Molecular Methods in Clinical Microbiology

Question 1

Mass Spectrometry

Answer 1

• Has become the new standard of most bacterial identification
• Provides identification of a PURE COLONY in 3-5 minutes!
• Only used for bacteria and yeast
• Not used for moulds, some bacteria, viruses or parasites

Question 2

Polymerase Chain Reaction (PCR)

Answer 2

• Used to amplify a single or several targets of nucleic acid (DNA or RNA) by doing so you are detecting the presence of the target nucleic acid (pathogen)
• Generates several thousand copies of the target sequence.

Question 3

Stages of PCR

Answer 3

1. Denaturing
2. Annealing
3. Extension
   - Increase temp. just enough that your primers still work, then decrease just slightly to allow DNA to bind to YOUR primers, but not low enough for DNA to come back together

Question 4

Ingredients in a PCR reaction?

Answer 4

• Template RNA/DNA
• DNA polymerase
• 2 primers
• DNA nucleotides
• Cations

Question 5

Caution about PCR?

Answer 5

PCR could pick up dead genes… thus need to always take it in the clinical sense!

Question 6

Why use PCR? Disadvantages?

Answer 6

• Can test for agents that cannot be cultured… ex. virus! Slow growing agents… high risk agents that are dangerous to grow, anti-viral drug susceptibility,.

Disadvantages: $$$, very sensitive.. false positives!