Capsule Stain- Differentiated between capsule, cell and background Capsule is a thick structured layer that protects the cell from phagocytes. Primary Stain- Congo Red Counter Stain- Crystal Violet Biochemical Tests- presence or absence of specific enzymes helps us to identify bacteria -used to evaluate metabolic capabilities of specific bacteria. 1 . Catalane Test- Enzyme: Catalane Hydrogen Peroxide Product: Oxygen (+)- Catalane… No Bubbles (-)- No Catalane 2.
Identification Test- Enzyme: Nitrate Reeducates Substrate: Nitrate Product: Nitrogen Gas Bubbles in Durham Tube (+)- Nitrate Reeducates Substrate: Reagent: Hydrogen Peroxide Medium: TTS Bubbles Reagent: None Medium: Nitrate Broth No Bubbles (-)- No Nitrate Reeducates 3. Hydrogen Sulfide (HAS)Production- Enzyme: Cytosine Decreases HAS) Black Color (+)- Cytosine declarers Substrate: Cytosine Product: Hydrogen Sulfide Medium: SIMI (Sulfur Indolent Motility) Deep No Color Change (-) – No Cytosine decreases 4.
Indolent Production Test- Enzyme: Transposable Substrate: Thyrotrophic Product: Indolent Indolent Motility) Deep Red Color (+)- Transposable Reagent: Kavas Medium: SIMI (Sulfur No Color Change (-) – No Transposable 5. Methyl Red Test- Enzyme: Multiple Enzymes Substrate: Glucose Product: Many Acids Reagent: Methyl Red Medium: Methyl Red Broth (MRS.) Red Color (+) – More Acidic and has Multiple Enzymes that breakdown sugar… Yellow Color (-) – Fewer Acids and does NOT have Enzymes that breakdown sugar 6.
Oxidation-Fermentation Test – Determines Oxygen requirements of sugar or Glucose Enzyme: Not Specified tort the breakdown Substrate: Dextrose Medium: Product: arroba or anaerobe O-F Dextrose (Green Tube) Inert- Both Tubes Remain Green Obligate Arroba- Tube with Oil remains Green and Tube without Oil Turns Yellow Facultative Anaerobe- Both Tubes Turn Yellow Microbial Control 1 . Active Ingredients of Surface Control Agents -“know what the bottles tell you”. 2.
Kirby- Bauer Disk Diffusion Test- used to select appropriate antibiotic shows Antibiotics-bacterial susceptibility Mueller-Hint Agar Zone of Inhibition- Area around the antibiotic where the bacteria is sensitive to drug Concentration Effectiveness- The concentration of antibiotic that diffuses into the media decreases with increasing distance from the source. Therefore, the more sensitive the bacteria are to a given antibiotic, the larger the clear bacteria-free zone that forms around the disk containing that antibiotic.
Evidence of Adaptive Resistance- Few colonies grow in the zone of inhibition show some cells have developed resistance to the drug. Spectrum of Activity- Inoculate plates with several antibiotics. The antibiotics that inhibit the growth of the most bacteria is broad spectrum, the least is narrow spectrum. Staining Procedures- Gram Stain- 1. Inoculate bacteria- Smear it into a drop of water- Let it Dry- Heat Fix it 2. Apply Primary Stain (Crystal Violet)- Let sit 60 Seconds- Rinse 3.
Apply Binding Agent(Mordant)- Let sit 60 Seconds- Rinse 4. Apply Decontrolling Agent (Ethanol)- Sit 30 Seconds or Less- Rinse 5. Apply the Counter Stain (Seafaring)- Sit 60 Seconds- Blot on Bibulous Paper 6. Observe on High Power and Oil Immersion Spore Stain- 1. Inoculate bacteria- Smear it into a drop of water- Let it Dry- Heat Fix it 2. Place on Hot Plate for 10 Minutes (Malachite Green)- Let sit 60 Seconds- Rinse 3. Apply Primary Stain 4.
Apply Counter Stain (Seafaring)- Sit 60 Seconds- Rinse- Blot on Bibulous Paper Capsule Stain- 2. Apply Primary Stain (Congo Red)- Let sit 60 Seconds- Rinse 3. Apply Counter Stain (Crystal Violet)- Sit 60 Seconds- Rinse- Blot on Bibulous Paper Proper Use of Report Form- Observation- Color Change, Bubbling, etc. Positive, Aerobic, Anaerobic or Inert is the enzyme present or not? Results- Negative, Interpretation- Enzyme Explanation-