Antibacterial potential of phytochemicals alone or in combination with antimicrobials against fish pathogenic bacteria.

AIMS: This study investigated the antibacterial activity of five phytochemicals (carvacrol, citral, eugenol, linalool and thymol) alone or in combination with florfenicol or oxytetracycline against bacteria isolated from silver catfish (Rhamdia quelen). We also analysed the potential of these compounds to inhibit biofilm formation and haemolysis caused by the bacteria. METHODS AND RESULTS: Bacteria were tested with antimicrobials to calculate the multiple antibiotic resistances. The checkerboard assay was used to evaluate a putative synergy between five phytochemicals and antimicrobials against the strains isolated. The biofilm formation inhibition assay was performed with phytochemicals and antimicrobials, and the haemolysis inhibition assay was performed with the phytochemicals. Carvacrol, eugenol and thymol were the most effective phytochemicals. Three combinations (linalool with florfenicol or oxytetracycline against Aeromonas hydrophila and citral with oxytetracycline against Citrobacter freundii) demonstrated synergy in the checkerboard assay. All phytochemicals inhibited biofilm formation and haemolysis activity. CONCLUSION: The tested phytochemicals showed satisfactory activity against fish pathogenic bacteria. SIGNIFICANCE AND IMPACT OF THE STUDY: The phytochemicals did not present antagonistic interactions with the antimicrobials, allowing their combined use, which may contribute to a decrease in the use of conventional drugs and their residues in aquatic environment.

The use of Ocimum americanum essential oil against the pathogens Aeromonas hydrophila and Gyrodactylus sp. in silver catfish (Rhamdia quelen).

UNLABELLED: The bactericidal activity (minimum inhibitory concentration (MIC)-test) of Ocimum americanum (inflorescences) essential oil (OAEO) against Aeromonas hydrophila was determined in this study. Also investigated was the potential of OAEO and the main compound found in the oil (linalool) at subinhibitory concentrations to be inhibitors of haemolysis caused by Aer. hydrophila in fish erythrocytes. An in vivo experiment was conducted to evaluate the survival of fish (Rhamdia quelen) experimentally infected with Aer. hydrophila and exposed to OAEO. A second experiment was conducted to evaluate the in vitro and in vivo activity of OAEO (mix from inflorescences and leaves) against the parasite Gyrodactylus sp. The OAEO showed weak in vitro activity against Aer. hydrophila (6400 µg ml(-1) ). Subinhibitory concentrations of OAEO (100 µg ml(-1) ) inhibited haemolysis (90%) caused by Aer. hydrophila in fish erythrocytes, however, linalool did not inhibit haemolysis activity. At the low concentrations (10 and 20 mg l(-1) ) added to the water, OAEO promoted the survival of fish experimentally infected with Aer. hydrophila. Lastly, the OAEO mix (50 mg l(-1) ) was effective against Gyrodactylus sp., significantly reducing (60%) the number of parasites in the fish. SIGNIFICANCE AND IMPACT OF THE STUDY: Phytochemicals, such as essential oils (EOs) are a great source of new molecules and have shown potential to be used in aquaculture systems. However, additional studies focused on the in vivo efficacy, mode of action and identification of the active compounds are needed. This study determined the potential of Ocimum americanum EO for use against two important fish pathogens, Aeromonas hydrophila and Gyrodactylus sp., as well as providing preliminary information about the role of the main EO compound (linalool) against Aer. hydrophila virulence.

In vitro effects of plant essential oils on non-specific immune parameters of red drum, Sciaenops ocellatus L.

Phytochemicals such as plant essential oils (EOs) have been reported to favour various activities in the innate immune system of fish. Thus, the aim of this study was to verify the in vitro effect of three different plant EOs (Ocimum americanum, Cymbopogon flexuosus and Melaleuca alternifolia) on non-specific immune parameters and erythrocyte osmotic fragility of red drum, Sciaenops ocellatus. Concentrations of each plant EO evaluated in preparations of head-kidney macrophages, blood leucocytes and blood plasma were as follows: 0.0 (control), 1.0, 2.0, 4.0, 8.0, and 16.0 µg/ml. Red drum head-kidney macrophages significantly increased extracellular superoxide anion production when exposed (20 h) to O. americanum EO (1.0-8.0 µg/ml) and C. flexuosus EO (2.0 and 4.0 µg/ml). The respiratory burst of blood leucocytes (NBT test) significantly increased in all concentrations when compared to the respective control group, for all EOs. At the highest concentration (16.0 µg/ml), C. flexuosus EO significantly inhibited the haemolytic activity of complement system in red drum blood after 1 h exposure. None of the tested concentrations significantly altered plasma lysozyme activity or erythrocyte osmotic fragility after exposing (1 h) red drum whole blood to each EO. This study demonstrated that these plant EOs are capable of triggering superoxide anion production in red drum leucocytes (head-kidney macrophages and/or blood leucocytes). In vivo studies are warranted to address their potential as immunostimulants in the diet of red drum and other aquacultured species.

Plant essential oils against Aeromonas hydrophila: in vitro activity and their use in experimentally infected fish.

AIMS: The aims of this study were to investigate the in vitro antibacterial activity of the essential oils (EOs) of Hesperozygis ringens (HREO), popularly known as 'espanta-pulga' and two different species of basil, Ocimum gratissimum (OGEO) and Ocimum americanum (OAEO), as well as, the potential of these products to be used in silver catfish (Rhamdia quelen) infected with Aeromonas hydrophila. METHODS AND RESULTS: OGEO and HREO showed better antibacterial activity in vitro. Subinhibitory concentrations of all EOs inhibited haemolysis caused by Aer. hydrophila in fish erythrocytes (100% reduction for OAEO at 100 µg ml(-1) and more than 90% for HREO and OGEO at 150 µg ml(-1) ). However, OAEO and HREO showed the best survival results (75 and 70% respectively) after their use as treatment (therapeutic baths-1 h daily/5 days) in silver catfish experimentally infected with Aer. hydrophila. A second in vivo assay using healthy fish was conducted to verify the potential of the EOs (preventive baths-1 h daily/5 days) to promote fish survival. Fish exposed to HREO and OAEO and their diluent (ethanol) showed significant lower haematocrit values and higher complement system activity compared to control. Plasma cortisol level was significantly higher in the groups exposed to both EOs. There was no significant difference in survival of silver catfish challenged with Aer. hydrophila after preventive baths with HREO, OAEO and control group. CONCLUSIONS: All tested EOs showed in vitro antibacterial properties against Aer. hydrophila and HREO and OAEO showed potential to be used in the treatment of infected fish. SIGNIFICANCE AND IMPACT OF THE STUDY: These products can be used in aquaculture as therapeutic and prophylactic agents against fish pathogens, with antimicrobial and/or immunostimulant properties.

Teores de proteína bruta em dietas práticas para juvenis de carpa capim / Crude protein levels in practical diets for grass carp juveniles

Avaliou-se a resposta de juvenis de carpa capim (Ctenopharyngodon idella) à inclusão de proteína bruta (PB) em dietas práticas, em um experimento com 120 juvenis e 80 dias de duração. As dietas eram compostas por farinha de carne suína, farelo de soja, milho e farelo de trigo com 22, 30, 36 ou 44 por cento de PB. Dez juvenis (153,0±1,5g) por tanque (850L) foram alimentados duas vezes ao dia (3 por cento da biomassa). Foi observado efeito linear positivo da PB para peso final, ganho em peso, taxa de crescimento específico, rendimento de carcaça e filé, coeficiente de retenção proteica, taxa de eficiência proteica, deposição de gordura e proteína e hematócrito. O mesmo efeito também foi observado para triglicerídeos e colesterol total no sangue, indicando que a proteína da dieta foi utilizada como fonte energética. Conversão alimentar aparente e gordura no peixe inteiro apresentaram efeito quadrático, com pontos de máxima em 40,6 e 37,1 por cento de PB, respectivamente. Pode-se concluir que a porcentagem mínima de PB para a carpa capim na fase de recria foi de 44 por cento e que a variação da proteína da dieta promoveu alterações no metabolismo dos juvenis de carpa capim, refletidos nos parâmetros sanguíneos e de carcaça.

The response of grass carp (Ctenopharyngodon idella) juveniles to crude protein (CP) levels in practical diets was evaluated in an experiment with 120 juveniles and 80 days of feeding. The diets containing porcine meat meal, soybean meal, corn and wheat meal, with 22, 30, 36, or 44 percent CP. Ten juveniles (153.0±1.5 g) per tank (850L) were fed twice daily with pelleted feed (3 percent of biomass). Linear effect of CP on final weight, weight gain, specific growth rate, carcass and fillet yield, coefficient of protein retention, protein efficiency rate, deposition of fat and protein, and hematocrit were observed. It was also observed effect on triglycerides and total cholesterol in blood, indicating that dietary protein was being used as energy source. Feed conversion ratio and fat content in whole fish showed quadratic effect, with maximum points at 40.6 and 37.1 percent CP, respectively. The best CP content for grass carp in growing phase was 44 percent, and the variation of protein content in diet changed the metabolism of grass carp juveniles, reflected in blood and carcass parameters.