The present study evaluated the assay to quantify the respiratory burst activity of blood leukocytes of pacu as an indicator of the innate immune system, using the reduction of nitroblue tetrazolium (NBT) to formazan as a measure of the production of reactive oxygen species (ROS). In order to assess the accuracy of the assay, fish were challenged by Aeromonas hydrophila and sampled one week after challenge. The A. hydrophila infection increased the leukocyte respiratory burst activity. The protocol showed a reliable and easy assay, appropriate to determine the respiratory burst activity of blood leukocytes of pacu, a neotropical fish, in the present experimental conditions. Keywords: immune methodology, fish immunology, cellular immunity.
Atividade respiratória de leucócitos como indicador de imunidade inata de pacu (Piaractus mesopotamicus)Resumo O presente estudo avaliou o ensaio para quantificar a atividade respiratória dos leucócitos do sangue de pacu como um indicador do sistema imune inato, usando a redução do nitroazul tetrazólio (NBT) a formazan como medida da produção de espécies reativas de oxigênio (EROs). Para avaliar a precisão do ensaio, peixes foram desafiados por Aeromonas hydrophila e amostrados uma semana após o desafio. A infecção com A. hydrophila aumentou a atividade respiratória dos leucócitos. O protocolo se mostrou confiável e de fácil aplicação, apropriado para determinar a atividade respiratória de leucócitos do sangue do pacu, peixe neotropical, nas condições experimentais apresentadas.Palavras-chave: metodologia, imunidade de peixe, imunidade mediada por células.
The oxygen consumption (MO2) of two groups of 10° C acclimated steelhead trout Oncorhynchus mykiss was measured for 72 h after they were given a 100 µl kg(-1) intraperitoneal injection of formalin-killed Aeromonas salmonicida (ASAL) or phosphate-buffered saline (PBS). In addition, plasma cortisol levels were measured in fish from both groups prior to, and 1 and 3 h after, they were given a 30 s net stress. The first group was fed an unaltered commercial diet for 4 weeks, whereas the second group was fed the same diet but with 0·5% (5 g kg(-1) ) Aloe vera powder added; A. vera has potential as an immunostimulant for use in aquaculture, but its effects on basal and acute phase response (APR)-related metabolic expenditures and stress physiology, are unknown. Injection of ASAL v. PBS had no measurable effect on the MO2 of O. mykiss indicating that the APR in this species is not associated with any net increase in energy expenditure. In contrast, incorporating 0·5% A. vera powder into the feed decreased routine metabolic rate by c. 8% in both injection groups and standard metabolic rate in the ASAL-injected group (by c. 4 mg O2 kg(-1) h(-1) ; 5%). Aloe vera fed fish had resting cortisol levels that were approximately half of those in fish on the commercial diet (c. 2·5 v. 5·0 ng ml(-1) ), but neither this difference nor those post-stress reached statistical significance (P > 0·05).
This study verified the effects of CaSO 4 on physiological responses of the tropical fish matrinxa˜Brycon amazonicus (200.2 ± 51.1 g) in water containing CaSO 4 after a 4-h transportation at concentrations of: 0, 75, 150, and 300 mg L )1 . Blood samples were collected prior to transportation (initial levels), immediately after packaging, at arrival, and 24 h and 96 h after transportation (recovery). Cortisol levels increased after packaging (118.2 ± 14.2 ng ml )1 ), and decreased slightly after transportation in water containing CaSO 4 (106.8 ± 14.1), but remained higher than initial levels (21.0 ± 2.6 ng ml )1 ). Fish kept at 150 mg L )1 CaSO 4 reached the pre-transportation levels at 24 h of recovery. Blood glucose increased after transportation in all treatments (8.2 ± 0.2 mmol L )1 ) and declined after full recovery to values below initial levels (4.8 ± 0.1 mmol L )1 ). Chloride levels did not change in CaSO 4 treatments; serum sodium concentrations decreased after packaging and after transportation. Serum calcium levels did not differ among treatments, but decreased after packaging and increased at 96 h of recovery. Hematocrit and the number of red blood cells were higher in all treatments after packaging and arrival, except in fish exposed to 300 mg L )1 CaSO 4 . Mean corpuscular volume increased in 75 mg L )1 CaSO 4 , which reached the higher VCM after transportation. Hemoglobin levels increased only after transportation, regardless of calcium sulfate levels. Handling before transportation and transportation itself were both stressful to fish; calcium sulfate at concentrations tested in the present work had a moderate influence in the reduction of stress responses.
Tested was the efficiency of dietary metyrapone (MTP) in the inhibition of cortisol biosynthesis, as a tool to elucidate the effects of this hormone in the tropical fish pacu (Piaractus mesopotamicus H.) during air-exposure-induced stress. Moreover, MTP efficiency was shown to be affected by both the dose and time of administration and that a daily dose of 30 mg kg À1 body weight for 10 days reduced the levels of circulating cortisol in stressed fish. The tested method of MTP administration, including the concentration and the time of administration, efficiently regulated cortisol synthesis in a non-invasive manner.U.S.
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