Yeast from the Iron City (YIC): Sampling and Plating

A friend and I decided  a while back to return to the roots of fermentation and try to go wrangle us up some home grown bugs. Well, really they grow on fruits. Ever notice the white powder on grape skins that you can wipe off to reveal a shiny skin underneath? What you just brushed off, in addition to the epicuticular wax of the fruit, were thousands of microorganisms, some of which were wild yeast (Figure 1). Grape skins aren't the only thing you'll find yeast on. Most fruits especially those that we think of as sweet are teeming with microorganisms. The skins of these fruits are acting like natural nutrient agar plates, growing microorganisms on any sugars they can get their hands on. It's pretty much harmless if you happen to eat it, unless you ingest so much that you create a miniature brewery in your gut. Don't believe me? Check out this story on NPR: Auto-Brewery Syndrome. Luckily for you and I, this has only been observed in a single case; so I don't think there is much to worry about.

Figure 1. Grapes with Epicuticular Wax (white powder)

Anyway, we set out to swab different fruits and bring back samples for culture with the following methods. Most of the stuff we used to take samples you can get from a drug store, the supplies for culturing it may require a click over to Amazon, or maybe your LHBS will have some of what you need.

Sample Collection

We used cotton swabs to swab the outsides of several types of fruit. The swabs were then placed into zip top bags or, if you have them pill pouches, (they are almost exactly the right size for a single cotton swab). We did not take the time to sterilize anything at this point because during the culturing step you should be able to form isolated colonies the different microorganisms present on a 4 quadrant plate. Culture media was made up from a recipe found on BKyeasts Blog. Essentially the materials needed are: something to boil your media in (we used a erlenmeyer flask inside a pot), petri-dishes (either sterile plastic or reusable glass) (Figure 2), agar (an extract from sea weed), malt extract (I've found that DME is easier to measure and store when using small quantities), yeast nutrient, and of course filtered tap water or DI water if you have it. You may also want to add in a hop pellet for its anti-microbial power to prevent some growth. Theoretically, the hop acids will help weed out some of the organisms that you definitely don't want. You can read more about that on a previous post IPA: Myth or Microbiology.

Figure 2. Glass Petri-Dishes (Left), 500ml Erlenmeyer Flask (Right)

Why go through the trouble of making agar media instead of making up a batch of dilute wort? One, you will be able to see the organisms you have much more easily on a plate. More importantly, culturing on agar media is the only way you will be able to isolate the yeast you want from all of the other nasty microorganisms that will just ruin your beer, but that is for a later post.

After boiling the malt agar to melt and sterilize it we let it cool to just above the temperature at which it starts to set up. This prevents a large amount of condensation from forming on the lids of the Petri-dish that could drip down onto the media and smear the microorganism colonies around. After letting the plates cool down for a day or so, you are ready for the streaking (of the microorganisms)! Before you begin the process of streaking your culture, you need to make sure you follow aseptic technique. A fancy term meaning keep everything as clean as possible. A diluted bleach solution or other antimicrobial cleaner should be used to clean any surfaces that you may come in contact with, and you should make sure to wash your hands very well. Gloves may be a good idea but are not absolutely necessary (we didn't use them and things turned out fine).

Figure 3. Swab Pattern for Microorganism Plating

For the initial transfer of the samples to the culture media we divided plates up into quarters, labeled them, and brushed the tips of each cotton swab on each quarter of a plate to save space (Figure 3). For now let the plates incubate at room temperature. You should start to be able to see some growth of various shapes, color, and texture. It took a little under a week for our samples to show much growth (Figure 4). Be watchful, after the initial growth started they seemed to grow exponentially leading me to place them in the fridge to slow them down until we could work with them again.

Figure 4. Plated Microorganisms After One Week of Incubation at Room Temperature.

There is pretty much a 100% chance that the samples swabbed will contain more than one type of microorganism (known as a mixed culture), so you will need to use a technique to isolate single colonies of microorganisms so you can pick and choose what you want to take to the next step. I'll write a brief overview of this isolation process and how we did it in a following post but for now, happy yeast hunting!