RU2013448C1 - Strain streptomyces bambergiensis - a producer of moenomycin a - Google Patents

Abstract

FIELD: microbiological industry. SUBSTANCE: strain Streptomyces bambergiensis VKPM S-1111 is proposed. Strain is a producer of antibiotic moenomycin A. The level of activity of this antibiotic after 120-140 h cultivation at 29 C under conditions of intensive aeration attains 3000-3500 U/ml that is 1.5-time more as compared with activity level of parent strain VKPM S-826. EFFECT: enhanced yield of antibiotic.

Description

 The invention relates to the microbiological industry and relates to a strain of a microorganism of the species Streptomyces bambergiensis producing moenomycin, a non-medical antibiotic.

 Moenomycin is a member of the moenomycin antibiotic group, the producers of which, in addition to S. bambergiensis, are microorganisms of the species S. ghanaensis, S. geysiriensis. S. ederensis. The antibiotics produced by these microorganisms are formed for the most part as long chains of molecules with complexes of very similar components with a relatively high molecular weight from 1,600 to 2,100 daltons and a phosphorus content of about 2%. The greatest biological activity among them is shown by moenomycin A produced by S. bambergiensis.

 Moenomycin A is a phosphoglycolipid antibiotic with a complex structure, consisting of a disaccharide with an amino group, an amino sugar, a phosphoric acid residue connected in the form of an ether bond with a polyhydric alcohol moenocinol; the composition of moenomycin also includes a UV chromophore, which is a disaccharide with an acetamino group connected to an aminocyclopentadione.

 Moenomycin is active against gram-positive bacteria, and in high concentrations - against gram-negative, not active against fungi and yeast. On sensitive cultures, the antibiotic has a bacteriostatic effect in low concentrations and bactericidal in high.

Due to the specificity of the action of moenomycin A, its toxicity to warm-blooded animals is very low. For mice, the lethal dose (LD ₅₀ ) is 738 mg / ml.

 The high specificity of moenomycin against staphylococci and streptococci resistant to tetracycline, penicillin, streptomycin, novobiocin and erythromycin, as well as its low toxicity to warm-blooded animals makes the use of this antibiotic especially valuable in animal husbandry, as it helps to reduce the incidence of animals and, consequently, increase survival young animals, improving feed digestibility and accelerating growth.

 Flavomycin (bambermicin) - commercial names of moenomycin preparations obtained on the basis of various streptomycetes.

 Flavomycin producing strains are known that belong to different types of streptomycetes: S. bambergiensis ATCC 13879, S. ghananensis ATCC 14672, S. ederensis ATCC 15304, S. geysirensis ATCC 15303 (Federal Republic of Germany patents NN 1113791, 1210514, 1236735, 1236726 Noeshst ", auth. St. NRB N 31014, registered in DIKLS under N 69/1980.

 The best indicators of the level of accumulation of moenomycin A were recorded on the producer strain S. bambergiensis VKPM S-826 (ed. St. USSR N 4417858). When using this strain, the maximum accumulation of moenomycin A is achieved after 140 hours of fermentation and averages 1600 units / ml with a maximum accumulation level of 2000 units / ml. The disadvantage of the strain of the prototype is the lack of synthesis of moenomycin.

 The aim of the invention is the creation of a new producer strain of moenomycin, which allows to obtain an antibiotic in a higher concentration in the growth medium.

 The goal is achieved by creating a new strain of S. bambergiensis 1-39, deposited in the All-Union collection of industrial microorganisms and having a registration number VKPM S-1111. A new strain of S. bambergiensis VKPM S-1111 was obtained by multistage selection from strain S-826 after mutagenesis.

 Characterization of the strain S. bambergiensis VKPM S-1111.

 Morphological and cultural characters.

 The cells of the mycelium are filiform, spore-bearing straight, spores oval, smooth, covered with hairs.

Growth on agar media (on the 12th day of growth at 28 ^about C).

 On Gauze-1 medium of standard composition, the strain forms slightly convex colonies with a diameter of 5-7 mm. The surface of the colonies is smooth, spores of gray-violet color, can be with exudate, at the edges a narrow (0.5 mm) rim of white color is possible. The edges of the colony are even or slightly wavy. Creamy aerial mycelium. The substrate mycelium is dark brown. Around the colony there is a starch hydrolysis zone with a diameter of 25-30 mm.

 On CP-6 medium of standard composition, the strain forms light cream colonies with a diameter of 5-7 mm with a folded surface and wavy edges. The center of the colony is slightly elevated. The spores are white, the substrate mycelium is red-brown.

 On a liquid fermentation medium (the composition is given below), strain S-1111 forms large plexuses of hyphae with pronounced basophilia. On the 3rd day of fermentation, the culture fluid acquires a dark purple color. The precipitate is gray-violet.

 Physiological and biochemical properties.

 Attitude to carbohydrate sources.

 Strain VKPM S-1111 weakly dilutes gelatin, hydrolyzes starch well. Like the original strain S-826, the new strain S-1111 grows well and synthesizes moenomycin on mannose, xylose. For the growth of culture, almost all carbohydrates are absorbed.

 Attitude to nitrogen sources.

 The new strain S-1111, like the original strain S-826, does not grow in the presence of L-cystine and L-tyrosine, absorbs nitrogen in the form of ammonium salts, does not grow on urea.

 Strain storage.

The culture of the strain S. bambergiensis VKPM S-1111 is stored on the jambs at 2-4 ^{° C} or freeze-dried state in glass sealed ampoules at room temperature.

 Obtaining moenomycin using a new producer strain.

 The cells of the producer strain S. bambergiensis VKPM S-1111 are grown on Gauze-1 agar medium, and then seeded in flasks with inoculum containing soy flour, corn extract, beet pulp or wheat bran as a source of organic nitrogen. As a carbon source, potato or corn extract, dextrin, molasses can be used.

The inoculum, grown for 30-40 hours at 28-30 ^{° C} in a volume of 10%, was transferred to a flask with a fermentation medium containing a carbon source in molasses, various starches, or dextrin, as well as the nitrogen source, soybean meal, corn steep liquor , wheat bran, beet pulp, various oils (soya, castor, olive, sunflower, corn).

The fermentation was carried 120-140 hours at 28-30 ° ^C. At the end of fermentation, the antibiotic is partially released into the growth medium, but the main part (over 90%) is found in the mycelium is extracted from ethyl or methyl alcohol in a ratio of 1: 4. The activity of the alcohol the extract is determined by agar diffusion using Vasillus cereus as a test culture. It is possible to carry out fermentation under the same conditions, bypassing the seed medium, sowing spores from the agar medium directly into flasks or tubes.

EXAMPLE EXAMPLE 1 An aqueous spore suspension of strain S. bambergiensis VKPM S-1111, obtained by flushing culture grown at ²⁸ ° C for 12 days on a medium Gauze-1, was inoculated into 750 ml flasks containing 50 ml of seed medium of the following composition, g / l: soy flour 20; starch 20; sodium chloride 2.0; potassium phosphate disubstituted 1.0; chalk 4.0; pH 7.0-7.2. The flasks are placed on a circular rocking chair (240 rpm) and the seed is grown for 30-40 hours at 29 ^about C.

Prepared seed material with a volume of 10% is seeded with 750 ml flasks containing 50 ml of fermentation medium of the following composition, g / l: soy flour 30; molasses 30; starch 30; ammonium sulfate 3.0; monosubstituted potassium phosphate 1.0; sodium chloride 2.0; chalk 4.0; soybean oil 0.5 ml; pH 7.0-7.2. The flasks are placed on a circular rocking chair (240 rpm) and grown 120-140 hours at 29 ^about C.

 Activity at the end of fermentation is 3500 units / ml.

 PRI me R 2. An aqueous suspension of spores obtained as in example 1, seeded flasks with a fermentation medium, bypassing the seed medium. The composition of the medium and the fermentation conditions are the same as in example 1.

 Activity at the end of fermentation is 3000 u / ml.

 PRI me R 3. Filmed spores of the strain VKPM S-1111 grown as in example 1, seeded in tubes with a diameter of 20-22 mm with 5 ml of fermentation medium. The composition of the medium and the fermentation conditions are the same as in example 1.

 Activity at the end of fermentation is 3000 u / ml.

 Thus, the claimed strain of S. bambergiensis VKPM S-1111 in terms of activity (3500 units / ml) is one and a half times higher than the strain prototype VKPM S-826 (2000 units / ml).

 Using the proposed strain will not only create a technology for the industrial production of moenomycin A, but also increase the efficiency of its production.

Claims (1)

 Strain Streptomyces bambergiensis VKPM S-1111 - producer of moenomycin A.