Influence of dexamethasone and levamisole on macrophage recruitment, giant cell formation and blood parameters in the tropical fish Piaractus mesopotamicus / Influência da dexametasona e levamisol no recrutamento de macrófagos, formação de células gigantes e parâmetros hematológicos no peixe tropical Piaractus mesopotamicus

The aim of this study was to evaluate the effect of parenteral administration of dexamethasone and levamisole on the accumulation of macrophages and formation of giant cells in chronic inflammation by foreign body and blood parameters in pacu (Piaractus mesopotamicus). It was used 50 mg/kg of levamisole and 2.0 mg/kg of dexamethasone and the combination of both drugs. Coverslips were implanted under the skin. After 2, 7, and 15 days postimplantation (DPI), fish were anesthetized for removal of coverslips and the number of macrophages and giant cells were count. It was also collected blood from the caudal vessel to evaluate: red blood cell count, hematocrit, hemoglobin concentration, leukocytes count, thrombocytes count, MCV, and MCHC. We observed that dexamethasone affects negatively the formation of giant cells in the chronic inflammation for foreign body. Levamisole, despite being immunostimulatory in several species, showed limited action. However, it was enough to counteract the effect of dexamethasone; the association of the drugs did not interfere significantly in erythrocyte and leukocyte number in most of the treatments and times studied. In dexamethasone group there was a reduction in the number of erythrocytes and hemoglobin associated with increased mean corpuscular volume, suggesting slight macrocytic anemia. At 15 DPI, most groups showed the recovery of hematologic response. As in mammals, dexamethasone affects negatively the inflammatory response. Levamisole showed little effect by itself. However, in some parameters the association of both drugs causes similar response to control and naïve groups, showing the antagonistic effect of these drugs.

O objetivo deste estudo foi avaliar o efeito da administração parenteral de dexametasona e levamisol na acumulação de macrófagos e formação de células gigantes na inflamação crônica por corpo estranho e avaliação de parâmetros sanguíneos em pacu (Piaractus mesopotamicus). Utilizou-se 50 mg/kg de levamisol e 2,0 mg / kg de dexametasona e a combinação de ambos fármacos. As lamínulas foram implantadas sob a pele. Depois de 2, 7 e 15 dias pós-implantação (DPI), os peixes foram anestesiados para a remoção das lamínulas e contagem do número de macrófagos e células gigantes. Foi coletado sangue da veia caudal para realizar contagem de células vermelhas, leucócitos, trombócitos, concentração de hemoglobina, MCV e MCHC. Observou-se que a dexametasona afeta negativamente a formação de células gigantes na inflamação crónica por corpo estranho. O levamisol, apesar de ser considerado imunoestimulante em várias espécies, mostrou ação limitada. No entanto, foi suficiente para neutralizar o efeito da dexametasona; a associação das drogas não interferiu significativamente no número de eritrócitos e de leucócitos na maioria dos tratamentos e períodos estudados. No grupo da dexametasona, houve redução do número de eritrócitos e concentração de hemoglobina associado ao aumento do volume corpuscular médio sugerindo leve anemia macrocítica. Aos 15 DPI, a maioria dos grupos mostrou recuperação na resposta hematológica. Como em mamíferos, a dexametasona afeta negativamente a resposta inflamatória. O levamisol mostrou pouco efeito por si só. No entanto, em alguns parâmetros a associação das duas drogas provoca resposta similar ao grupo controle e naive, mostrando o efeito antagonista de estas drogas.

Acute aerocystitis in Nile tilapia bred in net cages and supplemented with chromium carbochelate and Saccharomyces cerevisiae.

Oreochromis niloticus bred in net cages were supplemented with cell wall of Saccharomyces cerevisiae (Sc) (0.3%) or chromium carbochelate (Cr) (18 mg/kg of feed) or in association (Sc + Cr), for 90 days. After this period, acute inflammation was induced in the swim bladder by inoculation of 3 × 10(8) CFU of inactivated Streptococcus agalactiae, and another group received 0.65% saline solution (control). Twelve, 24, and 48 h after stimulation, the inflammation was evaluated through total and differential counting of accumulated cells, and through leukocyte respiratory burst in the blood, cortisolemia, glycemia and serum lysozyme concentration. The results showed that there were greater total numbers of cells in the exudate of fish inoculated with inactivated bacterium than in those injected with saline solution, with predominance of lymphocytes, thrombocytes, macrophages and granulocytes. Tilapia supplemented with Cr presented increased total numbers of cells with significant accumulation of lymphocytes and reductions in cortisolemia and glycemia, but the different treatments did not have any influence on leukocyte respiratory burst or serum lysozyme concentration. Tilapia supplemented with Sc and the Cr + Sc association did not present significant changes to the variables evaluated, despite higher accumulation of lymphocytes in the inflammatory exudate from fish treated with Sc. The results indicate that tilapia bred in net cages and supplemented with Cr presented higher total accumulation of cells at the inflammatory focus, thus indicating an increase in the inflammatory response induced by the bacterium, probably due to the reduction in cortisolemia and higher glucose consumption. Thus, supplementation with Cr had beneficial action, which facilitated development of acute inflammation induced by the bacterium, but did not affect neither leukocyte respiratory burst in the blood nor serum lysozyme concentration.

Lectinhistoquímica del granuloma inducido por el bacilo de Calmette Guérin en Piaractus mesopotamicus / Lectinhistochemical staining of granuloma induced by bacillus Calmette-Guerin in Piaractus mesopotamicus

Objetive. This study was conducted to evaluate, by means of lectinhistochemistry (LHC), the expression of carbohydrates in granulomas induced by the bacillus Calmette-Guerin (BCG) in muscle tissue of Piaractus mesopotamicus after 33 days. Material and methods. Histological sections with 3 μm thick were incubated with the following lectins :WGA (Wheat germ agglutinin), DBA (Dolichos biflorus agglutinin) and HPA (Helix pomatia agglutinin), and the results were evaluated by light microscopy. Results. Acid fast bacilli were stained by Ziehl Neelsen (ZN) and strong labeled by WGA in the cytoplasm of macrophages. Labeling with DBA was intense in fibroblasts and weak in macrophages. On the other hand, HPA binding was stronger in macrophages, especially in those that were in close contact with epithelioid cells, without evidence of binding to fibroblasts. The epithelioid cells were not labeled by the used lectins, but they were identified by Hematoxilin-Eosin (HE). The lectins labeled specific type saccharides in glycoproteins, as N-acetylglucosamine present in bacilli and macrophages, as well as N-acetyl-galactosamine in macrophages. The control group showed no inflammation or lectin binding. Conclusions. This technique may be useful in identifying receptors for WGA, DBA and the HPA lectins in epithelioid granuloma induced by BCG in P. mesopotamicus.

El presente estudio fue realizado para evaluar por medio de lectinhistoquímica (LHC), la expresión de carbohidratos en granulomas inducidos por el bacilo de Calmette-Guérin (BCG) en músculo de Piaractus mesopotamicus después de 33 días. Materiales y métodos. Cortes histológicos de 3 µm de grosor fueron incubados con las siguientes lectinas: WGA (Wheat germ aglutinin), DBA (Dolichos biflorus agglutin) y HPA (Helix pomatia agglutinin), y los resultados evaluados por medio de microscopia de luz. Resultados. Bacilos ácido resistentes fueron identificados por la tinción de Ziehl Neelsen(ZN). Se observó un marcaje intenso con WGA en el citoplasma de macrófagos. El marcaje con DBA fue intenso en fibroblastos y débil en macrófagos. Con la lectina HPA el marcaje fue intenso en macrófagos, principalmente en los que estaban en estrecho contacto con las células epitelióides, externamente se observó marcaje débil en fibroblastos. Las células epitelióides no fueron marcadas por las lectinas, pero fueron identificadas con la tinción de Hematoxilina-Eosina (HE). Las lectinas tuvieron un tipo de marcaje específico en algunos monosacáridos, como N-acetilglucosamina presente en los bacilos y en macrófagos, y N-acetilgalactosamina en macrófagos. En el grupo control no fue observada inflamación así como tampoco marcaje con las lectinas. Conclusiones. Esta técnica resultó eficiente en la identificación de receptores para las lectinas WGA, DBA y HPA en el granuloma epitelióide inducido por BCG en P. mesopotamicus.

Acute aerocystitis in Piaractus mesopotamicus: participation of eicosanoids and pro-inflammatory cytokines.

A total of 360 pacus (Piaractus mesopotamicus) were used to study vascular permeability (VP) and inflammatory cell component (CC) in induced aerocystitis in P. mesopotamicus through inoculation of inactivated Aeromonas hydrophila, and the effect of steroidal and nonsteroidal anti-inflammatory drugs. It was observed that after inoculation of A. hydrophila, the maximum VP occurred 180 min post-stimulus (MPS). Pretreatment with anti-inflammatory drugs inhibited VP, and the inhibitory effect of dexamethasone was seen earlier than the effects caused by meloxicam and indomethacin. Inoculation of the bacterium caused a gradual increase in the accumulation of cells, which reached a maximum 24 h post-stimulus (HPS). Pretreatment with dexamethasone, indomethacin and meloxicam reduced the accumulation of lymphocytes, thrombocytes, granulocytes and macrophages. There was no significant difference between the different doses of the drugs tested. The results suggest that eicosanoids and pro-inflammatory cytokines participate in chemical mediation in acute inflammation in pacus.

Myxosporidiosis in intensively-reared Piaractus mesopotamicus: Histopathological diagnosis by means of Ziehl-Neelsen staining / Mixosporidiose em Piaractus mesopotamicus de criação intensiva: diagnóstico histopatológico pela coloração Ziehl-Neelsen

Samples of different organs from intensively-reared Piaractus mesopotamicus were collected and processed using routine histological techniques in order to produce thin sections for staining with hematoxylin-eosin and with the Ziehl-Neelsen method. Through examination under an optical microscope, myxosporidians of the genera Henneguya sp. and Myxobolus sp. were identified, respectivelyin the gills and kidneys of P. mesopotamicus. Plasmodia with immature spores of Henneguya sp. were located along the secondary lamellae, with total length of 30.45±4.84µm and width of 3.52±0.33µm. Spores of Myxobolus sp. were located in the kidneys, with total length of 8.94±0.82µm and width of 5.59±0.39µm. Histopathological analysis of the gills showed plasmodia containing spores of Henneguya sp., at intralamellar and intravascular localities, at different stages of development. Spores of Myxobolus sp. were identified in the kidneys, in the peritubular region and in the interstices and glomerulus, surrounded by melanomacrophages. Focal hemorrhage was recorded in a few cases. Ziehl-Neelsen staining allowed to identify particular features of the spores and facilitated biometry and enabled classification in comparison with hematoxylin-eosin, thus demonstrating its usefulness for histopathological diagnosis of the parasitosis.

Amostras de diferentes órgãos de Piaractus mesopotamicus mantidos em criação intensiva foram coletadas e processadas mediante as técnicas histológicas usuais para obtenção de cortes que foram corados com hematoxilina-eosina e pelo método de Ziehl-Neelsen. Ao exame em microscopia de luz foi possível identificar mixosporídeos dos gêneros Henneguya sp. e Myxobolus sp. em brânquia e rim de P. mesopotamicus respectivamente. Plasmódios com esporos imaturos de Henneguya sp. foram localizados ao longo das lamelas secundárias e mensurados (comprimento total 30,45±4,84µm e largura 3,52±0,33µm) e no rim esporos de Myxobolus sp. (comprimento total 8,94±0,82µm e largura 5,59±0,39µm). Na análise histopatológica das brânquias observaram-se plasmódios contendo esporos de Henneguya sp., com localização intralamelar e intravascular, em diferentes estágios de desenvolvimento. No rim identificaram-se esporos de Myxobolus sp., na região peritubular e no interstício e glomérulo, circundados por melanomacrófagos. Em poucos casos foi registrada hemorragia focal. O uso da coloração de Ziehl-Neelsen permitiu identificar particularidades dos esporos, facilitou sua biometria e classificação em comparação com a hematoxilina-eosina, demonstrando sua utilidade no diagnóstico histopatológico da referida parasitose.