RU2168909C1 - Strain of bacterium lactobacillus plantarum for fodder ensilage - Google Patents

Abstract

FIELD: biotechnology, microbiology. SUBSTANCE: invention relates to preparing the novel strain of bacterium Lactobacillus plantarum 60 VNIISHM N 78D used for effective multiplication if green mass to be ensilaged and for lactic acid synthesis. Strain Lactobacillus plantarum 60 exhibits the enhanced multiplication activity for decreased time as compared with other known bacterial strains (for example, Streptococcus faecium N 500, Lactobacillus plantarum 52) to form lactic acid mainly. Strain shows the more expressed antagonistic properties with respect to putrefactive bacteria occurring in ensilage, for example, Bacillus megatherium, Bacillus subtilis, Bacillus mycoides. This strain exhibits efficiency for ensilage of different grasses, among them, goat's-rue (Galega officinalis), orchard-grass (Dactylis glomerata) and mixture of alfalfa (Medicago) with grasses. EFFECT: new strain of bacterium indicated above, improved ensilage quality. 3 tbl, 2 ex

Description

 The invention relates to agricultural microbiology in feed production, in particular to a preparation for silage of feeds, and relates to a new strain of bacteria to obtain said preparation.

 Known and closest to the claimed strain on the technical nature of the strain Streptococcus faecium N 500 described in ed. St. N 1514766, MKI C 12 N 1/20 (C 12 N 1/20, C 12 R 1/46, A 23 K 3/00), published in the Official Bulletin of Inventions of the USSR N 38 of 10/15/89. The strain was deposited in VNIISKhM culture collections under registration number 42.

 The strain of Streptococcus faecium N 500 isolated from silage (a mixture of sunflower, oats and peas) late ripening with benzoic acid, consists of bacteria Streptococcus faecium.

 The strain is intended for the silage of alfalfa, hedgehogs and mixtures of alfalfa with cereals. It has a high reproduction activity and the ability to form mainly lactic acid in a short time. So, for example, when siloing alfalfa, silage is obtained with an increase in the content of lactic acid by 0.11%, protein by 3.2%; the digestibility of the silo obtained in this way increases by 3.9% compared with the control.

 The strain Streptococcus faecium N 500 (prototype) obtained by the above method is able to effectively suppress putrefactive microflora and obtain high-quality silage from plant materials.

 The disadvantage of using the drug for the preparation of silage containing a strain of bacteria Streptococcus faecium N 500 is the low content of protein and water-soluble sugars in the silo.

 The objective of the invention is to obtain a new strain of bacteria that can more efficiently multiply in a silo green mass, synthesize lactic acid, suppress putrefactive microflora and get better silage from plant materials.

 This problem is solved due to the fact that for the silage of feed as a microorganism, a bacterial strain Lactobacillus plantarum 60 is used.

 The strain Lactobacillus plantarum 60 was obtained as a spontaneous mutant of the strain Lactobacillus plantarum 52 and is able to grow on a medium with 10% potassium chloride. Acquired tolerance to increased osmotic pressure, thus acquired, increased its ability to reproduce in a silted green mass.

 The strain Lactobacillus plantarum 60 was deposited in the collection of the All-Russian Research Institute of Agricultural Microbiology (VNIIISHM) on April 06, 2000 under N 78 D.

 The strain is characterized by the following morphological-cultural and physiological-biochemical characteristics.

 Cells are in the form of rods with rounded ends of 1.5-4.0 microns, do not form spores, motionless, gram-positive. Optional anaerobic.

 On meat-peptone agar grows weakly, forming a subtle drip growth along the line. On meat-peptone agar, growth is weak. On wort agar with chalk it forms characteristic colonies (with a chalk enlightenment zone) - lenticular with smooth edges or surface round, pasty consistency, white or cream. The diameter of the colony is 0.6 - 1.2 cm.

The optimum temperature for growth is 24-40 ^o C. At a temperature of more than 45 ^o C and less than 23 ^o C, growth is slow.

 The optimal pH of the medium is 5.5, and growth occurs at pH from 4.0 to 6.5.

 The strain grows well on media containing up to 10% NaCl. In milk containing methylene blue (0.1%), there is no growth.

The culture can withstand a temperature of 60 ^o C for 30 minutes, but the growth is weak.

 On media containing 40% bile, the strain grows well.

 The strain needs growth stimulants.

 Attitude to carbohydrates: ferments cellobiose, fructose, galactose, glucose, lactose, maltose, mannitol, mannose, raffinose, ribose, sorbitol, sucrose; weakly ferments arabinose; does not ferment ramnose.

 Catalase does not form. It exhibits antagonistic properties against putrefactive bacteria found in silos: Bacillus megaterium, Bacillus mesentericus, Bacillus subtilis, Bacillus mycoides.

 The energy of acid formation and the main products of fermentation of lactic acid bacteria are determined during their cultivation in the wort, followed by the determination of organic acids by the Wigner method.

 Table 1 presents the results of studying the growth and synthesis of organic acids by three strains of bacteria (Streptococcus faecium N 500 - prototype, Lactobacillus plantarum 52 - the original strain, Lactobacillus plantarum 60 - the proposed strain).

 As can be seen from table 1, the strain Lactobacillus plantarum 60 has a higher reproduction activity and the ability to form mainly lactic acid in a shorter time.

The strain Lactobacillus plantarum 60 is maintained on the must with chalk replanting after 3 months. It is stored in a refrigerator at + 4 ... + 5 ^o C.

 Table 2 presents the results of a study of the antagonistic relationships of the above three strains of lactic acid bacteria (Streptococcus faecium N 500 - prototype, Lactobacillus plantarum 52 - the original strain, Lactobacillus plantarum 60 - the proposed strain) in relation to putrefactive bacteria found in the silo.

 As can be seen from table 2, the strain Lactobacillus plantarum 60 in relation to putrefactive bacteria in all of the tests has more pronounced antagonistic properties.

 An example of obtaining a drug for silage.

Preparation of uterine culture. In a sterile nutrient medium containing (in vol.%) Molasses - 1.0, biotin - 0.5, meat and bone meal - 0.9, sodium chloride - 6.0, tap water - the rest, bacteria bring in the amount of 1 -2 vol.% And grown for 24-30 hours at a temperature of 25-30 ^o C.

Production cultivation of the strain in a fermenter. Prepare the medium of the specified composition, sterilize, cool to 30-32 ^o C and make 4 vol.% Of seed obtained by the specified method. Cultivation is carried out at 28 - 30 ^o C for 20-24 hours at a pressure of 0.3 - 0.5 bar without blowing air. The agitator is turned on immediately after applying the seed for 3-5 minutes, and then every 3-5 hours of cultivation and before sampling.

Fermentation for silage is a light brown liquid with a sediment of a nutrient medium and contains lactic acid bacteria in an amount of 900 - 1100 million / ml. The activity of lactic acid bacteria (Turner acidity) - 25-30 ^o C.

 An example of using the resulting bacterial starter culture.

 For silage use goatskin, hedgehog team and a mixture of alfalfa with cereals. The crushed plant mass in the amount of 1.5 kg is laid in glass vessels with a volume of 3 l and bacterial yeast in the amount of 0.01% by weight of the green mass is added. The fermented mass is thoroughly mixed, tamped, the jars are covered with plastic wrap and poured with molten wax. The quality of the silo is determined after 90 days.

 The results of the experiments are presented in table 3.

Claims (1)

 The bacterial strain Lactobacillus plantarum 60 VNIISCHM N 78D for silage feed.

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