Nutrient media for cultivation of v. cholerae

 

(57) Abstract:

The invention relates to Microbiology, and in particular relates to a method of obtaining nutrient media, providing optimal conditions for vital activity of V. cholerae can be used in Microbiology and biotechnology. Nutrient media for cultivation of V. cholerae contains sodium chloride, as a nutrition - sugar-molasses, water and sodium sulfite as a stimulator of growth of V. cholerae. The invention allows to simplify the method of preparation of the nutrient medium and reduce its cost.

The invention relates to Microbiology, in particular the production of nutrient media, which create optimal conditions for isolation and cultivation of V. cholerae.

Bacteriological examination for cholera using various nutrient medium: liquid medium enrichment, alkaline agar, elective differential diagnostic environment and a set of environments to identify (Labinsky A. S. Workshop on microbiological methods. - M.: Medgiz, 1963, S. 443). The basis of the known nutrient media are animal products - peptone, to the proposed nutrient medium is a liquid medium enrichment, containing 100 g of peptone, 50 g of sodium chloride, 1 g of potassium nitrate, 20 g of sodium bicarbonate in 1 l of distilled water (Birger M. O. Handbook of microbiological and virological methods. - M., 1982, S. 65).

The disadvantage of this environment is its high cost and difficulty in making.

The purpose of the present invention is the selection of the basic nutritional environments of plant origin, which would provide a cumulative effect and reducing its cost, and simplify the cooking method.

This objective is achieved in that the nutritional basis of the proposed nutrient medium syrup is sugar, sodium chloride in tap water with the addition of liquid culture medium stimulating supplements, and as a stimulating additives used sodium sulfite in the following ratio of ingredients, g/l:

Syrup refinery 20,0

Sodium chloride 5,0

Sodium sulfite 0,4

Tap water is the Rest

Syrup refinery (OST 18-233-75) in its composition contains at least 45% sucrose. Unlike the prototype of the proposed environment provides cumulative properties, optimal conditions and desealing basics is a distinctive feature of the proposed nutrient medium. The method is illustrated by the following example.

A nutrient medium is prepared as follows: molasses sugar dissolved in tap water, add sodium chloride, heated to boiling, bring the pH to 7.7 to 8.0 with 20% sodium hydroxide. The precipitation is filtered through a fabric filter with a filter paper, the solution add the required amount of sodium sulfite. The finished medium is poured into a graduated 100 ml flasks and sterilized in an autoclave at 0:5 ATM 30 minutes According to the guidelines for determining the quality of nutrient media for the cultivation of V. cholerae (instruction on infection control diagnostic culture media for V. cholerae), preparing a culture avirulent strain NAKED-cholerae R-9741. What culture stored on semisolid agar, seeded 17% peptone water or broth Martin (pH of 7.6 to 7.8) and incubated at 37°C for 3-5 h, and then plated on agar plate was assessed Martin or Hottinger (pH of 7.6 to 7.8). After 18-20 h of growth from the agar select smooth colony NAKED-cholerae R-9741 subcultured them on agar plates (2 passage). The culture is grown for 3-6 h at 37°C, and then used for testing. From the daily culture was prepared by suspension MT, is equal to enemy in physiological solution of 4.5 ml conveyed to the content in 1 ml of 100 and 10 m K. From these breeding suspension cultures were sown in 0.1 ml sterile bacteriological pipette 3 bottles of test and control environment. As a last used a pre-tested, high-quality 1% peptone water prepared in distilled water. Crops incubated at 37°C for 6 h, then from each vial surface sow loop # 5 Chaplasha on a Petri dish with a pre-tested, high-quality dense environment. The cultivation is carried out at 37°C. liquid Nutrient medium is suitable if at sowing 100 m as the test strain in 100 ml after 6 h of cultivation and subsequent plating on agar plates grow accordingly not less than 10 and 1 colonies. Evaluation of the growth properties of the nutrient medium is carried out by counting the number of grown colonies of V. cholerae in the crop contents of the bottles on the plate with a dense nutrient medium.

Example 1. The test strain was grown on a nutrient medium containing, g/l: molasses refinery 18,0; sodium chloride 3,0; sodium sulfite 0.3; the tap water - the rest. With this ratio of ingredients, the number of colonies grown on solid agar plates, for V. cholerae inoculated from flna nutrient medium containing, g/l: molasses refinery 20,0; sodium chloride 5,0; sodium sulfite 0,4; tap water - the rest. With this ratio of ingredients, the number of colonies grown on solid agar plates, for V. cholerae inoculated vials when sowing the dose of 100 and 10 M. K. were, respectively, 30 and 5.

Example 3. The test strain was grown on a nutrient medium containing, g/l: molasses refinery 22,0; sodium chloride 6,0; sodium sulfite 0.5; the tap water - the rest. With this ratio of ingredients, the number of colonies grown on solid agar plates, for V. cholerae inoculated vials when sowing the dose of 100 and 10 M. K. were, respectively, 13 and 2.

1% peptone water (control) number of grown colonies was 10 and 1 respectively.

The obtained results allowed to determine the optimal variant of the nutrient medium on the basis of refinery molasses (example 2).

Thus, on the basis of the above can be said about the obvious advantages of the proposed nutrient medium, consisting of molasses sugar-20,0 g/l; sodium chloride 5.0 g/l; sodium sulfite 0.4 g/l; water - the rest.

Nutrient medium is technically simple in prigotovlen in based on vegetable raw materials (molasses sugar with sodium sulfite) and for its preparation does not require distilled water the environment is prepared in tap water.

Nutrient media for cultivation of V. cholerae containing a nutrient basis, sodium chloride and water, characterized in that it additionally contains sodium sulfite as a growth promoter, as nutrition - sugar-molasses, water - tap water with the following ingredients, g/l:

Syrup refinery 20,0

Sodium chloride 5,0

Sodium sulfite 0,4

Tap water is the Rest



 

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FIELD: biotechnology, microbiology, medicine.

SUBSTANCE: invention relates to the strain Lactobacillus paracasei CNCM I-2116 used for diarrhea prophylaxis causing by pathogenic microorganisms. Supernatant of this strain culture elicits ability to prevent colonization of intestine with pathogenic microorganisms causing diarrhea also and this strain is designated for preparing agent used for prophylaxis and/or treatment of disorders associated with diarrhea. Agent for oral administration represents therapeutically effective dose of the strain L. paracasei CNCM I-2116 or supernatant of its culture and acceptable foodstuff. Invention provides the enhanced viability of the strain in its applying and effectiveness in prophylaxis of adhesion to intestine cells and invasion to intestine cells of pathogenic microorganisms causing diarrhea.

EFFECT: valuable medicinal properties of strain.

5 cl, 8 dwg, 10 ex

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