Hematological Evaluation of Three Common Teleosts in Relation to The Environmental Changes from Trang Province, Thailand.

Hematological evaluation of fish is essential to the assessment of their physiological status. This study describes the morphometric analysis and comparison of blood cell characteristics in Zanarchopterus sp., Gerres filamentosus Cuvier, 1829 and Leiognathus decorus (De Vis, 1884). The species were collected at two locations off the coast of Trang Province, Thailand. A comparative hematological evaluation was made to assess the effects of environmental conditions on the blood of the fish. Ten individuals of each species were collected from a seagrass bed at Libong Island, where human activities are increasing, and from a secluded sandy beach. Their blood samples were analysed using the blood smear technique. Erythrocytes of all the studied fishes were either elliptical or oval. The morphometric data from both locations showed that erythrocytes were of similar size, except for those of Zanarchopterus sp. Fish from both stations showed several types of leukocytes, including neutrophils and lymphocytes.The highest proportion of leukocytes was made up of lymphocytes, followed by neutrophils. However, monocytes were only observed in fish from Libong Island and the erythrocytic nuclei of fish collected from Libong Island were both reniform and lobate. Our results show the potential of hematological evaluation as an early warning signal of environmental impacts on aquatic animals. The determination of baseline parameters could provide a tool for the monitoring of environmental quality.

Immune response in sexual inverted and non-inverted Nile tilapia fingerlings supplemented with organic acid and essential oil.

The development and intensification of tilapia farming depends on the manipulation of some physiological functions, such as the sexual inversion of larvae using a synthetic androgen (17α-methyltestosterone). This inversion, however, may represent a potential oxidative stress factor and cause damage to animals in the short, medium, and long term. Dietary supplementation of natural antioxidant compounds is an interesting alternative to combat such damage. To test this hypothesis, an experimental trial was carried out involving sexual inverted and non-inverted Nile tilapia fingerlings, both supplemented and not supplemented with a blend of organic acids and essential oils protected by microencapsulation. Animals were divided into four experimental groups: NI (non-inverted animals), I (sexual inverted animals), NI + M (non-inverted animals supplemented with microcapsules), and I + M (sexual inverted animals supplemented with microcapsules). Blood parameters (WBC - white blood cells; LY - lymphocytes; RBC - red blood cells; HGB - hemoglobin; HCT - hematocrit number; MCH - mean corpuscular hemoglobin; MCV - mean corpuscular volume and MCHC - mean corpuscular hemoglobin concentration), as well as oxidative stress markers (enzymatic activity of superoxide dismutase - SOD and catalase - CAT; and total antioxidant capacity - 2,2-diphenyl-1-picryl-hydrazyl (DPPH)) and gene expression (heat shock protein 70 kDa - HSP70) were evaluated. The HGB (p < 0.001) and HCT (p = 0.005) parameters were reduced beyond the recommended limits for the animals in group I. The MCV varied statistically between the groups (p < 0.001). However, all values were within the recommended range for the species, jointly indicating normocytic anemia in group I fingerlings at the time of collection. The activity of CAT and SOD, as well as DPPH differed statistically between the experimental groups (p < 0.001), with the lowest SOD and CAT activity, as well as the highest DPPH registered in animals supplemented with microcapsules. The expression of HSP70 was lower in I + MI animals (p < 0.001). The synergistic evaluation of the results indicates that animals sexual inverted during the larval stage have a lower total antioxidant capacity in the fingerling stage, which reflects a worsening in hematological and enzymatic parameters related to immunity; and that dietary supplementation with blend of organic acids and essential oils protected by microencapsulation is sufficient to improve the immunological response both in sexual inverted and non-inverted fingerlings.

Dietary Brown Propolis Extract Modulated Nonspecific Immune System and Intestinal Morphology of Pacu Piaractus mesopotamicus

Abstract The immunomodulatory and growth promoter effects of brown propolis ethanolic extract (PEE) were determined in pacu. Fish (28.4±0.4 g) were randomly distributed into 12 polyethylene circular tanks (250 L, 30 fish per tank) and fed for 60 days with a commercial diet (32% crude protein) supplemented with 0.0, 1.5, 3.0 and 4.5% (v/w) of PEE in a totally randomized experimental design (n=3). Fish organic defense cells numbers such as thrombocytes and neutrophils increased (p<0.05) in fish fed 3.0% dietary PEE. Serum lysozyme concentration also increased (p<0.05) in fish fed 1.5 and 3.0% PEE supplementation when compared to unsupplemented fish. Growth parameters were not influenced (p>0.05) by treatments. Moreover, dietary PEE decreased (p<0.05) fish intestinal muscular thickness when compared to control treatment. Intestine villi height also significantly decreased in fish fed 3.0% PEE. Long term dietary PEE at 3.0% supplementation level modulated fish nonspecific immune system and it is a non-toxic substance for pacu.

Na2EDTA anticoagulant impaired blood samples from the teleost piaractus mesopotamicus / Anticoagulante Na2EDTA danifica amostras de sangue do teleósteo piaractus mesopotamicus

The present study aimed to evaluate the effects of Na heparin and Na2EDTA on blood of Piaractus mesopotamicus (360.7±42.4g, 26.4±1.0cm). Twenty fishes were sampled in two experiment trials, ten for erythrocyte fragility analysis and ten for hematologic and plasma biochemical study. The blood collected by venous-caudal puncture was fractioned and stored in anticoagulants solution: Na2EDTA 10%, Na2EDTA 3%, Na heparin 5000 IU and Na heparin 100 IU. Plasmatic levels of calcium presented in the Na2EDTA stored samples were about 80% lower than both heparin groups. Blood samples of P. mesopotamicus stored with Na2EDTA demonstrated increase in the hematocrit and MCV, and decrease in MCHC. The dose-response effect was observed in this study. The results are reinforced by the higher levels of plasmatic protein and hemolysis presented in the Na2EDTA 10% stored blood, confirming the deleterious effect of this anticoagulant treatment on the quality of blood samples. Na2EDTA is not indicated to store P. mesopotamicus blood samples, but sodium heparin at 100 IU is the most recommended anticoagulant, since this treatment presented the lower rate of alterations in the stored blood.

O presente estudo teve por objetivo avaliar os efeitos da heparina sódica e Na2EDTA no sangue de Piaractus mesopotamicus (360,7±42,4g, 26,4±1,0cm). Foram amostrados vinte peixes em dois ensaios experimentais, sendo dez peixes utilizados para análise da fragilidade dos eritrócitos e dez peixes para análise dos parâmetros hematológicos e estudo bioquímico do plasma. O sangue coletado por punção veno-caudal foi aliquotado e armazenado em diferentes soluções anticoagulantes: Na2EDTA 10%, Na2EDTA 3%, heparina sódica 5.000 UI e heparina sódica 100 UI. Níveis plasmáticos de cálcio apresentados nas amostras armazenados em Na2EDTA diminuíram cerca de 80% em relação aos dois grupos armazenados com heparina. Amostras de sangue de pacus armazenados com Na2EDTA demonstraram aumento do hematócrito e VCM, e diminuição na CHCM. O efeito dose-resposta foi observado neste estudo. Estes resultados são reforçados pelos níveis mais elevados de proteína plasmática e hemólise apresentado no sangue armazenado com Na2EDTA 10%, o que confirma o efeito deletério desse tratamento anticoagulante na qualidade de amostras de sangue. Na2EDTA não é indicada para armazenar amostras de sangue de P. mesopotamicus, e heparina sódica a 100 UI é o anticoagulante mais recomendado, uma vez que este tratamento apresentou a menor taxa de alterações no sangue armazenado.

Toxicological and hematological effect of Terminalia arjuna bark extract on a freshwater catfish, Heteropneustes fossilis.

Increasing demand for eco-friendly botanical piscicides and pesticides as replacements for harmful synthetic chemicals has led to investigation of new sources of plant materials. Stem bark of Terminalia arjuna, which has been used as a popular folk medicine since ancient time, was examined for its piscicidal activity. This study aims to determine toxicity of ethanol extract of T. arjuna bark on fresh water stinging catfish (Heteropneustes fossilis), along with evaluation of changes in hematological parameters of the fishes exposed to a lethal concentration. The percent mortality of fishes varied significantly in response to concentrations of the extract and exposure times (between exposure time F = 36.57, p < 0.001; between concentrations F = 39.93, p < 0.001). The lethal concentrations (LC50) of ethanol extract were found to be 12.7, 8.94, 5.63 and 4.71 mg/l for 24, 48, 72 and 96 h, respectively. During acute toxicity test, blood samples of treatment fishes showed significant decreases in the red blood cells count, hematocrit content, hemoglobin concentration, mean corpuscular hemoglobin concentration and plasma protein level when compared to those of the control group, while there were significant increases in the mean corpuscular volume, mean corpuscular hemoglobin, white blood cells count and plasma glucose concentration. These results suggest that T. arjuna bark extract could be considered as a potent piscicide due to its toxic effect on fish, particularly fish hematology.

Heparina e K3EDTA como anticoagulantes para tambaqui (Colossoma macropomum Cuvier, 1816) / Heparin and K3EDTA as anticoagulants for tambaqui (Colossoma macropomum Cuvier, 1816)

Este estudo avaliou a eficácia da heparina sódica e do EDTA tripotássico como anticoagulantes e seus efeitos sobre os parâmetros hematológicos de tambaqui (Colossoma macropomum). Foram utilizados dez indivíduos de tambaqui com médias de 384,9 ± 85,71 g de peso e 27,90 ± 2,10 cm de comprimento total para avaliação da heparina 5.000 UI e 100 UI, bem como do K3EDTA 10 por cento. Foram analisados a inibição da coagulação por 10 h, eritrograma e teste de fragilidade osmótica dos eritrócitos. Os resultados foram submetidos à análise de variância e as médias comparadas pelo teste de Tukey (P < 0,05). Heparina 5000 UI, heparina 100 UI e K3EDTA 10 por cento foram eficazes na prevenção da coagulação por mais de 10 h, no entanto o EDTA tripotássico causou hemólise desde os primeiros momentos. No eritrograma não foi observada diferença (P > 0,05) na contagem de eritrócitos, hematócrito, taxa de hemoglobina e CHCM, no entanto, houve aumento do VCM (P < 0,05) nas amostras acondicionados com K3EDTA 10 por cento. Este anticoagulante causou incremento (P < 0,01) na fragilidade osmótica dos eritrócitos quando comparado com a heparina pura, heparina diluída, e grupo controle. A utilização da heparina como anticoagulante é mais apropriada para tambaqui (Colossoma macropomum), visto que foi eficiente na prevenção da coagulação por mais de 10 h, sem ocasionar hemólise, ou alterações nos parâmetros hematológicos e na fragilidade osmótica dos eritrócitos.

The efficacy of sodium heparin and tripotassium EDTA as anticoagulant and their effect on the hematological parameters of tambaqui (Colossoma macropomum) were evaluated in this study. Ten fish weighing 384.9 ± 85.71 g and measuring 27.90 ± 2.10 cm were used for heparin 5.000 IU, heparin 100 IU and K3EDTA 10 percent evaluation. Clotting inhibition after 10 h, erythrogram and osmotic fragility of erythrocytes were observed. The results were submitted to variance analysis and means compared by Tukey test (P < 0.05). Heparin 5.000 IU, heparin 100 IU and K3EDTA 10 percent were effective in preventing coagulation for more than 10 h. However, tripotassium EDTA caused hemolysis since first moments. In erythrogram there was no difference (P > 0.05) in erythrocyte count, hematocrit, hemoglobin and MCHC. On the other hand, an increase in MCV (P < 0.05) in samples kept with K3EDTA10 percent was observed. This anticoagulant provoked a significant increase (P < 0.01) in the osmotic fragility of erythrocytes when compared to pure heparin, diluted heparin and the control group. Heparin as an anticoagulant is more appropriate for tambaqui since it was effective in preventing coagulation for more than 10 h, without causing hemolysis, changes on hematological parameters or osmotic fragility of erythrocytes.

Heparina e Na2EDTA como anticoagulantes para surubim híbrido (Pseudoplatystoma reticulatum x P. corruscans): eficácia e alterações hematológicas / Heparin and Na2EDTA as anticoagulants for hybrid surubim catfish (Pseudoplatystoma reticulatum x P. corruscans): efficacy and hematological changes

Este estudo avaliou o efeito da heparina 100UI e do Na2EDTA nas concentrações de 3, 5 e 10 por cento sobre a coagulação sanguínea e os parâmetros hematológicos de surubim híbrido. Foram utilizados 10 peixes, com peso médio de 386,7±24,3g e comprimento total médio de 38,7±6,4cm, para colheita das amostras sanguíneas e determinação do percentual do hematócrito, teor de proteínas plasmáticas totais e teste de fragilidade osmótica dos eritrócitos (FOE). Os dados foram submetidos à análise de variância, e as médias foram comparadas pelo teste de Tukey, a 5 por cento de probabilidade. A coagulação foi eficientemente inibida quando utilizado o Na2EDTA; já as amostras com heparina coagularam 10 horas após a coleta. Houve aumento na fragilidade osmótica dos eritrócitos (P<0,01), especialmente quando utilizado o Na2EDTA 10 e 5 por cento, não havendo diferença entre o controle e a heparina. No hematócrito e teor de proteínas plasmáticas totais, não foram verificadas diferenças significativas. O Na2EDTA 3 por cento é seguro e eficiente como anticoagulante para surubim híbrido, prevenindo a coagulação por mais de 10h e ocasionando discreto efeito sobre a fragilidade osmótica dos eritrócitos.

The effect of heparin 100IU and Na2EDTA in concentration of 3 percent, 5 percent and 10 percent on the blood coagulation and hematological parameters of hybrid surubim catfish were evaluated. We used ten fish weighing 386,7±24,3g and average length of 38,7±6,4cm for collection of blood samples and determination of the percentage of hematocrit, plasma total protein content and osmotic fragility erythrocytes test (OFE). Data were submitted to analysis of variance and averages compared by Tukey test at 5 percent probability. Coagulation was efficiently inhibited when used the Na2EDTA, while the samples with heparin coagulate ten hours after collection. There was an increase in osmotic fragility of erythrocytes (P<0.01), especially when using Na2EDTA 10 percent and 5 percent, with no difference between control and heparin. In hematocrit and total plasma protein content were not observed statistical differences. The Na2EDTA 3 percent is safe and effective anticoagulant for hybrid surubim catfish, preventing clotting for more than 10 hours causing slight effect on the osmotic fragility of erythrocytes.

Distúrbios morfológicos em leucócitos de exemplar de kinguio (Carassius auratus auratus Linnaeus, 1758) com septicemia hemorrágica bacteriana: relato de caso / Morphologic disorders in leukocytes of goldfish sample (Carassius auratus auratus Linnaeus, 1758) with bacterial hemorrhagic septicemia: case report

O presente estudo teve por objetivo relatar um caso sobre as alterações morfológicas em leucócitos de kinguio comsepticemia hemorrágica bacteriana. Um exemplar foi atendido no Laboratório de Patologia de Organismos Aquáticos doCAUNESP, Unesp de Jaboticabal, apresentando apatia, natação errática, anorexia e discreta perda de equilíbrio, buscandoconstantemente a superfície da água. Através de análise visual da superfície corpórea, foram observadas áreas de hemorragiapetequial na base das nadadeiras peitoral, pélvica e caudal, além da órbita bucal, periocular e anal e efusão celomática. Naextensão sanguínea foram observadas a presença de monócitos ativados, hiperativados, e fagócitos mononuclearesrealizando eritrofagocitose e leucofagocitose, neutrófilos com basofilia citoplasmática e com vacuolização citoplasmática,além de imunócitos e leucócitos imaturos com anisocitose e anisocariose. As alterações morfológicas descritas no presenteestudo revelaram parte da patogenia e resposta orgânica de kinguio com septicemia hemorrágica bacteriana. Dessa forma,as análises qualitativas das células sanguíneas devem ser também utilizadas como ferramenta complementar aos estudosdo estado geral da saúde de peixes em cultivo.

The morphological changes in leukocytes of goldfish with bacterial hemorrhagic septicemia were reported. One sample was served at the Laboratory of Pathology of Aquatic Organisms of CAUNESP, Unesp in Jaboticabal. The fish showed lethargy, erratic swimming and without force, slight loss of balance, trying constantly to the water surface. In the body inspection, areas of petechial hemorrhage at the base of the pectoral fins, pelvic fins and caudal fin were observed, beyond the periocular and oral orbitand anal. Blood smears presented hiperactivated monocytes, performing erythrophagocytosis and leukophagocytosis, the cytoplasm of neutrophils presented basophilia and vacuolization, beyond immunocytes, immature leukocytes with anisocytosis and anisokaryosis. The morphological changes described in this study revealed part of the pathogenesis and organic response of Kinguio with bacterial hemorrhagic septicemia. Then, the qualitative analysis of blood cells should also be used as a complementary tool to study the general health status of fish farmed