SU1514783A1 - Method of extracting campilobacteria - Google Patents

Description

The invention relates to medical Microbiology and can be used in the practice of laboratory diagnosis of acute intestinal diseases. The purpose of the invention is to improve the accuracy of the method. Is2

the following material is sown on a dried nutrient medium containing, g / l: hydrolyzed chicken hearts and stomachs 28 30, lysed blood (based on total nitrogen) 3.25-5.95; sodium phosphate acid disubstituted 2.8-3.0; rifampicin 0.008 0.01; amphotericin B 0.004 0.005;

agar-agar 10 12 and hepatic brain infusion the rest. pH of 7.4 - 7.6. Crops are incubated under anaerobic conditions at 28 44 ^e C. Using the medium allows you to isolate Campylobacter without prior enrichment, and a wide range of temperatures to detect various types of Campylobacter. The accuracy of the method increases on average by 10%.

The invention relates to medical Microbiology and can be used in the practice of laboratory diagnosis of acute intestinal diseases.

The purpose of the invention is to improve the accuracy of the method.

The method is as follows.

The test material is sown directly on a dense selective medium, bypassing the accumulation stage. The nutrient base of the medium is hydrolyzed from chicken hearts and stomachs. The medium is prepared as follows: 28-30 g of the hydrolyzate from chicken hearts and stomachs are dissolved in 950 1000 ml of brain-brain infusion, 10 12 g of agar are added, boiled until melted, 2.8-3 g of sodium phosphate dibasic is added, the pH is adjusted to 7 , 4 - 7.6. autoclaved at 1 atm 30 min. then add 0.008 - 0.01 g of rifampicin, 0.004- 0.005 g of amphotericin B, 3.25 5.95 g of lysed blood (based on total protein), poured into 20 ml Petri dishes. Then the frozen medium is dried for 1 day at -) - 37 ⁰ C in anaerostat. Sowing of the pathological material is carried out on several cups and incubated for 24-48 hours at 28 ° -44 ° C in anaerostats. in a gas atmosphere containing,%: Ν ₂ 75, CO ₂ 17, O ₂ 8.

Example 1. 0.1 ml of native pathological material (feces, biopsy specimens, blood) are rubbed with a spatula over the surface of a dried medium containing 28 g of chicken hearts and stomachs hydrolyzate, 10 g of agar, 2.8 g of phosphoric disubstituted sodium, 3, 25 g (by total} protein) of lysed blood, 0.008 g of rifamp, 0.004 g of amphotericin B. pH of the medium is 7.4. Hepatic brain infusion add up to 1 l.

Sowing is done on several cups.

simultaneously. Due to the fact that the sensitivity spectrum of different species of Campylobacter to temperature is different, the crops are cultivated ;,, 1514783

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four

tivuyu simultaneously for 24-48 hours

at 28, 35, 37 and 44 ^С С in anaerostats in

atmosphere containing,%: N5 75, CO "17,

eight.

After 48 hours, cultured, morphological, and biochemical properties are studied in grown bacteria. The purity of crops is checked by microscopy, stained with any dye. Learn mobility on semi-fluid agar without antibiotics. Identify by b test: morphology, mobility, positive samples for catalase, oxidase, negative test for aerotolerance, temperature test. Differentiation within the species is carried out according to 9 tests. 15

After 48 h of growth, 320–360 colonies grow with typical properties of campylobacter 3 types of Saprulobacter 1 1 of 1 ί I lI> 1 ί ί Iί5, C. 1ap <1g and S. bibi1 and 5 - small, racing, curved g ,, - sticks in the form of θ "gull wings" or 8-shaped, mobile; they grow in the form of a disk above the surface of the medium, hydrolyze hippurate, grow by 1% glycene, 1% bile, 0.25% cysteine, and do not grow at 8% glucose. Not all strains grow at 3.5% 25

! \ 1C1 (except C. 1E1 of 1P1P. 1W.S.Libic) are resistant to nalidoxic acid and sensitive to cephalothin. grow at 44 ° C.

Pp. 1e1i5 1p1ezchpnz grow on medium with 3.5%; ι (Ζ1, sensitive to cifalotine, grow only at + 37 ° С \ аС1, sensitive to 30 cifalotine, grow only at + 37 ° С. All strains possess catalase, oxidase, do not grow in the presence of oxygen above 10%.

The proposed method on a freshly prepared medium from 225 samples allocated 19 cultures of Campylobacter, which is 8% 35

seeding rate.

Example 2. 0.1 ml of the native material is inoculated on a medium containing 30 g of chicken hearts and stomachs hydrolyzate, 12 g of agar, 3 g of sodium phosphoric acid bifurcated, 5.95 g of lysed blood,

0.01 g, rifampicin, 0.005 g amphotericin B per 1 liter of brain-brain infusion, pH 7.7. On Wednesday grows 470—

570 colonies, bacterial growth after 24 hours

Of the 225 samples, 23 cultures of the exciter were isolated for 2–3 days, 5 species (C. ^ e ^ un!

C. coH, S. articulum, S. 1elis 1n1e5IpN5, S. liberia) Seeding out is 11%.

Example 3. The method is carried out in accordance with example 1, but using a nutrient medium of the following composition, g / l: hydrolyzed chicken hearts and stomachs 29, agar-agar 11, sodium phosphoric disubstituted 2.9, lysed blood 4.5, rifampicin 0.009 , amphotericin B 0.0045, hepatic brain infusion else, pH 7.5.

The use of the method allows to allocate a wider range of Campylobacter directly on the same medium with good growth characteristics of the environment (seeded by the prototype 1.6%).

Claims (2)

Claim The method of isolation of campylobacterium by planting the studied material on a nutrient medium containing a nutrient base, lysed blood, mineral salt, rifampicin, amphotericin B and agar-agar, incubation under anaerobic conditions, followed by selection of grown colonies, characterized in that, in order to improve the accuracy of the method , sowing is carried out on the dried surface of the nutrient medium, containing as a nutrient base hydrolyzed chicken hearts and stomachs and hepato-cerebral infusion, and as mineral saline s phosphoric acid disubstituted with the following ingredients quantitative ratio, g / l: 1 Chicken Hearts and Stomach Hydrolysis Lysed blood (based on total protein) Sodium phosphorus-acidic disubstituted Rifampicin Amphotericin B Agar-Agar Hepatic Infusion 28 30 3.25 -5.95 2.8--3.0 0.008 - 0.01 0,004—0,005 10-12 Up to 1 l. pH of the medium is 7.4-7.6, and incubation is simultaneously at 28–44 ° C. lead