Plating experiment - Bromocresol green differentiation

This one has been done to death by homebrewers, but I just wanted to see for myself about using bromocresol green to differentiate between Saccharomyces vs Brettanomyces. Also, I received some Cantillon dregs from Nick, and thought that I might plate the dregs out as well.

My usual petris were split into 2 batches - malt extract agar (MEA), and MEA with bromocresol green. MEA was made with 20g/L dried malt extract and 15g/L agar, and bromocresol green was added at a concentration of 0.022g/L. The media were sterilised in a pressure cooker for 30 mins before being poured into plates.

I streaked Brettanomyces bruxellensis "Trois", Wyeast 1056 (American Ale) and the Cantillon dregs onto the bromocresol green MEA plates. This was left to incubate aerobically at room temp for 2 weeks. Oddly enough, while published sources always state that Brett grows a tad more slowly, I find that this particular strain formed colonies quite quickly. By the end of Day 5, both the Brett and 1056 plates had formed nice colonies.

Day 5 - Brett b Trois

Brett is supposed to be able to metabolise bromocresol green, forming white colonies. On the other hand, "domesticated" brewer's yeast strains are reported to lack this ability and will accumulate the dye, resulting in green or blue colonies.

Day 5 - American Yeast (Wyeast 1056)

As you can see by Day 5, the 1056 looks like they are still white. OK, perhaps with the very slightest hint of green, but that's really pushing it. Even a side-by-side comparison at Day 8 yields a rather subjective interpretation. The Brett plate is secreting more acids, turning the bromocresol green into a yellow colour, while the 1056 plate seems to have less of the yellow tinge. Once again, the colour of the colonies themselves are quite difficult to discern.

Day 8 - side by side comparison

However, if you flip the plates over, that's where the results become much clearer. It's now really obvious that the Brett b colonies remain resolutely white, while the 1056 colonies are starting to accumulate the blue dye - at least on the underside, and especially on the margins.

Underside of the plates. Brett on left, 1056 on right.

I think a few more platings would be in order, to see if all brewer's yeasts respond in this way. Looking at the colony colours from the bottom-up may represent a more sensitive method of differentiating between Brett and Saccharomyces. I'd be interested in seeing how early in the course of incubation this difference comes up, as it would be quite promising as a more rapid way of differentiating the 2 rather than waiting for the full 14 days.

As for the Cantillon dregs, unfortunately no yeast grew after 14 days. I simultaneously plated some dregs onto MEA plates treated with nystatin though (that'll come in a future post), and managed to isolate some bacteria out of those plates. I'm thinking I'll try to put the dregs in a starter next time, perhaps to revive the yeasts.

Cantillon dregs on bromocresol green plates - nothing grew