Neisseria + AFB

Neisseria Gram negative cocci - diplococci (bean shaped)

Biochemical reaction Catalase +ve Oxidase +ve

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

culture Chocolate medium + 5% CO2 (capnophilic) Incubation: At 37ºC for 24-48 hrs. Colonial morphology: small, gray, translucent and raised.

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

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

AFB

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

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)

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

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

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

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