(Laboratory) Identification of Streptococcus and Enterococcus

Question 1

List down the step-by-step procedure in the Catalase Test of Streptococcus/Enterococcus:

Answer 1

1. Place 1 drop of hydrogen peroxide reagent in a glass slide
2. Emulsify a colony from the plate using applicator stick
3. Observe

Question 2

List down the step-by-step procedure in the Gram Staining of the Streptococcus/Enterococcus:

Answer 2

1. Using a sterile loop, place a loopful of saline into a labelled slide
2. Emulsify a colony from Plate into the saline
3. Air dry and heat fix
4. Stain with gram stain
5. View under microscope

Question 3

List down the step-by-step procedure in the Bacitracin disc test (Taxo A) of Streptococcus/Enterococcus:

Answer 3

1. Make a bacterial suspension (pure inoculum) by placing 3-5 colonies from plate into sterile distilled water. Match the turbidity with the MacFarland standard
2. Using lawn culture method, streak the pure inoculum into a blood agar plate
3. After drying for 10 minutes, place the 0.04 units bacitracin disc into the blood agar plate using sterile forceps
4. Incubate

Question 4

List down the step-by-step procedure in the CAMP Test for Streptococcus/Enterococcus:

Answer 4

1. Inoculate beta-toxin producing S. aureus straight on the center of BAP
2. Using bacteria from plate, inoculate a straight line at right angle to Staphylococcus streak. Make sure the two streak lines do not touch.
3. Incubate at 35 C for 18 hours in a candle jar
4. Observe

Question 5

List down the step-by-step procedure in the Colonial Morphology of Streptococcus/Enterococcus:

Answer 5

1. Using multiple interrupted streak, inoculate a BAP with bacteria from plate.
2. Incubate for 18-24 hours

Question 6

List down the step-by-step procedure in the Hippurate Hydrolysis:

Answer 6

1. Inoculate bacteria on sodium Hippurate substrate to produce a milky suspension
2. Incubate at 35 C for 2 hours
3. Add 0.2 mL ninhydrin reagent and further incubate for 15 minutes

Question 7

List down the step-by-step procedure in the Bile Esculin Test:

Answer 7

1. Using multiple interrupted streak, inoculate a BEA plate with bacteria from plate.
2. Incubate for 18-24 hours

Question 8

List down the step-by-step procedure in the Growth on 6.5% NaCl:

Answer 8

1. Inoculate loopful of bacteria to 6.5% NaCl broth
2. Incubate at 35 C for 18 hours

Question 9

Gram Stain Morphology of Streptococcus/Enterococcus:

Answer 9

Gram-positive cocci in pairs or chains

Question 10

What is the Catalase Test Result of Enterobacteriaceae:

Answer 10

Catalase Negative
No Effeverscence

Question 11

A test that differentiates Group A Streptococci from other Streptococci

Answer 11

Bacitracin Susceptibility Test (Taxo A)

Question 12

What is the positive result in Bacitracin Susceptibility Test (Taxo A)?

Answer 12

Zone of Inhibition
Group A Strep

Question 13

What is the negative result in Bacitracin Susceptibility Test (Taxo A)?

Answer 13

Negative Zone of Inhibition
Other Beta-hemolytic Streptococci spp.

Question 14

Test for Group B Streptococci except for Hippurate Test:

Answer 14

CAMP Test

Question 15

Positive Result in CAMP Test:

Answer 15

Arrowhead hemolysis [Group B Strep] or bowtie appearance [S. agalactiae]

Question 16

CAMP means:

Answer 16

Christie, Atkins, Munch and Petersen

Question 17

A diffusible extracellular hemolytic heat stable protein

Answer 17

CAMP Factor

Question 18

Test for Group A and Group D Streptococci

Answer 18

PYR Hydrolysis Test

Question 19

Positive Result in the PYR Hydrolysis Test:

Answer 19

Cherry Red/Pink

Question 20

Substrate of the PYR Hydrolysis Test:

Answer 20

L-pyrrolidonyl-beta-napthylamide

Question 21

Reagent of the PYR Hydrolysis Test:

Answer 21

p-dimethyl-aminocinnamaldehyde

Question 22

Enzyme of PYR Hydrolysis Test:

Answer 22

Pyrroglutamylamino peptidase or Pyrrolidomylaryl amidase

Question 23

The only species of streptococcus that is PYR positive

Answer 23

Streptococcus pyogenes

Question 24

A test used to detect the ability of bacteria to hydrolyze substrate hippurate into glycine and benzoic acid by the action of the hippuricase enzyme present in the bacteria.

Answer 24

Hippurate Hydrolysis Test

Question 25

A constitutive enzyme that hydrolyzes the substrate hippurate to produce the amino acid glycine

Answer 25

Hippuricase

Question 26

Medium of the Hippurate Test:

Answer 26

Broth with Hippurate

Question 27

Reagent Indicator in the Hippurate Test

Answer 27

Ninhydrin

Question 28

Substrate of the Hippurate Test

Answer 28

Sodium Hippurate

Question 29

What is the positive result of the Hippurate Hydrolysis Test?

Answer 29

Purple Color Streptococcus agalactiae

Question 30

_____ is detected by the oxidation with Ninhydrin which results in the production of a deep purple.

Answer 30

Glycine

Question 31

A test that differentiate Group D from Beta-hemolytic

Answer 31

Bile Esculin Test

Question 32

Enzyme of the Bile Esculin Test

Answer 32

Esculinase

Question 33

Media used for the Bile Esculin Test

Answer 33

Bile Esculin Agar

Question 34

What is the color of the Bile Esculin Agar?

Answer 34

Slightly Brown in color

Question 35

Substrate used in the Bile Esculin Test

Answer 35

Esculin

Question 36

What is the positive result of the Bile Esculin Test?

Answer 36

Blackening of Medium Group D

Question 37

Further test from Bile Esculin Test

Answer 37

Salt Tolerance Test (6.5% NaCl)

Question 38

What is the negative result of the Salt Tolerance Test (6.5% NaCl)?

Answer 38

Clear/Transparent Non-enterococci

Question 39

What is the positive result of the Salt Tolerance Test (6.5% NaCl)?

Answer 39

Turbid Enterococci

Question 40

Media used in the Salt Tolerance Test (6.5% NaCl)

Answer 40

6.5% NaCl Broth

Question 41

Substrate in the Leucine Aminopeptidase Test

Answer 41

Leucine-β-naphthylamide → β-naphthylamine

Question 42

Reagent in Leucine Aminopeptidase Test

Answer 42

Paradimethylaminocinnamaldehyde reagent

Question 43

Positive result in the Leucine Aminopeptidase Test

Answer 43

Red Color Viridans Streptococci

Question 44

A susceptibility test in conjunction with bacitracin is used for the presumptive identification of beta hemolytic Streptococci on blood agar.

Answer 44

SXT Susceptibility Test

Question 45

What specie/s is resistant to SXT Susceptibility Test?

Answer 45

Group A & B Strep

Question 46

What specie/s is susceptible in the SXT Susceptibility Test?

Answer 46

Other Strep spp.

Question 47

A test that detect carbohydrate component of the cell wall of Streptococci

Answer 47

Serological Test

Question 48

Principle of Serological Tests

Answer 48

Antigen (from colonies) - Antibody (from Reagent) Reaction

Question 49

Positive result in the Serological Tests

Answer 49

Agglutination/clumping [Group A Strep]

Question 50

Negative result in the Serological Tests

Answer 50

No agglutination [S. agalactiae]

Question 50

A rapid latex test system for use in the qualitative detection and identification of the lancefield group of Streptococci.

Answer 50

Serological Tests

Question 51

Incubation period of CAMP Test

Answer 51

35 C for 18 hours

Question 52

How many units of Bacitracin Disc is placed into plate?

Answer 52

0.04 units

Question 53

Incubation period of CAMP Test

Answer 53

35 C for 18 hours

Question 54

How many units of Bacitracin Disc is placed into plate?

Answer 54

0.04 units

Question 55

Incubation period of Hipurate Hydrolysis Test

Answer 55

35 C for 2 hours and then after adding 0.2 mL ninhydrin reagent, further incubate for 15 minutes

Question 56

How many ml of ninhydrin reagent is added into the tube for Hippurate Hydrolysis Test?

Answer 56

0.2 mL of ninhydrin reagent

Question 57

Incubation period for the Bile Esculin Test

Answer 57

18-24 hours

Question 58

Incubation period for the 6.5% NaCl growth

Answer 58

35 C for 18 hours