Re: how do I do simple incubation

Date: Mon Nov 28 07:53:18 2005
Posted By: Lynn Bry, MD/PhD, Dept. Pathology, Brigham & Women's Hospital, Harvard Medical School
Area of science: Microbiology
ID: 1133043917.Mi

Message:

Hi Travis-
There are a few things you can try to get the bacterial species in your mouth (the"oral flora") to grow. Your normal mouth flora consists of quite a variety of organisms. While some are areobic (like to grow in our oxygen-containing atmosphere) or "facultatively anaerobic" (can grow in the presence or absence of oxygen), many are anaerobic (cannot grow in the presence of oxygen gas), and so you will not be able to isolate these.

You should be using a nutritious medium such as tryptose soy base agar (TSA), Sheep's blood agar, chocolate agar or Todd-Hewitt agar.

Incubation at body temperature, 37°C (98.6°F), provides the optimal temperature at which oral flora grows. However, some species will be able to grow at room temperature (25°C), just a bit more slowly.

Many organisms like to have an atmosphere rich in CO2, streptococcal species in particular. Your mouth is filled with what we refer to as "Viridans streptococci", some species of which are involved in causing tooth decay. You can create a CO2-rich environment by creating a candle jar in which you'll culture your plates. Here's what you'll need for your jar:

A large sturdy, sealable metal tin or large glass jar with an air-tight screw-on lid. The container needs to be wide enough and tall enough to hold your plates and still have 2-3 inches of room to spare. Large tins that held popcorn can be good for this, or large glass jars that held pasta sauce. Don't use plastic containers, as they might burn.
A small tea or votive candle - the kind that's about 1/2 inch high and formed in a small metal container.
Matches.

Streak your plates and label them with the date, source of material and who streaked them out. For more information on how to streak plates for isolated colonies, try the information at How to streak an agar plate.
Place your plates in the jar or metal container.
Have an adult help you light the candle and place it on top of the plates in the container, or on the side in an open area where it won't contact anything.
Seal the lid immediately.

If you have a glass jar, you can watch the candle burn for a bit, then extinguish - it does this once the oxygen supply has gotten too low for it to continue burning. The oxygen is not completely gone, just much lower that in the air we breathe. In fact, you couldn't survive on the O2 left in the jar, but many species of bacteria find this environment quite welcoming! The burning reaction also increases the concentration of CO2, which will improve the conditions for isolation of streptococci.

If you can find an incubator at 37°C, that would be the best plate to put your candle jar. Incubation overnight, for 18-24h should give you reasonable growth. If you can't find an incubator, you can try leaving the jar at room temperature for 2-3 days, then check the growth.

Hope this helps..

-L. Bry, MadSci Admin

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