Coptobrycon bilineatus ( Ellis , 1911 ) ( Characiformes : Characidae ) : redescription and comments on its phylogenetic relationships

Coptobrycon bilineatus (Ellis, 1911) is redescribed on the basis of specimens from the District of Paranapiacaba, Municipality of Santo André, upper rio Tietê, and additional ones recently collected in a small coastal river system of Serra do Mar, very near the headwaters of the rio Tietê. The genus was compared to other Characidae lacking a supraorbital, and it seems to be more phylogenetically related to Grundulus based on the possession of various putative apomorphic character states related to: the absence of a rhinosphenoid and fourth, fifth (sometimes) and sixth infraorbitals; nasal pores separated; nares with up to six nasal lamellae; cephalic laterosensory system poorly developed on supraorbital and infraorbital series; and a globose scapula. Furthermore, Coptobrycon and Grundulus are characterized by the absence of the adipose fin, of the supraorbital laterosensory series on the parietal, and of the humeral spot, and by the reduction of lateral musculature in front of the first pleural rib and between the first and second pleural ribs. Biogeographic comments are also provided.


Introduction
Hasemania bilineata was described by Ellis (1911) based on the holotype and eight paratypes from Alto da Serra (Paranapiacaba and Mogi das Cruzes).Later, Géry (1966) erected the genus Coptobrycon for that species, justifying his decision in view of the considerable differences shown by this species when compared to other Hasemania species: "dorsal fin in advance of the middle of the body, a very short lateral line (only 3 perforated scales), a very short anal fin, and chiefly very aberrant mandibular teeth (Parodon-like multicuspid incisors) which number only 3 on each side".Géry (1966: 224) suggested that, contrary to Hasemania, Coptobrycon could be related to the "Hemibryconini," since it displayed some characters commonly found in the latter group, such as four teeth in the inner row of the premaxilla, third infraorbital well developed and, generally, irregularly implanted teeth in the outer row of the premaxilla.More recently, Mirande (2009Mirande ( , 2010) ) suggested a close phylogenetic relationship between Coptobrycon and Grundulus.
Until recently, only a few specimens of Coptobrycon from the upper rio Tietê were known: the type series of Hasemania bilineata, consisting of nine specimens, deposited at FMNH and CAS, plus four specimens in the MZUSP and six at the Museu de História Natural "Prof.Adão José Cardoso" of the Universidade Estadual de Campinas (ZUEC).The recent collection of a large series of Coptobrycon from the rio Itatinga (Serra et al., 2007), a tributary of the rio Itapanhaú, an independent small coastal river system in São Paulo State, prompted a redescription of this monotypic and enigmatic genus.

Material and Methods
Counts and measurements followed Fink & Weitzman (1974), with the following modifications or additions: longitudinal scale rows below lateral line counted to the pelvicfin insertion; scales along the anal-fin base, covering the basal portion of the rays; head depth, measured between the supraoccipital spine and the isthmus; predorsal distance, from the snout tip to the dorsal-fin origin; preventral distance, from the snout tip to the pelvic-fin origin; distance between the pelvic and anal fins, from the pelvic-fin posterior insertion to the anterior portion of the base of the anal fin.
All measurements were taken using an electronic caliper, point-to-point, from the left side of the specimens whenever possible, to the nearest 0.1 mm.In the description, the numbers in parentheses indicate absolute frequency.Specimens were cleared and stained (c&s) according to Taylor & van Dyke (1985) and dissected according to Weitzman (1974).Vertebrae (including those four in the Weberian apparatus), gill rakers, branchiostegal rays and small dentary teeth were counted in cleared and stained specimens.Institutional abbreviations followed Reis et al. (2003) with the addition of: Departamento de Zoologia e Botânica, UNESP at São José do Rio Preto (DZSJRP), and the Universidade Estadual de Maringá, Maringá (NUP).

Diagnosis.
Coptobrycon is easily distinguished from all other Characidae due to: a very peculiar dentary dentition, with only three, large, anterior, multicuspid and pedunculated, principal teeth in each half of the dentary (Fig. 5; vs. four or more conical or cuspidate and not pedunculated teeth, except in the Cheirodontinae and Iguanodectinae, which have more than four pedunculated teeth); the absence of the nasal bone (vs.present); the presence of a straight, dark brown stripe slightly above the base of the anal fin (Fig. 1), from vertical through its origin or slightly behind, continuing through the ventral portion of the caudal peduncle up to the base of the caudal fin (vs.absent).Furthermore, it can also be separated from all other Characidae, except Grundulus, by having the nasal pores separated by a non confluent space (Fig. 2; vs. nasal pores very close and confluent); infraorbitals 4 and 6 (5, sometimes) absent (Fig. 4; vs. present); infraorbital laterosensory canal greatly reduced, with pores inconspicuous or absent (Fig. 4; vs. present); supraorbital laterosensory canal absent in posterior portion of the frontal (vs.present).Other useful diagnostic characters include: up to six olfactory rosette lamellae, turned to the posterior portion of the nasal canal (Fig. 3; vs. more than ten lamellae, generally more than fifteen, concentrically arranged; except in Grundulus and Xenurobrycon); adipose fin absent (Fig. 1; vs. present; except in Grundulus, Gymnocharacinus bergi, Hasemania, some Hyphessobrycon species, and Spintherobolus).
Description.Morphometric data in Table 1.Small size, largest specimen examined 37 mm SL.Body moderately high, greatest body depth slightly ahead vertical through dorsal-fin origin.Dorsal profile of head straight or slightly convex from tip of snout to posterior portion of supraoccipital spine.Dorsal profile of body convex, from supraoccipital spine to dorsalfin origin; straight, or slightly convex, and descending along dorsal-fin base; straight and gradually descending from dorsal fin to caudal peduncle end.Head rounded in transverse section; predorsal portion slightly keeled mainly on posterior half; postdorsal portion slightly flattened anteriorly and slightly keeled posteriorly.Ventral profile of body convex from snout to pelvic-fin origin; straight and horizontal from pelvic fin to anal-fin origin (considerably more convex in specimens with replete digestive tube); straight, or slightly concave, and ascending along anal-fin base; straight and slightly ascending, nearly horizontal, from anal fin to caudal-fin base.
Ventral portion of head and body up to anal-fin origin rounded in transverse section; base of anal fin keeled; caudal-peduncle ventral portion rounded, slightly keeled near anal-fin base.
Head triangular from lateral view; snout rounded.Mouth terminal.Premaxilla short; maxilla short, extending to behind vertical passing through anterior margin of orbit.Anterior and posterior nasal pores completely separated (Fig. 2); anterior nasal pore slightly elevated, on apex of short cone, and posterior pore not elevated, at same level of snout surface; olfactory rosette with up to six lamellae, posteriorly directed and turned to posterior nasal pore (Fig. 3).Adipose eyelid underdeveloped, mostly inconspicuous, except in anteroventral portion of eye.
Frontoparietal fontanel from posterior portion of mesethmoid to base of supraoccipital spine, visible and conspicuous in c&s specimens, but not apparent and covered by thick skin in alcohol-preserved specimens.Rhinosphenoid absent, only cartilaginous tissue present between lateral ethmoid and vomer.Orbitosphenoid with posterior invagination limiting ventral projection, posterior and elongated, turned to pterosphenoid.Fourth and sixth infraorbitals absent; fifth infraorbital also absent in some specimens (Fig. 4); when present, separated from third infraorbital by space equivalent to half its depth (Fig. 4).Nasal and supraorbital absent; antorbital vertically elongated with anteroventral process crossing anterior process of maxilla and turning to premaxilla dorsoposterior portion.
Canals and pores of cephalic lateral line present and well developed along latero-anterior portion of frontal (extending only up to vertical passing through epiphyseal bar), behind anterior nasal pore, on dentary and preopercle; infraorbital bones with canals underdeveloped or absent and inconspicuous pores.Free neuromasts (sensory papillae) present along the supraorbital, infraorbital, maxilla, dentary, gular, and supraoccipital cephalic laterosensory series.
Four branchiostegal rays.Gill rakers elongated on first branchial arch; two on hypobranchial, eight to ten on ceratobranchial and five to six on epibranchial.
Outer row of premaxilla with one tri-to tetracuspid tooth (rarely with more than four cusps), between first and second teeth of inner row and slightly turned anteriorly; inner row with four (33) or five (2) pedunculated, penta-or octocuspid  teeth, with wide compressed crown, slightly curved posteriorly, with median cusp subequal, slightly more developed than others (Fig. 5).Maxilla without teeth.Dentary short, with single row of two (2), three (31) or four (2) anterior, large, hepta-to decacuspid, pedunculated teeth, with large, compressed, straight and turned posteriorly crown, with rounded subequal cusps, followed by one to four much smaller conical teeth (Fig. 5).Palatine approximately rectangular, without openings, perforations or thinning in its mid-anterior portion.
Hypaxialis muscle portion largely reduced, especially obliquus superioris muscle, leaving thin muscular layer, composed principally of obliquus inferioris, in front of first pleural rib, exposed anterior and proximally (Fig. 6); muscle reduction also between first and second pleural ribs.No direct contact of swim bladder with skin.
Coracoid little developed ventrally.Opening between coracoid and cleithrum nearly entirely visible laterally; scapula well developed, globose, with small scapular foramen, more central and bordered anteriorly by wide bony portion (Fig. 7).Pelvic fin with i,4(1), i,5(28) or i,6(3) rays; with posterior rounded edge, not reaching anal-fin origin; its posterior extremity beyond vertical passing through last third of distance between its origin and anal-fin origin; anal fin with iii,10(1), iii,11(19), or iii,12(13) rays; with straight or slightly concave edge; second unbranched ray and two first branched rays, longer and subequal, subsequent rays gradually shorter.Caudal fin with rounded lobes and i,17,i(33) principal rays.Mature males and females (dissected from DZSJRP 6610 and 6621) without hooks on pelvic and anal-fin rays.boundaries of this protected area.The rio Itatinga (Fig. 9) is a typical Atlantic Forest stream, possessing swift-flowing, transparent, cold waters, and alternating relatively deep pools and shallow riffles.Usually, the bottom of the more lotic portions is composed of sand, stones and boulders, whereas in the more lentic stretches there is an accumulation of leaves and branches from the riparian vegetation.The riparian vegetation is an old, abandoned eucalyptus plantation, with dense undergrowth of native shrubs and trees.
Coptobrycon bilineatus was common and abundant in most of the sampled stretches of the rio Itatinga.Specimens were easily observed forming large schools, often associated with schools of Astyanax paranae.The species was generally recorded in areas with a relatively slow water flow, among macrophytes, branches and submerged logs.The examination of some stomach contents revealed mainly fragmented terrestrial vegetation, although some digested items of animal origin were recorded in smaller quantities.Immature, maturing, mature, and empty gonads were recorded, which seems to indicate that the species reproduces in more than one season of the year.Other species collected at the rio Itatinga are Astyanax paranae, Glandulocauda melanogenys [= G. melanopleura Ellis, 1911  Until recently, Coptobrycon bilineatus was considered to be endemic to the upper course of the rio Tietê (Langeani, 1989), where it was collected for the last time in 1981 (ZUEC 4287).Extensive collecting in the upper rio Tietê basin, conducted from 1985 to date was not successful in finding any additional specimens of the species.Coptobrycon bilineatus is officially considered an endangered species in Brazil (Serra et al., 2008).The discovery of Coptobrycon bilineatus in the rio Itatinga is, from a conservation point of view, quite reassuring.However, its overall conservation status is still cause for concern and special attention to the species is required.

Color in life.
Body predominantly yellowish ventrally.In water, specimens appear golden, probably as a consequence of great quantities of guanine along portions of visceral cavity, infraorbital and opercular regions; lateromedial and lateroventral black conspicuous stripes.Fins yellowish orange, especially dorsal, anal, and caudal fins.

Color in alcohol.
Black chromatophores distributed all over body.Head with large concentration of small chromatophores along lateral portion; larger chromatophores on dorsal portion of operculum and below eye, on third infraorbital; regions of larger chromatophores bordered by clear areas, without pigment.Concentrated chromatophores near edges of scales, forming reticulate pattern.Humeral spot absent; mid-lateral stripe from vertical passing dorsal-fin origin, or posterior portion of head (more melanic specimens), up to caudal-fin base; shorter stripe, slightly above anal-fin base, from analfin origin, or little behind, through ventral portion of caudal peduncle up to caudal-fin base; caudal-fin base dark.Dorsal portion of body dark from head to caudal fin; head chromatophores small and numerous; from head to dorsal-fin origin as in rest of body; posterior to dorsal fin much more numerous, larger, conspicuous, and visible laterally.Ventral portions of head and body lighter, with small and numerous chromatophores along dentary, anterior portion of isthmus, and ventral portion of preopercle and interopercle; chromatophores slightly larger and more scattered from posterior portion of head to pelvic-fin origin, and numerous, dense and conspicuous from anal pore to caudal fin.Fins slightly dark, mainly along anterior and posterior edges of rays.
Distribution.Upper rio Tietê (upper rio Paraná drainage) and rio Itatinga, a tributary of the rio Itapanhaú, a small independent coastal river system, São Paulo State, Brazil (Fig. 8).

Ecology and conservation.
The RPPN Parque das Neblinas is a privately owned conservation unit within the Atlantic Forest, and most of the rio Itatinga basin is within the

Discussion
Although there has been recent progress in clarifying phylogenetic relationships within the Characidae (Calcagnotto et al., 2005;Mirande, 2009Mirande, , 2010;;Javonillo et al., 2010), there are still many limitations to a deeper and complete understanding of characid phylogeny, principally due to the enormous diversity of the group (Weitzman & Malabarba, 1998;Malabarba & Vari, 2000;Lima et al., 2003), with some species and genera poorly represented in collections and still some with great morphological modifications.
Nasal pores separated by a non confluent space (Fig. 2) (Mirande, 2010; character 351: 1).The presence of anterior and posterior nasal pores very close and confluent is a plesiomorphic condition in the Characidae.Coptobrycon and Grundulus, nevertheless, have anterior and posterior nasal pores individualized, separated by a space, and non confluent.Anostomids (pers.obs.) and crenuchids are the only other Characiformes having nasal pores separated and non confluent.Buckup (1993a: 233) commented that the separation of nasal pores in the Crenuchidae, as in a few other Characiformes (not listed), would be usually associated with miniaturization.
Few olfactory rosette lamellae turned to the posterior portion (Fig. 3).The olfactory rosette in the Characidae, and in most Characiformes, usually have more than ten lamellae (generally more than fifteen), concentrically arranged (pers.obs.).Coptobrycon and Grundulus, nevertheless, have up to six lamellae, turned to the posterior portion of the nasal canal.Similar conditions also occur in Xenurobrycon and Pyrrhulina, here interpreted as independent acquisitions, since Xenurobrycon is a highly derived member of the subfamily Glandulocaudinae (Weitzman & Fink, 1985), currently considered a clade within the Stervardiinae (Mirande, 2009(Mirande, , 2010)), while the lebiasinid Pyrrhulina is even more distantly related, belonging to a monophyletic clade that includes the Lebiasinidae, Erythrinidae, Hepsetidae and Ctenoluciidae (Vari, 1995), absent in Coptobrycon and Grundulus.Rhinosphenoid absent.Coptobrycon and Grundulus show no rhinosphenoid bone and the small space between the orbitospheroid and the midposterior portion of the lateral ethmoid and vomer is filled with a cartilaginous tissue.Among the Characidae sensu stricto most of the included taxa have a rhinosphenoid; the loss of this bone is secondary apomorphic.It was also observed by us in Carlastyanax aurocaudatus and Hasemania crenuchoides, and according to Chernoff & Machado-Allison (1990), in all species of Ceratobranchia and Astyanax festae, an unidentified Creagrutus species, and in some species of Bryconamericus.Gymnocharacinus bergii also does not have a rhinosphenoid (Miquelarena & Arámburu, 1983).The absence of the rhinosphenoid was proposed by Mirande (2010, character 47: 0) as a synapomorphy for the clade 280, including Gymnocharacinus, Coptobrycon, and Grundulus.
In the Characidae, infraorbitals 1 to 6 show a conspicuous laterosensory canal.In Coptobrycon and Grundulus, this canal is greatly reduced, with pores inconspicuous or absent.
Infraorbitals 4, 5, and 6 absent (Fig. 4).The Characidae usually have six infraorbitals and the loss or reduction of any of them is frequently associated with small size or miniaturization (Weitzman & Fink, 1983, 1985).Infraorbitals 4 and 6 are always, and 5 eventually, absent in Coptobrycon and Grundulus.
Supraorbital laterosensory canal partially absent in frontal and totally absent in parietal.The supraorbital laterosensory canal frequently extends up to the parietal; its absence in that bone is apomorphic within Characiformes and related to a miniaturization phenomenon (Buckup, 1993b), which may, nevertheless, also occur in normal-sized non-miniature species (Zanata & Vari, 2005).In our analysis, the supraorbital laterosensory canal in Grundulus and Coptobrycon, apart from being absent in the parietal, also does not occur in the posterior portion of the frontal.
Scapula globose, with a wider anterior edge and reduced foramen (Fig. 7).The main body of the scapula in the Characidae has an hourglass shape, with dorsal and ventral, narrow and anterior, projections fused anteriorly, forming a large bone ring which borders the scapular foramen (Weitzman, 1962: 41, fig. 20).In Coptobrycon and Grundulus, and also in Markiana, Metynnis and Triportheus, the scapula is more globose; the scapular foramen is reduced and the bone ring wider.This is probably apomorphic, albeit not homologous for all those taxa, since both Metynnis and Triportheus are not part of the Characidae sensu strictu because they have a supraorbital and share with other Serrasalminae (Machado-Allison, 1983;Mirande, 2009), the first, and Lignobrycon (Malabarba, 1998) and Brycon (Mirande, 2009, who did not examine Lignobrycon), the second, other apomorphic states absent in Coptobrycon and Grundulus.Markiana is according to Mirande (2009) included in his Astyanax clade, with various Astyanax species, and some of Hyphessobrycon, Bryconamericus, and Psellogrammus.
Mirande (2010) also proposed as synapomorphies uniting Coptobrycon and Grundulus: laterosensory canal of dentary reduced or absent (79: 1) and lateral line interrupted (91: 1), character-states also occurring independently in other characids.Finally, the four teeth in the inner row of the premaxilla and ii,7-8 rays in the dorsal fin, features shown by Coptobrycon, should be interpreted as an independent offshoot from similar conditions in the members of the socalled Characidae Clade A of Malabarba & Weitzman (2003) (= Stervardiinae sensu Mirande, 2009Mirande, , 2010)).In contrast to the taxa belonging to that clade, Coptobrycon has a postcleithrum 3 with a posterior convex projection (vs.simple), palatine without openings, perforation or thinning in its midanterior portion (vs.normally present), orbitosphenoid with a ventral and posterior invagination (vs.absent) and humeral spot absent (vs.normally present).
Biogeography.Coptobrycon and Grundulus, occurring respectively in the eastern Brazilian Shield and Colombian Andes, would fit better in the "Pattern A" of Ribeiro (2006), a consequence of ancient events, which resulted in sistertaxa geographically restricted and very distant from each other.The restricted occurrences of Coptobrycon and Grundulus only in the the Serra do Mar and,the highlands of the Andes, leaving a large area in the central portion of South America which lacks records of both genera, lead us to hypothesize a relictual pattern of occurrence for the clade that, probably, underwent a process of extinction in the lowland drainages (Román-Valencia et al., 2003, for Grundulus) situated in between, similar to the proposition of Bührnheim et al. (2008) for the distribution of the monophyletic clade composed of Spintherobolus and Amazonspinther (Cheirodontinae, Characidae).Furthermore, extinction seems to be currently threatening the clade; Coptobrycon bilineatus and Grundulus species are considered endangered (Román-Valencia et al., 2003, 2005;Serra et al., 2008, respectively), similar to the situation for Spintherobolus, as already addressed by Bührnheim et al. (2008).
In spite of their restricted geographic distribution, recent surveys in the northern Andes and Serra do Mar, respectively, have uncovered new species of Grundulus (Román-Valencia et al., 2003, 2005) and enlarged the area of distribution of Coptobrycon bilineatus (Serra et al., 2007), suggesting the possibility of discovering other new taxa, which may provide more information on the clade.Serra et al. (2007) suggested that the occurrence of Coptobrycon bilineatus together with Glandulocauda melanogenys [= G. melanopleura (Ellis, 1911) sensu Menezes & Weitzman, 2009], Pseudotocinclus tietensis, and Astyanax paranae, in a coastal drainage of the São Paulo State, was due to headwater capture of the upper Tietê by the rio Itatinga, similar to what happened in other drainages in that part of southeast Brazil (see Ribeiro et al., 2006, for a description of the tectonic processes and additional examples).Moreover, this discovery reinforces the statement made by Oliveira & Britski (2000) that a greater number of species endemic to the upper Tietê, as proposed by Langeani (1989), possibly occur in other drainage areas of southeast Brazil.

Fig. 6 .
Fig. 6.Coptobrycon bilineatus, DZSJRP 6610, 30.7 mm SL: humeral region with tegument removed, showing reduced muscular tissue in front of first pleural rib and between first and second pleural ribs.