RU2445363C2 - Method of producing culture medium for investigating microbial contamination of air - Google Patents

Abstract

FIELD: chemistry.

SUBSTANCE: dry enzymatic peptone, glucose, dialysate of baker's yeast and microbiological agar are mixed in given quantities. The obtained mixture is added to 1 litre of distilled water and boiled until complete dissolution of the agar. While hot, the mixture is poured into vials, sterilised in an autoclave and cooled to 47-55°C. The cooled medium is poured into Petri dishes and held until complete setting of the gel. Weighed zinc nanopowder in a physiological solution in amount of 0.005 mcg of zinc nanopowder per 0.1 ml of the physiological solution per Petri dish is deposited on the solid surface of the gel in aseptic conditions.

EFFECT: invention increases accuracy of results of investigating microbial contamination of air.

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Description

The invention relates to the field of microbiology, to sanitary microbiology.

A known method of obtaining a nutrient medium used for an approximate assessment of the degree of microbial air pollution, called meat peptone agar, which consists of meat water, enzyme peptone and sodium chloride (Polyak M.S., Sukharevich V.I., Sukharevich M.E. Nutrient media for medical microbiology, St. Petersburg, 2002, p.28).

The disadvantage of obtaining a nutrient medium is that unpretentious microorganisms grow on its surface and in the presence of mold its continuous growth is observed, so the bulk of the microorganisms present in the air under investigation do not grow, which leads to inaccurate research results.

There is a method of obtaining a nutrient medium for studying microbial air pollution by pathogenic microorganisms - blood agar, consisting of meat peptone agar and 10-25% of fresh or defibrinated animal blood (Rozanov NI Microbiological diagnosis of diseases of farm animals. Moscow, 1952, p. 85) .

The disadvantage of obtaining a nutrient medium is its high cost and the possible presence of microorganisms in the blood of an animal donor, which, along with existing microorganisms in the test air, can germinate on the medium, which leads to unreliable research results.

A known method of obtaining a nutrient medium is Saburo agar, used to control microbial (fungal) contamination of environmental objects, including to assess the degree of microbial air pollution, containing dry enzymatic peptone for bacteriological purposes, glucose, baker's yeast dialysate, microbiological agar and distilled water (Prototype - Appendix No. 2 to the order of the Ministry of Health of the Russian Federation of July 14, 2000 No. 263 “On permission of the medical use of culture media”). Release form - in the form of a gel placed in bottles, which are heated in a water bath until the gel of the medium is completely melted, the medium is cooled to a temperature of (47-2) degrees. C and poured into sterile Petri dishes. After the medium has solidified, the dishes are dried at a temperature of (33-2) degrees. C for (40 + 5) min. In addition, this medium is produced in the form of a dry nutrient medium, which contains the following ratios of ingredients at the rate of 1 liter of distilled water: dry enzymatic peptone for bacteriological purposes in the amount of 10 g / l, glucose in the amount of 40 g / l, baker's yeast dialysate - 5 g / l, microbiological agar - 10 g / l (medium producer Saburo - JSC BIOMED named after II Mechnikov).

The disadvantage of obtaining the Saburo medium is that it provides the growth of only microscopic fungi (mold, yeast-like, etc.) found in the air. With the growth of fungal colonies, they fill the entire surface of the nutrient medium and prevent the colonies from growing other types of microorganisms that got into this medium from the studied air, which leads to inaccurate results of microbiological studies and, as a result, to an incomplete picture of the sanitary assessment of air.

An object of the invention is to improve the accuracy of the results of studies of microbial air pollution with a view to its sanitary assessment.

The technical problem is achieved by a method of obtaining a nutrient medium for the study of microbial air pollution by mixing dry peptone for bacteriological purposes - 10 g / l, glucose - 40 g / l, dialysate of baker's yeast - 5 g / l, microbiological agar - 10 g / l, introducing the mixture into 1 liter of distilled water, boiling until the agar is dissolved, sterilizing and cooling the nutrient medium to a temperature of 47-55 ° C, keeping the gel completely solidified, followed by applying the gel to the surface sterile suspension of zinc nanopowder in physiological saline in an amount of 0.005 μg zinc nanopowder per 0.1 ml of physiological saline based on one Petri dish.

The difference between the proposed method of obtaining a nutrient medium for the study of microbial air pollution from the prototype is that a sterile suspension of zinc nanopowder in physiological solution in the amount of 0.005 μg zinc nanopowder per 0.1 ml of physiological solution per one petri dish.

A nutrient medium for microbiological evaluation of air is prepared as follows. Dry peptone for bacteriological purposes in an amount of 10 g / l, glucose in an amount of 40 g / l, dialysate of baker's yeast - 5 g / l, microbiological agar - 10 g / l are uniformly mixed. The mixed mixture is added to 1 liter of distilled water and boiled until the agar is completely melted. In hot form, the mixture is poured into bottles, sterilized in an autoclave, then cooled to a temperature optimal for transfusion into Petri dishes, in particular to a temperature of 47-55 ° C. Poured into sterile Petri dishes and incubated until gel solidification. As a rule, the gel hardens at a positive temperature. For example, at a temperature of 30-33 ° C, the gel hardens within 30 minutes. At temperatures below 30 ° C, but not lower than 0 ° C, the gel hardens faster. Under aseptic conditions, a suspension of zinc nanopowder in physiological saline is applied to the surface of the gel. To obtain this suspension, zinc nanopowder is preliminarily weighed in an amount of 0.005 μg (based on one Petri dish), sterilized, introduced into 0.1 ml of sterile physiological solution (based on one Petri dish). Saline in an amount of 0.1 ml is sufficient for its uniform distribution over the entire surface of one Petri dish. A suspension of zinc nanopowder in physiological saline is applied to the surface of the gel, evenly distributed over the surface, and the suspension is absorbed into the gel for an hour, after which the nutrient medium is ready for use. Ready nutrient medium can be stored for 3 days in a cold place.

Example. In the student canteen of the Saratov Agrarian University. N.I. Vavilova investigated microbial air pollution using a nutrient medium with the composition of the prototype and with the composition according to the invention. We counted the number of types of microorganisms and counted colony forming units (CFU) with the determination of the microbial number per 1 m ^{3 of} air according to the Omelyansky formula. Table 1 shows a comparative characteristic of the results of studies of microbial air pollution, carried out using a nutrient medium with a composition according to the prototype and according to the invention.

Table 1 Comparative characteristics of the results of studies of microbial air pollution after 72 hours of incubation. Indicators The composition of the medium according to the prototype The composition of the proposed environment Bacteria: - micrococci not detected discovered - staphylococci discovered discovered - streptococci not detected discovered - bacilli not detected discovered Mold mushrooms: - aspergils discovered discovered - penicils discovered discovered Yeast discovered discovered Actinomycetes not detected discovered Total Microbial Number (CFU) 263 ± 5 802 ± 10

As can be seen from table 1, when studying microbial air pollution using a nutrient medium with the composition of the prototype, 2 times less types of microorganisms were found, and, in addition, the total microbial number (CFU) was 3 times lower, using the composition of the proposed medium .

Table 2 shows a comparative characteristic of the concentration of zinc nanopowder with a particle size of 18-20 nm, suspended in 0.1 ml of physiological saline.

table 2 The number of species of microorganisms on a nutrient medium depending on the concentration of zinc nanopowder. Indicator The concentration of the solution of zinc nanopowder in 0.1 ml of physiological solution 0.001 0.002 0.003 0.004 0.005 0.006 0.007 The number of species of microorganisms four 5 6 6 8 8 8

As can be seen from table 2, when the concentration of zinc nanopowder in the amount of 0.005 μg found the largest number of species of microorganisms.

Claims (1)

A method of obtaining a nutrient medium for studying microbial air pollution by mixing dry peptone for bacteriological purposes - 10 g / l, glucose - 40 g / l, dialysate of baker's yeast - 5 g / l, microbiological agar - 10 g / l, making the mixture in 1 liter of distilled water, boiling until the agar is dissolved, sterilizing and cooling the nutrient medium to a temperature of 47-55 ° C, keeping the gel completely solidified, followed by applying a sterile suspension of zinc nanopowder to the surface of the gel that has set, in physiological saline in an amount of 0.005 μg zinc nanopowder per 0.1 ml of physiological saline based on one Petri dish.