WO2021015648A2

WO2021015648A2 - Protein feed additive for livestock animals and fish

Info

Publication number: WO2021015648A2
Application number: PCT/RU2020/050121
Authority: WO
Inventors: Елена Сергеевна БАБУСЕНКО; Валерий Алексеевич Быков; Маргарита Витальевна ЛАЛОВА; Леонид Евгеньевич Левитин; Александр Иванович Сафонов; Наталья Леонидовна Куликова
Original assignee: Общество с ограниченной ответственностью "ГИПРОБИОСИНТЕЗ"
Priority date: 2019-07-22
Filing date: 2020-06-15
Publication date: 2021-01-28
Also published as: JP2022541726A; EA202091614A1; JP7618246B2; RU2717991C1; WO2021015648A3

Abstract

The invention is intended for use in animal and fish farming, and in particular relates to feed bio-additives for feed when raising livestock animals and fish in artificial water bodies to improve productivity and natural resistance. The technical result obtained by using the proposed feed additive is the creation of an efficient technology for obtaining an additive, improved usage properties of the additive due to increased productivity of the livestock animals and fish. The technical result has been achieved by using a protein feed additive for livestock animals and cultured fish which contains a strain of the methane-oxidizing bacteria Methylococcus capsulatus (GBS-15) VKPM B-12549. The additive additionally contains a strain of the heterotrophic bacteria Cupriavidus gilardii (GBS-15-1) VKM B-3265D and/or a strain of the heterotrophic bacteria Stenotrophomonas acidaminiphila (GBS-15-2) VKM B-3264D and/or a strain of the heterotrophic bacteria Klebsiella pneumonia 1-17 "GKPM - OOBOLENSK" B-8465, taken in equal amounts, with the following ratio of components by wt.%: 85-94 of a strain of the methane-oxidizing bacteria Methylococcus capsulatus VKPM B-12549 : 6-15 of the strain mixture.

Description

PROTEIN FODDER ADDITIVE FOR AGRICULTURAL

ANIMALS AND FISH

The invention is intended for use in animal husbandry and fish farming, in particular, it relates to feed bioadditives for feed when growing farm animals and fish in artificial reservoirs to increase productivity, natural resistance.

Currently, there is a decrease in the quality of feed, including their energy and protein nutritional value. Almost all types of feed used belong to the low-protein group, therefore, the supply of protein (in particular for cattle) does not exceed 85 g, instead of 108 g according to the norms, which is one of the most important reasons for the low productivity of animals, increased feed consumption per unit of production and its high cost price.

The attention of researchers was drawn to the use of herbal ingredients, in particular to sources of vegetable protein, due to their availability and low cost. The main source of the protein product is soybean meal. However, the natural conditions of our country are not suitable for growing soybeans in sufficient quantities. Fishmeal has traditionally been the main source of protein for fish nutrition. However, the price of fishmeal is steadily increasing and the volume of aquaculture production is increasing.

Experts have to look for other ways to produce feed protein and alternative sources of their production.

The introduction of a protein feed additive into the mixture allows to optimize the nutritional composition of feed, improve their bioavailability and influence on the rate of development of animals and fish, provide a prophylactic effect, etc.

As a result of the conducted patent search, the following sources of information were selected.

Thus, technologies have been developed for the production of fodder protein on the basis of waste alcohol (distillery stillage) and milling (bran, flour) industries and a mixture of stillage with grain waste (RF patent N ° 2159287; RF patent N ° 2140449; USSR author's certificate N ° 1595900; USSR author's certificate JSfo 1571061; USSR author's certificate N ° 1532580; USSR author's certificate JSfo 1507787; RF patent N ° 2054881; N ° 2042713, etc.). Yeast-like imperfect fungi are used as producer crops in the listed inventions. genus Candida. The disadvantage of the known strains of microorganisms is the lack of the ability to actively grow under conditions of a long non-sterile cultivation process. In addition, the use of the yeast of the genus Candida as a producer culture requires the complication of the production technology by the mandatory introduction of the stage of heat treatment of the resulting feed biomass to ensure the absence of living cells in the finished product.

Various bacterial strains are known that are protein producers (RF patents N ° N ° 2244000, 2390554, 2244001). A common disadvantage of these strains is the insufficiently high concentration of biomass accumulated in the presence of 2.5% lactic acid, and, consequently, the insufficient amount of protein that could be formed by the culture.

One of the promising ways to obtain a complete protein feed product is methanotrophic bacteria that produce protein. Under suitable conditions, methanotrophic bacteria actively process natural gas, multiply rapidly and build up their biomass rich in valuable protein, vitamins and other biologically active substances.

The use of methane for protein production has a number of advantages over liquid hydrocarbons: large reserves of natural gas, good transportability, and the possibility of obtaining a finished product without additional purification from the substrate.

Considering that Russia has large natural gas reserves, according to some sources, they account for up to 40% of the world. The introduction of microbiological protein production at Russian enterprises promises not only an economic effect, but is also capable of ensuring the country's food security.

Various strains of bacteria are known that produce fodder protein belonging to different types of methane-oxidizing bacteria (methanotrophs), for example, for example, strains of the genus Pseudomonas methanica, Methylococcus capsulatus BKM B-2116, Methylocystis parvus BKM B-2129, MethylosMinpo sporium BKM BKM B-2129, MethylosMinpox sporium BKM BKM B-2117, Methylobacter acidophilus, Methylomonas rubra VSB-90, Methylococcus sp. ChM-9, Methylococcus capsulatus VSB-874, Methylococcus minimus, Methylomonas methanica, Methylomonas agile.

They are products of microbial synthesis obtained as a result of the cultivation of methane-oxidizing bacteria on natural gas. These strains are characterized by different growth rates and biomass yield. Known strain Methylococcus capsulatus VSB-874 - producer of biomass. It is kept in the Museum of Cultures of the VNIIgenetics Institute under the collection number TsMPM B - 1743 (author certificate of the USSR N ° 770200). Methane, both pure and in the composition of natural gas, and methanol are used as a carbon source. The product using this strain is produced under the GAPRIN trademark. The disadvantages of this strain are insufficiently high productivity (8-10 g / l h), low content of crude protein in biomass (no more than 70%), unstable biomass yield (30-60 g / l).

Known invention (RF2687137), which relates to biotechnology and can be used to obtain a microbial protein mass for animal feeding. The strain of heterotrophic bacteria Klebsiella pneumonia 1 -17, capable of using the products of co-oxidation of methane homologues present in natural gas, was deposited in the State Collection of Pathogenic Microorganisms and Cell Cultures "GKPM - OBOLENSK" under registration number B -8465. This strain provides an increase in the productivity of methane-oxidizing bacteria.

Known invention, which relates to biotechnology and can be used to obtain a microbial protein mass for animal feeding. The strain of heterotrophic bacteria Cupriavidus gilardii GBS-15-1, capable of using the products of co-oxidation of methane homologues present in natural gas, was deposited in the All-Russian collection of microorganisms of the Institute of Biochemistry and Physiology of Microorganisms of G.K. Skryabin RAS under registration number VKM B-3265D. This strain provides an increase in the productivity of methane-oxidizing bacteria (RF 2687135).

Known invention, which relates to the microbiological industry. The strain of heterotrophic bacteria Stenotrophomonas acidaminiphila GBS-15-2 has the ability to use the products of co-oxidation of methane homologues present in natural gas. The strain of heterotrophic bacteria Stenotrophomonas acidaminiphila was deposited in the All-Russian collection of microorganisms at the Institute of Biochemistry and Physiology of Microorganisms of G.K. Skryabin RAS under the registration number VKM B-3264D. The strain of heterotrophic bacteria Stenotrophomonas acidaminiphila VKM B-3264D can be used to obtain microbial protein mass (RF JV "2687136).

The closest of the analogs in technical essence and the claimed effect is a strain of methane-oxidizing bacteria Methylococcus capsulatus GBS-15. The area of application of the strain is the production of protein-vitamin complexes for balancing feed for agricultural animals. Deposited in the All-Russian Collection of Industrial Microorganisms under the number VKPM B -12549. The strain provides biomass accumulation up to 32 g / l. (RF N ° 2613365). The content of crude protein in biomass is not less than 79%.

The technical task is to expand the range of used protein feed additives based on methane-oxidizing bacteria Methylococcus capsulatus for farm animals and fish grown in artificial reservoirs, which have a higher technological potential and cost-effective industrial process, high protein content.

The technical result obtained from the use of the proposed feed additive is the creation of an effective technology for obtaining the additive, improved operational properties of the additive by increasing the productivity of farm animals and fish.

The technical result was achieved through the use of a protein feed additive for farm animals and cultured fish, including a strain of methane-oxidizing bacteria Methylococcus capsulatus (GBS-15) VKPM B-12549. The supplement additionally contains a strain of heterotrophic bacteria Cupriavidus gilardii (GBS-15-1) VKM B-3265D and / or a strain of heterotrophic bacteria Stenotropkomonas aeidanmupkila (GBS-15-2) VKM B-3264D and / or a strain of heterotrophic bacteria Klebsiella pneumonia " GKPM - OOBOLENSK "V-8465, taken in equal amounts, with the following ratio of components, wt.%:

- strain of methane-oxidizing bacteria Methylococcus

capsulatus VKPM B-12549 85 - 94

- a mixture of strains 6 - 15

Feeding the diet with a protein feed supplement containing the proposed mixture of strains contributes to an increase in body weight due to muscle tissue gain. Meat quality is improved by reducing muscle fat, reducing subcutaneous fat, and increasing the muscle eye area.

The invention will increase the productivity of animals and fish, their survival, the ecological safety of meat, and obtain meat with functional properties.

The claimed ratio of the components of the protein feed additive provides the creation of an easily digestible high-protein additive with a content of heterotrophic bacteria. A protein feed additive for farm animals and fish is obtained by mixing the components according to the stated ratio and then, if necessary, granulated or extruded. Table 1 shows the formulations of the proposed protein supplements.

The following experiments were carried out with variants of the protein feed additive:

- option 1 - two-component mixture - GBS-15 strain and GBS -15-2 strain;

- option 2 - three-component mixture - GBS-15 strain, GBS -15-1 strain and GBS -15-2 strain;

- option 3 - four-component mixture - strain GBS-15, strain GBS-15-1, strain GBS -15-2 and strain 1-17;

- option 4 - an additive based on one GBS-15 strain.

As shown by the experiments, the use of the proposed protein feed additive is essential in mixtures of feed additives to optimize the nutritional composition of feed, improve their bioavailability. In addition, the need for fish, animals and poultry for the necessary nutritional components is ensured, and feed costs per unit of production are reduced.

Methanotroph Methylococcus capsulatus GBS-15 is the producer of the protein feed additive. The strain was isolated in the process of long-term multistage selection by cultivation under aeration and stirring in a liquid mineral medium in an atmosphere of natural gas. Strain Methylococcus capsulatus GBS-15 is cultivated as part of an associative culture, including heterotrophic associates Cupriavidus gilardii, Stenotrophomonas acidaminiphila, Klebsiell apneumoni, which utilize the products of co-oxidation of methane homologues present in natural gas of Methylcoccus C, and metabolic products of the main methane strain, Methylloc.

The resulting protein feed additive is a dried inactivated biomass of the association of methane-oxidizing and heterotrophic bacteria.

Protein feed additive in the form of powder or granules is dosed in the required amount, which is determined by the norms of use in the preparation of feed mixtures. The protein feed additive has a high nutritional value and is harmless to fish, animals and poultry.

Strain Methylococcus capsulatus GBS-15 and its associates are non-pathogenic, non-toxic and harmless.

The amount of protein in the protein feed additive is equivalent to animal proteins (fish meal and meat and bone meal). This allows widespread use protein feed additive as a substitute for fish meal and soybean meal in the composition of feed rations.

The biological value of the protein feed additive for chemical Scor is 65-70%. The limiting amino acid is methionine. The relative biological value to casein on the tetrachimenapyriformis test organism is 65-75%.

Studies of the nutritional properties of the protein feed additive were carried out on animals for which the product is intended: pigs, poultry, calves, sheep, fish. The animals were fed according to the accepted norms. Bean meal was included in the control diets as protein sources. The amount of the claimed protein feed additive and control additive included 20% of the daily rate of digestible protein in the diet.

The essence of the invention is illustrated by the following examples.

GAPRIN was used as a control food in all examples.

Example 1 Test on fish

The tests of feed additives were carried out on carp fingerlings weighing 38-45 g under laboratory conditions in flow-through aquariums. Carp underyearlings, 20 pcs. kept in 70 l flow-through aquariums at a water temperature throughout the experiment in the range of 18-21 ° C, and with an oxygen content of 6.8 to 9 mg / l. The course of feeding the feed additive was 24 days.

Upon completion of the course of feeding the fish at all stages of the experiment, a comparative assessment of their physiological state was carried out by hematological parameters. All calculations of the absolute, relative, and average daily gain in average weight were carried out according to generally accepted methods.

For the test results, see table 2.

The protein feed additive can be added to fish feed without disturbing the amino acid balance of the feed. The proposed mixture of strains has a high level of productivity, which contributes to an increase in the rate of food consumption of fish and the rate of weight gain.

Studies of feeds with the addition of a protein feed additive on fish of different ages have shown that the inclusion of this component in the diet stimulates the growth rate of fish while reducing feed costs, without disturbing the physiological status of the fish. Larval survival is also related to the diet. The presence of a protein feed additive in experimental feed significantly increases the survival rate of carp, up to 93% versus 63% on the control feed. Thus, when the nutrients in the fish feed are properly balanced, the protein feed supplement shows results similar to other high protein components.

Example 2 Tests on pigs.

Tests of the biological product in the diets of pigs were carried out at different age periods and on different age and gender groups:

The animals were fed according to accepted standards. Soybean meal was included as protein sources in the control diets.

In the experimental groups, a dietary supplement was included in the amount of up to 20% of the daily rate of digestible protein in the diet. Animals were weighed individually before testing, at the end of each growing period. The physiological state and health of the animals throughout the experiment were normal. No negative effects were found throughout the experiment. For the entire period of testing, no differences were noted in the health status of animals, feed consumption and the value of average daily gains in experimental and control animals.

The level of protein feed additive in the experimental group was 20% of the daily rate of digestible protein. The nutritional value of the diets corresponded to generally accepted norms for the corresponding age of the animals and was identical for the control and experimental groups.

Throughout the experiment, no deviations in the physiological state and health of the animals were noted.

The use of a protein feed additive in the composition of mixed feed for pigs contributed to an increase in weight gain in comparison with control by 12%, did not cause metabolic disorders, contributed to the high safety of young animals by 7.5%, there were no cases of an increase in the incidence of pigs in the experimental groups.

Research results indicate the effectiveness and harmlessness of using a protein feed additive in the diets of growing and fattening pigs.

Example 3 Test on poultry

Productivity studies were carried out on broiler chickens. The poultry of the experimental group received feed, which included 5% protein feed additive by weight of the diet instead of 50% protein of animal origin. The chickens in the control group received complete standard feed.

The main physiological indicators in the control and experimental groups as a result of the experiment were: the safety of chickens in the experimental group was 99.2%, in control group 98.8%. The intensity of egg production in the experimental group is 79.4%, in the control group 72.4%.

Table 1

table 2

Claims

Claim

Protein feed additive for farm animals and cultured fish, including a strain of methane-oxidizing bacteria Methylococcus capsulatus VKPM B-12549, characterized in that the additive additionally contains a strain of heterotrophic bacteria Cupriavidus gilardii VKM B-3265D and / or a strain of heterotrophic acidaomonas VKM VKM B-3265D and / or a strain of heterotrophic bacteria StenKM-64 and / or the strain of heterotrophic bacteria Klebsiella pneumonia 1-17 "GKPM - OOBOLENSK" B-8465, taken in equal amounts, with the following ratio of components, wt%:

- strain of methane-oxidizing bacteria Methylococcus

capsulatus VKPM B-12549 85 - 94

- a mixture of strains 6 - 15