'Evolution under a microscope' going strong at MSU

Evolution is usually too slow to observe in a human lifetime. But in a Michigan State University lab, it plays out in days, years and now decades.

Beginning in 1988, MSU Hannah Distinguished Professor Richard Lenski shepherded the rapidly growing model organism Escherichia coli, or E. coli, through 75,000 generations. Lenski and researchers in the Department of Microbiology, Genetics and Immunology, or MGI, have witnessed evolution in real time for more than three decades.

Called the Long-Term Evolution Experiment, or LTEE, 12 small flasks of E. coli offer a glimpse behind the curtain of evolution — without the need to wait thousands or even millions of years. It’s a simple system that scientists around the world have learned from as they try to understand bacterial evolution in more complex environments, like within the lungs of a person battling cystic fibrosis, or inside the gut microbiome.

In 2022, the LTEE left MSU’s campus and was placed in the hands of Lenski’s former postdoctoral researcher, Jeffrey Barrick, at the University of Texas at Austin. His group continued the experiment there to 82,000 generations. Now, the E. coli are home again.

Barrick returned to MSU as a Hannah Distinguished Professor with joint appointments in MGI and the Department of Entomology. With his return, the LTEE is back at MSU, still ripe with possibilities. Barrick and Lenski say a new species could even emerge one day.

“I think this is part of continuing a long legacy of excellence in microbial evolution and ecology at Michigan State,” Barrick said. “We are historically strong in microbiology, including studying microbial communities associated with plants and animals.”

It all started with 12 flasks

Lenski started calling his 12 flasks of E. coli a long-term evolution experiment after about 10 months, when it reached 2,000 generations. At the time, that seemed like a lot.

Before he began studying bacteria, he was a field ecologist, but he grew tired of spending hours in the woods setting beetle traps and counting populations only to get seemingly straightforward results. He wanted to watch evolution happen in real time. That’s when microbes entered the picture.

Bacteria reproduce quickly, allowing scientists to watch entire generations be born and grow up in hours, rather than years. He realized that if he studied non-pathogenic E. coli, a common model organism, he could witness thousands of generations in under a year.

There was just one problem — Lenski wasn’t a microbiologist. Not letting that little detail get in the way, he wrote to Bruce Levin, an evolutionary biologist who had a lab at University of Massachusetts Amherst at the time. Before he knew it, he was a postdoctoral researcher in Levin’s lab.

“I was very worried about whether I could do microbiology and learn all the pipetting and basic skills,” Lenski said. “I quickly learned it was a lot of fun.”

Studying bacteria was as close to instant gratification as it got in evolutionary biology. Lenski could see changes in the number of E. coli cells in petri dishes from the day he started an experiment to the next morning. The more he learned, the more he kept coming back to the same question — can you repeat evolution? How does the tension between the power of natural selection and the randomness of mutations impact change over time?

That’s when Lenski had the idea to start an experiment.

“I thought, why not simplify things and just let the bacteria evolve?”

The experiment began in 1988 with 12 lineages of E. coli. Every day, he filled 12 flasks with fresh glucose medium and used a pipette to transfer 1% of each population into their new home. Then, they were left alone to grow until the next day’s transfer.

Day by day, the experiment went on. Lenski, first a faculty member at University of California Irvine, came to MSU in 1991, bringing the experiment with him.

With his arrival, Lenski joined the Center for Microbial Ecology, headed by University Distinguished Professor Emeritus James Tiedje. There, Lenski found an environment where microbes were viewed as real organisms, not sacks of DNA and other molecules, as many biologists tended to use them back then.

Unexpected evolution

Keeping the experiment going required constant care. Lenski and his lab didn’t miss a day, coming in on weekends and even holidays to keep the population growing. Every 500 days, they added an agent to protect the cells from damage and froze them at minus 80 degrees Celsius. Those frozen samples became a fossil record that scientists can revive to compare their progeny for generations to come.

The routine was predictable, but the evolution unfolding in the lab was not. Lenski and his colleagues assumed each beneficial mutation would drive its competitors extinct. Instead, they noticed several branches with different mutations often coexisted for long periods.

“One of the exciting things about the long-term lines is that every result we get raises as many questions as it answers,” Lenski said. “I like to call it the experiment that keeps on giving.”

At first, the bacterial cultures all evolved in similar ways. They grew larger than their parent cells, and they all fine-tuned how their DNA was coiled. In some lineages, the hot-dog shape of their ancestors morphed into something rounder. The bacteria also learned to spring into action as soon as they were added to fresh medium, instead of taking about an hour to wake up as the ancestor did.

Then one day, one of the flasks developed an unusual appetite.

Glucose is the primary food source for the LTEE cultures, but the flasks also contained citrate to help with iron availability. Normally, E. coli cannot consume citrate except under special circumstances. But in 2003, after about 31,000 generations, a group in one flask evolved to use citrate as a food source.

“It’s surprising, because there’s a huge advantage when they evolve to do this,” Barrick said. “It’s like their own private nutrient that’s there at a much higher concentration than they would normally have. The flask becomes noticeably different, with more cloudiness from the cells, which you can see without a microscope. It was a big innovation.”

Lenski’s lab watched as the citrate-eating population grappled with its newfound ability. Even though they had a private food source, they struggled to consume it efficiently. Their metabolism, which was used to running on glucose, faltered when it tried to produce its cellular building blocks from the new fuel.

Applications beyond the lab

Over time, scientists beyond Lenski’s lab used his experiment as a conceptual framework for thinking about microbial evolution in more complex environments, such as the human gut or the lungs of someone suffering from cystic fibrosis. They asked if the twists and turns of evolution within those flasks could explain what they saw in real world infections.

The LTEE is widely emulated in the biotechnology field too. E. coli is commonly used as a cell factory to produce enzymes, medicines, and ingredients for plastics and other materials. Scientists can use evolution to make those factories more stable and efficient.

“In a way, the long-term experiment has become a model system of evolution,” Lenski said. “It’s super simplified. It gives us a chance to dissect evolution and say, where are our misconceptions? Where are we seeing things that are unexpected? Where are things going as we expected. And then, other people can push this in new directions, like the lungs of people with infections.”

Not slowing down

As the LTEE approaches 100,000 generations, researchers are still looking to the flasks with new questions and new applications. Now in the hands of Barrick and his lab, the LTEE inspires their studies of the gut microbiome of honeybees and in the emerging field of synthetic biology.

Barrick and his team will continue the daily transfers as they monitor the populations for the next innovation.

This story originally appeared on the College of Natural Science website.