Wild Yeast Project: Lambic Plating Results

How awesome would it be to brew a beer from yeast that was floating around the famed rafters of the Cantillon brewery?

A couple of posts back I decided to isolate the dregs of a Belgian lambic and plate the slurry on selective media that will encourage the growth wild yeasts and bacteria. I opened a 2004 bottle of Cantillon’s Lou Pepe Frambiose and plated the dregs:

1. Transferred dregs to separate sterile 50 ml conical tubes and stored at 4°C.
2. Aliquoted 200 μls to a 1.5 ml centrifuge tube.
3. Made a ten-fold serial dilution of straight slurry all the way out to 1/1,000,000.
• Set up 6 more centrifuge tubes and added 180 μls of double distilled water to each.
• Took 20 μls of slurry and added it to next tube.
• Vortexed and took 20 μls and added it to the next tube (1:10 dilution)
• Repeated until sample was diltued 1/1,000,000
4. Plated 50 μls of the last dilution on MYPG, MYPG + Bromocresol Green, WLN, and WLD (cycloheximide) plates.

Conclusions: In short, I got lots of colonies and I have absolutely no idea what type of wild yeast or bacteria they are. Importantly, if anyone reading this blog (Chad Yakobson, I’m looking at you!) recognizes these colonies, please comment on what I might have.

Some important things to consider with the above results. The MYPG plates with the Bromocresol Green dye had colonies of different color, an off-white or blue. Although this is not true for all Brettanomyces species, Brett is capable of metabolizing the dye to an off-white color. More importantly, some species of Brett will secrete acid and create a zone of clearing around the colony where the blue color changes to white due to the pH change of the acid. Taken together, it seems as if the white colonies are acid-producing Brett colonies that are metabolzing the dye.

As for the yellow colonies, I’m not sure what they are but I suspect they are some species of coliform bacteria.  I did look at the dregs under a microscope and, as expected for a lambic, I saw many bacterial species including rod-shaped bacilli, smaller bacteria reminiscent of pediococcus, and different types of yeast.

Future: I just got this result today and I plan on letting the colonies grow larger. Giant colony morphology is a great way to determine differences in colonies that are very similar when they are small. Also, I plan to look at the cells from different colonies under the microscope and post some images to my blog. Although some colonies are similar, they may look completely different under the scope indicating different species. Of course, I plan on bringing my refractometer into lab and measuring the ability ferment wort under different conditions (aerobic versus anaerobic). I’m going to start by focusing on the colonies that metabolized the dye and generated acid. Hopefully, these will be a new species of Brett that I can use for a future batch of homebrew.