Lab Midterm

What are the four objective lenses on a light microscope and their magnifications?

Scanning lens (4x), Low power (10x), High power (40x), Oil immersion (100x)

What dye is commonly used for simple staining?

Methylene blue, crystal violet, or safranin.

What is the purpose of simple staining?

To visualize bacterial morphology using a single dye

What is the purpose of using a counterstain in the Gram stain procedure?

To visualize Gram-negative bacteria after decolorization.

What color do Gram-positive and Gram-negative bacteria appear after staining?

Gram-positive: Purple (think peptidglycan); Gram-negative: Pink (thin peptidglycan)

What is a negative stain primarily used for?

To stain the background and not the cells, preserving cell shape and size.

What is the main reagent used in a capsule stain?

Congo red or India ink

What characteristic indicates a positive capsule stain?

A clear halo around the cells.

What differentiates endospore positive and negative results in bacteria?

Positive shows green (Malachite green) endospores inside red cells, while negative shows no green spores.

What is used in the acid-fast stain to identify Mycobacterium?

Carbol fuchsin.

What is the key characteristic of acid-fast bacteria?

They have waxy cell walls and retain carbol fuchsin after decolorization

What does Tryptic soy agar/broth serve as in microbiology?

A general purpose growth medium.

What indicator is used in Fluid Thioglycollate Broth to show oxygen presence?

Resazurin (Pink = oxidized, Colorless = reduced oxygen).

Where do obligate aerobes grow in Fluid Thioglycollate Broth?

At the top, where oxygen is present.

Where do obligate anaerobes grow in Fluid Thioglycollate Broth?

At the bottom, where no oxygen is present.

Where do microaerophiles grow in Fluid Thioglycollate Broth?

Slightly below the surface – They require low levels of oxygen but can't tolerate high amounts.

How do aerotolerant anaerobes grow in Fluid Thioglycollate Broth?

Evenly throughout the tube – They do not use oxygen but can survive in its presence.

What is the purpose of Mannitol Salt Agar (MSA)?

Selective for Staphylococcus, differentiates mannitol fermenters

Which agar is selective for Staphylococcus due to high salt concentration?

Mannitol salt agar.

What makes Mannitol Salt Agar (MSA) selective?

High salt concentration (7.5%) selects for Staphylococcus species.

What makes Mannitol Salt Agar (MSA) a differential medium?

Contains mannitol (sugar) and a pH indicator (phenol red), which changes color based on acid production.

What indicates lactose fermentation on MacConkey agar?

Pink/red colonies.

What is MacConkey Agar selective for?

Gram-negative bacteria

What makes MacConkey Agar selective?

Bile salts and crystal violet inhibit Gram-positive bacteria, allowing Gram-negative growth.

What color indicates a positive lactose fermentation result on MacConkey Agar?

Pink or red colonies (e.g., E. coli).

What color do non-lactose fermenters appear on MacConkey Agar?

Colorless or pale colonies.

What makes Eosin Methylene Blue (EMB) Agar selective?

Eosin Y and methylene blue inhibit most Gram-positive bacteria, selecting for Gram-negative bacteria.

What does Eosin Methylene Blue (EMB) Agar differentiate?

Lactose fermenters (green metallic sheen for strong fermenters like E. coli)

What does a green metallic sheen indicate on EMB Agar?

Strong lactose fermentation, commonly seen with E. coli.

What color do weak lactose fermenters appear on EMB Agar?

Pink or purple colonies.

What is the main purpose of Xylose Lysine Deoxycholate (XLD) Agar?

To isolate enteric pathogens, especially Shigella and Salmonella

What color do Shigella colonies appear on XLD Agar?

Red colonies.

What result indicates Salmonella on XLD Agar?

Red colonies with black centers (H₂S production).

Why does E. coli appear yellow on XLD Agar?

It ferments lactose and sucrose, lowering pH.

Why does Salmonella produce a black precipitate on XLD Agar?

Because it reduces sulfur compounds like sodium thiosulfate.

What does Fluid thioglycollate broth help determine?

Oxygen sensitivity of bacteria.

How does Fluid Thioglycollate Broth determine oxygen tolerance?

By creating an oxygen gradient to distinguish aerobes, anaerobes, and facultative bacteria

What does DNase agar test for?

The ability of an organism to produce DNase enzyme.

What happens if an organism produces DNase on DNase Agar?

A clear zone forms around the bacterial growth.

What enzyme does Starch Agar test for?

Alpha-amylase (which breaks down starch).

What reagent is added to Starch Agar after incubation to test for starch hydrolysis?

Iodine.

What indicates a gram-positive result for starch hydrolysis on Starch Agar?

A clear zone around bacterial growth after adding iodine.

What is the result of a Gram-negative bacterium on Starch Agar after adding iodine?

Iodine is retained, resulting in a blueish/orange color around the bacterial growth.

What is the purpose of Blood agar?

Enriched medium that differentiates hemolysis of bacteria.

What type of hemolysis results in a clear zone around colonies?

Beta-hemolysis (complete lysis of red blood cells).

What type of hemolysis results in a greenish discoloration?

Alpha-hemolysis (partial lysis of red blood cells).

What type of hemolysis shows no change in the agar?

Gamma-hemolysis (no hemolysis).

What is the purpose of Chocolate Agar?

To grow Neisseria and Haemophilus influenzae

Why is Chocolate Agar brown?

Red blood cells are lysed by heating, releasing nutrients like hemin and NAD.

What additional factors do fastidious bacteria need that are provided by Chocolate agar?

NAD (V factor) and Hemin (X factor).