Exercise 6_ Bacterial Cell Wall

Question 1

primarily provides a protective shield to bacteria against hostile environment, allow communication with its surroundings to find optimal conditions for growth and development of the bacterial cell

Answer 1

bacterial cell wall

Question 2

role of bacterial cell wall

Answer 2

• protective shield to bacteria against hostile environment, - allow communication with its surroundings to find optimal conditions for growth and development of the bacterial cell - its rigidity maintains the shape of the bacterium - enables transport of large molecules into and out of the cell

Question 3

who developed gram stain

Answer 3

Danish scientist and physician, Hans Christian Joachim gram

Question 4

when was gram stain developed?

Answer 4

year 1884

Question 5

gram staining is what type of staining method

Answer 5

differential staining

Question 6

advantages of using gram staining

Answer 6

• rapid turnaround and generation of information about the organism’s gram rxn - available inexpensive reagents

Question 7

2 divisions of bacteria according to gram staining method

Answer 7

gram + gram -

Question 8

what can be seen under the light microscope after staining the cells with gram stain

Answer 8

cell shape cell size cell arrangement

Question 9

what differentiates the gram reaction of a bacterium when stained with gram stain

Answer 9

surface layer composition (cell wall)

Question 10

Gram ____ are decolorized by alcohol, losing the purple color of crystal violet

Answer 10

negative

Question 11

Gram ____ are not decolorized by alcohol, can retain the purple color of crystal violet

Answer 11

positive

Question 12

True or false: a bacterium with a thick peptidoglycan can retain crystal violet after decolorization of alcohol

Answer 12

true

Question 13

True or false: a bacterium with a thin peptidoglycan can retain crystal violet after decolorization of alcohol

Answer 13

false

Question 14

counterstain medium of gram stain

Answer 14

safranin

Question 15

what color does safranin impart to the cells of the decolorized gram negative bacterium?

Answer 15

red or pink

Question 16

which gram bacterium will manifest red/pink colored cells after decolorization

Answer 16

gram negative

Question 17

which gram bacterium will manifest violet or blue colored cells after decolorization

Answer 17

gram positive

Question 18

what method can be used to determine the gram rxn of an unknown bacterium

Answer 18

gregersen’s method

Question 19

if the bacterium mounted in KOH turns viscous it is gram

Answer 19

gram negative

Question 20

if the bacterium mounted in KOH turns watery it is gram

Answer 20

gram positive

Question 21

control organisms utilized in this exercise

Answer 21

B polymyxa E coli

Question 22

gram negative control organism

Answer 22

E coli

Question 23

E coli’s gram reaction

Answer 23

gram negative

Question 24

gram positive control organism

Answer 24

B polymyxa

Question 25

B polymyxa's gram reaction

Answer 25

gram positive

Question 26

percent concentration of KOH

Answer 26

0.03

Question 27

mounting medium of gram stai

Answer 27

distilled water

Question 28

mounting medium of gregersen's method

Answer 28

3% KOH

Question 29

3% KOH is the mounting medium of which method?

Answer 29

gregersen's method

Question 30

distilled water is the mounting medium of which method?

Answer 30

gram stain

Question 31

During gregersen's test, do you heat fix samples?

Answer 31

NO

Question 32

During gram staining, do you heat fix samples?

Answer 32

yes

Question 33

1st staining agent utilized during gram staining

Answer 33

crystal violet

Question 34

time requirement in staining cells with crystal violet

Answer 34

1 min

Question 35

after staining with crystal violet, what's next?

Answer 35

wash smear with tap water

Question 36

next stain after crystal violet

Answer 36

gram's iodine

Question 37

time requirement in staining cells with gram's iodine

Answer 37

1-2 min

Question 38

decolorizer of gram staining

Answer 38

95% ethanol

Question 39

percent concentration of the decolorizer

Answer 39

0.95

Question 40

time requirement in decolorizing cells

Answer 40

10-15 secs

Question 41

counter stain used in gram staining

Answer 41

safranin

Question 42

time requirement in staining cells with safranin

Answer 42

45 secs

Question 43

where does crystal violet binds to?

Answer 43

peptidoglycan

Question 44

gram's iodine serves as the ____ which combines the stain with the peptidoglycan of the bacterial cell wall forming a complex

Answer 44

mordant (fixing agent)

Question 45

false gram stain results can be because of

Answer 45

- excessive heat fixation - concentration of primary stain - smear is too thick - faulty smear prep - age of culture - use of antibiotics - insufficient iodine exposure - prolonged decolorization - excessive counterstain - lack of experience

Question 46

this bacteria cannot be stained using gram staining due to the lack of a peptidoglycan in the cell wall

Answer 46

Mycoplasma

Question 47

Mycoplasma cannot be stained by gram staining because of what

Answer 47

absence of peptidoglycan in the cell wall

Question 48

the viscous sample in gregersen's test contains

Answer 48

cell wall debris and DNA

Question 49

the outer membrane of the gram-negative bacteria interact with ____ ____ causing the hydrolysis of the ____ bonds breaking down the cell wall including the peptidoglycan

Answer 49

hydroxide ions, ester

Question 50

hydroxide ions interact with what part of the gram negative bacterium

Answer 50

outer membrane

Question 51

what does the gram negative has that is not found in gram positive bacteria

Answer 51

outer membrane thick peptidoglycan

Question 52

what aids the cell wall of gram positive bacteria to not be degraded during gregersen's method?

Answer 52

thick peptidoglycan

Question 53

as a control organism, what is expected to be the color of the cells of E coli after gram staining

Answer 53

pink/re

Question 54

as a control organism, what is expected to be the color of the cells of B polymyx after gram staining

Answer 54

purple/blue

Question 55

alternatives for crystal violet as the primary stain for gram staining

Answer 55

methyl violet/gentian violet

Question 56

methyl violet/gentian violet can be alternative of what stain in gram staining method?

Answer 56

primary stain

Question 57

staining alternatives for safranin as the counterstaining agent in gram staining

Answer 57

dilute carbol fuchsin or neutral red

Question 58

dilute carbol fuchsin or neutral red can be alternative of what stain in gram staining method?

Answer 58

counterstain/safranin

Question 59

effect of replacing iodine with other oxidizing agents (eg. bromine, chlorine/fluorine)

Answer 59

can cause decolorization unnecessary cell wall interaction not form complexes with the primary stain

Question 60

importance of using freshly prepared KO

Answer 60

-KOH degrades over time as it reacts with the environment which can affect its ability to differentiate the gram positive and gram negative bacteria -Remains pure from contaminants (eg carbon dioxide)

Question 61

KOH stands for

Answer 61

potassium hydroxide

Question 62

higher concentrations than the 3% optimum concentrations of the KOH it will produce false ____ results

Answer 62

negative

Question 63

lower concentrations than the 3% optimum concentrations of the KOH it will produce false ____ results

Answer 63

positive