The document discusses various biochemical tests used to identify bacteria, including sugar fermentation, catalase, coagulase, and oxidase tests. Sugar fermentation tests for gas production using Andrade's indicator in sugar media. The catalase test detects the production of oxygen bubbles from hydrogen peroxide, indicating a positive result. The coagulase test shows clot formation in plasma due to coagulase enzyme activity. The oxidase test turns purple within 1-2 minutes due to cytochrome oxidase activity. Biochemical reactions require pure bacterial cultures.

3. Bacteria vary in their metabolic &
enzymatic activities.
Used in identification of different
genera & species of bacteria.
Biochemical reactions are done on
bacteria grown in pure culture.

4. Biochemical Reactions
1. Sugar fermentation
2. Oxidase test
3. Catalase test
4. Coagulase test
5. API Test

6. 1. Sugar fermentation
Sugar media are composed of:
Peptone water
+ 1% test sugar
+ Andrade’s indicator
+ A small inverted tube (Durham’s tube) .

7. Sugar fermentation test
uninoculated
Not fermented
fermented with A only
fermented with A & G

8. 1. Sugar fermentation
Sugar fermentation can be indicated
by change colour of the medium
from yellow to red.
Glucose
+ve
Lactose
+ve
maltose
+ve
mannite
+ve
Sucrose
- ve

10. Catalase Test
Principle:
H2O2
Procedure:
1
2
Catalase enzyme
3
H2O2
H2 O
+
O2
Air bubbles

11. Results:
Catalase Test
Positive test: rapid appearance of gas bubbles.
Catalase +ve
Catalase –ve

12. Catalase Test

13. Coagulase Test
Principle:
Fibrinogen
Plasma
Procedure:
2
Coagulase enzyme
Fibrin
Visible Clot
1
1 ml rabbit plasma
3
Place at 37oC, observe for formation of visible clot for up to 4 hrs.

14. Coagulase Test
Coagulase +ve
Coagulase -ve

15. Oxidase Test
Principle:
Tetramethyl p-phenylene diamine hydrochloride
(oxidase reagent)
colourless
Cytochrome oxidase enzyme
Indophenol
(Purple colour)
within 1-2 min

16. Oxidase Test

17. API (Analytical Profile Index)

18. Incubation
Interpretation acc. to given charts

19. Bacterial
Typing

20. Classification of a certain species of bacteria into Types.
Importance: Epidemiological investigation (e.g. food
poisoning)
Examples of Typing methods:
 bacteriophage typing
 bio-typing
 serotyping
 plasmid analysis
 + others

21. Bacteriophage Typing
Bacterial strain on surface of media + drops of different phages
homogenous bacterial growth
+ areas of lysis (= susceptibility)

22. Questions

23. 1- In sugar fermentation, Andrade's indicator is added
to the sugar media to test for gas production:
( ) True
( ) False
2- In the coagulase test, coagulation of plasma occurs
in positive cases:
( ) True
( ) False
3- Biochemical reactions must be done on a pure
culture of the organism to be identified:
( ) True
( ) False

24. MCQ
4-Catalase test is:
a) negative if oxygen bubbles are released
b) positive if a visible clot is formed
c) done by adding a drop of H2O2
d) done to test for the presence of urease enzyme
5- Oxidase test is:
a) done using catalase reagent
b) positive when a deep purple color is produced
c) positive after a few hours
d) negative when a yellow color is produced

25. Thank you