Microbiology Lab Practical 1 March 2009

Front 1

what is the purpose of a four quadrant streak plate?

Back 1

to isolate bacteria or bacterial colony

Front 2

what are the three colony characteristics

Back 2

form (what is the basic shape - circular, irregular, or filamentous); elevation (what is the cross-sectional shape of the colony); margin (what is the magnified shape of the colony)

Front 3

name the three bacterial growth characteristics in broth

Back 3

pellicle, sediment, turbidity

Front 4

describe pellicle

Back 4

a growth characteristic of broth; thin film of bacteria growing at the surface of nutrient broth (strict aerobe)

Front 5

describe sediment

Back 5

Bacteria growing at the bottom (facultative
anaerobe and anaerobic bacteria)

Front 6

describe turbidity

Back 6

Growing throughout the broth (facultative anaerobe)

Front 7

simple stain

Back 7

1. basic dye 2. positive charge; note: the basic dye can be crystal violet, methylene blue, safranin, or malachite green

Front 8

how do we determine morphology of bacteria

Back 8

with the simple stain (basic, positive) we can determine shape and arrangement (morphology)

Front 9

what are the basic shapes and arrangements of bacteria?

Back 9

shapes: coccus, bacillus, spirilum; arrangements: staphylo, strepto, tetrad, sarcina

Front 10

for gram staining, name the following: primary stain, mordant, decolorizer, counter stain

Back 10

• Crystal Violet-Primary
Stain
• Iodine-Mordant
• Alcohol-acetone
decolorize
• Safranin-Counter stain

Front 11

what color is gram positive? gram negative?

Back 11

Gram Positive will retain
purple dye
Gram Negative will be
pink

Front 12

what is the name of the spore stain?

Back 12

there are two: malachite green and safranin; the spore stains green, the rest of the bacteria stain pink (from safranin)

Front 13

name the spore forming organisms

Back 13

– Clostridium
– Bacillus

Front 14

describe acid fast organisms

Back 14

mycobacterium; gram + rod; mycolic acid in cell wall

Front 15

dyes for acid fast organisms

Back 15

• Dyes
– Carbolfuchsin
– acid-alcohol
– methylene blue

Front 16

what are the results from acid fast staining?

Back 16

• Acid Fast + deep red
• Acid Fast - blue

Front 17

with negative staining what are we staining?

Back 17

Negative stain
• Background is stained
instead of the organism

Front 18

what types of dyes are used with negative staining? + or -?

Back 18

•Acidic dyes with a negative
charge are used

Front 19

what are the names of the dyes used for negative staining?

Back 19

•Acidic dyes used for negative staining are Nigrosin and
Congo red

Front 20

with negative staining, what happens to the capsule?

Back 20

Capsule remains clear since
no dyes can stain capsule

Front 21

does india ink have a + or - charge?

Back 21

india ink has no charge

Front 22

what is monotrichous?

Back 22

only one flagella

Front 23

what is amphitrichous?

Back 23

one flagella on each end of organism (2 flagella total)

Front 24

lophotrichous?

Back 24

multiple flagella coming from both ends.

Front 25

peritrichous?

Back 25

flagella are on all sides of organism; multiple flagella

Front 26

what is involved with the bacterial motility test?

Back 26

• Hanging drop method
• Motility medium
• Flagella stain

Front 27

what does the test tube look like if the bacteria are non-motile? motile?

Back 27

the motile bacteria spread the red dye all over the test tube. the non-motile bacteria leave the red dye in a line in the center of the test tube.

Front 28

what is the name of the tube used when testing fermentation of carbohydrates?

Back 28

durham tubes.

Front 29

purpose of durham tubes

Back 29

catch gas bubbles

Front 30

what is the medium used when determining if bacteria is a fermenter of carbohydrates?

Back 30

pH indicator: Phenol Red

Front 31

when using durham tubes, what is a red result? orange? yellow?

Back 31

• Red = basic
• orange= neutral
• yellow = acidic

Front 32

what is starch?

Back 32

complex polysaccharide

Front 33

when checking for digestion of starch using a complex polysaccharide, what is the medium. what enzyme are we testing for?

Back 33

we use iodine to test for the enzyme amylase

Front 34

what is a positive result when testing for digestion of starch?

Back 34

Positive for digestion of starch is Amylase positive -- a halo around the area
where starch is utilized

Front 35

what is a positive result when testing to see if the enzyme amylase is present?

Back 35

Positive for Amylase is
halo around the area
where starch is utilized

Front 36

what is a positive result when testing for digestion of starch using a complex polysaccharide?

Back 36

Positive for Amylase is
halo around the area
where starch is utilized

Front 37

when checking for DNA digestion, what plate do we use?

Back 37

DNAse Agar Plate with Methyl Green

Front 38

what dye do we use when checking for DNA digestion

Back 38

methyl green

Front 39

what enzyme are we testing for with DNA digestion testing?

Back 39

DNase

Front 40

what signifies a +DNAse test?

Back 40

clearing around the organism

Front 41

what does the enzyme DNAse do?

Back 41

• Enzyme DNAse
– breaks down long
chains of DNA

Front 42

catalase test: what are we testing for?

Back 42

testing to see if the catalase enzyme is present

Front 43

what does catalase do?

Back 43

breaks down hydrogen peroxide into H2O and O2

Front 44

what happens if catalase is present -

Back 44

this is a + test; bubbling occurs

Front 45

what happens if there is no catalase

Back 45

no bubbling; - test

Front 46

what types of bacteria are catalase +? catalase - ?

Back 46

• Streptococci
– Catalase Negative
• Staphylococci
– Catalase Positive

Front 47

when testing for hydrogen sulfide production, what enzyme are we testing for?

Back 47

– cysteine desulfurase

Front 48

what does cysteine desulfurase do?

Back 48

• Breaks down cysteine
into alanine and
hydrogen sulfide
(H2S)

Front 49

concerning hydrogen sulfide testing: Iron (Fe2++) + hydrogen
sulfide (H2S) = ?

Back 49

– Iron (Fe2++) + hydrogen
sulfide (H2S) =FeS

Front 50

concerning hydrogen sulfide testing: black precipitate = ?

Back 50

– black precipitate = H2S
production

Front 51

concerning hydrogen sulfide testing: SIM = ?, ?, ?,

Back 51

– SIM = sulfide, indole,
motility,

Front 52

concerning hydrogen sulfide testing, what does a positive test look like? negative test?

Back 52

H2S positive is brown/gold and cloudy. H2S negative is milky yellow

Front 53

look at micro review for...

Back 53

look at slide 4 to get a mental picture of all of the shapes of bacteria

Front 54

is staphylococcus epidermidis gram + or garm -

Back 54

gram +

Front 55

is citrobacter diversus gram + or gram -

Back 55

gram -

Front 56

when doing an acid fast test against mycobacterium smegmatis, describe what the results will look like

Back 56

mycobacterium smegmatis is acid-fast +, so it will be deep red/deep pink

Front 57

when doing an acid fast test against enterococcus faecium, describe what the results will look like

Back 57

enterococcus faecium is acid-fast negative so, it will be blue

Front 58

describe the stain of Klebsiella Pneumonia - using the negative staining

Back 58

the acidic stain colorizes the background while the basic stain colorizes the cell, leaving the capsules as unstained, white clearnings around the cells

Front 59

when doing fermentation of carbohydrates in the durham tubes and testing Escherichia Coli, what are the results?

Back 59

A/G (see lab manual and results for explanation of A/G, A/-, -/-, K)

Front 60

when doing fermentation of carbohydrates in the durham tubes and testing Staphyloccocus Aures, what are the results?

Back 60

A/- (see lab manual and results for explanation of A/G, A/-, -/-, K)

Front 61

when doing fermentation of carbohydrates in the durham tubes and testing Micrococcus Luteus, what are the results?

Back 61

- / - (see lab manual and results for explanation of A/G, A/-, -/-, K)

Front 62

when doing fermentation of carbohydrates in the durham tubes and testing Alcaligenes faecalis, what are the results?

Back 62

K (see lab manual and results for explanation of A/G, A/-, -/-, K)

Front 63

when testing for amylase on the starch agar plate, what is the result of E. Coli and Bacillus cereus

Back 63

E. Coli is negative; Bacillus cereus is positive

Front 64

On a DNAse agar plate, with Methyl green, what will the results be for: Staph aures and Staph epidermidis?

Back 64

S. aures is + (with a "clearing" around the bacteria); S. epidermidis is - (with no change)

Front 65

when testing for indole production, what enzyme are we testing for?

Back 65

tryptophanase

Front 66

what does tryptophanase do?

Back 66

• breaks down tryptophan
into indole, pyruvic acid
and ammonia

Front 67

what reagent is used for the indole production test?

Back 67

Kovac’s reagent

Front 68

what happens to indole if Kovac's is present?

Back 68

• Indole + Kovac’s = red
ring

Front 69

what happens if there is no indole present?

Back 69

• No Indole + Kovac’s =
amber or green color

Front 70

when the SIM medium is innoculated with Morganella morganii, what happens

Back 70

red ring forms; indole positive

Front 71

when the SIM medium is innoculated with enterobacter aerogenes, what happens

Back 71

no red (looks a little more cloudy than the control); indole negative

Front 72

when testing for urea digestion, what enzyme are we testing for?

Back 72

Urease

Front 73

when testing for urea digestion, what is the substrate?

Back 73

urea

Front 74

when testing for urea digestion, what is the pH indicator?

Back 74

Phenol Red

Front 75

what are four possible results from the urea digestion testing?

Back 75

– Red @ Basic pH
– Orange @ neutral
– Yellow @ acidic
– Hot pink = ammonia
released from urea

Front 77

describe form

Back 77

Form - What is the basic shape of the
colony? For example, circular, irregular or filamentous, etc.

Front 78

describe elevation

Back 78

Elevation - What is the cross sectional
shape of the colony?

Front 79

describe margin

Back 79

Margin - What is the magnified shape
of the edge of the colony?

Front 80

for the simple stain, why are the dyes a positive charge?

Back 80

all basic dyes for simple staining are (+) charge. they are attracted to the (-) charged cytoplasm

Front 81

with gram staining, what step is most critical

Back 81

the decolorizing step is the most critical, organism can be over decolorized or under decolorized if not careful

Front 82

with negative staining, india ink is too ___ to penetrate the cell wall

Back 82

too large

Front 83

with capsule staining, why doesn't the capsule stain?

Back 83

because it has no charge

Front 84

what makes spores difficult to stain?

Back 84

the dipicoloinic acid in the spore coat makes it difficult to stain

Front 85

why is mycobacterium difficult to stain?

Back 85

the cell wasll of mycobacterium contains mycolic acid making it difficult to stain

Front 86

what is involved with capsule staining?

Back 86

capsule staining is a negative stain slide overlaid with a simple stain

Front 87

which is not a growth characteristic in broth? (a) pellicle (b) color (c) sediment (d) turbidity (e) none of the above

Back 87

color is not a growth characteristic

Front 88

which is a margin characteristic? (a)filamentous (b)rhizoid (c)umbonate (d)circular

Back 88

filamentous is a margin characteristic

Front 89

simple stains are (a)acidic (b) neutral (c) basic

Back 89

simple stains are basic

Front 90

which is NOT a simple stain? (a)crystal violet (b)methylene blue (c)congo red (d)malachite green

Back 90

congo red is NOT a simple stain