Introduction
The Camallanidae Railliet & Henry, 1915 (Rhabditida: Camallanoidea) comprises a diverse and taxonomically complex group of nematodes that parasitize primarily fishes from both freshwater and marine environments, and less frequently amphibians and reptiles (Anderson et al. Reference Anderson, Chabaud and Wilmot2009; Moravec Reference Moravec1998). Members of this family are characterized by their distinctive reddish coloration in life and by the presence of a strongly sclerotized buccal capsule, a feature widely used in generic and subgeneric diagnoses (Anderson et al. Reference Anderson, Chabaud and Wilmot2009; Moravec and Thatcher Reference Moravec and Thatcher1997). In particular, the presence, structure and arrangement of internal ridges, spiral ornamentations, and trident-like structures within the buccal capsule have traditionally served as key criteria for delimiting genera within Camallanidae (Ailán-Choke and Pereira Reference Ailán-Choke and Pereira2021; Silva et al. Reference Silva, Santos, Trindade and Melo2026). Despite this apparently well-defined morphological framework, the systematics of Camallanidae remain contentious. Recent integrative studies incorporating molecular data have challenged the traditional reliance on buccal capsule morphology, suggesting that several genera may not represent monophyletic lineages (Ailán-Choke and Pereira Reference Ailán-Choke and Pereira2021; Ailán-Choke et al. Reference Ailán-Choke, Davies, Tavares and Pereira2019).
In the Neotropical region, camallanids are widely distributed and parasitize a broad range of freshwater fishes (Luque et al. Reference Luque, Aguiar, Vieira, Gibson and Portes Santos2011; Morey and Floríndez Reference Morey and Floríndez2018; Morey et al. Reference Morey, de Sousa, Carvalho, Cruz and Puicón2025; Pinheiro et al. Reference Pinheiro, Melo, Monks, Santos and Giese2018; Ramallo and Ailan Choke Reference Ramallo and Ailán-Choke2017; Ruffeil et al. Reference Ruffeil, Giese and Pinheiro2023). The Peruvian Amazon, one of the most biodiverse ichthyological regions worldwide (Albert et al. Reference Albert, Tagliacollo and Dagosta2020; Castello et al. Reference Castello, McGrath, Hess, Coe, Lefebvre, Petry, Macedo, Renó and Arantes2013; Dagosta and de Pinna Reference Dagosta and de Pinna2019; Ortega et al. Reference Ortega, Hidalgo, Correa, Espino, Chocano, Trevejo, Meza, Cortijo and Quispe2018; Reis et al. Reference Reis, Albert, Di Dario, Mincarone, Petry and Rocha2016), harbours a rich but still incompletely characterized nematode fauna. With respect to diversity of camallanids from Peruvian Amazon, some species have been reported parasitizing fishes from different families, including Spirocamallanus inopinatus (Travassos, Artigas & Pereira, 1928), Spirocamallanus pintoi (Kohn & Fernandes, 1988), Spirocamallanus juana Ramallo & Ailan Choke, Reference Ramallo and Ailán-Choke2017, Spirocamallanus iheringi Artigas & Pereira, 1928, Spirocamallanus krameri (Petter, 1974), and Paracamallanus amazonensis Ferraz & Thatcher, 1992.
Among siluriform hosts, species of the Auchenipteridae have received comparatively little attention regarding their helminth parasites. Centromochlus heckelii (De Filippi, 1853), commonly known as the spiny driftwood catfish and locally referred to in Peru as novia aceitera, is a small catfish widely distributed throughout the Amazon basin. This species is highly valued in the ornamental fish trade due to its manageable size, distinctive behavior, and adaptability to captivity (Ortega et al. Reference Ortega, Hidalgo, Correa, Espino, Chocano, Trevejo, Meza, Cortijo and Quispe2018).
During a parasitological survey of helminths infecting C. heckelii in the Peruvian Amazon, specimens of an undescribed nematode belonging to Denticamallanus were recovered from the intestine. Detailed morphological analyses using light and electron microscopy, together with molecular data, demonstrated that these specimens represent a new species. The present study describes this species and evaluates its implications for the monophyly and diagnostic consistency of Denticamallanus within Camallanidae.
Materials and methods
Collection of samples and morphological study
In May 2025, specimens of C. heckelii were collected using a “bolichera” net at two localities in the Itaya River, Loreto, Peru (Moena Caño, 3°45′S, 73°13′W; Cabo López, 3°47′S, 73°15′W). A total of 60 fish (30 per locality) were examined. Fieldwork and specimen collection were conducted under the permits Resolution N° 132-2014-GRL-DIREPRO, Resolution N° 21-2016 GRL-DIREPRO, and PTH-068-16-PECSANIPES. Hosts were subjected to systematic necropsy for the detection and isolation of helminths. Nematodes were recovered from the intestine, rinsed in saline solution (0.85%), and fixed in hot 70% ethanol. Host identification was carried out following the criteria of García-Dávila et al. (Reference García-Dávila, Castro-Ruiz and Vásquez2018).
Specimens intended for light microscopy were cleared in Amman’s lactophenol and mounted on temporary slides (Amato et al. Reference Amato, Boeger and Amato1991). The 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 number (n) of structures measured in parentheses. Taxonomic identification of the parasites was based on the morphological descriptions provided by Moravec (Reference Moravec1998). Parasitological indices were calculated according to the criteria established by Bush et al. (Reference Bush, Lafferty, Lotz and Shostak1997).
For scanning electron microscopy (SEM), selected specimens were post-fixed in 10% formalin, dehydrated through a graded ethanol series, subjected to critical point drying using carbon dioxide, sputter-coated with gold, and examined using an Inspect S50-FEI, at an accelerating voltage of 5–10kV.
Type specimens were deposited in the Helminthological Collection of the Museum of Natural History at the San Marcos University (MUSM-HEL), Lima, Peru.
Genetic procedures
Genomic DNA of two specimens 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 PhilonemaF (5′−GCC TAT AAT GGT GAA ACC GCG AAC−3′) and the reverse primer PhilPCRr (5′–CCG GTT CAA GCC ACT GCG ATTA–3′) (Černotíková et al. Reference Černotíková, Horák and Moravec2011). PCR assays were carried out in a total volume of 25 μl containing 12.5 μl of PCRBIO Taq Mix Red (PCR Biosystems), 0.5 μl of each primer with final concentration at 10 μM, 5 μl of DNA sample, and ultrapure water to complete. An annealing temperature of 56°C was used for amplification. PCR products were purified with sparQ PureMag Beads (Quantabio, USA) following the manufacturer’s instructions and sequenced in PSOMAGEN Inc. (Brooklyn, New York, USA) with the Sanger sequencing method.
Phylogenetic analyses of molecular data
Sequences were edited and contig was assembled using Geneious 2026.0.2. 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 sequences generated in this study were aligned with reference sequences of identified species of Procamallaninae available in GenBank (Table 1) using Molecular Evolutionary Genetics Analysis v.12 (MEGA 12) (Kumar et al. Reference Kumar, Stecher, Suleski, Sanderford, Sharma and Tamura2024). Dracunculus insignis (Leidy, 1858) was used as outgroup taxa (Table 1). The best-fitting nucleotide substitution model according to jModelTest 2 (Darriba et al. Reference Darriba, Taboada, Doallo and Posada2012) was SYM+G+I. Because SYM is not directly implemented in MrBayes, Bayesian inference analyses were conducted under the GTR model (nst = 6), with a proportion of invariable sites and gamma-distributed rate variation (invgamma), as GTR represents a more general model that accommodates the substitution scheme of SYM, using four discrete gamma categories. Base frequencies were fixed as equal. Bayesian analyses were conducted in MrBayes v3.2.6 (Ronquist et al. 2012) 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.
List of camallanids reported parasitizing fish species in the Peruvian Amazon
Results
Camallanidae Railliet & Henry, 1915
Procamallaninae Yeh, 1960
Denticamallanus Moravec and Thatcher, Reference Moravec and Thatcher1997
Denticamallanus peruensis n. sp.
Type-host: Centromochlus heckelii (De Filippi, 1853) (Siluriformes: Auchenipteridae).
Site in host: Intestine.
Type locality: Itaya River (3°46′S, 73°15′W), Loreto Region, Peru.
Type material: Holotype, male (MUSM-HEL 5671), allotype, female (MUSM-HEL 5672); 9 paratypes, males (MUSM-HEL 5673a-i); 9 paratypes, females (MUSM-HEL 5674a-i).
Prevalence: 60 of 60 hosts infected (100%) with a total of 160 worms.
Mean intensity: 8 nematodes per infected host (range 4–8).
Etymology: The specific epithet refers to the country where the species was found.
General
Medium-sized nematodes with almost smooth cuticles. Oral opening circular, surrounded by six low elevations, two lateral and four submedian; twelve cephalic papillae arranged in three circlets, and two lateral amphids present. Buccal capsule orange-brown, slightly wider than long in males and nearly as long as wide in females; basal ring indistinct. Muscular oesophagus nearly equal in length to the glandular portion; both regions clavate. Small deirids located just posterior to level of nerve ring; excretory pore slightly posterior to nerve ring. Females with conical tail bearing an elongated finger-like projection. Male tail laterally expanded into two distinct lobes (caudal alae) and terminating in a ventrally directed, finger-like projection.
Male [Based on 10 mature specimens; Figs. 1A, C, H and 2E–F]: Body 7.5–9.7 (8.5; n = 10) mm long, maximum width 0.23–0.36 (0.27; n = 10) mm. Buccal capsule 230–300 (270; n = 8) long, 260–340 (300; n = 8) wide. Inner surface of capsule lacking spiral thickenings (ribs), but bearing 10–12 large, single, anteriorly directed conical teeth in its posterior half. Muscular oesophagus 530–730 (630; n = 10) long, 90–150 (130; n = 8) wide; glandular oesophagus 400–690 (560; n = 9) long, 80–140 (110; n = 10) wide. Caudal alae present, forming broad lateral lobes that give posterior extremity bulbous, spade-like shape, 120–210 (150; n = 78) long, 50–100 (80; n = 8) wide. One pair of preanal and four pairs of postanal subventral papillae present; first pair of postanal papillae located a short distance posterior to anal opening, second pair situated slightly posterior to caudal alae; third pair small, located laterally near distal region of tail; four pair small, located at base of digitiform process. Spicules equal, well sclerotized, short, 50–100 (80; n = 10) long, distal ends rounded, provided with a small terminal membranous process. Gubernaculum absent. Tail 200–280 (240; n = 10) long, bearing a ventrally directed finger-like projection, 80–160 (120; n = 10).
Denticamallanus peruensis n. sp. (A) Anterior end of male, lateral view; (B) buccal capsule of gravid female, dorsoventral view; (C) optical section of lower part of buccal capsule of the female; (D) cephalic end of male, later view; (E) tail of male, lateral view; (F) spicules; (G) tail of male, ventral view; (H) tail of gravid female, lateral view.
Scanning electron micrographs of Denticamallanus peruensis n. sp. (A) Cephalic end of male showing three circles of cephalic papillae (a, b, c), and the amphid (am, arrowhead); (B) cephalic end of female; (C) excretory pore (ep); (D) tail of a male (lateral view) showing broad lateral lobes; (E) tail of male showing a finger-like projection and phasmids (*); (F) tail of a male (ventral view) shoving one pair of preanal, four pairs of postanal papillae (arrows) and lateral phasmids (*); (G) tail of a female (lateral view) showing anus.
Female [Based on 10 mature specimens; Figs. 1A, D–G and 2A–D]: Body 11.7–27.9 (17.2; n = 10) mm long, maximum width 0.21–0.48 (0.33; n = 10) mm. Buccal capsule 100–140 (120; n = 9) long, 90–110 (100; n = 10) wide. Inner surface of posterior half of buccal capsule bearing 11–14 spiral thickenings. Muscular oesophagus 0.60–1.02 (0.70; n = 10) mm long, 0.09–0.17 (0.13; n = 10) wide, glandular oesophagus 0.48–1.04 (0.70) long, 0.10–0.17 (0.13; n = 10) wide. Vulva located approximately at mid-body. Uterus containing numerous larvae. Tail short, 70–190 (100; n = 10) long, bearing an elongated finger-like appendage, 50–110 (80; n = 9) long.
Remarks
The newly collected specimens are assigned to Denticamallanus, as defined by Moravec and Thatcher (Reference Moravec and Thatcher1997), based on the marked sexual dimorphism observed in the structure of the buccal capsule. In females, the buccal capsule bears a few spiral ridges with only faintly outlined tooth-like structures. In contrast, males lack spiral ridges entirely and instead possess several large, sharply pointed, discrete teeth.
Traditionally, Denticamallanus comprised two species, Denticamallanus dentatus Moravec & Thatcher, Reference Moravec and Thatcher1997 (type species) from Bryconops alburnoides Kner, 1859 (Cypriniformes) in Brazil, and D. ana Ramallo, 2011 from Thoracocharax stellatus (Kner, 1858) (Gasteropelecidae) in Argentina. Denticamallanus peruensis n. sp. can be readily distinguished from its two congeners by the distinctive morphology of the male caudal region, which forms broad lateral lobes giving the posterior extremity a conspicuously bulbous, spade-like appearance. In addition, D. peruensis n. sp. differs from D. dentatus and D. ana in the number and arrangement of caudal papillae. The new species possesses one pair of preanal and four pairs of postanal papillae, whereas D. dentatus exhibits two pairs of preanal and three pairs of postanal papillae, and D. ana has two to three pairs of preanal papillae, one pair of adanal papillae, and two pairs of postanal papillae. Furthermore, the new species differs from both congeners in spicule length (50–100 in D. peruensis n. sp. vs 78–117 in D. dentatus and 40–46 in D. ana) and in the number of teeth in the male buccal capsule (10–12 in D. peruensis n. sp. versus about 10 in D. dentatus and 2 in D. ana).
Recently, Silva et al. (Reference Silva, Santos, Trindade and Melo2026) amended Denticamallanus to include species characterized by the presence of conical tooth-like structures at the basal ring or in the posterior region of the buccal capsule of at least one sex, with spiral ridges that may or may not be present. In that study, the author transferred the following species to Denticamallanus, P. (Procamallanus) annipetterae Kohn & Fernandes, 1988; P. (Spirocamallanus) chimuensis (Freitas & Ibáñez, 1968); P. (S.) iheringi (Travassos, Artigas & Pereira, 1928); P. (S.) krameri (Petter, 1974); P. (S.) rarus Travassos, Artigas & Pereira, 1928; P. (S.) saofranciscencis (Moreira, Oliveira & Costa, 1994); P. (P.) spiculastriatus Pinhiero, Melo, Monks, Santos & Giese, 2018; and P. (Spirocamallanus) huacraensis Ramallo, 2008. Denticamallanus peruensis n. sp. differs from all these species in the structure of the male buccal capsule. In the new species, the male buccal capsule bears only discrete teeth and completely lacks spiral ridges, whereas in the other transferred species the male buccal capsule exhibits both spiral rings and teeth.
Molecular analysis
Two partial 18S rDNA sequences (1113 bp) were obtained from one male (PZ366531) and one female (PZ366532) specimen of D. peruensis n. sp. collected from C. heckelii. 18S rDNA alignment consisted of 17 taxa and 799 characters, of which 613 were conserved, 181 variables, and 98 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 −2959.252 and −2958.861, respectively. In the resulting Bayesian phylogenetic tree (Fig. 3), the two sequences of D. peruensis n. sp. clustered together in a well-supported monophyletic clade. This clade was recovered as the sister lineage to Spirocamallanus inopinatus (MT901634) from Megaleporinus elongatus (Valenciennes, 1850) in Brazil, with strong support (PP = 0.99). In turn, this assemblage formed the sister group to a strongly supported clade (PP = 1) comprising Spirocamallanus pintoi (DQ442666) from Hoplisoma atropersonatum (Weitzman & Nijssen, 1970) in Peru.
Phylogenetic reconstruction of Procamallaninae nematodes based on 18S rDNA gene sequences using Bayesian inference. Scale bar indicates number of substitutions per site.
Denticamallanus was recovered as non-monophyletic in our analysis. Denticamallanus peruensis n. sp. was nested within a clade otherwise composed of Spirocamallanus spp., whereas Denticamallanus rarus (Travassos, Artigas & Pereira, 1929) (JF803912) and Denticamallanus huacraensis (Ramallo, 2008) (MK794615), the only congeners for which molecular data are currently available, formed a distinct lineage positioned basally relative to the remaining ingroup taxa, with low to moderate support at the corresponding node (PP = 0.55).
Discussion
The present study represents the first record and formal description of a species of Denticamallanus from Peru, increasing the number of freshwater camallanid species known in the country to seven (see Table 2). This finding expands the documented diversity of Camallanidae in the Peruvian Amazon and provides new data relevant to the systematics of the family.
Collection data and GenBank accession numbers for sequences analysed in this study; new sequences in bold
* Outgroup.
Our 18S rDNA phylogeny recovered D. peruensis n. sp. within an assemblage comprising representatives of Procamallanus and Spirocamallanus, whereas D. rarus and D. huacraensis were resolved in a separate lineage. This topology appears to indicate a lack of support for the monophyly of Denticamallanus under the emended diagnosis proposed by Silva et al. (Reference Silva, Santos, Trindade and Melo2026), suggesting that the expanded generic concept may include phylogenetically heterogeneous taxa. Similar discordances between morphology-based classification and molecular phylogenies have been documented for Camallanidae (Ailán-Choke and Pereira Reference Ailán-Choke and Pereira2021; Ailán-Choke et al. Reference Ailán-Choke, Tavares, Luque and Pereira2020; Černotíková et al. Reference Černotíková, Horák and Moravec2011; Wijová et al. Reference Wijová, Moravec, Horák and Lukeš2006), where genera were not consistently recovered as monophyletic across datasets. Thus, the result observed herein aligns with accumulating evidence suggesting instability in generic delimitations within the family.
The systematics of Camallanidae has traditionally relied almost exclusively on buccal capsule morphology (Anderson et al. Reference Anderson, Chabaud and Wilmot2009; Moravec and Thatcher Reference Moravec and Thatcher1997; Rigby and Rigby 2014), particularly the presence, distribution, and configuration of internal ridges and associated accessory structures (Rigby and Rigby 2014). As mentioned by Silva et al. (Reference Silva, Santos, Trindade and Melo2026), generic diagnoses remain largely grounded in capsule morphology and trident configuration. However, persistent taxonomic instability associated with variation in buccal ornamentation suggests that these characters may not consistently reflect natural evolutionary relationships.
Ailán-Choke and Pereira (Reference Ailán-Choke and Pereira2021) conducted a critical integrative assessment of Camallanidae systematics, demonstrating that morphology-based classification founded on buccal capsule characters may be artificial for certain camallanid lineages. In their analysis, the integration of morphological traits with molecular phylogenies revealed that buccal capsule features can display homoplasy and variable phylogenetic consistency depending on the taxon assemblage examined. Moreover, they found that no genus was consistently recovered as monophyletic across datasets and that the phylogenetic informativeness of 18S rDNA differs from that of more variable markers such as COI. In this framework, the placement of D. peruensis n. sp. separate from other congeners may reflect deeper evolutionary structure not captured by capsule-based taxonomy.
The species described herein conforms to the original diagnosis of Denticamallanus proposed by Moravec and Thatcher (Reference Moravec and Thatcher1997), defined by marked sexual dimorphism of the buccal capsule, with males bearing posterior dentiform structures and females exhibiting internal spiral ridges without such teeth. Nevertheless, the BI topology does not support this character configuration as diagnosing a single evolutionary lineage. The emended diagnosis proposed by Silva et al. (Reference Silva, Santos, Trindade and Melo2026) broadened the generic diagnostic characters to include species such as D. rarus and D. huacraensis, whose males possess both spiral ridges and basal dentiform structures, differing from the original diagnosis.
Although 18S is relatively conserved and more informative for deeper divergences, the recurrent recovery of non-monophyletic genera within Camallanidae indicates that the observed pattern cannot be attributed solely to limited resolution. Rather, it may reflect an inherent artificiality in traditional generic delimitations. The revised taxonomic key proposed by Silva et al. (Reference Silva, Santos, Trindade and Melo2026) represents an important contribution toward morphological standardization; however, further molecular corroboration remains necessary to evaluate the stability of generic boundaries. Our findings support the need for a comprehensive integrative revision of Denticamallanus and related genera within Camallanidae, incorporating expanding taxon sampling, multilocus datasets, and a reassessment of the polarity and homology of buccal capsule characters. Within this framework, the transfer of five species from Spirocamallanus and two from Procamallanus to Denticamallanus, as proposed by Silva et al. (Reference Silva, Santos, Trindade and Melo2026), should be regarded as provisional pending more robust phylogenetic evidence.
Data availability
The obtained sequences are deposited in GenBank under the accession numbers PZ366531–32.
Acknowledgements
This work was funded by the Consejo Nacional de Ciencia, Tecnología e Innovación Tecnológica (CONCYTEC) and the Programa Nacional de Investigación Científica y Estudios Avanzados (PROCIENCIA) under the call ‘E041-2024-03 Proyectos de Investigación Básica’ [PE501087644-2024].
Competing interests
The authors declare that they have no conflict of interest.
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