Exercise 7: The Gram Stain

Question 1

Who developed the Gram stain?

Answer 1

Hans Christian Gram

Question 2

What two groups does the gram stain identify?

Answer 2

gram positive

and gram negative

Question 3

What are antibiotics?

Answer 3

• chemicals that act very specifically to kill bacteria or inhibit bacterial growth

Question 4

Why do physicians rely on gram stain when prescribing antibiotics?

Answer 4

• some antibiotics kill only gram negative bacteria, some kill only gram positive, some are broad-spectrum and will kill/inhibit a wide array of bacteria (including members of the normal flora)

Question 5

How do gram positive and gram negative differ?

Answer 5

• because of the cell wall

Question 6

Describe gram positive cell wall.

Answer 6

• thicker wall
• multiple layers of peptidoglycan
• lipoteichoic and teichoic acids

Question 7

Gram positive cell walls are thought to what during decolorization?

Answer 7

retain the crystal-violet/iodine complex

Question 8

Describe gram negative cell walls.

Answer 8

• very thin cell walls
• usually only one layer of peptidoglycan
• ## outer lipid membrane: lipopolysaccharide

Question 9

During decolorization what happens to the gram negative cell walls?

Answer 9

usually stripped away. will not hold the crystal-violet complex but will hold the counterstain, safrain

Question 10

Do all bacteria stain readily with gram-positive or gram negative with this method?

Answer 10

• no several factors play role in getting reliable results

Question 11

What are the dependents on getting reliable results with the gram stain?

Answer 11

• age of culture
• naturally gram- variable
• The mycobacterium species
• resist decolorization

Question 12

What is a good age of culture for the gram stain procedure? and why?

Answer 12

• usually 18-24 hours old
• because after this time the cultures age, waste products begin to accumulate, and cell walls break down. So gram + could look gram -

Question 13

What genus will stain-variable no matter what the age of culture?

Answer 13

-Gram positive bacillus

Question 14

What species will appear barely visible after gram staining? and why?

Answer 14

• the Mycobacterium species

- because it contains a waxy mycolic acids that are impervious to gram-stain

Question 15

What species usually resists decolorization despite being gram- negative?

Answer 15

• the Neisseria species

Question 16

If you forget to add iodine or overdecolorize what will gram + look like following staining?

Answer 16

red

Question 17

If you underdecolorize what will gram + look like following staining?

Answer 17

purple

Question 18

If you forget to add iodine or overdecolorize what will gram- look like following staining?

Answer 18

red

Question 19

If you underdecolorize what will gram - look like following staining?

Answer 19

purple

Question 20

What does iodine act as during the gram-stain procedure?

Answer 20

a mordant, forms a molecular bond with with the crystal-violet

Question 21

What does the decolorizer do?

Answer 21

strips away the outer membrane of the gram negative cells (can also strip down gram + walls if left too long)

Question 22

What is safranin?

Answer 22

• a positively charged dye used as a counterstain

Question 23

What does purple cells mean at the end of the procedure?

Answer 23

• gram positive

Question 24

What does red cells mean at the end of the procedure?

Answer 24

• gram negative