Diagnostic Bacteriology

Question 1

What are the 5 main types of infection?

Answer 1

Bacteria
Viruses
Fungi
Protozoa
Helminth worms

Question 2

What are the three types of bacteriological tests and what does each test identify?

Answer 2

Microscopy (Gram stain) - Which type of bacteria?
Culture - Which species of bacteria?
Sensitivities - Which treatment should be used?

Question 3

What colour are gram negative bacteria and what does it consist of?

Answer 3

Pink

2 outer membranes with thin peptidoglycan layer in the middle of them

Question 4

What colour are gram positive bacteria and what does it consist of?

Answer 4

Purple

1 outer membrane with thick peptidoglycan around it

Question 5

Which part of a bacterial cell envelope “holds” the Gram stain?

Answer 5

Peptidoglycan

Question 6

In what order are the gram stain substances applied?

Answer 6

Crystal violet
Iodine (mordant)
Alcohol (decolorisation)
Safranin (counterstain)

Question 7

What is the purpose of the alcohol washes?

Answer 7

Dehydrates the peptidoglycan area resulting in it becoming tighter

Question 8

What makes gram positive purple?

Answer 8

Alcohol causes peptidoglycan layer to thicken and tighten hence the crystal and iodine complexes can’t be washed out hence it remains stained with crystal violet and appears purple

Question 9

Why does gram negative bacteria stain pink?

Answer 9

Alcohol washes out crystal violet and iodine as there is only a thin peptidoglycan layer
Safranin then stains bacteria pink

Question 10

What shape does coccus describe?

Answer 10

Sphere

Question 11

What shape does bacillus describe?

Answer 11

Round-ended cyclinder

Question 12

How would you describe the shape of coccobacilli?

Answer 12

Oval-like

Question 13

Describe the shape of diplococci

Answer 13

Spherical in pairs

Question 14

What is hemolytic activity useful in differentiating between?

Answer 14

Gram positive → Staphylococcus, Streptococcus, Enterococcus

Question 15

What does the hemolytic activity test for?

Answer 15

Capacity of microorganism to produce hemolysins (enzymes that damage red blood cells)

Question 16

List and describe the results of the haemolysis test

Answer 16

Gamma - no haemolysis
Alpha - partial (opaque zone)
Beta - complete haemolysis (transparent greenish zone)

Question 17

What is the major cause of haemolysis in new born babies?

Answer 17

Group B Streptococcus or Streptococcus agalactiae is the commonest cause of meningitis in babies under 3 months

Question 18

Outline how to complete the haemolytic activity test

Answer 18

1. Pick a bacterial colony from an agar plate
2. Streak out the bacterial colony onto medium containing 5% sheep red blood cells
3. Incubate plate overnight and assess activity next day
4. What we assess are the affect of the bacteria on the red blood cells

Question 19

What type of agar is used for the lactose fermentation test?

Answer 19

MacConkey agar - containing bile salts, crystal violet and lactose

Question 20

What is the lactose fermentation used for?

Answer 20

Differentiating gram negative bacteria (more specifically, between species of Bacilli)

Question 21

What does the lactose fermentation test measure?

Answer 21

Difference in pH to determine whether bacteria is fermenting or non-fermenting

Question 22

What are the results for a lactose-fermenter?

Answer 22

Agar plate shows pink colonies after being incubated overnight

Question 23

What are the results for a lactose non-fermenter?

Answer 23

Colourless (yellow) colonies on agar plate

Question 24

What does the catalase test determine?

Answer 24

Determines whether a bacteria produces catalase

Question 25

What does catalase break down?

Answer 25

Hydrogen peroxide

Question 26

What does the catalase test discriminate between?

Answer 26

Staphylococci (produces catalase)

Streptococci (does not produce catalase)

Question 27

What are the results for catalase positive and catalase negative?

Answer 27

Catalase + = Oxygen bubbles

Catalase - = No oxygen bubbles

Question 28

Outline how to complete the catalase test

Answer 28

1. Apply bacteria onto glass slide
2. Apply 2H2O2 (hydrogen peroxide)
3. Observe generation of bubbles

Question 29

What is coagulase?

Answer 29

Enzyme that cross-links fibrinogen in plasma to form a clot on the bacterial surface
(Processes factor 2 to 2a. Thrombin cleaves fibrinogen into fibrin. Factor VIIIa cross links fibrin to form fibrin clots.)

Question 30

What can coagulase be used to discriminate between?

Answer 30

S. aureus (produces coagulase) and other staphylococci (don’t produce coagulase)

Question 31

Describe the results of coagulase positive and negative

Answer 31

Coagulase + = clumps

Coagulase - = no clumps

Question 32

Outline the steps for the coagulase test

Answer 32

1. Apply bacteria onto slide
2. Apply plasma
3. Incubate for 15 seconds and gently rotate
4. Generation of clumps if coagulase +

Question 33

List the names of some diplococci and what gram stain they are

Answer 33

N. gonorrhoeae
N. meningitidis
M. catarrhalis

All Gram-negative

Question 34

Which of these species gives a positive coagulase test?

Staphylococcus epidermidis
Staphylococcus aureus
Staphylococcus lugdenesis
Staphylococcus hemolyticus

Answer 34

Staphylococcus aureus