lab pt 2

Why must a smear air-dry before heat-fixing?

To prevent boiling, splattering, and distortion of bacterial cells.

What is heat-fixing and why is it performed?

Passing an air-dried smear through a flame to kill bacteria and help them adhere to the slide.

How do stains work?

Dyes bind to cellular structures through chemical interactions, increasing contrast so cells can be seen.

How do basic and acidic dyes differ?

Basic dyes are positively charged and bind to negatively charged bacterial surfaces; acidic dyes are negatively charged and commonly stain the background.

What is the purpose of staining bacteria?

To increase contrast and make cells and structures easier to observe.

What is a Gram stain and why is it differential?

A differential staining technique that distinguishes bacteria based on differences in cell wall structure.

What are the four Gram stain steps in order?

Crystal violet → Gram's iodine → decolorizer → safranin.

What is the function of crystal violet?

The primary stain; initially colors all bacteria purple.

What is the function of Gram's iodine?

The mordant; forms a crystal violet-iodine complex that helps retain the primary stain.

What is the function of the decolorizer?

Alcohol or acetone-alcohol removes the crystal violet-iodine complex from Gram-negative bacteria.

What is the function of safranin?

The counterstain; colors decolorized Gram-negative bacteria pink or red.

Why do Gram-positive bacteria remain purple?

Their thick peptidoglycan layer retains the crystal violet-iodine complex during decolorization.

Why do Gram-negative bacteria become pink?

Their outer membrane is disrupted by alcohol, and their thin peptidoglycan cannot retain the crystal violet-iodine complex.

What colors are Gram-positive and Gram-negative bacteria?

Gram-positive: purple. Gram-negative: pink or red.

What happens if crystal violet is omitted?

The bacteria do not receive the primary purple stain.

What happens if iodine is omitted?

The crystal violet-iodine complex does not form properly, so Gram-positive bacteria may lose the purple stain.

What happens if the decolorizer is omitted?

Both Gram-positive and Gram-negative bacteria remain purple.

What happens if too much or too little decolorizer is used?

Too much: Gram-positive may appear falsely Gram-negative. Too little: Gram-negative may appear falsely Gram-positive.

What happens if safranin is omitted?

Gram-negative bacteria remain colorless after decolorization.

What is an endospore and why do bacteria form one?

A dormant, highly resistant structure formed by certain bacteria to survive unfavorable conditions.

What conditions can endospores resist?

Heat, drying, radiation, chemicals, and other environmental stresses.

Are endospores reproductive structures?

No. One bacterial cell produces one endospore, which can germinate into one vegetative cell.

Which bacterial genera commonly produce endospores?

Bacillus and Clostridium.

What happens when conditions become favorable?

The endospore germinates into an active vegetative cell.

What are culture characteristics?

Visible characteristics of bacterial growth on or in culture media.

What colony characteristics can be observed on solid media?

Size, shape, margin, elevation, color, texture, and opacity.

What is colony morphology?

The visible appearance of bacterial colonies on solid media.

What are colony form, margin, and elevation?

Form: overall shape; margin: colony edge; elevation: vertical profile.

What is colony opacity?

How much light passes through a colony: transparent, translucent, or opaque.

What is turbidity?

Cloudiness in liquid culture caused by suspended bacterial cells.