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This study evaluated the growth and physiological response of proactive and reactive Colossoma macropomum juveniles in a recirculating aquaculture system (RAS). In Phase 1 of the experiment (50 days of cultivation), juveniles, weighing 2.16 ± 0.52 g, were stocked in 12 28-L tanks to test the following treatments: proactive (PT), reactive (RT) and mixed (MT) composed of reactive (MRT) and proactive (MPT) animals. In Phase 2 of the experiment (40 days of cultivation), the animals were transferred to 175-L tanks with the same treatments as Phase 1. The animals were fed twice a day with commercial diet during both phases. After Phase 1, MPT animals showed higher growth than MRT animals (P < 0.05), and higher weight gain and daily weight than PT animals (P < 0.05). After Phase 2, PT animals showed higher weight gain and daily weight gain than RT and MT animals (P < 0.05), as did MPT animals compared to PT animals. Performance for RT animals was superior (P < 0.05) to that of MRT animals. Glucose (P < 0.04) and cholesterol (P < 0.01) were higher for RT animals compared to PT animals. Cholesterol was higher for MPT animals compared to MRT animals (P < 0.01), while plasma protein was lower (P < 0.001). Glucose (P < 0.001) and cholesterol (P < 0.01) were higher for MPT animals compared to PT animals and for MRT animals compared to RT animals (glucose P < 0.02, cholesterol P < 0.01). After 90 days of cultivation, proactive animals cultivated separately presented better performance. When cultivated together, reactive animals experienced a decrease in performance and both stress coping styles showed more signs of stress.

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Phylogenetic analyses of mitochondrial and nuclear data of 31 specimens of Cyphocharax from trans-Andean rivers support the presence of one lineage of Cyphocharax aspilos in Lago Maracaibo and three cryptic lineages of Cyphocharax magdalenae: (1) Cauca-Magdalena and Ranchería, (2) León and Atrato, and (3) Chucunaque-Tuira, Santa María, and Chiriquí basins of Central America. Results suggest that the Serranía del Perijá facilitated Late Miocene cladogenetic events, whereas post-Isthmian C. magdalenae expansion was enabled by gene flow across the lower Magdalena valley and Central American lowlands. Time-calibrated phylogenetics indicate that the C. magdalenae colonized lower Central America in the Pliocene (3.7 MYA; Ma), the divergence Atrato-Magdalena occurred in Late Pliocene (3.0 Ma) and the split Ranchería-Magdalena during the Middle Pleistocene (1.3 Ma). Updated geographic distribution data support the hypothesis that the Cordillera de Talamanca functions as a barrier to northward expansion of C. magdalenae in Central America.

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Constraints on phenotypic evolution can lead to patterns of convergent evolution, by limiting the 'pool' of potential phenotypes in the face of endogenous (functional, developmental) or exogenous (competition, predation) selective pressures. Evaluation of convergence depends on integrating ecological and morphological data within a robust, comparative phylogenetic context. The staggering diversity of teleost fishes offers a multitude of lineages adapted for similar ecological roles, and therefore, offers numerous replicated evolutionary experiments for exploring phenotypic convergence. However, our understanding of fish feeding systems has been primarily shaped by marine species, with the monolithic exception of freshwater cichlids. Here we use piranhas and pacus (Serrasalmidae) to explore the evolution of different feeding ecologies and their morphological proxies in Neotropical freshwater environments. Specifically, we explore whether convergence is more widespread among plant-eating fishes, arising from strong constraints on phenotypic evolution in herbivores. Using osteological micro-computed tomographic imaging (µCT), we describe the major axes of morphological variation in pacus and piranhas, regarding their diet and feeding behaviors. Next, we evaluated whether herbivorous niches are less labile than other dietary guilds and whether herbivorous species' phenotypes evolve at a slower evolutionary rate than other taxa. We then assess how convergent herbivorous taxa are, using three different suites of morphological characters (dental, jaw, and abdominal morphometrics). Ecologically, herbivory is not a dead end, exhibiting similar observed transition rates as those between carnivores and omnivores. However, we documented widespread convergence in herbivores and that herbivores have slower rates of phenotypic evolution than carnivores. Most instances of convergence are found in herbivorous taxa, specifically in frugivores and folivores. Moreover, instances of 'complete' convergence, indicated by positive convergence metrics observed in more than one morphometric dataset, were only found in herbivores. Herbivores do appear to evolve under constrained circumstances, but this has not limited their ecological lability.

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The first studies on ion regulation in fish exposed to low pH, which were inspired by the Acid Rain environmental crisis, seemed to indicate that ion transport at the gills was completely and irreversibly inhibited at pH 4.0-4.5 and below. However, work on characid fish native to the Rio Negro, a naturally acidic, blackwater tributary of the Amazon River, found that they possess ion transport mechanisms that are completely insensitive to pHs as low as 3.25. As more species were examined it appeared that pH-insensitive transport was a trait shared by many, if not most, species in the Order Characiformes. Subsequently, a few other species of fish have been shown to be able to transport ions at low pH, in particular zebrafish (Danio rerio), which show rapid recovery of Na+ uptake at pH 4.0 after initial inhibition. Measurements of rates of Na+ transport during exposure to pharmacological agents that inhibit various transport proteins suggested that characiform fish do not utilize the generally accepted mechanisms for Na+ transport that rely on some form of H+ extrusion. Examination of zebrafish transport at low pH suggest the rapid recovery may be due to a novel Na+/K+ exchanger, but after longer term exposure they may rely on a coupling of Na+/H+ exchangers and NH3 excretion. Further work is needed to clarify these mechanisms of transport and to find other acid-tolerant species to fully gain an appreciation of the diversity of physiological mechansisms involved.

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The Rio Negro basin of Amazonia (Brazil) is a hotspot of fish biodiversity that is under threat from copper (Cu) pollution. The very ion-poor blackwaters have a high dissolved organic carbon (DOC) concentration. We investigated the Cu sensitivity of nine Amazonian fish species in their natural blackwaters (Rio Negro). The acute lethal concentration of Cu (96 h LC50) was determined at different dilutions of Rio Negro water (RNW) in ion-poor well water (IPW), ranging from 0 to 100%. The IPW was similar to RNW in pH and ionic composition but deficient in DOC, allowing this parameter to vary 20-fold from 0.4 to 8.3 mg/L in tests. The Biotic Ligand Model (BLM; Windward version 3.41.2.45) was used to model Cu speciation and toxicity over the range of tested water compositions, and to estimate lethal Cu accumulations on the gills (LA50). The modeling predicted a high relative abundance of Cu complexes with DOC in test waters. As these complexes became more abundant with increasing RNW content, a concomitant decrease in free Cu2+ was observed. In agreement with this modeling, acute Cu toxicity decreased (i.e. 96 h LC50 values increase) with increasing RNW content. The three most sensitive species (Hemigrammus rhodostomus, Carnegiella strigatta and Hyphessobrycon socolofi) were Characiformes, whereas Corydoras schwartzi (Siluriformes) and Apistogramma agassizii (Cichliformes) were the most tolerant. These sensitivity differences were reflected in the BLM-predicted lethal gill copper accumulation (LA50), which were generally lower in Characiformes than in Cichliformes. Using these newly estimated LA50 values in the BLM allowed for accurate prediction of acute Cu toxicity in the nine Amazonian fish. Our data emphasize that the BLM approach is a promising tool for assessing Cu risk to Amazonian fish species in blackwater conditions characterized by very low concentrations of major ions but high concentrations of DOC.

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Primordial germ cells (PGCs) are embryonic pluripotent cells that can differentiate into spermatogonia and oogonia, and therefore, PGCs are a genetic source for germplasm conservation through cryobanking and the generation of germline chimeras. The knowledge of PGC migration routes is essential for transplantation studies. In this work, the mRNA synthesized from the ddx4 3'UTR sequence of Pseudopimelodus mangurus, in fusion with gfp or dsred, was microinjected into zygotes of three neotropical species (P. mangurus, Astyanax altiparanae, and Prochilodus lineatus) for PGC labeling. Visualization of labeled PGCs was achieved by fluorescence microscopy during embryonic development. In addition, ddx4 and dnd1 expressions were evaluated during embryonic development, larvae, and adult tissues of P. mangurus, to validate their use as a PGC marker. As a result, the effective identification of presumptive PGCs was obtained. DsRed-positive PGC of P. mangurus was observed in the hatching stage, GFP-positive PGC of A. altiparanae in the gastrula stage, and GFP-positive PGCs from P. lineatus were identified at the segmentation stage, with representative labeling percentages of 29% and 16% in A. altiparanae and P. lineatus, respectively. The expression of ddx4 and dnd1 of P. mangurus confirmed the specificity of these genes in germ cells. These results point to the functionality of the P. mangurus ddx4 3'UTR sequence as a PGC marker, demonstrating that PGC labeling was more efficient in A. altiparanae and P. lineatus. The procedures used to identify PGCs in P. mangurus consolidate the first step for generating germinal chimeras as a conservation action of P. mangurus.

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Characiformes is a diverse and evolutionarily significant order of freshwater fish encompassing over 2,300 species. Despite its diversity, our understanding of Characiformes' evolutionary relationships and adaptive mechanisms is limited due to insufficient genome sequences. In this study, we sequenced and assembled the genomes of four Characiformes species, three of which were chromosome-level assemblies. Our analyses revealed dynamic changes in gene family evolution, repeat sequences and variations in chromosomal collinearity within these genomes. With the assembled genomes, we were not only able to elucidate the evolutionary relationship of the four main orders in Otophysi but also indicated Characiformes as the paraphyletic group. Comparative genomic analysis with other available fish genomes shed light on the evolution of genes related to tooth development in Characiformes. Notably, variations in the copy number of secretory calcium-binding phosphoproteins (SCPP) genes were observed among different orders of Otophysi, indicating their potential contribution to the diversity of tooth types. Our study offers invaluable genome sequences and novel insights into Characiformes' evolution, paving the way for further genomic and evolutionary research in fish.

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Animal welfare is a key issue not only for aquaculture industry and food production, but also for daily husbandry practices in research topics related to physiology in wild and farmed animals. In this context, teleost fish constitute interesting models to assess alternative welfare indicators because of their wide diversity in reproductive and social structures. Any framework for assessing teleost fish welfare needs to account for the physiological mechanisms involved in each species as a first step. A comprehensive approach should also take into account how these physiological and behavioral parameters can be altered by environmental enrichment considering the specific requirements in each case and identifying intrinsic biological characteristics of individual species. This review will show how cortisol and sex steroids regulate social behavior in teleost fish, and how different aspects of social behavior can be employed as welfare indicators according to specific characteristics in each case. This article will consider evidence in teleost fish, including cichlids, characids and cyprinids with different reproductive strategies and social structures (e.g., territorial social hierarchies or shoaling behavior). Neotropical species will be particularly emphasized. The main laboratory-based animal welfare indicators are cortisol, a classical stress hormone, together with sex steroids. Considering that the endocrine landscape is intrinsically related to social behavior, reproductive and agonistic behavioral traits such as aggression, anxiety and courtship are key elements to assess welfare under housing and culture conditions. This review highlights the importance of assessing physiological mechanisms and identifying behavioral characteristics in teleost fish, especially in Neotropical species, as a baseline to understand which environmental enrichment can improve animal welfare in each individual species.

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The Neotropical region bears the most diverse freshwater fish fauna on the planet and is the stage for dramatic conservation struggles. Initiatives aiming for conservation of a single emblematic fish, a flagship species, to which different onlookers relate on a cultural/personal level, holds promise towards engagement and conservation actions benefiting whole biological communities and ecosystems. Here, we present the first comprehensive genomic resources for Salminus brasiliensis, a potential flagship Neotropical species. This fish faces pressing conservation issues, as well as taxonomic uncertainty, being a main species relevant to angling and commercial fisheries. We make available 178 million Illumina paired-end reads, 90 bases long, comprising 16 Gb (≈15X coverage) of filtered data, obtained from a primary genomic library of 500-bp fragments. We present the first de novo genomic assembly for S. brasiliensis, with ∼1 Gb (N 50 = 10,889), as well as the coding genome annotation of 12,962 putative genes from assembled genomic fragments over 10 kb, most of which could be identified from the Ostariophysi GenBank database. We also provide a genome-wide panel for more than 80,000 predicted microsatellite loci for low-cost, fast and abundant DNA marker development for this species. A total of 47, among 52 candidates, empirically assayed microsatellites were confirmed as polymorphic in this fish. All genomic data produced for S. brasiliensis is hereby made publicly accessible. With the disclosure of these results, we intend to foster general biology studies and to provide tools to be applied immediately in conservation and aquaculture in this candidate flagship Neotropical species.

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Characinae is one of the most species-rich subfamilies of Characidae and holds special taxonomic importance because it includes Charax, type-genus of Characidae and Characiformes. Currently, the monophyly and the hypotheses of intergeneric and interspecific relationships of Characinae are based on a few morphological and molecular studies but all with low species coverage. Given their diversity, taxonomic importance, and the lack of a taxon-dense phylogeny, we sought to buttress the systematic understanding of Characinae collecting DNA sequence data from ultraconserved elements (UCEs) of the genome from 98 specimens covering 57 species (61%) plus 17 characiforms as outgroups. We used maximum likelihood, Bayesian inference, and coalescent-based species tree approaches and the resulting phylogeny with 1,300 UCE loci (586,785 characters) reinforced the monophyly of the subfamily as well as of six genera: Acestrocephalus, Charax, Cynopotamus, Galeocharax, Phenacogaster, and Roeboides. The phylogeny provides a hypothesis of intergeneric and interspecific relationships for the subfamily with Phenacogaster sister to all genera, and Acanthocharax sister to Cynopotamini (Cynopotamus (Acestrocephalus Galeocharax)) and Characini (Charax Roeboides). We propose a new tribe Acanthocharacini to allocate Acanthocharax, two subclades for Phenacogaster, two for Cynopotamus, three for Charax, and reinforced the four subclades for Roeboides previously identified by morphological studies. Additionally, we generated a time-calibrated phylogeny for Characinae that suggested an initial diversification during the Miocene at around 19 million years ago and discussed historical biogeographic events for major subclades. The results obtained here will contribute to the development of further research on the evolutionary processes modulating species diversification in Characinae.

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The present study evaluated neurotoxic, biotransformation, genotoxic and antioxidant responses to relevant environmental concentrations of diclofenac (0.4 µg L-1) and caffeine (27.5 µg L-1), separate and combined, in adult males of the freshwater fish Astyanax altiparanae after a subchronic exposure (14 days). Fish exposed to diclofenac and caffeine, both separate and combined, revealed a neurotoxic effect through the inhibition of acetylcholinesterase activity in the muscle, while diclofenac alone and in combination caused cyclooxygenase inhibition. Caffeine alone produces genotoxicity on this species but, when combined with diclofenac, it potentiates hepatic lipoperoxidation and the inhibition of oxidative stress enzymes, while diclofenac alone or in combination produces a general inhibition of important enzymes. This study suggests that aquatic contamination produced by these pharmaceuticals has the potential to affect homeostasis and locomotion in A. altiparanae and compromise their immune system and general health.

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Crustaceans of the subclass Copepoda are an important component of the invertebrate aquatic fauna. They occur in all aquatic environments and include some representatives that are free-living organisms and others that have a parasitic lifestyle. The genus Therodamas comprises marine and freshwater copepods whose females are parasites of fish in their adult phase, with only seven species described so far. During a field survey of fish parasites in the Jari River, a large tributary of the Amazon River system, in Brazil, we found a new species of the genus Therodamas infecting Leporinus fasciatus. Therodamas longicollum n. sp. is the second strictly freshwater species known. Phylogenetic analysis showed that the new species is grouped in the family Ergasilidae, and divergence estimates showed that T. longicollum n. sp. diverged from its ancestor at around 66.34 Ma, in the late Upper Cretaceous. Therodamas longicollum n. sp. differs from its congeneric in that it does not have lobes and/or expansion of the anterior neck region. Besides describing a new Therodamas species, thereby increasing the diversity of the genus to eight species, this study points out the existence of a lineage of these copepods that has adapted to the freshwater environment of the Amazon. This study also corroborates the genus Therodamas as part of the family Ergasilidae.

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The present study describes Henneguya tietensis n. sp. parasitizing Prochilodus lineatus (Valenciennes, 1837) from Brazil, through morphological and molecular analysis. Myxospores were found in the gill tissue of P. lineatus with a prevalence of 10%. The myxospores consisted of two elongate, elliptical shell valves each with a long, tapering caudal appendage. The morphology was consistent with Henneguya and the myxospore measured as follows (mean ± SD): total length 55.5 ± 2.1 µm, body length 16.2 ± 1.1 µm, body width 5.5 ± 0.1 µm, caudal appendages length 39.0 ± 2.0 µm. The polar capsules were 7.3 ± 0.2 µm long, 1.7 ± 0.2 µm wide and contained a polar filament coiled 11 to 13 turns. Histological analysis showed the plasmodia developing in the middle region of each gill filament and caused a focally extensive distension by forming a space-occupying mass within the gill filament. The ultrastructural analysis allowed the observation of the presence of mature myxospores throughout the plasmodium. Phylogenetic analysis with the SSU rDNA gene showed Henneguya tietensis n. sp. as a sister species of the subclade formed by Henneguya piaractus and Henneguya brachypomus that infect fishes of the genus Piaractus. The genetically closest species was H. piaractus, which showed a similarity of 82.4%. Using molecular and morphological characterization, the myxozoan parasite described herein represents a new species of the genus Henneguya.

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In view of the established climate change scenario and the consequent changes in global temperature, it is essential to study its effects on animal spermatogenesis. Therefore, the aim of this study was to verify the duration of spermatogenesis at different temperatures. For this purpose, 96 male and adult specimens of Astyanax altiparanae were kept in a closed circulation system with water temperature stabilized at 27 °C and 32 °C. Subsequently, the specimens received pulses of BrdU (bromodeoxyuridine) at a concentration of 100 mg/kg/day for 2 consecutive days, and the samples were collected daily for a period of 15 days. Their testes were removed, fixed, processed in historesin, and sectioned in 3 µm, submitted to hematoxylin/eosin staining and to bromodeoxyuridine immunodetection. Partial results of the optimum temperature experiments allowed the classification of A. altiparanae spermatogenic cells in Aund, Adiff, and type B spermatogonia, spermatocytes, spermatids, and spermatozoa. The duration of spermatogenesis was determined as approximately 6 days for animals at a temperature of 27 °C and 1 day for animals at 32 °C. The elevated temperature was also responsible for increasing cell proliferation, resulting in an increase in the number of spermatocytes, spermatids, spermatozoa, and cell death (cell pyknotic). The duration of spermatogenesis in A. altiparanae was directly affected by the elevated water temperature, causing a reduction in the estimated time of spermatogenesis.

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Different studies suggest some social calls could be used in fish identification if their specificity is unambiguously assessed. Sounds of different populations of piranhas Serrasalmus maculatus Kner, 1858 were recorded to determine their homogeneity between rivers inside a single basin (Araguari and Grande River, upper Paraná River basin) and between separated basins (Amazon and Paraná basins). All fish from the different populations produced sounds with similar acoustic features. Consequently, the populations were not discernible based on individual sound characteristics. This high homogeneity between sounds from different populations indicates their usefulness for conservation projects using passive acoustic monitoring in piranhas. Moreover, it supports the use of acoustic features as complementary key characteristics in taxonomic studies.

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Abstract: Based on the information and evidence on the zoological expedition led by the senior Austrian zoologist Franz Steindachner in 1903, the examination of original species descriptions using material collected in this expedition, the scientific literature on this expedition, examination of specimens of Knodus victoriae, and the consultation of State decrees modifying the name of localities and municipalities, we propose herein updated and accurate type localities for Knodus victoriae and Loricaria parnahybae. The accurate type locality for both species is at the mouth of a stream that flows into the Parnaiba River, Alto Parnaíba municipality in the State of Maranhão, upper Parnaiba River basin, northeastern Brazil. This update in the type locality aims to avoid doubts and confusion that hinder biogeographic and taxonomic studies.

Resumo: Com base em informações e evidências da expedição zoológica liderada pelo zoólogo sênior austríaco Franz Steindachner em 1903, exame das descrições originais das espécies que utilizaram material coletado nessa expedição, literatura científica sobre essa expedição, exame de exemplares de Knodus victoriae, e consulta a decretos estaduais que modificaram o nome de localidades e municípios, nós propomos a atualização com acurácia das localidades tipos de Knodus victoriae e Loricaria parnahybae. A localidade tipo acurada para ambas as espécies é na foz de um riacho que desagua no rio Parnaíba, no município de Alto Parnaíba no Estado do Maranhão, bacia do alto rio Parnaíba, nordeste do Brasil. Esta atualização na localidade tipo visa evitar dúvidas e confusões que podem atrapalhar estudos biogeográficos e taxonômicos.

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In this study, the complete mitochondrial genome of Hyphessobrycon herbertaxelrodi is presented, and we also discussed its mitochondrial characteristics. The full length of the mitochondrial genome was 17,417 bp, including 13 protein coding genes (PCGs), 2 ribosomal RNAs (12S and 16S), 22 transfer RNA genes, 1 non-coding control region (D-loop), and 1 origin of replication on the light-strand. The total nucleotide composition of mitochondrial DNA was 29.76%A, 29.88%T, 25.35%C, 15.01%G, and AT was 59.64%. The phylogenetic tree suggested that H. herbertaxelrodi shared the most recent common ancestor with Astyanax giton, Grundulus bogotensis, Astyanax paranae, and Oligosarcus argenteus.

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A new species of Characidium is described from headwater tributaries of the upper rio Guaporé, Rio Madeira basin, Mato Grosso state, Brazil. The new species can be diagnosed from all congeners, except Characidium summus, for lacking the preorbital and postorbital stripes. It can be diagnosed from the latter species by having 12 circumpeduncular scales (vs. 14), by an adipose fin present (vs. adipose fin absent), and 10-12 bars along the body (vs. absence of bars), among other characters. The new species is so far only known from direct tributaries of the Rio Guaporé at the Serra da Borda, an isolated plateau 300-800 meters above sea level, and possess a behavior very uncommon within the genus, being a mid-water pelagic fish, instead of the benthic behavior of most congeners. Comments on the putative phylogenetic relationships of the new species, as well as some remarks on its unusual behavior within the genus, are presented.

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The presence of intermuscular bones in fisheries products limits the consumption and commercialization potential of many fish species, including tambaqui (Colossoma macropomum). These bones have caused medical emergencies and are an undesirable characteristic for fish farming because their removal is labor-intensive during fish processing. Despite the difficulty in identifying genes related to the lack of intermuscular bone in diverse species of fish, the discovery of individuals lacking intermuscular bones in a Neotropical freshwater characiform fish has provided a unique opportunity to delve into the genetic mechanisms underlying the pathways of intermuscular bone formation. In this study, we carried out a GWAS among boneless and wt tambaqui populations to identify markers associated with a lack of intermuscular bone. After analyzing 11 416 SNPs in 360 individuals (12 boneless and 348 bony), we report 675 significant (Padj  < 0.003) associations for this trait. Of those, 13 associations were located near candidate genes related to the reduction of bone mass, promotion of bone formation, inhibition of bone resorption, central control of bone remodeling, bone mineralization and other related functions. To the best of our knowledge, for the first time, we have successfully identified genes related to a lack of intermuscular bones using GWAS in a non-model species.

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Integrative taxonomy was used to evaluate two component populations of Procamallanus (Spirocamallanus) inopinatus in Brazil and the phylogeny Camallanidae. Parasite populations were collected in the characiform Anostomoides passionis from River Xingu (Amazon basin) and Megaleporinus elongatus from River Miranda (Paraguay basin). Morphology was analysed using light and scanning electron microscopy (SEM). Genetic characterization was based on partial sequences of the 18S and 28S rDNA, and COI mtDNA. Phylogenies were based on 18S and COI due to data availability. Generalized Mixed Yule Coalescent (GMYC), Poisson Tree Process (PTP) and *BEAST were used for species delimitation and validation. SEM revealed for the first time the presence of minute denticles and pore-like structures surrounding the oral opening, phasmids in females and confirmed other important morphological aspects. Statistical comparison between the two-component populations indicated morphometric variations, especially among males. The different component population of P. (S.) inopinatus showed variable morphometry, but uniform morphology and were validated as conspecific by the GMYC, PTP and *BEAST. Some camallanid sequences in GenBank have incorrect taxonomic labelling. Host, environment and geographic aspects seem to be related to some lineages within Camallanidae; however, their real phylogenetic meanings are still unclear.

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