1. Neisseria + AFB

2. Neisseria
Gram negative cocci
- diplococci (bean shaped)

3. Biochemical reaction
Catalase +ve
Oxidase +ve

4. Oxidase test
The Oxidase Test is used to identify bacteria containing the respiratory enzyme cytochrome c oxidase
TMPD

5. culture Chocolate medium + 5% CO2 (capnophilic)
Incubation: At 37ºC for hrs.
Colonial morphology: small, gray, translucent and raised.

6. Neisseria spp. : Carbohydrate Utilization test
only one carbohydrate source in the medium
Phenol red as pH indicator
Differentiate between N.gonorrhea & N.menengitidies

7. Neisseria spp. : Carbohydrate Utilization test
N. Gonorrhea
N. Meningitides
Glucose
Maltose
Lactose
N. Gonorrhea + -
N. Meningitides

8. AFB

9. Gram stain Gives positive or negative reaction
It does not applied to all bacteria
ex. Acid Fast Bacilli

10. Acid Fast Bacilli (AFB)
Why Gram stain is not used ?
- AFB cell wall has ~ 60% lipid (waxy). So, it makes gram stains impermeable.
Ex. Mycobacteria spp.
Mycobacterium tuberculosis (MTB)

11. AFB stain Two procedures : 1- Light microscope
- Ziehl-Neelsen and Kinyoun methods
2- Fluorescent microscope
- Fluorochrome method

12. Ziehl-Neelsen stain Procedure :-
1- Primary stain: carbol fuchsin , 5 minutes( basic fuchsin +phenol)
2- decolorizer : acid alcohol , 1-2 minutes
3- counterstain: methylene blue, 1 minutes

15. Ziehl-Neelsen (ZN)staining
Hot method by using heat
Cold method ( no heat ) is called Kinyoun
- increasing concentration of basic fuchsin and phenol

16. Auramine-Rhodamine Staining
Fluorescent staining (use UV light )
More sensitive than ZN staining
Auramine-Rhodamine binds to cell wall ( mycolic acid)
Procedure :-
1- primary stain: Auramine-Rhodamine, 15 minutes
2- Decolorizer : Acid Alcohol , 2 minutes
3- Counterstain : Potassium Permanganate , 3 minutes