Dear William,
Thanks for your email, this sounds like a really promising project. There are a few ways that you can try to detect S. aureus using different microbiological media as well as biochemical tests.
But the first thing you need to do, is prepare some nutrient agar plates (your college may already have some you can use). Then using a sterile swab, swab the inside of your nose (just run the tip of the swab around the inside of one nostril) and culture this on the nutrient agar. You will need to make sure that you can isolate pure colonies from the nose swab, so when you inoculate the agar plate you will have to prepare a streak plate – I am sure you biology teacher will be able to demonstrate this technique to you.
Incubate the plates at 37oC overnight, and the following day look to see if you have any colonies, and importantly to see if there are single colonies. Choose a few well isolated colonies and note their appearance i.e. colour, size, mucoid, shiny, matte, circular, entire or wrinkled edge.
Then subculture each of the colonies you have chosen onto a fresh nutrient agar plate (one for each colony selected) – again, prepare streak plates for single colonies (You could take photos of your plates to include in your project once they have grown) and incubate the plates overnight at 37oC.
Once the isolates have grown you can do some microscopy and biochemical tests. So the first thing to do is a Gram stain on each isolate – make sure you use well isolated colonies for this. Your biology teacher will be able to show you how to Gram stain. View the slides using x100 oil immersion if you can, if not x40 will do. You are looking for round shaped (coccus) purple coloured (Gram positive) bacterial cells – these are likely to be S. aureus. They might also grow in grape-like clusters, again indicative of S. aureus. If its possible, you could take pictures of the slides, you can just hold a camera up to the microscope eye-piece to do this, so you have nice images for your report.
So, next you need to record which of your isolates were Gram positive coccus shaped cells, because you will do some biochemical tests with these. Only test the ones that are Gram positive cocci, discard the other isolates.
For the Gram positive cocci you can do a catalase test, which determines the ability of the organism to produce the enzyme catalase which breaks down hydrogen peroxide into water and oxygen. To do the catalase test, remove one isolated colony from your agar plate using a wire loop transfer it onto a glass microscope slide. Put one or two drops of a 3-6% hydrogen peroxide solution onto a glass coverslip (use a Pasteur pipette to do this). Then then place the microscope slide, bacteria face down, onto the coverslip. If the organisms is catalase positive you will see bubbles of oxygen being produced. The reaction is quite fast, so you will need to examine it immediately.
Then record how many of your Gram positive isolates are catalase positive – S. aureus is a catalase positive organism.
Next, sub-culture the Gram positive, coccus shaped, catalase positive isolates (only) onto Mannitol salt agar (MSA) plates (your biology teacher will provide these for you). Again prepare streak plates. Incubate the plates over night at 37oC. It is important that you note what colour the agar is before you inoculate the plate.
The next day look at the plates and record any that have produced yellow coloured colonies with a yellowing of the surrounding agar (if you Google mannitol salt agar you will find images of what they might look like). Any organism that grows on MSA is a S. aureus. Any isolates that produce yellow coloured colonies are called coagulase positive S. aureus, and strains such as MRSA fall into this category. Make sure you don’t open these agar plates and that you do not touch any of the bacterial colonies on the plates. These plates look really nice, so take some pictures if you can.
Having completed this set of experiments you have determined whether you carry S, aureus in your nose or not.
Hope that helps you, drop me an email if you have any other questions and good luck!!
Sarah
Dr. Sarah E. Maddocks MSB CBiol
Lecturer – Microbiology
Cardiff School of Health Science
Cardiff Metropolitan University
Western Avenue
Llandaff
CF5 2YB
