Fake plants have real benefits for farmed tilapia - Part 2

Experimental design

The experiment started in May 2018 and lasted 100 days. The 640 juveniles were divided into 16 groups of 40 fish each, following a design of 4 treatments with 4 replications (Fig. 1). 

The first treatment was composed of environmental enrichment, using shelter (PVC pipes, 10 cm in diameter and 20 cm in length). 

The second treatment was also composed of environmental enrichment, using artificial water hyacinth (frayed nylon rope simulating the natural water hyacinth root) fixed to styrofoam structure or to the aquarium itself). 

The third treatment was outlined with food supplementation with tryptophan supplemented to the diet at 2.56 % by weight. 

The fourth treatment was a control tank without enrichment or supplemented food. Biometrics were performed on days 0, 30, 60 and 100 with 50 % of the individuals of each aquarium, to evaluate welfare and zootechnical parameters performance, as well as to adjust the amount of feed provided for fish as they grow.

Fig. 1. Experimental design: (a) control; (b) artificial water hyacinth treatment

Feed and feeding

The fish were manually fed four times a day at 8 a.m., 11 a.m., 2 p.m. and 5 p.m., following a feeding pattern in fish farms, with extruded 4 mm feed pellets formulated following the recommendation of the “Brazilian Table for Tilapia Nutrition” proposed by Furuya (2010), specifically for juvenile tilapia. The fish were fed until the apparent satiety. Specifically for the tryptophan supplemented treatment, the feed provided was produced using the same base formula as the other treatments, but with 2.56 % of tryptophan in the feed production, while the other treatments received pellets containing only 0.32 % tryptophan. The composition of the diets is presented in Table 1.

Table 1. Diet formulations and nutritional compositions (g kg−1): (a) feed provided to the animals of the environmental enrichment and control treatments; and (b) feed provided to the animals of the tryptophan-supplemented treatment.

(a)
Ingredients	%
Soybean meal-45	43.50
Soy protein concentrate	10.00
Poultry flour	5.00
Ground corn	33.04
Soybean oil	0.50
Wheat middlings	2.00
DL-methionine	0.22
L-Threonine	0.36
L-Tryptophan *	0.32
Dicalcium phosphated	2.01
Inert material (sand) *	2.24
BHT 1	0.02
Mineral vitamin mix2	0.60
Vitamin C	0.09
Salt	0.10
Total	100.00
Nutritional Composition	%
Dry matter	86.23
Crude energy (kcal kg−1)	3762
Crude protein	31.86
Digestible energy (kcal kg−1)	3044
Digestible protein	29.75
Crude fiber	3.31
Ether extract	3.03
Calcium	0.81
Available phosphorus	0.53

 

(b)
Ingredients	%
Soybean meal-45	43.50
Soy protein concentrate	10.00
Poultry flour	5.00
Ground corn	33.04
Soybean oil	0.50
Wheat middlings	2.00
DL-methionine	0.22
L-Threonine	0.36
L-Tryptophan *	2.56
Dicalcium phosphated	2.01
Inert material (sand) *	-
BHT 1	0.02
Mineral vitamin mix2	0.60
Vitamin C	0.09
Salt	0.10
Total	100.00
Nutritional Composition	%
Dry matter	88.45
Crude energy (kcal kg−1)	3908
Crude protein	33.74
Digestible energy (kcal kg−1)	3044
Digestible protein	29.75
Crude fiber	3.31
Ether extract	3.03
Calcium	0.81
Available phosphorus	0.53

Ingredients followed by (*) were modified among the diets.

Discussion

Fish welfare promotion by environmental enrichment is an incipient area. There are few studies investigating the effects of environmental enrichment in fish and none regarding fish welfare techniques and aquaculture. In this study, we have shown that by introducing environmental enrichment, significant improvements can be obtained in the parameters tested, which justifies the use of these techniques as a method of ensuring better quality of life in animals without affecting the commercial aspect linked to aquaculture production.

Animal growth is a measure used for analysis and is related to animal welfare, given that animals reared in good conditions demonstrate improved results. For such parameters, it was observed that the shelter, artificial water hyacinth, and control treatments presented better average final weight and average weight gain compared to the tryptophan treatment. The high percentage of such an amino acid in the composition of diet may have acted more intensely in the hypothalamus-hypophysis-interrenal axis, increasing the body’s serotonin levels. Serotonin is implicated in the modulation of different neural activities including stress reduction and food ingestion by promoting satiety for longer.

This study clearly shows that fish prefer environmentally enhanced sections before and after feeding. This result demonstrates the preference for the enrichment and its similar features to the natural environment, providing, in this case, a place of refuge and shelter for the fish. Previous studies in tilapia submitted for preference testing also showed a higher demand for sheltered and gravel enriched environments compared to non-enriched environments. Furthermore, Nile tilapia had enhanced learning and memory when an enrichible environment was offered. Thus, an enriched environment simulating natural resources is clearly beneficial for fish, even in intensive production systems, as supported by the results obtained in this study.

We found that tryptophan-supplemented food reduced confrontations. The low rate of confrontations that individuals showed under this treatment can be explained by the fact that tryptophan is a serotonin precursor, a neurotransmitter involved in the control of aggressive behavior and the susceptibility to stress. Similar results were demonstrated by Wolkers et al. (2014) with the matrinxa fish; aggressive behavior such as latency for the first attack and the number of bites and stalking were lower in the fish that received TRP. Other literature studies, in which the effect of this amino acid in the diet was analyzed, concluded that diets supplemented with TRP decreased the stress response in carps at different stocking densities, increased fish tolerance under saltwater conditions and decreased cortisol release .

Water hyacinth and shelter presented high levels of confrontation, and it is well-known that aggressive reactions frequently occur during fear, predation, disputes over territories, defense and situation of reproduction. Despite the benefits, enriched environments can generate competition between territorial fish, which means more confrontations. According to Barreto et al. (2011), Nile tilapia males showed higher aggressiveness in territories enriched by more resources to be defended, and this result reinforces those obtained in this study, where the artificial water hyacinth treatment presented the higher confrontation number.

However, environments enriched with artificial water hyacinth and shelter present higher latency to trigger confrontations, being less intense (mostly threats but not escalated fighting), and concentrated at the section with enrichment items. In the control situation (without enrichment), the occurrence of these confrontations presented lower latency, were more intense (pursuits, lateral confrontation, and bites) and occurred through the entire aquarium region. Some studies that observed fish confrontations showed similar results. Torrezani et al. (2013) analyzed the behavior of Congo tilapia in an enriched environment and the fish showed a decrease in latency to start fighting, and attack frequency was lower. 

The dominance hierarchy, also observed, was less explicit compared to animals kept in an environment without enrichment. In the study proposed by Mendonça et al. (2010), Nile tilapia, that were kept with different substrates and environments, presented equal frequency of total attacks without them, but, as this study shows, highly-intense attacks were fewer in animals kept in aquariums with the substrate. Mendonça and Freitas (2008), in which an enrichment nest, in this case, could have been used to signal territory, where dominant fish defend the nest or this resource, only attacking when similar individuals invade such a demarcation.

Confrontation occurs due to natural tilapia behavior when they seek resources to nest, establish territory and when they feel threatened by conspecifics or predators, seek ways to avoid harmful confrontations by fleeing or hiding. 

Therefore, considering the possible causes that lead to an increase or reduction in aggressive behavior, the high levels of confrontation in treatments influenced by water hyacinth and shelter are justified, since there was a dispute for the enrichment and a higher concentration of animals around the resource, leading to intensification in the number of confrontations. 

However, as already mentioned, these confrontations are inherent to the process and species, and fish may be in good condition even under stress. In animal welfare precepts, an important characteristic is a species manifesting natural behavior, which in breeding conditions can be abolished or subverted. 

In non-enriched environments, we can evaluate that the monotonous and resource-less environment induces the occurrence of confrontations, its intensity and the constant alertness of the fish. As such, concerning fish ethology, the presence of environmental resources becomes an important tool to be exploited by fish farms in the scope of productive growth, once fish can make decisions to benefit themselves, avoiding fights that can result in injuries and higher mortality.

When comparing environmental enrichment to food supplementation, the data from this study showed that food supplementation on its own may not be sufficient to act as an alternative to animal welfare. Here we showed that scratching, considered a stereotypical behavior, increased in tryptophan treatment relative to environmental enrichment treatments. The non-enriched environment caused changes in the behavior of individuals, such as repeated movements (stereotypical). Studies ascribed such stereotypical behavior to poor animal welfare, since this stress-relieving behavior represents a fish strategy for dealing with restrictive environmental .

Environments without apparatus to simulate natural habitat can be considered reducing tools to the fish welfare, causing anomalous behavior. In Brachydanio rerio, the use of artificial plants as a way to enrich the environment promoted increased telencephalon cell proliferation and decreased anxiety-type behavior. Including an artificial water hyacinth decreased stereotypical behavior. Stereotypical behavior is not desirable for production, as by repeatedly rubbing their body against the net-tank structure, fish can remove the protective mucus layer, become injured and more prone to fungal and bacterial contamination, resulting in high mortality and production losses.

Regarding stress indicators, we analyzed the opercular beat frequency (or ventilatory frequency – BO0), where high BO is related to increased stress levels, and is an important animal welfare indicator. Here we showed that after the management and introduction to new environmental situations, fish in the water hyacinth environment had a lower BO which was indicative of lower stress. 

This suggests a rapid adaptation to the new environment, where water hyacinth individuals returned to their basal state of BO, while the others maintained their alertness or failed to adapt. This adaptation to a new environment, when using an environmental resource that promotes stress relief, is ideal for production as fish are often handled when sorted, harvested and transported. Finally, a relatively simple environmental resource such as an artificial water hyacinth seems to act as a suitable tool for managing stress and environmental novelty.

In conclusion, a poor environment in fish farming has deficiencies that can be improved by including proposed enrichments, filling a fish need to express natural behavior, improving the quality of life for animals. We can highlight artificial water hyacinths as the treatment that presented the best and most consistent results. Such a means of environmental enrichment can be easily implemented in fish farms since it is low cost, easy to handle, and highly durable.