a new dwarf species of proceratophrys miranda-ribeiro, 1920

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Zootaxa 3551: 25–42 (2012) www.mapress.com/zootaxa/

Articleurn:lsid:zoobank.org:pub:BC6CB247-BCCA-49FD-A014-E3A5FDB366DC

A new dwarf species of Proceratophrys Miranda-Ribeiro, 1920 (Anura, Cycloramphidae) from the highlands of Chapada Diamantina, Bahia, Brazil

MAURO TEIXEIRA JUNIOR1,2, RENATA CECÍLIA AMARO1, RENATO SOUSA RECODER1, FRANCISCO DAL VECHIO1 & MIGUEL TREFAUT RODRIGUES1

1Laboratório de Herpetologia, Departamento de Zoologia, Instituto de Biociências, Rua do Matão, no 101, CEP 05508-090, Universi-dade de São Paulo, São Paulo, Brazil2Corresponding author. E-mail: [email protected]

Abstract

A new species of Proceratophrys is described from the highlands of northeastern Brazil. Molecular and morphological data suggests that Proceratophrys redacta sp. nov. is sister to P. minuta, and related to P. schirchi and P. cristiceps. The new species is diagnosed by its small size, absence of rostral and palpebral appendages, sagittal ridges interrupted, absence of postocular swellings, snout vertical in profile and dorsal coloration lacking distinct ocelli. The new species represents another example of endemism for the genus Proceratophrys in Chapada Diamantina region, and of another appendageless small-sized species associated with highlands. The phylogenetic results indicate that current morphological groupings in Proceratophrys may not represent natural groups.

Key words: Proceratophrys redacta sp. nov., mountain endemism, frog diversity

Introduction

After his travel through the Brazilian coast, the German naturalist Alexander Phillip Maximilian, Prinz zu Wied-Neuwied (see Myers et al. 2011), described the horned frog, Ceratophrys boiei (Wied 1824), which underwent taxonomic rearrangement (Gravenhorst (1825; 1829), ending up to be the first described species in the currently recognized genus Proceratophrys. After that several species were described (Peters 1872; Günther 1873; Müller 1883; Miranda-Ribeiro 1920; 1926; 1937; Jim & Caramaschi 1980; Izecksohn & Peixoto 1981; Weygoldt & Peixoto 1985; Giaretta & Sazima 1993; Mercadal de Barrio & Barrio 1993; Eterovick & Sazima, 1998; Izecsohn et al. 1999; Giaretta et al. 2000; Kwet & Faivovich 2001; Cruz et al. 2005; Prado & Pombal 2008; Cruz & Napoli 2010; Frost 2011; Martins & Giaretta 2011; Napoli et al. 2011; Cruz et al. 2012) and its taxonomy rearranged (Lynch 1971; Prado & Pombal 2008; Amaro et al. 2009).

Currently the genus comprises 27 species of small to medium/large-sized frogs, occurring in forests, savannas, grasslands and shrubby lands of eastern, central and southern South America, in Brazil, Argentina and Paraguay (Prado & Pombal 2008; Frost 2011; Martins & Giaretta 2011; Napoli et al. 2011). However, its present richness is likely underestimated, as indicated by the constant discovery of new species (Prado & Pombal 2008; Ávila et al.2011; Martins & Giaretta 2011; Napoli et al. 2011; Cruz et al. 2012). Furthermore, preliminary molecular phylogenies are showing that species formerly attributed to Odontophrynus, are indeed members of Proceratophrys (Amaro et al. 2009).

Recently, a dwarf species of Proceratophrys (P. minuta Napoli, Cruz, Abreu & Del Grande 2011) was described from the highlands of Chapada Diamantina, at the northern portion of Espinhaço range in state of Bahia (Napoli et al. 2011). This small-sized highland species seems to be restricted to the cloud forest patches of high elevations (above 800 m a.s.l., being very common at about 1170 m a.s.l.). Based on similarities in external morphology this new species was supposed to be allied to P. schirchi, (Napoli et al. 2011).

Both P. schirchi and P. minuta are not currently assigned to any species group, as well as P. rondonae, however the remaining species have been informally assigned to three or four species groups, based on overall

Accepted by M. Vences: 10 Oct. 2012; published: 16 Nov. 2012 25

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morphological similarity. The P. boiei species complex, characterized by a single and long uni-cuspidate palpebral appendage and lack of a triangular rostral appendage, is thought to include P. boiei (Wied, 1824); P. paviotii Cruz, Prado and Izecksohn, 2005; and P. renalis (Miranda-Ribeiro, 1920). The P. appendiculata species complex, sometimes referred to as a subset of P. boiei group, also with a single and long uni-cuspidate palpebral appendage, additionally characterized by the presence of a triangular rostral appendage: P. appendiculata (Günther, 1873); P. laticeps Izecksohn and Peixoto, 1981; P. melanopogon (Miranda- Ribeiro, 1926), P. moehringi Weygoldt and Peixoto, 1985; P. phyllostomus Izecksohn, Cruz and Peixoto, 1998; P. sanctaritae Cruz and Napoli, 2010; P. subguttata Izecksohn, Cruz and Peixoto,1998; and P. tupinamba Prado and Pombal, 2008. Four species have been associated to the P. bigibbosa group, characterized by the presence of postocular swellings (Kwet & Faivovich 2001): P. avelinoi Mercadal de Barrio & Barrio, 1993; P. bigibbosa (Peters, 1872); P. brauni Kwet & Faivovich, 2001; and P. palustris Giaretta & Sazima, 1993. Finally, eight species have been associated with the P. cristicepsgroup, which gathers species lacking both prominent supraocular appendages and postocular swellings, but does not have any recognized shared derived characters (Giaretta et al. 2000): P. aridus Cruz, Nunes and Junca, 2012; P. caramaschi Cruz, Nunes and Junca, 2012: P. cristiceps (Müller, 1883); P. concavitympanum Giaretta, Bernarde & Kokubum, 2000; P. cururu Eterovick & Sazima, 1998; P. goyana (Miranda-Ribeiro, 1937); P. moratoi (Jim and Caramaschi, 1980); P. strussmannae Ávila, Kawashita-Ribeiro & Morais, 2011; and P. vielliardi Martins & Giaretta, 2011.

Herein, based on a series of specimens recently collected at the highlands of northern Chapada Diamantina, at Morro do Chapéu municipality, we describe a new dwarf species of Proceratophrys providing data on its natural history, its phylogenetic placement, as well as those for P. minuta, discussing the evolution of body shape and the validity of the recognized species groups within Proceratophrys.

Material and methods

Field sampling. Fieldwork was carried out during the rainy season, from 22nd December 2011 to 2nd January 2012, at Morro do Chapéu municipality. Ten pitfall traps with drift fences were installed at a plain sandy Caatinga, and 25 others at a montane forest at the slopes of Morro do Chapéu hill. Unquantified active search was also performed at the surrounding areas and at Ferro Doido waterfall.Collected specimens were killed with lethal doses of anesthesics, fixed in 10% formalin, preserved in 70% alcohol and housed at the herpetological collection of Museu de Zoologia da Universidade de São Paulo, São Paulo, Brazil.

Morphological assessment. Morphological comparisons were based on voucher specimens housed at the herpetological collection of the Museu de Zoologia da Universidade de São Paulo (MZUSP) and specimens collected by the Laboratório de Herpetologia, Instituto de Biociências da Universidade de São Paulo (MTR) (Appendix I), and data available in the literature (Giaretta & Sazima 1993; Mercadal de Barrio & Barrio 1993; Eterovick & Sazima 1998; Giaretta et al. 2000; Kwet & Faivovich 2001; Cruz et al. 2005; Prado & Pombal 2008; Cruz & Napoli 2010; Santana et al. 2010; Ávila et al. 2011; Martins & Giaretta 2011; Napoli et al. 2011; Cruz et al.2012).

Diagnostic features and morphological descriptions follow Prado and Pombal (2008) and Napoli et al. (2011). Measurements were taken using a digital caliper to the nearest 0.01 mm and rounded to the first decimal unit to avoid pseudoprecision (Hayek et al. 2001). Twelve morphometric measurements were taken following Heyer et al. (1990) and Prado and Pombal (2008): snout-vent length (SVL); head length (HL); head width (HW); eye diameter (ED); eye to nostril distance (END); upper eyelid width (UEW); interorbital distance (IOD); internostril distance (IND); thigh length (THL); tibia length (TL); foot plus tarsus length (FTL); and hand length (HAL). All individuals were measured, as all were consider adults: males with vocal slits and females with vitelogenic eggs visible though skin. Moreover individuals were found in amplexus, attesting for their maturity.

Geographical distribution. The geographical distribution of Proceratophrys was assessed, to present the distribution of the new species in the genus context and to add some data to the discussion on validity of some species. For this purpose records were compiled, mostly from literature (Miranda-Ribeiro 1955; Jim & Caramaschi 1980; Giaretta & Sazima 1993; Mercadal de Barrio & Barrio 1993; Rossa-Feres & Jim 1996; Caramaschi & Velosa 1997; Eterovick & Sazima 1998; Machado et al. 1999; Giaretta et al. 2000; Brandão & Araujo 2001; Giasson et al.2001; Kwet & Faivovich 2001; Boquimpani-Freitas et al. 2002; de Sá & Langone 2002; Lavilla et al. 2002; Kwet & Baldo 2003; Silvano & Pimenta 2003; Hiert & Moura 2004; Pimenta & Carvalho-e-Silva 2004; Cruz et al. 2005;

TEIXEIRA JR. ET AL.26 · Zootaxa 3551 © 2012 Magnolia Press

Brusquetti & Lavilla 2006; Leite et al. 2006; Cacciali et al. 2007; Vieira et al. 2007; de Lima 2007; Zanella & Busin 2007; Brasileiro et al. 2008; Gonsales 2008; Lucas & Foster 2008; Prado & Pombal 2008; Vieira et al. 2008; Amaro et al. 2009; Cintra et al. 2009; Oda et al. 2009; Santos et al. 2009; Caldart et al. 2010; Carosini et al. 2010; Carvalho et al. 2010; Cruz & Napoli 2010; Fatorelli et al. 2010; Rolim et al. 2010; Santana et al. 2010; Ávila et al.2011; Barros et al. 2011; Maffei et al. 2011; Martins & Giaretta 2011; Napoli et al. 2011; Valdujo et al. 2011; Bernardo et al. 2012; Brandão et al. 2012; Cruz et al. 2012; Mangia et al. 2012), and from examined specimens (Appendix I).

Molecular data. Partial sequences of two mitochondrial (16S and cyt b) and one nuclear (Rag-1) genes of two individuals from the new species, and two individuals of Proceratophrys minuta were obtained (Appendix II) and added in a matrix including Proceratophrys appendiculata, P. avelinoi, P. bigibbosa, P. boiei, P. concavitympanum, P. cristiceps, P. cururu, P. goyana, P. laticeps, P. melanopogon, P. moratoi, P. renalis, P. schirchi, Odontophrynus americanus, O. carvalhoi, O. cultripes, Macrogenioglottus alipioi, Cycloramphus acangatan and Thoropa taophora from Amaro et al. (2009).

Total genomic DNA was extracted from liver and muscle according to Fetzner (1999) and amplification was carried using standard PCR protocols with primers 16SAR and 16SBR (Palumbi 1996) for amplification of 16S, primers CB1 and CB3 for cyt b (Palumbi 1996) and primers R1GFF and R1GFR (Faivovich et al. 2005) for Rag-1. The PCR cycle protocol consisted of one initial cycle of 94 °C for 5 min followed by 35 cycles of 94 °C for 40 sec, 48 °C for 40 sec, 72 °C for 40 sec and the products were directly purified with Exonuclease I and Shrimp Alkaline Phosphatase (USB or Fermentas). Automated sequencing was carried out using BigDye Terminator v3.1 Cycle Sequencing kit (Applied Biosystems), followed by analysis on ABI Prism 310, 3700 or 3170 Genetic Analyzer Sequencers (Applied Biosystems) according to the manufacturer’s instructions. Sequences were edited and initially aligned in CodonCode Aligner (CodonCode Corporation).

The three data sets were combined in a matrix of 1558 bp (536 bp form 16S, 594 bp from cyt b and 428 bp from Rag-1) and different modes of inference were used in phylogenetic analyses: Bayesian analysis (BA) implemented in MrBayes 3.0b4 (Ronquist & Huelsenbeck 2003) and Maximum Likelihood (ML) in Treefinder (Jobb 2008). Some heterozygous sites were found for Rag-1 and were treated as ambiguities. Two independent Bayesian analyses were performed for each partition, with a random starting tree, four incrementally heated Markov chains, and 10,000,000 generations, with trees sampled every 1000 generations to estimate likelihood and sequence evolution parameters. For Bayesian analyses, the best-fit model of nucleotide substitution for each data partition was selected using the Akaike information criterion (AIC) and we used a separate model for each gene (GTR+I+G for 16S; HKY+I+G for cyt b ; HKY+G for Rag-1). Stationarity for each run was detected by plotting the likelihood scores of the trees against generation time, and the topology, posterior probability values, and branch lengths inferences were estimated after discarding 25% of the initial trees of each run as burn-in samples. Nodes with posterior probability ≥ 95% on a 50% majority rule consensus tree from both runs were considered significant support for a given clade. Maximum likelihood (ML) bootstrapping (1000 replicates) was performed in Treefinder (Jobb 2008) using default settings and models GTR + G for cyt b and 16S and HKY + G for RAG-1 (models were also estimated in Treefinder). The majority 50% consensus trees saved with posterior probabilities and bootstrap values on the nodes were visualized using FigTree 1.3.1 (http://tree.bio.ed.ac.uk/). Uncorrected genetic distances (pdistances) were calculated using PAUP* 4.0b10 (Swofford 2003).

Results

Proceratophrys redacta sp. nov.(Figs. 1–3)

Holotype. MZUSP 150266, an adult male, field number MTR 22579, collected at Morro do Chapéu hill, Morro do Chapéu municipality, Bahia, Brazil (11°35'26.02"S, 41°12'32.87"W, 1254 m a.s.l. datum WGS-84), by M. Teixeira

Jr, F. Dal Vechio, I. Prates, R. P. Damasceno and M. T. Rodrigues, on 30th December 2011.Paratopotypes. MZUSP 150263, 150264, 150265, 150268, 150269, adult males, field numbers MTR 22474,

22554, 22564, 22582, 22583; MZUSP 150267, adult female, field number MTR 22580. Collected between 27th and

30th December 2011.

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Paratypes. MZUSP 150272, an adult male, field number MTR 22628; MZUSP 150270, 150271, 150273 adult females, field numbers MTR 22623, MTR 22626, MTR 22629. All from an old road (11°33'58.3"S, 41°07'32.3"W, 929 m a.s.l., datum SAD69), at Morro do Chapéu municipality, close to city town. Collected between 31st

December 2011 and 2nd January 2012.

FIGURE 1. Proceratophrys redacta sp. nov. holotype (MZUSP 150266) in dorsal (left) and ventral (right) views (21.9 mm in SVL, male)

Diagnosis. A Proceratophrys species (nuptial pads absent, enlarged glands absent, fingers webbing absent, supernumerary tubercles on hands and feet present, dorsal surfaces of fingers and toes wrinkled, and dorsum with sinuous longitudinal ridges outlining a spear-shaped ornamentation) characterized by: (1) small size (26.9–29.7 mm of SVL in males; 33.6–34.9 mm of SVL in females); (2) no rostral appendage; (3) no palpebral appendage; (4) snout rounded from above and vertical in profile; (5) rostral ridge sharp, slightly curved; (6) presence of interrupted sagittal ridges of warts from eyelids to sacral region; (7) absence of postocular swellings; (8) tympanic region indistinct; (9) dorsal coloration lacking distinct dark ocelli.

Comparisons with other species (data for the species in comparison are given in parenthesis).Proceratophrys redacta sp. nov. can be promptly distinguished from all species of Proceratophrys, with the exception of P. avelinoi, P. minuta, P. moratoi and P. palustris, by its smaller body size, SVL ranging from 26.9–29.7 mm in males, and 33.6–34.9 mm in females (P. appendiculata: 40.4–59.9 males, 33.5–63.2 females; P. aridus: 32.5–42.8 males, 43.2–46.3 females; P. bigibbosa: 35.5–43.8 males, 51.2–53.4 females; P. boiei: 39.8–61.9 males, 40–74.3 females; P. brauni: 30–34.6 males, 38.9–39.8 females; P. caramaschii: 42.7–56.3 males, 53.8–65 females; P. concavitympanum: 39.6–57.1 males, 53.6–60.3 females; P. cristiceps: 39.5–50.2 males, 43–53.8 females; P. cururu: 36.5–43.1 males, 47–53.9 females; P. goyana: 39.6–53.5 males, 54.6–60.0 females; P. laticeps: 59.5–78 males, 50.3–82.7 females; P. melanopogon: 34.9–49.9 males, 40.7–62.1 females; P. moehringi: 59.2–62.6 males, 63.6 the only female known; P. paviotti: 41.9–53.2 males, 50.9–51.8 females; P. phyllostomus: 55.4 the only male known, 40.9–76.6 females; P. renalis: 36.8–53 males, 38.5–71.9 females; P. rondonae: 62.9 the only male knwon; P. sanctaritae: 38.4–45.5 males, 60.8 female; P. schirchi: 31.6–40.4 males; P. strussmanae: 41.1–47.3 males, 52.6–59.8 females; P. subguttata: 36.7–41.8 males, 36.6–58.9 females; P. tupinamba. 52.6–63.4 males, 37.7–72.8 females; P. vielliardi: 39.1–41.9 males, 45.5 the only female known; P. redacta sp. nov. can be distinguished from P. minuta by its larger size, SVL ranging from 26.9–29.7 mm in males, and 33.6–34.9 mm in females, by having an incomplete sagittal ridge of warts extending from eyelid to sacral region, and having a row of small, rounded tubercles in the eyelid (SVL 19.2–25.2 mm males, 25.9–31.9 mm females; presence of a


complete sagittal ridge of warts from eyelids to urostile; a row of enlarged, pointed tubercles in the eyelid is present); from P. avelinoi, P. palustris, P. brauni and P. bigibbosa, it can be distinguished by lacking postocular swellings (present) a additionally from P. avelinoi and also P. schirchi by having a row of small, rounded tubercles in the eyelid (a row of enlarged, pointed tubercles in the eyelid is present); from P. moratoi, P. palustris and P. vielliardi it can be distinguished by having the dorsal and lateral surfaces of body covered with small granules with a few scattered larger tubercles, incomplete sagittal ridge of tubercles extending from eyelid to sacral region, and dorsal coloration variable, but never with dark irregular ocelli (dorsal surfaces with many large tubercles, no sagittal ridge of tubercles and dorsal coloration with irregular ocelli).

Additionally P. redacta sp. nov. can also be distinguished from P. appendiculata, P. boiei, P. laticeps, P. melanopogon, P. moehringi, P. paviotti, P. phyllostomus, P. renalis, P. rondonae, P. sanctaritae, P. subguttata, and P. tupinamba by lacking a single long horn-like palpebral appendage (present); from P. appendiculata, P. melanopogon, P. laticeps, P. phyllostomus, P. subguttata and P. tupinamba by lacking a rostral appendage (present); and from P. concavitympanum and P. strussmannae by having an indistinct tympanic region (tympanic region defined by a concave depression on the skin).

FIGURE 2. Details of the head in dorsal view (A), and profile (B), and ventral views of right hand (C) and foot (D) of the holotype (MZUSP 150266) of Proceratophrys redacta sp. nov. Scale bars correspond to 5 mm.


Description of holotype. Body small, stout, nearly ovoid (Fig. 1); 28.3 mm in SVL. Head wider than long; head length 87% of head width. Snout rounded in dorsal view, vertical to slightly oblique in profile (Fig. 2). Upper eyelids without appendages, covered with a row of small rounded tubercles. Canthal crests distinct, loreal region slightly concave. Interocular crests absent. Interorbital distance 88% of upper eyelid width, 23.9% of head width. No oblique rows of warts on head sides. Three prominent and yellowish postrictal tubercles posteriorly to mouth commissure. Pre-ocular crests present, short and vertical. Eyes large, eye diameter 34% of head length, and 1.52 times the eye to nostril distance. Tympanum hidden. Nares prominent, nostrils elliptical, nearly horizontal in profile, diverging posteriorly in dorsal view. Tongue cordiform, free posteriorly. Vomerine teeth in two short rows between choannae. Choannae large, rounded, separated by a distance of about 2.5 times its diameter. Vocal sac single, subgular; vocal slits present, lateral to tongue on the floor of mouth. Arms short and robust; finger length IV<II<I<III. Finger tips rounded, not expanded. Fingers not webbed, bordered laterally by distinct ridges of tubercles. Nuptial pads absent in males. Inner carpal tubercle elliptical. Outer carpal tubercle bifid, ovoid with a ventral invagination inwards. Subarticular tubercles large, rectangular with rounded corners; supernumerary tubercles well defined, rounded. Outer margin of forearms with enlarged tubercles, but not forming well defined rows. Legs short and moderately robust, leg length (THL+TL) 75% of SVL, foot plus tarsus 91% of SVL. Toe lengths I<II<V<III<IV. Toe tips rounded, smooth, without expansions, bordered laterally by distinct ridges. Toes basally webbed, web formula I 0–1- II 1½–2- III 1-–3+ IV 3-–1- V. Inner metatarsal tubercle large, globular, and semicircular. Outer metatarsal tubercle small, rounded. Subarticular tubercles large, rectangular with rounded corners; supernumerary tubercles present. Tarsal tubercles in a ventrolateral row, forming a lateral ridge on foot. Dorsal surfaces granulate with larger granules irregularly distributed in the dorsum, especially within dorsal ridges. Symmetrical sinuous longitudinal ridges of tubercles on dorsum, from posterior border of eyelids and scapulae, converging at middle of the body, and fading posteriorly, not reaching the urostile. Ventral surfaces shagreened. A pair of enlarged light tubercles below the anal region.

Color in preservative. Dorsal background color light brown. Dark brown interorbital bar projecting posteriorly. Dark brown triangular areas irregularly delimited along the outer margins of the dorsal sagittal crests projecting posteriorly over scapulae, curving inwards, reaching sacrum region. A small chevron medially located at the dorsum. Irregular dark brown markings distributed over the dorsum and along flanks. Limbs with transverse dark brown bars. Ventrally, ground color yellowish-brown; darker brown on palms, soles, anal region and throat. Three oblique dark brown stripes on sides of head, accompanying ridges of tubercles, and two less defined on snout to lips. Upper lip cream.

Measurements of holotype (in mm). SVL 28.3; HL 11.0; HW 12.6; ED 3.8; END 2.5; UEW 3.4; IOD 3.0; IND 2.1; THL 11.4; TL 9.9; FTL 15.9; HAL 12.4.

Variation. We found great variation on color pattern among specimens, the more common individuals have a similar pattern to that described for the holotype (Fig. 3A and C) as well as a more melanic-marbled pattern (Fig. 3B). Although less common, a pattern with a yellowish brown inner to the borders of the sagittal crests is also found (Fig. 3D). Although males and females share the same dorsal color variation, the throat in males present a dark brown color, and females a whitish coloration. Measurements for P. redacta sp. nov. and P. minuta are presented in Table 1.

Etymology. The name of the new species is derived from the Latin adjective redactus that means reduced, in allusion to the small size of the new species.

Natural history and distribution. Proceratophrys redacta sp. nov. is known only from the surroundings of type locality, at the montane region of Morro do Chapéu municipality, close to where P. minuta occurs (about 100 km away) (Fig. 4). The region is dominated by the Mesoproterozoic sandstones of Morro do Chapéu formation (Battilani et al. 1996), reaching up to 1300 m a.s.l., the climate is a sub-warm semi-humid with annual mean temperatures under 22oC, (4oC was already recorded), and precipitation is regular with a short dry season (Nimer 1972). Landscape in the area consists of an extensive and relatively flat plateau situated around 1200 m from which the hill of Morro do Chapéu (“hat hill”) emerges. At the plain sandy depositional areas grows an open shrubby vegetation, however at the richer soil areas a semi-deciduous forest is found; the bare rock at the mountain tops and eroded areas are covered by a typical rocky meadows vegetation, where Cactaceae, Bromeliaceae, and Velloziaceae are abundant. Steeper areas are covered by a lower open forest, the ground is sandy and covered with a deep layer of leaves and a few scattered small ponds are present, surrounded with mosses.


FIGURE 3. Individuals of Proceratophrys redacta sp. nov. in life. Adult male (A); melanic-marbled male individual (B); an adult female (C) and a couple in amplexus (D).


FIGURE 4. Distributional map Proceratophrys redacta sp. nov. (red stars) and P. minuta (blue circles) at the northeastern portion of Chapada Diamantina, Bahia, Brazil. White dots represent cities.


TABLE 1. Morphometric data for Proceratophrys redacta sp. nov. and P. minuta. Measurements abbreviations are explained in the text. Average values are given (in mm), followed by range, in parenthesis.

P. redacta sp. nov. was found active on the ground at night, within the montane forest and the adjacent open vegetation (Fig. 5). It was also captured in pitfall traps, along the Morro do Chapéu hill, about 1250 m a.s.l. and in the dense vegetation at the shores of streams running over bare rock outcrops, as those of Ferro Doido waterfall at about 890 m a.s.l. No individual was heard calling. Also none was found at the dry sandy scrublands, the most abundant vegetation at this area, indicating a restriction to this open mesic areas.

Molecular phylogeny. Our analyses recovered Proceratophrys as monophyletic, and consisting of two main clades. The first is composed by P. cristiceps, P schirchi, P. minuta and P. redacta sp. nov. All analyses recovered Proceratophrys redacta sp. nov. as sister to P. minuta, both being sister of a clade assembling P. cristiceps and P. schirchi (Fig. 6). The second clade is composed by the remaining species of Proceratophrys, however the intrageneric relationships are poorly resolved. Three different lineages were recovered in this clade: 1) P. appendiculata and P. melanopogon as sister species; 2) P. avelinoi and P. bigibbosa as sister species; and 3) a clade containing P. boiei, P. goyana, P. concavitympanum, P. moratoi, P. cururu, P. renalis, and P. laticeps. In the latter Proceratophrys moratoi is sister to P. concavitympanum and both sister to P. boiei and P. goyana. This latter clade is sister to a clade where Proceratophrys laticeps is basal to P.cururu and P. renalis, which are recovered as sister species.

Uncorrected p-distances ranged from 12 to 21% for cyt b, 3 to 11% for 16S and 1 to 5% for Rag-1 among Proceratophrys species (Table 2). Genetic distances between P. minuta and P. redacta sp. nov. were 12-13% for cyt b, 4% for 16S and 2% for Rag-1 and are compatible with interspecific distances found among Proceratophrysspecies (Table 2). It is important to note that none of the differences found on Rag-1 between P. minuta and P. redacta sp nov. were related to heterozygous sites.

Discussion

Among the Proceratophrys species, the ones lacking eyelid appendages such as those included in the P. cristicepsgroup, are difficult to identify confidently. The species here described as well as P. minuta, are good examples; they are morphologically very similar to P. cristiceps, but as they are much smaller, the size helps to diagnose them. However, among the species with similar sizes the distinction is not so clear. Such problematic diagnoses might result in possible misidentifications as those visualized on our map (Fig. 7). Individuals identified as P. cristiceps(Cintra et al. 2009; Oda et al. 2009), thought to be a Caatinga dweller, are reported deep into the Cerrado biome, where P. goyana is widespread and thought to be endemic. Similarly individuals occurring deep in to the Caatinga biome, were identified as P. goyana (Ávila et al. 2011). Highlighting this confusion Cruz et al. (2012) redescribed P. cristiceps, and two additional related new species, restricting the distribution of P. cristiceps to the coastal

Proceratoprys redacta sp.n. Proceratophrys minuta

Males (N=6) Females (N=4) Males (N=10) Females (N=11)

SVL 28.4 (26.9–29.7) 34.4 (33.5–34.9) 20.9 (19.2–22.6) 28.9 (25.9–31.6)

HL 11.6 (10.8–12.4) 13.3 (12.7–14.1) 9.1 (8.6–9.5) 11.2 (10.5–12.2)

HW 13 (12.2–13.8) 15.7 (15.2–16.2) 10.0 (9.2–10.8) 13.2 (12.2–14.1)

ED 4.1 (3.7–4.4) 4.6 (4.5–4.7) 2.6 (2.4–2.9) 3.2 (2.9–3.6)

END 2.5 (2.4–2.6) 2.9 (2.7–3.1) 2.1 (1.8–2.3) 2.6 (2.4–3.2)

UEW 3.4 (3.3–3.7) 4 (3.8–4.2) 3.1 (2.8–3.4) 3.8 (3.4–4.3)

IOD 3 (2.4–3.4) 3.4 (3.1–3.8) 2.5 (2.3–3.0) 3.2 (3.1–3.4)

IND 1.9 (1.7–2.2) 2.1 (2–2.5) 1.8 (1.6–1.9) 2.2 (2.1–2.3)

THL 12.1 (11.3–12.8) 13.9 (13.1–14.5) 9.5 (8.6–10.4) 11.9 (11.3–12.4)

TL 10.2 (9.8–10.5) 12 (11.9–12.2) 8.5 (8.1–9.1) 10.9 (10.5–11.3)

FTL 17.2 (15.9–18.4) 20.1 (19.7–20.4) 13.1 (11.9–14.1) 17.2 (15.8–17.8)

HAL 13.3 (12.3–14.2) 23.9 (15.9–46.4) 11.1 (10.4–12.2) 14.5 (14.1–14.8)


lowland and suggesting that individuals from Espinhaço range should be considered P. cururu, while those west from it, at the Cerrado, should be considered P. goyana. Unfortunately the authors did not examined specimens from those localities, and other areas at the Caatinga, thus leaving Caatinga specimens without a specific name. Thus these dubious identifications from some localities mentioned above, and the recent results on Proceratophrystaxonomy (Prado & Pombal 2008; Ávila et al. 2011; Martins & Giaretta 2011; Napoli et al. 2011; Cruz et al. 2012) strongly indicate that a rich cryptic diversity is still uncovered, highlighting the urgently need for a taxonomic revision of several species, especially on the widespread appendageless ones.

FIGURE 5. Habitat in which Proceratophrys redacta sp. nov. was found. (A) general view of Morro do Chapéu hill, which reaches 1300 m a.s.l., showing the shrubby vegetation at the lower lands and the montane forest at the slopes close to the top; (B) General view of Ferro Doido waterfall; (C) detailed view of the shrubby vegetation over bare rock at Ferro Doido waterfall; (D) and detailed view from the inside of the forest at the top of Morro do Chapéu hill.


FIGURE 6. Bayesian tree topology obtained from the combined molecular data set (16S, cyt b and Rag-1); figure shows the 50% majority-rule consensus phylogram. Posterior probability and ML bootstrap values are shown above the branches.

Despite the taxonomic clarity, the geographical distributions of Proceratophrys morphotypes bring some insights on the evolution of body shape. Although intermediate forms can also be found, two patterns can be visualized: (i) large-bodied species with median to long palpebral and rostral appendages are generally found at the warm/ wet forests, and (ii) small-bodied species with very short or no appendages are generally found in open and/or dryer habitats, at high altitude and/or latitude.

This switch in body shape could be the result of a process such as Allen’s rule, in which reducing body surfaces (e.g. by reducing appendages) (Alho et al. 2011) would be adaptive for maximize heat gain and minimize water loss at cold and/or dry areas, as the open environments are generally dryer and/or cooler than the forest ones. However physiological analyzes, together with further morphological and phylogenetic data are needed to attest this suggestion.

The dwarf species P. redacta sp. nov. and P. minuta are currently known only from a few highland areas at Espinhaço range, and seems to be sister to either a Caatinga dweller (P. cristiceps) or a Atlantic Forest one (P.

schirchi). Nonetheless they are found in broad sympatry with P. cristiceps, however not syntopically. The environmental climatic conditions as well as microhabitat availability at both above and below 800 m a.s.l. at the Espinhaço range may be representing an eco-physiological barrier (Lomolino et al. 2006) for those species, where P. cristiceps do not climb up, and P. redacta sp. nov. and P. minuta do not go down, been therefore restricted to the highland habitats.


FIGURE 7. Distributional records of all current recognized Proceratophrys species, including the new species described here: P. redacta sp. nov.

Considering this montane restriction, the disjunct distribution of P. minuta at Miguel Calmon and Palmeiras, having P. redacta sp. nov. in its middle (Fig. 4), deserves further investigation as these areas are presently separated by lowlands covered with Caatinga vegetation. It will be important in the future to analyze population variation through both molecular and morphological approaches to verify their specific identity.

As in the former results (Amaro et al. 2009; Pyron & Wiens 2011), in our phylogenetic analysis the traditional morphological groups are not recovered as monophyletic. Nonetheless the phylogenies published to date (Amaro et al. 2009; Pyron & Wiens 2011) are likely to change considerably in the future as the new recently described Proceratophrys species (Cruz et al. 2005; Prado & Pombal 2008; Cruz & Napoli 2010; Ávila et al. 2011; Martins & Giaretta 2011; Napoli et al. 2011; Cruz et al. 2012) are to be included, what would confirm that the diversity within the genus is clearly underestimated.


Acknowledgements

We are grateful to Ivan Prates, Roberta Pacheco Damasceno, Caroline Garcia, Milena Ferreira dos Santos and Eric Cazetta for assistance in the field work and to Sabrina Baroni for technical assistance, Hussam Zaher and Carolina Castro Mello for access to specimens, and Instituto Chico Mendes de Conservação da Biodiversidade (ICMBio) for collection authorization (permit number 19754-1). We would also like to thank Miguel Vences, Paula Valdujo and an anonymous reviewer for valuable comments, corrections and suggestions on early versions of this manuscript. We thank Fundação de Apoio à Pesquisa do Estado de São Paulo (FAPESP), Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq), and National Science Foundation (NSF) for support.

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APPENDIX I. Material examined.

Proceratophrys concavitympanum—Brazil: Tocantins: Palmas, MRT 6826–6827.Proceratophrys cristiceps—Brazil: Pernambuco, São José dos Cordeiros, MRT 9986–9995; Ceará: Macuripe, MZUSP

88427–88434; Rio Grande do Norte: Angicos, MZUSP 88427–88434.Proceratophrys cururu—Brazil: Minas Gerais: Santana do Riacho, Parque Nacional da Serra do Cipó, MTR 19628;

MTR19629; MTR 19639. Itabira, Serra dos Alves, MTR 21764, MTR 21853.Proceratophrys minuta—Brazil: Bahia: Miguel Calmon, Parque Estadual de Sete Passagens, MZUSP 146197–146536.Proceratophrys goyana—Brazil: Goiás: UHE Serra da Mesa, MZUSP 89104–89113.Proceratophrys schirchi—Brazil: Bahia: Amargosa, MZUSP 142721–14724; Minas Gerais: Jequitinhonha: Reserva Biológica

da Mata Escura, MTR 17320–17321; MTR 17351.


APP

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F (L

abor

atór

io d

e C

itoge

nétic

a de

Ver

tebr

ados

, Ins

titut

o de

Bio

ciên

cias

, Uni

vers

idad

e de

São

Pau

lo, B

razi

l), JC

(fie

ld n

umbe

r of J

osé

Cas

sim

iro, U

nive

rsid

ade

de S

ão P

aulo

, Bra

zil),

FSF

L (F

ield

num

ber o

f Fel

ipe

Sá F

orte

s Lei

te,

Pont

ifíci

a U

nive

rsid

ade

Cat

ólic

a de

Min

as G

erai

s, B

razi

l), C

H (C

élio

F.B

. Had

dad,

tiss

ue c

olle

ctio

n , U

nive

rsid

ade

Esta

dual

Pau

lista

, Rio

Cla

ro, S

ão P

aulo

, Bra

zil),

DB

(D

iego

Bal

do, U

nive

rsid

ad N

acio

nal d

e M

isio

nes,

Arg

entin

a), M

ZUSP

(Mus

eu d

e Zo

olog

ia, U

nive

rsid

ade

de S

ão P

aulo

, Bra

zil)

ZUFR

J (D

epar

tam

ento

de

Zool

ogia

, Ins

titut

o de

Bio

logi

a, U

nive

rsid

ade

Fede

ral d

o R

io d

e Ja

neiro

, Bra

zil).

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enB

ank

acce

ssio

n nu

mbe

r Sp

ecie

s V

ouch

er Id

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calit

y cy

t b

16S

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mph

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cang

atan

A

F 16

05

Cau

caia

do

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ia, S

P FJ

6856

63

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FJ

6857

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Mac

roge

niog

lottu

s alip

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AF

919

Serr

a do

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mos

o, Ju

ssar

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A

FJ68

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6856

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FJ68

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acro

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F 16

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FJ

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FJ68

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FJ

6857

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O. c

arva

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JC

122

4 M

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A

FJ68

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FJ

6856

87

FJ68

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O

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tripe

s FS

FL 8

75

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ginh

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G

FJ68

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6856

88

FJ68

5708

O

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atoi

C

H 5

11

Itira

pina

, SP

FJ68

5669

FJ

6856

89

FJ68

5709

Pr

ocer

atop

hrys

app

endi

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taA

F144

6 Sa

ntos

, SP

JX98

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JX

9870

66

JX98

7067

P.

ave

linoi

D

B 1

246

Mis

ione

s, A

rgen

tina

FJ68

5671

FJ

6856

91

FJ68

5711

P.

boi

ei

AF

1587

Se

rra

da C

anta

reira

, São

Pau

lo, S

P FJ

6856

72

FJ68

5692

FJ

6857

12

P. b

igib

bosa

D

B 2

313

Mis

ione

s, A

rgen

tina

FJ68

5673

FJ

6856

93

FJ68

5713

P.

con

cavi

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panu

m

AF

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U

sina

Hid

relé

trica

de

Laje

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mas

, TO

FJ

6856

74

FJ68

5694

FJ

6857

14

P. c

ristic

eps

AF

887

Parq

ue N

acio

nal d

a Se

rra

das C

onfu

sões

, PI

FJ68

5675

FJ

6856

95

FJ68

5715

P.

cur

uru

FSFL

580

C

arde

al M

ota,

MG

FJ

6856

76

FJ68

5696

FJ

6857

16

P. g

oyan

a A

F 11

88

Petro

lina

de G

oiás

, GO

FJ

6856

77

FJ68

5697

FJ

6857

17

P. la

ticep

s A

F 19

00

Res

erva

Nat

ural

da

Val

e do

Rio

Doc

e, L

inha

res,

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6856

78

FJ68

5698

FJ

6857

18

P. m

elan

opog

on

CH

362

M

arin

gá, I

tatia

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J FJ

6856

79

FJ68

5699

FJ

6857

19

P. m

inut

a A

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8 M

igue

l Cal

mon

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JX

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61

JX98

2965

JX

9829

69

P. m

inut

a A

F200

9 M

igue

l Cal

mon

, BA

JX

9829

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JX98

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JX

9829

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P. re

dact

a sp

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TR22

579

Mor

ro d

o C

hapé

u, B

A

JX98

2963

JX

9829

67

JX98

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P.

reda

cta

sp. n

ov.

MTR

2258

2 M

orro

do

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péu,

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JX

9829

64

JX98

2968

JX

9829

72

P. re

nalis

ZU

FRJ 8

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jo M

adre

de

Deu

s, PE

FJ

6856

80

FJ68

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6857

20

P. sc

hirc

hi

CH

371

Sã

o Lo

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ço, S

anta

Ter

esa,

ES

FJ68

5681

FJ

6857

01

FJ68

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Th

orop

a ta

opho

ra

AF

1434

Sa

ntos

, SP

FJ68

5662

FJ

6856

82

FJ68

5702


a new dwarf species of proceratophrys miranda-ribeiro, 1920

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