Wild Yeast Isolated From Cantillon Blabaer

I have been wanting to post this for a while but research, parenthood, and preparing for the yeast class this coming Wednesday, has been keeping me busy. Two posts back I mentioned that I was able to isolate some wild yeast from the dregs of a bottle of Cantillon Blabåer (2010 vintage).

To recap, I diluted the dregs to get single colonies plated on regular MYPG agar but I saw no growth. I then plated straight dregs form the bottle and also saw no growth for two weeks. I found this to be strange since an earlier experiment with Lou Pepe dregs at least grew coliform bacteria. I then decided to inoculate a very small amount of Blabåer sediment into 10 mls of sterilized wort and after four days in a shaking incubator I saw growth.

I plated what grew onto three different selective agar:

  1. WLN media – allows for the growth of anything
  2. WLD media – contains cycloheximide which blocks S. cerevisiae growth, but not Dekkera/Brett
  3. MYPG + Bromocresol green - S. cerevisiae  will not metabolize the dye and colonies will look green.

Check out the Wild Yeast Project main page to see the preparation and controls for the above mentioned growth mediums.

Here are my results:

WLN media

WLD media

MYPG + Bromocresol Green

A couple of important things to note. First, the wild yeast that grew did not obey the selective media similar to Brett B and Brett L. Specifically, this yeast grew less under cycloheximide selection and did not metabolize the dye. They looked like S. cerevisie. Secondly, the colonies are almost uniform. This suggests that one yeast strain dominated the wort culture. There is an exception to this however, as one (yes one!) yeast cell grew into a colony that looked different from the others:

Next, I picked two colonies, one normal one and the different colony, and looked at the cells using an 100x immersion oil objective. For simplicity, I named the flower-like colony CB1 and the other colony CB2.

CB-1

CB-2

Conclusions:

These two strains definitely look like wild yeast, especially CB-2. The long cells are reminiscent of Brett L. It is interesting to note that the CB-1 does not have the same cellular morphology (elongated cells) as CB-2 and suggests these are two independent strains. Only PCR to look at genetic markers will identify these strains as different and wild. Another curiosity is why I got no bacteria in this culture but the opposite in my Lou Pepe experiment. Whatever strain(s) this wild yeast is, it outcompeted all other wild yeast and bacteria. Also, the wild yeast isolated behaved just like S. cerevisie. The yeast were largely inhibited by cycloheximide and did not metabolize the Bromocresol green dye. Chad Yakobson of the Brettanomyces project and Crooked Stave has mentioned that non-commercial wild yeast might behave completely different since hardly anything is known about them. For now I’ll consider this a success in isolating wild yeast and brewing a batch of homebrew with this yeast is in my future!

The next step in characterizing these strains are some forced fermentation tests.

21 Comments

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21 responses to “Wild Yeast Isolated From Cantillon Blabaer

  1. CB-2 looks like Brett. L I pulled out of WY Old Ale Blend. Then again you can never tell with Brett because the cell morphology is anything but consistent.

    • Yeah I agree, anything but consistent. Even wild yeast selective agar is inconsistent. Though Chad Yakobson of the Brett Project suggests that not all wild yeast will follow a pattern when plated. I have yet to make CuSO4 plates however.

  2. Oh also do you have any particular genetic markers in mind? Seems like an awful advanced way to differentiate strains on a homebrew level. Though I admit getting oligos and PCRing Brett stains to tell the difference is about as cool and advanced as homebrewing gets!

  3. Pingback: Wild Yeast Isolated from Cantillon Blabaer! - Home Brew Forums

  4. Looks awesome Jason! Both are certainly Brett strains. and they seem to the Brettanomyces L. type strains (although only B. bruxellensis exists as all lambicus are now reclassified.. geneticist always mucking things up..)

    Re streak the single colonies which differ in morphology so you can get pure colonies, even though you have great uniformity now and can see the colonies very well! Then let the colonies grow for a while. You’ll get some very beautiful large colonies. Or just let what you have keep going. Also you may have more strains. All the small colonies on some of the first photos could grow up and be a different strain. Also looks like lots of small colonies in the WLN photo?

    Cheers,
    Chad

    • HI Chad! Good to hear from you.

      That’s what thought too about Brett L. I hope, however, that this is not the same strain that is commercially available from Yeast companies.

      You know – the small colonies on WLN could very well be another strain. I still have the plate so I’ll take a look.

      Let me know if you would like any of these strains. I can always ship them to you on dry-ice as frozen stocks.

      Jason

      • Chances are pretty good it’s not a commercial strain that the labs already cultured. I think of all the “wild” strains I collect as unique. Best way to find out is to compare it to the others if you get a chance just so you know what they look like. I might have some photos of the lambicus type strains at http://www.brettanomycesproject.com

        Check the plates out over the next couple of days..

        I’m actually not a fan of freezing or even storing Brett in the cooler. Seems too finicky.. If it survived for the 3 plus years in a barrel and however long in your bottle, why not store it at room temp? Thats how I store my strain in liquid culture at room temp. I know it goes against the norm but not everything is text book. If you did want to ship, ship’em standard maybe in some solution.

        I’ve got more bottles for you as we have released a few more since we last spoke… New caps too ;) Have you cultured any Crooked Stave bottles yet?

      • I’ll have to agree on storing Brett in the freezer, at least at -80C. I isolated Brett from Santification and Supplication and froze down what grew. Unfortunately, the viability went to zero… I’m have plates and cultures at 4C so it should be good for a while.

        I haven’t had time to culture WWB yet, I still have a bottle in my fridge. I opened one bottle that did not leak at a tasting and it got lots of great comments. I’ll make sure to plate the last bottle and will let you know when I do.

        And yes! Send me more tasty brew from Crooked Stave!! ;)

        Give me your address and I’ll send you yeast.

        Jason

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  6. Thank you for your great work. I really appreciate it. I just looked at bugs from a dreg and got similar results as you did (http://eurekabrewing.wordpress.com/2012/01/13/microscopy-pictures-of-bfm-la-torpille-dregs/). It seems that I isolated B. lambicus as well. And I store my wild dregs in bottles as well and put them into the refrigerator to slow down the bacterial growth.

    Samuel

    • Samuel,

      Thanks for the comments. Actually, taking a closer look at your pics makes me think you have something else. I’ll comment more on your blog, which is great by the way. There is not enough homebrewers doing the stuff we do!

      J

      • Hi Jason,

        It is me again. A while back I talked to you about looking for wild yeast in the Arizona desert. It seems that I have found some. To my surprise they appear to be Saccharomyces. My initial thought is that it appears to be a good flocculator as well. I will try to reduce the quantity of bacteria in a high IBU wort and cool fermentation, then plate to isolate. I am still working on my plating technique with White Labs and Wyeast strains I have around the house. I have also been photo documenting any strains that passed through my house. The results can be found on my Flickr page.

        http://www.flickr.com/photos/brydoty6/

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