Introduction
Peru harbours one of the greatest levels of biological diversity in the world, particularly within its Amazonian region, which supports an exceptional variety of amphibian species, many of which remain poorly studied (Catenazzi and Von May Reference Catenazzi and von May2014; Von May et al. Reference Von May, Catenazzi, Angulo, Brown, Carrillo and Chávez2008). The country ranks fourth worldwide in amphibian species richness, with 688 species recorded, comprising 668 frogs and toads (Anura), 4 salamanders (Caudata), and 16 caecilians (Gymnophiona) (AmphibiaWeb, 2025). The highest species richness is found in the Peruvian Amazon, particularly in the lowland forests, followed by the Andean foothills and premontane zones, where local richness is lower but endemism and species turnover are high (Catenazzi et al., Reference Catenazzi, Lehr and von May2013). In contrast, the coastal zone exhibits the lowest amphibian diversity in the country (Catenazzi et al., Reference Catenazzi, Lehr and von May2013; Penhacek et al., Reference Penhacek, Souza, Santos, Guerra, Castro-Souza and Rodrigues2024). Despite this remarkable diversity, most research on Peruvian amphibians has primarily focused on taxonomic descriptions and distribution patterns (Herrera-Alva et al. Reference Herrera-Alva, Catenazzi and Aguilar-Puntriano2023; Penhacek et al. Reference Penhacek, Souza, Santos, Guerra, Castro-Souza and Rodrigues2024; Von May et al. Reference Von May, Diaz, Ttito, Santa-Cruz and Catenazzi2024), whereas parasitological studies remain scarce, despite their relevance for understanding host-parasite interactions and evaluating ecosystem health (Chero et al. Reference Chero, Cruces, Cacique, Ponce, Iannacone and Alvariño2023).
Nematodes represent the most diverse groups of amphibian parasites, particularly those belonging to the Cosmocercidae, such as the genus Aplectana, which is widely distributed among anurans across tropical regions. Currently, about 58 valid Aplectana species are recognized (Santos et al. Reference Santos, Borges, Willkens, Santos, Costa-Campos and Melo2023), with the Neotropical region exhibiting the greatest known richness, approximately 28 recorded species (Santos et al. Reference Santos, Willkens, Silva, Melo and Fernandes2023). Several of these have been described over the past three decades, mainly from anurans (Bursey et al. Reference Bursey, Goldberg and Parmelee2001; Castillo et al. Reference Castillo, González Rivas and Acosta2023; De Oliveira et al. Reference De Oliveira, de Sousa Machado, de Castro Araújo, De Sousa Silva and Ávila2024; Gómez et al. Reference Gómez, Sánchez, Ñacari and Espínola-Novelo2020; Villegas-Ojeda et al. Reference Villegas-Ojeda, Fernández-Marinone, Jofré and González2025), though representatives have also been reported in Gymnophiona (Ubelaker Reference Ubelaker1966) and Squamata (Amorim et al. Reference Amorim, Silva, Morais, Silva and Avila2017; Bursey et al. Reference Bursey, Goldberg and Telford2006; Da Silva et al. Reference Da Silva, Teixeira, Teles, Araujo Filho, Mesquita and Oliveira Almeida2018; Ramallo et al. Reference Ramallo, Goldberg, Ruiz and Stazzonelli2023; Vieira et al. Reference Vieira, Goncalves, Lima, Sousa and Muniz-Pereira2020).
In Peru, three species of Aplectana have been recorded (Bursey et al. Reference Bursey, Goldberg and Parmelee2001; Chero et al. Reference Chero, Cruces, Cacique, Ponce, Iannacone and Alvariño2023; Iannacone Reference Iannacone2003). The most widely distributed is Aplectana hylambatis (Baylis, Reference Baylis1927), recorded from the Andes to the Amazon, parasitizing the gastrointestinal tract of various amphibian hosts such as Leptodactylus bolivianus Boulenger, 1898 (Leptodactylidae) (Bursey et al. Reference Bursey, Goldberg and Parmelee2001), Rhinella marina (Linnaeus, 1758) and R. spinulosa (Wiegmann, 1834) (Bufonidae) (Chero et al. Reference Chero, Cruces, Iannacone, Sáez, Alvariño, Luque and Morales2016; Toledo et al. Reference Toledo, Fonseca, Cárdenas-Callirgos, Vidaurre and Silva2017), Telmatobius marmoratus (Duméril & Bibron, 1841) (Chero et al. 2014; Recharte Reference Recharte1995), Telmatobius jelskii (Peters, 1873) (Telmatobiidae) (Iannacone Reference Iannacone2003), and Gastrotheca marsupiata (Duméril & Bibron, 1841) (Hemiphractidae) (Gómez et al. Reference Gómez, Sánchez, Ñacari and Espínola-Novelo2020). The other species reported in the country include Aplectana membranosa (Schneider, Reference Schneider1866) Miranda, Reference Miranda1924, associated with Gastrotheca marsupiata in northern and central Andean regions (Gómez et al. Reference Gómez, Sánchez, Ñacari and Espínola-Novelo2020), and Aplectana vellardi Travassos, Reference Travassos1926, found in R. arequipensis (Vellard, 1959), and R. marina from the regions of Ancash, Arequipa, Lima, and San Martín (Gómez et al. Reference Gómez, Sánchez, Ñacari and Espínola-Novelo2020; Sarmiento et al. Reference Sarmiento, Tantaleán and Huiza1999; Toledo et al. Reference Toledo, Fonseca, Cárdenas-Callirgos, Vidaurre and Silva2017). However, despite these records, knowledge of the helminth fauna of Amazonian anurans in Peru remains limited (Bursey et al. Reference Bursey, Goldberg and Parmelee2001; Campião et al. Reference Campião, Morais, Dias, Aguiar, Toledo, Tavares and Silva2014; Goldberg et al. Reference Goldberg, Bursey, Caldwell, Vitt and Costa2007).
As part of a parasitological survey of amphibian helminths conducted at the Kawsay Biological Station, Madre de Dios region, in the southern Peruvian Amazon, nematodes representing an undescribed species of Aplectana were recovered from the intestine of L. bolivianus. In this study, this species is described based on an integrative approach combining detailed morphological observation (combining light microscopy and scanning electron microscopy), with molecular data from the partial 18S rDNA gene.
Materials and methods
Collection, processing, and morphological analyses of parasites
Between August and September 2024, 71 adult individuals of L. bolivianus were captured through visual encounter surveys (VES) at the Kawsay Biological Station in the Madre de Dios region (12°31′S, 69°W), Peru. The anurans were handled, identified in situ by a specialist, and euthanized using 20% benzocaine (Arteaga-Navarro and Guayasamin Reference Arteaga-Navarro and Guayasamin2011). The anuran collection was authorized by the Servicio Nacional Forestal y de Fauna Silvestre (SERFOR) under Directorial Resolution N°D000234-2024-MIDAGRI-SERFOR-DGGSPFFS-DGSPFS. For parasitological examination, the guts were removed, placed in a petri dish with 0.9% saline solution, and examined under a stereoscopic microscope. The nematodes found were transferred to petri dishes with 0.9% saline solution to be cleaned of organic residues and mucus, fixed with hot 70% ethanol, and preserved in 80% ethanol. In the laboratory, nematodes were cleared in lactophenol (Andrade Reference Andrade2000; Portela et al. Reference Portela, Santos and Dos Anjos2020). Specimens were examined and photographed using a Nikon™ Eclipse Ni-L compound photomicroscope equipped with normal light and differential interference contrast microscopy (DIC), and drawings were made with the aid of a drawing tube. Measurements were taken in micrometres, unless otherwise indicated, straight-line distances between extreme points of the structures measured and are expressed as the range, followed by the mean and the number (n) of structures measured in parentheses. The terminology and pattern of the caudal papillae followed those proposed by González et al. (Reference González, Gómez and Hamann2019). For SEM observation, 16 specimens (5 females and 11 males) were post-fixed in 10% formaldehyde, dehydrated through a graded ethanol series, critical-point dried with carbon dioxide, coated with gold, and examined using an Inspect S50-FEI at an accelerating voltage of 5kV to 10kV (González et al., Reference González, Hamann, Laurenti and Kazmiruk2012). Type specimens were deposited in the Helminthological Collection of the Museum of Natural History at the National University of San Marcos (MUSM-HEL), Lima, Peru.
Authorship of the new taxa is attributed to the first and senior authors following the Recommendation 50.1 (Identity of authors) of the International Code of Zoological Nomenclature (ICZN, 2012). To comply with the regulations set out in article 8.5 of the amended 2012 version of the International Code of Zoological Nomenclature (ICZN, 2012), details of the new taxon have been submitted to ZooBank. The Life Science Identifier (LSID) is reported in the taxonomic summary.
Molecular analysis
Genomic DNA of the nematodes was extracted using a TIANamp Genomic DNA Kit (TianGen, Beijing, China) following the manufacturer’s protocol. The partial 18S rDNA was amplified by polymerase chain reaction (PCR) using forward primer Nem_18S_F (5′-CGCGAATRGCTCATTACAACAGC-3′) and reverse primer Nem_18S_R (5′-GGGCGGTATCTGATCGCC-3′) (Floyd et al. Reference Floyd, Rogers, Lambshead and Smith2005). PCR assays were carried out in a total volume of 20 μL containing 1× QIAGEN PCR buffer (with MgCl₂ at 1.5 mM), 0.2 mM dNTPs, 0.5 μM of each primer, Taq DNA polymerase (QIAGEN) at 0.025 U/μL, 1 μL of DNA (2–10 ng), and ultrapure water to complete. PCR cycling conditions consisted of an initial denaturation at 95°C for 1 min, followed by 35 cycles of 95°C for 15 s, 55°C for 15 s, and 72°C for 20 s, with a final extension at 72°C for 1 min. PCR products were purified with sparQ PureMag Beads (Quantabio, USA) following the manufacturer’s instructions and sequenced in Psomagen Inc. (United States) with the Sanger sequencing method.
Sequences were edited and contig was assembled using Geneious R9.1 (Kearse et al. Reference Kearse, Moir, Wilson, Stones-Havas, Cheung, Sturrock and Drummond2012). Similar sequences were retrieved from the National Center for Biotechnology Information (NCBI) database (henceforth ‘GenBank’) using the Basic Local Alignment Search Tool (BLAST). The 18S sequence generated in this study was aligned with reference sequences of identified species and putative lineages of Cosmocercidae available in GenBank (Table 2) using Clustal W (Larkin et al. Reference Larkin, Blackshields, Brown, Chenna, McGettigan, McWilliam, Valentin, Wallace, Wilm, Lopez, Thompson, Gibson and Higgins2007). Cucullanus grandistomis (Ferrez & Thatcher, 1988) was used as outgroup taxa (Table 2).
The best-fitting nucleotide substitution model according to jModelTest2 (Santorum et al. Reference Santorum, Darriba, Taboada and Posada2014) was K80+G. Because K80 is not explicitly implemented in MrBayes, Bayesian inference analyses were conducted under the HKY+G model (nst = 2 with gamma-distributed rate variation), allowing base frequencies to be estimated from the data. Analyses were performed in MrBayes v3.2.6 (Ronquist et al. Reference Ronquist, Teslenko, Van Der Mark, Ayres, Darling, Höhna, Larget, Liu, Suchard and Huelsenbeck2012) using two independent runs of four Markov chains each, starting from random trees. The Markov chain Monte Carlo (MCMC) analyses were run for 1,000,000 generations, with trees sampled every 1,000 generations. The first 25% of the sampled trees were discarded as burn-in, and the remaining trees were used to construct a majority-rule consensus tree.
The proportion (p) of absolute nucleotide sites (p-distance) (Nei and Kumar Reference Nei and Kumar2000) was calculated to compare the genetic distance between sequences used.
Results
Cosmocercoidea Railliet, 1926
Cosmocercidae Railliet, 1925
Cosmocercinae Railliet, 1916
Aplectana Railliet & Henry, 1916
Aplectana peruensis n. sp. Miranda & Chero
Type-host: Leptodactylus bolivianus Boulenger, 1898 (Anura: Leptodactylidae)
Site in host: Small intestine.
Type locality: Kawsay Biological Station (12°31′S, 69°W), Madre de Dios region, Peru.
Type material: Holotype, male (MUSM-HEL 5630), allotype, female (MUSM-HEL 5631); 10 paratypes, males (MUSM-HEL 5632); 20 paratypes, females (MUSM-HEL 5633).
Prevalence: 58 of 71 hosts infected (81.69%) with a total of 1096 worms.
Mean intensity: 19 nematodes per infected host (range 1–114).
Etymology: The specific epithet refers to the country where the species was found.
General
Small-sized, whitish nematodes. Body cylindrical, maximum width at about region of middle body. Cuticle with fine transverse striations. Somatic papillae small, distributed irregularly over body surface. Lateral alae extending from 100 to 190 (145; n = 20) posterior to the base of the lips; in females, the alae reach approximately the mid-tail region, whereas in males they extend only to the level of the third precloacal papilla (Fig. 1C). Oral aperture simple, triangular, surrounded by 3 small lips, each with inner flanges (Figs. 2A, 3B). Dorsal lip with one pair of papillae; subventral lips with single large double cephalic papilla and amphid each (Figs. 1B, 3A). Oesophagus divided into anterior short pharynx, cylindrical corpus, slightly narrower isthmus and terminal posterior bulb with valves (Fig. 1A). Nerve ring located at about 1/2 of oesophageal length. Excretory pore slightly anterior to oesophageal bulb (Figs. 1A, 3C). Tail of both sexes conical, with long filamentous tip (Figs. 1D-E).
Line drawings of Aplectana peruensis n. sp. (A) Female, anterior region, lateral view; (B) male, anterior extremity, apical view; (C) male, posterior region showing the pattern of caudal papillae, ventral view; (D) male, spicule and gubernaculum, lateral view; (E) female tail, lateral view; (F) female, vagina, lateral view; (G) male, spicules, ventral view; (H) male, gubernaculum, ventral view; (I) larvated egg. Scale bars: C = 100 μm; A, D, E, F, I = 50 μm; G, H = 40 μm.
Scanning electron micrographs of males of Aplectana peruensis n. sp. (A) Male showing the pattern of precloacal papillae (white arrows), the adcloacal papilla (ad), the postcloacal papillae (arrowhead), and the unpaired papilla (up); (B) unpaired papilla (up) and papillae on the upper lip of the cloaca (*); (C) lateral view showing the lateral ala (la); (D) posterior region showing the bifurcated spicule (white arrows).
Scanning electron micrographs of females of Aplectana peruensis n. sp. showing: (A) anterior region with large double cephalic papillae (white arrow), amphids (am), and somatic papillae (asterisks); (B) posterior region with the vulva (vu) and tail; (C) excretory pore (ep); (D) detail of the vulva (vu) with mamelon-like cuticular protuberances (white arrow).
Male (Based on 62 mature specimens; Figs. 1–2): Body 1.20–3.17 (2.08; n = 62) mm long, maximum width 42–192 (107; n = 62). Oesophagus 313–676 (394; n = 62) long, representing 21.3–26.1 (18.9)% of body length; pharynx + corpus + isthmus 233–478 (353; n = 62) long, bulb 58–85 (70) × 51–87 (67; n = 62) (Fig. 1A). Nerve ring 124–273 (176; n = 62) and excretory pore 248–371 (308; n = 62) from anterior extremity, respectively. Posterior end of body distinctly curved ventrally. Spicules similar in shape and length, 152–194 (169; n = 62) μm long, chitinised, bearing a proximal capitulum and a clearly defined distal bifurcation, representing 6.1–12.7 (8.1)% of body length (Figs. 1G, 2C). Gubernaculum present, V-shaped, 60–89 (70; n = 62) long (Fig. 1H). Three pairs of precloacal papillae arranged in a single ventral row, larger than the somatic papillae and with variable spacing that decreases towards the cloaca; five pairs of adcloacal papillae, divided into two groups, three ventral pairs located on the anterior cloacal lip arranged in a 1 + 3 pattern (one large unpaired median papilla flanked by three pairs of papillae) and two lateral pairs situated on each side of the cloaca; and five pairs of postcloacal papillae, distributed ventrally and laterally along the tail, with the anterior pairs larger and the posterior pairs progressively smaller (Figs. 1C, 2A). Tail 127–301 (172; n = 62) long, representing 9.5–10.6 (8.3)% of body length (Fig. 1C).
Female (Based on 41 mature specimens; Figs. 1–3): Body 1.42–4.19 (3.06; n = 41) mm long, maximum width 70–378 (191; n = 41). Oesophagus 236–560 (435; n = 41) long, representing 13.4–16.6 (14.2)% of body length; pharynx + corpus + isthmus 178–555 (389) long, bulb 67–110 (87) × 64–126 (87). Nerve ring 88–231 (189; n = 41) and excretory pore 227–427 (340; n = 41) from anterior extremity, respectively. Vulva transverse slit, 0.48–2.03 (0.94; n = 41) mm from posterior extremity, at 33.8–48.4 (30.7)% of body length (Figs. 1F, 3D), with three mamelon-like cuticular protuberances on the anterior lip of the vulva. Ovaries two, located anterior to vulva. Vagina muscular. Uteri amphidelphic, full of eggs in different stages of development; egg oval, large, with smooth surface, 59–115 (88) × 58–102 (83; n = 41) (Fig. 1I). Tail 114–323 (177; n = 41) long, representing 7.7–8.0 (5.8)% of body length (Fig. 1E).
Remarks
The specimens are assigned to Aplectana based on the presence of a suit of diagnostic characters, including: (1) well-developed lateral alae, (2) numerous minute somatic papillae, (3) 3 lips bearing 4 large submedian cephalic papillae, (4) 1 pair of lateral amphids, (5) an oesophagus with a short cylindrical pharynx, (6) a cylindrical corpus and a well-developed oesophageal bulb, (7) an excretory pore located anterior to the oesophageal bulb, (8) 2 equal spicules in males, and (9) male tail lacking rosettes or plectanes (Baker Reference Baker1981; Bursey et al. Reference Bursey, Goldberg, Siler and Brown2018; Ramallo et al. Reference Ramallo, Goldberg, Ruiz and Stazzonelli2023; Vieira et al. Reference Vieira, Goncalves, Lima, Sousa and Muniz-Pereira2020).
Aplectana peruensis n. sp. is distinguished from its congeners by a unique combination of characters, including the presence of 13 pairs of caudal papillae plus a single unpaired papilla, a well-developed gubernaculum, and relatively long spicules (152–194) bearing a clearly defined distal bifurcation. Among Neotropical species of Aplectana, A. peruensis n. sp. most closely resemble A. albae Adamson & Baccam, Reference Adamson and Baccam1988 and A. hylambatis (Baylis, Reference Baylis1927), as these species share males bearing a gubernaculum, 13 pairs of caudal papillae, and a single unpaired papilla anteriorly to cloaca (Table 1). However, the new species can be readily distinguished from A. albae by the arrangement of the caudal papillae (3 precloacal, 5 adcloacal, and 5 poscloacal in A. peruensis n. sp. vs 6–7 precloacal, 1 adcloacal, and 5 postcloacal in A. albae), as well as by the larger spicules (150–190 in the new species vs 61–63 in A. albae) and gubernaculum (60–190 in the new species vs 41–46 in A. albae). Aplectana peruensis n. sp. differs from A. hylambatis in the arrangement of the caudal papillae (3 precloacal, 5 adcloacal, and 5 poscloacal in A. peruensis n. sp. vs 3–5 precloacal, 4–5 adcloacal, and 5–8 postcloacal in A. hylambatis), and in spicule morphology, which bear a pointed distal end in the new species, in contrast to the articulated distal end observed in A. hylambatis.
Comparative measurements of Neotropical Aplectana species with presence of a gubernaculum
* Precloacal papillae: adcloacal papillae: postcloacal papillae + unpaired papilla.
Genetic information of the species analyzed in phylogenetic study
Molecular analysis
A partial sequence of the 18S rDNA gene (1112 bp) was obtained from a single specimen of A. peruensis n. sp. collected from L. bolivianus. 18S rDNA alignment comprised 15 taxa and 747 characters, of which 651 were conserved, 95 variable, and 39 parsimony-informative. Bayesian inference showed good convergence, with effective sample size (ESS) values exceeding 600 for all parameters. Mean and median log-likelihood values were −1671.789 and −1671.458, respectively. In the resulting Bayesian phylogenetic tree (Fig. 4), A. peruensis n. sp. formed a sister lineage to A. hylambatis from Pithercopus azureus (Cope, 1862) in the Neotropical region, with full support (PP = 1). This sister pair was, in turn, sister to a strongly supported clade (PP = 0.91) comprising A. pella and a separate clade including A. chamaeleonis and a clade formed by two Oriental species of Aplectana (A. dayaoshanensis Chen, Ni, Gu & Li, 2021 + A. xishuangbannaensis Chen, Ni, Gu & Li, 2021), as well as Cosmocerca ornata, and Cosmocerca simile. Genetic distances between A. peruensis n. sp. and these species ranged from 1% to 4.5%, corresponding to 7–30 bp (Table 3).
Phylogenetic reconstruction of cosmocercid nematodes based on 18S rDNA gene sequences using Bayesian inference. Scale bar indicates number of substitutions per site.
Pairwise sequence divergences for the 18S rDNA gene among Aplectana peruensis n. sp. and other nematode species. The proportion (p) of absolute nucleotide sites (p-distance) is shown below the diagonal, and the raw number of base pairwise differences is shown above the diagonal
Discussion
The morphological characters observed in A. peruensis n. sp. clearly support its separation from all previously described congeners worldwide. Within Cosmocercidae, species delimitation has traditionally relied on a set of stable and taxonomically informative traits, particularly the number and arrangement of caudal papillae (Gibbons Reference Gibbons2010), spicule length and morphology (Piñeiro-Gómez et al. Reference Piñeiro-Gómez, González and Sanabria2017), the configuration of the caudal extremity (Ramallo et al. Reference Ramallo, Goldberg, Ruiz and Stazzonelli2023), and the presence or absence of a gubernaculum (Chen et al. 2021). These characters have repeatedly proven reliable for distinguishing closely related species and remain central to the description of new taxa within the family (Bursey and Goldberg Reference Bursey and Goldberg2004; Bursey et al. Reference Bursey, Goldberg, Siler and Brown2018; Ramallo et al. Reference Ramallo, Goldberg, Ruiz and Stazzonelli2023; Vieira et al. Reference Vieira, Goncalves, Lima, Sousa and Muniz-Pereira2020).
Among these traits, the pattern of caudal papillae represents one of the most consistent diagnostic characters within Aplectana. Several studies have demonstrated that the arrangement of precloacal, adcloacal, and postcloacal papillae enables the separation of morphologically similar species, even in cases where other characters exhibit partial overlap (Bursey and Goldberg Reference Bursey and Goldberg2015; Felix-Nascimento et al. Reference Felix-Nascimento, Vieira, Muniz-Pereira and Moura2020; Piñeiro-Gómez et al. Reference Piñeiro-Gómez, González and Sanabria2017). In A. peruensis n. sp., the combination of 3 precloacal, 5 adcloacal, and 5 postcloacal papillae, together with the presence of a single unpaired precloacal papilla, does not correspond to any pattern previously documented for Neotropical species (Ramallo et al. Reference Ramallo, Goldberg, Ruiz and Stazzonelli2023; Vieira et al. Reference Vieira, Goncalves, Lima, Sousa and Muniz-Pereira2020). This distinctive papillae arrangement alone provides strong support for the recognition of this taxon as a new species. Spicule morphology further reinforces the distinctiveness of A. peruensis n. sp. Recent integrative studies have emphasized that modifications of the distal end of the spicule, such as bifurcations, expansions, or membranous structures, are highly informative for species-level delimitation within the genus (Bursey et al. Reference Bursey, Goldberg, Siler and Brown2018; Chen et al. Reference Chen, Ni, Gu, Sinsch and Li2021b). Distally bifurcated spicules, as observed in A. peruensis n. sp., have also been reported in the original descriptions of A. pusilla (Miranda, Reference Miranda1924) and A. membranosa (Miranda, Reference Miranda1924), with subsequent clarification of this character in A. membranosa by Santos et al. (Reference Santos, Cardoso, Tavares-Costa, Figueredo, Rebêlo, Müller, Alcantara, Adriano, Morais, Freire, Dos Santos and de Vasconcelos Melo2025). In A. peruensis, the bifurcation occurs directly at the distal tip of the spicule, whereas in A. membranosa it is associated with a hyaline membrane, and in A. pusilla the distal modification is less pronounced. This distinctive character, together with the number and arrangement of caudal papillae and the lengths of the spicule and gubernaculum, reinforces the validity of A. peruensis n. sp. as a new taxon.
The gubernaculum also provides important diagnostic information. Whereas some species of Aplectana exhibit a reduced or absent gubernaculum, others possess a well-developed structure. The taxonomic relevance of the gubernaculum as a diagnostic character in Aplectana is supported not only for Neotropical species but also for Oriental (Asian) taxa. Several descriptions of Asian species distinguished taxa based on the presence or absence of the gubernaculum in combination with the arrangement of caudal papillae (Bursey et al. Reference Bursey, Goldberg, Siler and Brown2018; Chen et al. 2021; Sou et al. Reference Sou, Sow and Nandi2018).
The description of A. peruensis n. sp. increases the number of known Cosmocercidae species in the Neotropical region, which currently records approximately 65 species in nine genera: Aplectana (28 species; Santos et al., Reference Santos, Borges, Willkens, Santos, Costa-Campos and Melo2023), Cosmocerca (13 species; Velázquez-Brito et al., Reference Velázquez-Brito, Garduño-Montes de Oca, García and León-Règagnon2023), Cosmocercoides (4 species; Velázquez-Brito et al., Reference Velázquez-Brito, Garduño-Montes de Oca, García and León-Règagnon2023), Cosmocercella (3 species; Baker & Vaucher, Reference Baker and Vaucher1983), Raillietnema (6 species; Moravec et al., Reference Moravec, Maldonado and López1990), Oxysomatium (1 species; Moravec et al., 2020), Neocosmocercella (2 species; Dos Santos et al., Reference Dos Santos, Rodrigues, Dos Santos, González and Melo2017), Oxyascaris (7 species; Dos Santos et al., Reference Santos, Willkens, Silva, Melo and Fernandes2023), and Paradollfusnema (1 species; Gonçalves et al., Reference Gonçalves, Vieira, Felix-Nascimento, Ribeiro, Silva and Muniz-Pereira2025). In addition, the description of the new species increases the number of recognized Aplectana species to 59 worldwide (Santos et al., Reference Santos, Borges, Willkens, Santos, Costa-Campos and Melo2023). Several authors have highlighted that Neotropical amphibians host diverse and still insufficiently documented parasite assemblages (Chero et al. Reference Chero, Cruces, Cacique, Ponce, Iannacone and Alvariño2023; González et al. Reference González, Hamann and Duré2021; Olivera and Campião Reference Olivera and Campião2024), particularly in regions where altered and preserved habitats coexist (Draghi et al. Reference Draghi, Drago, Saibene and Agostini2020; Euclydes et al. Reference Euclydes, Dudczak and Campião2021; Portela et al. Reference Portela, Santos and Dos Anjos2020).
In the present study, the partial 18S rDNA sequence of A. peruensis n. sp. from the Neotropical region was positioned as a sister species to A. hylambatis within a well-supported clade (Figure 4), indicating a close evolutionary relationship. A. pella (Neotropical) and A. chamaeleonis (Ethiopia) occupy distinct subclades, showing that A. peruensis represents an independent lineage within the Neotropical clade. The phylogenetic tree also reveals a geographic pattern in the diversification of Aplectana: Neotropical lineages (A. peruensis n. sp., A. hylambatis, A. pella) are clearly separated from Asian species (A. dayaoshanensis and A. xishuangbannaensis), reflecting a geographically structured diversification of the genus, consistent with observations by Neves et al. (Reference Neves, Rebêlo, Félix, Gardner, Maldonado, Costa-Campos and Melo2025).
Morphologically, both A. peruensis and A. hylambatis possess 13 precloacal papillae, five adcloacal papillae, and one unpaired papilla anterior to the cloaca. However, A. peruensis can be clearly distinguished by spicule morphology: in A. peruensis, the spicules exhibit a well-defined distal bifurcation at the tip, whereas in A. hylambatis, the spicules are articulated, with a pointed distal end and a capitulum at the proximal end (Castillo et al. Reference Castillo, González-Rivas and Acosta2023; Villegas-Ojeda et al. Reference Villegas-Ojeda, Fernández-Marinone, Jofré and González2025).
The parasitic fauna of Leptodactylidae has been extensively investigated in several Neotropical regions, particularly in Brazil, Argentina, Paraguay, and Mexico, where a high diversity of nematodes and other helminths has been from species of Leptodactylus and related genera (Goldberg et al. 2009; González et al. Reference González, Hamann and Duré2021; González and Hamann Reference González and Hamann2015; Velázquez-Brito et al. Reference Velázquez-Brito, Garduño-Montes de Oca, García and León-Règagnon2023). In contrast, parasitological studies on L. bolivianus are remarkably scarce. To date, the only available report corresponds to Bursey et al. (Reference Bursey, Goldberg and Parmelee2001) from Peru, who recorded the following gastrointestinal helminths: A. hylambatis, Cosmocerca brasiliense Travassos, Reference Travassos1925 (Cosmocercidae), Oswaldocruzia lopesi Freitas & Lent, Reference Freitas and Lent1938 (Molineidae), Physalopteroides venancioi (Lent, Freitas, and Proença, Reference Lent, Freitas and Proença1946) (Physalopteridae), Brevimulticaecum sp. (larvae) (Heterocheilidae), and Physaloptera sp. (larvae) (Physalopteridae). Thus, the description of A. peruensis n. sp. not only expands current knowledge of nematode diversity infecting L. bolivianus but also underscores the importance of integrative morphological and molecular approaches for uncovering the hidden diversity of helminths in Neotropical amphibians.
Data availability
All data that support the findings of this study are available in the main text.
Acknowledgements
The authors would like to thank Eduardo Almora, Tomás Villena, and Johanna Gomez, staff at the Laboratory of Invertebrate Zoology, Faculty of Biological Sciences (FCB), National University of San Marcos (UNMSM), Lima, Peru, for their technical support during the study.
Author contribution
NM: Investigation, processed the methodology of collecting parasites, formal analysis, writing – original draft, drawing pictures, writing – review and editing, prepared figures. JPG: Investigation, collected the hosts, and processed the methodology of collecting parasites. JDC: Investigation, formal analysis, writing – original draft, prepared figures, writing − review and editing, supervision. All authors have read and agreed to the published version of the manuscript.
Financial support
This research was funded by the Universidad Nacional Mayor de San Marcos – RR No. 005446-2025-R and project number B25100271.
Competing interests
The authors declare that they have no conflict of interest.
Ethical statement
No ethical statement was reported.
