some bacteria is giving the positive result as expected i.e. blue to green but some is giving color change to yellow from green. I taken care of pH while preparing the media. I am attaching the picture for better understanding of my problem.
please check the pH of the medium after incubation which turned green to yellow.
It will be acidic as BTB is yellow under acidic conditions.
Do not bother about the colour but see for any growth in the agar medium, if there is growth it indicates that the bacterium is utilizing citrate, a positive test.
Simon citrate test is employed for the differentiation of Enterobacteriaceae and aerogenes group on the basis of citrate utilization as sole carbon source. Bromothymol blue is the pH indicator dye incorporated in the medium.
Ammonium dihydrogen phosphate and sodium citrate serve as the sole nitrogen and carbon source respectively. Microorganisms also use inorganic ammonium salts as their sole nitrogen source. Utilization of citrate involves the enzyme citritase, which breaks down citrate to oxaloacetate and acetate. Oxaloacetate is further broken down to Pyruvate and Carbon dioxide. Carbon dioxide that is released subsequently reacts with water and the sodium ion in the medium to produce sodium carbonate, as well as ammonia from utilization of sodium citrate and ammonium salt respectively results in alkaline pH. This results in change of medium’s color from green to blue.
At pH 7.0 and below, lactate and acetoin are also produced. If acidic conditions prevails the slant turns yellow.
The medium should be freshly prepared because in dry conditions, changes in colour may appear even before inoculation, especially at the bottom of the slant.
Do anyone know any such type of bacteria which shows this type of results in citrate utilization?
I think your substance is pH dependent. In alkaline situation being green and under low or ~pH5 it is giving yellow color. the bug is producing lactic or some acidic product. So
I guess that a n electron charge transfer complex was formed or ph changes can be one of the explanations, but you may test othe opportunity by adding adding ecectron donor
Hai,
I am also facing this kind of problem while isolating Bacillus spp. you are not sattisfied with this method there are Several other citrate utilization media have been available. One distinguishing factor amongst the methods is that some require that the organism be able to use citrate in order to grow, while others have alternative carbon sources available. Koser's citrate medium (3), Simmons citrate medium, and Christensen's citrate medium are all commercially available.
even i am facing such problem. My enterococcus aerogenes ATCC culture which gives blue color gave yellow color on slant. this was unexpected, cause never had this happen before. even i would like to know the reason for the same.
can you please tell which organism is streaked on the slant with yellow color.
Which organism did you use?
Normally, pH range of the Bromothymol blue is green to yellow (pH 7.0 to below ) and green to blue (pH 7.0 to up).
When an organic acid such as citrate (remember Krebs cycle) is used as a carbon and energy source, alkaline carbonates and bicarbonates are produced ultimately. In addition, ammonium hydroxide is produced when the ammonium salts in the medium are used as the sole nitrogen source.
Utilization of exogenous citrate requires the presence of citrate transport proteins (permeases). Upon uptake by the cell, citrate is cleaved by citrate lyase to oxaloacetate and acetate. The oxaloacetate is then metabolized to pyruvate and CO2. The carbon dioxide that is released will subsequently react with water and the sodium ion in the medium to produce sodium carbonate, an alkaline compound that will raise the pH. In addition, ammonium hydroxide is produced when the ammonium salts in the medium are used as the sole nitrogen source. With an increase in medium pH to above 7.6, bromothymol blue changes to blue color.
Citrate negative means, trace or no growth will be visible. No color change will occur; the medium will remain the deep forest green color of the uninoculated agar. Only bacteria that can utilize citrate as the sole carbon and energy source will be able to grow on the Simmons citrate medium, thus a citrate-negative test culture will be virtually indistinguishable from an uninoculated slant.
If it gives yellow color mean, I think it was an error of the preparation of medium or organism has ability to utilize produced CO2 during krebs cycle. Yellow color due to drop of the pH below 7.0. It means, the organism should be produce acids by utilizing of C source. But in this medium whole C source is Citrate. Breakdown of citrate gives CO2, Acetate and lactate. This CO2 helps to give blue color. Think about the organism can utilize CO2. Then CO2 of the medium become decreases and pH goes below to 7 due to acidic environment. Then color change from green to yellow.
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