Research Article |
Corresponding author: Rommel R. Rojas ( rrojaszamora@gmail.com ) Academic editor: Franco Andreone
© 2016 Rommel R. Rojas, Juan Chaparro, Vinicius Carvalho, Robson Avila, Izeni Farias, Tomas Hrbek, Marcelo Gordo.
This is an open access article distributed under the terms of the Creative Commons Attribution License (CC BY 4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Citation:
Rojas RR, Chaparro JC, De Carvalho VT, Ávila RW, Farias IP, Hrbek T, Gordo M (2016) Uncovering the diversity in the Amazophrynella minuta complex: integrative taxonomy reveals a new species of Amazophrynella (Anura, Bufonidae) from southern Peru. ZooKeys 563: 43-71. https://doi.org/10.3897/zookeys.563.6084
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A new species of the genus Amazophrynella (Anura, Bufonidae) is described from the departments of Madre de Dios, Cusco and Junin in Peru. An integrative taxonomy approach is used. A morphological diagnosis, morphometrics comparisons, description of the advertisement call, and the phylogenetic relationships of the new species are provided. Amazophrynella javierbustamantei sp. n. differs from other species of Amazophrynella by: intermediate body-size (snout-vent length 14.9 mm in males, n = 26 and 19.6 mm in females, n = 20), tuberculate skin texture of body, greatest hand length of the Amazophrynella spp. (3.6 mm in males, n = 26 and 4.6 mm in females, n = 20), venter coloration yellowish, tiny rounded black points covering the venter, and thirteen molecular autapomorphies in the 16S RNA gene. Its distribution varies from 215 to 708 m a.s.l. This discovery highlights the importance of the remnant forest in preserving the biodiversity in Peru, and increase in seven the species formally described in the genus Amazophrynella.
Resumen
Describimos una nueva especie del género Amazophrynella (Anura, Bufonidae) del Perú de los Departamentos de Madre de Dios, Cusco y Junin de Peru. Utilizamos un método de taxonomía integrativa. Obtuvimos la diagnosis morfológica, comparaciones morfométricas, descripción del canto de reproducción y las relaciones filogenéticas de la nueva especie. A. javierbustamantei sp. n. difiere de las otras Amazophrynella spp. por poseer tamaño medio (Hocico-cloaca en machos 16.9 mm, n = 26 y en hembras 19.6 mm, n = 20); textura de la piel tuberculada; tamaños de las manos mayores (3.6 mm en machos, n = 26 y 4.6 mm en hembras, n = 20); coloración ventral amarillento-pálida, pequeños puntos redondos de color negro en el vientre y por trece autopomorfias moleculares en el gen 16S RNA. Su distribución varía desde 215 m hasta 708 m a.s.n.m. Este descubrimiento resalta la importancia de los remanentes de la selva Peruana en términos de conservación, e incrementa en siete las especies formalmente descritas en del género Amazophrynella.
Resumo
Descrevemos uma nova espécie do gênero Amazophrynella (Anura, Bufonidae) dos departamentos de Madre de Dios, Cusco e Junin do Peru. Utilizamos um método de taxonomia integrativa. Apresentamos a diagnose morfológica, comparações morfométricas, descrevemos o canto de anúncio e geramos uma hipótese filogenética da nova espécie. Amazophrynella javierbustamantei sp. n. difere das outras Amazophrynella spp. por possuir tamanho médio (Comprimento rostro-cloacal 16.9 mm em machos, n = 26 e 19.6 mm em fêmeas, n=20); textura da pele tuberculada; tamanhos das mãos maiores (3.6 mm em machos, n = 26 e 4.6 mm em fêmeas, n = 20); coloração ventral amarelo-clara, coberta por pequenos pontos redondos pretos e por treze autapomorfias moleculares no gene 16S RNA. Sua distribuição varia entre os 215 m até os 708 m a.n.m. Nossa descoberta aumenta a importância dos remanescentes da floresta Peruana em termos de conservação e incrementa em sete as espécies formalmente descritas no gênero Amazophrynella.
Amphibian, Tree Toad, conservation, Southern Peru, integrative taxonomy
Anfibios, Sapo del árbol, conservación, Sur del Perú, taxonomía integrativa
Anfíbios, Sapo do arvore, conservação, Sul do Peru, taxonomia integrativa
Until 2012, two species of Amazophrynella were placed in the genus Dendrophryniscus Jimenez de la Espada, 1868.
In the following years, an additional four new species of the genus were described: A. vote Ávila, Carvalho, Gordo, Ribeiro & Morais, 2012 and A. manaos Rojas, Carvalho, Gordo, Ávila, Farias & Hrbek, 2014 based on morphology; A. amazonicola and A. matses Rojas, Carvalho, Gordo, Ávila, Farias & Hrbek, 2015, based on morphology and genetic data (
Using a phylogenetic analysis based on mitochondrial and nuclear genes (
Given this scenario, herein is described an additional new species of Amazophrynella from the departments of Madre de Dios, Cusco and Junin, Peru, founded on the principles of integrative taxonomy. Morphological, morphometric, bioacoustic and phylogenetic relationships are provided as evidence for the existence of the new taxon.
Forty eight specimens previously identified as Amazophrynella minuta (Melin, 1941), deposited at the
Additionally five preserved specimens of Amazophrynella bokermanni (Izecksohn, 1993) from near the type locality (Juruti, 30 Km from type locality), the holotype and paratypes of A. manaos deposited in the Collection of Amphibians and Reptiles of the
Morphological character analyses were carried out according to
Measurements were carried out with a digital caliper following
Statistical analysis. We used a total of 80 adult males of the Amazophrynella minuta species complex (numbers of individuals and populations of origin in parentheses): A. minuta sensu stricto (n = 23, from Taracuá), A. amazonicola (n = 15, from Puerto Almendras and Fazenda Zamora); A. matses (n = 13, from Nuevo Salvador) and the new species of Amazophrynella (n = 29, from Tambopata, Nuevo Arequipa, Candamo and Inambari).
All morphometric measures were log10 transformed to conform to requirements of normality (
Laboratory procedure. Total DNA was extracted from muscle tissue using standard phenol/chloroform extraction (
Phylogenetic analysis. We obtained 16S rDNA sequence data from two specimens of the new species (Accession numbers: KR905184, KR905185), two paratypes of A. vote (Accession numbers: KF433970, KF433971), two specimens of A. bokermanni (Accession numbers: KF433975, KF433976), two topotypic specimens of A. minuta (Accession numbers: KF792834, KF792836), two paratopotypes of A. matses (Accession number: KP681688, KP681689), the holotype and one paratopotype of A. amazonicola (Accession number: KP681868, KP681669) and two paratypes of A. manaos (Accession number: KF433954, KF433957) deposited in the tissue collection of the Laboratório de Evolução e Genética Animal of the Universidade Federal do Amazonas (CTGA-ICB/UFAM). The dataset also included two sequences of A. sp. aff. minuta (Accession number: AY326000, DQ158420) from
Sequences were aligned using the Clustal W algorithm (Thompson et al. 1996) implemented in BioEdit (Hall 1999) and alignment was adjusted as necessary against the secondary structure of the 16S rDNA. The existence of lineages in a phylogenetic tree-based context (
Molecular species delimitation. Evolutionary lineages are diagnosed by discontinuities in character variation among lineages, and correspond to phylogenetic species. The existence of lineages is therefore a necessary and sufficient prerequisite for inferring the existence of a species under the different conceptualizations of the Phylogenetic Species Concept (PSC) (
We analyzed one advertisement call obtained from the CD of Frogs of Tambopata, Peru (Macauly Library of Natural Songs and Cornell Laboratory of Ornithology) by the authors
In the resulting phylogeny, the six nominal species of Amazophrynella were recognized as monophyletic (Fig.
In the first clade the Amazophrynella species: A. manaos is sister taxon of the possible new specie from the Guiana Shield: A. sp. aff. manaos (bootstrap support= 91), and both are sister to A. bokermanni (bootstrap support= 98). Amazophrynella vote is sister of A. bokermanni + (A. manaos + A. sp. aff. manaos) with a bootstrap support of 81.
The second clade corresponding to the A. minuta “species complex”, A. amazonicola is sister of A. minuta + A. sp. aff. minuta from western Amazonia (bootstrap support= 99). Our analysis further highlighted the occurrence of a new monophyletic lineage (A. javierbustamantei sp. n.) showing sister relationship with A. matses (bootstrap support = 96), both being in turn sister group of A. amazonicola + (A. minuta + A. sp. aff. minuta) with a bootstrap support of 99.
Smallest uncorrected 16S rDNA p-distances estimated between phylogenetic linages was observed between A. minuta and A. sp. aff. minuta (= 3%). Greatest interspecific distance (= 14%) was observed between Amazophrynella javierbustamantei sp. n. and A. bokermanni and was comparable to divergence observed between A. manaos and A. minuta. Within the “A. minuta” species complex, the new species shows a high degree of genetic divergence from A. minuta (= 7%), A. amazonicola (= 9%) and minor genetic distance with their sister taxon A. matses (= 3%) (see all pairwise genetic distance values summarized in Table
Comparative analysis of quantitative morphological data allowed us to distinguish Amazophrynella sp. n. from the other members of the A. minuta “species complex”. The first two principal components extracted by the PCA account for 48.56% of the variation found in the dataset. The first component (PC1) explained 24.93% of total variation. In the first principal component axis, A. amazonicola is distinguished from the other species due to its larger size (SVL = 14.9 ± 0.7 mm, see Table
Measurements (mm) of adult male specimens (including the holotype) in the type series Amazophrynella spp. Mean ± standard deviation, with ranges in parentheses. Abbreviations are defined in material and methods.
Variable |
A. minuta sensu stricto (n = 15) |
A. manaos (n = 29) |
A. bokermanni (n = 5) |
A. vote (n = 14) |
A. amazonicola (n = 15) |
A. matses (n = 13) |
A. javierbustamantei sp. n. (n = 26) |
---|---|---|---|---|---|---|---|
SVL | 13.5 ± 0.6 (12.5–14.2) |
14.2 ± 0.7 (12.3–15.0) |
16.8 ± 1.4 (14.6–18.2) |
13.1 ± 0.7 (12.0–14.1) |
14.5 ± 0.7 (13.3–15.4) | 12.1±0.6 (11.5–13.5) | 14.9 ± 0.9 (12.7–16.4) |
HW | 4.2 ± 0.2 (4.0–4.3) | 4.2 ± 0.3 (3.7–4.7) | 3.2 ± 0.3 (2.5–3.3) | 4.0 ± 0.7 (3.3–4.4) | 4.4 ± 0.3 (4.2–4.6) | 3.6 ±0.2 (3.1–3.8) | 4.2 ± 0.2 (3.5–4.7) |
HL | 4.9 ± 0.2 (4.8–5.3) | 5.3 ± 0.3 (4.7–5.6) | 3.4 ± 0.4 (2.8–3.8) | 4.6 ± 0.3 (4.0–5.2) | 5.2 ± 0.3 (5.0–6.2) | 4.3 ± 0.3 (3.9–4.8) | 5.1 ±0.3 (4.4–5.6) |
SL | 2.3 ± 0.1 (2.2–2.5) | 2.7 ± 0.2 (2.3–2.7) | 3.0 ± 0.4 (2.2–3.1) | 2.1 ± 0.2 (1.9–2.6) | 2.4 ± 0.2 (2.2–2.5) | 2.0 ± 0.3 (1.6–2.3) | 2.2 ± 0.2 (1.7–2.6) |
ED | 1.4 ± 0.1 (1.3–1.5) | 1.3 ± 0.1 (1.2–1.6) | 1.8 ± 0.2 (1.5–2.0) | 1.3 ± 0.1 (1.2–1.5) | 1.2 ± 0.1 (0.9–1.2) | 1.1 ± 0.1 (0.9–1.2) | 1.3 ± 0.1 (1.0–1.6) |
IND | 1.2 ± 0.1 (1–1.3) | 1.1 ± 0.1 (1.0–1.4) | 1.4 ± 0.2 (1.0–1.5) | 1.1 ± 0.1 (1.0 –1.3) | 1.2 ± 0.1 (1.0–1.3) | 1.0 ± 0.1 (0.8–1.2) | 0.9 ± 0.1 (0.8–1.2) |
UAL | 3.8 ± 0.2 (3.2–4.1) | 3.6 ± 0.4 (2.9–4.1) | 5.5 ± 0.6 (5.0–5.6) | 3.9 ± 0.5 (2.8–3.9) | 4.5 ± 0.3 (4.2–5.3) | 3.5 ± 0.4 (2.9–4.2) | 4.5 ± 0.4 (3.8–5.7) |
HAL | 2.8 ± 0.2 (2.6–3.0) | 2.8 ± 0.6 (1.9–2.9) | 3.4 ± 0.6 (2.8–4.2) | 2.7 ± 0.3 (2.3–3.2) | 3.2 ± 0.2 (2.8–3.3) | 2.7 ± 0.2 (2.3–3.1) | 3.6 ± 0.4 (2.5–4.5) |
THL | 6.8 ± 0.2 (6.4–7.2) | 6.7 ± 0.3 (2.3–3.1) | 8.7 ±1.4 (7.2–8.9) | 6.5 ± 0.7 (5.4–7.2) | 7.7 ± 0.6 (6.3–8.0) | 6.2 ± 0.4 (5.1–6.3) | 7.6 ± 0.7 (6.2–9.2) |
TAL | 6.7 ± 0.3 (6.3–7.1) | 6.9 ±0.6 (4.2–7.3) | 8.3 ± 1.0 (6.7–9.2) | 5.7 ± 0.7 (4.8–7.0) | 7.2 ± 0.6 (6.1–7.9) | 5.8 ± 0.3 (5.1–6.3) | 7.6 ± 0.7 (6.2–8.8) |
TL | 4.1 ± 0.2 (3.8–4.6) | 4.6 ± 0.4 (4.3–6.3) | 5.4 ± 1.4 (2.9–6.2) | 3.8 ± 1.0 (4.2–7.0) | 4.2 ± 0.6 (6.3–8.0) | 3.8 ± 0.2 (3.6–4.3) | 4.7 ± 0.8 (3.9–8.7) |
FL | 4.8 ± 0.4 (4.2–5.2) | 5.2 ± 0.5 (4.7–6.1) | 6.3 ± 1.3 (3.9–7.6) | 4.4 ± 0.6 (3.2–5.4) | 5.1 ± 0.4 (4.7–6.0) | 4.3 ± 0.4 (5.5–3.0) | 5.7 ± 0.6 (4.5–7.2) |
Individuals of the genus Amazophrynella (A) and Dendrophryniscus (D) used in the molecular analyses. Information includes samples, collecting locality, GenBank accession number for the 16S rDNA fragment, voucher number and specimen status.
Sample | Locality | Accession Number | Voucher number | Specimen status |
---|---|---|---|---|
A. javierbustamantei | Quebrada Guacamayo, Peru | KR905184 | MHNC 8331 | Holotype |
A. javierbustamantei | Quebrada Guacamayo, Peru | KR905185 | MHNC 8363 | Paratype |
A. matses | Nuevo Salvador, Peru | KF681688 | MZUNAP 928 | Paratopotype |
A. matses | Nuevo Salvador, Peru | KF681689 | MZUNAP 941 | Paratopotype |
A. minuta sensu stricto | Taracuá, Brazil | KF792834 |
|
Topotype |
A. minuta sensu stricto | Taracuá, Brazil | KF792835 |
|
Topotype |
A. amazonicola | Puerto Almendras, Peru | KF681868 | MZUNAP 901 | Holotype |
A. amazonicola | Puerto Almendras, Peru | KF681669 | MZUNAP 915 | Paratopotype |
A. vote | Parque Nacional Nascentes do Lago Jari, Brazil | KF433970 |
|
Paratype |
A. vote | Parque Nacional Nascentes do Lago Jari, Brazil | KF433971 |
|
Paratype |
A. bokermanni | Juruti, Pará, Brazil | KF433975 |
|
|
A. bokermanni | Juriti, Pará, Brazil | KF433976 |
|
|
A. manaos | Mineração taboca, Brazil | KF433954 |
|
Paratype |
A. manaos | Mineração taboca, Brazil | KF433957 |
|
Paratype |
A. sp. aff. manaos | Mitaraka, French Guiana | JN867570 | 296MC | |
A. sp. aff. manaos | Mitaraka, French Guiana | EU201057 | 3035T | |
A. sp. aff. minuta | Rio Lagarto Cocha, Peru | AY326000 |
|
|
A. sp. aff. minuta | Equador | DQ158262 | QCAZ833 | |
D. proboscideus | Mata Escura, Brazil | JN867566 | MTR17173 | |
D. proboscideus | Mata Escura, Brazil | JN867564 | MTR17171 | |
D. oreites | Serra das lontras, Brazil | JN867567 | MTR16368 | |
D. carvalhoi | Parna Caparão, Brazil | JN867568 | MTR15755 | |
D. carvalhoi | Parna Caparão, Brazil | JN867569 | MTR15757 | |
D. leucomyxtas | Ilha grande, Brazil | JN867558 | MTR15547 | |
D. leucomyxtas | Ilha grande, Brazil | JN867557 | MTR15548 | |
D. berthalutzae | Treviso, Brazil | JN867551 | CFBH10322 | |
D. brevipollicatus | Estação Biológica de Boracia, Brazil | JN867554 | AF1541 | |
D. brevipollicatus | Estação Biológica de Boracia, Brazil | JN867553 | AF1175 |
Uncorrected p-distances between Amazophrynella (A), species and the sister genus Dendrophryniscus (D). Molecular distances are based on the 480-bp fragment of 16S rDNA. We included A. minuta sensu stricto from its type locality and two candidate species, Amazophrynella sp. aff. manaos and A. sp. aff. minuta mentioned in
16S rDNA | 1 | 2 | 3 | 4 | 5 | 6 | 7 | 8 | 9 |
---|---|---|---|---|---|---|---|---|---|
1 A. amazonicola | |||||||||
2 A. matses | 0.08 | ||||||||
3 A. sp. aff. minuta | 0.06 | 0.07 | |||||||
4 A. minuta | 0.05 | 0.08 | 0.03 | ||||||
5 A. javierbustamantei sp. n. | 0.09 | 0.03 | 0.06 | 0.07 | |||||
6 A. vote | 0.12 | 0.12 | 0.12 | 0.12 | 0.13 | ||||
7 A. bokermanni | 0.12 | 0.12 | 0.11 | 0.11 | 0.13 | 0.10 | |||
8 A. manaos | 0.12 | 0.12 | 0.12 | 0.14 | 0.12 | 0.10 | 0.08 | ||
9 A. sp. aff. manaos | 0.12 | 0.11 | 0.12 | 0.13 | 0.12 | 0.10 | 0.07 | 0.04 | |
10 D. leucomystax | 0.19 | 0.21 | 0.17 | 0.18 | 0.20 | 0.22 | 0.18 | 0.20 | 0.20 |
Species level diagnostic characters observed in the 16S rDNA gene of Amazophrynella javierbustamantei sp. n. and other species of genus Amazophrynella. First line indicates position of the character within the 16S rDNA gene; (-) indicates a deletion.
Species | 213 | 232 | 271 | 276 | 470 | 471 | 473 | 474 | 476 | 477 | 478 | 479 | 480 |
---|---|---|---|---|---|---|---|---|---|---|---|---|---|
A. manaos | A | C | A | C | A | T | G | T | C | A | A | A | A |
A. vote | A | T | A | C | C | C | C | T | T | A | A | A | G |
A. minuta | C | T | A | A | C | C | C | T | T | A | A | A | G |
A. bokermanni | A | T | A | C | A | T | G | T | C | A | A | A | A |
A. amazonicola | C | C | A | C | C | C | C | T | T | A | A | T | G |
A. javierbustamantei sp. n. | T | G | G | T | T | G | T | G | A | G | C | C | - |
A. matses | C | T | A | C | C | C | C | T | T | A | A | T | T |
Character loadings, eigenvalues, and percentage of explained variance for Principal Components (PC) 1–2. The analysis was based on eleven morphometric variables of adult males: Amazophrynella minuta complex (A. minuta sensu stricto; A. amazonicola; A. matses and A. javierbustamantei sp. n.).
Variables | PC1 | PC2 |
---|---|---|
HW | 0.462 | -0.146 |
HL | 0.455 | -0.104 |
SL | 0.374 | -0.244 |
ED | 0.261 | 0.052 |
IND | 0.369 | -0.271 |
UAL | 0.139 | 0.258 |
HAL | -0.032 | 0.484 |
THL | 0.311 | -0.295 |
TAL | 0.314 | 0.350 |
TL | 0.116 | 0.364 |
FL | 0.063 | 0.433 |
% of variation | 24.93 | 23.63 |
% | 24.93 | 48.56 |
All the species of the group are significantly different in shape (MANOVA, F24.3, Pillae´s trace < 0.001). The discriminate function analysis (DFA) found specimens correctly classified in 56.6% of cases and a moderate prior probabilities of groups (A. minuta = 28.75%, A. amazonicola = 18.75%, A. matses = 16.25% and A. javierbustamantei sp. n. = 36.25%). The variables that contributed most to the classification were HAL, SVL and TAL (Table
Character loadings of explained variance for Discriminant Function Analysis (DFA). The analysis was based on twelve morphometric variables of adult males of the Amazophrynella minuta complex (A. minuta sensu stricto; A. amazonicola; A. matses and A. javierbustamantei sp. n.).
Variables | Discriminant Function |
---|---|
SVL | 6.343 |
HW | -7.628 |
HL | 0.146 |
SL | -5.479 |
ED | -1.175 |
IND | -6.015 |
UAL | 1.313 |
HAL | 5.744 |
THL | -3.871 |
TAL | 13.944 |
TL | -1.250 |
FL | 1.016 |
(Fig.
(Fig.
The new species is part of Amazophrynella based on molecular phylogenetic relationships (Fig.
Amazophrynella javierbustamantei sp. n. is characterized by: (1) skin on dorsum tuberculate, with many subconical tubercles disperse on arms, legs, head and body; ventral skin coarsely areolate, throat and chest aerolate; (2) tympanic membrane and tympanic annulus not apparent through the skin; (3) snout long, subacuminated, protruding in lateral views; (4) upper eyelid with smaller tubercles, cranial crests absent; (5) dentigerous process of vomers absent; (6) vocal sac, vocal slits and nuptial pads absent; (7) finger I shorter than finger II, tips of digits rounded; (8) fingers lacking lateral fringes; (9) ulnar tubercles present; (10) heel bearing eight or more small low tubercles, tarsus with small tubercles and lack of folds; (11) plantar surfaces of feet bearing one metatarsal tubercle, the inner 2.5x larger than the outer, outer subconical; supernumerary plantar tubercles round and low; (12) toes lacking lateral fringes; webbing basal; toe III equal than toe V, tips of digits rounded; (13) dorsally is dark brown to light brown, and gray to black in some, ventrally, cream with yellow to orange marks, with black to dark brown spots; (14) SVL 16.39–22.25 mm in females, 12.79–16.42mm in males; (15) hand length is the greatest of all species of Amazophrynella: 3.6 mm in males (n= 26) and 4.6 mm in females (n=20), see Fig.
Amazophrynella javierbustamantei sp. n. (Figs
Body slender, head triangular, slightly longer than wide; head length 35.5% of SVL, head width 30.9% of SVL. Snout long, subacuminate in dorsal view, protruding in lateral view; canthus rostralis straight and loreal region vertical; without papilla; snout length 39.0% of head length; tympanic membrane and tympanic annulus not apparent through the skin, skin of the tympanic area covered by round sub-conical warts; vocal sac externally not visible, vocal slits absent; eyes prominent 23.8% of head length; upper eyelid covered with small tubercles; those close to the external margin aligned in a more or less distinct row; nostril closer to snout than to eyes; internarial distance smaller than eye diameter; presence of a line of small spiny granules from the outer edge of the mouth to upper arm, choanas small and circular.
Dorsal skin finely tuberculate with several large tubercles scattered sub-conical tubercles on upper arm; texture of ventral skin granular, covered by rounded granules. Dorsolateral surfaces, granular, with presence of large rounded tubercles. Forelimbs slender, upper arm length 29.6% of SVL; edges of lower arm and upper arm finely tuberculate with several large sub-conical and spiny granules; hand length 76.5% of upper arm length; fingers slender, tips not expanded; relative length of fingers I<II<IV<III; supernumerary tubercles and accessory palmar tubercles present, palmar large and rounded, supernumerary tubercles low, small rounded; subarticular tubercles rounded and small, one tubercle on fingers I, II and IV and two on finger III; fingers I and II basally webbed; indistinct nuptial pad. Hind limbs slender; ventral skin from thigh to tarsus covered by spiny tubercles, foot length 66% of thigh length; relative length of toes I<II<V<III<IV: inner metatarsal tubercle oval, 2.5× larger than outer; outer metatarsal tubercles small, rounded; subarticular tubercles present, rounded, present one on fingers I, II, and two on fingers III, V and three on finger IV; and tip of toes not expanded.
(in millimeters). SVL 15.1; HW 4.6; HL 5.3; SL 2.1; ED 1.2; IND 1.0; UAL 4.4; HAL 3.4; THL 8.1; TAL 8.1; TL 4.5; FL 5.3.
In life: dorsum of the holotype mostly light brown with dark brown in the dorsum; dorsolaterally creamish-brown with scattered black blotches; dorsal surfaces of hands and feet creamish-brown, and gray on arms and legs; belly creamish-gray with black dots, and the throat gray; fingers, toes and plantar surfaces reddish-black; groin with orange marks; iris with a bronze ring; cloaca with orange flap, black pupil and bronze iris. In alcohol: dorsum brownish-grey; venter cream with black and brown dots; orange surfaces turned cream, with a white longitudinal stripe on upper jaw extending from nostril to forearm.
The new species is phenotypically variable. In some individuals (e.g. MHNC 8245 and MHNC 11002, see Fig.
The following values are presented as: min-max (average ± SD, number of notes). The call is a trill type call issued during continuous and regular intervals (Fig.
Advisement call of Amazophrynella javierbustamantei sp. n. from the Tambopata National Reserve, Madre de Dios, Peru (207 meters a.s.l.) (Macauly Library of Natural Songs and Cornell Laboratory of Ornithology) by the authors Crocoft, Morales and Mc Diarmid (2007). A Oscilogram and spectrogram by one note B Oscilogram and spectrogram of notes from the advisement Call.
Amazophrynella javierbustamantei sp. n. is known from the Department of Cusco, in the lower Urubamba river basin and Department of Madre de Dios (Inambari, Candamo and Nueva Arequipa) in Peru (Fig.
Distribution map of Amazophrynella javierbustamantei sp. n. in Peru. Holotype locality in square orange, 5 Guacamayo Creek, Department Madre de Dios. Paratypes localities in white circles 1 Tsoroja, Department Junin 2 Mapi, Department Cusco 3 Camana, Department Cusco 4 Inambari, Department Madre de Dios 6 Los Amigos Biological Station, Department Madre de Dios 7 Explorer’s Inn, Department Madre de Dios.
The species is named after Dr. Javier Bustamante, a Peruvian residing in United States, to whom we dedicate this species in recognition of his friendship and support of herpetological taxonomy and systematics research and amphibian conservation in Peru.
Taxonomic reviews of Amazonian amphibians suggests that morphological characters are too conservative to permit delimiting species since closely related species share similar morphologies, and amphibians in general are morphologically conservative (e.g.,
The taxonomic ambiguity surrounding the name A. minuta and to a lesser extent A. bokermanni resulted in a severe underestimation of the taxonomic diversity of this genus. Since the descriptions of A. minuta in 1941 and A. bokermanni in 1993, the taxonomy of the genus has not been revised, leading to misdiagnoses of other species as either A. minuta or A. bokermanni due to the relatively generalized descriptions of these taxa. Three publications since 2012 (
The descriptions by
While part of our evidence for the existence of the new species as well as those described previously by
The new species A. javierbustamantei sp. n. is clearly differentiated in multivariate morphometric space from the other members of the Amazophrynella minuta “species group” (A. minuta, A. amazonicola and A. matses). Together with the description of Amazophrynella javierbustamantei sp. n. we also provide advertisement call. Amazophrynella javierbustamantei sp. n. is only the second species of the genus for which an advertisement call is known and recorded (see
The threats to the biological conservation of A. javierbustamantei sp. n. are evident, with uncontrolled exploration for gold, illegal mining and the destruction of habitat in the Departments of Madre de Dios and Cusco, probably causing a significant reduction in the population sizes of the species and fragmenting its distribution. For these reasons is necessary to analyze the current population status and trends of this and another amphibian species in this Department of southern Peru.
RRRZ was supported by a fellowship from Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES), RRRZ also thanks Nora M. Rojas Ruiz, Silvia Rojas, Marco Rojas Ruiz for hospitality in Lima, Elda Zamora, José Armas, Micaela Perea, Ronald Rojas and Miguel Rojas in Iquitos and Edson Caceres, Fababa and Douglas Giardini in Puerto Maldonado, Peru. For allowing access to the respective herpetological collections under their care, we are grateful to Richard C. Vogt (Instituto Nacional de Pesquisas da Amazonônia -
Amazophrynella minuta—BRAZIL: Taracuá, Uaupés River:
Amazophrynella bokermanni—BRAZIL: Juriti, Pará:
Amazophrynella vote—BRAZIL: Fazenda São Nicolau, Cotriguaçu, Mato Grosso (Holotype:
Amazophrynella manaos—BRAZIL: Campus da Universidade Federal do Amazonas, Amazonas (Holotype:
Amazophrynella amazonicola—PERU: Puerto Almendra San Juan Bautista, Loreto (Holotype: MZUNAP 901, paratopotypes: MZUNAP 906; MZUNAP 915; MZUNAP110; MZUNAP 907, MZUNAP 917; MZUNAP 889; MZUNAP 910; MZUNAP 911; MZUNAP 916; MZUNAP 913; MZUNAP 914; paratypes: MZUNAP 906; MZUNAP 915; MZUNAP 110; MZUNAP 907, MZUNAP 917; MZUNAP 889; MZUNAP 910; MZUNAP 911; MZUNAP 916; MZUNAP 913; MZUNAP 914); 58 km of Iquitos–Nauta highway on Fundo Zamora, San Juan Bautista, Loreto (Paratypes: MZUNAP 908, MZUNAP 924, MZUNAP 886, MZUNAP 900, MZUNAP 888, MZUNAP 919, MZUNAP 902, MZUNAP 887, MZUNAP 905, MZUNAP 920); Nauta, Maynas (Paratypes: MZUNAP 918, MZUNAP 909); Fundo UNAP, Maynas, Loreto (Paratype: MZUNAP 242)
Amazophrynella matses—PERU: Nuevo Salvador, Requena, Loreto (Holotype: MZUNAP 921, paratopotypes: MZUNAP 934, MZUNAP 955 MZUNAP 940, MZUNAP 948 MZUNAP 943, MZUNAP 952, MZUNAP 953, MZUNAP 958, MZUNAP 922, MZUNAP 923, MZUNAP 925, MZUNAP 926, MZUNAP 927, MZUNAP 944, MZUNAP 938, MZUNAP 936); Jenaro Herrera, Requena, Loreto (Paratypes: MZUNAP 928, MZUNAP 929, MZUNAP 930, MZUNAP 931, MZUNAP 933, MZUNAP 955, MZUNAP 935, MZUNAP 950, MZUNAP 937, MZUNAP 939, MZUNAP 941, MZUNAP 942, MZUNAP 946, MZUNAP 947, MZUNAP 949).
Amazophrynella javierbustamantei sp. n. —PERU: Quebrada Guacamayo, Tambopata, Madre de Dios (Holotype: MHNC 8331; MHNC 8363, MHNC 8245, MHNC 8238, MHNC 8316, MHNC 8484, MHNC 8362); La Pampa, Tambopata, Madre de Dios (MHNC 11101, MHNC 11102, MHNC 11103, MHNC 11104); Inambari, Manu, Madre de Dios (MHNSM 17993); Nuevo Arequipa, Tambopata, Madre de Dios (MHNC 8363, MHNC 8245, MHNC 8331, MHNC 8238, MHNC 8354, MHNC 8316, MHNC 8484, MHNC 8362); Rio Tambopata, Tambopata, Madre de Dios (MHNSM 9635, MHNSM 9641, MHNSM 9647, MHNSM 9642, MHNSM 9648, MHNSM 9633, MHNSM 9644, MHNSM 9646, MHNSM 9657, MHNSM 9640); Camana, La Convencion, Madre de Dios (MHNSM 25651); Mapi, La Convencion, Madre de Dios (MHNC 9939, MHNC 9940); Tambo Poyeni, Junin (MHNC 9387); Tsoroja, Junin (MHNC 9754, MHNC 9756, MHNC 9626, MHNC 9679, MHNC 9680, MHNC 9757). Rio Urubamba, Urubamba, Cusco (MHNC 9939, MHNC 9626, MHNC 9686, MHNC 9679, MHNC 9940, MHNC 9757, MHNC 9387, MHNC 9754, MHNC 9756).