Effects of chlorantraniliprole insecticide on innate immune response of silver catfish (Rhamdia quelen) naturally infected with Aeromonas hydrophila.

It is well documented today that pesticides, used in crop production, may modulate the immune system of healthy fish. However, there is still only limited information regarding the effects of these anthropogenic stressors in conjunction with natural stressors (pathogens), on the innate immune responses of freshwater fish. Thus, the aim of this investigation was to compare the innate immune response of two groups of fish (Rhamdia quelen), naturally infected with Aeromonas hydrophila, exposed and unexposed to a non-lethal concentration of chlorantraniliprole (CAP) insecticide (0.0 and 1.3 µg/L/24 h). Unhealthy fish exposed to CAP showed significant higher total leukocyte counts and neutrophils percentage compared to non-exposed infected fish). However, the monocytes and eosinophils percentage significantly decreased in fish exposed to CAP. Furthermore, lysozyme activity values measured in plasma, skin mucus, gill and intestine significantly reduced in fish exposed to CAP. The CAP-induced immunomodulation may interfere on the ability of the animal to heal or fight the infection, and possible contribute to the spread of bacterial infection in fish production or environment.

The effect of ß-1,3-glucan derived from Euglena gracilis (Algamune™ ) on the innate immunological responses of Nile tilapia (Oreochromis niloticus L.).

Algamune™ is a commercial additive produced from Euglena gracilis, providing a rich source of the ß-1,3-glucan paramylon. Isolated kidney phagocytes of Nile tilapia were incubated with graded doses (0, 8, 16, 32, 64 and 128 µg/ml) of Algamune™ and purified paramylon to gauge their ability to elicit the production of reactive oxygen species. A linear response was observed for extracellular superoxide anion for both sources but only Algamune™ for intracellular superoxide anion. After corroborating the immunostimulant properties ex vivo, a feeding trial was conducted to evaluate the dietary supplementation of Algamune™ (0, 100, 200, 400 and 800 mg/kg of diet) for Nile tilapia. Fish were fed for 3 weeks, after which, fish were sampled for blood and head kidney phagocytes. The remaining fish were challenged with Streptococcus iniae. Macrophage extracellular superoxide anion production was significantly elevated in fish fed diets with 200 mg of Algamune™ kg-1 when compared to fish fed the basal diet. Even though the disease challenge did not show statistical differences, it is worth mentioning that fish fed intermediate doses of Algamune™ had lowest numerical mortality values. Therefore, Algamune™ was demonstrated to enhance some immunological responses of tilapia both in ex vivo and in vivo.

Aloysia triphylla essential oil as additive in silver catfish diet: Blood response and resistance against Aeromonas hydrophila infection.

The essential oil of Aloysia triphylla (EOAT) is a promising product with potential use in aquaculture systems. This study evaluated hematological/biochemical responses and survival of silver catfish (Rhamdia quelen) fed a diet containing EOAT and infected by Aeromonas hydrophila. After 21 days of feeding trial, fish were infected with A. hydrophila following a 10-day period of observation. Blood collection was performed before and after the bacterial challenge. Dietary EOAT by itself seems to affect some blood parameters, decreasing total leukocyte, lymphocyte, and neutrophil counts and increasing total protein values. However, 2.0 mL EOAT/kg diet showed a possible potential protective effect after A. hydrophila infection, maintaining the evaluated parameters similar to basal values (from healthy fish before the feeding trial) and promoting survival of silver catfish.

S-(+)- and R-(-)-linalool: a comparison of the in vitro anti-Aeromonas hydrophila activity and anesthetic properties in fish.

Linalool is the main compound of many essential oils and occurs in two isomeric forms: S-(+)- and R-(-)-linalool. This study aimed to determine if linalool isomers have different antimicrobial and anesthetic properties in fish. For this purpose, these compounds were previously isolated from Lippia alba (Mill.)N. E. Brown and Ocimum americanum L. essential oils. Antimicrobial effects were evaluated through the microdilution test against Aeromonas hydrophila, an important fish disease etiologic agent. Induction time until sedation, anesthesia and recovery time were determined in silver catfish (Rhamdia quelen) through bath exposure (60, 180, 300 or 500 µL L-1). The results showed different biological properties for the isomers being S-(+)-linalool the only active against A. hydrophila at 3.2 mg mL-1. The sedation was induced without differences between the compounds, however R-(-)-linalool promoted faster anesthesia. There were no differences regarding the recovery time of the animals exposed to the linalool isomers. Although both S-(+)- and R-(-)-linalool can be used for sedative purposes, their use in A. hydrophila infection is inadvisable due to the high effective concentration. Considering anesthesia as the main objective, the R-(-)-linalool demonstrated clear advantages at lower concentration.

Evaluation of Ocimum americanum essential oil as an additive in red drum (Sciaenops ocellatus) diets.

This study evaluated productive parameters, whole-body composition, non-specific immune responses and pH and microbiota of digestive tract contents of red drum (Sciaenops ocellatus) fed diets supplemented with Ocimum americanum essential oil (OAEO) (0 - control, 0.25, 0.5, 1.0 and 2.0 g/kg diet). After 7 weeks no significant differences in productive parameters and whole-body composition were observed. Plasma and intestinal lysozyme measurements and pH of the stomach and intestine (6 h after feeding) did not show significant differences among groups. Intestinal microbial community in fish fed the basal and OAEO diets (all concentrations) were identical. However, red drum fed the diet with OAEO at 1.0 g/kg had significantly increased intraperitoneal fat deposition and stomach pH (2 h after feeding) and decreased superoxide ion production (NBT-test) compared to the control group. Hemolytic activity of the complement system increased in fish fed diets containing OAEO. Red blood cells from fish fed the lowest OAEO concentration (0.25 g/kg) showed significant lower fragility in erythrocyte osmotic fragility assay, but fish fed 0.5 and 1.0 g/kg showed significant higher erythrocyte fragility. Lysozyme measurement in the supernatant of stomach content was significantly higher in fish fed the diet supplemented at 0.5 g/kg. Based on these various results, OAEO at different supplementation levels did not influence growth performance and intestinal microbial community; however, the EO added to the diet showed effects on immunological responses of red drum.

Lippia alba essential oil promotes survival of silver catfish (Rhamdia quelen) infected with Aeromonas sp.

In vitro and in vivo activity of the Lippia alba essential oil (EO) against Aeromonas sp. was evaluated. In the in vitro assay the minimum inhibitory concentration (MIC) and a minimum bactericidal concentration (MBC) of EO for Aeromonas cells were determined using the microdilution method. Twenty five strains of Aeromonas sp. isolated from infected fish obtained from local fish farms were used. MIC and MBC values were 2862 and 5998 µg mL-1 for L. alba EO and 0.5 and 1.2 µg mL-1 for gentamicin, respectively. In the in vivo assay silver catfish juveniles (Rhamdia quelen) (7.50 ± 1.85 g and 10.0 ± 1.0 cm) with typical injuries associated to Aeromonas infection were divided into four treatments (in triplicate n=10): untreated fish (negative control), 10 mg L-1 of gentamicin, and 20 or 50 µL L-1 of EO. Fish were maintained in aerated 20 L plastic boxes. After 10 days survival of silver catfish infected with Aermonas sp. and treated with essential oil (50 µL L-1) was greater than 90%.

Humic acid of commercial origin causes changes in gill morphology of silver catfish Rhamdia quelen exposed to acidic water: Response of silver catfish gills to low pH and humic acid.

The effect of dissolved organic carbon (DOC) against the burden of low pH was assessed in silver catfish Rhamdia quelen through the gills. A commercial humic acid (HA) was used as the source of DOC at 0 (control), 10, 25, and 50 mg/l. For each HA concentration, two pH levels were tested: 6.5 (control) and 5.5. After 40 days of exposure, the gills were removed and morphological variables were analyzed through light and scanning electronic microscopy. The low water pH caused a reduction in the length of filaments, number of lamellae, and gill respiratory surface area (GRSA) and an increase in chloride cells (CCs) number in the filament epithelium. When HA was added to the test water, GRSA increased and CCs proliferated in lamellae, suggesting ionic and respiratory disturbances. Scanning electronic microscopy revealed that CC morphometric variables, that is, apical area, fractional area, and density, were undisturbed by the pH reduction, but increased in the presence of HA. This study indicates that the commercial HA did not protect silver catfish against low pH stress. Instead, it caused changes that may affect vital processes such as ion regulation and ventilation and, consequently, reduce growth.

The Essential Oil of Hyptis mutabilis in Ichthyophthirius multifiliis Infection and its Effect on Hematological, Biochemical, and Immunological Parameters in Silver Catfish, Rhamdia quelen.

This study evaluated the activity of leaf essential oil (EO) from Hyptis mutabilis as well as its major constituent, (-)-globulol, in infections by the parasite Ichthyophthirius multifiliis (ich). Effects on hematological, biochemical, and immunological parameters of silver catfish, Rhamdia quelen, exposed to the same samples also were evaluated. In the first experiment, naturally infected fish were treated with EO (0, 10, and 20 mg · L-1) and ethanol, using several methods of exposure. Fish mortality and the number of trophonts per fish were assessed after 48 and 96 hr. Hour-long daily baths resulted in optimal survival, so this methodology was used for the second experiment, in which infected animals were exposed to (-)-globulol at 2.5 and 5 mg · L-1. The most effective concentrations in Experiments 1 and 2 were chosen for Experiment 3, in which healthy animals were subjected to hour-long daily baths with EO (20 mg · L-1) or (-)-globulol (2.5 mg · L-1). Additionally, an in vitro experiment was performed with EO and globulol at the same concentrations of the in vivo test. EO and (-)-globulol increased the survival of fish infected with ich and altered certain hematological and biochemical parameters. After 4 days, levels of hematocrit, erythrocytes, and leukocytes increased significantly in healthy animals exposed to EO. Exposure to (-)-globulol increased leukocyte number alone. No significant differences in nonspecific immunological parameters were detected when treated groups were compared to controls, but the leukocytosis observed in EO- and globulol-treated healthy animals indicates that EO and (-)-globulol increased innate immunity in these fish. An in vitro antiparasitic effect was observed in both samples.

The use of nitazoxanide against the pathogens Ichthyophthirius multifiliis and Aeromonas hydrophila in silver catfish (Rhamdia quelen).

The antiparasitic and antibacterial activities of nitazoxanide (NTZ) against Ichthyophthirius multifiliis (Ich) and Aeromonas hydrophila, respectively, were evaluated in silver catfish juveniles (Rhamdia quelen). In the first experiment, fish naturally infected by Ich were treated with different concentrations of NTZ. Fish mortality and the number of trophonts/juveniles were evaluated. In the second experiment, the antimicrobial potential in vitro (MIC test) and preventive potential in vivo against A. hydrophila was evaluated. The treatment with 1.5mgL(-1) NTZ showed the best result against Ich, with a significant reduction of trophonts and fish mortality of 10%. NTZ did not show in vitro activity against A. hydrophila at concentrations tested, but showed a probable preventive activity in vivo. However, these results are preliminary and more studies should be conducted.

S-(+)- and R-(-)-linalool: a comparison of the in vitro anti-Aeromonas hydrophila activity and anesthetic properties in fish

ABSTRACT Linalool is the main compound of many essential oils and occurs in two isomeric forms: S-(+)- and R-(-)-linalool. This study aimed to determine if linalool isomers have different antimicrobial and anesthetic properties in fish. For this purpose, these compounds were previously isolated from Lippia alba (Mill.)N. E. Brown and Ocimum americanum L. essential oils. Antimicrobial effects were evaluated through the microdilution test against Aeromonas hydrophila, an important fish disease etiologic agent. Induction time until sedation, anesthesia and recovery time were determined in silver catfish (Rhamdia quelen) through bath exposure (60, 180, 300 or 500 μL L-1). The results showed different biological properties for the isomers being S-(+)-linalool the only active against A. hydrophila at 3.2 mg mL-1. The sedation was induced without differences between the compounds, however R-(-)-linalool promoted faster anesthesia. There were no differences regarding the recovery time of the animals exposed to the linalool isomers. Although both S-(+)- and R-(-)-linalool can be used for sedative purposes, their use in A. hydrophila infection is inadvisable due to the high effective concentration. Considering anesthesia as the main objective, the R-(-)-linalool demonstrated clear advantages at lower concentration.