Discussion

Based on the result that we have obtained, P.vulgaris show positive result while the other
show negative result. The reason E.coli did not show positive result is because the culture is
kept too long. For Indole test it is to prove the ability of certain bacteria to decompose the
amino acid tryptophone to indole, which accumulate in the medium. Indole production test
is significant in the identification of Entrobacteria. P.vulgaris and E.coli break down amino
acid tryptophan and released indole. This is performed by the chain of a number of
intracellular enzymes, a enzyme generally referred to as “tryptophanase”. it is used as part
of the IMViC procedures, a test designed to duistinguish among members of family
Entrobacteriacaea. Tryptophan is an amino acid that can undergo deamination and
hydrolysis by bacteria that express tryptophanase enzyme. Indole is produce by reductive
deamination from tryptophan via the intermediate molecule indolepyruvic acid.
Tryptophanase catalyst the deamination reaction, during which the amine (-NH2) group of
tryptophan molecule is removed. Final product of the reaction are indole , pyruvic acid and
ammonium (-NH4+) group and energy. Pyridoxal phosphate is required as a coenzyme.
When indole is combined with Kovac’s reagent the solution turn from yello to cherry red.
Because amyl alcohol is not water soluble, the red coloration will form in oily layer on top of
the broth.

For Methyl Red Test, the function is to determine whether the microbe performs mixed acid
fermentation when added with glucose. Types and propotion of fermentation product
produced by anaerobic fermentation of glucose is one of yhe key taxanomic characteristics
which help to differentiate various genera of enteric bacteria. The pH at which methyl red
detect acid is considerably lower than the pH for other indicator used in bacteriologic
culture media. Thus, to produce a color change, the test organism must produced large
quantities of acid from carbohydrate substrate being used. Theoritically, only Klebsilla shows
negative result while other show positive result. Based on the result, Klebsilla show positive
result because the pH is at or below 4.4 fro the fermentation of glucose.

Voges-Proskauer is a double eponym, named after two microbiologists working at the

beginning of the 20thcentury. They first observed the red color reaction produced by
appropriate culture media after treatment with potassium hydroxide. It was later discovered
that the active product in the medium formed by bacterial metabolism is acetyl methyl
carbinol, a product of the butylenes glycol pathway. Pyruvic acid, the pivotal compound in
the fermentative degradation of glucose, is further metabolized through various metabolic
pathways, depending on the enzyme systems possessed by different bacteria. One such
pathways result in the production of acetion, a neutral-reacting end product. Organisms
such as members of the Klebsiella-Enterobacter-Hafnia-Serratia group produce acetoin as
the chief end product of glucose metabolism and form smaller quantities of mixed acids. In
the presence of atmospheric oxygen and 40% potassium hydroxide, acetoin is converted to
diacetyl, and alpha-naphthol serves as a catalyst to bring out a red complex. Based on the
result, only Klebsilla show positive result.
Citrate utlilization test is used to determine the ability of bacteria to utilize sodium citrate as
its only carbon source and inorganic (NH4H2PO4) is the sole fixed nitrogen source. As for the
result that have been obtained, all culture show positive result. The agar slant change from
green to blue color indicate that the alkaline carbonate and bicarbonate produces as a by
product of citrate catabolism increase the pH of the medium to above 7.6, making the
bromothymol blue to change from the original green color to blue.

Conclusion

As a conclusion, the objective which is to identify the characteristics of coliform, Gram

negative and rod shape bavteria have been obtained.

References

1. Madigan, M. T., Martinko, J. M., Dunlap, P. V., & Clark, D. P. (2009).

2. Brock Biology of Microorganisms, 12thed., San Francisco, CA: Pearson Benjamin

Cummings.Cappuccino, J. G. & Sherman, N. (2008).

3. Microbiology. A Laboratory Manual , 8thed.,San Francisco, CA: Pearson Benjamin

Cummings.
Result

Culture / Test Indole M-Red V.Proskauer Citrate

Pseudomonas Negative Positive Negative Positive

B. subtilis Negative Positive Negative Positive

P. vulgaris Positive Positive Negative Positive

E. coli Negative Positive Negative Positive

klebsilla Negative Positive Positive Positive