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IPC classes for russian patent Nutrient medium for determining antibiotic susceptibility of legionella pneumophila (RU 2542390):
 Bacteria bacillus subtilis producing 5`-aminoimidazole-4-carboxamideriboside (aicar), and method of microbiological synthesis of aicar by culturing this bacterium / 2542387
Bacterium Bacillus subtilis is proposed, producing 5′-aminoimidazole-4-carboxamideriboside (AICAR). The said bacterium comprises deregulated pur-operon against the background of inactivated gene purH, the modified heterologous genes prs and purF E.coli under the control of a strong promoter PrpsF as part of chromosome. At that it has one of the following characteristics: comprises the gene zwf under control of a strong promoter PrpsF, comprises the heterologous gene udhA under the control of a strong promoter PrpsF, comprises the deletion of the gene sacB. The method of synthesizing AICAR by culturing the said bacteria under proper conditions is also proposed. At that culturing is carried out on the medium of the following composition, wt %: fodder yeast 0.5-1.0, sucrose 10-13, soy isolate 2.5-3.5, corn-steep extract 3.0-5.0, urea 0,6-0,8, (H34)23CO4 0.8-1.6, propinol 0.4-0.5, water - the rest.
 Using bottom seawater of hydrogen sulphide pool of black sea as sea algae culture medium / 2541448
Invention "Using bottom sea water of hydrogen sulphide pool of the Black Sea as sea algae culture medium" refers to marine culture. Technical substance of the invention consists in using bottom seawater of the Black sea both containing hydrogen sulphide, and oxidised as a sea algae culture medium. Studying the biogenic properties of the aqueous medium from a reduction zone of the Black sea implemented by the invention has shown that the bottom seawater has no bad influence on the Black Sea plankton algae in the presence of high initial concentrations of hydrogen sulphide. After complete oxidation of this xenobiotic, the Black Sea bottom water can be used as a nutrient fertile culture medium of unicellular plankton and multi-cellular benthoalgae in laboratory or industrial environment, and in marine culture farms.
 Saccharomyces cerevisiae vkpm y-3973 yeast strain for fruit-and-berry wine production / 2540023
Invention relates to winemaking industry. "Quince -D" Saccharomyces cerevisiae yeast strain is deposited in the All-Russian Collection of Industrial Microorganisms (VKPM), Federal State Unitary Enterprise State Research Institute for Genetics, under Registration No Y-3973. Y-3973 strain has spore formation capability, ferments and digests glucose, sucrose, maltose, galactose, 1/3 of raffinose. Saccharomyces cerevisiae VKPM Y-3973 yeast strain has a high fermentative activity, well ferments quince wort containing sugar in an amount of 14.75 g/100 cm3 accumulating 8.79 % vol. of ethanol. The strain can more completely digest carbohydrates with formation of ethanol increased by 0.79 % vol. as compared to a conventionally known strain during the same period of fermentation. The specificity of the wine material produced with usage of Saccharomyces cerevisiae Y-3973 strain in a finer aroma and taste.
 Strain of lactobacillus delbrueckii, reducing content of cholesterol in blood / 2539785
Group of inventions relates to biotechnology. Claimed is strain of Lactobacillus delbrueckii subspecies lactis CNCM I-3741, reducing content of cholesterol in blood. Strain is applied for obtaining fermented dairy products.
Arthrospira platensis (nordst) geitl rsemsu t/05-117 strain - lipid-containing biomass producer / 2539766
Invention relates to biotechnology. The Arthrospira platensis (Nordst.) Geitl. rsemsu T/05-117 strain has high content of neutral lipids. The strain is stored in the collection of the research laboratory of renewable energy sources of the Faculty of Geography of Moscow Lomonosov State University.
 Bacillus subtilis strain capable of splitting sugars and having antagonistic effect on pathogenic and opportunistic pathogenic bacteria and fungi, and use thereof / 2539762
Group of inventions relates to biotechnology. Disclosed is a Bacillus subtilis VKPM V-11353 strain, capable of splitting a wide range of mono- and di-sugars and a wide range antagonistic effect on pathogenic and opportunistic pathogenic bacteria and fungi, which cause diseases in plants and farm animals. Also disclosed are versions of using the Bacillus subtilis VKPM V-11353 strain as a bacterial preservative for silos, for producing agents for normalising intestinal microflora of animal farms and for producing agents for protecting plants from diseases.
Strain of bacteria paenibacillus sp. for obtaining biological product against diseases of wheat caused by phytopathogenic fungi / 2539738
Strain of bacteria Paenibacillus sp. IB-1 has antagonistic activity against phytopathogenic fungi. The strain is deposited in the Russian National Collection of Microorganisms under the registration number VKM B-2823D and can be used to produce the biological product for protection of plants against diseases caused by phytopathogenic fungi.
Method of cleaning soils from oil under conditions of low positive temperatures with psychrotolerant bacteria pseudomonas sp. ib-1.1 / 2539148
Method comprises application into soil of suspension of microbial preparation on the basis of suspension of psychrotolerant bacterial strain Pseudomonas sp. IB - 1.1 with a titre of not less than 2.0·108 CFU/ml.
Bacillus subtilis b 93 vizr bacterial strain for protection of potatoes from illnesses during storage / 2538157
Bacillus subtilis strain - I5-12/23 has the antagonist activity with reference to phytopathogenic funguses and bacteria. The strain is reposited in GNU VIZR State Collection of Microorganisms under registration number Bacillus subtilis 93 VIZR also can be used for protection of potatoes against illnesses during storage.
Method for suppressing viability of pathogenic leptospira / 2537271
Invention relates to biotechnology and can be used for suppressing viability of pathogenic leptospira. The method includes growing strains of bacteria "Bacillus subtilis TNP-3-DEP" and "Bacillus subtilis TNP-5-DEP". The grown strains are used for the preparation of a suspension. The obtained suspension, containing the strains of bacteria "Bacillus subtilis TNP-3-DEP" and "Bacillus subtilis TNP-5-DEP" in equal ratios with the concentration of 5×108 CFU in 1 ml, is introduced into vials with cultures of leptospira, incubated at specified process parameters with the registration of a result by the absence of leptospira in the sample.
 Method for preparing pertussis component of combined vaccines / 2540014
Invention refers to biotechnology and concerns a method for preparing a pertussis component of complex vaccines. The presented method involves the B. pertussis culture growth on dense Bordet-Gengou and/or CCA nutrient media, the grown colonies selection according to the morphological characteristics, the selected colonies passage, the bacterial mass growth, the culture washout, the extinction reduction of the prepared pertussis suspensions to 10 international optical units, the agglutination reaction to measure agglutinogens 1, 2, 3 and the selection of pertussis suspensions, wherein the agglutinogen content is determined by type-specific serum dilution 1:3200 and more. The detected B. pertussis culture expressing agglutinogens 1, 2, 3 actively after the lyophilisation is used to prepare the pertussis component of complex vaccines.
Method for microorganism adhesive potential rapid test / 2537128
Method involves the preliminary sorption of a specific ligand in polystyrene tray wells; the above ligand is specified in: haemoglobin, myoglobin, collagen, fibrinogen, fibronectin, immunoglobulin A; that is followed by adding a microorganism cell suspension into the tray wells with the sorbed ligand, incubating the cell suspension in the tray wells for 15 minutes, sampling a suspension aliquot from the well and adding it to wells of another or the same tray containing 0.5% sodium chloride or 0.2 M sodium phosphate; the bacterial cell adhesion is assessed by measuring a decrease of the optical density of the prepared diluted suspensions at wave length 600 nm as compared to the well references free from ligands.
Vibrio aquamarinus strain, method of determining sample toxicity using same and testing culture for determining sample toxicity / 2534819
Group of inventions relates to biotechnology and can be used for biotesting of toxicity of environmental media. The invention discloses a Vibrio aquamarinus bacterial strain, a method of determining toxicity of samples using said strain and use of the strain as a testing culture for determining chemical toxicity of samples. The method comprises measuring luminescence of the Vibrio aquamarinus VKPM V-11245 strain in the presence of at least one chemical toxicant of a sample, said toxicant being ZnSO4 or CuSO4 or K2Cr2O7 or oil or diesel fuel or bilge water or phenol or a heavy metal. Luminescence of said strain is measured without a chemical toxicant and the measured luminescence levels are compared. Change in luminescence indicates toxicity of the analysed sample.
Method of quality control of disinfection of poultry premises / 2534356
Invention relates to the field of quality control of disinfection. The method comprises disinfection of premises, selection of test objects which are used as a suspension Bacillus subtilis of the strain ATCC PTA-6737 (PB6) sorbed on the carrier - the filter paper with an adhesive surface part. The test objects are placed in the premises of disinfection and added to the nutrient medium. The tubes are heated to 60-80°C and cultured at 37°C for 12-24 hours followed recording the result by recording the growth of bacteria after 12 hours. Clouding of the nutrient medium to form a mucous film is indicative of inadequate quality of disinfection, absence of clouding and formation of a film after 24 hours is indicative of the satisfactory disinfection.
Method for identifying staphylococcal exotoxin genes with high level of structural homology with superantigens by multiplex polymerase chain reaction and test system for implementing it / 2532225
Invention refers to biotechnology and microbiology and represents a method for identifying a cluster of antigens coding staphylococcal proteins that are exotoxins. The present method is implemented by performing a single multiple-primer polymerase chain reaction in two reaction mixtures, first of which contains primers to genes coding such staphylococcal proteins, as thermonuclease, beta-glucosidase in the species S.aureus, S.epidermidis, S.haemolyticus, S.lugdunensis, S.saprophyticus, and the second one - to set1, set2, set3, set4, set5 genes coding the exotoxins. That is followed by a comparative analysis of amplified gene fragments prepared in two sample aliquots and a positive control reference according to the identification table. The present invention also discloses a test system for implementing the above method, which contains DNA recovery components, PCR components, and result analysis components. The PCR components contain a 10-merous buffer solution, pH 8.4, deionised sterile water, one positive control reference, Taq polymerase, mixture of four dNTPs, mixture of primers No.1 containing epi-F, epi-R, aur-F, aur-R, hae-F, hae-R, lug-F, lug-R, sap-F, sap-R in a ratio of 1:1:1:1:1:1:1:1:1:1, and a mixture of primers No. 2 containing set1-F, set1-R, set2-F, set2-R, set3-F, set3-R, set4-F, set4-R, set5-F, set5-R in a ratio of 1:1:1:1:1:1:1:1:1:1.
 Method of generating starter culture, starter culture and fermentation method with its application / 2531343
Claimed solutions deal with a method of generating a starter culture, the starter culture and a fermentation method with an application of the said starter culture. The claimed method of generating the starter culture includes impact on a parent bacterial strain, which contains, at least, a part of the locus CRISPR, with a bacteriophage to obtain a mixture of bacteria, which contains a bacteriophage-resistant variant strain, containing the modified locus CRISPR, which contains, at least, one additional spacer in the said modified locus CRISPR; independent impact on the same parent bacterial strain with the same bacteriophage to obtain the mixture of bacteria, which contains other bacteriophage-resistant variant strain, containing the modified locus CRISPR, which contains, at least, one additional spacer in the said modified locus CRISPR, different from the additional spacer in the first bacteriophage-resistant variant strain, selection of the said bacteriophage-resistant variant strains from the mixtures of bacteria and their separation.
Method of increasing sensitivity of microorganisms to antimicrobial agents / 2529367
Two suspensions are prepared. Clinical polyantibiotic-resistant strains Escherichia coli are added to an isotonic solution of NaCl to achieve the concentration of 30-40 thousand CFU/ml. Copper nanoparticles are added to the solution of NaCl to achieve the concentration of 0.01-0.05 mg/ml. The suspension of ethylenediaminetetraacetic acid - EDTA is prepared by its dilution in distilled water at the rate of 0.1-0.2:1, respectively. NaOH is added to the prepared suspension to obtain the solution of EDTA with pH=7.6-8. The prepared suspensions are connected with the solution of EDTA in the following ratios by wt %: suspension of copper nanoparticles - 70-85, suspension of microorganisms - 10-20, EDTA - 5.10. It is incubated in the shaker at 100-150 rev/min and a temperature of 36-38°C for 40-60 minutes. The resulting biomass is inoculated on the solid nutrient medium with the volume of 20-25 ml in the amount of 0.1-0.12 ml. It is incubated in the thermostat at a temperature of 36-38°C for 18-24 hours. The sensitivity of E.coli strains to antibiotics is determined.
 Method of specific differentiation of viable rhodococcus immobilised in gel carrier / 2525934
Granule of biocatalyst with the cells contained in it of one or more species of Rhodococcus is placed in a well of a 96-well plate, a colorant iodine-nitro-tetrazolium (INT) is added. After the appearance of specific purple staining formazan the optical density of granule is measured using a tablet photometer. In the case of a stained substrate or preliminary presence of the immobilised biocatalyst in the soil the extraction of formazan is carried out with 96% ethanol and the optical density of the extract is measured. The amount of viable Rhodococcus in the granule of the gel is determined using the calibration dependency diagram of the optical density on the number of living cells in suspension as determined by traditional method of plating on solid growth medium. Then, from the granule stained by the colorant of the biocatalyst or from the granule after extraction of the colorant the DNA is isolated and PCR is performed using sets of species-specific primers. The specific position of the immobilised Rhodococcus is determined.
Method of evaluating detoxification activity of black soils in farming ecosystems / 2525677
Invention relates to biotechnology. The method comprises collecting soil samples, adding metsulfuron-methyl (MSM) to the sample in a given amount, followed by incubation and determining specific metabolic activity of the saprophytic soil complex by multisubstrate testing and counting the number of colony-forming units on a diluted agarised medium. The level of the detoxification activity is estimated based on the relative increase in specific metabolic activity of the saprophytic microbial community of the soil when MSM in comparison with characteristics of the soil without adding MSM (Kd factor).
 Method of growing colonies of microbial cells and device for its realisation / 2522005
Group of inventions relates to biotechnology. Claimed is a method of growing colonies of microbial cells on a surface of a porous plate. The method includes supply of a nutrient solution from bottom to top through the porous plate into zones of growth of colonies of the microbial cells on its upper surface, supply of a suspension of the microbial cells onto the upper surface of the porous plate, creation of controlled conditions for the colony growth, performing observation of the colony growth, separation of the grown colonies of the microbial cells from the zones of growth and their transfer into external means of identification. The nutrient solution is supplied into the zones of growth of the colonies of the microbial cells by creation of a pressure difference between the hole input and output. Holes are made in the plate from an anode aluminium oxide orthogonally to its large plane and are topologically coded. The said zones of growth are formed in them in the form of porous membranes. The porous membranes are located at the same level as the upper surface of the plate or with formation of a hollow and do not pass the microbial cells. After supply of the nutritional solution, the suspension of the microbial cells of a specified concentration is supplied onto the upper surface of the plate until their homogenous distribution is achieved. Between the zones of growth on the surface of the plate a film, preventing attachment of the microbial cells, is formed. Separation of the grown microcolonies from the zones of growth is performed by hydroblow. A hydroblow is directed from the side of the input of cylindrical holes of the plate and spreads along them and farther through the pores of the porous membranes with force, which does not destroy the microcolonies but is sufficient for their separation from the growth zones. Also claimed is a device for growing the colonies of the microbial cells by the claimed method.
 Method for production of bacterial concentrate and its application as probiotic biologically active food additive / 2541778
Inventions group relates to biotechnology and may be used for preparing bacterial preparations applicable as probiotic biologically active additives. The bacterial concentrate production method envisages preparation of a nutritional medium, sterilisation and cooling. Inoculum introduction, cell propagation, bacterial mass separation from culture liquid, dispensing and closuring. Into the nutritional medium composition one introduces cedar or linseed oil of fish or ringed seal fat in an amount of 1-1.5% of the medium weight; the inoculum is represented by a Bifidobacterium longum DK-100 strain. The produced bacterial concentrate is used as a probiotic biologically active food additive.
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FIELD: biotechnology. SUBSTANCE: nutrient medium comprises enzyme-autolyzate of cattle spleen, monosubstituted potassium phosphate, disubstituted potassium phosphate, potassium gluconate, mesoinosite, soluble starch, gelatin, L-cysteine, iron (III) sodium salt, high-grade ink and distilled water in a predetermined ratio of components. EFFECT: invention enables to improve the accuracy of determining of antibiotic susceptibility of cultures of legionellosis causative agent. 3 tbl 8 ex
The present invention relates to medical Microbiology, namely to obtain a dense nutrient medium through which you can determine the antibiotic sensitivity of the cultures of the pathogen regionaliza - Legionella pneumophila. Common environment to determine the antibiotic sensitivity of the vast majority of pathogenic bacterial species is medium Mueller-Hinton (The Oxoid Manual, 1976, p.193) containing, g/l:. agar, Difco - 10; infusion from beef - 300; casein hydrolysate - 17,5; soluble starch - 1,5; distilled water to 1 liter at a pH of 7.4. However Legionella in the environment, due to the nature of their metabolism, even with the introduction of various additives grow poorly, so the medium Mueller-Hinton for the study of antibiotic sensitivity Legionella is not applied. A prototype of the selected nutrient medium for the cultivation and selection Legionella environment ά (see WHO. Legionella and the prevention of legionellesis. WC 200. WHO. 2007), which has the following composition, g/l: yeast extract - 10,0; agar, Difco 13,0; charcoal Norit N - 2,0; ACES - 10; alpha Ketoglutarate acid - 1,0; L-cysteine - 0,4; pyrophosphate of iron - 0,25; glycine - 3,0; cycloheximide - 80,0-100.0 mg/l; vancomycin - 1.0 mg/l; polymyxin In 80-100 thousand IU/l; distilled water to 1 liter at a pH of 7.1. The disadvantage of the prototype is that Legionella are very sensitive to peroxide compounds and the growth of a single virulent cells of the pathogen in the environment is only possible when the environment is highly active adsorbent, blocking the action of free radicals, is the activation of coal, made by special technology. However, coal is strongly adsorbs many of the antibiotics currently in use, such as ofloxacin, ciprofloxacin, rifampicin, clindamycin, and others (see K. Nilsen, J. V. Bangsborg, N. Hoiby Susceptibility of Legionella to five antibiotics and development of resistance by exposure to erythromycin, ciprofloxacin assay and rifampicin. Diagn. Environ. Infect. Dis., 36 (2000), 43-48). Therefore, for this reason, the results characterizing antibiotikoustoichivosti cultures of Legionella received on the environment, unreliable as indicators of inhibition Legionella in most cases artificially low. The technical problem of the invention was to create a new nutrient medium with a high degree of accuracy in determining antibiotikoustoichivosti Legionella pneumophila. This object is achieved in that the culture medium to determine the antibiotic susceptibility of Legionella pneumophila, including microbiological agar and L-cysteine, added EDTA iron(III) sodium salt, growth stimulants - potassium phosphate one-deputizing and potassium phosphate (disubstituted), and as osmoprotection - metainit and potassium gluconate, at the same time as nutrition in the environment used fermentational spleen cattle what about the livestock and food gelatin, in the following ratio of components, g/l:
A nutrient medium is prepared as follows. The source materials for the proposed environment (denoted B-2): agar microbiological 14,0-16,0; -cysteine 0.3 to 0.5 and starch soluble 2,4-2,6; as the growth fundamentals take fermentational spleen of cattle of 6.0-8.0 and gelatin food 2,4-2,6; as a buffer, a mixture of phosphate salts: potassium phosphate one-deputizing (KH2PO4) 1.5 to 1.7 and potassium phosphate disubstituted (K2HPO4) 3,3-3,6. Osmoprotection selected metainit 2,4-and 2,6 potassium gluconate 2,4-2,6, and as the source of iron used complex EDTA iron(III) sodium salt 0,026-0,028. To give a medium dark shade, which is clearly visible zone of inhibition growing culture Legionella, use high-quality fine sterile ink 4-6 ml All of the above components, except for L-cysteine and EDTA iron(III)sodium salt, bring in a container and add to 1 liter of cold distilled water. The mixture is heated in a steam sterilizer and boil for 30 minutes at 100°C with careful stirring. The liquid is then autoclave at 0.5 psi for 20 minutes the result is a Foundation that is stored at a temperature (2-8°C in the refrigerator for 2 years. If necessary, the preparation of the working environment based melt and enter the number of the other two additives: L-cysteine and EDTA iron(III) sodium salt. The environment in the melted state (45°C) is poured into plastic Petri dishes with a diameter of 90 mm in a volume of 30 ml and thickness is Noah layer of 0.7 see Cups store in an airtight package when the temperature of the refrigerator for 1-2 months. Basic biological performance of the proposed environment B-2 is determined in comparison with the best for cultivation of Legionella environment BCYEAά. First characterize its sensitivity, germination colonies of Legionella, as well as the effectiveness of the environment (the amount of grown colonies and the development of turf culture) - examples 1, 2, 3, 4, and 5. Then, using for comparison the same environment ά determine the fundamental suitability of the environment B-2 to determine the antibiotic susceptibility of Legionella cultures using discs with antibiotics (example 6), strips with gradations of antibiotics (example 7) and using the method of serial dilutions of antibiotics in the environment (example 8). In the experiments used the virulent and avirulent strains of Legionella, obtained from the Museum of living cultures Rostov-on-don, anti-plague Institute: 1) L. pneumophila Philadelphia ATS 33152, serogroup 1, LD50 - 4 105M. K. for seawater. pigs (virulent) 2) the same, but avirulent strain 3) L. pneumophila Blumington ADS 33155, 3 serogroup (avirulent) 4) L. pneumophila Chicago ATS 332 215, serogroup 6 (avirulent) Example 1 For cooking environment using ingredients in the following minimum concentrations, g/l: fermentational spleen of cattle - 5,0; microbiological agar - 2,0; potassium phosphate one-deputizing - 1,0; potassium phosphate disubstituted - 3,0; potassium gluconate - 2,0; soluble starch - 2,0; gelatin - 2,0; metainit - 2,0; L-cysteine - 0,1; EDTA iron(III) sodium salt of 0.01; high quality ink 3 ml; distilled water to 1 l, pH of 7.1. The composition of the medium comparison ά similar to the above (see prototype). In this work, we used the standard methods of determining the sensitivity of nutrient media for Legionella. Used standard strain of Legionella - Legionella pneumophyila Philadelphia - 1 - virulent strain, LD 50 - 4 105M. K. for Guinea pigs and three avirulent strain. Preparation of inoculum: crops grown for 2 days on medium ά at 35°C, then washed with potassium phosphate buffer (pH of 7.1). After that prepare a suspension of cells with a concentration of 1 billion, M. K./ ml, which was diluted to the working concentration of 103m/ml and seeded in a Cup of 0.1 ml, i.e., 100 M. K. Crops incubated for three days at 35°C, count the number of grown colonies, determine the time of their appearance, the degree of atypical morphology legionellas colonies and compare their diameter in different environments at different times of incubation. The sowing was done three times, for comparison using average values. Statistical significance of the similarities and differences of the values determined using Chi-Quadra is. The results revealed that chuvstvitelnosti both environments statistically differed from 100 sown M. K. Legionella all four strains on the third day of incubation in the medium B-2 composition of example 1 was increased to 20 colonies of the pathogen on the control environment - 50-60 typical colonies of Legionella (p is 0.05, Chi-square 6,0), i.e., the degree of sensitivity of the first environment is insufficient. Similar differences in the time of appearance of colonies on the media - in all cases, the proposed environment they were recorded using a stereomicroscope on the fifth day of incubation, visually on the sixth - in the form of small rosinate colonies up to 1 mm in diameter, stereomicroscopes with typical fine grain and melkopyatnistoy with a greenish glow. On the prototype environment stereomicroscopes colonies of Legionella were registered on the second day, visually - third that meets the standards. Efficiency environment ά was much better than Wednesday B-2 of example 1, i.e., the amount of colonies on 5 days of growth on average in the first environment was 4-5 mm, the second maximum diameter of colonies on 6 days of growth was 2-3 mm. Growth of lawn culture on the prototype environment at inoculation of 0.1 ml of suspension culture Legionella all strains tested after one day of incubation at 35°C was sufficient for its visual check on proposed was extremely weak. T is thus, Wednesday, prepared according to example 1, does not satisfy the requirements as a diagnostic of nutrient medium for Legionella pneumophila, and cannot be used to determine its antibiotic sensitivity. Example 2 To prepare the environment used the above ingredients in the following concentrations, g/l: fermentational spleen of cattle - 6,0; microbiological agar - 14,0; potassium phosphate one-deputizing - 1,3; potassium phosphate disubstituted - 3,2; potassium gluconate - 2.3; soluble starch - 2,3; gelatin - 2.3; metainit - 2,3; L-cysteine - 0,3; EDTA iron(III) sodium salt - 0,025; a high quality ink - 4 ml; distilled water to 1 l, pH of 7.1. Methods to determine and compare the sensitivity, time of germination and the efficiency of the proposed nutrient medium B-2 and ά were identical to those described in example 1. The results of the comparison of the sown 100 M. K. virulent and avirulent strains of Legionella in both environments was raised on the third day 50-60 typical colonies (p is 0.05, Chi-square - 0,31), the period of formation of colonies - 2 nights when observed through a stereomicroscope and 3 days by visual observation. But the efficiency of growth of colonies on the 5th day of incubation environment ά was somewhat better prepared according to example 2. At the same time, the severity of lawn culture on the environment was quite the full visual check on the 2 days of growth. Thus, the composition of the medium, prepared according to example 2, provides the growth of Legionella to the extent that this environment was considered as corresponding to generally accepted standards of the who. Example 3 To prepare the environment used the above ingredients in the following concentrations, g/l: fermentational spleen of cattle - 7,0; microbiological agar - 15,0; potassium phosphate one-deputizing - 1,6; potassium phosphate disubstituted - 3,4; potassium gluconate - 2,5; soluble starch - 2,5; gelatin - 2,5; metainit - 2,5; L-cysteine - 0,4; EDTA iron(III) sodium salt - 0,027; ink high-quality - 5 ml; distilled water to 1 l, pH of 7.1. Methodology a comparative study of the sensitivity and efficiency of both environments consistent with the above. The obtained results (growth 45-65 typical colonies of all tested strains in both environments, the appearance of colonies on 2-3 day incubation at 35°C, sufficient efficiency of colony growth and turf crops with some exceeding these indicators on environment comparisons) indicate that the environment is taken in this part, fully complies with the requirements of diagnostic reference nutrient medium BCYEAά used for the cultivation and selection of Legionella pneumophila. Test environment specified in part on these grounds may be used for the detection of the of the antibiotic susceptibility of Legionella cultures and for normal growth. Example 4 To prepare the environment used the above ingredients in the following concentrations, g/l: fermentational spleen of cattle - 8,0; microbiological agar - 16,0; potassium phosphate one-deputizing - 1,7; potassium phosphate disubstituted - 3,6; potassium gluconate - 2,6; soluble starch - 2,6; gelatin - 2,6; metainit - 2,6; L-cysteine - 0,5; EDTA iron(III) sodium salt - 0,028; ink high quality 6 ml; distilled water to 1 l, pH of 7.1. Methodology of comparative study of the above indicators of both environments corresponded described. The obtained results (growth 50-65 typical colonies of all tested strains in both environments, the appearance of colonies on 2-3 day incubation at 35°C, sufficient efficiency of colony growth and turf crops with some exceeding these indicators on environment comparisons) indicate that the environment is taken in this part, fully complies with the requirements of diagnostic reference nutrient medium BCYEAά used for culturing and selection Legionella. The test environment in this composition on the above grounds may be used for detection of antibiotic susceptibility of Legionella pneumophila cultures and for normal growth. Example 5 The environment in this example is prepared with maximum concentrations of ingredients, g/l: EN zymes is autolist spleen of cattle - 9,0; microbiological agar - 17,0; potassium phosphate one-deputizing - 1,8; potassium phosphate disubstituted - 4,0; potassium gluconate - 2,8; soluble starch - 2,8; gelatin - 3,0; metainit - 3,0; L-cysteine - 0,8; EDTA iron(III) sodium salt - 0,03; a high quality ink - 10 ml; distilled water to 1 l, pH of 7.1. Methodology of comparative study of the above indicators of both environments corresponded described (see example 1). Results: growth of up to 20 typical colonies of all tested strains in the test environment with significantly better growth environment comparison (50-60 colonies with reliable statistical difference P - 00,5 Chi-square - 8,0), longer periods of appearance of colonies on the environment of this part for 4-5 day incubation at 35°C. Conclusion: insufficient efficiency of colony growth and turf crops on the environment show that the environment is taken in the present composition does not meet the requirements for diagnostic reference nutrient medium BCYEAά used for culturing and selection Legionella. The test environment in this composition on the above grounds may not be used for detection of antibiotic susceptibility of Legionella cultures and for normal growth. Example 6 The suitability of the proposed environment in the determination of the antibiotic susceptibility of crops Le is jonell method of disks in comparison with the environment ά Prepare the environment for the recipe of example 3 and Wednesday ά according to the common recipe. First environment has a dark color, the other black. The volume of media in Petri dishes - 30 ml Prepare odnostoechnye culture of these strains of Legionella, which is seeded on agar plates in both environments 0.1 ml of the suspension of Legionella cells (4×109M. K.), distribute them over the surface and dried agar. Then on agar impose disks with antibiotics, most commonly used to treat legionellosis: rifampicin, ofloxacin, clindamycin, erythromycin, doxycycline, moxifloxacin, azithromycin, levofloxacin, and ciprofloxacin (antibiotic disc 1-1,25 µg). For every Cup of post no more than three disks. Crops incubated for 48 h at 35°C, the first accounting carried out after 24 h by measuring the zone of inhibition of growth of culture and compare their diameters in both environments. On the second day these indicators measure again. The results of comparative tests in determining antibiotikoustoichivosti cultures of Legionella are presented in table 1. As can be seen from table 1, the zones of inhibition of growth of lawns cultures of Legionella all tested strains around eight discs of antibiotics (rifampicin, ofloxacin, clindamycin, erythromycin, amoxicillin, levofloxacin and ciprofloxacin) on the environment proposed in 2-4 times statisticslecture (with an average Chi-square = 6,3) and higher than similar areas, received on the control environment BCYEAά, explained by the adsorbing action of activated charcoal in the past. A small zone of inhibition of growth of Legionella drive around with doxycycline depends on relatively weak sensitivity of Legionella to the agent, accompanied by complete absence of adsorbing effect. Thus, the proposed environment sensitivity to antibiotics Legionella detected more accurately and completely than in the medium BCYEAά. Example 7 This example presents the results of a comparative test environment, taken in the optimal composition (as in example 3), and environment BCYEAά to determine the antibiotic susceptibility of Legionella cultures using strips containing a concentration gradient of antibiotic - erythromycin and levofloxacin (produced by Oxoid, F - test). Culture Legionella prepared as described in example 6. Strips nakladyvat planted on the lawn culture at number two on the Cup. Analysis carried out over night incubation at 35°C, the other day the light of the results of repeat. In the presence of culture sensitivity to the subject an antibiotic around the strip formed Racecourse zone of inhibition of growth of a culture of Legionella, the lower edge of the zone of inhibition was consistent with MIC - minimum inhibitory concentration. The results are presented in table the CE 2. As can be seen from table 2, the zones of inhibition of growth of lawns cultures of Legionella in the environment proposed is approximately 5 times larger than the MICK similar performance to the control environment (average Chi square = 6,3). Therefore, the results of determination of antibiotic susceptibility Legionella using strips are more accurate for the proposed environment. Example 8 Determining the suitability of the proposed environment to determine the antibiotic susceptibility of Legionella cultures by the method of serial dilutions of antibiotics in comparison with the environment ά Prepare a Cup with authors and α, as indicated in the previous case, except that on Wednesday injected antibiotic in stepwise increasing concentrations: 0,01, 0,05, 0,1, 0,25, 05, 0,75, 1 mcg/ml are Seeded above culture Legionella on the surface of the agar with antibiotics in a dose - big loop suspension with a concentration of 4×109M. K., forming sowing in the form of a disk with a diameter of 1 see the Crops dried and incubated under the same conditions as in the previous case. The incubation period is 2 days, the account after 24 and 48 hours Results take into account the presence or absence of growth of the culture (see table 3). The parameters are compared for both environments. As can be seen from table 3, the minimum inhibitory concentration of antibiotics specific for Legionella pneumophila, the proposed environment in 5-10 times less than that in the medium containing activated charcoal. Therefore, testing of the two environments to determine the antibiotic sensitivity Legionella different methods, namely using discs, strips and serial dilutions, confirmed that the most accurate and reliable working environment B-2. Therefore, when Troubleshooting a strong adsorbent is activated charcoal, which binds a number of antibiotics, reducing this sensitivity, the new environment provides the ability to accurately determine antibiotikoustoichivosti Legionella pneumophila, not understating the performance inhibition, which is of great importance in clinical practice in the treatment of regionaliza.
Nutrient medium for determining the antibiotic susceptibility of Legionella pneumophila, including microbiological agar and L-cysteine, characterized in that the composition of the medium was additionally introduced EDTA iron(III) on rieva salt, growth promoters - potassium phosphate one-deputizing and potassium phosphate (disubstituted), and as osmoprotection - metainit and potassium gluconate, at the same time as nutrition in the environment used fermentational spleen of cattle and food gelatin, in the following ratio of components, g/l:
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