SU1602865A1 - Strain of vibrio cholerae eltor bacteria used as test culture for identification of hemolysine - Google Patents

Abstract

This invention relates to medical microbiology and can be used in solving the taxonomy of Vibrio cholerae. The purpose of the invention is to obtain a Eltor cholera vibrio strain producing hemolysin type II subtype β. The strain obtained by selection of cholera vibrio Eltor on nutrient media containing red blood cells of various animal species. The strain deposited under the number CM N81. The strain of Vibrio cholerae Eltor KM N81 does not lyse erythrocytes of sheep and humans, however, has a pronounced lytic activity in relation to guinea pig erythrocytes. In order to detect hemolysin, strain KM N81 is grown on m-septon broth for 18-20 hours at 37 ° and the Greig test is made, which is read out after 30 minutes. In the presence of type II hemolysin in the test material, subtype β occurs, the guinea pig erythrocyte lysis occurs.

Description

This invention relates to medical microbiology and can be used in solving the taxonomy of Vibrio cholerae.

The goal from gaining nirami of Vibrio cholerae Eltor, the product of hemolysin type II subtype B.

strain obtained by passaging. a strain of Vibrio cholerae Eltor 149 on nutrient media containing 1.5% of sheep erythrocytes. On these media, a clone with the desired properties was selected by the selection method — not lysing. sheep erythrocytes producing hemolysin active against guinea pig erythrocytes. The strain of Vibrio Cholerae el tor 149 Hly - type II subtype | 3 depon fowan number KM

No. 81 and is characterized by the following features.

Cultural and morphological features In smears. - vibrios, in the field of phase-KOHTpacTfioro microscope typical for vibrio mobility.

Prototroph

On Zh1SCHK1GH and dense nutrient media, typical growth of cholera vibrio is formed: on 1% peptone water, diffuse cloudiness,

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on the surface there is a gentle blue film, on alkaline agar - large, round with smooth edges, flat, transparent in transmitted light colonies Biochika properties. The strain decomposes glucose under aerobic and anaerobic conditions, mannoz, sucrose, mannitol, starch - by the formation of acid without gas. Produces acetylmethylcarbinol, indole. Lekarboksiliry lysine, ornithine Does not break down arabinose, does not dehydrolyze arginine. Does not form hydrogen sulfide.

Possesses proteolytic activity

Serolopgichesky properties.

The strain is agglutinated about cholera serum to titer, serum Ogawa

about titer, with Inaba serum does not give half-glasses results. Phage sensitivity.

Is lysed with the Eltor phage to-titer, with phage 3,4,5 to DRT, refers

to the 10th phage type according to the 7th Ozhevkina-Arutyunov, not lietfuyus phage C.

Sensitivity to antibiotics.

MZK tetracycline 0,015 µg / ml. It grows on polymyxin - 30 units / ml. Hemagglutination reaction is positive The reaction Voges-Proskauer is positive. Virulence for suckling rabbits. Causes a choleragenic reaction in a dose of 10 m.

The method, storage conditions and composition of the storage medium for the Martin agar strain, pH 7.2, inoculation, are kept under liquid paraffin at room temperature. .

Example 1. Whether hemolysin production was detected, strain KM No. 81 (149) was grown on m septon broth, pH 7.2, for 18 hours, which was later used to issue a Graigur test, titrate in wells of polystyrene plates or sediment. t spots on agar plates containing 1.5% marine erythrocytes

pigs. In the Greig test, a 1% suspension of guinea pig erythrocytes is used, and in the wells of the plates one drop of 2.5% of the same erythrocytes is added in one drop. These objects for testing hemolysin are placed in a thermostat at

37 for 30 minutes and read the result on the hemolysis of eri-groocytes.

To determine the activity of hemolysin obtained by the cultzf centrifuge

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the presence of hemolysin and its titer are determined with a 1% suspension of guinea pig erythrocytes in physiological saline in the supernatant liquid. Hemolysin causes lysis of guinea pig erythrocytes at 37 ° C for 30 minutes. Hemolysin titer 1: 800.

The resulting HEM: hemolysin produced by the strain is filtered through ceramic candles, lyses the guinea pig erythrocytes for 30 min at 37 ° C as in. liquid (1%: e suspension of erythrocytes), and on dense (1.5% - suspension of erythrocyte suspension) nutrient-media. Does not destroy human erythrocytes and sheep.

Example 2. The studied strain of cholera vibrio Eltor is seeded on solid nutrient media containing 1.5% of sheep erythrocytes, and: after incubation for 18-20 hours at 37 ° C, a colony is selected that does not form lysis zones. At the same time, the test culture was sown — shlmm 149. Next, a Greig test was performed with both cultures with a 1% suspension of guinea pig erythrocytes. The presence of lysis of guinea pig erythrocytes and the absence of lysis with sheep erythrocytes indicates the presence of a hemolysin type II subtype | 1 in the strain. In the study of strains of classical biovar, the stage of seeding on dense media with sheep erythrocytes is excluded, since the cholera vibrios of the classical biovar produce hemolysin type II of the p subtype, with these cultures directly put the Gregu test with guinea pig erythrocytes. As control use strain 149.

The use of the strain permits the identification of Vibrio cholerae by a hemolysin formation test of the appropriate type. In addition, the strain can be used as a carrier of a stable marker when studying the structure of the cholera vibrio gene.

Claims (1)

Claims. The strain of the bacterium Vibrio Cholerae eltor KM P 81 used as a test culture for the identification of hemolysin type II subtype fi cholera vibrioes.