Wednesday Kessler-GRM Obolensk

INSTRUCTIONSon the use of a set of reagents for bacteriological studies"Nutritional medium for the detection of bacteria of the Escherichia coli group dry" Kessler-GRM medium Obolensk

1. PURPOSE

The Kessler-GRM Obolensk environment is designed to detect bacteria of the Escherichia coli group during the sanitary examination of environmental objects.

1. set characteristic

Medium Kessler-GRM Obolensk is a fine, hygroscopic, light-sensitive grayish-yellow powder.

Produced in polyethylene cans of 250 g.

2.1. OPERATING PRINCIPLE

The set of components included in the kit provides the nutritional requirements for the growth of bacteria of the Escherichia coli group and the inhibition of certain types of microorganisms.

2.2. COMPOSITION OF THE SET

The Kessler-GRM Obolensk medium is a mixture of dry components at the rate, g/l:

1. ANALYTICAL AND DIAGNOSTIC CHARACTERISTICS

Kessler-GRM Obolensk medium should provide visually detectable growth of Escherichia coli 675, Klebsiella pneumoniae 418, Citrobacter freundii 101/57 test strains in all inoculated test tubes in the form of diffuse turbidity of the medium and gas formation, test strain Enterobacter aerogenes 10006 - in the form of diffuse turbidity of the medium and weak gas formation, the Pseudomonas aeruginosa 27/99 test strain in the form of a slight turbidity of the medium without gas formation after 22-24 hours of incubation at a temperature of (37 ± 1) ° C, as well as the E. coli 675 test strain in the form of a diffuse turbidity of the medium and gas formation after 22-24 hours of incubation at a temperature of (43 ± 1) ° C with inoculation of 0.5 ml of a microbial suspension of each test strain from a dilution of 10 -6:

The Kessler-GRM Obolensk medium should completely suppress the growth of the test strain Proteus vulgaris HX 19 222 in all inoculated tubes when inoculated with 0.5 ml of microbial suspension from a dilution of 10 -4 and the test strain Staphylococcus aureus Wood-46 when inoculated with 0.5 ml of microbial suspension from a dilution of 10 -1 after 44-48 hours of incubation at a temperature of (37±1) °C.

1. PRECAUTIONARY MEASURES

Compliance with the "Rules for the arrangement, safety precautions, industrial sanitation, anti-epidemic regime and personal hygiene when working in laboratories (departments, departments) of sanitary and epidemiological institutions of the Ministry of Health" (Moscow, 1981).

1. EQUIPMENT AND MATERIALS

• Thermostat providing temperature 37±1 °C
• Scales laboratory 2 classes of accuracy
• Autoclave
• Glass test tubes
• Glass pipettes allowing to take liquid volumes of 1 and 2 ml
• Glass measuring cylinder with a capacity of 1000 ml
• Petri dishes sterile
• Distilled water
• flasks
• Glass funnels

1. SAMPLES ANALYZED

Objects of research in sanitary microbiology, scientific research.

1. Carrying out the ANALYSIS

Sample studies are carried out in accordance with the relevant Guidelines and GOSTs.

7.1. PREPARATION FOR ANALYSIS

Preparation of Kessler-GRM medium.

23.0 g of the drug is mixed in 1 liter of distilled water. Boil for 2-3 minutes, filter through a paper filter, pour 5 ml into sterile test tubes with floats and sterilize by autoclaving at a temperature of 112 ° C for 20 minutes.

Kessler's medium - medium for the isolation of enterobacteria on the basis of lactose fermentation, dry, is a finely dispersed homogeneous, hygroscopic, photosensitive powder of light cream color.

Composition (in terms of 1 liter of finished medium):
Enzymatic peptone, dry - 4.0 g.
Soy flour hydrolyzate - 4.0 g.
D(+)-lactose - 8.0 g.
Bile, dry - 4.0 g.
Gentian violet - 0.024 g.
Sodium carbonate - 0.2 g.

Liquid medium for the isolation of enterobacteria (E. coli group) on the basis of lactose fermentation in the sanitary and bacteriological study of food products and environmental objects (water, wastewater, etc.).

Dissolve 20 g of the drug in 1 liter of purified water, bring to a boil, boil for 2 minutes, if necessary, filter through a paper filter, pour 3 ml into sterile test tubes with floats (Durham tubes). Sterilize at 112°C for 20 minutes.

The prepared medium should be clear, purple in color with slight opalescence.

The prepared medium can be stored in a dark place for no more than 7 days at a temperature of 2-8°C until use. Incubate the inoculations of the studied samples for 24-48 hours at a temperature of 37°C. Growth of microorganisms capable of utilizing lactose changes the color of the medium from purple to pale purple. Durham tubes must accumulate gas. At the same time, the growth of gram-positive microflora is absent or weakly expressed.

The invention relates to sanitary microbiology. The medium for the detection of bacteria of the Escherichia coli group has the following composition, g/l of tap water: dry enzymatic peptone 3.0-5.0, sterile cattle bile 50.0-55.0, lactose 4.0-5.0, gentian violet 0.036-0.04, pH 7.51. The invention allows to reduce the cost of the nutrient medium. 3 tab.

The invention relates to sanitary microbiology and can be used in sanitary microbiological studies of food products and washings from environmental objects to detect bacteria of the Escherichia coli group. Known liquid selective Kessler medium for determining the fermenting ability of coliform bacteria to form acid and gas (Methods of Microbiological control of baby products, clinical nutrition and their components. Guidelines MUK 4.2.577-96. Ministry of Health of Russia. Moscow, 1998, p. 20 containing, g/l: Dry enzymatic peptone - 10.0 Sterile bovine bile - 50.0 cm 3 Lactose - 2.5 Tap water - 950.0 cm 3 1% Aqueous solution of gentian violet (crystal violet) - 4 cm 3 pH 7.51 There is evidence in the literature that micro-organisms capable of persisting in the environment grow effectively on media with a reduced amount of peptone.Biol.15 USA Univ.Maryland et Baltim.666.04 Biology Consolidated Volume 1995. Similar data were obtained by the authors (see Tables 1, 2, 3). ukt in a ratio of 1:10, which in themselves are an additional nutrient substrate for bacteria. The aim of the invention is to reduce the cost of the nutrient medium by 2-3 times with equivalent growth qualities of the proposed and known medium. The goal is achieved when using a medium containing, g/l of tap water: Dry enzymatic peptone - 3.0-5.0 Sterile cattle bile - 50.0-55.0
Lactose - 4.0-5.0
Gentian violet - 0.036-0.044
pH 7.51
Example 1. Preparation of the environment. Peptone, cattle bile is added to 1 dm 3 of tap water. The mixture is boiled with stirring in a water bath for 20-30 minutes. Then it is filtered through a cotton-gauze filter, lactose is added, the volume is adjusted to 1 dm 3 with water, the active acidity is set to 7.51 pH units using 1N NaOH or HCL solutions and checking the pH value on the potentiometer. 4 cm 3 of a 1% aqueous solution of gentian violet are added and poured in the required volumes into test tubes or flasks, floats are placed, sterilized for (151) minutes at (1211) o C. Example 2. Medium with minimal ingredients. The medium has the following composition, g/l of tap water:
Peptone enzymatic dry - 3.0
Sterile cattle bile - 50.0
Lactose - 4.0
Gentian violet - 0.036
pH 7.51
Growth qualities of the environment are presented in table. 1, 2, 3. Example 3. Medium with optimal values ​​of ingredients, g/l tap water:
Peptone enzymatic dry - 4.0
Sterile cattle bile - 52.5
Lactose - 4.5
Gentian violet - 0.038 cm 3
pH 7.51
Growth qualities of the environment are presented in table. 1. Example 4. Environment with maximum values ​​of ingredients. The medium has the following composition, g/l of tap water:
Peptone enzymatic dry - 5.0
Sterile cattle bile - 55.0
Lactose - 5.0
Gentian violet - 0.04
pH 7.51
Growth qualities of the environment are presented in table. 1, 3. Example 5. Determination of the growth qualities of the medium by the method of serial dilutions. The culture of E. coli 221 is inoculated into a test tube with 5 ml of meat-peptone broth. After 4 hours of incubation of the culture at 37 o C, one microbiological loop of the broth culture was inoculated into test tubes with 10 ml of Kessler medium with lactose, prepared from a dry commercial medium (TOO "Quadra" TU 10.02.875-90) and the proposed medium with various concentrations of peptone one%; 0.5%; 0.4%; 0.3%; 0.2%. From each tube after incubation of crops at 37 o With during the day produce a number of serial dilutions, starting from 10 -1 to 10 -12 . For dilution, Kessler medium is used, prepared in accordance with the instructions from a dry medium. All test tubes with dilutions are incubated for a day at 37 o C and turbidity and gas formation are noted at the end of the period. From all the test tubes, including the remaining transparent ones, seeding is done on the sectors of the cups with the Endo medium. The next day after the incubation of the crops at 37 o With account for the presence of the growth of characteristic red colonies with a metallic sheen in the appropriate dilutions, thus determining the growth quality of the environment with a certain content of peptone. Example 6. Determination of the growth qualities of the medium by the method of dosed seeding. The culture of Citrobacter freundii 227 is inoculated on beveled meat-peptone agar, cultivated at 37 o C for 18 hours. Using the turbidity standard, 1 ml is prepared. a suspension of culture in saline and produce a series of serial dilutions in saline to a concentration of 1000 microns. bodies in 1 ml, 500 b.w.; 100 bw; 10 b.w. in 1 ml. In test tubes with 10 ml of the proposed medium with a peptone concentration of 1%; 0.5%; 0.4%; 0.3%; 0.2% and 10 ml of Kessler's medium with lactose, prepared from a dry commercial medium, inoculate 1 ml of each of the prepared dilutions of the bacterial suspension. Crops are incubated at 37 o C for a day at 37 o C and take into account the growth in the respective sectors of characteristic dark pink colonies (see table. 3).

Claim

Modified Kessler medium for the detection of bacteria of the Escherichia coli group, containing dry enzymatic peptone, sterile cattle bile, lactose, gentian violet, tap water, characterized in that it contains the above-mentioned components in the following ratio, g/l of tap water:
Dry enzymatic peptone - 3.0 - 5.0
Sterile cattle bile - 50.0 - 55.0
Lactose - 4.0 - 5.0
Gentian violet - 0.036 - 0.04
pH 7.51.

Similar patents:

The invention relates to the field of medicine, and more specifically to medical microbiology, and can be used for laboratory diagnosis of gastroduodenal pathology associated with Helicobacter pylori, based on the detection of these microorganisms in biopsy specimens extracted during endoscopy, and the determination of their bacterial contamination

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Description 7974-00039

Kessler's medium Timing dry growth medium description

designed to detect bacteria of the Escherichia coli group on the basis of lactose fermentation during sanitary and bacteriological examination of food products and environmental objects.

Kessler's medium timing dry nutrient medium is a finely dispersed, hygroscopic, light-sensitive grayish-yellow powder.

Kessler's medium Timing dry nutrient medium specifications

Compound:

Enzymatic dry peptone, pancreatic hydrolyzate of fish meal, lactose, dry purified bile, crystal violet, sodium carbonate.

Preparation:

23.0 g of the drug is mixed in 1 liter of distilled water. Boil for 2-3 minutes, filter through a paper filter, pour 5 ml into sterile test tubes with floats and sterilize by autoclaving at a temperature of 112 ° C for 20 minutes.

The finished medium is purple.

The sterile medium can be used for 4 weeks if stored at a temperature
2-8C, in a dark place.

Operating principle:

The combination of components that make up the medium provides the nutritional requirements for the growth and detection of bacteria of the Escherichia coli group on the basis of lactose fermentation and inhibition of certain types of microorganisms.

Analyzing:

Sample studies are carried out in accordance with the relevant Methodological Guidelines and State Standards.

Incubate at a temperature of (37 ± 1) ° C for 24-48 hours, E. coli additionally at a temperature of (43 ± 1) ° C for 24 hours.

Quality Control:

Best before date: 2 years.

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