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    Home > Biochemistry News > Microbiology News > Multi-tube fermentation method to determine the E. coli in water

    Multi-tube fermentation method to determine the E. coli in water

    • Last Update: 2021-01-21
    • Source: Internet
    • Author: User
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    multi-tube fermentation method to determine the E. coli groups inwater, the purpose requirements1. Learn the multi-tube fermentation method to determine the number of E. coli groups in water.2. Understand the importance of E. coli populations in drinking water., basic principles multi-tube fermentation method includes primary (step) fermentation test, flat plate separation and re-fermentation test three parts.1. Initial (step) fermentation testthe fermentation tube is equipped with lactose proteinliquid
    culture
    -base, and upside down a Dehan's small sleeve. Lactose can play a selective role, because many bacteria can not ferment lactose, and the E. coli group can ferment lactose and produce acid gas. In order to facilitate the observation of the acid production of bacteria,
    fleter
    in addition of bromophenol purple as a pH indicator, bacteria produce acid, the culture base from the original purple to yellow. Bromophenol purple also inhibits the growth of other bacteria such as bud bacteria.Water sample inoculated in the fermentation tube, cultured at 37 degrees C, within 24 hours in the small casing gas formation, and the culture base turbidity, color changes, indicating the presence of colium bacteria in the water, positive results, but there are other types of bacteria in this condition may also produce gas; In the case of low quantities, gas production may also be delayed until 48 hours later, which should be considered suspicious results, therefore, both results need to continue to do the following two parts of the experiment to determine whether it is E. coli. After 48 hours, the negative results were still not gas produced.2. Tablet separationtablet mediums generally use red sodium sulphate
    agar
    (Farto's medium, Endo's medium) or eosin methylene blueagar (EMB agar), the former containing alkaline redness Dye, in this as an indicator, it can be decolored by sodium sulphate in the culture base, so that the culture base is pale pink, E. coli group fermented lactose produced by the acid and acetaldehyde and redness reaction, forming a crimson complex, so that the E. coli colony colony into a metallic luster of crimson. Sodium sulphate also inhibits the growth of other bacteria. Yihong Mei blue agar plate contains Yihong and Mei blue dye, in this also as an indicator, E. coli fermented lactose caused acidic environment, the two dyes are combined into a compound, so that the E. coli group produces similar to the Far Vine culture base, with a core, metallic luster of dark purple (purple purple of dragon bile purple) colonies. The initial fermentation tube within 24 hours of acid production gas and 48 hours of acid production gas are required to draw a line on the above plate to separate the bacteria.3. Referment test more thanE. coli group positive colonies, smeared with Glomin-negative Bacillus aureus, through this test and further confirmed. The principle is the same as the initial fermentation test, after 24 hours of culture and gas production, and finally determined to be positive results of E. coli.3. Equipmentlactose protein fermentation tube (with inverted small casing), three times concentrated lactose proteinfermentation tube (bottle) (with inverted small sleeve), Irotty Blue agar plate, sterilized water;glass slides, sterilized glass plug empty bottle, sterilized
    straw
    , sterilized
    test tube
    etc., the operation step 1. Water-like2. Tap water inspection(1) initial (step) fermentation test in 2 containing 50 ml of triple concentrated lactose protein ferment
    smosting
    , each added 100 ml of water samples. In 10 fermentation tubes containing 5 mlconcentrated lactose protein, 10 ml of water samples were added (as shown X.IV.-1). After mixing, 24 hours of culture at 37 degrees C and 48 hours after 24 hours of non-production.(2) flat plate separation after 24 hours of culture, the production of acid gas and 48 hours of acid production of fermentation tube (bottle), respectively, line inoculated on the Yihongmei blue agar plate, and then cultured at 37 degrees C 18-24 hours, will meet the following characteristics of a small part of the bacterios, smear, Grady dyeing, mirror examination.(a) dark purple and black with a metallic sheen.(b) Purple black, without or slightly metallic gloss.(c) pale fuchsia and darker center color.(3) Referment test by smearing, dyeing, mirror testing, such as Gradococcal-negative spore-free bacteria, then pick another part of the colony, re-inoculated in the ordinary concentration of lactose proteinconfirmed the presence of E. coli, then according to the number of positive tubes (bottles) of the initial fermentation test to check table X.IV.-2, that is, the number of E. coli groups.
    3. Inspections of pool water, river water, or lake water(1) dilute water samples into 10-1 and 10-2.(2) absorbed 1 ml of diluted water samples and 1 ml of raw water samples, respectively, into a fermentation tube containing 10 mlconcentration of lactose protein. Another 10 ml and 100 ml of raw water samples were injected into test tubes (bottles containing 5 ml and 50 ml of triple concentrated lactose proteinfermentation fluid, respectively. (3) the following steps with the above-mentioned tap water plate separation and re-fermentation test. (4) will be 100, 10, 1, 0. 1 (10-1) ml water-like fermentation tube results check the table XI.V.-3, will be 10, 1, 0. 1(10-1 )、0. 01 (10-2) ml water sample fermentation tube results check table XI.V.-4, that is, the number of colium sterilization per liter of water sample 5, experimental report 1. Results (1) tap water 100 ml of water sample positive tube number? number of positive tubes with 10ml water samples? the number of E. coli groups per liter of water samples on the table XI.V.-2? (2) pool water, river water or lake water positive results remember ""; negative results remember "-". the number of E. coli colonies per liter of water samples on the table XI.V.-3? the number of E. coli groups per litre of water samples on the table XI.V.-4?
    2. Question: (1) What is the definition of coli group? (2) Why should E. coli be chosen as an indicator of contamination of intestinal pathogens as a water source? (3) EMB culturer contains some of the main ingredients? What role do they play in examining E. coli? (4) have been checked for water samples that meet drinking standards?
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