This experiment hopefully will finish Aim 1 of my Wild Yeast Project which is to isolate different strains of wild yeast. Aim 2 of the project would be to use these strains in forced fermentations tests to see which strains are ideal for fermenting homebrew. I have tried plating the dregs from other American Wild ales, most notably Russian River’s Temptation and Sanctification. Unfortunately, while these yeast grew fine if plated directly from the bottle, they did not survive the freezing process in glycerol at -80°C.
My attention then started to turn to other sources of wild yeast. Namely, Chad Yakobson of Crooked Stave and the Brettanomyces Project mentioned that Vinnie (brewmaster at Russian River) inoculates his barrels and brews with commercially available strains of Brett. To get something truly unique, he mentioned I should look to lambics brewed in Belgium.
Lambics are spontaneously fermented beers predominantly from the region of Belgium, though spontaneous fermentation could happen anywhere. These beers are made with a significant amount of unmalted wheat in a process known as a “turbid mash”. This results in a wort with a high percentage of protein, allowing for the beer to have a foamy head and distinct wheat taste. The hops that are employed are also unique. Lambic brewers use aged hops that have lost their bitterness potential but not their antimicrobial properties. High IBUs will inhibit the growth of lactic acid producing bacteria which is a strongly desired quality in lambics. The wort is then transferred to a vented attic and placed in large open tanks called coolships (see picture above). This is the critical moment and birth of a lambic – once cooled, the wort is inoculated with local microflora found in the air. Once inoculated, the wort is racked to used wine barrels to finish fermentation (which could take 2-3 years). For more reading on lambics I suggest a great blog called Lambic and Wild Ale.
Fermentation of this style of beer occurs in stages depending on the growth of the fermenting organism. In the first stage, wild yeast (Kloeckera apiculata) enterobacteria contribute little to the beer’s final taste but are important in breaking down proteins in the beer. The second stage is where Saccharomyces cerevisae rapidly multiply and ferment most of the sugars into alcohol and some esters. In the third stage, lactobacillus becomes the focus and produces lactic acid giving the beer a twangy sour character. The fourth and final stage involves numerous strains of Brettanomyces which give lambics the characteristic funky character. It is the yeast from this final stage that I am interested in and can be found in commercial lambic bottles.
I utilized the beer trading forum over at Beer Advocate to hunt down a great lambic from Belgium. While I could get Belgian lambics from off the shelves, I wanted something unique and hard to find. I traded for a bottle from the famous Belgian lambic brewery, Brasserie Cantillon. While I originally traded for a bottle of Cantillon’s famed Bläbær (a lambic brewed with blueberries), the generous trader found out about my project and threw in a bottle of Lou Pepe Framboise from 2004 as well. This lambic was brewed with raspberries and is wonderfully tart with an aggressive sourness and classic barnyard notes. I have tried a bottle from 2006 which was less astringent, suggesting that the yeast present in the bottle are still active and will sour the beer for years to come.
I shared the bottle with some friends and isolated the yeast sediment at the bottom of the bottle. I used a sterile 10 ml pipette and transferred the dregs into a sanitized mason jar. As of now the dregs are still in my fridge but I will bring them in the lab for testing after the July 4th holiday. Here will be my protocol for plating:
- Transfer to a sterile 50 ml conical.
- Centrifuge the sample at 1000 rpm to clarify yeast from raspberry material. If there are multiple bands resulting from the centrifugation, I may need to isolate each of those separately.
- Count cells under a microscope
- Dilute cells enough to plate on selective media (might need to dilute to 100,000 fold).
- Plate on WLN/WLD and MYPG with and without Bromocresol green.