The study of the effect of the black soldier fly larvae (Hermetia illucens) added feed on tilapia.

Director of the Innovation Center «Bioakvapark –
Science and Technology Center for mariculture» AGTU
Doctor of Biological Sciences,
Professor Ponomarev S.V.

 

Experimental works have been carried out on the premises of “Bioakvapark – Scientific and Technology Center for mariculture” FSBEI of HE “Astrakhan State Technical University”. The object of the study was the fingerling of the red tilapia (a hybrid of albinic Mozambican females and Nile tilapia males). Heated aquariums of 400 l with artificial aeration and filtration were used for the maintenance of tilapia.

The study of the effect of the black soldier fly larvae (Hermetia illucens) added feed on tilapia.

Picture 1 – Aquariums for red tilapia

When assessing the effect of feed on the status of farmed fish, a mix of fish-biological and physiological-biochemical methods have been used (Ponomarev et al., 2002).

For control weighing and measurement, a random sampling of 25 individuals of each kind was made. Experimental feeds were produced in laboratory conditions using domestic-made feed components (quality and safety certificates and certificates of conformity were obtained for all components) by the method of wet pressing at low pressure. Herbal ingredients were pre-micronized for starch gelatinization (a process similar to extrusion), which increases its digestibility and nutritional value. After that, all the components were weighed on electronic scales, and then thoroughly mixed with water until a homogeneous mass was obtained, after which the wet mixture was passed through a meat grinder, dried in a thermostat at a temperature of 600 °C. The finished feed was crushed and scattered in accordance with the required size of grains, which was set in accordance with the mass of fish grown. The content of the experimental feed is presented in table 1.

Table 1 – Composition of produced feed stuff

Component Content
Control group Option with 30% replacement Option with 70% replacement Option with 100% replacement
1. Soybean Meal 15 15 15 15
2. Sunflower Meal 7 7 7 7
3. Wheat 15 15 15 15
4. Fish flour 45 31,5 13,5
5. Protein concentrate from black soldier fly larvae 13,5 31,5 45
6. Fish fat 7 7 7 7
7. Wheat gluten 5 5 5 5
8. Meat flour 5 5 5 5
9. Premix PM-2 1 1 1 1
  Content of nutrients,%
10 Crude protein 48,36 46,32 43,61 41,57
11 Crude fat 12,23 12,58 13,54 13,4

 

The oxygen content in water was measured daily three times a day (to prevent unwanted oscillations) using a CyberScanDO 300 thermal oximeter. The pH values were determined using a Hanna pH meter. In addition, water temperature was recorded 3 times a day.

The measurement design of fish was carried out according to the method of Pravdin I.F. (1966).

The average daily growth rate of the older age groups was calculated using the compound interest formula (Castell, Tiews, 1979):

А =[( mk /mо )1/t – 1] х 100 (%)

where mk и mо are the mass of fish at the end and at the beginning of the experiment;

t is the duration of the experiment, days.

The absolute incremental growth was calculated by the formula (Pravdin, 1966):

Gab = mc – m0

where mk  – the final mass of young, grams;

m0 is the initial mass of fingerling, gram.

Average daily growth was calculated by the formula (Pravdin, 1966):

Gav/day. = (mk – m0)/t

where mk  – the final mass of young, grams;

m0 is the initial mass of fingerling, gram.

t is the duration of the experiment, days.

To determine the growth rate more accurately, the mass accumulation coefficient was calculated (Reznikov et al., 1978; Kupinsky et al., 1986).

Cm = ((Mk1/3 – Мо1/3)*3) / t

where Cm is the total production coefficient of the growth rate;

Mk и Mo – final and initial mass of fish, g;

t is the time of cultivation, days.

Feed costs were calculated by the formula (Ponomarev et al., 2002):

Fc= Ck/( mc – m0),

where Ck Ck – the amount of feed spent on growing fish (feed costs per unit of growth).

Сk=R*m.average start*t

where R is the daily rate of feed,%; m.average start is the average initial mass, g; t-growing period.

Survival was expressed as a percentage of the total number of fish observed.

When determining the norms of feeding, as well as the size of grain, recommendations on feeding were used, based on the optimal water quality and the temperature of cultivation of 270C. Food was set manually 3-4 times a day. The stocking density of the young was established based on the weight of the fish grown.

All data were subjected to statistical processing according to G.F. Lakin (1990) using the Excel statistical analysis toolbar. At the same time, elements of statistical analysis were used with the definition of the mean, the error of the mean. The level of differences was assessed using Student t-test.

When assessing the physiological state of the fish, hematological parameters are important, the changes of which depend on the species and age characteristics of the fish grown. In addition, blood biochemical parameters are an adequate indicator of the balance of the food consumed.

Blood was collected in vivo from the tail vein into Eppendorf tubes. For hematological analysis (hemoglobin concentration, erythrocyte sedimentation rate, leukocyte formula), ethylenediaminetetraacetic
acid disodium salt (EDTA) was used as an anticoagulant.

The hemoglobin concentration in the blood was determined photometrically using an Agat-Med reagent kit (vanKampen, Zijlstra, 1961), the ESR was determined by the Panchenkov method. Blood smears were prepared using a fixer-dye according to May-Grunwald of the firm Olvex-Diagnosticum (Kozinets, 1998; Abramov, 1985). Leukocyte identification was performed using a LOMO microscope and an immersion objective (100 / 1.25). On each smear, 200 leukocytes were determined taking into account the stages of their cytogenesis according to N.T. Ivanova (1983).

For biochemical analysis of blood (total protein, total lipids, cholesterol, glucose), blood samples were collected in tubes without EDTA, left to coagulate, then centrifuged at 3000 rotations per minute to separate the serum.

The content of whey protein was determined on an IRF-22 refractometer (Filippovich et al., 1975), the level of cholesterol in the blood was determined by an enzymatic method (Trinder, 1969; Fishbach, Dunning, 2004). Diagnostic kit of reagents from the company PLIVA – Lachema (Zollneretal., 1962; Knichtetal, 1972) was used to determine the total serum lipids. The serum glucose concentration was determined by an enzymatic colorimetric method without deproteinization (the Trinder reaction) (TrinderP., 1969). To measure the optical density of the obtained samples, a Unico 2100 spectrophotometer was used.

Results are presented as mean value of the indicator and its standard error (M ± m). The assessment of reliability was performed using Student t-test.

Hematological studies were carried out in accordance with the “Guidelines for conducting hematological examination of fish” (Approved February 2, 1999, №13-4-2/1487).

For histological analysis, the female reproductive products were taken, fixed in a Buen mix, embedded in paraffin, and sections of 7 μm thick were made (Mikodina et al., 2009). The obtained sections were stained with hematoxylin-eosin and photographed on a microscope OlimpusBX 53.

 

2 Study results

During the experiment, the survival rate in aquariums with red tilapia was 100%. The best and statistically reliable indicators of the cultivation results were obtained for a group of experimental fish that consumed food with a 30% replacement of fish meal with protein concentrate from black soldier fly larvae (Table 2).

 

Table 2 – Fish and biological indicators of growing red tilapia on experimental feed

Indicators Experiment samples
Control group Option with 30% replacement Option with 70% replacement Option with 100% replacement
Initial weight, g 109,27+5,26 109,27+5,26 109,27+5,26 109,27+5,26
Final weight, g 138,07±2,2 157,9±2,59* 150,31±3,06 139,11±1,02
Initial length, cm 15,03±1,09 15,03±1,09 15,03±1,09 15,03±1,09
Final length, cm 17,1±0,74 18,6±1,01** 18,2±1,02 17,9±1,03
Absolute gain, g 28,8 48,63 41,04 29,84
Average daily gain, g 0,96 1,62 1,37 0,99
Average daily growth rate,% 0,77 1,22 1,06 0,80
Mass accumulation coefficient, units 0,038 0,061 0,052 0,039
Feed ratio 1,4 1,1 1,1 1,3
Survival, % 100 100 100 100
The duration of the experiment, days. 30 30 30 30

Note: * the differences are significant at p≥0.001; ** -r≥0.05

For 30 days, the absolute increase in all experimental groups exceeded the indicators of the control group, and in the best experimental sample – by 1.68 times. Weight growth when feeding tilapia with food with a replacement with a black soldier fly was more intense than in control group, as well as linear.

The increase in weight during the consumption of feed additive by tilapia also reliably demonstrates the average daily growth in all experimental samples. While in the control group it was lower than the best option with a 30% replacement of fish meal at 0.66 g. The feed coefficient, which characterizes the efficiency of digestion and digestibility of feed, in the experimental samples was higher – in the group with a 30% replacement of fish flour – 0.3 units. Thus, growth indices indicate a positive effect on tilapia fingerlings for the replacement of fish meal with protein concentrate from black soldier fly larvae.

When studying the main biochemical parameters of the grown young tilapia, no significant differences in the protein content were noted (Table 3).

 

Table 3 – Biochemical parameters of red tilapia (content,% in dry matter)

Indicators Experiment samples
Control group Option with 30% replacement Option with 70% replacement Option with 100% replacement
Moisture,% 72,04 70,02 69,99 71,64
Protein, in dry matter,% 60,01±1,04 65,22±1,52 63,24±1,79 57,91±0,98
Lipids, in dry matter,% 20,3±0,76 19,31±0,39 21,41±1,21 22,97±0,97
Ash, in dry matter,% 19,02±0,36 18,74±0,84 19,16±0,21 18,72±0,78

When assessing the biochemical composition of the body of tilapias involved in the experiment, no significant differences were found. However, an increase in the protein level is worth noting in the control and experimental groups with a 30% and 70% replacement of fish meal for a protein concentrate from black soldier fly larvae. 100% replacement of fishmeal is not strongly reflected in the biochemical parameters.

The state of the fish in the proposed environmental conditions can be objectively assessed on physiological and biochemical parameters of blood, which act as specific indicators of physiological or pathological changes in the fish. Fish-breeding indicators, as a rule, correlate with the physiological state of the fish, which is confirmed by hematological parameters. As can be seen from table 4, there is a positive trend in the use of 30% fish meal replacement, which affects the hemoglobin level, maintain the glucose level at the level of normative values, decrease the ESR, increase whey proteins and slightly lower blood cholesterol.

Table 4 – Physiological and biochemical blood parameters of tilapia

Option Hemoglobin, g/l ESR mm/h Total protein, g/l Cholesterol, mmol/l Glucose, mmol/l
Experiment samples Control group 81,05±3,09 4,4±0,09 23,01±0,63 3,41±0,03 3,2±0,03
Option with 30% replacement 85,36±2,41 3,8±0,08 27,25±1,41 3,29±0,04 3,1±0,02
Option with 70% replacement 82,29±3,12 4,4±0,12 23,73±1,97 3,81±0,02 3,1±0,06
Option with 100% replacement 78,63±1,48 4,7±0,11 23,64±2,30 4,11±0,02 3,8±0,05

 

To date, the effect of feed with the addition of black soldier fly larvae when growing rainbow trout, canal catfish and blue tilapia has been studied the most. Initial studies have shown that in the case of rainbow trout, the larvae of the black soldier fly Hermetia Illucens can replace 25% of fish meal or 38% of fish oil in feed without any side effects. Thus, fly larvae are an ideal substitute for fish meal. The data obtained in the course of our studies allows us, first of all, to recommend the replacement of fishmeal by 30% in tilapia production feeds for their commercial cultivation, which increases growth rates, reduces feed costs, and also maintains the physiological state of the fish at the appropriate standards. It can also be recommended to use a 70% replacement of fish meal. A 100% replacement of fish meal by reducing protein and increasing fat in the recipe does not significantly increase growth rates compared to other options for industrial cultivation, nor has a clear negative effect on fish-biological and physiological indicators.

 

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