In Vivo Trials With Fish
Global aquaculture production has grown rapidly during past decades, contributing significant quantities to the world's fish supply for human consumption. As aquaculture expands and develops, an increase in problems and challenges arises including a widespread occurrence of diseases, such as parasitic infestations, bacterial and viral infections. This can cause heavy losses to the industry. Different strategies have been implemented to counteract bacterial and viral threats; chemotherapy being the most used approach, uses large amounts of antibiotics and chemical products. Development of drug resistance in bacteria, accumulation of such chemicals in the environment and carry-over in shrimp and fish tissues has led to strict regulations that limit use of antibiotics and other chemicals in aquaculture. Thus, the industry has been looking for safe and effective approaches that help improve aquaculture production. Biomin® P.E.P. is one of the few phytogenic feed additives of which the mode of action is well described for terrestrial animals (Kroismayr et al., 2008a,b). A series of research trials was carried out to gain more insight into the application of this product in aquatic species. These trials showed that addition of Biomin® P.E.P. to fish diets improved growth performance in different species.
1. Trials at the Aquaculture Centre of Applied Animal Nutrition (ACAN)
In order to evaluate phytogenics application possibility, several trials were carried out in BIOMIN's Aquaculture Centre of Applied Animal Nutrition (ACAN). The ACAN is a state-of-the-art research facility in Bangkok, Thailand, which allows for testing feeding strategies and feed additives in standardized conditions with different aquatic species.
1.1 Trial design:
In this trial facility, Biomin® P.E.P. 125 was tested in Tra catfish (Pangasius hypothalamus) according to the following complete randomized block design with 3 replications per treatment:
Table 1: Effect of organic acids on performance parameters of cat fish
|Treatment||Supplementation with Biomin® P.E.P. 125 (g/t)|
|Trial Group 1||
|Trial Group 2||
Thirty juvenile catfish averaging 150 g were stocked in each of the 12, 500-L tanks of system 3. The fish were fed pellet diets to near satiety 3 times daily for a period of 8 weeks.
Although the density of the rearing conditions was high (12 kg/ m3 in the beginning and reaching 20 kg/m3 at end) survival rate was 100% and the fish almost doubled their weight during the trial period. The FCR obtained during the trial for all diets was good (FCR=1.18 DM basis) and bellow that observed in the industry for this species (approx. 1.5-1.6). Even under these excellent rearing conditions body weight gain and daily growth coefficient, as well as FCR were improved by addition of Biomin® P.E.P. (Figures 1-3). Similar positive results were obtained with Red Tilapia (Orechromis niloticus × O. Mussambicus) in additional ACAN trials,which confirms the potential of this phytogenics feed additive in aquatic species.
Figure 1. Total gain of Pangasius catfish (ACAN)
Figure 2. Daily Growth Coefficient of Pangasius catfish (ACAN)
Figure 3. Feed Conversion Ratio of Pangasius catfish (ACAN)
2. Field trial with Pangasius (Vietnam)
In a field trial carried out under the surveillance of Vasafeed Co., Ltd. (Ho Chi Minh City, Vietnam), Biomin® P.E.P. 125 was used in Pangasius feeds under commercial conditions.
2.1 Trial Design
The trial was conducted during the normal production cycle for Pangasius (6 months) in the production ponds of a Vasafeed farm in Vietnam Tay Nhin province. Groups of 2,000 juvenile Tra catfish (Pangasius hypothalamus) with an average initial weight of 50±1 g were stocked into 4 cages (5 x 10 x 2.5 m) in a production pond. The trial design consisted of two treatments with four replicates per treatment. The control group was fed a commercial diet containing no growth-promoting additives, whereas in the trial group the commercial diet was supplemented with 150 g/ton with Biomin® P.E.P. 125. Fish were fed the extruded diet three times daily to near satiety.
- Control group: Commercial diet
- Trial group: Commercial diet + Biomin® P.E.P. 125 (150 g/kg)
The trial results from the 6 month period (October 12, 2008 to April 10, 2009) showed a significant improvement in growth performance for fish fed diet supplemented with Biomin® P.E.P. 125 (Table 1). Compared to the control group, fish fed the Biomin® P.E.P. supplemented feed had 14.2 % higher weight gain. Improvement in weight gain was observed after the first month of sampling (87.2 g vs. 79.6 g) and continued throughout the production period.
Table 1. Effect of Biomin® P.E.P. on growth performance in catfish
|Control Group||Biomin® P.E.P. 125|
|No. of fish||
|Initial body weight (g)||
|1st month body weight (g)||
|2nd month body weight (g)||
|6th month body weight (g)||
|Weight gain (d 1-180) (g)||
|Feed conversion rate||
* 1 hapa was broken and the fish removed from the experiment
a,b Significant difference (P < 0.05)
The feed conversion rate (FCR) for this trial period was higher than normal farming conditions and possibly related to the low water temperature of winter months and over feeding by the farm manager. Feed supply was similar to every cage and results show that fish fed the Biomin® P.E.P. 125 supplemented feed had a better feed efficiency resulting in lower FCR (1.83 vs. 2.12).
The economical calculation was carried out by calculating the costs and revenue from the fish produced in the trial hapas (Table 2). Inclusion of Biomin® P.E.P. 125 at 150 g/t resulted in extra revenue of US$393 per ton of fish for the farmer and a return on investment (ROI) of 1:6.
Table 2. Economical benefits from inclusion of Biomin® P.E.P. 125 in Pangasius diets under the trial conditions
|Control Group||Biomin® P.E.P. 125|
|Feed consumed (kg)||7,792||7,764|
|Sales (0.85 US$/kg)||3,125||3,567|
|Feed costs (0.37 US$/kg)||2,883||2,932|
|Feeding costs (US$/t fish)||784||699|
|Gross Revenue (US$)||242||635|
3. In vitro immune-stimulating effects of phytogenics
Disease problems have emerged as constrains to the growth of aquaculture. In some cases the uncontrolled and repeated use of therapeutic antibiotics to treat bacterial infections has resulted in the development of antibiotic resistant pathogens. Considering the potential human and animal health issues associated with the misuse of antibiotics, future disease management will focus more on environmental-friendly and preventative approaches. The prevention and treatment of infectious diseases by applying immuno-modulating substances appears to be a possible and sustainable option. Such substances may enhance the non-specific, innate immune system and may be used from hatching, and thus are attractive for use in intensive juvenile production. Stimulation of the immune system by essential oils was observed in the non-specific immune system of common carp (Cyprinus carpio). Comparing several natural extracts in vitro, nitric oxide production (a potent indicator of macrophage activity) was significantly enhanced by Biomin® P.E.P. in head kidney cells for different concentrations (0.1, 1.0 and 5.0 ?g/ml), thus, indicating enhanced macrophage activation. This effect was even more pronounced in comparison with that observed for yeast extracts at doses of 1.0 and 5.0 ?g/ml (Figure 4).
Figure 4. Changes in nitric oxide production in macrophages isolated from head kidney following in vitro incubation with different doses of test substances. *Significant differences (P < 0.05) from negative control are shown by asterisks.
Moreover, the respiratory burst was significantly increased in the treatment with Biomin® P.E.P. at the three lowest concentrations (0.1, 1.0 and 5.0 ?g/ml), while no significant changes were measured in other treatments. These in vitro results attest the capacity of essentials oils to stimulate the non-specific immune system in fish, which has practical application in commercial challenging conditions.
Sustainable aquaculture requires the use of safe and effective solutions to tackle industry challenges of the present and future. In this regard, phytogenics represents a promising group of modern feed additives. There is growing evidence that natural products, such as phytogenics, have application in aquaculture, thus helping to improve performance and minimize the use of preventative and therapeutic chemicals.
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