Cannabidiol improves Nile tilapia cichlid fish welfare.

Cannabidiol (CBD) is a substance derived from Cannabis sativa, widely studied in medicine for controlling neural diseases in humans. Besides the positive effects on humans, it also presents anxiolytic proprieties and decreases aggressiveness and stress in mammals. Therefore, CBD has the potential to increase welfare in reared animals, as it seems to reduce negative states commonly experienced in artificial environments. Here, we tested the effect of different CBD doses (0, 1, 10 and 20 mg/kg) on aggressiveness, stress and reproductive development of the Nile tilapia (Oreochromis niloticus) a fish reared worldwide for farming and research purposes. CBD mixed with fish food was offered to isolated fish for 5 weeks. The 10 mg/kg dose decreased fish's aggressiveness over time, whereas 20 mg/kg attenuated non-social stress. Both doses decreased the baseline cortisol level of fish and increased the gonadosomatic index. However, CBD 1 and 10 mg/kg doses decreased the spermatozoa number. No CBD dose affected feeding ingestion and growth variables, showing that it is not harmful to meat production amount. Despite the effect on spermatozoa, CBD supplementation exhibits high potential to benefit animals' lives in artificial environments. Therefore, we showed for the first time that CBD could be used as a tool to increase non-mammal welfare, presenting a great potential to be explored in other husbandry and captivity species.

Differences in the Alcohol Preference Assessment of Shy and Bold Zebrafish.

Individuals differ in their preference for alcohol and propensity to develop alcoholism, where the behavioral profile, such as the bold-shy axis, plays an important role for such a difference. However, literature is limited and conflicting on the causes and consequences of this relationship. Translational studies using animal models, such as zebrafish, can help identify behavioral traits that predispose individuals to drink alcohol compulsively. Here, the preference for alcohol was investigated in two distinct traits in zebrafish: shy and bold. For this purpose, fish were separated into shy and bold traits and then a conditioned place preference paradigm was used, a strategy that allows the rewarding effects from alcohol to be assessed by the ability to enhance the animal's preference for an environment that initially was not preferred. It was found that bold zebrafish actively searched for the environment that was paired to alcohol after one acute exposure, whereas, shy fish changed their place preference even without alcohol administration, showing that the conditioned place preference protocol, given the short amount time to assess place preference, is not ample enough for shy fish to choose. Our results show that behavioral profiles must be considered in further studies since differences between shy and bold individuals on preference behavior can strongly interfere in the assessment of drug preference, mainly when using the conditioned place preference paradigm.

Toxicity of spike fragments SARS-CoV-2 S protein for zebrafish: A tool to study its hazardous for human health?

Despite the significant increase in the generation of SARS-CoV-2 contaminated domestic and hospital wastewater, little is known about the ecotoxicological effects of the virus or its structural components in freshwater vertebrates. In this context, this study evaluated the deleterious effects caused by SARS-CoV-2 Spike protein on the health of Danio rerio, zebrafish. We demonstrated, for the first time, that zebrafish injected with fragment 16 to 165 (rSpike), which corresponds to the N-terminal portion of the protein, presented mortalities and adverse effects on liver, kidney, ovary and brain tissues. The conserved genetic homology between zebrafish and humans might be one of the reasons for the intense toxic effects followed inflammatory reaction from the immune system of zebrafish to rSpike which provoked damage to organs in a similar pattern as happen in severe cases of COVID-19 in humans, and, resulted in 78,6% of survival rate in female adults during the first seven days. The application of spike protein in zebrafish was highly toxic that is suitable for future studies to gather valuable information about ecotoxicological impacts, as well as vaccine responses and therapeutic approaches in human medicine. Therefore, besides representing an important tool to assess the harmful effects of SARS-CoV-2 in the aquatic environment, we present the zebrafish as an animal model for translational COVID-19 research.

Welfare in Nile Tilapia Production: Dorsal Fin Erection as a Visual Indicator for Insensibility.

In aquaculture, to ensure animal welfare in pre-slaughter and slaughter stages, it is fundamental that fish are insensible. A method for evaluating fish insensibility is based on visual sensibility indicators (VSI) assessment (i.e., self-initiated behavior, responses to stimuli and reflexes). However, many stimuli used to assess fish responses are painful. Therefore, this study verifies whether the presence/absence of a dorsal fin erection (DFE) response can be used as a painless VSI in Nile tilapia (Oreochromis niloticus). Three stunning protocols were applied to fish: benzocaine anesthesia (40 mg/L and 80 mg/L), ice water immersion (0-1, 2-3 and 5-6 °C) and CO2 stunning. After these stunning methods were applied in fish, the time of loss and return of DFE was observed, along with the vestibulo-ocular reflex (VOR). All fish stunned using benzocaine and ice water immersion lose both VSIs, while 95% of fish stunned using CO2 lose these VSIs. In all treatments, DFEs return quicker than VOR. Therefore, DFE can be used as a VSI in Nile tilapia, which is simple for producers to assess and does not require a painful stimulus. However, the DFE alone does not totally ensure fish insensibility and must be used together with other well-established VSIs at fish farms.

The attraction trick: males in early stages of disease become more chemically attractive to Nile tilapia (Oreochromis niloticus) females.

In aquatic systems, olfaction plays an important role in acquiring information about the social environment and influences important behaviours in various contexts, including predator avoidance, foraging, aggressive and reproductive behaviour and mate selection. As the presence of diseases might modify individual odour, fish may use the variability in conspecifics' odours as an indicator of the health status and infectious load of potential mates. Here, female Nile tilapia were tested for their ability to detect infected males and discriminate between bacterial infected and uninfected individuals by means of chemical cues. Females were allowed to choose between the odours of males infected by Aeromonas hydrophila bacteria and uninfected males. The findings show that female Nile tilapia initially showed a preference for infected males in terms of their first choice in a dichotomous choice test, but the total duration of time spent with the stimulus from infected males was not longer than that for the uninfected males. This may indicate that males at early stages of infection, i.e., without advanced clinical signs of infection, emit odours that allow them to enjoy the benefits of socialization when the infection is not yet detected by conspecifics. Thus, in the context of reproduction, males might attract female partners and have some chance of reproducing, before being avoided.

Low cholesterol is not always good: low cholesterol levels are associated with decreased serotonin and increased aggression in fish.

The inverse relationship between serum cholesterol and levels of aggression led to the cholesterol-serotonin hypothesis. According to this hypothesis, low dietary cholesterol intake leads to depressed central serotonergic activity, which is associated with increased aggression. Here we present the hypothesis about the evolutionary origins of low cholesterol and aggressive behavior, investigating the relationship between low levels of plasma cholesterol and aggressive behavior in fish. We used Nile tilapia (Oreochromis niloticus), a species of aggressive fish with a clear dominant subordinate relation, as an experimental model. The fish were treated with statin, a cholesterol-lowering drug. Aggressive behavior, brain serotonin (5-HT) concentrations, 5-hydroxyindoleacetic acid (5-HIAA, the major 5-HT metabolite) and plasma cholesterol were analyzed after chronic administration of statin. Our results show that fish treated with statin exhibited reduced plasma cholesterol, reduced telencephalic indexes of 5-HIAA/5-HT and increased aggressive behavior compared to control fish. These results indicate that changes in plasma cholesterol may affect neurochemical processes underlying aggressive behavior in fish, suggesting an evolutionary mechanism conserved among vertebrates. Such mechanisms may be important for the control of aggression in many vertebrate species, not just mammals, as has been demonstrated so far.

Effects of Glyphosate-Based Herbicide Sub-Lethal Concentrations on Fish Feeding Behavior.

Glyphosate-based herbicides are widely used in agricultural systems. Although the target organism are particularly plant organisms, there are numerous studies showing adverse effects in aquatic animals, such as inhibition of acetyl cholinesterase, effects on kidney, liver, and gill and stressors effects. This study analyzed the effects of commercial formulation of glyphosate on feeding behavior in Pacu (Piaractus mesopotamicus). Fish were exposed to three glyphosate concentrations (0.2, 0.6, and 1.8 ppm) for 15 days. At concentrations of 0.2 and 0.6 ppm, food intake decreased on day 13 and then returned to normal on day 15. At the highest glyphosate-based herbicide concentration, 1.8 ppm, food consumption decreased dramatically and did not recover on day 15. This study showed that glyphosate-based herbicide at sub-lethal concentrations can affect feed intake in pacu and consequently inhibits its growth.

Bile acids as potential pheromones in pintado catfish Pseudoplatystoma corruscans (Spix & Agassiz, 1829): eletrophysiological and behavioral studies

Bile acids are potent olfactory and gustatory stimulants for fish. Electro-olfactogram recording was used to test whether the olfactory epithelium of pintado catfish Pseudoplatystoma corruscans is specifically sensitive to bile acids, some of which have been hypothesized to function as pheromones. Five out of 30 bile acids that had been pre-screened for olfactory activity in fish were selected. Cross-adaptation experiments demonstrated that sensitivity to bile acids is attributable to at least 3 independent classes of olfactory receptor sites. The taurocholic acid (TCA) and taurochenodeoxycholic acid (TCD) were the most potent compounds. By using avoidance/preference tests, we found that P. corruscans prefers water containing TCA. Bile acids are discriminated by olfactory epithelium of pintado, supporting that these compounds could function as pheromones.

Os ácidos biliares são potentes estimulantes olfatórios e gustatórios em peixes. Registros em eletro-olfactograma foram usados para testar se o epitélio olfatório de Pseudoplatystoma corruscans, pintado, é sensível aos ácidos biliares, alguns dos quais têm sido propostos como feromônios. Foram selecionados cinco de uma lista de trinta ácidos biliares previamente testados em atividade olfatória em peixes. Testes de adaptação cruzada demonstraram que a sensibilidade aos ácidos biliares se dá por 3 classes independentes de sites de receptores olfatórios. O ácido taurocólico (TCA) e o ácido tauroquenodesoxicólico (TCD) foram os compostos mais potentes. Em testes de evasão/preferência, P. corruscans prefere água contendo o ácido TCA. Os ácidos biliares são discriminadas por epitélio olfatório de pintado, evidenciando que estes compostos podem funcionar como feromônios.

Does sex influence intraspecific aggression and dominance in Nile tilapia juveniles?

Although sex of mature fish is known to influence aggression, this issue has so far been neglected in juveniles. Here, we tested this sex effect and showed that it does not significantly affect intraspecific aggression in juveniles of the cichlid Nile tilapia. To reach this conclusion, we measured the latency period before onset of confrontation, the frequency and types of aggressive interactions, the duration of a dispute, and the probability of becoming dominant. This was done on pairs of Nile tilapia that varied by sex: females×females, males×males, and females×males. In a double blind approach, after pairing, the sex of each individual was histologically verified and contrasted with behavioral data.

Blood cues induce antipredator behavior in Nile tilapia conspecifics.

In this study, we show that the fish Nile tilapia displays an antipredator response to chemical cues present in the blood of conspecifics. This is the first report of alarm response induced by blood-borne chemical cues in fish. There is a body of evidence showing that chemical cues from epidermal 'club' cells elicit an alarm reaction in fish. However, the chemical cues of these 'club' cells are restricted to certain species of fish. Thus, as a parsimonious explanation, we assume that an alarm response to blood cues is a generalized response among animals because it occurs in mammals, birds and protostomian animals. Moreover, our results suggest that researchers must use caution when studying chemically induced alarm reactions because it is difficult to separate club cell cues from traces of blood.