diagnostic bacteriology

Question 1

What library diagnostic tests are used and what samples can be taken?

Answer 1

Laboratory investigations
○ Microbiological tests
○ Immunological tests
○ Molecular biology tests

Samples
○ Saliva, blood. Urine, faeces
○ Sputum, cerebrospinal fluid
○ Tissue biopsies, pus

Question 2

Why is diagnosis important?

Answer 2

Improves effectiveness of treatment - Avoid long term complications

Identify and prevent outbreaks
Prevent transmission of virus/bacteria
Prevent further cases or an outbreak

Prevent overuse or mis-use of treatments - Reduce use of antibiotics when unnecessary

Question 3

What is the gram stain test based on and what does it identify?

Answer 3

• Gram-negative bacteria - 2 outer membranes with thin peptidoglycan
Stays purple

• Gram-positive bacteria - 1 outer membrane with thick peptidoglycan
Appears pink

Question 4

Explain the process of gram staining

Answer 4

• use heat to fix bacteria to surface of slide
• Apply crystal violet solution to the bacteria, crystal violet molecules enter cell wall
• Iodine is applied, leading to the formation of crystal violet and iodine complexes that are larger in size
• Series of washes with alcohol, alcohol dehydrates the peptidoglycan layer resulting in it becoming tightened

In gram positive bacteria the peptidoglycan later is thick complexes are unable to wash out so stay stained purple

gram negative bacteria- a thinner peptidoglycan layer the complexes are washed out

Counter stain with safranin and this stains all bacteria pink so gram negative stains pink

Question 5

What are the two classifications of bacterial shape?

Answer 5

Spheres - coccus

Round ended cylinders - bacillus

Can also have oval shaped - coccobacilli

Question 6

What are the four growth tests done to diagnose bacteria?

And what gram bacteria are they used for?

Answer 6

+ve
Hemolytic activity - whether they produce hemolysins - enzymes to damage RBCs

Catalase - determines whether bacteria produces catalase (breaks down H2O2)

coagulase test - discriminate gram positive bacteria - staphylococci (produce catalase) and streptococci (do not produce catalase).

-ve
Lactose fermentation - tests the capacity of a microorganism to ferment lactose

Question 7

method to test for haemolytic activity?

Answer 7

• Method

	1. Pick out a colony from an agar plate 
	2. Strike out the colony on an agar plate that contains five percent sheep red blood cells 
	3. Incubate the plate overnight and assess hemolytic activity the next day 
	4. See whether the red blood cells have been lysed

Question 8

method to test for fermentation?

Answer 8

1. Pick a bacterial colony off an agar plate
   2. Streak out the colony onto a Conkey agar
   that contains an indicator, at neutral agar should be
   red
   3. Incubate overnight and assess lactose fermentation
   the day after
   4. If lactose fermenter bacteria, lactic acid will lower
   ph and the colonies will be pink if not the colonies
   appear colourless

Question 9

method of coagulate test|?

Answer 9

1. Bacteria applied to a glass slide
2. Plasma applied on top
3. Short incubation with gentle rotation
4. Coagulase positive forms clumps whereas negative will not

Question 10

Method of catalase test?

Answer 10

1. Bacteria is applied to a glass slide
2. Hydrogen peroxide is applied to bacteria
3. Short incubation
4. Observe whether bubbles are produced

Question 11

the two methods to test for antibiotic resistance?

Answer 11

Zone of inhibition

E-test to measure minimum inhibitory concentration