Legendary Bacterial Evolution Experiment Enters New Era

E. coli cells used in the long-term evolution experiment (LTEE) on an agar plate.Credit: Jeffrey Barrick

On February 24, 1988, evolutionary biologist Richard Lenski filled 12 flasks with sugary growing medium and seeded each with Escherichia coli bacteria. For the past 34 years, Lenski, at Michigan State University in East Lansing, and his colleagues have nurtured the bacterial cultures, changed the growth media daily, and frozen samples every few months for future research.

The long-term evolution experiment (LTEE) has become a cornerstone in evolutionary biology where researchers continue to seek insights. Over their 75,000 generations, the bacteria have made tremendous strides in fitness — how fast they grow relative to other bacteria — and have developed some surprising properties.

Last month, Lenski and his lab provided the LTEE for the last time. the 12 E coli lines are now frozen in cryoprotective media and will soon be revived to start a new life in the lab of Jeffrey Barrick, an evolutionary biologist at the University of Texas at Austin who first worked on the experiment in the 2000s as a postdoc in Lenski’s laboratory.

The two spoke to Nature about the past, present and future of one of the longest running experiments in biology.

Composite image of two portraits of Jeffrey Barrick and Richard Lenski.

Jeffrey Barrick (left) will take charge of the E. coli experiment from mentor Richard Lenski, who began the experiment in 1988.Credit: Laura Gerson/UT College of Natural Sciences, Sipa US/Alamy

What inspired you to start the experiment?

Richard Lenski: I like big open questions. I wanted a very simple, long experiment to see how repeatable evolution was. The original target was 2000 generations. And I thought that deserved the nickname “the long-term evolution experiment.” I had no idea it would last as long as it actually did – and hopefully will last much longer.

Why did you keep going – and going and going?

Lenski: It’s actually a very easy experiment to keep going. The amount of work for a person in a typical day is perhaps half an hour. It’s 365 days a year, at least in principle, but the amount of work per day isn’t huge.

And then, of course, the bacteria did some really interesting things over time. New technologies emerged, such as the ability to cheaply sequence genomes. People like Jeff came to the lab and brought new ideas and questions. And the bacteria continued to do interesting things. I realized it just had to go on as long as humanly possible.

How many generations had passed when Jeff started the experiment?

Lenski: Did you join the lab around 2007? It was probably about 40,000 generations.

Jeffrey Barrick: Sounds about right. I knew less about the history of the experiment—perhaps less than many of Rich’s postdocs and graduate students. But I was at a point in my career where I had studied evolution at the molecular level, looking at all kinds of bacterial genomes. I wanted to study the evolution of whole organisms in the laboratory and follow the evolution. It’s just something I’ve been fascinated with since my bachelor’s degree.

Was it ever a challenge to keep the experiment going?

Barrick: Like Rich said, it’s pretty simple. There are some big snow storms and other things going on, and other infrastructure anomalies that can make it challenging at times, but you can always go back to the freezer, which is a lot of fun about the experiment and makes it much more feasible than other experiments. Humans have tried long-term experiments with mice and flies and other organisms, where it’s very difficult when something goes wrong.

Lenski: One of the advantages of the long-term experiment is that everything is so simple. We work with a chemically defined medium, we can freeze logs and revive them so that wood knocks, I think it should go on pretty smoothly in the new house.

What did the experiment teach us about the repeatability of evolution?

Lenski: My bias in the experiment was that all species would go in very different directions. I thought the role of chance and contingency in evolution would have been greater than they were. And over the years, we’ve really only seen striking amounts of reproducibility. So while a typical line has improved its relative fitness compared to its ancestor by perhaps 70% or 80%, the difference in competitive fitness between most lines is more than just a few percent. So they are all greatly increased, but very similar.

But over the years, we’ve also seen some rather striking differences between the rules. Thirty thousand generations after the experiment, one of the 12 lines developed the ability to consume citrate instead of just glucose. And that attracted a lot of attention, and even some, shall we say, hostility from some people who are skeptical of the power of evolution. And after 75,000 generations, it is still the only one of the 12 lines to have developed that ability.

Are there any big questions about evolution that you hope to answer by going longer?

Barrick: For many bacteria that have moved into simple, constant environments — especially simple endosymbionts that live in the cells of insects — their genomes gradually shrink over time. And I would say that one of the most surprising things to me is that this E coli have been in a very constant environment, but their genomes have not shrunk very much.

Lenski: I think part of the problem with genome shrinkage is that it’s a slow process. Thirty years and 75,000 generations – it’s a drop in the evolutionary bucket. So I’m guessing that if we could come back — in a million years or whatever, the bacteria would probably have extremely reduced genomes. That’s one reason to keep going.

Photo of Richard Lenski's long-term evolutionary experiment with E. coli, taken June 25, 2008.

The E. coli has now multiplied about 75,000 generations.Credit: Brian Baer and Neerja Hajela via Wikimedia Commons (CC BY-SA 1.0)

Why did you decide to pass the torch to the LTEE?

Lenski: I won’t be around forever. I think it’s better to do those things now, plan them carefully and thoughtfully. So it just made sense. I am 65 years old and although I have no plans to retire in the next few years, the lab is getting smaller and smaller. And one of the important things to keep the long-term lines going is this daily rhythm. I think a lab with half a dozen or more people is ideally suited for the weekend and holiday coverage that the experiment benefits from.

So I asked Jeff, maybe in 2018 or 2019. I have a National Science Foundation grant to conduct the experiment, and Jeff is now a co-principal investigator in that area. And when we wrote the most recent renewal, we set up this plan for the transfer and that it would happen about now.

Why did Jeff record it?

Barrick: I’m a big proponent of open science. This is a great resource that I want to support, share and continue. It has become sort of a common touchstone for many stories of bacterial evolution. And something that people can use in so many ways. I am excited about supporting the community.

Are you afraid to take responsibility for such a long-running experiment?

Barrick: As Rich said, it’s not super hard to keep the experiment going. What gets difficult is to organize all kinds in the freezer, make sure you can send them to people, and all the paperwork that goes into that sort of thing.

I don’t want the experience of the experiment coming to my lab and then getting contaminated, and I’m setting it back two or three years. I want to make sure I’m really moving things forward. But a lot of the pressure is off because it’s frozen, not just in Rich’s lab and my lab, but in France and other places as well. So that takes the pressure off that I could be causing an irreparable problem. So I’m more excited about educating my colleagues about the experiment.

Rich, what’s your advice for Jeff?

Lenski: Keep calm and keep going. Frustrating things will happen. But the experiment is quite forgiving. As long as my brain is working, I’ll be very excited to see what new spin-off experiments he and his collaborators generate, what new analytics he and the wider community generate to understand what’s going on with the long-term lines. Probably the most important thing Jeff will have to think about in 20 or 30 years is who’s next?

Barrick: Your work isn’t done yet, Richard. You are still the best science communicator and person to bring the long-term evolution experiment to people and build that community. That’s actually the most intimidating thing that no one can replace right now. Doing the experiments is very easy.

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