Wild Sourdough Yeast

Isolating wild populations of Saccharomyces cerevisiae from sourdough starters

Outline/Schedule

• Today: Intro lecture and preparing basic sourdough starters
• 6/15: Lecture on Yeast 23-and-Me and species ID by DNA sequencing, and basic cooking with sourdough
• 6/22: Isolating microbes from starters and hands-on species ID from DNA sequences
• 6/29: Microbial analysis and sharing of sourdough recipes Week 1
• 7/6: Microbial analysis and sharing of sourdough recipes Week 2
• 7/13: DNA extraction (from strawberries/kiwis/saliva)
• 7/20: Closing lecture, sharing of results and analysis

What are Yeasts?

• Yeasts are fungi, meaning they’re not plants, animals, or bacteria.
• Yeasts are microbes, meaning that they are single-celled organisms that, at the single-celled level, are only visible with a microscope.
• Yeasts are recognized by their shape and the way they reproduce, by division of a single cell to produce two single cells.
• Yeasts are non-motile, meaning that they don’t have ways of moving around.

Pictures of Yeast (Saccharomyces cerevisiae)

Cells (Electron microscope)

Cells (Light microscope)

Colonies

Saccharomyces cerevisiae is (almost) everywhere!

• “Saccharomyces” means “sugar yeast”
• “cerevisiae” means “beer” (cerveza, anyone?)
• S. cerevisiae loves sugar, and turns it into various other things (alcohol, CO₂ gas, flavorful compounds)
• Humans first started using wild yeast to make tasty and/or alcoholic foods and beverages over 5,000 years ago.
• Bread, beer, and wine strains became “domesticated”, but some people still use wild strains (sourdough!!!)
• Wild yeast is used in coffee and chocolate to convert sugar into flavors

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S. cerevisiae as a probiotic

S. cerevisiae is not normally a resident of the gut microbiome, passes through within a few days of consumption.

S. cerevisiae are bigger than the types of bacteria in your gut

Which thing is a yeast, and which is a bacterium?

Videos of Yeast cells budding

From YouTube video “The Life of Yeast”

What are Chromosomes?

Chromosomes contain genes (DNA) wrapped up with proteins in a compact form.

Humans have 23 chromosomes, yeast have 16

Yeast cells can be haploid or diploid

Haploid: one copy of each of the chromosomes per cell

Diploid: two copies of each of the chromosomes per cell

Haploid or diploid?

Yeast or Human?

Haploid yeast cells can fuse to form diploid cells

Haploid cells are one of two “mating types”, a or alpha.

Only a and alpha haploids can fuse/mate to form diploids.

From YouTube video “The Life of Yeast”

Diploid yeast cells can sporulate to form haploids

When diploid cells are starved in the right way, they can sporulate and produce up to four haploid cells (two a, two alpha) within each diploid cell. This is called an ascus

From YouTube video “The Life of Yeast”

Some haploid yeast can switch mating types

From a to alpha, or alpha to a.

This only happens after the haploid cell has budded at least once.

Now you have cells of opposite mating type! and they can fuse.

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From YouTube video “The Life of Yeast”

The HO gene is required for mating-type switching

• If yeast cells have functional HO gene, then spores can switch mating types after budding, and an isolated haploid spore can produce a colony that contains both haploid and diploid cells.
• If yeast cells have a mutant HO gene, then spores can’t switch mating types, and an isolated haploid spore will produce a colony containing only haploid cells
• Mysteriously, about 1/4th of “wild” isolates carry mutations in HO that prevent mating-type switching!

Germination

Just like with plant seeds, “germination” refers to the process by which, upon re-exposure to nutrients, yeast spores break out of their tough coat and start to grow.

Despite over a century of studying S. cerevisiae

as a model for human biology, there have been shockingly few studies of yeast germination.

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My lab got a 5-year, $1m grant in 2019 from the NSF to study this process: “Molecular and cellular mechanisms of germination in Saccharomyces”

Part of every NSF grant is a “Broader Impact” component that involves spreading knowledge to a broader segment of society...

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Where do Saccharomyces species live in the “wild”?

• Rotting fruit and the legs and guts of insects who feed on rotting fruit

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• Bark of trees with sugary “exudates”

• Wheat?

Yeast isolated from Oaks on my campus

…but all oak samples so far are able to mating-type switch, and (amazingly) all have the exact same DNA sequence of the HO gene! So I want to find other sources of wild yeast…

Let’s go find some wild Saccharomyces!

(from http://robdunnlab.com/projects/wildsourdough/#makeastarter)

Materials:

• Mason jar or other glass jar
• Cloth or paper napkins to use as jar covers
• Rubberband
• Ruler
• Measuring spoons
• Spoon for mixing

Basic Sourdough Starter

(from http://robdunnlab.com/projects/wildsourdough/#makeastarter)

Method:

• Add 2 Tbsp of flour and 2 Tbsp of water into a jar and mix to make a paste
• Mix thoroughly, scraping down the sides of the jar with your spoon.
• Cover the mouth of the jar with a paper towel and secure it with a rubber band. This keeps out any large debris or insects, but microbes do filter in from the environment.
• Place your jar in a warm location out of direct sunlight for 24 hours before moving on to the next step.

Basic Sourdough Starter

(from http://robdunnlab.com/projects/wildsourdough/#makeastarter)

Refreshing/feeding your starter:

• Remove the paper towel and use a spoon to mix your starter thoroughly.
• Smell your starter. This may sound odd, but by giving your starter a good sniff each day, you are training your nose for the subtle shifts happening in the microbial community. Use our Sourdough Aroma Wheel as a reference.
• Remove 1 Tbsp of the starter and dump it into the trash or compost. (or transfer to a new jar to make another!)

Basic Sourdough Starter

(from http://robdunnlab.com/projects/wildsourdough/#makeastarter)

Refreshing/feeding your starter:

• Add 4 teaspoons (1 1/3 Tbsp) of flour and 1 Tbsp of water and mix well, scraping down the sides of the jar.
• Cover the jar with a paper towel.
• Set aside for 24 hours.
• Continue to repeat the refreshing/feeding every 24 hours until you have refreshed the starter at least 14 times.

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Basic Sourdough Starter

(from http://robdunnlab.com/projects/wildsourdough/#makeastarter)

Adjusting the feeding schedule:

After 4 or 5 days, you may notice that your starter reliably rises but then falls again and develops a layer of liquid on the surface within the 24 hour period after feeding.

This layer is called “hooch” and, combined with the reliable rising and falling of the starter, indicates that your starter is hungry. It is time to switch up the schedule! Instead of refreshing your starter every 24 hours, you will need to refresh it every 12 hours instead. Not everyone’s starter will need this.

Science time: Characterize your starter!

(from http://robdunnlab.com/projects/wildsourdough/#makeastarter)

Once you have refreshed/fed your starter at least 14 times, you should have your very own wild sourdough starter(s)! Or, perhaps you won’t. We actually have very little data about how often sourdough starters fail.

So regardless of whether you were successful or plan to throw this one out and try again (or give up on wild sourdough altogether) please help us by characterizing your starter (or failed starter) and submitting your data.

Characterize your starter!

(from http://robdunnlab.com/projects/wildsourdough/#makeastarter)

For some of you, you will finish refreshing/feeding your starter for the 14th time after 14 days (one feeding every 24 hours).

For those of you with particularly hungry starters that shifted to a twice-a-day feeding cycle, you will get to your 14th feeding sooner.

Either way is OK!

Just characterize your starter whenever you have refreshed/fed your starter 14 times and are about to feed it for the 15th time.

Characterize rise time and height

(from http://robdunnlab.com/projects/wildsourdough/#makeastarter)

We want to know how long it takes for your starter to reach its maximum rise, and the extent (i.e,. height) of this rise.

When it is time to refresh your starter for the 15th time, rather than discarding a tablespoon of the starter as you usually do, transfer 2 Tbsp of your starter to a new jar. Depending on how much starter you have, this may be all of it. That is ok, you will still have it at the end. We just want to make sure that everyone is starting with the same amount.

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Characterize rise time and height

(from http://robdunnlab.com/projects/wildsourdough/#makeastarter)

Add 3 Tbsp flour and 2 Tbsp water and mix thoroughly, scraping down the sides. The starter will be a little thicker than usual.

Before setting it aside, draw a line with a sharpie on the jar to indicate the height of the starter.

Using a ruler, measure the height of the starter from the base of the jar. This is your “Baseline” height.

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Characterize rise time and height

(from http://robdunnlab.com/projects/wildsourdough/#makeastarter)

Every few hours, check on your starter and indicate the height with a new mark. Ideally, this would be every 3 hours. If you set it up in the evening, it is OK to leave it overnight. Just check on it first thing in the morning.

Keep checking on your starter every few hours until it is no longer growing in size.

Measure the height of the highest mark from the base of the jar. This is your “High Tide” mark.

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Characterize aroma

(from http://robdunnlab.com/projects/wildsourdough/#makeastarter)

After you measure its height, remove the paper towel lid and give your starter a good sniff.

What does your sourdough starter smell like?

Use the Sourdough Aroma Wheel to the right for reference.

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Take two photos of your starter

(from http://robdunnlab.com/projects/wildsourdough/#makeastarter)

Photo #1: Take a photo of the side view of the starter in its jar against a solid background.

Photo #2: Take a second photo from an aerial view, looking into the jar.

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Submit your data!

https://amyinthekitchen.com/how-to-make-sourdough-starter-recipe/

What’s next?

• Try out various ways to isolate and characterize yeast at home
• Try to exploit household items (Jello, food coloring, Instant Pots)
• Learn about using DNA sequence for identification of fungal species
• Practice using actual DNA sequences for this purpose
• For fun, practice extracting DNA from household items