Reduced stress and increased immune responses in Nile tilapia kept under self-feeding conditions

ABSTRACT: Stress and immune responses were studied in Nile tilapia Oreochromis niloticus under both conditions of self-feeding and scheduled feeding. Self-feeding fish had a significantly lower cortisol level in their blood plasma than that of scheduled feeding fish. Furthermore, skin color was found to be paler in the self-feeding fish compared with that of the scheduled feeding fish. Thus, the self-feeding fish had a less stressful physiological status than the other experimental fish. Immunological analyses revealed that the self-feeding fish significantly exceeded the scheduled feeding fish in the phagocytic activity of their macrophages, antibody production, and the number of blood-circulating lymphocytes. The higher immune responses of self-feeding fish seem to be attained through a less stressful environment. This self-feeding behavior could be exploited to create reduced stress for disease control in fish farming.     

Silver catfish Rhamdia quelen immersion anaesthesia with essential oil of Aloysia triphylla (L'Hérit) Britton or tricaine methanesulfonate: effect on stress response and antioxidant status

Responses to anaesthesia with essential oil (EO) of Aloysia triphylla (135 and 180 mg L^?1) and tricaine methanesulfonate (MS222) (150 and 300 mg L^?1) were assessed in silver catfish. Exposure to the anaesthetics elicited a stress response in the species. In the case of MS222, it was displayed as a release of cortisol into bloodstream, elevation in hematocrit and plasma ion loss. The EO presented cortisol‐blocking properties, but increased haematocrit and disturbances of hydromineral balance were observed. Liver antioxidant/oxidant status of EO and MS222‐anaesthetized silver catfish was also estimated. The synthetic anaesthetic induced lipoperoxidation, notwithstanding increased catalase contents, whereas the naturally occurring product was capable of preventing the formation of lipid peroxides, possibly due to combined actions of catalase and glutathione‐S‐transferase. Anaesthetic efficacy was also tested via induction and recovery times. Overall, the promising results obtained for the physiological parameters of the EO‐treated fish counterbalanced the slight prolonged induction time observed for 180 mg L^?1. As for 135 mg L^?1, both induction and recovery times were lengthy; despite that, the EO was able to promote oxidative protection and mitigate stress. None of the MS222 concentrations prompted such responses concomitantly.     

Nutrient composition and sensory evaluation of fillets from wild-type Nile tilapia (Oreochromis niloticus, Linnaeus) and a red hybrid (Florida red tilapia × red O. niloticus)

A comparison of composition and sensory evaluation were performed on fillets of Oreochromis niloticus (wild type) and red hybrid tilapia (RHT) (Florida red tilapia ♂× red O. niloticus♀). Fifty male tilapia, 25 O. niloticus (initial weight 159.3 g) and 25 red hybrid (initial weight 132.4 g), were placed in each of three 2.0 m³ volume tanks. The fish were fed a commercial feed containing 35.9% crude protein during the 120‐day experimental period and then killed in cold water (3°C). All the fish were filleted. Twelve fillets from each treatment were used for proximate analyses, five fillets from each treatment were used for fatty acids analyses and the remaining fillets were used for sensory evaluation. The compositional analyses showed similar moisture, true protein and ash content in both genetic groups, but a lower crude fat content was measured in the red hybrid fillets (0.70%) compared with that of O. niloticus fillets (0.97%). No differences between O. niloticus and the red hybrid were observed in their fillet profile of fatty acids. In the sensory evaluation test, a difference in fillet flavour between both genetic groups was perceived. Of the 112 consumers in the preference test, 81.2% perceived a difference in fillet flavour between the two tilapias, with a general preference for the red hybrid over the wild‐type O. niloticus. The benefits of cultivating a RHT with a low lipid content are discussed.     

Anesthetic activity of the essential oil of Aloysia triphylla and effectiveness in reducing stress during transport of albino and gray strains of silver catfish,Rhamdia quelen

This study investigated the efficacy of the essential oil (EO) of Aloysia triphylla as an anesthetic for albino and gray strains of silver catfish, Rhamdia quelen. Juveniles were exposed to concentrations between 20 and 800 μL L^?1 EO of A. triphylla to evaluate time of induction and recovery from anesthesia. In another experiment, both strains were divided into four groups such as 0 (control), 30, 40, or 50 μL L^?1 EO and transported for 5 h. The longest time for anesthetic induction and recovery was observed in the albinos. Both strains reached anesthesia in the 100–800 μL L^?1 (11.1–1.24 min) range, without mortality, being 200 μL L^?1 the best response considering time to anesthesia (5.35 min). Albinos transported with all EO concentrations showed higher values of carbon dioxide in the water of transport, but lower levels were observed in grays transported with 40 and 50 μL L^?1 EO when compared to control fish. The same concentrations did not prevent significant whole-body cortisol rise at the end of transport in the albino strain. Juveniles of both strains transported with EO presented lower ion loss to the water compared to control fish. The EO of A. triphylla is an effective anesthetic for albino and gray silver catfish. This EO increases whole-body cortisol levels in the albino strain, but as it reduces net ion loss as in the gray strain, it can be also recommended for transport.     

Chemical composition of the essential oil of Aloysia triphylla under seasonal influence and its anaesthetic activity in fish

Plants are sources of secondary metabolites, and some are rich in essential oils (EOs), which possess a bioactive pharmacological importance as sedatives and anaesthetics that have been experimentally demonstrated in fish. In search of products with lower residual effects and optimization of the production based on the local flora, the properties of the EO of Aloysia triphylla (EOAT) have been investigated. The samples of two years were analysed for the percentage variability of the chemical composition and the totality of the major components according to the season. Limonene, citral isomers (α and β), caryophyllene, caryophyllene oxide and spathulenol were the main constituents of the EOAT throughout these two years. In the second year, caryophyllene predominates over its oxide in all seasons; however, in the first year, it occurs only in autumn and winter. Seasonal pools of EOs were made for the biennium obtaining four samples, and silver catfish (Rhamdia quelen) juveniles were subjected to a concentration range of 20–800 μl/L EOAT in order to assess induction and recovery times of anaesthesia. The relationships between percentages of individual components in EOAT and time to induce deep anaesthesia indicate that its activity may be attributed to the association of several of its components.     

Essential oils in diets for Nile tilapia juveniles: Productive performance and plasmatic biochemistry

The objective of this study was to evaluate the inclusion of increasing levels of a commercial mix of essential oils (EOs) from cashew (Anacardium occidentale) and castor bean (Ricinus communis) as an additive in diets for Nile tilapia. Thus, 240 juveniles with an initial weight of 35.06 ± 1.02 g were distributed in a completely randomized design (four treatments and six replicates) in 24 cylindrical‐tapered tanks; the fish were fed for 60 days, four times a day, diets containing increasing levels of EO (0.10, 0.15 and 0.20%) and a control diet (without the additive). After the experimental period, the animals were desensitized, and an aliquot of blood was collected for the biochemical analysis; they were subsequently euthanized for the evaluation of productive performance and fillet chemical composition. No significant differences (p > .05) were observed in the analyses of productive performance, fillet chemical composition, and triglycerides, VLDL, total proteins, urea and glucose. However, significant differences (p < .05) were observed in the analyses of total cholesterol, HDL and LDL. Inclusion above 0.15 of the additive showed an improvement in the levels of lipoprotein carriers, increasing HDL levels and decreasing LDL values, without interfering with the fish productive performance.     

Partial replacement of fish oil by flaxseed oil in Atlantic halibut (Hippoglossus hippoglossus L.) diets: effects on growth,nutritional and sensory quality

Three isonitrogenous (520 g protein kg^?1 DM) and isoenergetic (25 MJ kg^?1 DM) diets containing increasing levels of flaxseed oil (FxO; 0%, 40% and 70% of total added oil) at the expense of fish oil (FO) were tested for 33 weeks in groups of 61 individually PIT‐tagged halibut (initial weight, 849 ± 99 g). Effects on fish growth performance, fillet nutritional and sensory quality were determined. Specific growth rate (0.2% day^?1), feed conversion ratio (1.2–1.3) and nitrogen and energy retention were not affected by dietary treatments. Dietary fatty acid composition was reflected in fatty acid profiles of halibut muscle, liver and heart. Muscle of fish fed FxO diets contained higher 18:2n‐6 and 18:3n‐3 concentrations whereas 20:5n‐3 and 22:6n‐3 levels were significantly reduced. However, increasing FO replacement induced preferential retention of 22:6n‐3 especially in heart, and a trend for 20:5n‐3 conservation in heart and muscle was observed. FO replacement did not affect colour, texture and the characteristic fish odour and flavour of cooked fillets. By selectively retaining long‐chain polyunsaturated fatty acids halibut can adapt to a lower dietary supply without adverse effects on growth, feed conversion ratio, survival, and fillet nutritional and sensory quality.     

The use of clove oil, metomidate, tricaine methanesulphonate and 2-phenoxyethanol for inducing anaesthesia and their effect on the cortisol stress response in black sea bass (Centropristis striata L.)

Juvenile and adult black sea bass (Centropristis striata L.) were exposed to various concentrations of four anaesthetics to determine practical dosages for handling as well as for procedures such as bleeding, ovarian biopsy or tag implantation. In experiment 1, juveniles exposed to either 2.0 mg L^?1 metomidate, 15 mg L^?1 clove oil, 70 mg L^?1 tricaine methanesulphonate (TMS) or 200 mg L^?1 2‐phenoxyethanol (2‐PE) reached stage II of anaesthesia in 3–5 min and could be handled for weighing and measuring. All fish had completed recovery to stage III within 6 min. In experiment 2, the established concentrations of each anaesthetic were tested on juveniles to determine their ability to prevent a reflex to a subcutaneous needle puncture. All of the fish exposed to clove oil (20 mg L^?1) and 40% of the TMS‐treated (70 mg L^?1) fish reacted while none of the fish anaesthetized in metomidate (2.0 mg L^?1) or 2‐PE (200 mg L^?1) responded to the needle puncture. In experiment 3, metomidate (5.0 mg L^?1), clove oil (30 mg L^?1) TMS (125 mg L^?1) or 2‐PE (300 mg L^?1) were all effective for performing an ovarian biopsy or tag implantation on adults. In experiment 4, TMS (125 mg L^?1) exacerbated the cortisol response to a short handling stressor during a 30 min exposure. Fish anaesthetized in 2‐PE (300 mg L^?1), metomidate (5.0 mg L^?1) or clove oil (40 mg L^?1) had increased cortisol levels associated with the handling stressor but there were no further increases during the remainder of the experimental period. The results demonstrate that these anaesthetics are effective for sedation and anaesthesia of black sea bass and that the best choice is dependant upon the procedures to be performed.     

High stocking density produces crowding stress altering some physiological and biochemical parameters in gilthead seabream, Sparus aurata, juveniles

High stocking density has been shown to produce a wide variety of effects on cultured fish populations, such as alterations in behavior and poor feed utilization, resulting in mortality and poor growth. High stocking density has also been reported to produce chronic stress and mobilization of energy sources in fish. There are few studies focusing on the effect of high stocking density on one of the most important marine fish species for Mediterranean aquaculture, gilthead seabream (Sparus aurata). Consequently, this study investigates the effect of high stocking density on juveniles of this species, focusing on effects of stocking density on growth, biochemical composition, immune status and hematology.The results of this study showed that high stocking density produced a chronic stress situation. Plasma cortisol levels increased up to 16.25 ng ml–1 in fish held at high stocking; this value was 4 fold that of fish held at low stocking; density (3.91 ng ml–1 ). As a consequenceof this stressful situation, there was both haemoconcentration and a decrease in alternative complement pathway (ACP), an important component of the immune system of fish (from 167.23 U ml–1 down to 146.37 U ml–1). Haematocrit, haemoglobin concentration and red blood cell count were significantly higher in fish held at high stocking density (43.87%, 10.76 9dl–1 and 3.36 ×106 mm–3, respectively) compared with those fish held at low stocking density (37.21%, 9.32 g/dlg–1 and 2.82 ×106 mm–3, respectively).In addition, high stocking density produced a decrease in hepatosomatic index (from 2.26 down to 2.04) and altered liver fatty acid composition. Oleic acid (18: 1n-9) decreased in liver total lipids of fish held at high stocking density and arachidonic acid (20: 4-n6) and n-3 high unsaturated fatty acids (n-3 HUFA) were reduced in liver polar lipids of those fish. These alterations reflect the effect of stocking density on lipid metabolism to help meet the increased energy demand.     

Three strategic feeding during hatchering of Nile tilapia: effects on organs integrity,parasitism and performance parameters

The effect of natural feed, artificial feed, and artificial feed with hormone 17 alpha-methyltestosterone on gill and liver integrity, parasite prevalence and performance parameters during the masculinization phase of Nile tilapia Oreochromis niloticus were evaluated. The experiment consisted of the three feeding strategies in triplicate. Water parameters, histopathological, parasitological and performance analyses were performed on day 7, 14, 28 and 60. Both artificial feeds (with and without hormone) were harmful to the hepatic and branchial integrity. The most significant change in liver was lipid degeneration, while in gills a significantly higher lamellar edema was observed when compared to fish that received the natural feed. Fish fed with the artificial diet (with and without hormone) showed a high prevalence (p < 0.05) of trichodinids and Cryptobia, while fish that received natural feed showed no parasites after 28 days of study. Fries that received diet with hormone experienced damage to their health, which decreased the survival of this group. The anabolic effect of this molecule, however, improved significantly (p < 0.05) their productive performance. Despite the absence of deleterious effects to fish health, the use of natural feed has a high risk of introducing predators. Introduction of odonate larvae drastically affected the survival of this group at the end of the study. All feeding strategies investigated have positive and negative effects that were discussed in the present study.     

Development and evaluation of a stress challenge testing methodology for assessment of Nile tilapia (Oreochromis niloticus, Linn.) fry quality

A decline in the quality of the freshwater fish seed available to small-scale farmers in Asia has been identified as an important constraint on the development of aquaculture in the region. There is a need for simple and reliable methods of determining the quality of seed at the farm gate or hatchery. The present study was carried out, in the field, to develop and evaluate a stress challenge test for Nile tilapia (Oreochromis niloticus, Linn.) fry as the effects of stress on fish performance have been well documented and similar tests are used elsewhere in the sector. Formalin and iodized salt were used as reference toxicants. LC₅₀ values were estimated using mortality figures resulting from exposure of marketable-sized fry to saline and formalin solutions. The two test concentrations, 24 ppt for saline and 500 p.p.m. for formalin, were used as reference toxicants in acute, static bioassays to challenge fry reared under different conditions during the post sex-reversal, nursery period. Using existing farm practice as controls the conditions were: 50% increased stocking density, formalin treatment, 40% reduction in feed rate and no supplementary feed. Fry length proved to be a very significant co-variable; adjusted means showed no significant difference between treatments. The importance of only marketing tilapia fry weighing over 0.2 g was emphasized. It was recommended that the challenge testing approach could be modified, but that it was vulnerable to unquantifiable and uncontrollable factors arising from management and the environment, which may have altered responses to the stressor.     

Growth response and acquired resistance of Nile tilapia, Oreochromis niloticus (L.) that survived Streptococcus iniae infection

This study determined the growth performance and acquired resistance of Nile tilapia, Oreochromis niloticus (L.) that survived Streptococcus iniae infection. Tilapia were challenged with three doses of S. iniae (8.8 × 10³, 8.8 × 10⁴ and 8.8 × 10⁵ CFU fish^?1 for low, medium and high challenges respectively). Groups of non‐injected and tryptic soy broth‐injected fish were maintained as controls. Significantly (P<0.05) higher mortality (45.0%) occurred in the high challenge treatment than in the low challenge treatment group (29.6%). The medium challenge group had mortality (36.3%) that did not differ significantly from the high or low treatment. Few fish died in the non‐injected and broth‐injected treatments (3.4% and 0.8% respectively). The tilapia that survived S. iniae infection used to assess growth performance were selected from survivors without gross clinical signs of disease. These fish were randomly stocked at a rate of 30 fish into each 57 L aquarium in triplicate and fed to apparent satiation for 8 weeks. No significant differences were detected in weight gain, feed intake, feed efficiency ratio or survival between S. iniae‐survived tilapia and the control treatments following the 8‐week growth performance trial. Following the 8‐week feeding study, tilapia were challenged with 1 × 10⁶ CFU fish^?1 of S. iniae to assess acquired immunity. Mean cumulative mortality was significantly higher (P<0.05) in the control treatments (41.7% for the non‐injected and 43.3% for the broth‐injected fish) than in the low, medium and high challenge treatments (7.4%, 3.3% and 8.3% respectively). Serum protein was significantly (P<0.05) elevated in the S. iniae‐survived tilapia that were subsequently challenged when compared with controls challenged for the first time. Agglutinating antibody titre was significantly higher in the fish in the medium and high challenge treatments, compared with the control fish challenged for the first time. The results suggest tilapia that survive S. iniae challenge without showing overt disease signs performed as well as non‐infected tilapia. Further, the S. iniae‐survived tilapia challenged following the 8‐week growth performance trial gained acquired resistance to homologous S. iniae challenge.     

Effect of L‐tryptophan supplemented diets on serotonergic system and plasma cortisol in Totoaba macdonaldi (Gilbert, 1890) juvenile exposed to acute stress by handling and hypoxia

Brain serotonin and cortisol play a central role in integrating the neuroendocrine response to stress. Previous studies in aquatic species show that dietary supplementation with the 5‐HT precursor L‐tryptophan amino acid is able to modulate this response. The aim of this study was to evaluate the effect of dietary tryptophan (Trp) on the serotonergic system and acute stress response in Totoaba macdonaldi. Four groups of fish were fed with one of four isonitrogenous and isocaloric diets formulated with different Trp content: 0.5% (1X TRP, control diet), 1.1% (2X TRP), 1.7% (3X TRP) and 2.3% (4X TRP). After a feeding period of 21 days, groups were exposed to acute stress by handling with a net or hypoxia. Feeding with Trp‐enriched diets resulted in increased 5‐HT content in telencephalon of undisturbed fish. Handling increased the 5‐hydroxyindole acetic acid (5‐HIAA) and serotonin turnover (5‐HIAA/5‐HT) in telencephalon of fish fed with 1X TRP and 2X TRP but was not significant for the fish fed 3X TRP and 4X TRP, and no significant increase was observed by hypoxia in both telencephalon and hypothalamus in any of the diets. An inverted U‐shaped response was observed in plasma cortisol increasing with 2X TRP after both handling and hypoxia and decreasing with diets rich in Trp, 3X TRP and 4X TRP. A higher increase was observed in haematocrit, plasma glucose and lactate in hypoxia than the handling groups. Our results indicate that Trp supplementation modulated cortisol response and mediated negative feedback of 5‐HT system in telencephalon after acute stress.     

Transport stress in Atlantic salmon (Salmo salar L.): anaesthesia and recovery

The effects of metomidate anaesthesia on levels of plasma cortisol, glucose, haematocrit and chloride in Atlantic salmon (1⁺) (Salmo salar L.), after a 2‐h transport and during a 48‐h recovery period were investigated. The use of metomidate anaesthesia during transport led to a reduced release of cortisol and significantly lower levels of plasma cortisol after a 48‐h recovery period. Plasma glucose did not return to basal level after a 48‐h recovery period, indicating that even longer recovery may be needed for the fish to return to a pre‐stress state. The results show that metomidate anaesthesia combined with a recovery period lessens the stress burden imposed by hauling and transport.     

Comparison of faecal collection method with high- and low-quality diets regarding digestibility and faeces characteristics measurements in Nile tilapia

The need for unbiased digestibility estimation has led to the development of a number of faeces collection methods. However, there is still a large variation in apparent digestibility coefficient (ADC) of diet/ingredients between these methods. This study investigated the impact of dietary quality on ADC and faeces recovery measurements obtained by two faecal collection methods (Choubert and settling tank). The fish were fed five diets: a control diet as high‐quality diet, two levels of dried duckweed (20% and 40%) and two levels of fresh duckweed (20% and 40%), which were added to the control diet, as low‐quality diets. Apparent digestibility coefficient estimates were highly correlated (r>0.95) between both faecal collection methods. For all diets the ADC estimates were higher when using settling tanks compared with Choubert collectors. For the control diet differences in ADC between two collectors were small, but by inclusion of duckweed these differences increased. The recovery percentage of faeces was not correlated between both faecal collection methods (r=0.22, P=0.41). The estimated recovery percentage of faeces by settling tanks was about three times higher than by Choubert. In conclusion, the differences in ADC and faeces recovery measured by two faecal collection methods depend on diet quality.     

Effects of cortisol on growth and development in tilapia larvae,Oreochromis mossambicus

Treatment of one-day-old yolksac larvae of tilapia (Oreochromis mossambicus), by immersion in cortisol solution for two weeks, significantly enhanced growth. The standard length, tail length, head width and wet weight showed dose-dependent increases at concentrations of cortisol (hydrocortisone) from 0.05 ppm to 0.5 ppm, with no further increase at 1.0 ppm. However, cortisol at 5.0 ppm retarded the growth of the larvae without causing any morphological abnormality. Yolk absorption and the onset of free-swimming activity in the larvae were also accelerated by cortisol treatment.     

Effects of different photoperiods on growth, stress and haematological parameters in juvenile great sturgeon Huso huso

The effects of constant light (24L:00D), no light (00L:24D) and two light–dark periods (18L:06D;12L:12D) on the growth, stress and haematological variables were assessed in juvenile great sturgeon, Huso huso . During the 8-week experimental period, juveniles (22.5 ± 0.6 g) were kept under a 150 lx light intensity in fibreglass tanks (0.8 m², 500 L). Differences in growth were insignificant during the experiment, but lactate levels were higher in the 00L:24D and 24L:00D photoperiods compared with 12L:12D and 18L:06D photoperiods. Cortisol levels did not show differences among the various photoperiods. At the end of the experimental period, fish reared under a 12L:12D photoperiod had higher haemoglobin values and erythrocyte numbers than in the other photoperiods, while no differences were found between groups with regard to haematocrit values or leucocyte numbers. The highest survival rate (89%) was observed in the 12L:12D period in which the levels of lactate and cortisol as stress indicators were minimal. The results indicate that various photoperiods cause different stress levels in juvenile great sturgeon and have no significant effects on growth, at least in short time periods.     

Digestibility, faeces recovery, and related carbon, nitrogen and phosphorus balances of five feed ingredients evaluated as fishmeal alternatives in Nile tilapia, Oreochromis niloticus L.

This study shows that alternatives for fishmeal in a fish diet affect not only fish growth but also faeces stability and nitrogen (N) and phosphorus (P) waste production. Wheat gluten diet (WGD), soybean meal extract diet (SBE), soybean meal diet (SBM), duckweed diet (DWD) and single‐cell protein diet (SCP) were evaluated as a fishmeal replacement on a 15% weight weight^?1 basis in tilapia diets. Fishmeal replacement affected dry matter (dm), protein, ash and P digestibility significantly. Faeces recovery (6.8–11.2%) was not significantly affected, although the amount of non‐recovered faeces and total faeces showed significant differences. Duckweed diet and SCP resulted in the largest amounts of non‐recovered and total faeces (199–210, 224–225 g dm kg^?1 feed dm). Compared with fishmeal diet (FMD), the WGD and SBE resulted in similar growth, but higher non‐faecal N losses (471–495 vs. 416 g N kg^?1 N). Soybean meal diet, DWD and SCP resulted in lower growth but less non‐faecal loss (409–450 g N kg^?1 N). The DWD and FMD had the highest N retention (480 g N kg^?1 N) compared with the other diets (431–451 g N kg^?1 N). Carbon retention, faecal and non‐faecal losses and P retention were similar for all diets (302–358, 142–176 and 489–523g C kg^?1 C, 606–704 g P kg^?1 P). Phosphorus faecal loss was lower for all diets (329–381 g P kg^?1 P) than for the FMD (401 g P kg^?1 P).