1. Lab. No. 3

3. Gram’s +ve Bacilli Spore forming Non spore forming AerobicAnaerobic Bacillus Clostridium Corynebacterium

4. Most Important Species: Most Important Species:  B. anthracis causes anthrax.  B. cereus causes food poisoning.

5. Identification of Bacillus Species (I) Morphology (microscopical examination): A- Gram Stain:  Gram’s +ve spore- forming bacilli, arranged in chains.  Diameter of spores equal to that of the vegetative cells.

6. B- Spore Stain:  It has oval central spores.  Using the Spore staining technique (Malachite green & safranin), the spores appear green while the vegetative cells appear red.

7. (II) Cultural charcteristics: No special requirements for growth → grow on simple nutrient media. Growth on Blood Agar:Growth on Blood Agar:  Bacillus species grow well on blood agar showing a double zone of hemolysis Except B. anthracis, Except B. anthracis, which grows well on blood agar without any hemolytic effect.

8. (III) Biochemical reactions: A- Catalase test:  All bacillus species are catalase +ve.

9. B- Starch Hydrolysis Test: Principle: Starch amylase enzyme glucose Blue colour No colour Procedure: 1. Inoculate starch agar plate with the test organism. 2. Incubate the plate at 35 o C for 24 hrs. 3. Flood the plate with iodine solution. I2I2 I2I2

10. Results: Amylase activity is indicated by a clear zone around the growth while the rest of the plate gives blue color after addition of iodine solution.