

• Paleogeographic and hydrogeological events have likely driven Megaleporinus speciation.

Abstract

A new genus of Anostomidae (Characiformes) is described to include ten valid extant species previously classified in Leporinus or Hypomasticus and distributed throughout most major river basins in South America: L. brinco, L. conirostris, L. elongatus, H. garmani, L. macrocephalus, L. muyscorum, L. obtusidens, L. piavussu, L. reinhardti, and L. trifasciatus. The monophyly of Megaleporinus is well-supported in a phylogenetic analysis based on two mitochondrial and three nuclear genes, as well as its sister group relationship to Abramites. Megaleporinus is diagnosed by having the exclusive combination of three unicuspid teeth on each premaxillary and dentary bone and a color pattern composed of one to four dark midlateral blotches. Additional distinguishing features and possible synapomorphies include a unique ZZ/ZW sex chromosome system confirmed for six congeners and a drumming apparatus wherein the first rib is elongated and associated with hypertrophied intercostal muscles, which was confirmed for three congeners as exclusive to mature males. Furthermore, our study identified at least four undescribed cryptic species, emphasizing the need for further taxonomic work and genetic analyses. A time-calibrated phylogenetic and biogeographical analysis of the new genus suggests that speciation in the proto-Amazon-Orinoco lineage was primarily driven by paleogeographic processes, such as the formation of the Orinoco and Tocantins basins. Dispersal and diversification of the genus in coastal basins draining the Eastern Brazilian Shield appears to have been facilitated by connections between paleo-basins during low sea level periods and headwater captures between coastal and inland watersheds. The present contribution demonstrates the importance of integrating data from morphology, DNA sequences and cytogenetics to advance the taxonomy and systematics of any complex species group.

Introduction

The neotropical freshwater ichthyofauna is perhaps the most diverse in the world but remains poorly known, as the total number of extant species is uncertain (Reis et al., 2016). In the last three decades, molecular data have significantly expanded our knowledge of neotropical fish diversity (Albert and Reis, 2011, Pereira et al., 2013). Analyses of mitochondrial and nuclear genes have added missing pages to the evolutionary history of many large and complex neotropical fish groups (e.g., Oliveira et al., 2011, Roxo et al., 2014, Thompson et al., 2014, Ramirez et al., 2016). In addition, DNA barcoding has disclosed taxonomic uncertainties and hidden diversity within the major groups of neotropical fishes (Pereira et al., 2013, Ramirez and Galetti, 2015).

The family Anostomidae is broadly distributed in South America and comprises approximately 150 described species distributed in 14 genera (Garavello and Britski, 2003, Sidlauskas and Vari, 2008). Leporinus is the most species-rich genus with approximately 90 valid species (Birindelli et al., 2013, Burns et al., 2014) showing wide morphological diversity, especially in snout shape, mouth position and dentition (Sidlauskas, 2007, Sidlauskas and Vari, 2008).

The taxonomy of Anostomidae was substantially improved by Myers (1950), who defined eight genera (Leporellus, Gnathodolus, Synaptolaemus, Rhytiodus, Leporinus, Abramites, Schizodon, and Laemolyta) based on tooth morphology and mouth position to allocate the 75 species known at the time (Fowler, 1950). Myers (1950) considered previous attempts to split Leporinus to be inconsistent, as for example Myocharax (created for L. desmotes) and Hypomasticus (created for species with a distinctly ventral mouth such as H. mormyrops) (Borodin, 1929, Fowler, 1914). The only comprehensive taxonomic assessment of Leporinus was performed by Garavello (1979), who defined species groups within Leporinus based on coloration patterns. Garavello’s groups were adopted by some authors to aid in species-level identification (e.g., Sidlauskas and Vari, 2012), even though most of them proved to be artificial units (Sidlauskas and Vari, 2008). Only two comprehensive morphological studies have addressed the evolutionary history of Anostomidae, one focused on small-sized species with an upturned mouth, previously assigned to Anostominae (Winterbottom, 1980), and another focused on the familial relationships within the broader Anostomoidea (Sidlauskas and Vari, 2008). The latter study recovered Leporinus as paraphyletic, and possibly including Abramites. A study based on molecular data (Ramirez et al., 2016) obtained similar results with species of Leporinus interleaved among other genera (Abramites, Hypomasticus, Laemolyta, Rhytiodus, and Schizodon), revealing that it is feasible and imperative to split Leporinus.

Cytogenetic studies on Leporinus have shown that most species lack sex chromosomes (Galetti et al., 1991, Galetti et al., 1981). Six species, however, have a distinctive ZZ/ZW sex chromosome system (Galetti et al., 1995, Galetti et al., 1981, Molina et al., 1998, Venere et al., 2004): Leporinus conirostris, L. macrocephalus, L. obtusidens [cited as L. elongatus (Paraná basin) and L. cf. elongatus (São Francisco basin)], L. piavussu (cited as L. obtusidens), L. reinhardti, and L. trifasciatus. In the ZZ/ZW system, the W chromosome is the largest one in the female karyotype and is absent from the male complement. Galetti et al. (1995) proposed the ZZ/ZW sex chromosome system as a synapomorphy for a putative monophyletic species group. However, the cytogenetics of most anostomid species remains unstudied, suggesting that potential ZW Leporinus might still be described. A recent phylogenetic analysis based on molecular data also grouped species bearing the ZW chromosome system as monophyletic (Clade C, Ramirez et al., 2016).

The species group treated herein includes the largest body-sized species of the family (up to 600 mm standard length (SL)), which are exploited in commercial and subsistence fisheries throughout tropical South America (Garavello and Britski, 2003). Its members are mostly herbivorous (Alvim and Peret, 2004) and potentially good seed dispersers (Galetti et al., 2008). This fish is characterized by a dental formula of 3/3. This number of teeth is an important feature for Leporinus species diagnoses (Britski and Birindelli, 2008). As a monophyletic group of ecologically equivalent species (Galetti et al., 1995) occupying all major hydrographic basins in South America, the systematics of this group may serve as an excellent model for studies of biogeography and the geology underlying watershed history (Hubert et al., 2007).

In the present study, two mitochondrial and three nuclear genes were used to reconstruct the evolutionary relationships of Anostomidae species bearing ZW sex chromosomes, as well as other potentially closely related species. We tested whether molecular and anatomical synapomorphies support the monophyly of the ZW clade, describing a new genus. We also inferred intrageneric relationships and the evolutionary processes involved in the diversification of this fish.