Question

Dichotomous key showing which selective media and biochemical tests you would need if you were to test your unknown bacteria: Providencia stuartii. Please explain the procedure on how you would use biochemical tests and selective media to ID the specific bacteria.

Answer 1

Identifying the bacterium -

*The first step will be to perform a gram stain and observe the colony appearance and individual features.

If the colony stains with the gram dye, it is Gram +ve

If the colony does not stain with the gram dye, it is gram negative.

* Observe the shape of the bacteria under a microscope. According to shape, they are placed under three categories-

Cocci - spherical shape

Bacilli - rod shaped

Coccobacilli - intermediate shape like comma shaped or spiral shaped.

* Observe if the colony consists of individual bacterium, form chains or stay in squares of four bacteria oe are seen as clusters.

* Grow the given bacteria on different culture media that encourages the growth of some species and inhibits the growth of others. The sample is compared to variuos known bacteria by observing the growth pattern on the various media.

* Finally, DNA sequencing will have to be done to determine if the given sample is of known or unknown species or strain. Compare the findings with bacterial species whose genome has already been sequenced to determine if the given sample is of a new species or not.

Providencia stuarti -

* The given sample does not stain with Gram stain, that is, it is Gram -ve.

* It appears rod shaped under the microscope with flagella and is motile. Hence it is a bacillus.

* The sample grows best in Nutrient agar or Columbia agar. It does not ferment lactose.

* It is oxidase negative, which means that it does not have cytochrome c oxidase that oxidises the test reagent. The given sample may be aerobic, anaerobic or facultative.

* It tests positive for catalase which means it is aerobic or maybe facultative.

* It is urease producing as it changes the color of the medium from yellow to red. It hydrolyses urea to ammonia, which raises the pH of the medium.

Biochemical test of P stuarti

* Gram staining - negative

* Capsule - non-capsulated

* Flagella - present

* Spore producing -no

* Gas from glucose - negative

* Gelatin hydrolysis - negative

* Indole - positive

* Methyl red - positive

* Nitrate reduction - positive

* Oxidative fermentation - yes

* Citrate - positive

* Catalase - positive

* Urease - positive

* Voges proskauer - negative

These tests confirm the presence of P stuarti