SU1541251A1 - Method of testing nutrient media used for vibrinocynotyping cholera germs - Google Patents

Abstract

This invention relates to medical microbiology and can be used in assessing the quality of nutrient media used for vibriocinotyping of cholera vibrios. The purpose of the invention is to improve the accuracy of the method. To test nutrient media, three test strains of VIBRIO CHOLERAL ELTOR NN KM-13, KM-14 and KM-15 are used. Test strains are plated on the test medium, the cultures are incubated and then the indicator cultures of enterobacteria are seeded. Crops are incubated and the spectrum of inhibitory activity of all test strains in relation to indicator cultures is determined. By the nature of the spectrum, the suitability of the test medium for vibriocinotyping of cholera vibrios is determined. 4 tab.

Description

The invention relates to medical microbiology and can be used in assessing the quality of culture media used for vibriocinotyping of cholera vibrios.

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

For carrying out the method, culture cultures of bacteria Vibrio cholerae eltor P-10588, P-9952 and P-10117, which are deposited as CM-13, CM-14 and CM-15, and respectively, are used as test strains. characterized by the following features. one.

Cultural, morphological, physiological and biochemical characteristics. Gram-negative polymorphic curved sticks. Relate to group i

Heiberg

Dissolve glucose in Hugh-1 Lifeson's medium to form an acid without 1 gas under aerobic and anaerobic conditions.

x Form acetylmethylcarbinol.

Oxidase positive, possess lysine and ornithine decarboxylase. Do not have arginine dihydrolase. Form indole. Do not form hydrogen sulfide.

Strains KM-13 and KM-14 are agglutinated to titer with 0m and Inaba sera and are not agglutinated with Ogawa serum. Strain K-15 is agglutinated to titer with O and Ogawa sera and is not agglutinated with Inaba serum. All strains are sensitive to

HSKFi.l

Sssshi

ascafcs

phages Eltor and not sensitive to phages

The KM-14 and KM-15 strains grow well on polymnixin agar (50 units / ml), the KM-13 strain on this medium forms single colonies and has a broad spectrum of vibriocynogenic activity, the KM-14 strain has a moderate spectrum, the KM strain -15 - narrow spectrum.

The method is carried out as follows.

Daily broth cultures of each KM-13, KM-14 and KM-15 test strains are seeded in the middle of three dried plates with 1.5% nutrient agar with citrate-phosphate buffer. Crops are incubated for 48-72 hours. “After the expiration of the incubation period, the cultures of the test strains are subjected to cold shock, then treated with chloroform. To the bands of grown test strains, broth cultures of indicator strains of enterobacteria are sown: 1 - Shigella flexne-ri 3189; 2 - Shigella flexneri 38/40; 3 - Shigella dysenteriae 43; 4 - Shigella sonnei M 2/2 (22); 5 - Shigella sonnei M-56 (31); 6 - Shigella sonnei 17 (2); 7 - Escherichia coli C (l); 8 - Escherichia coli G (l); 9 - Escherichia coli K-12; 10 - Vibrio cholerae 541 (54); 11 -Vibrio cholerae 296 / .1 (20); 12 -Vibrio cholerae 24 Shein 1077.

Crops are incubated for 18 hours. Sensitivity of indicator cultures to vibriocins is recorded by the absence of their growth on the band or outside of the growth of the test strains. The sensitivity spectrum is set according to the table. one.

Test agar is considered suitable for vibriocinotyping when reproducing the spectrum of the inhibitory activity of test strains with respect to indicator cultures in accordance with Table. one

Example 1. Control of 1.5% Martin-agar, pH 7.6, series 15 for suitability for vibriocin-typing. According to the test results, the test strain P-1 (prototype) agar of this series was recognized as suitable for the diagnosis of cholera.

First day „Daily agar cultures of test strains KM-13 (P-10588).

0

five

0

five

0

five

0

KM-14 (P-9952) ICM-15 (P-10117) are seeded in Martin's broth, pH 7.6.

Second day. Ingredients of citrate-phosphate buffer kz per 100 ml of 2.0 ml of a 25% aqueous solution of dibasic potassium phosphate and sodium citrate and 0.5 ml of 0.6% are added to the agar melted and cooled to 45 ° C. solution of ammonium chloride.

One drop of daily broth cultures of each of the test strains is applied to the edge of a thoroughly dried plate taken for the study of agar and allowed to flow in the middle of the plate in the form of a strip. After the seed has dried, the cups are placed in a thermostat at 37 ° C for 48 hours.

Fourth day Petri dishes with cultures of test strains grown on them are transferred to a refrigerator at +4 C.

On the fifth day, the growth of test strains on the sowing strip is cleaned with a glass slide, treated with chloroform vapors for 1 hour, and then aired for 1-2 hours. Daily indicator broth cultures (4 cultures per plate) are seeded perpendicularly to the producer growth band also as lanes.

The sixth day Records of the results are given in table. 20

Example 2, Control of agar VNIIVS, series 8. Work on the first-sixth days is carried out analogously to example 1, Accounting results are given

in tab. 3

I

Example 3. Control of yeast agar, series 3. Work on the first days is carried out analogously to Example 1. The results are shown in Table 4.

Using the proposed method makes it possible to increase the accuracy of the selection of media suitable for vibriocinotyping of cholera vibrioes, which is impossible when using a known method.

rmula of invention

The method of controlling nutrient media used for vibriozinotyping of Vibrio cholerae, including the seeding of test strains on the nutrient medium and the incubation of crops, characterized in

that, in order to improve the accuracy of the method, as test strains, cultures of bacteria Vibrio cholerae eltor, strains KM-13, KM-14 and KM-15 are used, the medium is seeded in parallel with the test strain, after incubation strains sow in1251

6

The culture of enterobacteria and Vibrio cholerae cultures are incubated and the suitability of the medium for vibriocinotyping is determined from the spectrum of inhibitory activity of all test strains in relation to indicator cultures.

Table 1

Vibrio cholerae eltor KM-13 (P-10588) Vibrio cholerae eltor KM-14 (P-9952) Vibrio cholerae eltor KM-15 (P-10117)

Wide

Moderate

Narrow

+ ++++++++ t

+ - + + +

+ - +

+ +

+ f

e

+ + + + + + + + +

+ + +

j1sh

+ +

+ f

table 2

Fit

Table 3

Claims (1)

Claim A method for controlling the nutrient media used for vibriocinotyping of cholera vibrios, including sowing test strains on the studied nutrient medium and incubating the crops, characterized in that, in order to increase the accuracy of the method, bacterial cultures of Vibrio cholerae eltor, KM-strains are used as test strains 13, KM-14 and KM-15, the medium is sown in parallel each test strain, after incubation, test cultures are seeded with indicator cultures of enterobacteria and cholera vibrios, cultures are incubated and the suitability of the medium for vibriocinotyping is determined by the spectrum of inhibitory activity of all test strains in the ratios of indicator cultures. Table 1 Test strain The spectrum of inhibitory Availability zones 3 growth inhibition indicator of cultures 1 2 4 5 6 7 8 ----- 9 ------ 10 eleven 12 nosti Vibrio cho- Shirolerae eltor cue KM-13 (P-10588) + ¢ + + + + + + + + + + Vibrio cho- Able lerae eltor ren KM-14 ny (P-9952) + - + + + - + + - Vibrio cho- Narrow lerae eltor KM-15 e (P-10117) + - + - - - - - - + + - table 2 Test strain from. The presence of zones of growth inhibition of indicator cultures 1 2 3 4 5 - 6 7 8 9 10 eleven 12 6 Conclusion on the suitability of agar for vibriocinotyping Vibrio chole- gae eltor KM-13 (P-10588) + + + + + + Vibrio. cholerae eltor KM-14 (P-9952) + + + + - Vibrio cholerae eltor KM-15 (P-10117) + + + + + + + Good for - - - ♦ + - + -. ^and - 1541251 ’ Table 3 Test strain Availability zones of inhibition growth indicator crops Concluded - statement about- ' 1 2 3 4 5 6 7 8 9 10 eleven 12 suitability for vibrio-typing ί I vibrio 'cholerae eltor KM-13 (P-10588) V. cholerae eltor KM-14; (P-9952) V. cholerae eltor KM-1 5 (P-10117) + + + - - + - +. - f + - - + - - + + ** · * · - 4. 4He is good It Ji Table 4 Test strains The presence of zones of growth inhibition of indicator cultures - ---_ 2 3 4 5 6 7 8 9 10 eleven 12 Conclusion on the suitability of agar for vibrio ¹ - ·. cytotyping V. cholerae eltor KM-13 (P-10588) + V. cholerae eltor KM-14 (P-9952) + V. cholerae eltor KM-15, (P-10117) + Unfit II II