alkalinizes the medium, and the pH shift is identified by the color
change of phenol red from light orange at pH 6.8 to magenta
(pink) at pH 8.1. Immediate urease-positive organisms turn the
entire medium pink within 24 h.
3.9.1 Materials Required
l
Media: Urea agar/brain–heart infusion broth.
l
Organisms: Bacteria Xenorhabdus and Photorhabdus.
3.9.2 Procedure
1. Take a portion of well-isolated colony or few drops from an
overnight brain–heart infusion broth and streak the urea agar.
2. Close the cap loosely and incubate the tube at temperature of
37
C in ambient air for 48 h to 7 days.
3. Visualize for the development of a pink color for as long as
7 days.
3.9.3 Expected Result
Positive: Presence of pink color.
Negative: No color change.
3.10 Lactose
Fermentation Test
(Acid Gas Production)
3.10.1 Materials
Required
l
Media: Lactose broth.
l
Others: Durham’s tube.
l
Organisms: Bacteria Xenorhabdus and Photorhabdus.
3.10.2 Procedure
1. Evaluate the bacterial culture for acid and gas generation by
inoculating 5 ml of the sterile lactose broth in test tubes containing Durham’s tube.
2. Incubate the test tubes for 7 days at 30
C.
3. Accretion of gas in these Durham’s tube is considered positive
for gas production and change in color of the medium is taken
as positive for acid production.
3.10.3 Expected Result
Acid Production
Positive: Succeeding incubation, the liquid in the tube turns yellow
(indicated by the change in the color of the phenol red indicator). It specifies that there is drop in the pH due to the generation of the acid by the fermentation of the carbohydrate (sugar)
present in the media.
Negative: The tube comprising medium will persist red, indicating
the bacteria cannot ferment that specific carbohydrate source
present in the media.
Biochemical Characterization
127