Abstract
We describe two new species of poison frog from central and southern Peru that have been referred to as Ameerega picta, A. hahneli, or A. altamazonica throughout the past thirty years. Our phylogenies generated with genomic data provide strong support that the two new species are successive sisters to two described taxa, A. rubriventris and A. altamazonica, and collectively comprise the Ameerega rubriventris complex. The first new taxon, Ameerega panguana sp. nov., can be distinguished from all other Ameerega by its combination of a unique white venter and an advertisement call of 1–2 notes per second. The second new taxon, Ameerega imasmari sp. nov., is the only cryptically colored Ameerega species that is disttributed across the Fitzcarrald Arch in Southern Peru which possesses a ‘peep’ advertisement call consisting of 3–4 notes per second and a dominant frequency of 4.3–4.5 kHz. Within the Ameerega rubriventris complex, we observed differences between species in their ventral coloration, tympanum diameter, and call, which suggest that these taxa are reproductively isolated from each other.
References
Audacity Team (2017) Audacity(R): Free Audio Editor and Recorder. Computer Application. Version 2.1.3 Retrieved 4 October 2017. Available from: https://audacityteam.org/ (accessed 20 August 2019)
Bioacoustics Research Program (2014) Raven Pro: Interactive Sound Analysis Software. Version 1.5. Computer software. The Cornell Lab of Ornithology. Ithaca, New York. Available from: http://www.birds.cornell.edu/raven (accessed 20 August 2019)
Bolger, A.M., Lohse, M. & Usadel, B. (2014) Trimmomatic: a flexible trimmer for Illumina sequence data. Bioinformatics, 30, 2114–2120.
https://doi.org/10.1093/bioinformatics/btu170
Boulenger, G.A. (1884 “1883”) On a collection of frogs from Yurimaguas, Huallaga River, Northern Peru. Proceedings of the Zoological Society of London, 1883, 635–638, 631 colour pls., 631 black and white pls.
https://doi.org/10.1111/j.1469-7998.1883.tb06669.x
Brown, J.L., Schulte, R. & Summers, K. (2006) A new species of Dendrobates (Anura : Dendrobatidae) from the Amazonian lowlands in Peru. Zootaxa, 1152 (1), 45–58.
https://doi.org/10.11646/zootaxa.1152.1.2
Brown, J.L. & Twomey, E. (2009) Complicated histories: three new species of poison frogs of the genus Ameerega (Anura: Dendrobatidae) from north-central Peru. Zootaxa, 2049 (1), 1–38.
https://doi.org/10.11646/zootaxa.2049.1.1
Bushnell, B. (2014) BBTools software package. Available from: http://sourceforge. net/projects/bbmap (accessed 20 August 2019)
Crusoe, M.R., Alameldin, H.F., Awad, S., Boucher, E., Caldwell, A., Cartwright, R., Charbonneau, A., Constantinides, B., Edvenson, G., Fay, S.O, Fenton, J., Fenzl, T., Fish, J., Garcia-Gutierrez, L., Garland, P., Gluck, J., González, I., Guermond, S., Guo, J., Gupta, A., Herr, J.R., Howe, A., Hyer, A.O, Härpfer, A., Irber, L., Kidd, R., Lin, D., Lippi, J., Mansour, T., McA’Nulty, P., McDonald, E., Mizzi, J., Murray, K.D., Nahum, J.R., Nanlohy, K.O., Nederbragt, A.J., Ortiz-Zuazaga, H., Ory, J., Pell, J., Pepe-Ranney, C., Russ, Z.N., Schwarz, E., Scott, C., Seaman, J., Sievert, S., Simpson, J., Skennerton, C.T.O., Spencer, J., Srinivasan, R., Standage, D., Stapleton, J.A., Steinman, S.R., Stein, J., Taylor, B., Trimble, W., Wiencko, H.L., Wright, M., Wyss, B., Zhang, Q., Zyme, E. & Brown, C.T. (2015) The khmer software package: enabling efficient nucleotide sequence analysis. F1000Research, 4 (900), 1–12.
https://doi.org/10.12688/f1000research.6924.1
De la Riva, I., Márquez, R. & Bosch, J. (1996) The advertisement calls of three South American poison frogs (Amphibia:Anura: Dendrobatidae), with comments on their taxonomy and distribution. Journal of Natural History, 30, 1413–1420.
https://doi.org/10.1080/00222939600771311
Edgar, R.C. (2004) MUSCLE: multiple sequence alignment with high accuracy and high throughput. Nucleic Acids Research, 32, 1792–1797.
https://doi.org/10.1093/nar/gkh340
Faircloth, B. (2013) Illumiprocessor: a trimmomatic wrapper for parallel adapter and quality trimming. Available from: http://dx. doi. org/10.6079/J9ILL (accessed 4 November 2016)
Faircloth, B.C., McCormack, J.E., Crawford, N.G., Harvey, M.G., Brumfield, R.T. & Glenn, T.C. (2012) Ultraconserved elements anchor thousands of genetic markers spanning multiple evolutionary timescales. Systematic Biology, 61, 717–726.
https://doi.org/10.1093/sysbio/sys004
French, C.M., Deutsch, M.S., Chávez, G., Almora, C.E. & Brown, J.L. (2019) Speciation with introgression: phylogeography and systematics of the Ameerega petersi group (Dendrobatidae). Molecular Phylogenetics and Evolution, 138, 31–42.
https://doi.org/10.1016/j.ympev.2019.05.021
Grabherr, M.G., Haas, B.J., Yassour, M., Levin, J.Z., Thompson, D.A., Amit, I., Adiconis, X., Fan, L., Raychowdhury, R., Zeng, Q., Chen, Z., Mauceli, E., Hacohen, N., Gnirke, A., Rhind, N., di Palma, F., Birren, B.W., Nusbaum, C., Lindblad-Toh, K., Friedman, N. & Regev, A. (2011) Full-length transcriptome assembly from RNA-Seq data without a reference genome. Nature Biotechnology, 29, 644.
https://doi.org/10.1038/nbt.1883
Grant, T., Frost, D.R., Caldwell, J.P., Gagliardo, R., Haddad, C.F.B., Kok, P.J.R., Means, D.B., Noonan, B.P., Schargel, W.E. & Wheeler, W.C. (2006) Phylogenetic systematics of dart-poison frogs and their relatives (Amphibia, Athesphatanura, Dendrobatidae). Bulletin of the American Museum of Natural History, 299, 1–262.
https://doi.org/10.1206/0003-0090(2006)299[1:PSODFA]2.0.CO;2
Guillory, W.X., French, C.M., Twomey, E.M., Chávez, C., Prates. I., von May, R., De la Riva, I., Lötters, S., Reichle, S., Serrano-Rojas, S.J., Whitworth, A. & Brown, J.L. (2020) Phylogenetic relationships and systematics of the Amazonian poison frog genus Ameerega using ultraconserved genomic elements. Molecular Phylogenetics and Evolution, 142, 106638.
https://doi.org/10.1016/j.ympev.2019.106638
Haddad, C.F.B. & Martins, M. (1994) Four species of Brazilian poison frogs related to Epipedobates pictus (Dendrobatidae): Taxonomy and natural history observations. Herpetologica, 50, 282–295.
Hahn, C., Bachmann, L. & Chevreux, B. (2013) Reconstructing mitochondrial genomes directly from genomic next-generation sequencing reads—a baiting and iterative mapping approach. Nucleic Acids Research, 41, e129–e129.
https://doi.org/10.1093/nar/gkt371
IUCN, S.S.C. (2001) IUCN red list categories and criteria: version 3.1. Prepared by the IUCN Species Survival Commission. IUCN, Gland, 30 pp.
Jacob Machado, D., Janies, D., Brouwer, C. & Grant, T. (2018) A new strategy to infer circularity applied to four new complete frog mitogenomes. Ecology and Evolution, 8, 4011–4018.
https://doi.org/10.1002/ece3.3918
Kalyaanamoorthy, S., Minh, B.Q., Wong, T.K., von Haeseler, A. & Jermiin, L.S. (2017) ModelFinder: fast model selection for accurate phylogenetic estimates. Nature Methods, 14, 587.
https://doi.org/10.1038/nmeth.4285
Katoh, K. & Standley, D.M. (2013) MAFFT multiple sequence alignment software version 7: improvements in performance and usability. Molecular Biology and Evolution, 30, 772–780.
https://doi.org/10.1093/molbev/mst010
Kim, D., Langmead, B. & Salzberg, S.L. (2015) HISAT: a fast spliced aligner with low memory requirements. Nature Methods, 12, 357.
https://doi.org/10.1038/nmeth.3317
Lötters, S., Debold, P., Henle, K., Glaw, F. & Kneller, M. (1997) Ein neuer Pfeilgiftfrosch aus der Epipedobates pictus-gruppe vom Osthang der Cordillera Azul in Per. Herpetofauna, 19, 25–34.
Lötters, S., Jungfer, K.H., Schmidt, W. & Henkel, F.W. (2007) Poison Frogs: Biology, Species and Captive Husbandry. Edition Chimaira, Frankfurt am Main, 668 pp.
Lötters, S., Schmitz, A. & Reichle, S. (2005) A new cryptic species of poison frog from the Bolivian Yungas. Herpetozoa, 18, 115–124.
Lötters, S., Schmitz, A., Reichle, S., Rödder, D. & Quennet, V. (2009) Another case of cryptic diversity in poison frogs (Dendrobatidae: Ameerega): description of a new species from Bolivia. Zootaxa, 2028 (1), 20–30.
https://doi.org/10.11646/zootaxa.2028.1.2
Minh, B.Q., Nguyen, M.A.T. & von Haeseler, A. (2013) Ultrafast approximation for phylogenetic bootstrap. Molecular Biology and Evolution, 30, 1188–1195.
https://doi.org/10.1093/molbev/mst024
Morales, V.R. (1992) Study of the vocalization of some arrow poison frogs in Peru. Acta Zoológica Lilloana, 41, 107–119.
Myers, C.W. (1982) Spotted poison frogs: Description of Three new Dendrobates from Western Amazonia, and resurrection of a lost species from “Chiriqui”. American Museum Novitates, 2721, 23.
Myers, C.W., Daly, J.W. & Malkin, B. (1978) A dangerously toxic new frog (Phyllobates) used by Ember indians of Western Colombia, with discussion of blowgun fabrication and dart poisoning. Bulletin of the American Museum of Natural History, 161, 307–366, 302 pls.
Neves, M.O., da Silva, L.A., Akieda, P.S., Cabrera, R., Koroiva, R. & Santana, D.J. (2017) A new species of poison frog, genus Ameerega (Anura: Dendrobatidae), from the southern Amazonian rain forest. Salamandra, 53, 485–493.
Nguyen, L.-T., Schmidt, H.A., von Haeseler, A. & Minh, B.Q. (2014) IQ-TREE: a fast and effective stochastic algorithm for estimating maximum-likelihood phylogenies. Molecular Biology and Evolution, 32, 268–274.
https://doi.org/10.1093/molbev/msu300
R Core Team (2019) R: A language and environment for statistical computing. R Foundation for Statistical Computing, Vienna, Austria. Available from: http://www.R-project.org/ (accessed 20 September 2019)
Raposo do Amaral, F., Neves, L.G., Resende Jr, M.F., Mobili, F., Miyaki, C.Y., Pellegrino, K.C.M. & Biondo, C. (2015) Ultraconserved elements sequencing as a low-cost source of complete mitochondrial genomes and microsatellite markers in non-model amniotes. PLoS One, 10, e0138446.
https://doi.org/10.1371/journal.pone.0138446
Roberts, J.L., Brown, J.L., von May, R., Arizabal, W., Schulte, R. & Summers, K. (2006) Genetic divergence and speciation in lowland and montane peruvian poison frogs. Molecular Phylogenetics and Evolution, 41, 149–164.
https://doi.org/10.1016/j.ympev.2006.05.005
Schlüter, A. (1980) Bio-akustische Untersuchungen an Dendrobatiden in einem begrenzten Gebiet des tropischen Regenwaldes von Peru (Amphibia: Salientia: Dendrobatidae). Salamandra, 16, 149–161.
Schlüter, A. (2005) Amphibien an einem Stillgewässer in Peru: mit einer illustrierten Checklist der Amphibien und Reptilien des unteren Río Llullapichis. Vol. 22. Edition Chimaira, Frankfurt am Main, 347 pp.
Schulte, R. (1999) Pfeilgiftfrösche Artenteil—Peru“. INBICO, Wailblingen, 292 pp.
Siu Ting, K. (2012) Evaluación de la diversidad críptica en anfibios de amplia distribución en la Amazonía peruana. Master’s Thesis, Universidad Nacional Mayor De San Marcos, Lima, 124 pp.
Serrano-Rojas, S.J., Whitworth, A., Villacampa, J., Von May, R., Gutierrez, R.C., Padial, J.M. & Chaparro, J.C. (2017) A new species of poison-dart frog (Anura: Dendrobatidae) from Manu province, Amazon region of southeastern Peru, with notes on its natural history, bioacoustics, phylogenetics, and recommended conservation status. Zootaxa, 4221 (1), 71–94.
https://doi.org/10.11646/zootaxa.4221.1.3
Silverstone, P.A. (1976) A revision of the poison-arrow frogs of the genus Phyllobates Bibron in Sagra (Family Dendrobatidae). Natural History Museum of Los Angeles County Science Bulletin, 1976, 1–53, 52 color pls.
Tamura, K., Stecher, G., Peterson, D., Filipski, A., & Kumar, S. (2013) MEGA6: Molecular Evolutionary Genetics Analysis version 6.0. Molecular Biology and Evolution, 30, 2725–2729.
https://doi.org/10.1093/molbev/mst197
Twomey, E. & Brown, J.L. (2008) A partial revision of the Ameerega hahneli complex (Anura: Dendrobatidae) and a new cryptic species from the East-Andean versant of Central Peru. Zootaxa, 1757 (1), 49–65.
https://doi.org/10.11646/zootaxa.1757.1.3
Vaz-Silva, W. & Maciel, N.M. (2011) A new cryptic species of Ameerega (Anura: Dendrobatidae) from Brazilian Cerrado. Zootaxa, 2826 (1), 57–68.
https://doi.org/10.11646/zootaxa.2826.1.3
Zhang, C., Rabiee, M., Sayyari, E. & Mirarab, S. (2018) ASTRAL-III: polynomial time species tree reconstruction from partially resolved gene trees. BMC bioinformatics, 19, 153.