Bio-chemical identification of bacteria

various
microbial
show great differences in metabolic types. This difference is even more pronounced due to the unique single-celled primary nuclear biological properties of bacteria. Different bacteria have different enzyme systems, so different bacteria decomposition, the use of sugars, fats and protein substances have different ability, their metabolic spectrum and decomposition products of substances are also different, the use of this characteristic to establish bacteria
national
reaction, in the classification of strains has the main significance.1. To understandprinciples of physiological, bio-chemical and experimental reactions commonly used in bacterial identification.
2. Master techniques and methods for determining the physiological, bio-chemical response of bacteria.Experimental Principles Because of their different enzyme systems, bacteria can use different substrates, or, although they can use the same substrates, produce different metabolites, so they can use a variety of physiological and bio-chemical reactions to identify bacteria.
reagents
and equipment 1. Bacillus E. coli, Enterobacter aerogenes, Proteus vulgaris, Bacillus subtilis
2.
Culture
bicular glucose water
culture base
, protein water culture base, sugar fermentation culture (glucose, lactose or sucrose)
3. Reagents 40% NaOH solution, creatinate, methyl red reagents, argastin reagents, ether, 1.6% bromometrol purple indicator.
4. Appliance ultra-clean work table,
string temperature
culture box, autocultension pot,
tit tube
, pipetting tube, Dushi small casing.
5. Process Sugar fermentation test → V-P → and methyl red test → sterilite test.Experimental Methods and Steps 1. Sugar Fermentation Experiment
(1) Test tube labeling test tubes containing different sugars (e.g. glucose, sucrose, lactose, etc.) fermentation culture, etc., and labeling the bacteria cultured as needed (e.g. E. coli, E. coli, common deformed bacteria, etc.) and blank control).
(2) Inoculation culture with sterile operation to inoculated a small amount of moss to the corresponding test tubes above, not inoculated in the blank control, placed 37 degrees C
thermostat box
culture, respectively, in culture 24h, 48h and 72h observation results.
(3) Observation record Compared with blank control after culture, if the inoculation culture remains the original color, the reaction result is negative, indicating that the bacteria can not use the sugar, recorded with "-" indicated; The positive test of air bubbles in the duchy tube in the culture fluid indicates that the bacteria break down the sugar to produce acid and gas, and the record is expressed with a "plus", and if there are no bubbles in the duct as a negative reaction, the record is represented by "-".
2. V.P.
(1) Mark the test tube take the test tube containing glucoseculture, and mark the cultured bacteria as needed.
(2) Inoculation culture with sterile operation to inoculation of a small amount of moss to the corresponding test tube above, blank control tube is not inoculated, placed in a temperature box of 37 degrees C, culture 24 to 48h.
(3) observation record Take out the above test tube and oscillate 2min. Another 5 empty test tube corresponding marker bacteria name, respectively, add 3 to 5 ml or more of the corresponding tube culture, and then add 40% NaOH solution 10 to 20 drops, and add about 0.5 to 1 mg trace crea acid, oscillating test tube, so that oxygen in the air Dissolved, placed in a 37-degree C thermostat insulation 15 to 30min, if the culture liquid is red, recorded as V.P. test positive (indicated by "plus"), if not red, recorded as V.P. test negative reaction (indicated by "-").
: The culture left in the original test tube is used as a methyl red test.
3. The methyl red test (M.R. test)
takes a test tube containing glucose protein water culture fluid, inoculates the bacteria that need to be cultured, after night culture, adds 2 to 3 drops of methyl red indicator, pays attention to the addition along the pipe wall, carefully observes the upper layer of the culture fluid, if the upper layer of the culture liquid turns red, it is positive;
4. Test tube
(1) Test tube mark take a test tube containing proteinwater culture fluid, and mark the bacteria cultured as needed.
(2) Inoculation culture with sterile operation to inoculated a small amount of moss to the corresponding test tube above, the 5th tube as a blank control not inoculated, placed 37 degrees C thermostat box culture 24 to 48h.
(3) Observation record Add 5 to 10 drops of lychee reagents to the culture solution, so that the reagent floats on the surface of the culture, forming two layers, the observation results: if there is radon, ether layer appears rose red, this is a positive test for radon, otherwise it is negative, positive with "plus", negative "-" indicated.