THERAPEUTIC USES OF AN INSECT POWDER

Abstract

The invention relates to a Coleoptera powder for using in the treatment of vibriosis, which particularly affects the white shrimp.

Claims

1. A method for preventing or treating vibriosis and/or symptoms thereof, comprising administering beetle powder to a subject in need thereof.

2. The method according to claim 1, wherein said subject is in a bivalve and or a decapod.

3. The method according to claim 2, wherein administering the beetle powder prevents or treats shrimp enteritis.

4. The method to claim 3, wherein administering the beetle powder prevents or treats shrimp early mortality syndrome/acute hepatopancreatic necrosis syndrome in shrimps.

5. The method according to claim 1, wherein said beetle powder comprises at least 67% by weight proteins and at least 0.1% by weight chitin, percentages being given with respect to the total weight of beetle powder.

6. The method according to claim 5, wherein said beetle powder comprises at least 67% by weight proteins and at least 5% by weight chitin, percentages being given with respect to the total weight of beetle powder.

7. The method according to claim 5, wherein said beetle powder comprises at least 71% by weight proteins and comprises between 0.1 and 2% by weight chitin, percentages being given with respect to the total weight of beetle powder.

8. The method according to claim 1, wherein said beetle powder is obtained from the species Tenebrio molitor.

9. Nutrition regime for molluscs and/or crustaceans, comprising at least 5% by weight beetle powder, the beetle powder comprising at least 71% by weight proteins and comprising between 0.1 and 2% by weight chitin, percentages being given with respect to the total weight of beetle powder.

10. Nutrition regime according to claim 9, comprising 20% by weight beetle powder.

11. Use of a beetle powder comprising at least 67% by weight proteins and comprising at least 0.1% by weight chitin, percentages being given with respect to the total weight of beetle powder, as a nutrition supplement in human or animal nutrition.

12. Use of the beetle powder according to claim 11, as a nutrition supplement in the nutrition of molluscs and/or crustaceans.

Description

[0121] Other characteristics and advantages of the invention will become apparent from the following examples, given by way of illustration, with reference to:

[0122] FIG. 1 is a diagram showing the final weight of the white shrimps fed according to different nutrition regimes, either including or not including insect powder in place of fishmeal;

[0123] FIG. 2 is a diagram showing the weight gain of the white shrimps fed according to different nutrition regimes, either including or not including insect powder in place of fishmeal;

[0124] FIG. 3 is a diagram showing the activity of phenol oxidase as a function of the bacterial reduction of the pathogen, in white shrimps infected with Vibrio parahaemolyticus and fed according to different nutrition regimes, either including or not including insect powder in place of fishmeal; and

[0125] FIG. 4 is a diagram showing the survival rate of the shrimps infected with Vibrio parahaemolyticus as a function of the nutrition regime provided, either including or not including insect powder in place of fishmeal.

EXAMPLE 1: METHOD FOR THE PREPARATION OF AN INSECT POWDER

[0126] The composition according to the invention is prepared from Tenebrio molitor larvae. Upon receipt of the larvae, the latter can be stored at 4° C. for 0 to 15 days in their rearing tanks without major degradation before being killed. The weight with respect to age of the larvae used is variable and as a result their composition can vary, as illustrated in Table 1 below:

TABLE-US-00001 TABLE 1 Biochemical composition of the larvae of Tenebrio molitor according to the weight thereof. Biomass (Insects) mg 23 35 58 80 108 154 Dry matter %* 34 34 34.2 37.9 39.6 39.5 Ash %* 1.59 1.52 1.6 1.75 1.67 1.43 Crude protein %* 22.6 22.2 22 23.2 23.1 23.2 Lipids %* 6.62 6.88 7.98 10.3 10.9 11.7 *The % s are expressed in dry weight with respect to the wet weight of larvae.

[0127] Step 1: Blanching the Insects

[0128] Living larvae (+4° C. to +25° C.) are conveyed in layers with a thickness comprised between 2 and 10 cm, on a perforated conveyor belt (1 mm) to a blanching chamber. The insects are thus blanched with steam (steam nozzles or bed) at 98° C. or with water at 100° C. (spray nozzles) or in mixed mode (water+steam). The residence time in the blanching chamber is comprised between 1 to 15 minutes, ideally 5 min.

[0129] The temperature of the larvae after blanching is comprised between 75° C. and 98° C.

[0130] Step 2: Pressing

[0131] Once blanched, the larvae are conveyed to the feed hopper of a continuous single-screw press. While passing into the press, the larvae are maintained at a temperature above 70° C. in order to increase the de-oiling yields. The principle of de-oiling is to pressurize the material inside a cylindrical cage by means of an arrangement of screws and rings arranged on the central shaft. The cage is lined inside with bars distributed in sections and kept apart by spaces of different thicknesses depending on the work area. The interstices thus arranged allow the flow of an oil fraction and limit the passage of the so-called “dry” matter, the protein fraction, which will be called “press cake”, thus being involved in the pressurization.

[0132] The pressing yields obtained are comprised between 48 and 55%.

Y.sub.cake=(mass.sub.cake/mass.sub.juice+mass.sub.cake)

[0133] The press cake obtained contains 35 to 40% dry matter, 67 to 75% proteins and 13 to 17% fats, percentages by weight being given with respect to the dry weight of press cake.

[0134] Step 3: Drying

[0135] The press cake is then arranged on a tray in a thin layer (approximately 2 cm) and is dried in ventilated/stirred air at 90° C. for 5 hours in order to obtain a press cake having a dry matter content greater than 92%.

[0136] This step makes it possible to guard against any contamination having occurred since the killing.

[0137] The Wa (water activity) after drying is 0.35. The microbiological results show an absence of Salmonella spp (method: IRIS Salmonella BKR 23/07-10/11) and Enterobacteria values less than 10 CFU/g (method: NF ISO 2128-2, December 2004, 30° C. and 37° C.).

[0138] Step 4: Grinding

[0139] The dried press cake, comprising mainly proteins, is finally ground using a continuous hammer mill (6 reversible moving parts—thickness 8 mm). The grinder is fed by a hopper with a flow rate control flap (180 kg/h). The perforated grill used to control the output granulometry is 0.8 mm. The speed of rotation of the motor is 3000 rpm (electric motorization, absorbed power 4 kW (5.5 CV)).

EXAMPLE 2: CHARACTERIZATION OF THE INSECT POWDER OBTAINED IN EXAMPLE 1

[0140] The insect powder prepared in Example 1 was characterized.

[0141] 1. Analyses

[0142] 1.1 Determination of the Moisture Content

The moisture content is determined according to the method originating from EC Regulation 152/2009 of 27 Jan. 2009 (103° C./4 h).

[0143] 1.2 Determination of the Quantity of Crude Proteins

The crude proteins are determined according to the method called Dumas method, and corresponding to the standard NF EN ISO 16634-1 (2008).

[0144] 1.3 Determination of the Quantity of Chitin

Dietary fibres from insect meal are essentially composed of chitin, the latter was therefore measured according to the ADAC 991.43 method. The values thus obtained are consequently slightly overestimated.

[0145] 1.4 Determination of the Quantity of Fat

The fat was determined according to the method of EC Regulation 152/2009.

[0146] 1.5 Determination of the Quantity of Ash

The crude ash was determined according to the method under EC Regulation 152/2009 of 27 Jan. 2009.

[0147] 1.6 Determination of the Quantity of Phosphorus

The phosphorus is measured by ICP (induced coupled plasma) with internal calibration.

[0148] 1.7 Determination of Energy

The energy value is obtained with the coefficients of EU Regulation 1169/201.

[0149] 1.8 Determination of the Quantities of Amino Acids and Fatty Acids

This determination was carried out by gas chromatography after hydrolysis and derivatization of the amino acids and fatty acids respectively.

[0150] 1.9 Determination of Pepsin Digestibility

The pepsin digestibility is measured by the method described in Directive 72/199/EC.

[0151] 2. Results

[0152] The insect powder is detailed in Table 2 below.

TABLE-US-00002 TABLE 2 Composition of the insect powder Unit Composition Macronutrient Moisture %* 5.32 Protein %* 67.09 Chitin %* 8.0 Fat %* 13.6 Ash %* 3.21 Total phosphorus %* 0.75 Energy MJ/kg 23.74 Amino acids Arginine %* 2.56 Histidine %* 1.39 Isoleucine %* 2.11 Leucine %* 3.99 Lysine %* 3.32 Threonine %* 1.87 Valine %* 2.91 Methionine %* 1.43 Cysteine %* 0.63 Phenylalanine %* 1.98 Tyrosine %* 2.68 Taurine %* 0.42 Aspartic acid + asparagine %* 4.51 Glutamic acid + glutamine %* 6.36 Alanine %* 3.83 Glycine %* 2.54 Proline %* 3.18 Serine %* 2.94 Fatty acids C12:0 %* 0.03 C14:0 %* 0.22 C15:0 %* 0.01 C16:0 %* 1.33 C16:1 %* 0.05 C16:1n-7 %* 0.16 C17:0 %* 0.02 C17:1 %* 0.01 C18:0 %* 0.35 C18:1n-9 %* 3.03 C18:1n-7 %* 0.04 C18:2n-6 %* 2.96 C18:2tn-6 %* 0.02 C18:3n-3 %* 0.14 C20:0 %* 0.02 C20:1n-9 %* 0.01 C20:2n-6 %* 0.01 C22:0 %* 0.01 *Percentages by weight are expressed with respect to the total weight of powder composition. Moreover, a pepsin digestibility of 90 +/− 2% is obtained.

EXAMPLE 3: ALTERNATIVE METHOD FOR THE PREPARATION OF AN INSECT POWDER

[0153] 200 g of T. molitor larvae are introduced into a beaker, placed in a water bath at 100° C. and containing 200 mL of water brought to the boil beforehand. After 5 minutes, the beaker is removed from the water bath, the larvae are drained, then mixed with a volume of water of 200 mL. The liquid thus obtained is passed into a twin-screw-type press. The press cake thus obtained is dried for 24 hours in an oven at 70° C., then ground to 250 μm. An insect powder is thus obtained.

EXAMPLE 4: EFFECTS OF THE INSECT POWDER OBTAINED IN EXAMPLE 1 ON THE WHITE SHRIMPS, IN PARTICULAR ON THEIR RESISTANCE TO THE PATHOGENIC BACTERIUM VIBRIO PARAHAEMOLYTICUS

[0154] Three experiments were conducted in order to assess the appetence of the insect powders in shrimps (Experiment 1), the digestibility of the proteins, the lipids and the energy of the insect powders in shrimps (Experiment 2) and the efficacy of the insect powder on the growth and the immunity of white shrimps (Experiment 3).

1. MATERIAL AND METHODS OF THE EXPERIMENTS CONDUCTED

[0155] The fishmeal used for all of these experiments is a fishmeal originating from Thailand and comprising a crude protein content of 56%, expressed with respect to the total weight of said fishmeal.

[0156] Similarly, the insect powder used is identical for all of these experiments and corresponds to the insect powder obtained in Example 1.

Statistics

[0157] These experiments were designed in a completely random manner (CMCA). All the data were analysed by ANOVA (variance analysis). Duncan's multiple-range test was used in order to determine the differences between the means of the different nutrition regimes. Alphabetical notation was used for marking the differences at an alpha 0.05 significance level. All research for Experiments 2 and 3 was conducted in the Aquatic Animal Food and Nutrition Laboratory, Department of Aquaculture, Faculty of Fisheries, Kasetsart University, Bangkok, Thailand.

Experiment 1: Test of Appetence of Insect Powders on White Shrimps with Respect to a Usual Nutrition Regime Constituted by 15% Fishmeal

[0158] a. The Different Nutrition Regimes of the Experiment

[0159] The attraction capacity of the insect powders to white shrimps was assessed in a completely random manner using the 5 following regimes, each repeated four times: [0160] Regime 1 (R1): Control 30% fishmeal (hereinafter called FM); [0161] Regime 2 (R2): Negative control 15% FM; 0% insect powder (hereinafter called INSM); [0162] Regime 3 (R3): Negative control 10% FM; 5% INSM; [0163] Regime 4 (R4): Negative control 5% FM; 10% INSM; [0164] Regime 5 (R5): Negative control 0% FM; 15% INSM;

[0165] Four replicates of each regime were used in order to assess appetence.

[0166] b. Formulations and Production of the Nutrition Regimes

[0167] The regimes were formulated with ingredients that are useful in response to the known nutritional needs of white shrimps, described in particular in the reference manual of the National Research Council (NCR) published in 2011.

[0168] The insect powder used in the experimental nutrition regimes is that obtained in Example 1.

[0169] The attraction capacity of the insect powders to white shrimps was assessed in five nutrition regimes detailed below, the composition of which is presented in Table 3 below.

[0170] The approximate compositions of the feedstuffs tested such as the moisture, proteins, lipids, fibres, ash, were analyzed according to the description of the AOAC (2000) which is the international reference method.

TABLE-US-00003 TABLE 3 Compositions of the nutrition regimes of the white shrimps containing insect powder in place of fishmeal R1 R2 R3 R4 R5 INSM INSM INSM INSM INSM Ingredient of 0% −FM 0% −FM 5% −FM 10% −FM 15% −FM the formula 30% 15% 10% 5% 0% Fishmeal 30.0 15.0 10.0 5.0 0.0 Soya 15.0 15.0 15.0 15.0 15.0 Wheat gluten 8.0 8.0 8.0 8.0 8.0 Squid liver 8.0 8.0 8.0 8.0 8.0 powder Wheat flour 24.0 24.0 24.0 24.0 24.0 Broken rice 2.2 1.2 1.2 1.2 1.2 Soya protein 0.0 15.0 15.0 15.0 15.0 concentrate Insect 0.0 0.0 5.0 10.0 15.0 powder Maize gluten 5.0 5.0 5.0 5.0 5.0 Tunny fish 0.5 0.5 0.5 0.5 0.5 oil Soya oil 2.0 2.0 2.0 2.0 2.0 Soya lecithin 2.0 2.0 2.0 2.0 2.0 Mono- 0.0 1.0 1.0 1.0 1.0 calcium phosphate Binder 1.7 1.7 1.7 1.7 1.7 Pre-mix of 1.6 1.6 1.6 1.6 1.6 vitamins Total 100.0 100.0 100.0 100.0 100.0 Chemical composition by AOAC (2000) Moisture (%) 7.38 7.14 7.09 7.25 7.18 Proteins (%) 37.38 36.88 37.06 37.84 38.81 Lipids (%) 7.18 6.50 6.97 7.23 7.60 Fibre (%) 2.69 2.95 3.09 3.33 3.68 Ash (%) 10.39 8.02 7.70 6.58 5.96 Energy 4560.14 4467.36 4477.59 4487.82 4498.05 (Kcal/Kg)

[0171] c. Nutrition Protocol of the White Shrimps

[0172] The white shrimps were fed with pellets containing 36% crude proteins and 7% crude lipids, at a quantity of approximately 5% of the average bodyweight of the shrimps/day (i.e. 1.5-2% bodyweight per feed) by placing the feedstuffs on a tray and providing 3 feeds per day.

[0173] For 10 days and only for the feed at 13.00 hours, the time necessary for the shrimps to ingest the feed initially, the number of shrimps reaching the food within the 15 minutes following deposit of the feedstuffs on the tray and the consumption of the feedstuffs per feed were recorded.

[0174] Four replicates of each regime were used in order to assess attraction capacity.

[0175] Unconsumed feed was collected from the tray one hour after its deposit and freeze-dried at −40° C., then weighed in order to determine the total nutrition consumption.

[0176] Over a study of 10 days, the food intake of each tray in each regime was calculated in order to determine the appetence, focussing on the time necessary for ingesting the feed, the quantity of feedstuffs consumed and the number of shrimps consuming the feedstuffs.

Experiment 2: Study of the Digestibility of the Proteins, the Lipids and the Energy of the Insect Powders in White Shrimps

[0177] The in vivo digestibility of the proteins, the lipids and the energy of the insect powder in white shrimps were determined by an indirect method. Three hundred shrimps with an average weight of 5 to 8 g were randomly distributed in each of the twenty aquariums. Ten repetitions with ten shrimps, each fed with a reference nutrition regime composed of 25% fishmeal and 5% squid meal, 63.7% wheat flour, gluten and soya isolate and soya lecithin, 3% mixture of tunny oil and soya oil, 3.3% premix of vitamins and 1% chromic oxide as digestibility marker.

[0178] Another nutrition regime was provided to the other 200 shrimps (20 repetitions of 10 shrimps), composed of 30% insect powder plus 1% chromic oxide as marker, 63.7% wheat flour, gluten and soya isolate and soya lecithin, 3% of a mixture of tunny oil and soya oil, 3.3% of premix of vitamins and 1% of chromic oxide as digestibility marker.

[0179] The acclimatization period was one week. After this period, the shrimps were fed with the allocated nutrition regimes in excess (4%) three times per day. Two hours after the feed, the aquariums were cleaned, and half an hour before the following feed, the faecal matter was collected, for 1 to 2 weeks.

[0180] The samples originating from each nutrition regime were collected together at the end of the experiment, so as to have enough material for analysis. They were dried in a hot air oven at 65° C. The samples of the feedstuffs and the faecal matter were analyzed for lipids and proteins (AOAC, 2000). The chromic oxide was determined (Scott, 1978). The apparent digestibility coefficient (ADC) was calculated as described by Cho et al. (1985).

Apparent digestibility coefficient (ADC %)=100−(Indicator in the nutrition regime in %)/Indicator in the faecal matter in %

Apparent digestibility coefficient of nutrients (ADC Nu %)=100−(Indicator in the nutrition regime in % X nutrients in the faecal matter in %)/Indicator in the faecal matter in % X nutrients in the nutrition regime in c/o)

Experiment 3: Efficacy of the Insect Powder on the Growth Performance and the Immunity of White Shrimps (Litopenaeus vanammei)

[0181] a. Design of the Tests

[0182] The experiment was designed in a completely random manner (CMCA) with 5 regimes with 4 replicates.

[0183] The different nutrition regimes are indicated in Table 4 below.

TABLE-US-00004 TABLE 4 The different nutrition regimes Level of Insect powder Nutrition fishmeal in the replacing the regime Description nutrition regime fishmeal proteins T1 INSM replaces .sup. 25% 0% 0% of FM T2 INSM replaces 18.75%  25% (5.15% INSM)  25% of FM T3 INSM replaces 12.5% 50% (10.30% INSM) 50% of FM T4 INSM replaces 6.25% 75% (15.40% INSM) 75% of FM T5 INSM replaces   0% 100% (20.50% INSM)  100% of FM

[0184] The experiment was carried out in 20 glass aquariums each having 100 litres capacity, with 70 litres of salt water at 15 ppt (parts per trillion). Juvenile white shrimps (approx. 3 to 4 g) were stocked at a density of 40 shrimps/m.sup.2 (10 per aquarium). The insect powder was mixed into the nutrition while the fish oil was gradually reduced. The feedstuffs thus prepared were provided to the shrimps 3 times per day at a quantity comprised between 3 and 5% of the average bodyweight of the shrimps for 8 weeks. Every 2 to 3 days, the feed waste was siphoned off and 15-20% of the water renewed.

[0185] The ingredients of the different nutrition regimes were ground to 150-250 microns, mixed together, then 25% water was added before passing through the Hobart chopper. The pellets thus obtained were dried in a hot air oven.

[0186] The approximate compositions of the nutrition regimes tested such as the moisture, proteins, lipids, fibres, ash and energy were analyzed according to the description of AOAC (2000). This chemical composition as well as the composition of ingredients are summarized in Table 5 below.

TABLE-US-00005 TABLE 5 Composition of the experimental nutrition regimes T1 T2 T3 T4 T5 INSM INSM INSM INSM INSM replaces replaces replaces replaces replaces 0% of 25% of 50% of 75% of 100% of Matter FM FM FM FM FM Fishmeal 25 18.75 12.5 6.25 0 Soya 25 25 25 25 25 Wheat gluten 10 10 10 10 10 Liver powder 5 5 5 5 5 Wheat flour 26.7 26.6 26.65 26.15 25.55 Insect powder 0 5.15 10.3 15.4 20.5 Tunny fish oil 2 2 2 2 2 Soya oil 1 0.85 0.75 0.65 0.55 Soya lecithin 2 2 2 2 2 Lysine 0 0.1 0.2 0.3 0.4 Methionine 0 0.05 0.1 0.15 0.2 Monocalcium 0 1.2 2.2 3.3 4.5 phosphate Limestone 0 0 0 0.5 1 rock/oyster shell Carbamide 1.7 1.7 1.7 1.7 1.7 Pre-mix of 1.6 1.6 1.6 1.6 1.6 vitamins Total 100.00 100.00 100.00 100.00 100.00 Chemical composition by AOAC (2000) Moisture (%) 9.38 8.82 8.75 9.02 9.45 Proteins (%) 35.30 35.66 35.73 36.39 36.37 Lipids (%) 7.18 7.16 7.18 7.06 7.06 Fibre (%) 2.86 2.86 2.85 2.79 2.78 Ash (%) 8.25 8.02 7.35 6.65 6.03 Energy 4470.38 4434.08 4404.79 4362.88 4318.97 (Kcal/Kg)

[0187] b. Parameters

[0188] The data necessary for this experiment relate to the growth performance of the shrimps under test conditions. These include:

[0189] Growth Performance [0190] The live weight or average weight of the shrimp, the weight gain [0191] The specific growth rate (SGR: {(Ln weight Week 8)−(Ln weight Week 0)}×100)/(period of culture in days) [0192] The feed conversion rate (FCR: feed consumption/shrimp production) [0193] Survival (according to the usual laboratory research method)/mortality rate

[0194] Immune Status

[0195] The immune status is determined at the end of the study by counting the haemocyte total, the phenol-oxydase activity, the haemolymph proteins.

[0196] Morphology of the Intestinal Villi and Hepatopancreas.

[0197] At the end of the study, histopathology of the intestinal villi and hepatopancreas was carried out.

[0198] Test of Resistance to the Pathogen Vibrio parahaemolyticus.

[0199] The resistance of the Pacific white shrimps to Vibrio parahaemolyticus was studied in order to determine the ability of the shrimps to resist the pathogenic bacteria.

[0200] After having fed the shrimps with an experimental nutrition regime for 8 weeks, thirty shrimps on each regime were sampled and were the subject of an injection of pathogens, namely the bacterium Vibrio parahaemolyticus, 4.3/105 CFU (colony-forming unit)/ml (4.6 log CFU/ml) injected intramuscularly. The mortality was recorded for 10 days. The bacteria reduction capacity was determined.

2. RESULTS OF EXPERIMENTS 1, 2 AND 3

Results of Experiment 1

[0201] The appetence of the insect powder in the nutrition regime of the white shrimp (Litopenaeus vannamei) was studied, focussing on the start time of consumption of the feedstuffs by the shrimps, the number of shrimps consuming the feedstuffs within 15 minutes and the quantity of feedstuffs consumed during the feed observed. The results of the attraction capacity of the insect powder have been presented in Table 6 below. The attraction capacity, in terms of the quantity of feedstuffs consumed during the feed observed, the start time of consumption of the feedstuffs and the number of shrimps consuming the feedstuffs do not differ significantly between the different nutrition regimes (p>0.05). The daily nutrition consumption per feed of the shrimps fed with 30% fishmeal (R1) appears to be stable, while the groups of shrimps fed with 15% fishmeal with and without insect powder of Tenebrio molitor (R2-R5) show a wide variation in the consumption of feedstuffs per feed. The time necessary for the shrimps to start consumption of the feedstuffs and the number of shrimps consuming the feedstuffs are completely stable after five days of the study. These elements may mean that the feed based on insect powder of Tenebrio molitor did not show greater attraction in the nutrition regimes of the white shrimps. In addition, this also demonstrates that the reduction in fishmeal in the nutrition regimes of the white shrimps does not have a significant effect on the nutrition consumption by the shrimps.

TABLE-US-00006 TABLE 6 Attraction capacity of the feed based on insect powder in the nutrition regime of white shrimps with a rate of inclusion of 15% in fishmeal R1 R2 R3 R4 R5 INSM INSM INSM INSM INSM 0% −FM 0% −FM 5% −FM 10% −FM 15% −FM P- Attraction 30% 15% 10% 5% 0% value Nutrition 1.32 1.25 1.18 1.12 1.19 0.340 consumption (g per feed) Start time of 31.35 30.18 27.35 31.13 28.78 0.685 consumption of feedstuffs (seconds) Number of 5.40 5.28 4.85 4.38 4.80 0.307 shrimps consuming feedstuffs within 15 minutes

Results of Experiment 2

[0202] The in vivo digestibility of the insect powder of Tenebrio molitor in white shrimps was determined by the indirect method according to Cho et al. (1985). The apparent digestibility of nutrients is presented in Table 7 below. The results indicate that the insect powder of Tenebrio molitor has a high proteins and lipids content and an energy digestibility of 96 to 97%.

TABLE-US-00007 TABLE 7 Apparent digestibility of the nutrients in the insect powder according to Example 1 (average ± TE). Apparent digestibility Insect powder Digestibility of proteins (%) 97.47 + 1.66 Digestibility of lipids (%) 97.85 + 0.36 Energy digestibility (%) 96.05 + 1.20

Results of Experiment 3

[0203] Efficacy of the Insect Powder on the Performance of White Shrimps

[0204] The effects of the insect powder on the performance of white shrimps (L. vannamei) were studied for 8 weeks. The results on the growth performance of the shrimps were presented in Table 8, as well as in FIGS. 1 and 2. The results showed that from the 8th week, the growth performance of the shrimps in terms of final weight, weight gain, average daily gain and specific growth rate were significantly greater with respect to the control (p<0.05). The shrimps fed with insect powder had a better growth rate than the control (said control having 25% fishmeal, without a supplement of insect powder). The nutrition consumption over the 8 weeks was however not significantly different (p>0.05). Nor was the survival rate over 8 weeks significantly different (p>0.05). The feed conversion rate was significantly lower from week 8 for T2-T5 with respect to the control regime (p<0.05). The protein efficiency ratio is not significantly different (p>0.05).

[0205] Consequently, the insect powder may promote the growth rate of shrimps in a nutrition regime reducing fishmeal up to a replacement of 100% (rate of inclusion of fishmeal 25% in the control regime). The reason is the high digestibility of the proteins and lipids of the insect powder.

TABLE-US-00008 TABLE 8 Growth performance of the white shrimps fed with a nutrition regime comprising insect powder in place of fishmeal and comparison with the control (without insect powder). T1 T2 T3 T4 T5 INSM INSM INSM INSM INSM replaces replaces replaces replaces replaces Growth 0% of 25% of 50% of 75% of 100% of P- performance Periods FM FM FM FM FM value Production 0 weeks 24.03 .sup.a  22.48 .sup.a  24.00 .sup.a  24.95 .sup.a  21.87 .sup.a  0.094 (g/aquarium) 2 weeks 32.33 .sup.a  32.48 .sup.a  33.73 .sup.a  32.18 .sup.a  32.12 .sup.a  0.910 4 weeks 53.28 .sup.a  55.10 .sup.a  54.75 .sup.a  59.62 .sup.a  56.13 .sup.a  0.493 6 weeks 62.75 .sup.a  65.52 .sup.a  68.82 .sup.a  72.30 .sup.a  63.20 .sup.a  0.340 8 weeks 63.77 .sup.b  72.22 .sup.ab  79.00 .sup.a  78.55 .sup.a  71.62 .sup.ab  0.025 Number of 0 weeks 10  .sup.  10  .sup.  10  .sup.  10  .sup.  10  .sup.  / shrimps (per aquarium) Final weight 0 weeks 1.60 .sup.a 1.50 .sup.a 1.60 .sup.a 1.66 .sup.a 1.46 .sup.a 0.098 (g/individual) 2 weeks 2.21 .sup.a 2.22 .sup.a 2.29 .sup.a 2.17 .sup.a 2.17 .sup.a 0.831 4 weeks 3.76 .sup.a 3.86 .sup.a 3.85 .sup.a 4.11 .sup.a 3.93 .sup.a 0.662 6 weeks 4.88 .sup.a 5.08 .sup.a 5.29 .sup.a 5.42 .sup.a 4.97 .sup.a 0.537 8 weeks 5.54 .sup.b 6.43 .sup.a 6.87 .sup.a 6.65 .sup.a .sup. 6.23 .sup.ab 0.030 Weight gain 2 weeks 0.60 .sup.a 0.72 .sup.a 0.69 .sup.a 0.51 .sup.a 0.71 .sup.a 0.213 (g/individual) 4 weeks 2.16 .sup.a 2.36 .sup.a 2.25 .sup.a 2.45 .sup.a 2.47 .sup.a 0.592 6 weeks 3.28 .sup.a 3.58 .sup.a 3.69 .sup.a 3.76 .sup.a 3.51 .sup.a 0.665 8 weeks 3.94 .sup.b 4.93 .sup.a 5.27 .sup.a 4.99 .sup.a 4.77 .sup.a 0.021 Daily weight 2 weeks 0.04 .sup.a 0.05 .sup.a 0.05 .sup.a 0.04 .sup.a 0.05 .sup.a 0.019 gain (g/ 4 weeks 0.08 .sup.a 0.08 .sup.a 0.08 .sup.a 0.09 .sup.a 0.09 .sup.a 0.628 individual/day) 6 weeks 0.08 .sup.a 0.09 .sup.a 0.09 .sup.a 0.09 .sup.a 0.08 .sup.a 0.672 8 weeks 0.07 .sup.b 0.09 .sup.a 0.09 .sup.a 0.09 .sup.a 0.09 .sup.a 0.007 Specific growth 2 weeks 2.29 .sup.a 2.81 .sup.a 2.56 .sup.a 1.88 .sup.a 2.82 .sup.a 0.087 rate (%/day) 4 weeks 3.05 .sup.a 3.38 .sup.a 3.12 .sup.a 3.24 .sup.a 3.52 .sup.a 0.254 6 weeks 2.66 .sup.a 2.89 .sup.a 2.84 .sup.a 2.81 .sup.a 2.91 .sup.a 0.577 8 weeks 2.22 .sup.b 2.59 .sup.a 2.60 .sup.a 2.47 .sup.a 2.59 .sup.a 0.016 Nutrition 2 weeks 0.53 .sup.a 0.50 .sup.a 0.53 .sup.a 0.55 .sup.a 0.48 .sup.a 0.097 consumption 4 weeks 1.67 .sup.a 1.63 .sup.a 1.72 .sup.a 1.65 .sup.a 1.60 .sup.a 0.695 (g/individual) 6 weeks 4.04 .sup.a 4.04 .sup.a 4.07 .sup.a 4.15 .sup.a 4.14 .sup.a 0.977 8 weeks 6.17 .sup.a 6.40 .sup.a 6.26 .sup.a 6.46 .sup.a 6.27 .sup.a 0.975 Daily 2 weeks 0.04 .sup.a 0.04 .sup.a 0.04 .sup.a 0.04 .sup.a 0.03 .sup.a 0.068 nutrition 4 weeks 0.06 .sup.a 0.06 .sup.a 0.06 .sup.a 0.06 .sup.a 0.06 .sup.a 0.605 consumption 6 weeks 0.10 .sup.a 0.10 .sup.a 0.10 .sup.a 0.10 .sup.a 0.10 .sup.a 0.946 (g/individual/ 8 weeks 0.11 .sup.a 0.11 .sup.a 0.11 .sup.a 0.12 .sup.a 0.11 .sup.a 0.985 day) Nutrition 2 weeks 0.90 .sup.a 0.80 .sup.a 0.84 .sup.a 1.21 .sup.a 0.69 .sup.a 0.119 conversion 4 weeks 0.78 .sup.a 0.71 .sup.a 0.78 .sup.a 0.68 .sup.a 0.66 .sup.a 0.270 rate 6 weeks 1.24 .sup.a 1.16 .sup.a 1.11 .sup.a 1.13 .sup.a 1.20 .sup.a 0.683 8 weeks 1.59 .sup.a 1.32 .sup.b 1.20 .sup.b 1.30 .sup.b 1.32 .sup.b 0.052 Survival 2 weeks 97.78 .sup.a  97.78 .sup.a  97.78 .sup.a  98.89 .sup.a  98.89 .sup.a  0.924 rate (%) 4 weeks 94.44 .sup.a  95.56 .sup.a  94.44 .sup.a  96.67 .sup.a  95.56 .sup.a  0.909 6 weeks 85.56 .sup.a  86.67 .sup.a  86.67 .sup.a  88.89 .sup.a  84.44 .sup.a  0.719 8 weeks 76.67 .sup.a  75.56 .sup.a  76.67 .sup.a  78.89 .sup.a  76.67 .sup.a  0.896 Protein 10 weeks  1.80 .sup.a 2.13 .sup.a 2.33 .sup.a 2.12 .sup.a 2.06 .sup.a 0.106 efficiency ratio Comment: The means with the same exponents within the rows are not significantly different (p ≥ 0.05).

[0206] Immunity of the Shrimps

[0207] The immunity of the shrimps under normal conditions was studied over 8 weeks of experiment. The results presented in Table 9 have shown that there was no significant difference with respect to the control (p>0.05) on the total number of haemocytes, the protein content of the haemolymph and the phenol-oxidase activity. Consequently, insect powder can replace fishmeal up to 100% (rate of inclusion of fishmeal 25% in the nutrition formula) without any undesirable effect on the immunity of the shrimps.

[0208] The resistance status after injection of Vibrio parahemolyticus is studied over 6 hours. The immunity of the shrimps in Table 9 shows a high (p<0.05) activity of phenol-oxidase in the group of shrimps fed with the insect powder replacing 50 to 100% of the fishmeal (rate of inclusion of the fishmeal in the nutrition regime from 10.3 to 20.6%). More particularly, in FIG. 3, a negative correlation is noted between the presence of bacteria and phenol oxidase activity. In addition, there is a strong negative correlation between the reduction of bacteria and increasing inclusion of insect powder in the nutrition regime (p<0.05).

TABLE-US-00009 TABLE 9 Immunity of the white shrimps fed with insect powder of Tenebrio molitor as replacement for fishmeal and comparison with the control (without insect powder). T1 T2 T3 T4 T5 INSM INSM INSM INSM INSM replaces replaces replaces replaces replaces 0% of 25% of 50% of 75% of 100% of P- Immunity Status FM FM FM FM FM value Total number of Normal 1.01 .sup.a 1.24 .sup.a 1.16 .sup.a 1.28 .sup.a 1.31 .sup.a 0.160 haemocytes Injection 0.89 .sup.a 1.16 .sup.a 1.03.sup.a  1.16 .sup.a 1.10 .sup.a 0.299 (×10.sup.6 cells/ml) Protein content Normal 4.54 .sup.a 5.71 .sup.a 4.93 .sup.a 6.24 .sup.a 6.01 .sup.a 0.674 of the haemolymph Injection 5.83 .sup.a 6.21 .sup.a 4.99 .sup.a 3.53 .sup.a 4.06 .sup.a 0.066 (g/dl) Phenol-oxidase Normal 113.24 .sup.a  141.70 .sup.a  135.43 .sup.a  137.93 .sup.a  121.98 .sup.a  0.961 activity (unit/ Injection 29.29.sup.c  30.24.sup.c  66.29.sup.b  74.32 .sup.ab  104.64 .sup.a  0.002 mg protein) Bacteria reduction Injection 4.33 .sup.a .sup. 4.25 .sup.ab .sup. 4.03.sup.bc .sup. 3.82.sup.cd 3.72.sup.d  0.001 capacity (or clearance) (Log UFC/ml) Comment: The means with the same exponents within the rows are not significantly different (p ≥ 0.05).

[0209] Mortality of the Shrimps

[0210] The mortality of the shrimps after exposure to Vibrio parahaemolyticus, 4.3×10.sup.5 ufc/ml, has been presented in Table 10, as well as in FIG. 4. The shrimps fed with nutrition regimes in which fishmeal was reduced (by 50 to 100%) and supplemented by insect powder of Tenebrio molitor (replacing fishmeal by 50 to 100%) showed a significantly lower mortality rate (P<0.05) than the control nutrition regime with 25% fishmeal (T1) and than the group of shrimps fed with nutrition regimes in which the fishmeal was reduced and supplemented by insect powder (25% fishmeal replaced). This may result from the high lipid and protein digestibility of the insect powder and from a possible better quality of the lipids with respect to soya oil, allowing the shrimps to obtain more nutriments.

TABLE-US-00010 TABLE 10 Cumulative mortality rate after injection of the pathogen Vibrio parahaemolyticus in white shrimps fed with nutrition regimes comprising insect powder in place of fishmeal and comparison with the control (without insect powder) T1 T2 T3 T4 T5 INSM INSM INSM INSM INSM Day(s) after replaces replaces replaces replaces replaces injection of 0% of 25% of 50% of 75% of 100% of P- the pathogen FM FM FP FM FM value Day 1 0.00.sup.  0.00  .sup. 0.00  0.00 .sup.ab 0.00  * Day 2 10.00 .sup.a  3.33 .sup.ab .sup. 0.00 .sup.b  6.67 .sup.ab .sup. 0.00 .sup.b 0.029 Day 3 23.33 .sup.a 13.33 .sup.ab .sup. 3.33 .sup.b 13.33 .sup.ab .sup. 3.33 .sup.b 0.008 Day 4 30.00 .sup.a 23.33 .sup.ab .sup. 6.67 .sup.b 13.33 .sup.ab 10.00 .sup.b  0.048 Day 5 40.00 .sup.a 30.00 .sup.ab  10.00 .sup.b 13.33 .sup.ab 13.33 .sup.ab 0.014 Day 6 40.00 .sup.a 30.00 .sup.ab  10.00 .sup.b 13.33 .sup.ab 13.33 .sup.ab 0.014 Day 7 40.00 .sup.a 30.00 .sup.ab 10.00.sup.c 16.67 .sup.bc 13.33 .sup.bc 0.017 Day 8 40.00 .sup.a 30.00 .sup.ab 10.00.sup.c 16.67 .sup.bc 13.33 .sup.bc 0.017 Day 9 40.00 .sup.a 30.00 .sup.ab 10.00.sup.c 16.67 .sup.bc 13.33 .sup.bc 0.017 Day 10 43.33 .sup.a 30.00 .sup.ab 10.00.sup.c 16.67 .sup.bc 13.33 .sup.bc 0.005 Mortality 43.33 .sup.a 30.00 .sup.ab 10.00.sup.c 16.67 .sup.bc 13.33 .sup.bc 0.005 rate (%) Survival 56.67.sup.c  70.00 .sup.bc  90.00 .sup.a 83.33 .sup.ab 86.67 .sup.ab 0.005 rate (%) Comment: The means with the same exponents within the rows are not significantly different (p ≥ 0.05). * no statistical analysis, the value is the same.

[0211] Histology of the Hepatopancreas and the Intestinal Villi of the Shrimps

[0212] The hepatopancreatic histology of the shrimps fed with the control nutrition regime and the regimes with the insect powder showed larger B cells and R cells. These cells produce nutriment regulation enzymes. In the nutrition regimes T2-T5, the B (secretory) and R (reserve) cells enlarged more than in the control regime T1. This means that the hepatocyte cells, having accumulated more nutrients, can be reserves of glycogen and/or lipids, as a result of the high protein or lipid digestibility of the insect powder. The density and length of intestinal villi were high under all the nutrition regimes.

3. CONCLUSION

[0213] Insect powder, in particular from Tenebrio molitor, can replace fishmeal, up to 100% replacement of the fishmeal, without any negative effect on the growth performance, the health of the shrimps and the histology of the hepatopancreas and the intestinal villi in small-sized shrimps (1-5 g) with an equivalent nutritional profile between the nutrition regimes comprising fishmeal and/or insect powder. When the insect powder replaces between 25 and 100% of the fishmeal in the nutrition regime in shrimps during more than 6 weeks, the shrimp may have the best growth rate and a high digestibility. The group of shrimps fed with insect powder from Tenebrio molitor as replacement for fishmeal at 50 or 100% showed a better resistance to the pathogen Vibrio parahaemolyticus.