RU2067616C1 - Strain of bacterium bacillus subtilis carrying property antibioticoresistance used for preparation "bactisporin" making - Google Patents

Abstract

FIELD: biotechnology. SUBSTANCE: probiotic, antibioticoresistant strain of Bacillus subtilis 3H "ГИСК" N 248 is obtained. Cell sizes: 1.5-4.0 mcm on 0.4-0.6 mcm. Cells are gram-positive, form spores but not - capsules, aerobic respiration. Strain grows on the different nutrient media. Strain splits glucose, sucrose, mannitol, maltose; it hydrolyzes starch, casein, urea, peptonizes milk, liquefies gelatin, reduces nitrates to nitrites, utilizes citrate, forms ammonia and hydrogen sulfide; strain does not split tyrosine, does not form hemolysin and fibrinolysin. Strain inhibits the growth of staphylococci, proteus, klebsiellae, Escherichia coli, yeast fungi, streptococci and pyocyanic bacterium. Strain shows resistance to polymixin and rifampicin, low sensitivity to penicillin, lincomycin and streptomycin. Strain can be used for preparation making from antibiotic group which can be used together with antibiotics. Preparation can be used for treatment of different infections, food allergy, disbacteriosis, enzyme insufficiency. EFFECT: preparing strain indicated above. 3 tbl

Description

 The invention relates to the field of biotechnology, in particular the production of therapeutic and prophylactic drugs used for disorders of the gastrointestinal tract, dysbiosis, digestive enzyme deficiency, purulent infections, food allergies.

 A known strain of Bacillus subtilis 534, used to obtain the drug "Sporobacterin", which is intended for the prevention and treatment of diseases of the gastrointestinal tract, dysbacteriosis.

 However, the known strain is sensitive to antibiotics with the exception of polymyxin. This property of the strain limits the scope of the drug for severe forms of purulent-inflammatory diseases, the treatment of which requires the joint administration of a bacterial drug and an antibiotic.

 The aim of the invention is the creation of a new antibiotic-resistant strain to obtain a drug of the above purpose.

 The technical result obtained from the use of the new strain is expressed in increasing the effectiveness of treatment due to the possibility of combined use of a bacterial preparation with antibiotics. The strain Bacillus subtilis 3H deposited in the collection of GISK them. L.A. Tarasevich under N 248. The strain Bacillus subtilis 3H was obtained as a result of repeated reseeding of the strain Bacillus subtilis 534 through nutrient agar with an increasing concentration of the antibiotic.

 The strain of Bacillus subtilis 3H is characterized by the following properties.

 Cultural and morphological properties.

 Sticks. The size of the cells of a one-day agar culture (1.5-4.0) x (0.4-0.6) microns, often located in chains. Cells stain positively according to Gram, capsules do not form, form internal central spores.

 Colonies on meat-peptone agar are more often rough with scalloped edges from white to gray and pale pink, less often smooth with smooth edges.

On meat-peptone broth at 37 ^o C after 18-24 hours form a film on the surface and bottom sediment.

 Physiological signs.

Aerob, optimal growth conditions at a temperature of 37 ^o C and a pH of 6.0-7.5.

 Biochemical properties.

 Breaks down glucose, sucrose, mannitol, maltose. Gives a positive reaction of Voges-Proskauer to acetoin. Hydrolyzes starch, casein, urea, peptones milk, liquefies gelatin. Does not break down tyrosine. Produces catalase, lecithinase. Does not form hemolysin and fibrinolysin. Restores nitrates to nitrites. Disposes of citrate. Forms ammonia and hydrogen sulfide, does not form indole.

 Antagonistic properties.

 It inhibits the growth of staphylococci, protea, Klebsiella, Escherichia coli, yeast fungi, streptococci and Pseudomonas aeruginosa.

 The strain is resistant to polymyxin, rifampicin, insensitive to penicillin, lincomycin, streptomycin.

 Comparative data on the antibiotic sensitivity and antagonistic activity of strains of Bacillus subtilis 3H and known are presented in table. 1 and 2 (respectively).

The data table. 1 show that the new strain has high antibiotic resistance
As can be seen from the table. 2, the antagonistic activity of the strain of Bacillus subtilis 3H becomes higher compared to the original strain of Bacillus subtilis 534.

 The use of the strain is illustrated by the following examples.

 Example 1. To grow a strain of Bacillus subtilis 3H, a synthetic medium containing peptone, maltose, mineral salts and trace elements is used.

The strain Bacillus subtilis 3H is sown on mattresses with an agarized synthetic nutrient medium containing peptone, maltose, mineral salts, trace elements and placed in a thermostat at 37 ^° C for 18-20 hours. Next, the agar-washed culture is seeded in a fermenter to grow on the same medium composition in a volume of 5 l and grow 3-4 hours until the culture reaches the phase of exponential growth. This culture is used as seed for inoculation of the Bior reactor with a capacity of 100 liters. Cultivation is carried out for 16-20 hours before the transition of the culture into a spore form. The resulting bacterial mass is concentrated to (100-120) • 10 ⁹ cells / ml, washed from the nutrient medium with a sterile 0.9% sodium chloride solution. Poured into ampoules of (1.0 ± 0.1) ml and dried. The resulting preparation is called "Baktisporin", is a mixture of living vegetative cells and spores.

The drug "Baktisporin" is harmless, does not contain extraneous microflora. Residual humidity does not exceed 3.5%. It has specific activity: the number of cells in one dose of the drug (1-5) • 10 ⁹ cells / ml, antagonistic activity (growth inhibition zone of test strains of microorganisms) is 25-40 mm.

 Thus, the strain we obtained can be used as a production strain to obtain a drug from the group of probiotics, which can be used in conjunction with antibiotics.

Claims (1)

 The bacterial strain Bacillus subtilis GISK N 248, bearing the antibiotic resistance property, used to obtain the drug "Baktisporin".

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