Culturing wild yeast, identifying yeast

So my first try at culturing wild yeast was less than stellar, to put it lightly. I just did the leave some wort sitting out and see what starts eating it with which I fermented a beer, but it turned out to be an abomination. For my next try I'm going super geek, so I bought some petri dishes, a 100x microscope, some DME and some gelatin. From some instructions I found online I'm going to prepare the wort and add gelatin to make it gel in the petri dish. I'm going to let the petri dishes sit out for a few hours and then cover them to start growing whatever landed on it. The only question I have is, how do I differentiate yeast from other bugs? I don't know if 100x is enough for my microscope to do any good, but it's what I've got. Thanks for any help.

I'm working on this for an independent project. For me, yeast didn't look like much until I Gram stained. They should be circles/ovals and much larger than bacteria and they Gram stain positive. Bacteria will be various shapes and usually smaller than your yeast. Also when they grow on your plates yeast should grow cream/brown and opaque. 100x might be good enough but no guarantees as I found it difficult without 100x and oil immersion.

Is gelatin solid at room or elevated temps? I would order some agar if not.

Thanks for the reply. I'll look into Gram staining since I'm not entirely sure what that is. I have an idea based on the name, but I've never heard of the technique. The site I found said they used gelatin, so that's what I am going off of. I'll do a test before I go full bore and get some agar if it doesn't work.

You should be able differentiate bacteria from yeast with a 40x objective. (With a 10x eyepiece for a total magnification of 400x) Differentiating mold from yeast can be more difficult. The shape of the colony on the petri dish is normally a good indicator. It should be creamy white and uniform in appearance. Next you'll want to select a colony and culture it in a small amount of wort. Make sure it smells right while culturing.

I've got info on my blog with more details.
http://woodlandbrew.blogspot.com/2012/12/isolating-single-yeast-strain-by-plating.html

Yeah you're going to want a pure culture. I'd look into streaking for isolation that way you can select a lone colony and keep plating it until you feel its pure (and see under your microscope).

WoodlandBrew said:

You should be able differentiate bacteria from yeast with a 40x objective. (With a 10x eyepiece for a total magnification of 400x) Differentiating mold from yeast can be more difficult. The shape of the colony on the petri dish is normally a good indicator. It should be creamy white and uniform in appearance. Next you'll want to select a colony and culture it in a small amount of wort. Make sure it smells right while culturing.

I've got info on my blog with more details.
http://woodlandbrew.blogspot.com/2012/12/isolating-single-yeast-strain-by-plating.html

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Thanks. My microscope only goes up to 100x total, so it sounds like I might run into some trouble there, but I'll use your tips about checking the shape and smell of the colony. I appreciate the link.

Mojzis said:

Yeah you're going to want a pure culture. I'd look into streaking for isolation that way you can select a lone colony and keep plating it until you feel its pure (and see under your microscope).

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I'll check it out, thanks.

I've started a similar project, and have a few blog posts on the subject:
http://suigenerisbrewing.blogspot.ca/p/wild-sour-brewing.html#A4

As for your specific question, your microscope should be sufficient to ID yeast (all the pics on my blog are taken with 40X), however, depending on the optical system of your microscope it may be hard to see them.

Yeast/etc don't have a lot of internal contrast, so unless your scope is equipped with phase-contrast, DIC, or other contrast-enhancing optics it will be hard to make out the cells. Wild yeast can take on a range of morphologies - anything from spheres, to lemon-shaped, to strings of cells. They all are, however, much larger than bacteria.

As for identifying what you have, Woodland's post is spot-on; collect your wild bugs, streak out to get single colonies (example of streaking), and then pick/test single colonies fr morphology (under your scope) and small-ferments to see if they make something good.

I'm in the process of setting up a higher-tech way of IDing yeast; its beyond the capabilities of most home labs (I'm a biology prof, so I have toys at work), but if you find something good and want more info on it, I'm happy to accept a sample an run it through my identification system.

Bryan

I am currently trying to get a better way than "wait two weeks and that should be yeast" also.There should be an easier way than PCR. I cultured a bunch from organic apples and now trying to determine which genre they are. I wanted to avoid the PCR and genetics and focus on chemical and physiological identification. Most people don't have access to lab equipment. Hoping that tomorrow a malachite green stain will let me see the ascospores which could really help in identification.

A good book is The Yeasts: a taxonomic study, which is helping me through it. It would be interesting to send samples to you to see if they were correct

Edit: don't mean to hijack

Mojzis said:

I am currently trying to get a better way than "wait two weeks and that should be yeast" also.There should be an easier way than PCR.

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PCR is dead easy, and these days, is actually cheaper than making all of the various selective medias you need for classical identification...if you have access to a machine. At $4K+, it isn't exactly something most people will want to buy for home.

Mojzis said:

A good book is The Yeasts: a taxonomic study, which is helping me through it. It would be interesting to send samples to you to see if they were correct

Edit: don't mean to hijack

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"The Yeasts" is a good book. But almost as expensive as a PCR machine!

Warthaug said:

PCR is dead easy, and these days, is actually cheaper than making all of the various selective medias you need for classical identification...if you have access to a machine. At $4K+, it isn't exactly something most people will want to buy for home.


"The Yeasts" is a good book. But almost as expensive as a PCR machine!

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Only two times I used PCR it took me forever with no clear end product (i'm more of an ecologist). Being a senior bio undergrad I won't have access to the machine anymore so I'm working on finding a way of classical identification. I used cupric sulphate to discourage bacterial growth and it worked out pretty well. I'm just having a hard time figuring out which tests to run when the culture is finally pure. Hopefully the spore stain will help get rid of some possibilities.

That damn book costs almost 400$. Lucky we have a library here.

I have access to it on-line via my work library; otherwise I wouldn't use it. We use PCR all the time in my lab, but the first trial of ribosome PCR is happening tomorrow (Thursday), so we'll see how well those particular primers work soon...

Bryan