稱取本品 41.5g,加熱溶解于1000ml 蒸餾水中,煮沸不要超過2 分鐘。取適宜稀釋度樣品液1ml,加入到無菌平皿中心,將冷至 45 ± 0.5℃ 的 VRBA 10-15ml傾注于平皿中。小心旋轉平皿將培養基與樣液充分混勻。凝固后，再加 3-4ml VRBA 覆蓋 平板表層 ,翻轉平板 ,置于 36 ± 1℃培養 18-24 小時。無需高壓滅菌。臨用時制備,不得超過3 小時。
稱取本品 5.7g,加熱攪拌溶解于 100ml 蒸餾水中,不要過分加熱 , 冷至 50℃左右時,傾入無菌平皿,部分開蓋干燥 2h,然后蓋上，備用。無需高壓滅菌。在 24小時內使用。
稱取本品 109.0g,加入 1000ml 蒸餾水中,加熱溶解并不停攪拌煮沸 1 分鐘。121℃高壓滅菌15 分鐘，冷至 45-50℃左右時,傾入無菌平皿。
稱取本品 46.0g,加入 1000ml 蒸餾水中,115℃高壓滅菌 20分鐘,備用。滅菌好的培養基僅可重復加熱熔化一次。
稱取本品 32.3g,煮沸溶解于1000ml純化水中,分裝三角瓶,121℃高壓滅菌 15 分鐘, 備用。
稱取本品 48g,加熱溶解于1000ml 蒸餾水中,分裝三角瓶,每瓶 100ml,121℃高壓滅菌 15 分鐘,冷至 50-55℃時,每瓶加入 1 支復達欣(3mg),混勻,傾入無菌平皿備用。
Colony characteristics of common microorganisms on common selective media
A brief introduction to the medium of culture:
The culture medium is artificial feed for the growth and maintenance of microorganism, plant and animal tissue. It usually contains carbohydrates, nitrogen containing substances, inorganic salts (including trace elements), vitamins and water. Some media also contain antibiotics and pigments, which are used for the cultivation and identification of various microorganisms.
Basic medium contains peptone, beef dip powder, yeast extract, sodium chloride, agar and other basic nutrients.
Some selections and identification media are different according to their uses. They include bacteriostat, indicator, blood, sugar and other reagents to facilitate the growth and identification of the needed bacteria.
Some media that need to grow and propagate bacteria are mostly amino acids, nucleotides, inorganic salts, growth factors, etc.
Agar is a kind of alginate extracted from gelatinous or hedgerow (red algae). It is a long chain polysaccharide compound composed of agarose and agarose. The advantage is that it can not be decomposed and utilized by bacteria, and the gel strength is stable at the temperature of bacterial culture.
Beef extract is a nutritive substance obtained from fresh beef, which is extracted by digestion and filtration. It contains water soluble substances of creatine, creatinine, polypeptide, amino acids, nucleotides, organic acids, minerals and vitamins. The main role is to supplement peptone and other nitrogen sources of undernourishment.
Yeast powder is rich in B vitamins, organic nitrogen and carbohydrates.
Bile salt is a sodium cholic acid, a mixture extracted from the bile of animals, containing cholic acid, conjugated cholic acid and bile alcohol. Bile salts are used as selective inhibitors in the culture medium, which mainly inhibit the growth of Gram-positive bacteria.
Culture medium classification:
According to the nutrient substance, it can be divided into:
1. Natural medium: a kind of culture medium made from animal, plant or microorganism, including its extract. Such as beef paste peptone medium, wort medium, etc.
2. The combination medium, also known as the synthetic medium or the comprehensive medium, is a kind of medium prepared with a variety of high purity chemical reagents after the precise design of the nutritional requirements of microbes. Such as glucose ammonium salt medium, starch nitrate medium and so on.
3, half combination medium: a kind of medium which is mainly prepared with chemical reagents, plus some or some natural ingredients. For example, the potato sucrose medium.
According to the physical character:
1. Liquid medium: a liquid medium.
2. Solid medium: a medium with solid appearance. According to the nature, it is divided into. Solidified medium, non reversible curing medium, natural solid medium and filter membrane.
3. Semisolid medium: a semisolid medium made up of a small amount of coagulant in a liquid medium.
4, dehydrating medium: also known as prefabricated dry medium, refers to contain all ingredients except water for commercial media.
According to the function, it is divided into:
1, selective medium: a kind of medium based on the special nutritional requirements of a microorganism or its resistance to some chemical and physical factors. It has the function of making the inferior bacteria in the mixed bacteria into the dominant bacteria, and is widely used in the field of strain screening.
2, differential medium: a class of indicators that add color reaction to the colorless metabolites of the target bacteria in the composition, so as to find the medium of the colony of the target bacteria easily from the approximate colony only with the naked eye color. For example, eosinylene lactose medium (EMB).
3. Transport medium
4. Preservation medium
5, resuscitation medium
6, increase the culture medium
7, identification medium
8, multipurpose medium
Several ISO, GB, SN common medium and color medium culture medium formula and colony characteristics analysis:
1. Colony characteristics of Escherichia coli on crystal violet Neutral Red Bile Glucose Agar (VRBGA).
Peptone and yeast powder provide carbon and nitrogen sources and trace elements; lactose is fermentable sugar; sodium chloride can maintain a balanced osmotic pressure; bile salts and crystal violet inhibit Gram-positive bacteria, especially gram positive bacilli and Streptococcus faecalis; neutral red is a pH indicator.
Take this product 41.5g and dissolve it in 1000ml distilled water and boil it for no more than 2 minutes. The suitable dilution sample 1ml was added to the sterile dish center, and the VRBA 10-15ml, which was cold to 45 + 0.5 C, was poured into the pan. Carefully rotate the dish and mix the medium with the sample. After solidification, 3-4ml VRBA was used to cover the surface of the flat plate, and the plate was placed at 36 + 1 degrees for 18-24 hours. No autoclave is needed. The preparation shall not exceed 3 hours at the time of use.
Crystal violet neutral red gall salt glucose agar (VRBGA) is also called intestinal bacteria count agar. According to the SN and FDA BAM methods, Enterobacteriaceae bacteria can produce pink to red colonies with or without precipitation ring on this medium, with a diameter of 0.5mm or larger.
Two. Colony characteristics of Salmonella on xylose lysine deoxycholate agar (XLD)
Xylose lysine deoxycholic acid agar (XLD):
Yeast extract provides nitrogen sources, vitamins and growth factors; sodium chloride maintains a balanced osmotic pressure; wood ponds, lactose and sucrose are fermentable sugars; acids make the phenol red indicator yellow; sodium deoxycholate inhibits Gram-positive bacteria, but does not affect the growth of Salmonella; sodium thiosulfate can be reduced to hydrogen sulfide by certain bacteria. The ferric salt of ferric ammonium citrate is used to form black iron sulfide; agar is the coagulant of the medium; phenol red is pH indicator.
Call this product 5.7g, heated and dissolve in the 100ml distilled water, do not overheat, cold to about 50 degrees, dip into the sterile flat, partially open the dry 2h, and then cover, spare. No autoclave is needed. It is used in 24 hours.
The yeast powder in the xylose lysine deoxycholic acid agar (XLD) provides vitamins and cofactors for the growth of bacteria, and xylose, lactose and sucrose as a fermentable carbon source. In addition to Shigella, most other Enterobacteriaceae ferment xylose, and lysine is added to identify Salmonella. The acid environment of xylose fermentation is beneficial to the production of decarboxylase by the formation of acid environment, which makes the increase of pH in the medium to alkaline, but can be neutralized by a large amount of acid produced by the fermentation of lactose and sucrose by other bacteria. Under alkaline conditions, the reaction of sodium thiosulfate and ammonium citrate with Salmonella thiosulphic reaction causes colony color. It is black, and the reaction under acidic condition is inhibited. Phenol red as an indicator, NaCl maintains osmotic pressure of the medium, sodium deoxycholate inhibits the growth of common gram-positive bacteria, and inhibits the growth of E. coli. According to the ISO and GB and FDA BAM methods, most of Salmonella in xylose lysine deoxycholic acid agar (XLD) can form a colony of red with black heart, about 2mm, smooth, moist and tidy, Arizona fermented lactose, and can form a yellow colony with black heart.
Three. Staphylococcus aureus colonies on mannitol high salt agar:
Mannitol high salt agar:
Peptone and beef powder provide nitrogen sources, vitamins and growth factors; D - mannitol is fermentable sugar; sodium chloride maintains a balanced osmotic pressure; agar is the coagulant of culture medium; phenol red is an indicator of pH.
This product is called 109.0g, added to 1000ml distilled water, heated and dissolved, stirring and boiling for 1 minutes. 121 degree high pressure sterilization for 15 minutes, cold to 45-50 degrees Celsius, pour into sterile dish.
NaCl in mannitol high salt agar can inhibit non salty non staphylococcal growth, D- mannitol as a carbon source, phenol red indicator, coagulase positive bacteria can make use of D- mannitol to produce acid to make red from red to yellow.
Four. Colony characteristics of Clostridium perfringens on Blood Agar Base in Columbia:
Columbia Blood Agar Base:
The casein trypsin digests, meat and stomach enzyme digests, cardiac pancreatin digests and yeast extract powder provide carbon and nitrogen sources, vitamins and amino acids; starch can promote the growth of Neisseria bacteria and enhance the hemolytic characteristics of streptococcus; sodium chloride maintains a balanced osmotic pressure; agar is the coagulant of culture medium.
39 grams of this product, heat dissolved in 1000ml distilled water, separate, 121 degrees centigrade sterilization 15 minutes, cold to 45-50 degrees, add the equivalent of 20mg gentamicin, sterile gentamicin sulfate, mixed, into a sterile flat dish.
Most of the Clostridium perfringens have double hemolytic ring on the blood plate, and the inner ring is completely hemolytic. It is due to the effect of theta toxin. The incomplete hemolysis of the outer ring is caused by alpha toxin.
Five. Colony characteristics of Candida albicans on potato dextrose agar medium:
Potato glucose agar medium:
Potato leach powder helps the growth of various molds; glucose provides energy; agar is the coagulant of the medium.
The product is called 46.0g, added to 1000ml distilled water, and sterilized at 115 C for 20 minutes. The medium with good sterilization can be reheated and melted once.
The colony characteristics of Candida albicans on the potato glucose agar medium were milky white colonies, and were protruding. On the potato glucose agar medium with chloramphenicol, the colony surface was coloured with milk oil.
Six. The colony characteristics of Pseudomonas aeruginosa on sixteen alkane three methyl ammonium bromide agar:
Sixteen alkanes and three methyl ammonium bromide agar:
Peptone and beef extract provide carbon and nitrogen sources, vitamins and growth factors; sodium chloride maintains a balanced osmotic pressure; sixteen alkanes and three methyl ammonium bromide are selective bacteriostat, which can release nitrogen and phosphorus in bacterial cells and inhibit bacteria from Pseudomonas aeruginosa as a quaternary ammonium salt decontamination agent; agar is a coagulant of culture medium.
This product is called 32.3g, boiled in 1000ml purified water, separated into triangular bottles, and sterilized at 121 C for 15 minutes.
Pseudomonas aeruginosa was on sixteen alkane three methyl ammonium bromide agar, the colony of Pseudomonas aeruginosa flattened and amorphous, spread to the periphery or slightly spread, the colony was gray white, and there were water soluble pigments around it.
Seven. Single colony Lester's colony characteristics on Lester selective agar.
Selective agar of Lester bacteria:
Tryptone, polypeptone and beef powder provide nitrogen sources, vitamins and growth factors; glucose provides carbon source; sodium chloride maintains a balanced osmotic pressure; sodium hydrogen phosphate two is a buffer; glycine, lithium chloride, benzyl alcohol and dahdahin can inhibit some gram-positive and Gram-negative bacteria other than Listeria. Agar is the coagulant of the medium.
The product was called 48g. The heating was dissolved in 1000ml distilled water and divided into a triangulation. Each bottle was sterilized at 121 degrees centigrade for 15 minutes. When cold to 50-55, each bottle was added to 1 Fu Daxin (3mg).
On the selective agar plate of Lester's bacteria, mononuclear cells grew on mononuclear cells, and Lester grew into blue colonies. There was an opaque ring around the colony of List Rand.