Ideal Method for swabbing surfaces and plating samples - (Jan/18/2010 )

Hi,

I was wondering if anyone has experience in surface swabbing with the purpose of identifying all bacterial contaminants present.

This is part of a research i am going to start. I will be swabbing surfaces using sterile swabs moistened with sterile water. It will be done in a hospital setting so i will most likely be finding common nosocomial pathogens.

Now, a question i have been asking myself is:
1. Is it ok to swab a blood agar plate and a mac plate (and a sabouroud plate) using the same swab, or should each plate be swabbed with a unique swab?
2. Is it more practical to plate all swabs initially on blood agar and then subculture on mac OR is it more accurate to use the 2 plates (will blood agar ensure growth of gram negative organisms present?)


Hope someone can provide me with some feedback plz

I would use the same swab for all plates. I used to work in public health as a microbiologist and that's really best, although of course you'll have a lot of overgrowth

also, this is important - if you use only sterile water, you will get lysis. I'd recommend methodology more or less like this, if you want to get ALL the bacteria (and please note you'll still miss many things like mycobacterium, anaerobes, and things that are very picky if you don't use the correct media to allow their growth - C difficile is fairly common in hospital environments and the Clostridia are anaerobic)

I'd use a sterile swab and aseptically dip first into something like THB to enrich slow-growers. then I'd dip the same swab into sterile SALINE (0.8% NaCl in water) and mix the daylights out of it. make 3 or 4 10^-1 dilutions in saline. use these to streak your plates. the dilutions are because you might get growth so heavy you can't make out individual colonies.

oh, and if this is homework, there IS a subforum for that.

First this is not HW!

I am a grad student undergoing a research. I am currently working with infection control in a hospital. I wish that you had replied a little sooner, bcz thats great advice. Unfortunately,I began my first cycle 2 days ago: I used sterile water to moisten my swabs and directly plated them directly on three types of agar (Blood, Mac and sabaroud). Out of 30 different samples (cultured on a total of 90 plates), 9 blood agar plates (from 9 different sites) showed a form of growth that ranged from one or two pinpoint colonies to a plate full of small seperate colonies. I was expecting to recover some gram negative organisms, but not even one MAC plate showed growth...

Can u please elaborate on the steps u gave me: were swabs moistened with anything? Can u please give me all the "boring" little details
PS I am collecting samples after disinfection/cleaning and not before.

Thanks a million in advance