Bioaccumulation, genotoxicity, and risks to native fish species from inorganic contaminants in the Pantanal Sul-Mato-Grossense, Brazil.

The Aquidauana River is one of the most important rivers in the Pantanal region, Brazil. However, its waters have been contaminated by nearby anthropogenic activities, threatening native fish species. In this study, our objectives were: 1) to determine the concentrations of Al, As, Cd, Cu, Fe, Mn, Ni, Pb, and Zn in water and sediment samples from the Aquidauana River and to assess the risks posed to aquatic biota; 2) to quantify the concentration of these elements in muscle and liver tissue samples from four native fish species; 3) to evaluate the potential bioaccumulation of inorganic elements in the muscles and liver; and 4) to investigate genotoxicity biomarkers and their association with the inorganic element concentrations present in the muscle tissue. Water and fish samples were collected in November 2020. The concentrations of Al, As, Cd, Cu, Fe, and Pb in the water samples were in disagreement with the Brazilian legislation and presented risks to the aquatic biota. In terms of mixtures of inorganic elements, there was a great increase in the risk to biota. The As concentration did not meet the Brazilian standard for sediments in the sample collected at sampling site 6. The concentrations of Cd and Pb in the muscle tissue of Hypostomus regani, Prochilodus lineatus, Brycon hilarii, and Mylossoma duriventre exceeded the Brazilian standards for human consumption. H. regani showed greater genotoxic damage, and the higher the Al and Fe concentrations in the muscle tissue, the higher the frequencies of lobulated nuclei and nuclear invaginations. Together, our results demonstrate the negative impacts on native fish species from the Aquidauana River contamination and indicate risks to Pantanal biodiversity.

The spawning method affects the reproductive efficiency of piracanjuba fish (Brycon orbignyanus) in restocking programs / El método de desove afecta la eficiencia reproductiva de piracanjuba (Brycon orbignyanus) en programas de repoblación / O método de desova afeta a eficiência reprodutiva de piracanjuba (Brycon orbignyanus) em programas de repovoamento

Abstract Background: Preserving the genetic diversity of wild fish is an important consideration for restocking programs, as inbreeding can compromise progeny survival as well as impact the resilience of natural populations. Objective: To evaluate the influence of spawning method: semi-natural (SN) or strip-spawning (ST), and the number of breeders (1♀:3♂ and 2♀:6♂) on the reproductive efficiency and genetic diversity of B. orbignyanus progeny destined for restoration of wild stocks. Methods: Rates of fertilization, hatching and broodfish mortality were recorded. For genetic evaluations (allele frequency, observed and expected heterozygosity, Shannon index, inbreeding coefficient, molecular variance analysis, and genetic differentiation), breeders (n=24), and their progenies (90 larvae/treatment) were sampled and analyzed using eight microsatellite markers. Results: Higher fertilization and hatching rates, and lower broodfish mortality were observed for the SN method (p<0.05), whereas the number of breeders did not affect these parameters (p>0.05). Interaction between spawning method and number of breeders was not significant (p>0.05). The amplified microsatellite loci produced a total of 30 alleles, with sizes between 80 and 225 bp and their frequencies indicated an increase (p<0.05) of genetic diversity in the progenies, but low genetic differentiation between treatments (p>0.05). Conclusion: The spawning methods and number of breeders tested increased equally the genetic diversity of the progeny, with low genetic differentiation between treatments. In contrast, rates of fertilization, hatching and brood fish mortality revealed that the SN method resulted in the best reproductive efficiency due to the handling stress and injuries caused by ST. Thus, SN proves to be the most suitable spawning-method for B. orbignyanus in restocking programs.

Resumen Antecedentes: Mantener la diversidad genética de los peces salvajes es una consideración importante para los programas de repoblación, ya que la endogamia puede comprometer la supervivencia de la progenie y afectar la supervivencia de las poblaciones naturales. Objetivo: Evaluar la influencia del método de desove (seminatural - SN o en franjas - ST) y el número de reproductores (1♀:3♂ y 2♀:6♂) sobre la eficiencia reproductiva y la diversidad genética de progenies de B. orbignyanus destinados a la restauración de poblaciones silvestres. Métodos: Se monitorearon las tasas de fertilización, eclosión y mortalidad de reproductores. Para las evaluaciones genéticas (frecuencias alélicas, heterocigosidad observada y esperada, índice de Shannon, coeficiente de endogamia, análisis de varianza molecular y diferenciación genética) los reproductores (n=24) y su progenie (90 larvas/tratamiento) se muestrearon y analizaron utilizando ocho marcadores microsatélites. Resultados: La interacción entre el método de desove y el número de reproductores no fue significativa (p>0,05). Se obtuvieron mejores tasas de fecundación y eclosión (p<0,05), y una menor mortalidad de reproductores (p<0,05) con el método SN, mientras que el número de reproductores no tuvo efecto (p>0,05). Los loci de microsatélites amplificados produjeron un total de 30 alelos con tamaños entre 80 y 225 pb, y sus frecuencias indicaron un aumento (p<0,05) en la diversidad genética de las progenies, pero una baja diferenciación genética entre tratamientos (p>0,05). Conclusión: Los métodos de desove y el número de reproductores evaluados aumentaron de la misma manera la diversidad genética de las progenies, con baja diferenciación genética entre tratamientos. En contraste, las tasas de fecundación, eclosión y mortalidad de peces reproductores revelaron que el SN tuvo la mejor eficiencia reproductiva, un hecho relacionado con el estrés del manejo y las lesiones causadas por el ST. Por lo tanto, el SN demuestra ser el método de desove más adecuado para B. orbignyanus en los programas de repoblación.

Resumo Antecedentes: A manutenção da diversidade genética dos peixes selvagens é uma importante consideração para programas de repovoamento, já que a endogamia pode comprometer a sobrevivência da progênie, além de impactar na resiliência das populações naturais. Objetivo: Avaliar a influência do método de desova (semi-natural - SN ou por extrusão - ST) e número de reprodutores (1♀:3♂ e 2♀:6♂) na eficiência reprodutiva e na diversidade genética de progênie de B. orbignyanus destinados à recuperação de estoques selvagens. Métodos: Taxas de fertilização, eclosão e mortalidade de reprodutores foram monitorados. Para avaliações genéticas (frequências alélicas, heterozigosidade observada e esperada, índice de Shannon, coeficiente de endogamia, análise de variância molecular e diferenciação genética), reprodutores (n=24) e sua progênie (90 larvas/tratamento) foram amostrados e analisados utilizando oito marcadores microssatélites. Resultados: Interação entre método de desova e número de reprodutores não foi significante (p>0,05). Melhores taxas de fertilização e eclosão (p<0,05), e menor (p<0,05) mortalidade de reprodutores foram observados para o método SN, enquanto que o número de reprodutores não afetou esses parâmetros (p>0,05). Os loci microssatélites amplificados produziram um total de 30 alelos, com tamanhos entre 80 e 225 pb e suas frequências indicaram aumento (p<0,05) da diversidade genética nas progênies, mas baixa diferenciação genética entre os tratamentos (p>0,05). Conclusão: Os métodos de desova e números de reprodutores avaliados aumentaram igualmente a diversidade genética das progênies, com baixa diferenciação genética entre tratamentos. Em contraste, as taxas de fecundação, eclosão e mortalidade de reprodutores revelaram que SN obteve a melhor eficiência reprodutiva, fato relacionado com o estresse de manipulação e injurias causadas por ST. Por isso, SN se mostrou como o método de desova mais adequado para B. orbignyanus em programas de repovoamento.

Increasing the known biodiversity of cnidarian parasites of bryconid fishes from South America: two novel Myxobolus species with ultrastructure and ssrDNA-based phylogeny.

This study increases the known biodiversity of cnidarian parasites in neotropical bryconid fishes. Two novel Myxobolus species are described based on morphology, ultrastructure and small subunit ribosomal DNA (ssrDNA) sequencing: Myxobolus vetuschicanus n. sp. infecting fins of Salminus franciscanus and Myxobolus mineirus n. sp. infecting the mesentery of Brycon orthotaenia from the São Francisco River basin, Minas Gerais State, Brazil. Ultrastructural analysis of the two species revealed an asynchronous sporogenesis process, with germinative cells and young developmental stages of myxospores in the periphery of the plasmodia. In M. vetuschicanus n. sp., the plasmodia were surrounded by a layer of fibroblasts and in M. mineirus n. sp., the plasmodial membrane had direct contact with the host tissue. The phylogenetic analysis based on the ssrDNA of Henneguya/Myxobolus species showed that the two novel Myxobolus species grouped in subclades together with other parasite species of bryconid fishes.

Taxonomy, phylogeny and host-parasite interaction of two novel Myxobolus species infecting Brycon orthotaenia from the São Francisco River, Brazil.

Two new Myxobolus species were described infecting Brycon orthotaenia from the São Francisco River, in the state of Minas Gerais, Brazil. From a total of 39 B. orthotaenia collected, two specimens (5.1%) exhibited infection of the ovary and 12 specimens (30.8%) displayed infection of the liver. The plasmodia of both Myxobolus species were white and spherical measuring around 1 mm in length. The plasmodium found in the ovary showed mature myxospores, which were oval shaped from the frontal view and measured 9.2-11.0 (9.8 ± 0.4) µm in length, 5.9-6.9 (6.5 ± 0.3) µm in width and 4.6-5 (4.9 ± 0.1) µm in diameter. The two polar capsules were the same size and measured 3.9-6.2 (4.7 ± 0.5) µm in length and 1.8-2.4 (2.1 ± 0.2) µm in width. The polar tubules had 9 coils. The plasmodium found in the liver showed mature myxospores which were ellipsoidal in shape from the frontal view and measured 10.0-11.4 (10.7 ± 0.5) µm in length, 7.3-8.6 (8.1 ± 0.4) µm in width and 5.3-7.0 (6.8 ± 0.4) µm in diameter. The two polar capsules were the same size and measured 4.2-5.4 (4.9 ± 0.3) µm in length and 1.9-2.9 (2.7 ± 0.3) µm in width. The polar tubules had 8 coils. Ultrastructural analysis revealed an asynchronous sporogenesis process, with young developmental myxospore stages more often found in the periphery of the plasmodium and mature myxospores in the centre of the plasmodium. The plasmodial wall was formed by a single membrane which was not surrounded by a layer of host tissue. A thick layer of fibrous material was found in the peripheral ectoplasm close to the plasmodial wall of the plasmodium found in the ovary. Phylogenetic analysis based on the small-subunit ribosomal DNA - ssrDNA sequences and using the closest myxozoan sequences to each one of the species studied here based on previous GenBank data and Henneguya/Myxobolus/Thelohanellus species parasitizing fish from South American, revealed that the new species are grouped in a subclade together with other Myxobolus species parasitizing bryconid hosts.

Argulus elongatus (Branchiura, Argulidae) in fish in the upper São Francisco river, Brazil / Argulus elongatus (Branchiura, Argulidae) de peixes do alto rio São Francisco, Brasil

Abstract Among 164 fish from the upper São Francisco river, caught in the Três Marias reservoir (18º 12' 59" S; 45º 17' 34" W) or downstream from this reservoir (18º 12' 32" S; 45º 15' 41" W) in 2007, 2008, 2016 and 2017, four specimens of Argulus elongatus Heller, 1857 were found, one specimen per fish, in the following host species: Brycon orthotaenia Günther (two fish parasitized out of 38 examined) and Salminus hilarii Valenciennes (one fish parasitized out of 45 examined) (both in Bryconidae); and Metynnis lippincottianus (Cope) (one fish parasitized out of 81 examined) (Serrasalmidae). This opportunistic ectoparasite deserves attention even in the case of low infestation in fish in a natural water system. This is the first record of a branchiuran species in B.orthotaenia; the host list of A.elongatus is extended through addition of the three fish species examined in this study, and the known Brazilian geographical distribution of this argulid is enlarged to the São Francisco river basin.

Resumo De 164 peixes do alto rio São Francisco, capturados no Reservatório de Três Marias (18º 12' 59" S; 45º 17' 34" O) ou à jusante dele (18º 12' 32" S; 45º 15' 41" O), em 2007, 2008, 2016 e 2017, quatro espécimes de Argulus elongatus Heller, 1857 foram encontrados, sendo um branquiúro por peixe nas seguintes espécies de hospedeiros: Brycon orthotaenia Günther (dois peixes parasitados de 38 peixes examinados), Salminus hilarii Valenciennes (um peixe parasitado de 45 examinados) - ambos Bryconidae, e Metynnis lippincottianus (Cope) (um peixe parasitado de 81 examinados) - Serrasalmidae. Esse ectoparasito oportunista merece atenção, mesmo no caso de infestações baixas em peixes de ambientes hídricos naturais. Esse é o primeiro registro de Branchiura em B.orthotaenia. A lista de hospedeiros de A. elongatus é ampliada com a adição das três espécies de peixes examinadas neste estudo. A distribuição geográfica conhecida desse argulídeo no Brasil, é ampliada para a bacia do rio São Francisco.

Genetic diversity of Piracanjuba populations in fish stocking programs in the Tietê River, Brazil / Diversidad genética de poblaciones de Piracanjuba del programa de repoblamiento del Río Tietê, Brasil / Diversidade genética de populações de Piracanjuba do programa de repovoamento do Rio Tietê, Brasil

Abstract Background: Piracanjuba (Brycon orbignyanus) is a fish species highly affected by anthropogenic actions such as overfishing, water pollution, and hydroelectric developments. This species is currently considered in danger of extinction. Objective: To analyze the genetic diversity of a natural population (NP) and two captive broodstocks (SA and SB) of B. orbignyanus. Methods: Samples of caudal fins (NP: 24, SA: 30, and SB: 30) were collected. DNA was extracted and amplified for six RAPD primers and four microsatellite loci. Results: Sixty polymorphic fragments and 17 microsatellite alleles were detected. High intrapopulation heterozygosity (NP: 0.692, SA: 0.724, and SB: 0.686) was observed. Thirty-eight fragments and six alleles were shared among NP, SA, and SB. The FIS and Shannon's Index of diversity revealed a lack of inbreeding within groups. AMOVA analyses and FST indicated very high (NP vs SA and SB) and small (SA vs SB) genetic differentiation, confirmed by genetic distance and identity, number of migrants and a dendrogram, which revealed the formation of two genetic groups. Conclusions: The two marker types showed similar variability. The groups have adequate genetic variability, with high differentiation between NP and SA-SB, and similarity between broodstocks.

Resumen Antecedentes: Piracanjuba (Brycon orbignyanus) es una especie de pez fuertemente impactada por acciones antrópicas como sobrepesca, contaminación del agua y proyectos hidroeléctricos. Esta especie está considerada en peligro de extinción. Objetivo: Analizar la diversidad genética de una población natural (NP) y de dos lotes de reproductores (SA y SB) de B. orbignyanus en cautiverio. Métodos: Se colectaron 84 muestras de aleta caudal (NP: 24, SA: 30 y SB: 30). El ADN fue extraído y amplificado para seis cebadores RAPD y cuatro loci microsatélites. Resultados: Se obtuvieron 60 fragmentos polimórficos y 17 alelos microsatélites. Se observó alta heterocigosidad intra-poblacional (NP: 0,692; SA: 0,724 y SB: 0,686). Treinta y ocho fragmentos y seis alelos fueron compartidos entre NP, SA y SB. Los valores de FIS e índice de Shannon mostraron ausencia de endogamia entre los grupos. Los análisis de ANOVA y FST indicaron alta (NP vs SA y SB) y pequeña (SA vs SB) diferenciación genética; resultados confirmados por la distancia e identidad genética, número de migrantes y dendograma, evidenciando la formación de dos grupos genéticos. Conclusiones: Los grupos poseen adecuada variabilidad genética, con alta diferenciación entre NP vs SA-SB y similitud entre los lotes de reproductores.

Resumo Antecedentes: Piracanjuba (Brycon orbignyanus) é uma espécie peixe fortemente impactada por ações antrópicas como sobrepesca, poluição e construção de hidrelétricas. Atualmente, essa espécie engloba a lista de peixes que correm perigo de extinção. Objetivo: Analisar a diversidade genética de uma população natural (NP) e de dois estoques de reprodutores em cativeiro (SA e SB) de B. orbignyanus. Métodos: Foram coletadas amostras de nadadeira caudal de 84 indivíduos (NP: 24, SA: 30 e SB: 30). O DNA foi extraido e amplificado para seis primers RAPD e quatro loci microssatélites. Resultados: Foram obtidos 60 fragmentos polimórficos e 17 alelos microssatélites. Foi observada uma alta heterozigosidade intra-populacional (NP: 0,692; SA: 0,724 e SB: 0,686). Trinta e oito fragmentos e seis alelos foram compartilhados entre NP, SA e SB. Os valores de FIS e índice de Shannon demonstraram ausência de endogamia entre os grupos. As análises de AMOVA e FST indicaram alta (NP vs SA e SB) e pequena (SA vs SB) diferenciação genética, resultados confirmados pela distância e identidade genética, número de migrantes e dendrograma, que evidenciaram a formação de dois grupamentos genéticos. Conclusões: Os grupos possuem adequada variabilidade genética, com alta diferenciação entre NP e SA-SB e similaridade entre os estoques de reprodutores.

Larval development of Brycon amazonicus (Teleostei, Bryconidae) with a focus on locomotory, respiratory and feeding structures.

In the present work we describe the larval development of Brycon amazonicus, with emphasis on structures linked to swimming, respiration and feeding. We monitored from larval hatching (13 h post-fertilization) to juvenile stage. Formation of the oral cavity and differentiation of gill arches began within 4 h post-hatching (hph) and within 11 hph the primordium of the pectoral fin appeared. At 25 hph, the head was in a rectilinear position with a terminal mouth, when the onset of tooth formation appeared. At 35 hph, gill arches were covered by the operculum and branchiostegal membrane and cusps of teeth pierced the epithelium of the premaxilla and anterior dentary region. Sharp teeth were observed in the maxilla at 171 hph and the pelvic fin primordium appeared at 243 hph, with taste buds on the tongue and the roof of the buccopharyngeal cavity. At 579 hph, all structures related to swimming and food capture were formed. Exogenous feeding of the larvae started at 29 hph, before the endogenous reserves were used up and the development of efficient swimming ability, reflecting the species' need to increasingly specialize on prey capture.

Efecto de la restricción de alimento y posterior realimentación sobre algunas variables fisiológicas y metabólicas en yamú (Brycon amazonicus) / Effects of food restriction and refeeding on some physiological and metabolic variables of yamú (Brycon amazonicus)

RESUMEN Se evaluó el efecto de la restricción de alimento y posterior realimentación sobre algunas variables fisiológicas en yamú (Brycon amazonicus) utilizando 1115 juveniles (peso inicial 217,75 g ± 14,74 g) distribuidos en tres estanques y asignados a los siguientes tratamientos: 1) Control (C), alimentación todos los días; 2) Restricción moderada (RM), restricción por 2 días y realimentación por 3 y 3) Restricción severa (RS), restricción total durante 5 semanas y realimentación a partir de la semana 6 hasta la 12. Se realizaron colectas de sangre los días 1, 35, 42, 56, 70 y 84, de 7 animales por tratamiento para determinación del hematocrito, proteína, glucosa, lactato, triglicéridos, colesterol, cortisol, T3 e insulina. Adicionalmente, los animales se sacrificaron para retirar el hígado y determinar el glucógeno hepático. El modelo experimental fue mixto de efectos fijos, con 3 tratamientos y 6 muestreos, con anidamiento del muestreo en los tratamientos. Los datos se analizaron a través de análisis de varianza y en los casos en que hubo diferencias significativas se empleó la prueba de Tukey-Kramer. No hubo diferencias significativas (p > 0,05) para hematocrito, proteína, glucosa, cortisol y lactato entre los tratamientos en ninguna de las muestras tomadas. La insulina y T3 mostraron diferencias entre tratamientos en el día 35, pero no en el 84, igual a lo ocurrido con colesterol y glucógeno hepático. Los triglicéridos mostraron diferencias a lo largo del estudio, en el día 84 RS presentó menores niveles. Los resultados permiten concluir que pese a la restricción parcial de alimento, no se generaron efectos negativos sobre el metabolismo de Brycon amazonicus.

ABSTRACT The effect of food restriction and refeeding on some physiological variables in yamú (Brycon amazonicus) was evaluated using 1115 juveniles (initial weight 217.75 g ± 14.74 g), distributed in three ponds. The groups were randomly assigned one of the following treatments: 1) Control (C), fed every day; 2) Moderate restriction (MR), restriction during 2 days and refeeding during 3; and 3) Severe restriction (SR), total restriction during 5 weeks and refeeding from week 6 to 12. Blood samples from 7 animals from each treatment were taken on days 1, 35, 42, 56, 70, and 84 to determine hematocrit, proteins, glucose, lactate, triglycerides, cholesterol, cortisol, T3, and insulin. The animals were sacrificed and their livers removed to calculate the hepatic glycogen. The experimental design used was a fixed effects mixed model of 3 treatments and 6 samples, with sampling nesting in the treatments. A variance analysis was performed to analyze the data; the Tukey-Kramer test a variance analysis was used in the cases with significant differences. No significant differences (p> 0.05) were found for hematocrit, protein, glucose, cortisol, and lactate. The insulin and T3 showed differences among treatments on day 35, but not on day 84, the same as cholesterol and hepatic glycogen. Triglycerides showed differences throughout the study, SR showed the lowest levels on day 84. The results allow us to conclude that, despite the partial feed restriction, there were no adverse effects on the metabolism of Brycon amazonicus.

The influence of the photoperiod on productive performance and survival of piabanha-of-Pardo (Brycon vonoi ), hatchery / Influência do fotoperíodo no desempenho produtivo e sobrevivência na criação de larvas de piabanha-do-Pardo (Brycon vonoi)

ABSTRACT: This research aimed to verify the influence of the photoperiod on the performance and survival piabanha-of-Pardo (Brycon vonoi) larvae. Piabanha-of-Pardo larvae 24 hours post-hatch, with body weight of 2.44 ± 0.02mg, and total length of 7.82 ± 0.46mm, were subjected to five different photoperiods: 0L:24D, 6L:18D, 12L:12D, 18L:6D and 24L:0D (871.12 ± 92.65 lux). Daily, the water temperature, dissolved oxygen (DO), percentage of oxygen saturation, pH, and water conductivity were monitored, and presented similar water quality values among tanks with distinct photoperiods, and within the range considered appropriate for the genus Brycon larvae. Piabanha-of-Pardo larvae have better productive performance and survival rate at longer photoperiods, over 6 hours of light and below 12 hours of light per day. The 9L:15D photoperiod, specified by the derived equation, is indicated for the piabanha larviculture, but researches with the species and the photoperiod need to be better studied.

RESUMO: Este trabalho apresenta como objetivo verificar a influência do fotoperíodo sobre o desempenho e sobrevivência de larvas piabanha-do-Pardo (Brycon vonoi). As larvas de piabanha-do-Pardo com idade de 24h pós-eclosão, com peso corporal de 2,44 ± 0,02mg, e comprimento total de 7,82 ± 0,46mm, foram submetidas a cinco diferentes fotoperíodos: 0L:24E, 6L:18E, 12L:12E, 18L:6E e 24L:0E (871,12 ± 92,65 lux). Diariamente a temperatura da água, oxigênio dissolvido, percentagem de saturação de oxigênio, pH, e condutividade da água foram monitorados. Estes apresentaram valores similares entre os aquários com distintos fotoperíodos, e dentro da faixa considerada, adequada para as larvas do gênero Brycon. As larvas de piabanha-do-Pardo apresentam melhor desempenho produtivo e taxa de sobrevivência em fotoperíodo mais longos, acima de 6 horas de luz e abaixo de 12 horas de luz. O fotoperíodo de 9L:15E, especificado pela equação derivada, é indicado para a larvicultura da piabanha, mas as pesquisas com a espécie e o fotoperíodo precisam ser melhor estudadas.

Cross-amplification of heterologous microsatellite markers in Piracanjuba / Amplificação cruzada de marcadores microssatélites heterólogos em piracanjuba

ABSTRACT: Brycon orbignyanus, popularly known in Brazil as piracanjuba, is a fish with great economic value but whose natural population drastically decreased in number during the last years. In this context, genetic variability studies of natural stocks and in restocking programs are fundamental for the adoption of conservation measures. Current analysis verifies the cross-amplification of heterologous primers in B. orbignyanus. Fifty-two primers of the species Brycon opalinus, Brycon hilarii, Brycon insignis, Prochilodus sp., Piaractus mesopotamicus, Colossoma macropomum and Oreochromis niloticus were tested. Primers with the best reproducibility were applied to a sample of 20 individuals and the genetic parameters were calculated. Nine primers provided good results for cross-amplification with B. orbignyanus, involving (BoM5 and BoM13) of Brycon opalinus, (Bh5, Bh6, Bh8, Bh13 and Bh16) of Brycon hilarii, (Bc48-10) of Brycon insignis and (Par80) of Prochilodus argenteus. Primers of Piaractus mesopotamicus, Colossoma macropomum and Oreochromis niloticus failed to provide amplification or provided non-specificity. Results demonstrated the possibility of using primers of different species and genera of B. orbignyanus, facilitating genetic studies on the species.

RESUMO: A piracanjuba (Brycon orbignyanus) é um peixe de grande valor econômico que nos últimos anos tem apresentado uma redução drástica em suas populações naturais. Nesse contexto, estudos de variabilidade genética dos estoques naturais e nos programas de repovoamento são fundamentais para adoção de medidas conservacionistas. O objetivo do presente trabalho foi verificar a amplificação cruzada de primers heterologos em B. orbignyanus. Foram avaliados um total de 52 primers das espécies Brycon opalinus, Brycon hilarii, Brycon insignis, Prochilodus sp., Piaractus mesopotamicus, Colossoma macropomum e Oreochromis niloticus. Os primers com melhor reprodutibilidade foram aplicados a uma amostra de 20 indivíduos e os parâmetros genéticos foram calculados. Nove primers apresentaram resultados satisfatórios de amplificação cruzada com B. orbignyanus, sendo das espécies Brycon opalinus (BoM5 e BoM13), Brycon hilarii (Bh5, Bh6, Bh8, Bh13 e Bh16), Brycon insignis (Bc48-10) e Prochilodus argenteus (Par80). Os primers de Piaractus mesopotamicus, Colossoma macropomum e Oreochromis niloticus não apresentaram amplificação ou apresentaram inespecificidade. Os resultados revelaram a possibilidade da utilização de primers de diferentes espécies e gênero em B. orbignyanus, o que facilita a realização de estudos genéticos nessa espécie.

Effects of exercise training on excitation-contraction coupling, calcium dynamics and protein expression in the heart of the Neotropical fish Brycon amazonicus.

Matrinxã (Brycon amazonicus) is a great swimming performance teleost fish from the Amazon basin. However, the possible cardiac adaptations of this ability are still unknown. Therefore, the aim of the present work was to investigate the effects of prolonged exercise (EX group - 60days under 0.4BL·s-1) on ventricular contractility by (i) in-vitro analysis of contractility comparing the relative roles of sodium/calcium exchanger (NCX) and sarcoplasmic reticulum (SR) in the excitation-contraction (E-C) coupling and (ii) molecular analysis of NCX, sarcoplasmic reticulum Ca2+ ATPase (SERCA2) and phospholamban (PLB) expression and quantification. The exercise training significantly improved twitch tension, cardiac pumping capacity and the contraction rate when compared to controls (CT). Inhibition of the NCX function, replacing Na+ by Li+ in the physiological solutions, diminished cardiac contractility in the EX group, reduced all analyzed parameters under both high and low stimulation frequencies. The SR blockage, using 10µM ryanodine, caused ~50% tension reduction in CT at most analyzed frequencies while in EX, reductions (34-54%) were only found at higher frequencies. SR inhibition also decreased contraction and relaxation rates in both groups. Additionally, higher post-rest contraction values were recorded for EX, indicating an increase in SR Ca2+ loading. Higher NCX and PLB expression rates and lower SERCA2 rates were found in EX. Our data indicate that matrinxã presents a modulation in E-C coupling after exercise-training, enhancing the SR function under higher frequencies. This was the first study to functionally analyze the effects of swimming-induced exercise on fish cardiac E-C coupling.

Microcystin-LR leads to oxidative damage and alterations in antioxidant defense system in liver and gills of Brycon amazonicus (SPIX & AGASSIZ, 1829).

Microcystin's (MCs) are toxins produced by several groups of cyanobacteria, in water bodies throughout the world, in a process which is being intensified by human action. Among the variants of MCs, MC-LR stands out for its distribution and toxicity. MCs are potent inhibitors of protein phosphatases 1 and 2 A, which causes disruption of the cytoskeleton and consequent cell death. They can also alter the antioxidant system and induce oxidative stress in various organs of many species. There is, however, a lack of information about the effects of MCs on the antioxidant system and oxidative damage in Brazilian fishes. This study evaluated the effect of microcystin-LR on the antioxidant system in liver and gills of the Brazilian fish Brycon amazonicus, after 48 h of i.p injection of 100 µg MC-LR.kg-1 body mass. The liver exhibited increases in the activity of GST (74%) and GPx (217%), and a 47% decrease in SOD activity, with no changes in CAT values. In the gills of fish exposed to MC-LR, CAT and GPx activities did not show significant changes, while SOD and GST activity decreased by 66% and 37%, respectively. The GSH content did not change significantly in the liver, however, a decrease of 43% was observed in the gills. Oxidative damage measured by protein oxidation (PC) and lipoperoxidation (LPO) showed significant effects in both tissues. In hepatic tissue, there was no change in PC levels but LPO increased by 116%. Conversely, in the gills LPO levels did not change but PC increased by 317%. In conclusion, these data show that MC-LR induces oxidative damage in both tissues but in different ways, with being liver most sensitive to LPO and gills to PC. This also suggests that the gills are most sensitive to oxidative stress than liver, due to the inhibition of its antioxidant responses following MC-LR exposure.

Evaluación genotóxica del agua del Río Grande (Antioquia, Colombia) mediante frecuencia de eritrocitos micronucleados de Brycon henni (Characiformes: Characidae) / Genotoxic evaluation of Río Grande (Antioquia, Colombia) water using micronucleus frequency in erythrocytes of Brycon henni (Characiformes: Characidae)

ResumenLa producción agrícola de monocultivo permanente de banano y plátano en la parte media del Río Grande (Turbo - Antioquia) requiere de la aplicación de diferentes plaguicidas. Prácticas inapropiadas en la producción y cultivos de plátano realizadas en esta región a menudo conllevan a contaminación con agroquímicos que llegan a este cuerpo de agua por procesos de lixiviación y escorrentía. Los peces son los vertebrados más utilizados como especies bioindicadoras de la calidad del agua porque son muy sensibles a la presencia de contaminantes en el ambiente. El objetivo de esta investigación fue comparar la frecuencia de eritrocitos micronucleados (EMN) en sangre periférica de ejemplares de peces sabaleta (Brycon henni) procedentes de dos estaciones del Río Grande que difieren en su grado de contaminación por agroquímicos. Se evaluó la frecuencia de EMN en sangre periférica de peces B. henni procedentes de cada estación durante dos épocas de lluvia de 2010 y dos época secas de 2011. Las muestras de sangre fueron obtenidas de la vena caudal, fijadas durante 24 horas y luego teñidas con Giemsa. La frecuencia de EMN fue significativamente mayor en el área impactada por agroquímicos. La frecuencia media de EMN fue mayor en la estación con impacto directo de agroquímicos (0.15±0.18) que en la estación poco alterada (0.06±0.08). Además, la frecuencia de EMN en B. henni fue significativamente mayor en la época seca. Los resultados de este estudio indican que el análisis de las EMN en B. henni puede ser recomendada como un método adecuado para la detección in situ de agentes genotóxicos en el ambiente.

Abstract:The permanent monoculture of bananas and plantains farming in the middle of Río Grande (Turbo - Antioquia) requires the application of a variety of pesticides. Inappropriate banana production practices in this region, have often led to waterbody pollution by agrochemicals from leachate and runoff processes. Currently, fish are the most common vertebrates used as bioindicators of water quality, because they are very sensitive to the presence of contaminants. Our main goal with this study was to compare the frequency of micronucleated erythrocytes (MNE) in peripheral blood specimens of fish Brycon henni, from two locations (polluted and unpolluted) in the Rio Grande. We evaluated the frequency of MNE in peripheral blood samples of fish B. henni from each location during two rainy seasons in 2010 and two dry seasons in 2011. Blood samples were collected, fixed for 24 h, and then were stained with Giemsa. Among results, we found that the median frequency of MNE was higher in the polluted site by agrochemical discharges (0.15±0.18), than in the unimpacted site (0.06±0.08). Furthermore, the frequency of MNE in B. henni during the dry season was highly significant for both locations. The results of this study indicated that the analysis of MNE in B. henni could be recommended as a suitable method for in situ detection of environmental genotoxins. Rev. Biol. Trop. 65 (1): 405-414. Epub 2017 March 01.

A new species of Auriculostoma (Trematoda: Allocreadiidae) from the intestine of Brycon guatemalensis (Characiformes: Bryconidae) from the Usumacinta River Basin, Mexico, based on morphology and 28S rDNA sequences, with a key to species of the genus.

We describe a new species of Auriculostoma Scholz, Aguirre-Macedo & Choudhury, 2004 based on several sources of information including morphology (light and scanning electron microscopy [SEM]), sequences of two nuclear genes, host association, and geographical distribution. Morphologically, the new species most closely resembles Auriculostoma astyanace Scholz, Aguirre-Macedo & Choudhury, 2004, but differs by having deeply lobated testes and cirrus-sac extending posteriorly to seminal receptacle level. Auriculostoma lobata n. sp. can be readily distinguished from all the other congeners by the combination of the following characters: testes located in tandem, testes deeply lobated, and larger body size. A phylogenetic analysis using 28S rDNA sequences along with those available for other allocreadiid trematodes, revealed that the new species is a sister taxon of A. astyanace, a species described from the banded astyanax, Astyanax fasciatus (Cuvier) in Nicaragua. Auriculostoma totonacapanensis Razo-Mendivil, Mendoza-Garfias, Pérez-Ponce de León & Rubio-Godoy, 2014 from the Mexican tetra, Astyanax mexicanus (De Filippi) in Mexico is the sister taxon of A. astyanace plus the new species. Genetic divergence levels for the 28S rDNA and ITS2 were estimated among the Middle-American species of Auriculostoma infecting characiforms. The validity of the new species is then established by reliable morphological differences, its host association to bryconids (Brycon guatemalensis Regan), restricted geographical distribution (Usumacinta and Lacantun River basins), and genetic divergence levels, albeit relatively low. A morphometric comparison between the new species and the other seven congeneric species was undertaken and, in addition, a taxonomic key to identify the species contained in the genus Auriculostoma, widely distributed across the Americas, is provided.

Physiological and biochemical changes of females of Piracanjuba, subjected to induced reproduction.

The aim of this experiment was to compare the level of fish female stress during induced reproduction with pituitary extract by two different methods, natural and semiextruded. The reproductive efficiency was 62.5% in the seminatural treatment and 100% in the extruded. Obtained egg volume was 5200 ml and 4000 ml, for seminatural and extruded treatments respectively. The mean number of eggs was 46.7 for the seminatural and 52.0 and for the extruded treatment. The percentage of viable eggs was, respectively, 87.2% and 8.17% for the natural treatment and extruded semimethods. Blood samples were collected to quantify cortisol and glucose levels, as well as red cell series and lymphocyte count. Fishes submitted to induction procedures showed elevated cortisol and glucose levels, compared to the control animals. The results for haematocrit, haemoglobin concentration and red blood cell count showed no significant differences among groups. Significant differences found in the number of lymphocytes and monocytes suggest the general adaptation syndrome. Our results suggest the reproductive induction process with extrusion of gametes as a more stressful method than seminatural reproduction process.

Morphological, ultrastructural and phylogenetic analyses of Myxobolus hilarii n. sp. (Myxozoa, Myxosporea), a renal parasite of farmed Brycon hilarii in Brazil.

Myxobolus hilarii n. sp. was described, based on morphology, histology, ultrastructure and 18S rDNA sequencing, infecting the kidney of Brycon hilarii (Valenciennes 1850) (Characiformes: Bryconidae) taken from fish farms in the state of São Paulo, Brazil. Thirteen specimens of B. hilarii were examined and 100% had round, white plasmodia in the kidney. The mature myxospores were rounded, measuring 11.5 ± 0.8 (9.8-13.4) µm in length, 11.0 ± 0.7 (9.7-12.4) µm in width and 7.6 ± 1.0 (6.7-9.0) µm in thickness. Polar capsules were elongated and of equal size, with 6.5 ± 0.4 (6.0-7.2) µm in length and 4.0 ± 0.2 (3.6-5.3) µm in width and their polar filaments had 5 to 7 coils. Histological analysis revealed plasmodial development in the renal tubules, causing compression and deformation of adjacent tissues and destruction of renal tubule cells. Ultrastructural analysis showed direct contact between the plasmodial wall and the host tissue and asynchronous plasmodial development. The phylogenetic analysis of South American myxobolids, based on 18S rDNA sequencing, showed the myxosporeans grouping into two main clades. M. hilarii n. sp. appears as sister species of Myxobolus piraputangae.

Seminal characteristics of piabanha before and after induction with different hormones / Características seminais de piabanha antes e após a indução com diferentes hormônios

The migratory species piabanha does not reproduce in lentic environments since it requires environmental stimuli for the maturation and extrusion of gametes, and therefore hormonal induction is mandatory. Current study compares the seminal characteristics of Brycon insignis without any hormonal induction (Control - Ctrl) and with two types of hormonal inductors, or rather, carp pituitary extract (T1 - 2.5 mg kg-1 body weight) and GnRH analogues, the latter applied in two different concentrations (T2 - 0.7 mg kg-1 body weight and T3 - 1.4 mg kg-1 body weight). Post-induction analyses showed that the hormones increased the motility rate - Ctrl (95%), T1 (100%), T2 (100%) and T3 (98%), although sperm concentration - Ctrl (11.52 x 109); T1 (4.37 x 109); T2 (4.34 x 109); T3 (4.01 x 109) decreased. Assessments for sperm vigor, motility time and spermatic morphology did not vary with hormonal induction. Hormonal inducer does not alter negatively the seminal characteristics of the piabanha, and the choice for the proper hormone depends on the preference of the dispenser.

A espécie migradora piabanha não possui a capacidade de reproduzir em ambientes lênticos devido à necessidade de estímulos ambientais para a maturação e extrusão dos gametas, por isso a necessidade da indução hormonal. No presente estudo, as características seminais do Brycon insignis foram comparadas sem indução hormonal (Ctrl) e utilizando dois tipos de indutores hormonais - Extrato de Hipófise de Carpa (T1 - 2,5 mg kg-1 de peso vivo) e Análogos de GnRH, sendo este último aplicado em duas concentrações distintas (T2 - 0,7 mg kg -1 de peso vivo e T3 - 1,4 mg kg-1 de peso vivo). As análises realizadas após a indução mostraram que os hormônios utilizados produziram um aumento da taxa de motilidade - Ctrl (95%), T1 (100%), T2 (100%) e T3 (98%), porém houve uma diminuição na concentração espermática - Ctrl (11,52 x 109), T1 (4,37 x 109), T2 (4,34 x 109) e T3 (4,01 x 109). Os restantes das avaliações, vigor espermático, tempo de motilidade e morfologia espermática não apresentaram variações com a indução hormonal. Portanto, a utilização do indutor hormonal não altera negativamente as características seminais de piabanha, e a escolha do mesmo se deve à preferência do manipulador.

Comparative morphology of the oocyte surface and early development in four characiformes from the São Francisco River, Brazil.

Early development from the egg fertilization to complete resorption of the yolk-sac is a critical period in the life cycle of teleost fish. Knowledge of this process provides essential parameters for aquaculture and identification of spawning sites in the wild. In the present study, a comparative morphological analysis of the oocyte surface as well as early development was performed in four commercially valuable species from the São Francisco River: Brycon orthotaenia, Leporinus obtusidens, Prochilodus argenteus, and Salminus franciscanus. Stripped oocytes, embryo, and yolk-sac larvae were analyzed by scanning electron microscopy (SEM) and histology. A set of 10 lectins was used for investigation of lectin-binding pattern in oocytes. In the four species, the outer layer of the zona radiata reacted to most lectins, indicating complex polysaccharides at the oocyte surface while no reactivity was detected in the inner zona radiata and yolk globules. Typical structural arrangements were recognized at the micropylar region by SEM. The four species showed nonadhesive eggs, short embryonic period (18-20 h at 24 ± 1°C), and poorly developed larvae at hatching. At 24 h posthatching (hph), larvae of the four species had neuromasts on the body surface. Rudimentary cement glands for larval attachment were identified on the cephalic region at 24 and 48 hph in B. orthotaenia and S. franciscanus, and following they were in regression. The time for whole yolk resorption varied among species from 48 to 120 hph, occurring earlier in S. franciscanus, followed by B. orthotaenia, P. argenteus, and L. obtusidens. The formation of the digestive tract and the mouth opening indicated initiation of exogenous feeding 24 h before complete resorption of the yolk. Together, our data indicate similarities in the early development among species that may be related to the life cycle strategies and phylogeny.

Aloe vera bathing improved physical and humoral protection in breeding stock after induced spawning in matrinxã (Brycon amazonicus).

In this study, we show that induced spawning causes stress, an intense loss of epithelia and immunosuppression, decreasing physical and humoral protection in fish, effects that were prevented or improved in fish bathed with Aloe vera. A. vera has several medicinal properties, including wound healing and immunostimulatory effects, which we observed in this study. Fish bathed with A. vera had a higher number of epidermal goblet cells and, in general, an improved wound healing rate compared with the control after induced spawning. These effects might be related to (1) the stimulation of leukocyte activity, represented here by the increased leukocyte respiratory activity triggered by A. vera (leukocytes are recognized as playing an important role in wound repair); (2) the antimicrobial properties of A. vera, which decrease wound infection and accelerate the healing process; and (3) several mechanisms that explain the healing effect of A. vera (increased collagen synthesis, rate of epithelialization, and anti-inflammatory and moisturizing effects). Our results also suggest that caution is necessary during the induced spawning process, especially during stripping, and A. vera bathing is recommended after intensive aquaculture operations.

Evaluation of a recirculation system for Dorada (Brycon moorei-Steindachner 1878) larvae in the Magdalena River / Evaluación de un sistema de recirculación para la etapa de larvicultura de dorada (Brycon moorei-Steindachner 1878) del Río Magdalena / Avaliação de um sistema de recirculação para a fase de larvicultura de dourada (Brycon moorei-Steindachner 1878) de Rio Magdalenao

Recirculating aquaculture systems allow increased productivity during larval stages due to a greater control of water quality parameters. Dorada larvae (Brycon moorei) were used 12 hours post hatching (HPE) to evaluate three planting densities with four replicates per treatment and 10, 20 and 30 larvae L-1 (T1, T2 and T3, respectively). The larvae were fed with Bocachico (Prochilodus magdalenae) larvae, Artemia salina nauplii and a 40% crude protein (CP) commercial diet for 15 days. Results showed significant differences between treatments (p<0.0001) for weight gain and specific growth rate (TCE), where T1 performed better (73.24±19 mg and 14.21±5.02%, respectively); followed by T3 (67.15±24.3 mg and 13.94±7.51%) and T2 (55.67±18.33 mg and 13.36±5.75%). Survival rate was inversely related to density, with highly significant difference (p <0.0001) (39.37±6.84%, 28.00±2.80% and 14.87±6.47%) for T1, T2 and T3, respectively. Simultaneously, cannibalism was directly related to planting density. The highest profitability was obtained with T2.

Los sistemas con recirculación para acuicultura permiten el incremento de la productividad en la etapa de larvicultura, debido al mayor control de los parámetros de calidad de agua. Larvas de dorada (Brycon moorei) de 12 horas post eclosión (HPE) fueron utilizadas para evaluar tres densidades de siembra con cuatro replicas por tratamiento, 10, 20 y 30 larvas L-1 (T1, T2 y T3, respectivamente). Las larvas fueron alimentadas durante 15 días, con larvas de bocachico (Prochilodus magdalenae), nauplios de Artemia salina y una dieta comercial de 40% de proteína cruda (PC). Al final del experimento, los resultados muestran diferencias altamente significativas entre los tratamientos (p<0.0001) para ganacia de peso y tasa de crecimento específica (TCE), obteniendo un major desempeñoen T1 (73,24±19 mg y 14,21±5,02%, respectivamente); seguido por T3 (67,15±24,3 mg y 13,94±7,51%) y T2 (55,67±18,33 mg y 13,36±5,75%). La sobrevicencia se relacionó de manera inversa con la densidad con diferencia altamente significativa (p<0.0001) (39,37±6,84%; 28,00±2,80% y 14,87±6,47%) para T1, T2 y T3, respectivamente y, simultáneamente, el canibalismo fue directamente relacionado con la densidad de siembra. Por otro lado, la mayor rentabilidad fue obtenida con T2.

Os sistemas de recirculação de aquicultura permitir o aumento da produtividade na fase de larvicultura devido ao melhor controle dos parâmetros de qualidade da água. Larvas Bream (Brycon moorei) 12 horas após a eclosão (HPE) foram utilizados para avaliar três densidades de estocagem, com quatro repetições por tratamento, 10, 20 e 30 larvas L-1 (T1, T2 e T3, respectivamente). As larvas foram alimentadas durante 15 dias com bocachico larvas (Prochilodus magdalenae), náuplios Artemia salina e uma dieta comercial de proteína bruta de 40% (CP). No final da experiência, os resultados mostraram diferenças altamente significativas entre tratamentos (p < 0,0001) para o ganho de peso e taxa específica de crecimento (TEC), obtendo-se um grande desempeño en T1 (73,24±14,21 mg e 19±5,02%, respectivamente); seguido de T3 (67,15±24,3 mg e 13,94±7,51%) e T2 (55,67±18,33 mg e 13,36±5,75%). O sobrevicencia está inversamente relacionada com a densidade, com diferença altamente significativa (p<0,0001) (39,37±6,84%; 28,00%±2,80 e 14,87±6,47%) para T1 , T2 e T3, respectivamente e, simultaneamente, o canibalismo foi diretamente relacionada à densidade de estocagem. Por outro lado, o maior rendimento foi obtido com T2.