32 Aims

To understand how to identify unknown microorganisms using molecular and media-based techniques
To understand how to isolate and analyse plasmid DNA from microorganisms
To understand how to carry out PCR and interpret the results from multiplexed reactions

32.1 Learning Outcomes

At the end of this laboratory session, you should understand:

How to use differential media to identify microorganisms
How to use PCR and MVLA typing to identify microorganisms
How to use biochemical tests (coagulase) to identify microorganisms

32.2 Competencies

Using aseptic technique.
Pipetting
Analysing genotypes using PCR
Performing basic lab calculations
Documenting and reporting on experimental results and conclusions.

32.3 Importance

This laboratory session aligns with lectures and reinforces material relating to circulatory system and skin pathogens, bacterial identification and phylogeny, bacterial virulence and clinical microbiology.

32.4 Overview of Lab 4

In this lab session, you will be learning (or further developing your skills in) a number of key microbiology skills:

You will analyse the results from the Mannitol Salt Agar plates you streaked in the previous lab session (Task 4A)
You will perform a biochemical assay to help identify pathogens of the cardiovascular system/skin (a coagulase test, Task 4B)
You will perform Multiple Locus Variable number of tandem repeats Analysis (MLVA) to help identify your unknown Staphylococcus aureus strain.
You will analyse your results from the prevous lab (growth of your unknown organism on MacConkey agar; oxygen requirement of your unknown organism (fluid thioglycollate medium and anaerobically-incubated plates))
You will set up plates for the next lab (analysing the antibiotic resistance of Pseudomonas isolates)

To prepare for Lab 4

Before attending the practical lab, you should ensure that you have read and understood:

The introductory sections covering the theory on Identification of staphylococci, Biochemical assays: lysostaphin and coagulase, MLVA typing, and Antibiotic resistance assays
The Protocols we will be following in the lab
You should also make sure you have performed the calculations required for your antibiotic resistance experiment (Table 36.1)

You also must ensure that you have read and signed the safety forms, or you will not be admitted to the lab.