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Bacterial proteins (peptidoglycan) are coagulated and fixed to the glass surface.

Heat fixation


Positively charged Chromagen (nucleic acid's and certain cell wall components carry a negative charge) strongly bind to the cationic chromogen (simple staining indicates morphology and arrangement) Crystal violet, methylene blue.

Basic stains


Negatively charged chromogen will not penetrate cells because of the negative charge on the surface of bacteria. Unstained cells against colored background. Negrosin

Negative stains


Alcohol increases the porosity of the cell wall by dissolving the lipids in the outer layer and dehydrating proteins. CV – I complex can be removed from the thinner and less highly cross-linked peptidoglycan layer. Which facilitates release of the unbound CV – I complex. Grams positive cells with their thicker peptidoglycan retain CV – I.

Graham negative cells


Dehydrating effect of alcohol reduces pores so CV – I is retained. Stain is difficult to remove.

Gram-positive cells


Morphology and arrangement – basic dyes positively charged chromogen

Simple stain


Smear, mix with water, air dry, heatfix, stain, one minute, water, blot.

Also simple stain


These are metabolically inactive, highly resistant to unfavorable conditions (exhaustion of carbon source). Impervious layers called spore coats. Resist damage by heat, freezing, desiccation, radiation, and microbiological stains. May germinate later into vegetative cell. Not reproduction.Malachite green

Spore stain


# of Colonies x milliliters x dilution factor = cells per milliliter

Calculating cells per mill


Thick peptidoglycan (cell wall) lattice over plasma membrane. Peptidoglycan = Nam plus nag carbohydrates. Proteins cross-link and anchor peptidoglycan into plasma membrane.

Gram + bacterial cell wall


Cell wall consists of (outer membrane & peptidoglycan) over plasma membrane. Phospholipid/peptidoglycan sandwich.

Graham - bacteria cell wall


Obligate aerobes. Obligate anaerobes. Facultative anaerobes.

Be familiar with test tube arrangement.


Produces a characteristic change in the appearance of bacterial growth and medium surrounding the colonies which permits differentiation. Eosin – methylene blue auger

Differential media


Designed to isolate specific groups of bacteria. incorporates chemical substance that inhibits the growth of one type of bacteria while permitting the growth of another Facilitating bacterial isolation. 7.5% sodium chloride auger

Selective media


In blood agar (and enriched media) alpha hemolysis is the incomplete lysis of red blood cells with reduction of hemoglobin to methemoglobin resulting in a greenish halo around the bacterial growth. Beta hemolysis is the lysis of red blood cells with the complete distruction and use of hemoglobin resulting in a clear zone surrounding the colonies.

The difference between alpha and beta hemolysis


Fermentation is a biooxidative process not requiring oxygen in which an organic substrate serves as the final electron acceptor. Aerobic respiration is biooxidation in which molecular oxygen can serve as the final electron acceptor.

Main difference between aerobic respiration and fermentation


The bending power of light passing through air from the glass slide to the objective lens, immersion oil minimizes the amount of refracted (lost light).

Refractive index