Amphibians of the largest inland Atlantic Forest fragment, Iguaçu National Park, Paraná State, southern Brazil

109 Amphibians of the largest inland Atlantic Forest fragment, Iguaçu National Park, Paraná State, southern Brazil Emerson A. da Rosa1,2, Rodrigo Lingnau3, Michel V. Garey2 1 Programa de PósGraduação em Biodiversidade Neotropical – UNILA, Av. Tarquínio Joslin dos Santos, 1000, Polo Jardim Universitário, Foz do Iguaçu, PR, Brazil 2 Laboratório de Ecologia de Metacomunidades – LEMET, Av. Tarquínio Joslin dos Santos, 1000, Polo Jardim Universitário, Foz do Iguaçu, PR, Brazil 3 Universidade Tecnológica Federal do Paraná – UTFPR, Rua Gelindo João Folador, 2000, Bairro Novo Horizonte, Francisco Beltrão, PR, Brazil Corresponding author: Emerson A. da Rosa (emersonnhr[email protected]) Copyright: © Emerson A. da Rosa et al. This is an open access article distributed under terms of the Creative Commons Attribution License (Attribution 4.0 International – CC BY 4.0). Research Article Abstract The present study aimed to provide a species list of amphibians from Iguaçu National Park, the largest forest fragment of inland Atlantic Forest, located in the westernmost region of Paraná state, recognized as a UNESCO Natural Heritage Site. The park’s vegetation is predominantly composed of two types: Semideciduous Seasonal Forest (SSF) and Mixed Ombrophilous Forest (MOF). To compile the species list, primary data obtained through sampling adult anurans and tadpoles in 21 water bodies was used, complemented by search for historical records based on specimens deposited in scientific collections. 31 anuran species belonging to seven families were recorded, with the Hylidae being the richest, comprising 17 species. Two species, Boana curupi and Physalaemus carrizorum, represent the first records in the state of Paraná. The anuran species recorded in the Iguaçu National Park present nine reproductive modes, reflecting a diversity similar to that observed in other areas of Semideciduous Seasonal Forest. Twelve species were recorded exclusively in the SSF, eight species exclusively in MOF, and 11 species occurred in both vegetation types. The anuran assemblage of Iguaçu National Park is most similar to that Iguazú National Park in Argentina, with which it shares 24 species. Although the Iguaçu River is the only geographic barrier separating these two protected areas, the complementarity in amphibian species composition highlights the importance of conserving both parks for maintaining regional biodiversity. Key words: Anurans, inventory, Mixed Ombrophilous Forest, Semideciduous Seasonal Forest, species list Introduction Brazil harbors at least 1,188 amphibian species, including 1,144 anurans, 39 gymnophionans, and five caudatans (Segalla et al. 2021). The Atlantic Forest domain exhibits the highest amphibian species richness in the country (Campos et al. 2014; Rossa-Feres et al. 2017), with more than 625 anuran species recorded and more than 77% anuran species are endemic (Rossa-Feres et al. 2017). Given its exceptional species richness, combined with high endemism, and severe anthropogenic pressures, has led to the classification of the Atlantic Forest as one of the world’s biodiversity hotspots (Myers et al. 2000). Within the Atlantic Academic editor: Uri García-Vázquez Received: 10 July 2025 Accepted: 13 November 2025 Published: 15 December 2025 ZooBank: https://zoobank. org/9F9B81FE-2B92-4600-8E70BE8AEB76715F Citation: da Rosa EA, Lingnau R, Garey MV (2025) Amphibians of the largest inland Atlantic Forest fragment, Iguaçu National Park, Paraná State, southern Brazil. ZooKeys 1264: 109–128. https://doi.org/10.3897/ zookeys.1264.164796 ZooKeys 1264: 109–128 (2025) DOI: 10.3897/zookeys.1264.164796 110 ZooKeys 1264: 109–128 (2025), DOI: 10.3897/zookeys.1264.164796 Emerson A. da Rosa et al.: Amphibians of the largest inland Atlantic Forest fragment, southern Brazil Forest, areas of Dense Ombrophilous Forest contain the highest richness of anuran species (Rossa-Feres et al. 2017) and still remain as extensive forest remnants with relatively high connectivity (Rosa et al. 2021). In contrast, more degraded regions of the Atlantic Forest domain are dominated by phytophysiognomies that support lower species richness of anurans, such as the Semideciduous Seasonal Forest (SSF) and Mixed Ombrophilous Forest (MOF) (Rossa-Feres et al. 2017). In the western region of Paraná, originally covered by SSF with some patches of MOF, recent trends indicate increasing forest cover and reduced habitat isolation (Rosa et al. 2021). Despite these reforestation efforts, only 9% of the region remain forested (Instituto Água e Terra 2021), and most of these forest fragments are small and highly isolated (Rosa et al. 2021). In the state of Paraná, surveys of anuran assemblages have been conducted primarily in the coastal region and the metropolitan area of Curitiba, where Dense Ombrophilous Forest is the dominant vegetation type (e.g., Conte and Rossa-Feres 2006; Armstrong and Conte 2010; Garey and Hartmann 2012), as well as the central-northern region, characterized by SSF and MOF (e.g., Conte and Machado 2005; Conte and Rossa-Feres 2007; Affonso and Delariva 2012; Nazaretti and Conte 2015). In contrast, the anuran assemblages in the MOF and SSF of western, southern, and southwestern regions of Paraná state remain understudied (Santos-Pereira et al. 2018). In the western region of Paraná state, only few remnants have been inventoried, including areas near Rio Guarani State Park in the municipality of Três Barras do Paraná (Bernarde and Machado 2001), the Bela Vista Biological Refuge in Foz do Iguaçu (Vicente-Ferreira et al. 2024), and some forest remnants and anthropogenic landscapes (farmlands and urban areas) in western of Paraná state (Leivas et al. 2018). Notably, Iguaçu National Park (INP) the largest forest fragment of inland Atlantic Forest, still lacks a systematic inventory of its amphibian assemblage (Santos-Pereira et al. 2018). The INP is considered the largest and most important fragment of the Atlantic Forest in southern Brazil. It is the largest protected area in Paraná state and the second largest in the Atlantic Forest domain (Paviolo et al. 2016). The INP has been recognized as a UNESCO World Natural Heritage Site since 1986 (D’Oliveira et al. 2002). Part of INP is located in a transboundary region, sharing more than 60 kilometers of border with Iguazú National Park in Argentina, thus forming part of one of the most significant biological corridors in the south-central region of South America (Instituto Chico Mendes de Conservação da Biodiversidade 2018). Despite its ecological importance, knowledge of the INP anurans remains largely anecdotal in the scientific literature. Although some undergraduate theses and dissertations have addressed amphibians in INP (e.g., Jeziorny 2015; Nazaretti 2016), peer-reviewed articles include only sparse records: five species reported by Lutz (1973), two species by Heyer (1979, 1994), one species by Conte et al. (2009), and four species recorded by Leivas et al. (2018). This severe data deficiency is reflected in the INP Management Plan, which currently recognizes just 12 amphibian species for the entire park (Instituto Chico Mendes de Conservação da Biodiversidade 2018). Given the lack of systematic inventories in the largest remaining forest fragment of the Atlantic Forest, and considering that such inventories are essential for providing baseline data to support the development of effective conservation and management strategies, this study aims to: (i) present a 111 ZooKeys 1264: 109–128 (2025), DOI: 10.3897/zookeys.1264.164796 Emerson A. da Rosa et al.: Amphibians of the largest inland Atlantic Forest fragment, southern Brazil comprehensive list of amphibian species occurring in Iguaçu National Park; (ii) provide information on species’ natural history and habitat use; and (iii) compare the amphibian species composition in INP with assemblages from other regions that share the same vegetation types present in the park. Materials and methods Study area We conducted amphibians survey in the Iguaçu National Park, located in the western region of Paraná state, southern Brazil (Fig. 1). The INP is considered the main fragment of the Atlantic Forest in southern Brazil (Paviolo et al. 2016) and comprises two main vegetation types: Semideciduous Seasonal Forest (SSF) and Mixed Ombrophilous Forest (MOF) (Ribeiro et al. 2009). The vegetation type variation follows the altitudinal gradient across the park, which ranges from 100 to 750 meters above sea level, with MOF occurring at higher elevations and SSF at lower elevations (Cervi and Borgo 2007). The INP covers 185,262.5 hectares (Instituto Chico Mendes de Conservação da Biodiversidade 2018) and includes portions of six municipalities within its boundaries: Foz do Iguaçu, São Miguel do Iguaçu, Matelândia, Serranópolis do Iguaçu, Céu Azul, and Capanema. The surrounding region includes eight municipalities: Santa Terezinha de Itaipu, Santa Tereza do Oeste, Santa Lúcia, Lindoeste, Capitão Leônidas Marques, Medianeira, Vera Cruz do Oeste, and Ramilândia (Instituto Chico Mendes de Conservação da Biodiversidade 2018). The surrounding buffer area is occupied by intensive livestock farming and mechanized agriculture (Instituto Chico Mendes de Conservação da Biodiversidade 2018). The climate in the INP is humid subtropical mesothermal (Cfa, under Köppen-Geiger classification) (Alvares et al. 2013). Mean annual temperature ranges between 20 °C and 22 °C, with January being the warmest month (mean 26.3 °C) and July the coldest (mean 16.6 °C). Annual precipitation ranging from 1600 to 1900 mm, distributed throughout the year without a distinct dry season. Monthly rainfall peaks in October, with an average of 186 mm, and reaches its minimum in July, with only 92 mm (Alvares et al. 2013). Sample design To compile the amphibian species list for Iguaçu National Park, we combined data from field survey, scientific collections, and the literature. We only considered literature records when voucher specimens were deposited in a scientific collection. We obtained primary data through sampling lentic and lotic water bodies within the INP in the municipalities of Foz do Iguaçu and São Miguel do Iguaçu. We conducted three sampling campaigns between November 2023 and March 2024, maintaining a minimum interval of 30 days between them to maximize the likelihood of detecting species with different reproductive periods (e.g., Bolzan et al. 2019; Dubos et al. 2019). We sampled amphibians, adult anurans and tadpoles, in 21 water bodies (18 lentic and three lotic). In each campaign, we surveyed each water body twice: once during the day for tadpole collection and once at night for adult anurans, with both diurnal and nocturnal sampling sessions lasting one hour. 112 ZooKeys 1264: 109–128 (2025), DOI: 10.3897/zookeys.1264.164796 Emerson A. da Rosa et al.: Amphibians of the largest inland Atlantic Forest fragment, southern Brazil We captured tadpoles using dip nets with a mesh size of 3 mm (adjusted in diameter according to water body features). We euthanized collected tadpoles by immersion in a lidocaine solution and preserved them in a 1:1 solution of 10% formalin and 70% ethanol for later identification under a stereomicroscope. We sampled adult anurans at night using active search methods at breeding sites (Scott et al. 1994). We euthanized some adult individuals through topical lidocaine application, fixed them in 10% formalin, and then preserved in 70% ethanol. We deposited all collected individuals (tadpoles and adults) in Bertha Lutz Herpetological Collection at Unila (CA-Unila) (Suppl. material 1). We obtained secondary data on amphibian occurrences in the INP from scientific collections and literature. We visited the scientific collections to verify and confirm specimen identifications at the Museu de História Natural Capão da Imbuia in Curitiba (MNHCI-Herpeto), Paraná, and the Bertha Lutz Herpetological Collection at the Universidade Federal da Integração Latino-Americana) in Foz do Iguaçu (CA-Unila), Paraná. Additionally, we conducted a systematic literature review using the search terms [“Amphibia” OR “Anura” OR “Gymnophiona”] AND [“Parque Nacional do Iguaçu” OR “Iguaçu National Park”] across Scopus, Web of Science, and Google Scholar platforms. We followed the taxonomic nomenclature proposed by the Amphibian Species of the World (Frost 2024). Figure 1. Geographic location of the study area: Iguaçu National Park, located in the extreme western region of the state of Paraná, Brazil. The centroids (larger circles) represent the average distances of the sampling points. Circle sizes vary according to anuran amphibian species richness and the number of sampling points within each phytophysiognomy: Semideciduous Seasonal Forest (SSF) and Mixed Ombrophilous Forest (MOF). 113 ZooKeys 1264: 109–128 (2025), DOI: 10.3897/zookeys.1264.164796 Emerson A. da Rosa et al.: Amphibians of the largest inland Atlantic Forest fragment, southern Brazil For each species recorded in the INP, we compiled data on: (i) location of occurrence, (ii) phytophysiognomy, (iii) population status, (iv) reproductive mode, (v) habit, (vi) semaphoront, and (vii) habitat type. Location refers to the municipality where the individual was found. We classified phytophysiognomy as either SSF or MOF. We determined population status (increasing, decreasing, or stable) according to the IUCN assessments (International Union for Conservation of Nature and Natural Resources 2024). We categorized reproductive modes based on field observations and literature sources (i.e., Toledo et al. 2021), following the classification proposed by Haddad and Prado (2005). We defined the habit as the environment in which adult individuals typically occur: arboreal, rheophilic, terrestrial, cryptozoic, or fossorial (Toledo et al. 2021). The semaphoront refers to the life stage of the individuals observed, classified as either adult or tadpole. We classified habitat type based on the water body where individuals (tadpoles or adults) were recorded, classified as lotic (e.g., streams) or lentic (e.g., temporary and permanent ponds, and marsh), following field observations and literature sources (e.g., Toledo et al. 2021). Data analysis We compared the anuran assemblage of Iguaçu National Park with 18 published inventories from Seasonal Forests (Deciduous or Semideciduous) and MOF using similarity index. We selected inventories based on two criteria: (i) the presence of similar phytophysiognomies and (ii) geographic distance less than 1000 km from the study area. The following inventories were included in the analysis: Refúgio Biológico Bela Vista (Vicente-Ferreira et al. 2024), Parque Estadual Rio Guarani (Bernarde and Machado 2001), Mata dos Godoy (Machado et al. 1999), São José dos Pinhais (Conte and Rossa-Feres 2006), Floresta Nacional de Piraí do Sul (Foerster and Conte 2018), Parque Municipal São Luís de Tolosa (Santos and Conte 2014), Fazenda Rio Grande (Conte and Rossa-Feres 2007), and Área de Proteção Ambiental da Serra da Esperança (Instituto Água e Terra 2009) in Paraná state; Estação Ecológica dos Caetetus (Brassaloti et al. 2010), Parque Estadual do Morro do Diabo (Santos et al. 2009), Floresta Estadual Edmundo Navarro de Andrade (Toledo et al. 2003) in São Paulo state; Parque Nacional da Serra da Bodoquena in Mato Grosso do Sul state (Uetanabaro et al. 2007); Parque Estadual Fritz Plaumann (Bastiani and Lucas 2013), Parque Nacional das Araucárias (Lucas and Marocco 2011), Floresta Nacional de Chapecó (Lucas and Fortes 2008) in Santa Catarina state; Parque Municipal de Sertão (Zanella et al. 2013), Parque Estadual do Papagaio Charão (Potrich et al. 2020) in Rio Grande do Sul state; Mbaracayú, Alto Paraná Department, in Paraguay (Cacciali et al. 2015); and Parque Nacional Iguazú, Misiones Province in Argentina (López and Garey 2021). Due to methodological variations among inventories, we restricted our analysis to presence-absence data. We calculated community similarity using Jaccard index and performed a non-Metric Multidimensional Scaling (nMDS) to graphically evaluate the variation in the species composition among anuran inventories. All analyses were performed in the R software (R Core Team 2024) using the package ‘vegan’ (Oksanen et al. 2022). 114 ZooKeys 1264: 109–128 (2025), DOI: 10.3897/zookeys.1264.164796 Emerson A. da Rosa et al.: Amphibians of the largest inland Atlantic Forest fragment, southern Brazil Results We recorded 31 anuran amphibian species in the Iguaçu National Park, including 19 species recorded during field sampling and 12 species from scientific collections. All recorded species belong to the Anura and are distributed across seven families: Hylidae (17 species, 54.8% of total), Leptodactylidae (n = 8, 25.8%), Bufonidae (n = 2, 6.5%), and four families with one species each (Alsodidae, Hylodidae, Microhylidae, and Odontophrynidae; 3.2% each) (Figs 2, 3). Among the municipalities within INP, Foz do Iguaçu had the highest species richness, with 22 species (71%) recorded. Céu Azul had 16 species (52%), followed by São Miguel do Iguaçu with ten species (32%). Only two species (6%) were recorded in Serranópolis do Iguaçu. Although Matelândia and Capanema are also part of the INP, we did not found records of amphibian collections from these areas. We recorded 11 species (39%) exclusively in the Semideciduous Seasonal Forest (SSF), eight species (26%) exclusively in the Mixed Ombrophilous Forest (MOF), and 11 species (36%) in both vegetation types within INP (Fig. 4). The anuran amphibians recorded in the INP exhibit nine reproductive modes. The family Hylidae exhibited the highest richness of reproductive modes (six reproductive modes), followed by Leptodactylidae with three. All other families exhibited a single reproductive mode. All species of Alsodidae, Bufonidae, and Microhylidae recorded in the INP reproduce by lay eggs in still water, where tadpoles develop (mode 1). The only representative of Odontophrynidae lay eggs in flowing water, which hatch into exotrophic tadpoles (mode 2). The single Hylodidae species reproduced exhibits mode 3, in which eggs and larvae develop in submerged chambers, and exotrophic tadpoles inhabit streams. Species of Leptodactylidae exhibit three reproductive modes: floating foam nests in ponds, with exotrophic tadpoles developing in ponds (mode 11); foam nests in constructed basins, with tadpole development in ponds (mode 13); and foam nests in underground cavities, where flooding facilitated tadpole migration to ponds (mode 30). Hylidae species exhibited six reproductive modes: eggs deposited in natural or constructed basins that, upon flooding released exotrophic tadpoles into ponds or streams (mode 4); eggs deposited in constructed nests that, after flooding, allowed exotrophic tadpoles to develop in ponds and streams (mode 5); floating foam nests in ponds with tadpoles remaining in the same environment (mode 11); eggs hatching into exotrophic tadpoles in lentic water bodies (mode 24). Additionally, Hylidae included modes 1 and 2, as previously described. Among the amphibians recorded in the INP, 17 species (55%) are arboreal, nine (29%) are terrestrial, two (6%) are rheophilic, two (6%) are cryptozoic, and one species (3%) is fossorial. All species in the family Hylidae are arboreal. Most Leptodactylidae species are terrestrial, with only one exception (Leptodactylus podicipinus) classified as cryptozoic. Both Bufonidae species are terrestrial. The species from the families Alsodidae and Hylodidae are rheophilic, while those from the Microhylidae and Odontophrynidae are fossorial and cryptozoic, respectively (Fig. 5). In terms of habitat type, 14 species (45%) occurred exclusively in lentic environments, six (19%) were restricted to lotic environments, and ten (33%) inhabited both habitat types. 115 ZooKeys 1264: 109–128 (2025), DOI: 10.3897/zookeys.1264.164796 Emerson A. da Rosa et al.: Amphibians of the largest inland Atlantic Forest fragment, southern Brazil Figure 2. Anuran amphibians recorded in the Iguaçu National Park, western Paraná, southern Brazil. A. Rhinella diptycha; B. Boana curupi; C. Trachycephalus typhonius; D. Elachistocleis bicolor; E. Boana faber; F. Proceratophrys avelinoi; G. Boana punctata; H. Aplastodiscus perviridis; I. Boana caingua; J. Rhinella ornata; K. Scinax squalirostris; L. Phyllomedusa tetraploidea; M. Leptodactylus mystacinus; N. Leptodactylus elenae; O. Boana raniceps. 116 ZooKeys 1264: 109–128 (2025), DOI: 10.3897/zookeys.1264.164796 Emerson A. da Rosa et al.: Amphibians of the largest inland Atlantic Forest fragment, southern Brazil Figure 3. Anuran amphibians recorded in the Iguaçu National Park, western Paraná, southern Brazil. A. Physalaemus cuvieri; B. Limnomedusa macroglossa; C. Scinax perereca; D. Leptodactylus luctator; E. Itapotihyla langsdorffii; F. Boana albopunctata; G. Leptodactylus podicipinus; H. Scinax fuscovarius; I. Dendropsophus nanus; J. Dendropsophus minutus; K. Leptodactylus fuscus; L. Crossodactylus schmidti. 117 ZooKeys 1264: 109–128 (2025), DOI: 10.3897/zookeys.1264.164796 Emerson A. da Rosa et al.: Amphibians of the largest inland Atlantic Forest fragment, southern Brazil The nMDS analysis (Fig. 6) revealed two distinct groups based on similarity in anuran assemblage composition: one composed by assemblages from SSF and the other from MOF. The anuran composition in the Iguaçu National Park showed greater similarity to that of the Iguaçu National Park in Misiones, Argentina, and the Refúgio Biológico Bela Vista in Foz do Iguaçu, western Paraná state, Brazil. The complete list of anuran species recorded, including their taxonomic authorities and relevant ecological information, is presented in Table 1. Discussion We recorded 31 anuran amphibian species in the Iguaçu National Park, representing approximately 20% of the species known for the state of Paraná (Santos-Pereira et al. 2018), 12% of those reported for the Semideciduous Seasonal Forest (SSF), and 4.8% of the species known from the Atlantic Forest Figure 4. Venn diagram showing the distribution of anuran amphibian species between the two phytophysiognomies of the Iguaçu National Park: Semideciduous Seasonal Forest (SSF) and Mixed Ombrophilous Forest (MOF). Figure 5. 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The Open Database License (ODbL) is a license agreement intended to allow users to freely share, modify, and use this Dataset while maintaining this same freedom for others, provided that the original source and author(s) are credited. Link: https://doi.org/10.3897/zookeys.1264.164796.suppl1