Comparative cytogenetics of Serrasalmidae (Teleostei: Characiformes): The relationship between chromosomal evolution and molecular phylogenies.

Serrasalmidae has high morphological and chromosomal diversity. Based on molecular hypotheses, the family is currently divided into two subfamilies, Colossomatinae and Serrasalminae, with Serrasalminae composed of two tribes: Myleini (comprising most of pacus species) and Serrasalmini (represented by Metynnis, Catoprion, and remaining piranha's genera). This study aimed to analyze species of the tribes Myleini (Myloplus asterias, M. lobatus, M. rubripinnis, M. schomburgki, and Tometes camunani) and Serrasalmini (Metynnis cuiaba, M. hypsauchen, and M. longipinnis) using classical and molecular cytogenetic techniques in order to understand the chromosomal evolution of the family. The four species of the genus Myloplus and T. camunani presented 2n = 58 chromosomes, while the species of Metynnis presented 2n = 62 chromosomes. The distribution of heterochromatin occurred predominantly in pericentromeric regions in all species. Tometes camunani and Myloplus spp. presented only one site with 5S rDNA. Multiple markers of 18S rDNA were observed in T. camunani, M. asterias, M. lobatus, M. rubripinnis, and M. schomburgkii. For Metynnis, however, synteny of the 18S and 5S rDNA was observed in the three species, in addition to an additional 5S marker in M. longipinnis. These data, when superimposed on the phylogeny of the family, suggest a tendency to increase the diploid chromosome number from 54 to 62 chromosomes, which occurred in a nonlinear manner and is the result of several chromosomal rearrangements. In addition, the different karyotype formulas and locations of ribosomal sequences can be used as cytotaxonomic markers and assist in the identification of species.

Hyperspectral data as a biodiversity screening tool can differentiate among diverse Neotropical fishes.

Hyperspectral data encode information from electromagnetic radiation (i.e., color) of any object in the form of a spectral signature; these data can then be used to distinguish among materials or even map whole landscapes. Although hyperspectral data have been mostly used to study landscape ecology, floral diversity and many other applications in the natural sciences, we propose that spectral signatures can be used for rapid assessment of faunal biodiversity, akin to DNA barcoding and metabarcoding. We demonstrate that spectral signatures of individual, live fish specimens can accurately capture species and clade-level differences in fish coloration, specifically among piranhas and pacus (Family Serrasalmidae), fishes with a long history of taxonomic confusion. We analyzed 47 serrasalmid species and could distinguish spectra among different species and clades, with the method sensitive enough to document changes in fish coloration over ontogeny. Herbivorous pacu spectra were more like one another than they were to piranhas; however, our method also documented interspecific variation in pacus that corresponds to cryptic lineages. While spectra do not serve as an alternative to the collection of curated specimens, hyperspectral data of fishes in the field should help clarify which specimens might be unique or undescribed, complementing existing molecular and morphological techniques.

Chromosomal Analysis of Ctenolucius hujeta Valenciennes, 1850 (Characiformes): A New Piece in the Chromosomal Evolution of the Ctenoluciidae.

Ctenoluciidae (Characiformes), a family of freshwater fishes, comprises 2 genera, Ctenolucius and Boulengerella, with 7 recognized species. Up to now, only species of the genus Boulengerella have been subjected to cytogenetic studies. Here, we investigated the karyotype and other cytogenetic features of pike characin, Ctenolucius hujeta, using conventional (Giemsa staining, C-banding, Ag-NOR staining) and molecular (rDNA, telomeric sequences, and fiber-FISH mapping) procedures. This species has a diploid chromosome number of 2n = 36, and a karyotype composed of 12m + 20sm + 4a and FN = 68, similar to that found in Boulengerella species. However, differences regarding the number and distribution of several chromosomal markers support a distinct generic status. Colocalization of the 18S and 5S rDNA genes is an exclusive characteristic of the C. hujeta genome, with an interspersed distribution in the chromosomal fiber, an unusual phenomenon among eukaryotes. Additionally, our results support the view that Ctenoluciidae and Lebiasinidae families are closely related.

Cast netting new species: Integrative taxonomy of emDistichodus/em emnotospilus/em (Characiformes: Distichodontidae) discovers new species and overlooked areas of endemism in Central Africa.

Distichodus notospilus was described from the Ogooué River and is considered to occur throughout the Lower Guinea ichthyofaunal province and the western tributaries of the middle and lower Congo River. Recent expeditions in Equatorial Guinea collected D. notospilus specimens in the Mbini River drainage and the Mbia River; a small coastal river that is located between the Ntem and Mbini river drainages. Detailed morphological analyses and multilocus molecular analyses confirm that these two populations are distinct from one another. Topotypic populations of D. notospilus were included in the analyses and demonstrated that populations in the Mbini and Mbia rivers are distinct and these two new species are described herein. Distichodus microps sp. nov. is endemic to the Mbia River drainage and is distinguished from D. notospilus in having more scales along the lateral line (41, rarely 40 versus 37-39, rarely 40), a nearly inferior mouth versus subterminal in D. notospilus, a curved posterolateral margin of the opercle versus straight in D. notospilus, a smaller eye (56.7-80.4 versus 70.1-104.3 % of snout length), and a less prominent elongated spot at the base of the caudal fin. Distichodus mbiniensis sp. nov. is endemic to the upper Mbini River drainage and distinguished from D. notospilus in having more scales along the lateral line (41-42, rarely 40 versus 37-39, rarely 40), a much less prominent elongated dark spot at the base of the caudal fin, and a shorter dorsal fin (21.4-27.2 versus 22.7-34.2% standard length). Distichodus microps is distinguished from D. mbiniensis in having a shallower body (usually six scales from lateral line to the pelvic fin versus seven), fewer anal-fin rays (usually 12 total rays versus 13 or 14), a more inferior mouth, a deeper and longer caudal peduncle, a smaller eye, and differences in several features associated with the head. In addition to the two new species described this study also revealed potential undescribed diversity in the D. notospilus species complex in the Ntem River and Dja River (Congo R. basin) in Cameroon. The biogeography of these fishes in the rivers of Lower Guinea suggests that the Mbini River and smaller coastal rivers are overlooked areas of endemism. Studies of other reported widespread species will likely reveal additional diversity and further elucidate the processes promoting and maintaining freshwater diversity in Central Africa.

High Genetic Diversity despite Conserved Karyotype Organization in the Giant Trahiras from Genus Hoplias (Characiformes, Erythrinidae).

In the fish genus Hoplias, two major general groups can be found, one of which is formed by the "common trahiras" (Hoplias malabaricus group) and the other by the "giant trahiras" (Hoplias lacerdae group, in addition to Hoplias aimara), which usually comprises specimens of larger body size. Previous investigations from the giant trahiras group recovered 2n = 50 meta/submetacentric chromosomes and no sex chromosome differentiation, indicating a probable conservative pattern for their karyotype organization. Here, we conducted comparative cytogenetic studies in six giant trahiras species, two of them for the first time. We employed standard and advanced molecular cytogenetics procedures, including comparative genomic hybridization (CGH), as well as genomic assessments of diversity levels and phylogenetic relationships among them. The results strongly suggest that the giant trahiras have a particular and differentiated evolutionary pathway inside the Hoplias genus. While these species share the same 2n and karyotypes, their congeneric species of the H. malabaricus group show a notable chromosomal diversity in number, morphology, and sex chromosome systems. However, at the same time, significant changes were characterized at their inner chromosomal level, as well as in their genetic diversity, highlighting their current relationships resulting from different evolutionary histories.

Adaptation to herbivory and detritivory drives the convergent evolution of large abdominal cavities in a diverse freshwater fish radiation (Otophysi: Characiformes).

Convergent evolution is often interpreted as evidence of natural selection favoring an optimal phenotype during adaptation. Morphological convergence is frequently found among lineages that converge on diet, but most studies have focused on morphological traits that relate exclusively to food handling and processing. In vertebrates, there is a strong inverse relationship between intestine length and trophic level. However, little is known about whether adaptation to a low trophic level influences the evolution of abdominal cavities that can accommodate larger intestines. Here, I reconstruct the evolutionary history of trophic ecology and examine abdominal cavity shape across 157 species of the fish order Characiformes to determine whether adaptation to an herbivorous-detritivorous diet drives convergent evolution of large abdominal cavities. Herbivorous-detritivorous species evolved significantly larger abdominal cavities than other trophic groups and repeatedly converged on a similar abdominal cavity morphology. Other trophic groups evolved abdominal cavity morphologies either stochastically or by selective pressures from an untested ecological character. These findings demonstrate that the selective demands of a larger intestinal tract promote the repeated convergence of a large abdominal cavity within herbivorous-detritivorous characiform fishes, while allowing other lineages to evolve randomly or adapt in response to other selection pressures, contributing to the overall body shape diversity of the order.

A new species of Characidium Reinhardt (Characiformes: Crenuchidae) from the Río Huallaga, central Peruvian Amazon, with a review of secondary sexual characters in the genus.

Characidium pumarinri n. sp. is described from the affluents of Río Huallaga, a tributary of Río Marañón that drains the eastern slopes of the Andes, in the Peruvian Amazon. The new species has a deep body (24-31% in LS ), 11-17 well-marked dark bars and remarkable sexual dimorphism related to colour pattern, in which males have darker anal, dorsal, pectoral and pelvic fins than females. C. pumarinri resembles some species allocated in Clade C4, mainly by having a high number of transversal bars and a deep body. In addition, a thorough review and further descriptions of the secondary sexual characteristics in Characidium are provided.

Adjustments in population and reproductive dynamics of native and non-native congeneric species during 26 years after invasion

We aimed to improve the understanding of the establishment of Serrasalmus marginatus (non-native), which was followed by a decrease in the abundance of Serrasalmus maculatus (native) in the upper Paraná River floodplain. We estimated age, mortality rate, length and age at first maturity and variations in gonad development in three time-periods along a 26-year truncated time scale for both species. Population and reproduction parameters of both species showed substantial fluctuations among periods. Most age classes were sampled in all time-periods, but with considerable difference in abundance, with predominance of older individuals in the second time-period and younger individuals in the third time-period for both species. The mortality rates decreased for both species in the second time-period, but increased for the native in the third time-period. Length and age at first maturity decreased in the second time-period for both species, increasing the number of mature individuals on their populations. In the third time-period, the number of immature individuals increased for both species. We suggest that species experienced stressful conditions during cooccurrence and this have resulted in physiological responses in both species, reflecting in population and reproductive adjustments that may have relaxed competitive interactions between them, optimizing survival, reproductive effort and coexistence.(AU)

Nosso objetivo foi aumentar o conhecimento sobre o estabelecimento de Serrasalmus marginatus (não nativa), que foi seguido pela diminuição na abundância de Serrasalmus maculatus (nativa) na planície de inundação do alto rio Paraná. Estimamos a idade, taxa de mortalidade, comprimento e idade de primeira maturação e variações no desenvolvimento gonadal em três períodos ao longo de 26 anos para as espécies. Os parâmetros populacionais e de reprodução das espécies mostraram flutuações substanciais. A maioria das faixas etárias foi amostrada em todos os períodos, mas com diferenças consideráveis na abundância, predominando indivíduos mais velhos no segundo período e indivíduos mais jovens no terceiro período para ambas as espécies. A taxa de mortalidade diminuiu para as espécies no segundo período, mas aumentou para a espécie nativa no terceiro período. O comprimento e a idade de primeira maturação diminuíram no segundo período para as espécies, aumentando o número de indivíduos adultos. No terceiro período, o número de indivíduos imaturos aumentou para ambas as espécies. Sugerimos que as espécies passaram por condições estressantes durante a coocorrência, resultando em respostas fisiológicas que desencadearam ajustes populacionais e reprodutivos que podem ter minimizado a interação competitiva entre elas, otimizando a sobrevivência, o esforço reprodutivo e a coexistência.(AU)

A new, narrowly distributed, and critically endangered species of Characidium (Characiformes: Crenuchidae) from the Distrito Federal, Central Brazil

A new species of Characidium is described from the Cerrado biome, in Brasília, Distrito Federal, Central Brazil. The new species can be readily diagnosed by the presence of two or three longitudinal rows of dots along the body sides, absence of bars, by the short pectoral fin, which does not reach the vertical through the dorsal fin origin. Additional useful diagnostic characters are the scaled isthmus, absence of the adipose fin, and the terminal mouth. The new species is only known from the córrego Taquara and its tributaries, a tributary of ribeirão do Gama, upstream from lago Paranoá, in the upper rio Paraná basin. In accordance to the IUCN Red List Categories and Criteria, the new species is categorized as Critically Endangered.(AU)

Uma nova espécie de Characidium é descrita do bioma Cerrado em Brasília, Distrito Federal, Brasil Central. A nova espécie pode ser diagnosticada pela presença de duas ou três fileiras longitudinais de pontos no corpo, ausência de barras e nadadeira peitoral curta, que não alcança a vertical que passa pela origem da nadadeira dorsal. Caracteres adicionais úteis na diagnose são o istmo escamado, a ausência da nadadeira adiposa e a boca terminal. A nova espécie é conhecida apenas do córrego Taquara e seus tributários, tributário do ribeirão do Gama a montante do Lago Paranoá, na bacia do alto rio Paraná. De acordo com os Critérios e Categorias para Listas Vermelhas da IUCN, a nova espécie é categorizada como uma Criticamente Ameaçada.(AU)

Use of bioacoustics in species identification: Piranhas from genus Pygocentrus (Teleostei: Serrasalmidae) as a case study.

The genus Pygocentrus contains three valid piranha species (P. cariba, P. nattereri and P. piraya) that are allopatric in tropical and subtropical freshwater environments of South America. This study uses acoustic features to differentiate the three species. Sounds were recorded in P. cariba, two populations of P. nattereri (red- and yellow-bellied) and P. piraya; providing sound description for the first time in P. cariba and P. piraya. Calls of P. cariba were distinct from all the other studied populations. Red- and yellow-bellied P. nattereri calls were different from each other but yellow-bellied P. nattereri calls were similar to those of P. piraya. These observations can be explained by considering that the studied specimens of yellow-bellied P. nattereri have been wrongly identified and are actually a sub-population of P. piraya. Morphological examinations and recent fish field recordings in the Araguari River strongly support our hypothesis. This study shows for the first time that sounds can be used to discover identification errors in the teleost taxa.

A new species of Characidium (Characiformes: Crenuchidae) from the rio Madeira basin, Brazil.

This study describes Characidium nambiquara, a new species from the upper rio Guaporé, rio Madeira basin, Brazil. The new species differs from most congeners by the presence of isthmus and area between the contralateral pectoral-fin bases completely naked. From congeners with some degree of scaleless ventral surface of the body C. nambiquara differs by having 10 circumpeduncular scales. The new species is also distinguished from congeners by the spotted colour pattern on body of the large-sized specimens and by having black dashes on all fins and conspicuous midlateral longitudinal dark stripe or conspicuous vertical bars absent. Characidium nambiquara further differs from most congeners by the presence of 34-36 pored scales on the lateral line, 3 horizontal scale rows above the lateral line and 3 horizontal scale rows from the lateral line to the midventral scale series. Remarks on intraspecific colour variation within the genus, not related to sexual dimorphism, are also provided.

The unknown diversity of the genus Characidium (Characiformes: Crenuchidae) in the Chocó biogeographic region, Colombian Andes: Two new species supported by morphological and molecular data.

Two new species of Characidium, C. tatama and C. dule, are described from the biogeographic Chocó region in western Colombia. Both new species are supported by both morphological and molecular data. C. tatama from the San Juan River in the Pacific and C. dule from the Atrato River in the Caribbean portion of Colombia are both distributed in the upper and lower portions of these basins. An extensive comparison with other trans- and cis-Andean species of Characidium was made, in addition to species delimitation, using COI sequences by distinct methods (GMYC, ABGD, bPTP).

Integrative taxonomy reveals hidden diversity in the southern African darters genus Nannocharax Günther 1867 (Characiformes: Distichodontidae).

The present study explored the diversity of Nannocharax within southern Africa by implementing three species delimitation methods for a data set consisting of 37 mitochondrial cytochrome oxidase subunit I sequences. Two unilocus coalescent methods, the General Mixed Yule Coalescent (GMYC) and the Bayesian implementation of the Poisson Tree Processes (bPTP), and a genetic distance method, the Automatic Barcode Gap Discovery (ABGD), were applied. Both GMYC and bPTP delimited the same operational taxonomic units (OTUs), revealing a higher diversity for the genus in the region than previously recognised, whereas the ABGD failed to delimit the same candidate species. All methods delimited two species groups, and these are supported based on colouration patterning and morphology; the Nannocharax multifasciatus and the Nannocharax macropterus species groups and the delimited OTUs were assigned to each. Two putative new species were identified, Nannocharax cf. lineostriatus "Okavango" from the Okavango River in Angola and N. cf. lineostriatus "Kwanza" from the Kwanza River system in Angola. The distribution of Nannocharax dageti was confirmed for the Upper Zambezi and extended to the Okavango system, and an identification key for the southern Africa Nannocharax species is provided.

Molecular phylogeny and species delimitation of the genus Schizodon (Characiformes, Anostomidae).

The genus Schizodon is part of a group of headstanders and relatives (Family Anostomidae) that are widespread and ecologically important fishes in South American rivers. Schizodon includes 15 nominal species but their taxonomy has been challenging due to paucity of decisive characters to diagnose species. We present new molecular data to assess species boundaries or molecular operational taxonomic units (MOTUs), and to infer phylogenetic relationships among species. Evidence from two mitochondrial and three nuclear genes was used in these analyses. Mitochondrial DNA data for 112 specimens (from 11 nominal species) supported 13 consensus MOTUs, six of which matched valid nominal species (Schizodon borellii, S. fasciatus, S. intermedius, S. isognathus, S. knerii and S. scotorhabdotus). The nominal species Schizodon vittatus, S. nasutus, and S. dissimilis were subdivided into two MOTUs each, revealing either cryptic species or strong population structuring. In contrast, S. platae and S. jacuiensis constituted a single MOTU, indicating a possible case of synonymy. Our phylogenetic analysis subdivided the genus Schizodon into two large clades that are compatible with observed color patterns and biogeographic distribution. The first clade includes species with three to four conspicuous dark vertical bars on the flanks that originated in the Amazonas region (S. borellii, S. dissimilis, S. intermedius, S. fasciatus, S. scotorhabdotus, S. vittatus, and a cryptic species, Schizodon aff. vittatus). The second clade includes species with a conspicuous dark caudal blotch on the caudal peduncle, with vertical bars absent or inconspicuous, with a biogeographic origin in the La Plata drainage (S. isognathus, S. jacuiensis, S. knerii, S. nasutus and S. platae). Our results reinforce the importance of using molecular analyses to accelerate the study of diversity, particularly in groups with a wide distribution, few variable meristic characters, and high morphological plasticity, which may hide still unknown or underestimated diversity.

Hidden diversity in Prochilodus nigricans: A new genetic lineage within the Tapajós River basin.

Highly spread through the Amazon River basin, Prochilodus nigricans have had its taxonomic validity recently questioned, when genetic differences between Western and Eastern Amazon populations from the Brazilian shield were detected. This area has been seeing as a region of high ichthyofaunal diversity and endemism, in which the hybrid origin of the Tapajós River basin has been raised. In this paper, we report a new molecular lineage within P. nigricans of Tapajós River, highlighting this region still hides taxonomically significant diversity. Haplotype networks were reconstructed using the mitochondrial COI and ATP6/8 markers, which were also used to calculate genetic distances among clusters. We additionally conducted a delimiting species approach by employing a Generalized Mixed Yule-Coalescent model (GMYC) with COI sequences produced here, and previous ones published for individuals sampled across the Amazon River basin. In addition to the genetic differentiation within P. nigricans, our findings favor the hypothesis of hybrid origin of the Tapajós River basin and reaffirm the importance of studies aiming to investigate hidden diversity to address taxonomic and biogeographic issues, that certainly benefit better biodiversity conservation actions.

A new rheophilic South American darter (Crenuchidae: Characidium) from the rio Juruena basin, Brazil, with comments on morphological adaptations to life in fast-flowing waters.

Characidium iaquira, a new species from the upper rio Juruena, rio Tapajós basin, Brazil, is described. The new species can be promptly distinguished from all congeners by having a unique v-shaped dark mark lying along the caudal-fin extension, in medium- and large-sized specimens, and a remarkable iridescent green colouration in life. Characidium iaquira is closely related to Characidium crandellii and Characidium declivirostre by sharing unambiguous synapomorphies such as branchiostegal membranes united to each other across the isthmus, a scaleless area extending from the isthmus to the pectoral girdle, and dermal flaps surrounding anterior and posterior naris independent, but touching each other distally. Morphological specializations of the paired fins in the three riffle-dwellers species are discussed, including the wing-like shape, robustness, and inclination of the pectoral fin.

Phylogenomics of the Neotropical fish family Serrasalmidae with a novel intrafamilial classification (Teleostei: Characiformes).

The Neotropical fish family Serrasalmidae comprises 16 extant genera and 101 species widespread through major Neotropical rivers with relevant importance for regional fisheries and aquaculture. The monophyly of Serrasalmidae and the recognition of three main clades are recurrent between morphological and molecular phylogenies. However, both intergeneric and interspecific relationships within each of those clades remain uncertain. Here, we used 81 terminals of 69 species (68%) and all 16 genera of Serrasalmidae to sequence 1553 loci of ultraconserved elements (UCEs), multiple nuclear loci widely applied in phylogenetic studies, and performed maximum likelihood, Bayesian, and species tree analyses. We obtained highly supported phylogenies in all applied methods corroborating the monophyly of Serrasalmidae and the three-clade hypotheses herein proposed as two subfamilies and two tribes: (Colossomatinae (Serrasalminae (Myleini + Serrasalmini))). Morphological features for each subfamily involve the absence (Colossomatinae) or presence (Serrasalminae) of a pre-dorsal spine. Morphological diagnoses among tribes include the pre-dorsal spine being continuous (Myleini) or discontinuous (Serrasalmini) relative to the first unbranched dorsal-fin ray. Our results highlight the complexity of the relationships especially the non-monophyly of Myleus, Mylesinus, Myloplus, Tometes, and Utiaritichthys within Myleini, as well as of Serrasalmus and Pristobrycon within Serrasalmini.

Mitochondrial genomes of four American characins and phylogenetic relationships within the family Characidae (Teleostei: Characiformes).

Mitochondrial genome sequencing has become widely used in numerous fields, including systematics, phylogeny, and evolutionary genomics. To elucidate phylogenetic relationships among members of the family Characidae, we sequenced the mitogenomes of four species within this family, namely, Aphyocharax rathbuni, Hyphessobrycon herbertaxelrodi, Hyphessobrycon megalopterus, and Prionobrama filigera. The mitogenomes were found to be 16,678-16,841 bp and encode 37 typical mitochondrial genes (13 protein-coding, 2 ribosomal RNA, and 22 transfer RNA genes). Gene arrangements in the studied species are consistent with those in the inferred ancestral fish. Most protein-coding genes in these mitogenomes have typical ATN start codons and TAR or an incomplete stop codon T-. Phylogenetic relationships based on Bayesian inference and maximum-likelihood methods indicated that A. rathbuni, H. herbertaxelrodi, H. megalopterus, and P. filigera belong to the Characidae family. Of the 15 Characidae species studied, three pairs were of the same genus, but the results for only one pair were well supported. This phylogenetic classification is inconsistent with those described in previous morphological and taxonomic studies on this family. Thus, systematic classification of the Characidae requires further examination. Our findings yield new mitogenomic data that will provide a basis for future phylogenetic and taxonomic studies.

The true identity of Bryconops cyrtogaster (Norman), and description of a new species of Bryconops Kner (Characiformes: Iguanodectidae) from the Rio Jari, lower Amazon basin.

Bryconops cyrtogaster, a poorly known species endemic from the Oyapock River at the border between French Guyana and Brazil, is redescribed herein based on examination of available type material, as well as newly collected material. Additionally, a new rheophilic species from the rio Jari rapids, lower Amazon basin, Brazil, is described. The two species belong to the subgenus Creatochanes and are unique among the congeneres for possessing a posteriorly positioned humeral blotch at the level of the sixth and seventh lateral line scales. They differ from each other by meristic and morphometric characters. The list of endemic species in the rio Jari basin is revised.

Localization and distribution of CCK-8, NPY, Leu-ENK-, and Ghrelin- in the digestive tract of Prochilodus lineatus (Valenciennes, 1836).

This study describes the histological characteristics and distribution of gastrointestinal tract endocrine cells (ECs) of Prochilodus lineatus (detritivorous fish) using immunohistochemical procedures. The digestive tract of P. lineatus was divided into seven portions: stomach (cardial and pyloric), pyloric caeca, and intestine (anterior, glandular, middle and posterior). A pool of specific antisera against cholecystokinin (CCK-8), -neuropeptide Y (NPY), -ghrelin (Ghre) and -leu-enkephalin (Leu-ENK) to identify ECs were used. According to the morphological characteristics of ECs, two different types were identified and classified as open or closed-type. The number of ECs varied throughout the gastrointestinal tract, though a high abundance was found in the anterior intestine and pyloric caeca. A large number of ECs immunoreactive to CCK-8 and NPY were recorded in the anterior, glandular and middle intestine. ECs immunopositive to Leu-ENK were distributed in the stomach and pyloric caeca. For Ghre, immunopositive ECs were restricted to the glandular intestine. The results of the present study indicate that P. lineatus presents an ECs distribution pattern with species-specific particularities. However, CCK showed a distribution similar to that of omnivores, which is possibly related to local signaling functions in order to achieve the correct digestion of the various organisms found in the detritus.