CN110564808A - Selective chromogenic culture method for gluconacetobacter, acetobacter aceti and gluconobacter in fermented milk and special culture medium thereof - Google Patents
Selective chromogenic culture method for gluconacetobacter, acetobacter aceti and gluconobacter in fermented milk and special culture medium thereof Download PDFInfo
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/20—Bacteria; Culture media therefor
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
- C12Q1/02—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving viable microorganisms
- C12Q1/04—Determining presence or kind of microorganism; Use of selective media for testing antibiotics or bacteriocides; Compositions containing a chemical indicator therefor
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
- C12Q1/02—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving viable microorganisms
- C12Q1/04—Determining presence or kind of microorganism; Use of selective media for testing antibiotics or bacteriocides; Compositions containing a chemical indicator therefor
- C12Q1/045—Culture media therefor
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N2333/00—Assays involving biological materials from specific organisms or of a specific nature
- G01N2333/195—Assays involving biological materials from specific organisms or of a specific nature from bacteria
Abstract
the invention discloses a selective chromogenic culture method aiming at acetobacter gluconicum, acetobacter aceti and gluconobacter in fermented milk, which comprises the following steps: A. the special chromogenic medium is prepared as follows: 40-60g/L of glucose, 4-6g/L of beef extract, 8-12g/L of yeast powder and 8-12mL/L of ethanol, and additionally adding 0.5-1.5g/L of bile salt and 0.5-1.5mL/L of Swiss dye solution; the pH value is 6.6-7.2; B. pretreating fermented milk to be detected; C. selective chromogenic culture: inoculating the culture solution into the special chromogenic culture medium in the step A according to the inoculation amount of 3-8% v/v, and carrying out streak culture; D. and (3) color development identification of three target strains. The outstanding characteristic of the invention is that a method for rapidly identifying three groups of mixed bacteria simultaneously is creatively obtained, selective culture and color development are combined, identification is rapid, stable and reliable, and the invention has very important scientific research and practical value.
Description
Technical Field
The invention relates to the technical field related to dairy inspection, in particular to a method for quickly culturing and identifying color development of mixed bacteria in fermented milk and a special culture medium thereof.
Background
the species of the pollution bacteria and pathogenic bacteria in the fermented milk are various, the pollution bacteria and the pathogenic bacteria comprise salmonella, staphylococcus aureus, shigella and other pathogenic bacteria and bacteria affecting the product quality such as acetobacter in the family of acetobacteriaceae and lactobacillus plantarum, the salmonella, the staphylococcus aureus, the shigella and the cronobacter sakazakii all have corresponding selective chromogenic culture media which can meet the requirement of specific screening, and the acetic acid bacteria serving as common pollution bacteria in the fermented milk lack of specific chromogenic culture media and become a key factor restricting the quality control of the fermented milk.
The identification of microorganisms by the chromogenic culture medium is a technology developed on the basis of biochemical reaction and is a hot spot of research and development at home and abroad. The principle is that the species and reaction conditions of microbial intracellular enzymes are used as the basis for microorganism classification identification, and a substrate of microbial specific enzymes is added into a separation culture medium, wherein the substrate consists of chromophoric groups and microbial metabolizable substances, is usually colorless, but is separated from the chromophoric groups under the action of the specific enzymes and shows a certain color, and the bacteria can be identified by directly observing the color of a bacterial colony, so that the steps of pure culture and further biochemical identification of the bacterial strain are reduced.
At present, the color development identification method for one-time multidimensional culture of various bacteria has important scientific research and practical value, but is still to be developed and researched.
Disclosure of Invention
the invention aims to solve the technical problem of providing a selective chromogenic culture method aiming at gluconacetobacter acetobacter, acetobacter aceti and gluconobacter in fermented milk and a special culture medium thereof.
In order to solve the technical problems, the technical scheme adopted by the invention is as follows.
A selective chromogenic culture method aiming at acetobacter gluconicum, acetobacter aceti and gluconobacter in fermented milk comprises the following steps:
A. Preparation of a series of culture media:
A-1, a special chromogenic medium, which is prepared as follows: 40-60g/L of glucose, 4-6g/L of beef extract, 8-12g/L of yeast powder and 8-12mL/L of ethanol, and additionally adding 0.5-1.5g/L of bile salt and 0.5-1.5mL/L of Swiss dye solution; the pH value is 6.6-7.2;
A-2, preparing a liquid basal culture medium as follows: 80-120 parts of glucose, 4-6 parts of beef extract, 4-6 parts of yeast powder, 15-25 parts of ethanol, 1000 parts of distillation by volume and 6.5-7.5 of pH;
B. Pretreatment of fermented milk to be detected:
B-1, first-generation culture: taking fermented milk to be detected, standing to restore the fermented milk to normal temperature, and inoculating the fermented milk to a liquid basal culture medium for first-generation activation culture;
b-2, second generation culture: after the basic culture is finished, adding the culture solution into a liquid basic culture medium to perform second generation activation culture;
C. Selective chromogenic culture:
B, inoculating the culture solution obtained by the second generation culture into the special chromogenic culture medium in the step A according to the inoculation amount of 3-8% v/v, and carrying out streak culture;
D. Color development identification of three target strains:
d-1, identifying three target strains according to the following typical standards: gluconacetobacter: the colony color is purple black, and an obvious transparent ring is arranged around the colony;
D-2, Acetobacter aceti: the colony is wine red, and transparent rings are not arranged around the colony;
d-3, Gluconobacter: the colony color is pink, and no transparent ring is arranged around the colony color; and (5) finishing the identification.
In a preferred embodiment of the present invention, in step A-1, the dedicated chromogenic medium is configured as follows: 50g/L of glucose, 5g/L of beef extract, 10g/L of yeast powder and 10mL/L of ethanol, and additionally adding 1g/L of bile salt and 1mL/L of Swiss dye solution; the pH value is 6.8.
In a preferred embodiment of the present invention, in step A-1, the dedicated chromogenic medium is configured as follows: 40g/L of glucose, 4g/L of beef extract, 12g/L of yeast powder and 12mL/L of ethanol, and additionally 0.7g/L of bile salt and 1.3mL/L of Swiss dye liquor are added.
In a preferred embodiment of the present invention, in step A-1, the dedicated chromogenic medium is configured as follows: 60g/L glucose, 6g/L beef extract, 8g/L yeast powder and 8mL/L ethanol, and additionally adding 1.2g/L bile salt and 0.8mL/L Swiss dye solution.
In a preferred embodiment of the present invention, in step a-2, the liquid basal medium is prepared as follows: 100 parts of glucose, 5 parts of beef extract, 5 parts of yeast powder, 20 parts of ethanol, 1000 parts of distillation by volume and pH 6.8.
As a preferred technical scheme of the invention, the specific operating parameters of the step B-1 are as follows: b-1, first-generation culture: taking fermented milk to be detected, standing to restore the normal temperature, taking 1mL, inoculating into 10mL of liquid basal culture medium, and carrying out shake culture at 30 ℃ and 200r/min for 48 h.
As a preferred technical scheme of the invention, the specific operating parameters of the step B-2 are as follows: b-2, second generation culture: after the basic culture is finished, 10mL of culture solution is taken out and added into 200mL of liquid basic culture medium, and shaking culture is carried out for 36h at 30 ℃ and 200 r/min.
As a preferred technical scheme of the invention, the specific operating parameters of the step C are as follows: C. selective chromogenic culture: b, inoculating the culture solution obtained by the second generation culture into the special chromogenic culture medium in the step A according to the inoculation amount of 3-8% v/v, carrying out streak culture, and carrying out shake culture at 30 ℃ for 36h at 200 r/min; and detecting the pH value and the viable count of the bacteria.
The invention also comprises a special chromogenic medium in the method.
the invention also comprises the application of the special chromogenic medium in culturing, distinguishing and identifying the gluconacetobacter acetobacter aceti and the gluconobacter gluconicum in the fermented milk.
Adopt the produced beneficial effect of above-mentioned technical scheme to lie in: the invention researches common acetobacter aceti in acetobacter family in fermented milk, optimizes the components of the culture medium by taking acetobacter gluconi, acetobacter aceti and gluconobacter as target strains, screens and researches the selective culture medium by utilizing the characteristics of bile salt and Swiss dye solution, and provides necessary technical support for the process control of fermented milk production enterprises.
The outstanding characteristic of the invention is that a method for rapidly identifying three groups of mixed bacteria simultaneously is creatively obtained, selective culture and color development are combined, identification is rapid, stable and reliable, and the invention has very important scientific research and practical value.
Drawings
FIG. 1A bacterial colony morphology of gluconacetobacter liquefaciens.
FIG. 2 Acetobacter aceti colony morphology.
FIG. 3 Escherichia coli colony morphology.
FIG. 4 the colonial morphology of Gluconobacter thailand.
FIG. 5 Salmonella typhimurium colony morphology.
FIG. 6 Salmonella typhi colony morphology.
FIG. 7 Proteus vulgaris colony morphology.
FIG. 8 Shigella sonnei colony morphology.
Detailed Description
The following examples illustrate the invention in detail. The raw materials and various devices used in the invention are conventional commercially available products, and can be directly obtained by market purchase. Wherein each strain is from a collection center; the bile salt reagent is purchased from chemical reagents of national drug group, ltd; swiss dye liquor (A, B liquor) was purchased from the institute of optochemical and fine chemistry in Tianjin.
Example 1 strains
The strains used in this experiment are detailed in the table below and are all from the strain collection.
Test strains
Strains in this study
Example 2 culture Medium & reagents
Basal liquid medium: 100g of glucose, 5g of beef extract, 5g of yeast powder, 20mL of ethanol, 1L of distilled water and pH 6.8.
solid medium: 1.5% agar was added on a liquid medium basis.
The collection center provides media: 100g of glucose, 20g of anhydrous calcium carbonate, 10g of yeast powder and 1L of distilled water, and the pH value is natural.
The main reagents are as follows: beef extract, yeast powder, agar, glucose and anhydrous calcium carbonate are all biochemical reagents, sodium hydroxide, sodium chloride and absolute ethyl alcohol are analytically pure, and the No. 3 bile salt is purchased from national medicine group chemical reagent company Limited; swiss dye liquor (A, B liquor) was purchased from the institute of optochemical and fine chemistry in Tianjin.
EXAMPLE 3 apparatus and Equipment
Model BS-124S electronic balance, beijing sumatris; ES2315 autoclave sterilizer, TOMY, Japan; HWY 22112 type constant temperature culture shaking table, Shanghai Zhicheng analytical instruments manufacturing Co., Ltd; MIR-254 incubator, SANYO, Japan.
Example 4 culture activation method
First-generation seed liquid: taking out the strain cryopreservation tube from an ultralow temperature refrigerator, inoculating 1mL of the strain to 10mL of liquid basal culture medium after the strain is melted, and carrying out shake culture at 30 ℃ and 200r/min for 48 h;
the second generation of seed liquid: after the first generation culture, 10mL of culture solution is taken and added into 200mL of liquid basal medium, and shake culture is carried out for 36h at 30 ℃ and 200 r/min.
example 5 development of Selective Special chromogenic Medium
5.1 optimization design: 3 levels of orthogonal test L with 4 factors of glucose content, beef extract content, yeast powder content and ethanol content9(34) Experiments were performed to determine the proper growth medium composition of the strains.
5.2 Medium validation test: the defined optimal growth medium for the strain is compared with the medium of the strain collection. Preparing two culture media according to the formula, inoculating the second generation seed solution into the culture media according to the inoculation amount of 5%, performing shake culture at 30 ℃ for 36h at 200r/min, and detecting the pH value and viable count of the second generation seed solution.
5.3 bile salt bacteriostasis test: based on a proper growth medium, the addition amounts of bile salts are respectively 0.1%, 0.2%, 0.3%, 0.4% and 0.5%, gram-positive bacteria are staphylococcus aureus ATCC6538, bacillus cereus CMCC63302, streptococcus thermophilus CICC6223, lactobacillus plantarum CICC6076 and lactobacillus delbrueckii subspecies lactis CICC 6077; the gram-negative bacteria are selected from Acetobacter aceti CICC22519, Gluconobacter Thailand CGMCC1.637, Escherichia coli ATCC25922, Salmonella typhimurium ATCC14028, Salmonella typhi CMCC50071, Proteus vulgaris CMCC49027 and Shigella sonnei CMCC 51592. Adding 1.5% agar after preparing liquid culture medium, sterilizing at 121 deg.C for 15min, pouring into sterilized empty plate, coating the above experimental strains and reference strains after solidification, culturing at 30 deg.C for 36 hr, and observing the result.
5.4 Switzerland dye solution color development test: the morchella staining solution is often used as a color developing agent of gram-negative bacteria, and consists of a staining solution (solution A) and a buffer solution (solution B). The experiment selects 2.1.2.4 determined proper culture medium as the basis, the addition amount of the Reynold staining solution in 200mL culture medium is respectively set to be 0.5% (0.5 mL of the solution A and the solution B respectively), 1% (1 mL of the solution A and the solution B respectively), 2% (2 mL of the solution A and the solution B respectively), and 1.5% agar is added to prepare the plate. Acetobacter aceti CICC22519, Gluconobacter Thailand CGMCC1.637, Escherichia coli ATCC25922, Salmonella typhimurium ATCC14028, Salmonella typhi CMCC50071, Proteus vulgaris CMCC49027 and Shigella sonnei CMCC51592 are used as controls, the strains are coated on a plate, inverted and cultured at 30 ℃ for 36h, and the results are observed.
Example 6 results and analysis
5.1 optimize the experimental results, see table below.
Factor level meter
Analysis of Experimental protocols and Experimental results
from the visual analysis of the table above, it can be seen that the significance primary and secondary sequence of the 4 factors on the results is D > A > C > B, the ethanol content has the greatest influence on the experimental results, the glucose and yeast powder have the secondary influence, and the beef extract has the least influence on the experimental results. From table 2.3 it can be seen that the theoretical optimal medium formulation is A1B1C3D1, but this combination was not found in the 9 experiments designed, and the actual optimal medium formulation was A3B2C3D1, so it was also necessary to compare the theoretical and actual optimal media. The results are as follows:
Comparison of theoretical optimal culture medium and actual optimal culture medium
Compare the results with actual best medium theory is the best medium
as can be seen by comparing the two indexes of pH value and viable count, A1B1C3D1 is obviously superior to A3B2C3D1, so that the optimal culture medium of the gluconacetobacter xylinus liquefaciens can be finally determined to be A1B1C3D1, namely: 50g/L of glucose, 5g/L of beef extract, 10g/L of yeast powder, 10mL/L of ethanol and 6.8 of pH value.
5.2 culture Medium validation test results
the culture medium determined by the orthogonal design experiment was compared with the culture medium provided by the strain collection (culture conditions were the same), and the results were as follows:
Comparison result of experimental optimal culture medium and culture medium of strain preservation center
Thecomparison results of optimum culture medium and culture collection center medium
As can be seen from the above table, the best culture medium determined by the experiment is obviously better than the culture medium provided by the strain preservation center, thereby further proving the superiority of the best culture medium determined by the orthogonal design experiment. The higher pH value of the culture medium provided by the strain preservation center is mainly due to the fact that the culture medium contains calcium carbonate, and the calcium carbonate has the function of continuously neutralizing acidic substances generated in the growth and reproduction processes of the strains.
5.3 results of bile salt bacteriostasis test
effect of different amounts of bile salts on the test and Standard strains
Different bile salts added quantity for experimental strain and standard strain
Note "+" indicates colony growth on the plate and "-" indicates colony growth on the plate
As can be seen from the above table, bile salts have a significant inhibitory effect on gram-positive bacteria and cannot grow even if the added amount of bile salts is small; for gram-negative bacteria, the addition amount of bile salt has no obvious influence on the growth of the bacterial strain, and the only difference is that a less obvious transparent ring appears around the colony of the experimental bacterial strain, because the bacterial strain can utilize the bile salt in a culture medium during the growth process to decompose and utilize the bile salt around the bacterial strain. From the viewpoint of cost saving, the content of the selected bile salt is determined to be 0.1%.
5.4 results of color development test with Swiss staining solution
the Swiss stain solution is added into the chromogenic culture medium according to the proportion of 0.5 percent, 1 percent and 2 percent. The experimental result shows that the results of the culture medium with the addition of 1% of the Swiss stain solution are closer to the results of the culture medium with the addition of 2%, the transparent circles around the bacterial colony are obvious (see the attached drawing), and the transparent circles on the culture medium with the addition of 0.5% of the stain solution are not obvious, so that the addition of the Swiss stain solution in the culture medium can be determined to be 1%.
Results of addition of Swiss stain
The add results of Wright's stain
The remarks "+" indicate positive and "-" indicates negative
Since gram-positive bacteria cannot grow on the medium containing bile salts, color development experiments were performed only on CGMCC1.24 and gram-negative bacteria CICC22519, CGMCC1.637, ATCC25922, ATCC14028, CMCC50071, CMCC49027, and CMCC51592, and the results are shown in the attached drawing.
Referring to the attached drawing, the 8 strains of bacteria are subjected to streak culture in a culture medium with the addition of 1% of the Switzerland stain, so that the bacterial colony of the acetobacter gluconicum is small and purple black, and a transparent ring is formed around the acetobacter gluconicum, the acetobacter aceticus and the gluconobacter which belong to the acetobacter aceti family and belong to the acetobacter gluconobacteriaceae are similar to the experimental strains in size, but are wine red and pink in color, and no transparent ring is formed, so that 3 strains of the acetobacter gluconicum, the acetobacter aceticus and the gluconobacter gluconicum can be obviously distinguished; the other 5 kinds of bacteria are common pathogenic bacteria, the bacterial colony is obviously larger than the experimental strain, the colors are different, and no transparent ring is formed.
in conclusion, the embodiment of the invention can be seen that the research is carried out on the common acetobacter aceti in fermented milk, the acetobacter gluconicum, the acetobacter aceti and the gluconobacter are taken as target strains, the components of the culture medium are optimized, the selective culture medium is screened and researched by utilizing the characteristics of bile salt and Swiss dye solution, the method for rapidly identifying the three groups of mixed bacteria simultaneously is creatively obtained, the selective culture and the color development are combined, the identification is rapid, stable and reliable, and the method has very important scientific research and practical values.
The above description is only presented as an enabling solution for the present invention and should not be taken as a sole limitation on the solution itself.
Claims (10)
1. A selective chromogenic culture method aiming at acetobacter gluconicum, acetobacter aceti and gluconobacter in fermented milk is characterized in that:
A. Preparation of a series of culture media:
a-1, a special chromogenic medium, which is prepared as follows: 40-60g/L of glucose, 4-6g/L of beef extract, 8-12g/L of yeast powder and 8-12mL/L of ethanol, and additionally adding 0.5-1.5g/L of bile salt and 0.5-1.5mL/L of Swiss dye solution; the pH value is 6.6-7.2;
A-2, preparing a liquid basal culture medium as follows: 80-120 parts of glucose, 4-6 parts of beef extract, 4-6 parts of yeast powder, 15-25 parts of ethanol, 1000 parts of distillation by volume and pH 6.5-7.5;
B. pretreatment of fermented milk to be detected:
B-1, first-generation culture: taking fermented milk to be detected, standing to restore the fermented milk to normal temperature, and inoculating the fermented milk to a liquid basal culture medium for first-generation activation culture;
B-2, second generation culture: after the basic culture is finished, adding the culture solution into a liquid basic culture medium to perform second generation activation culture;
C. Selective chromogenic culture:
b, inoculating the culture solution obtained by the second generation culture into the special chromogenic culture medium in the step A according to the inoculation amount of 3-8% v/v, and carrying out streak culture;
D. Color development identification of three target strains:
D-1, identifying three target strains according to the following typical standards: gluconacetobacter: the colony color is purple black, and an obvious transparent ring is arranged around the colony;
D-2, Acetobacter aceti: the colony is wine red, and transparent rings are not arranged around the colony;
d-3, Gluconobacter: the colony color is pink, and no transparent ring is arranged around the colony color; and (5) finishing the identification.
2. The method for selective chromogenic culture of gluconacetobacter aceti, acetobacter aceti and gluconobacter in fermented milk according to claim 1, wherein: in the step A-1, the special chromogenic medium is configured as follows: 50g/L of glucose, 5g/L of beef extract, 10g/L of yeast powder and 10mL/L of ethanol, and additionally adding 1g/L of bile salt and 1mL/L of Swiss dye solution; the pH value is 6.8.
3. The method for selective chromogenic culture of gluconacetobacter aceti, acetobacter aceti and gluconobacter in fermented milk according to claim 1, wherein: in the step A-1, the special chromogenic medium is configured as follows: 40g/L of glucose, 4g/L of beef extract, 12g/L of yeast powder and 12mL/L of ethanol, and additionally 0.7g/L of bile salt and 1.3mL/L of Swiss dye liquor are added.
4. The method for selective chromogenic culture of gluconacetobacter aceti, acetobacter aceti and gluconobacter in fermented milk according to claim 1, wherein: in the step A-1, the special chromogenic medium is configured as follows: 60g/L glucose, 6g/L beef extract, 8g/L yeast powder and 8mL/L ethanol, and additionally adding 1.2g/L bile salt and 0.8mL/L Swiss dye solution.
5. The method for selective chromogenic culture of gluconacetobacter aceti, acetobacter aceti and gluconobacter in fermented milk according to claim 1, wherein: in the step A-2, the liquid basal medium is prepared as follows: 100 parts of glucose, 5 parts of beef extract, 5 parts of yeast powder, 20 parts of ethanol, 1000 parts of distillation by volume and pH 6.8.
6. The method for selective chromogenic culture of gluconacetobacter aceti, acetobacter aceti and gluconobacter in fermented milk according to claim 1, wherein: the specific operating parameters of step B-1 are as follows: b-1, first-generation culture: taking fermented milk to be detected, standing to restore the normal temperature, taking 1mL, inoculating into 10mL of liquid basal culture medium, and carrying out shake culture at 30 ℃ and 200r/min for 48 h.
7. the method for selective chromogenic culture of gluconacetobacter aceti, acetobacter aceti and gluconobacter in fermented milk according to claim 1, wherein: the specific operating parameters of step B-2 are as follows: b-2, second generation culture: after the basic culture is finished, 10mL of culture solution is taken out and added into 200mL of liquid basic culture medium, and shaking culture is carried out for 36h at 30 ℃ and 200 r/min.
8. the method for selective chromogenic culture of gluconacetobacter aceti, acetobacter aceti and gluconobacter in fermented milk according to claim 1, wherein: the specific operating parameters of step C are as follows: C. selective chromogenic culture: b, inoculating the culture solution obtained by the second generation culture into the special chromogenic culture medium in the step A according to the inoculation amount of 3-8% v/v, carrying out streak culture, and carrying out shake culture at 30 ℃ for 36h at 200 r/min; and detecting the pH value and the viable count of the bacteria.
9. A dedicated chromogenic medium for use in the method of claim 1.
10. Use of the dedicated chromogenic medium as claimed in claim 9 for the culture, differentiation and identification of gluconacetobacter gluconicum, acetobacter aceti and gluconobacter in fermented milk.
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Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US5464755A (en) * | 1994-04-29 | 1995-11-07 | Biolog, Inc. | Microbiological medium and method of assay |
| US5726031A (en) * | 1996-03-26 | 1998-03-10 | Rcr Scientific, Inc. | Test media and quantitative method for identification and differentiation of biological materials in a test sample |
| US20030170773A1 (en) * | 2000-06-29 | 2003-09-11 | Anna Tsoraeva | Nutritional mixture and method for early identification and count of gram-negative organisms |
| CN109439725A (en) * | 2018-11-29 | 2019-03-08 | 上海申启生物科技有限公司 | Chromogenic culture medium and its application for Escherichia coli identification |
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Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US5464755A (en) * | 1994-04-29 | 1995-11-07 | Biolog, Inc. | Microbiological medium and method of assay |
| US5726031A (en) * | 1996-03-26 | 1998-03-10 | Rcr Scientific, Inc. | Test media and quantitative method for identification and differentiation of biological materials in a test sample |
| US20030170773A1 (en) * | 2000-06-29 | 2003-09-11 | Anna Tsoraeva | Nutritional mixture and method for early identification and count of gram-negative organisms |
| CN109439725A (en) * | 2018-11-29 | 2019-03-08 | 上海申启生物科技有限公司 | Chromogenic culture medium and its application for Escherichia coli identification |
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