A new species of Bryconamericus (Characidae: Stevardiinae: Diapomini) from the upper rio Paraná basin, Brazil

Bryconamericus is the most diverse genus within Stevardiinae, comprising 61 valid species distributed in Cisand TransAndean basins from Panama in Central America to northern Argentina in South America. Three species are known from the upper rio Paraná basin: B. exodon, B. iheringii, and B. turiuba. Herein we describe a new species of Bryconamericus from the upper rio Paraná basin inhabiting tributaries of Ivaí, Piquiri, and Tibagi basins, Paraná State, Brazil. The new species differs from its congeners by the presence of unaligned teeth in the outer tooth row of the premaxilla; a single, vertical, dorsally expanded and rounded humeral spot; 36-39 pored scales in the longitudinal series; body depth 31.6-37.9% SL; anal-fin base length 24.8-30.1% SL; number of branched anal-fin rays 19-22, and bony hooks on pelvicand anal-fin rays of sexually dimorphic males. The new species is syntopic with other Stervadiinae in the upper rio Paraná basin such as B. iheringii, B. turiuba, Piabarchus stramineus, and Piabina argentea.

The systematics of Bryconamericus has been investigated for decades, as well as its doubtful monophyly within Characidae (Eigenmann, 1927;Fink, 1976;Vari, Géry, 1980;Vari, Siebert, 1990). More recently, morphology and molecular based studies have shown that the genus is certainly polyphyletic (Malabarba, Malabarba, 1994;Malabarba, Kindel, 1995;Malabarba, Weitzman, 2003;Javonillo et al., 2010;Mirande, 2010;Oliveira et al., 2011). Although Bryconamericus sensu stricto lacks a morphological diagnosis from other Stevardiinae, Bryconamericus is still diagnosed by a combination of morphological characters proposed by Eigenmann (1927) and updated by Vari, Siebert (1990): premaxilla bearing two series of teeth; inner series bearing four teeth larger than outer series teeth; a single series of teeth in the dentary; few maxillary teeth; caudal fin scaleless; third infraorbital well developed, contacting both ventral and posterior arms of the preopercle; gill rakers arrow-shaped; lateral line completely pored, and the absence of glandular tissue and/or pouch scales in the caudal fin of males.

Material and Methods
Counts and measurements were made preferably on the left side of the specimens following Vari, Siebert (1990) and , except for distance from dorsal-fin origin to anal-fin origin and distance from dorsalfin origin to pectoral-fin origin, which followed Harold, Vari (1994). Counts of vertebrae, ribs, dentary teeth, tooth cusp number, supraneurals, and procurrent caudal-fin rays were made on cleared and stained specimens (c&s) prepared with an adapted protocol from Taylor, van Dyke (1985). Vertebral count included the four vertebrae forming the Weberian apparatus, and the terminal centrum as single element (Weitzman, Malabarba, 1999). Counts were made under microscope and measurements were taken with a caliper with 0.01 mm precision. In the description, counts are followed by the number of specimens presenting such value between parenthesis, and the value of the holotype followed by an asterisk. Body measurements are presented as percentages of Standard Length (SL) and subunits of the head as percentages of the Head Length (HL). Fish classification follows Eschmeyer et al. (2017). Acronyms of institutions follow Sabaj Pérez (2014).
Geometric morphometrics analysis (GM). GM was herein used to address the morphological variation among the new species of Bryconamericus and five sympatric species of Stervadiinae: B. exodon, B. iheringii, B. turiuba, Piabarchus stramineus, and Piabina argentea. Specimens of B. iheringii and P. argentea from their type-locality basins (Laguna dos Patos and rio São Francisco basins, respectively) were also included in the analysis to encompass intraspecific morphological variation. Lots used in geometric morphometrics analysis are listed as (GM) in the list of type-material and material examined. Individual photographs were taken with 10 Mpixels resolution digital camera. Photograph files were grouped and transformed in *.tps files using TPSUtil V. 1.86 program (Rohlf, 2015). Landmarks were traced with the assistance of TPSDig V.2.2 program (Rohlf, 2015). The landmarks were aligned with procrustes superimposition 2D option of the program PAST (Hammer et al., 2001), and the same program was used for the multivariate canonical variate analysis (MANOVA/CVA option) and principal components analysis (Principal Components option) (Hammer et al., 2001). The procrustes superimposition eliminates variations in position, scale, and orientation of landmarks (Klingenberg, 2002), superimposing all individuals, adjusting and centering each configuration between homologous landmarks, and generating a reference configuration. The following landmarks were traced on each specimen: 1) contacting point of anterior tip of dentary with premaxilla; 2) tip of snout; 3) anterior border of snout; 4) posterior tip of maxillary bone; 5) anterior margin of eye; 6) posterior margin of eye; 7) posterior tip of supraoccipital process; 8) contacting point between posterior margin of third and fourth infraorbitals; 9) posterior margin of opercle; 10) pectoral-fin base; 11) dorsal-fin origin; 12) posterior insertion of dorsal fin; 13) adipose-fin origin; 14) posterior insertion of adipose fin; 15) tip of penultimate dorsal procurrent ray; 16) tip of penultimate ventral procurrent ray; 17) posterior insertion of anal fin; 18) anal-fin origin; and 19) pelvic-fin origin. Description. Morphometric data presented in Tab. 1. Small sized, largest specimen presenting 71.3 mm SL. Body laterally compressed. Body deepest point at dorsal-fin origin. Dorsal profile of head convex from tip of snout to vertical through posterior limit of nostril, slightly concave to straight from that point to tip of supraoccipital spine. Dorsal profile of body convex along predorsal region, straight to slightly convex along dorsal-fin base and from terminus of dorsal-fin base to adipose fin, and straight to slightly concave along caudal peduncle. Ventral profile of head strongly convex from tip of ventral lip to pelvic-fin insertion, slightly convex from that point to anal-fin origin, straight along anal-fin base, and straight to slightly concave along caudal peduncle (Fig. 1). Mouth opening subterminal, lower jaw shorter than upper jaw. Premaxillary teeth arranged in two series: inner series with 4*(30) aligned penta-to heptacuspid teeth, decreasing gradually in size laterally; outer series with 4*(22) to 5(8) tri-to pentacuspid teeth. Outer series teeth unaligned. Maxillary teeth 2(10), 3*(14), 4(5), or 5(1), tricuspid, rarely uni-or pentacuspid. 4*(30) large dentary teeth pentacuspid, followed by 2(1), 4(2), 5(2), or 6(1) conical to tricuspid teeth decreasing in size posteriorly (Fig. 2). Branchiostegal rays 4(6); first gill arch with 1(1) or 2(5) rakers on hypobranchial, 6(3) or 7(3) rakers on ceratobranchial, 1(6) raker on intermediate cartilage, and 5(2) or 6(4) rakers on epibranchial.
Supraneurals 5(6). Precaudal vertebrae 17 (6), caudal vertebrae 18 (1), 19(3) or 21(2); total 35 (1), 36(3) or 38(2). Coloration in alcohol. Overall body coloration pale yellowish to dusk (Fig. 1), occasionally with intense deposition of guanine over infraorbitals, opercular, and gular areas of head, longitudinal dark lateral stripe and abdominal region. Head following same overall coloration of body, darker over dorsal surface, upper jaw components, orbital margin of infraorbitals and opercular apparatus, due to higher concentration of dark chromatophores. Midline dorsal scales from posterior tip of supraoccipital to origin of caudal-fin upper lobe darker than lateral of body due to higher concentration of dark chromatophores. Scales from all series above lateral line with proximal region darkened, due to high concentration of dark chromatophores contrasting to clear hyaline distal margin; resulting in diffuse reticulated pattern. Humeral spot single, vertically elongate, extending from two scale series dorsal to lateral line to one scale series ventral to lateral line. Humeral spot darker and wider dorsal to lateral line, two scales-wide and slightly rounded in that region. Dark lateral longitudinal stripe anteriorly diffuse, variably from region between humeral spot and vertical through dorsal-fin origin to end of caudal peduncle; approximately one-scale deep. Humeral spot and dark lateral stripe occasionally omitted by intense deposition of guanine. Fins mostly hyaline with few scattered dark chromatophores on interradial membranes and along margins of rays. Anal fin and middle caudal-fin rays more densely pigmented than remaining fins.

Coloration in life.
Overall body dark chromatophores distribution on head, body and fins as described above. Dorsal region of eye with dark chromatophores, occasionally with red tone (Fig. 3). Overall color of body olive brown to yellow on dorsal region of body; ventral region of body white to light yellow. Lateral of body with silver stripe extending from head to end of caudal peduncle. Body with higher concentration of guanine over infraorbitals, opercular apparatus and ventral region of body. Dorsal and posterior region of body with iridescent blue to greenish tint. Intense orange to red coloration occasionally over proximal region of dorsal and anal fins, also on mid-length of caudal-fin rays. Base of caudal fin lobes yellowish. Distal tip of dorsal and pelvic fins, anterior margin of anal fin, and dorsal, and ventral margins of caudal fin white in some adult specimens (Fig. 3).
Sexual dimorphism. Secondary sexual dimorphism observed on males with or larger than 40.5 mm SL. Sexually mature males with hooks on pelvic-and analfin rays. Hooks distributed over first unbranched to 11 th branched anal-fin rays; along the posterolateral margin of lepidotrichia in both unbranched and branched regions of rays; one pair of hooks per ray segment. Hooks usually placed over posterior margin of posterior branches, scarcely over posterior margin of anterior branches. All branched pelvic-fin rays bearing one pair of hook per segment, on medial margin of both lateral and medial ray branches. Hooks rarely present on unbranched pelvicfin ray. Sexually mature males bearing gill-gland on the first branchial arch, formed by fusion of five to 11 most anterior gill filaments.

Geographic distribution.
The new species is found in tributaries of the rio Tibagi, rio Piquiri and rio Ivaí basins, all from the upper rio Paraná basin (Fig. 4). Ecological notes. In the rio Taquara basin, Bryconamericus coeruleus was collected in small to medium sized streams, with clear water and bottom composed by rocks and sandy areas. Although found in both slow and rapid waters, the species was more frequently found in pools just after rapids, and also close to the vegetation along the river banks. Syntopic species from rio Taquara are listed in Galves et al. (2007).
Etymology. The specific name, from the Latin coeruleus, is an adjective meaning sky color, in reference to the species bluish iridescence.

Conservation status. Following to the IUCN criteria (IUCN, 2016) and up to date information about
Bryconamericus coeruleus, the species can be categorized as Least Concern (LC). The known extent of occurrence is restricted to less than 20,000 km 2 (Extent of Occurrence -EOO of approximately 16,592 km 2 ) in a region with continuing decline of habitat quality due to riparian forest degradation. Furthermore, the species is known to exist at no more than nine locations (rio Barra Grande, rio Formoso, rio Itacolomi, rio Keller, rio Maria Flora, rio Muquilão, rio Piquiri, rio Taquara and rio Ubazinho). However, additional studies on population dynamics might bring more detailed information about the conservation status of the species.
Sympatric species and multivariate morphometric analysis. Bryconamericus coeruleus is sympatric to four species of Stervadiinae in its area of occurrence: B. iheringii, B. turiuba, Piabina argentea, and Piabarchus stramineus. The geometric morphometrics analysis by mean of CVA evidenced four morphological distinct groups on first (39.1%) and second (30.3%) canonical variates axis (Fig. 5). Bryconamericus coeruleus formed a group with B. iheringii from Laguna dos Patos basin and from upper rio Paraná. A second grouping was formed by B. exodon and Piabachus stramineus. Piabina argentea from rio São Francisco basin were distinguished from all other species including the P. argentea from upper rio Paraná basin, which formed another group with B. turiuba, evidencing morphological similarities. These unnatural groupings evidenced convergence of body form among different taxa. In a separate analysis, when comparing Bryconamericus coeruleus with B. iheringii, the species with most similar body shape among the sympatric species of Stevardiinae, the PCA showed distinction of Bryconamericus coeruleus on first axis (Fig. 6).The first axis retained 33.4% of data variation, while the second axis retained 17.7%.    The consensus landmarks of geometric morphometrics evidenced the highest body depth to Bryconamericus coeruleus and B. iheringii, and the smallest body depth to Piabina argentea, that also presented the longest head and longest snout (Fig. 7). The origin of dorsal fin of Bryconamericus coeruleus is positioned similarly to B. iheringii from upper rio Paraná basin, but the posterior insertion is similar to B. iheringii from Laguna dos Patos basin, consequently the new species present a smaller dorsal-fin base length. The location of the pelvic-fin origin in Bryconamericus coeruleus is anterior when compared to specimens from both populations of B. iheringii. The same is observed in the origin of the anal fin, which is anterior to the vertical line at the end of dorsal-fin base compared to both populations of B. iheringii (at same line in B. iheringii from upper rio Paraná and posterior in B. iheringii from Laguna dos Patos). The body depth is mostly evidenced by the distance from the origin of dorsal fin to the pelvic-fin insertion, and the end of dorsal-fin base to the origin of anal fin. The anal-fin base length in Bryconamericus coeruleus is one of the longest among the analyzed species, except to Piabarchus stramineus.

Discussion
We consider the generic assignment of the new species as Bryconamericus as a necessary, and perhaps provisory, attempt facing the unknown phylogenetic position of B. coeruleus in Stevardiinae and the recognition of the unnatural condition of Bryconamericus lato sensu (Thomaz et al., 2015). We also consider this assignment the most conservative approach in behalf of taxonomic stability (Dagosta, Netto-Ferreira, 2015), since B. coeruleus possesses the morphological characters that, in combination, traditionally define the genus (Eigenmann, 1927;Géry, 1977;Vari, Siebert, 1990) and still differentiate this grouping from all allied genera in Stevardiinae. Attempts to include this new species in other genera would require deep changes in the definition and current taxonomy of the Stevardiinae.
The presence of two tooth rows in the premaxilla is the generalized condition observed among the Stevardiinae (Mirande, 2010), with derivate reduction to one tooth row (e.g., Monotocheirodon, Othonocheirodus, and Xenurobryconini (in part)) and/or derived teeth arrangements (e.g., Creagrutus and Piabina, see Vari, Harold, 2001) in some groups. The majority of Bryconamericus is conservative in presenting two tooth rows in the premaxilla (except one tooth row in B. lethostigmus), however there is certain variation in the arrangement of teeth in the outer row. Among the species of Bryconamericus the outer premaxillary teeth may be arranged in two distinct conditions: aligned in a shallow convex arch, as observed in B. iheringii (FCJ pers. obs.), B. ikaa and B. uporas (Casciotta et al., 2002 or unaligned, as present in B. coeruleus. In the latter condition, the misalignment is due to the anterior displacement of either the tooth vertical axis or the tooth base relative to the arch line. Langeani et al. (2005) described the outer teeth in B. turiuba as unaligned, "having first and last teeth projecting anteroventrally" without a conspicuous displacement of the unaligned tooth base from the arch line, a condition also present in the type-species of the genus B. exodon and herein observed in B. coeruleus (Fig. 2). Unaligned teeth is present in all analyzed specimens of B. coeruleus regardless of body size (range 16.4-71.3 mm SL), and is not likely influenced by ontogenetic factors or tooth replacement process.