SU1752772A1 - Method of pseudomonas aeruginosa isolation - Google Patents

Abstract

Use: medical microbiologists, laboratory diagnostics of infections caused by Pseudomonas aeruginosa. Essence: the studied material is sown on a dense nutrient medium containing a protein Base - a source of nitrogen (peptone, yeast hydrolyzate), sodium chloride in distilled water and selec-. The active agent is a 1% solution of phenyl (4-hydroxyphenyl) sulfide in DMSO, pH 7.2-7.4. The medium is sterilized before use at 120 ° C for 20 minutes. Crops are incubated at 37 ° C for 18-24 hours and take into account the results. The presence of colonies indicates the presence in the test material of the pathogen. The method allows the detection of both pigment-forming and non-pigmental strains of P. aeruginosa. The method is highly sensitive (isolation of an exciter from a mixture of cultures taken in a ratio of 1: 1000), 2 tab. with C

Description

This invention relates to medical microbiology and can be used to detect infections caused by Ps.aeruginosa.

A known method for isolating Ps.aeruginosa on a selective medium consisting of dry Dagestan peptone (20.6 g), K2S04 (10.0 g). MgCLa (1.5 g), araga 15.0 g, as a selective agent, 0.2-0.5% cetylperidine chloride (CLC), water - up to 1 l.

However, the medium used does not provide high selectivity with respect to Ps.aeruginosa, since it is associated with an increase in associates, and also does not have sufficient sensitivity.

A method for isolating Ps.aeruginosa by sowing on a nutrient medium containing sources of nitrogen, carbon, mineral salts, agar and a selective agent, in which p-hydroxyphenylsalicylamide is used in a certain concentration.

However, the used selective agent with increasing its concentration within the specified values inhibits the growth of Ps.aeruginosa, which reduces the sensitivity of the method. Growth Inhibition Index Ps.aeruginosa 2.6.

The aim of the invention is to increase the sensitivity of the method by reducing the inhibitory effect of the selective agent.

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Example 1. The sample, wt.%: Dry peptone SC 1,086; agar 1,130; Sodium chloride, 0.495, is dissolved in distilled water to 100 ml and heated until the agar is completely melted. The solution is adjusted to volume with water and 0.3 ml of a 1% solution of phenyl (4-hydroxyphenyl) sulphide is added as a selective agent (1% solution is prepared by dissolving 0.03 g selective agent in 2.8 ml of dimethyl sulfoxide), the medium is poured into vials and sterilized in an autoclave at 120 ° C for 20 minutes (pH 7.2-7.4), the prepared nutrient medium is melted in a water bath and poured into Petri dishes, Material from a patient with a diagnosis of chronic osteomyelitis seeded with a cotton swab on the surface of the nutrient medium, incubated settling at 37 ° C for 18-24 hours, after which the results are taken into account. The presence of grown colonies indicates that they belong to the species Ps.aeruginosa. Colonies are up to 2 mm in size, convex, gray, growth of other microorganisms is absent.

Example 2. The sample, wt.%: Dry peptone 1,146; agar 1.170; Sodium chloride 0.510 is dissolved in distilled water to 100 ml and heated until the agar is completely melted. Make up to volume with water and add 0.31 ml of 1% phenyl (4-hydroxyphenyl) sulfide solution, pour into vials and sterilize in an autoclave at 120 ° C for 20 minutes (pH 7.2-7, four). The prepared nutrient medium is melted in a water bath and poured into Petri dishes. A material from a patient with a diagnosis of pararectal fistula is sown with a cotton swab on the surface of the nutrient medium, the cultures are incubated for 18-24 h, after which the result is taken into account. Colonies have a size of up to 2.0 mm, convex, gray. The growth of other microorganisms is absent.

Example 3. Suspension, May%: dry peptone 1,160; agar 1,208; Sodium chloride 0.532 is dissolved in distilled water to 100 ml and heated until the agar is completely dissolved, brought to water to the original volume and 0.32 ml of 1% phenyl (4-hydroxyphenyl) sulfide solution is added. Poured into vials and sterilized in an autoclave at 120 ° C for 20 minutes (pH 7.2-7.4) Prepared nutrient medium

melted in a water bath and poured into Petri dishes. Material from a patient with a diagnosis of appendectomy and a cotton pad is planted on the surface of the nutrient medium,

incubated for 18-24 hours seeding, after which the result is taken into account. Colonies are up to 2.0 mm in size, gray, convex. The growth of other microorganisms is absent,

Results of sensitivity studies

the proposed environment are given in table. one.

The sensitivity of the medium was studied in five Ps.aeruginosa strains isolated from clinical material. One hundred microbial cells are seeded in an amount of 1 ml per 2 cups of dilution, incubated at 37 ° C for 18-24 hours and the number of grown colonies is counted. The results show that the proposed selective agent has a lower inhibitory effect on the growth of Ps.aeruginosa (1.5 times less compared with the known).

The tribute given in table. 2 indicate that when using phenyl (4-hydroxyphenyl) sulfide and oxzphenamide, it is possible to isolate the main pathogen from a mixture of cultures taken in a 1: 1000 ratio,

Instead of peptone in the composition of the medium, you can use hydrolyzed fodder yeast in a similar concentration with obtaining a similar positive result.

The results of the studied selective properties of the proposed environment are given in table. 2

Thus, the use of the proposed method allows to increase the selectivity of the release while reducing the inhibitory effect on the growth of Ps.aeruginosa.

Claims (1)

Invention Formula The method of isolating Pseudomonas aeruginosa by sowing the test material on a nutrient medium containing a source of nitrogen, agar, sodium chloride, selective agent and distilled water, followed by incubation of the crops and examining the grown colonies, characterized in that, in order to increase the sensitivity of the method by reducing the inhibitory effect of the selective agent, as a selective agent, use 1% G1 solution of phenyl (4-hydroxyphenyl) sulfide in dimethyl sulfoxide. Table