A new species of the catfish Neoplecostomus ( Loricariidae : Neoplecostominae ) from a coastal drainage in southeastern Brazil

A new species of loricariid catfish is described from the rio Perequê-Açú and surrounding basins, Parati, Rio de Janeiro State. The new species has the accessory process of ceratobranchial 1 more slender than the main body of the ceratobranchial, and a very large sesamoid ossification, markedly greater in size than the interhyal. Additionally, the new species presents a distinct dorsal color pattern consisting of a conspicuous horseshoe shaped light blotch with a central dark area posterior to the supraoccipital.

Neoplecostomus currently comprises 15 species, with most occurring in streams draining the Brazilian Crystalline Shield.The present work describes a new species endemic to four small coastal drainages in the State of Rio de Janeiro.

Material and Methods
Measurements were taken point to point with digital calipers to the nearest 0.1 mm, on the left side, following Zawadzki et al. (2008).Specimens smaller than 50.0 mm of SL, were not included in the morphometric and meristics analysis and are assigned as not measured.Plate counts followed Langeani (1990) and Zawadzki et al. (2008), with additional counts of the lateral plate series, following Schaefer (1997).Plates just below the pteroticsupracleithrum, surrounded by a naked area, were not included in the counts of the lateral series.Plates were counted on the left side in both cleared and stained (c&s) and alcohol-preserved specimens.The former were prepared according to Taylor and Van Dyke (1985).Measurements are presented as percentages of standard length (SL), head length (HL) or other measurements (e.g.snout length/orbital diameter, interorbital lenght/orbital diameter, interorbital lenght/mandibullary width, predorsal length/first dorsalfin ray length, caudal peduncle length/caudal peduncle depth, pelvic-fin length/caudal peduncle depth, lower caudal-fin spine/caudal peduncle depth).The presentation of measurements includes minimum, maximum, mean and standard deviation.Counts are presented as ranges with modes in parentheses.Vertebral counts included five from the Weberian apparatus and one from the hypural plate.Osteological analysis followed Pereira (2008).
Fragments of the mitochondrial cytochrome oxidase c subunit I (COI) gene were sequenced from N. microps from rio Paraíba do Sul basin at Silveiras, São Paulo State (four specimens), Cunha, São Paulo State (one), the rio Guapi-Açu basin, Rio de Janeiro State (seven), and from rio Macaé basin, Rio de Janeiro State (five).The same region was sequenced from three specimens of the new species from rio Pereque-Açú, Parati in Rio de Janeiro State.Total DNA was extracted from muscle tissue using the salting out method adapted from Miller et al. (1988).The extracted DNA was precipitated with sodium acetate and ethanol, re-suspended in 50 μL of ultrapure water, and stored at -20°C.
The polymerase chain reaction (PCR) profile was as follows: one initial cycle of 4 min at 94° C, followed by 35 cycles of 1 min at 94° C, 1 min at 46° C, 1 min at 72° C, and a final extension at 72° C for 5 min.The PCR products were purified and both strands were sent for sequencing by MACROGEN Korea.
The vouchers of the new species used in molecular analyses are deposited at DZSJRP under catalog numbers 13914-1, 13914-2 and 20433-1.Mitochondrial DNA sequences from this work were deposited in GenBank (accession codes KU550707 for N. paraty from rio Perequê-Açú basin, KU608294 and KU608295 for N. microps from rio Paraíba do Sul basin, KU608296 for rio Guapi-Açu basin, and KU608297 to KU608298 for those from the rio Macaé basin).

Fig. 1
Neoplecostomus sp.Pereira et al., 2003: 8 (rio      Head wide and moderately depressed.Head and snout weakly rounded in dorsal view.Interorbital space slightly convex in frontal view.One median ridge from snout tip to area between nares, another one from posterior naris to anterior margin of orbit.Snout convex in lateral profile.Eye moderately small (7.8-10.7% of HL), dorsolaterally placed.Iris operculum present.Lips well developed and rounded, covered by papillae.Lower lip not reaching pectoral girdle.Two or three irregular rows of papillae posterior to dentary teeth; papillae large, conspicuous and transversally flattened.Maxillary barbel short, coalesced with lower lip, and generally bifurcated in free portion (some specimens only with fold of skin instead).Teeth long, slender and bicuspid; mesial cusp longer than lateral.Dentary rami forming an angle of approximately 120°.Dorsal-fin ii,7; origin posterior to vertical passing through pelvic-fin origin.Nuchal plate not covered by skin.Dorsalfin spinelet generally present, half-moon shaped and wider than dorsal-fin first ray base, absent in some specimens; dorsal-fin locking mechanism absent.Dorsal-fin posterior margin slightly falcate, reaching or surpassing vertical through end of pelvic-fin rays when adpressed.Adipose fin present and well developed, preceded by none, one or two azygous plates.Pectoral-fin i,6; with depressed and inward curved unbranched ray, shorter than longest branched ray.Pectoral-fin posterior margin slightly falcate, reaching or nearly reaching half pelvic-fin length when adpressed.Pelvic-fin i,5; posterior margin nearly straight, reaching or nearly reaching anal-fin insertion when adpressed.Pelvicfin unbranched ray ventrally flattened, with dermal flap on dorsal surface in males.Pectoral and pelvic-fin unbranched rays with odontodes on lateral and ventral portions.Analfin i,5; posterior margin nearly straight.Anal-fin unbranched ray only with ventral odontodes.Caudal-fin i,7,7,i; bifurcate; lower lobe longer than upper.Vertebrae 31-32 (32).
Coloration.Dorsal surface ground color yellowish with light or dark brown blotches.Head with straight yellowish line from snout tip to anterior nares.Another large, less conspicuous and more laterally placed line running from snout border to slightly posterior of nares transverse line.Three other light areas between and around eyes and posterior to opercle and preopercle.Body with four transverse dark brown stripes at anterior portion of dorsalfin base and a little posterior; at posterior portion of dorsalfin base; from vertical through posterior portion of anal-fin base to adipose-fin spine; and at caudal peduncle.
Posterior to supraoccipital, a conspicuous light and horseshoe shaped spot with dark center (Fig. 4a), slightly faded in some specimens (Fig. 4b).Between dorsal dark stripes, three other lighter areas with dark blotches.Juveniles lacking dark blotches or with blotches only slightly demarcated (Fig. 4a).Ventral surface of head and body yellowish medially; light brown laterally from snout tip to region just anterior of anus; light brown posterior of anus to the caudal peduncle.All fins with irregular dark brown areas; sometimes forming inconspicuous transverse stripes.Adipose fin with dark brown spine and hyaline membrane.Etymology.The specific epithet refers to "Paraty", the original spelling of the municipality of Parati, Rio de Janeiro.Paraty (or Paratii) derived from the Tupi "parati" (the mullet Mugil curema Valenciennes, 1836) and "i" (river).A noun in apposition.
Conservation status.Neoplecostomus paraty occurs in four small and independent coastal drainages which run through unprotected land and also some conservation areas, such as Serra da Bocaina National Park and Serra do Mar State Park.The extent to which these conservation areas protect this species is unknown.Outside these parks (and to some extent also within these parks) streams and rivers are threatened by a range of anthropogenic threats associated with urbanization and the growth of the tourist industry, including Parati-Cunha road construction (Avena, 2003).Although N. paraty does not meet the criteria for any category of threat (IUCN, 2016), its highly endemic status demands special attention regarding conservation actions, and highlights the importance of the maintenance and even expansion of the protected areas in the region.
Molecular analyses.Partial sequences of the COI gene were taken from 21 specimens, resulting in a matrix with 663 base pairs (bp) from which 559 sites were invariant, 97 were variable, and 35 were parsimony informative.The nucleotide frequencies were, on average, 23.3% adenine, 25.7% thymine, 30.7% cytosine and 20.3% guanine.A single haplotype was found in both N. paraty and the population of N. microps from rio Guapi-Açu basin, whereas two haplotypes were found in both populations of N. microps from rio Paraíba do Sul and rio Macaé, totaling 6 haplotypes throughout the specimens sequenced.Molecular distances among all assessed populations are given in Table 2.
Considering the specimens from each basin as separate species or evolutionary units, the intraspecific distance values were zero for N. paraty and N. microps from Guapi-Açu and 0.15% for both N. microps from rio Paraíba do Sul and rio Macaé, due to a single nucleotide different on one specimen each.
A maximum-likelihood analysis showed that haplotypes from each basin are separate lineages well-suported by high bootstrap values (Fig. 7).Lower bootstrap values occur for the clades including more than one lineage within N. microps, suggesting that it may be a species complex.The analysis indicates, however, that N. paraty is the sister taxon to a clade containing all populations of N. microps.

Discussion
The new species possesses rare osteological features for the genus Neoplecostomus.The slender condition of the accessory process of ceratobranchial 1 is otherwise observed only in N. corumba, N. jaguari and N. ribeirensis, and the very large sesamoid ossification only in N. ribeirensis and N. yapo.Despite being internal, both characteristics are easily observable and distinguishable, and the latter shows a small intraspecific variability.Furthermore, the dorsal color pattern of N. paraty is very conservative and conspicuous, and not seen in any other species within the genus.Other Neoplecostomus species have the general pattern proposed by Langeani (1990), or show a diffuse pattern of smaller blotches, both of which are completely different patterns from that displayed by N. paraty.Thus, the color pattern is a reliable characteristic which easily distinguishes N. paraty from all other species of Neoplecostomus.
Neoplecostomus paraty (as Neoplecostomus P sp.n.) and N. microps were proposed as sister species by Pereira (2008).Although some uncertainty remains regarding the relationships between these species (and the overall phylogenetic relationships within Neoplecostomus fall beyond the scope of the present paper), the molecular analyses performed herein strongly support that N. paraty is a unique evolutionary lineage (Fig. 7).The genetic distance (>2%) between N. paraty and the remaining populations of N. microps (Table 2) provides evidence for a long isolation time and supports N. paraty as a new species.Additional phylogenetic analyses are necessary to refine our understanding about the relationships and historical biogeography of N. paraty and the N. microps species complex.

Fig. 5 .
Fig. 5. Southeastern Brazil showing geographic distribution of Neoplecostomus paraty (red circle) and Neoplecostomus microps (white circle).Type locality indicated by yellow circle.Symbols can represent more than one locality.Ecological notes.Neoplecostomus paraty was found in streams draining the Serra da Bocaina mountain range in the Parati Municipality, with clear and cold water, direct sunlight, fast flow, large rocks at the bottom, depths between 10 to 50 cm and moderate riparian vegetation (Fig. 6).Some sites in the Parati Municipality are probably among the lowest altitudes (e.g.38 m) at which Neoplecostomus species have been collected, since the genus commonly occurs at altitudes higher than 700 m.Other loricariids collected together with N. paraty are: Kronichthys heylandi (Boulenger 1900), Pareiorhina sp., Hemipsilichthys nimius Pereira, Reis, Souza & Lazzarotto 2003 and Schizolecis guntheri (Miranda Ribeiro 1918).
Description.Measurements and counts given in Table1.Body elongated and depressed.Greatest body width at cleithrum, narrowing to caudal peduncle.Dorsal body profile gently convex, elevating from snout tip to dorsalfin origin and descending to first caudal-fin ray.Greatest body depth at dorsal-fin origin.Trunk and caudal peduncle dorsally rounded in cross section.Body ventrally flattened to anal-fin origin, flattened to slightly ascending towards caudal fin.Dorsal surface of body completely covered by dermal plates, except for naked area around dorsal-fin base.Snout tip with small naked area.Ventral head surface naked except by canal plate ahead of gill openings.Abdomen with conspicuous and small dermal platelets between insertions of pectoral and pelvic fins, forming thoracic shield (heptagonal-or hexagonal-shaped), surrounded by naked areas.One or two plates, arranged linearly and surrounded by naked areas (rarely with one to three small plates in front of them), between humeral process of cleithrum and first plate of lateral series (only exception in left side of holotype -four plates in line and two small ones in front of them).

Table 1 .
Morphometric and meristic data of Neoplecostomus paraty.H= holotype, n= number of specimens and SD = standard deviation.