This Week's Citation Classic: The Fluctuation Test

Luria S. and Delbruck M. 1943. Mutations of bacteria from virus sensitivity to virus resistance. Genetics 8: 491.

This week's citation classic comes from Nobelists Salva Luria and Max Delbruck and is one of the most famous experiments in biology. Luria and Delbruck wondered about the nature of mutations. Are mutations spontaneous? Or do they occur in response to environmental conditions? The latter view, common to scientists of the day (e.g. Cyril Hinshelwood), was one of the last vestiges of Lamarckism in evolutionary biology.

Since the time of d'Herelle, it was known that a culture of bacteria exposed to bacteriophage would eventually become clear, as if all the bacteria in the culture were killed. However, eventually the culture would grow cloudy again. It was surmised that the bacteria acquired resistance to the phage, and were able to repopulate the culture. The question was, how can the system be used to demostrate the role of chance in mutations?

Luria struggled with the problem for several months, trying to devise a test to show that mutations were spontaneous. Then at a faculty dance at Indiana University, Luria had his eureka moment.

"During a pause in the music I found myself standing near a slot machine, watching a colleague putting dimes into it. Though losing most of the time he occasionally got a return. Not a gambler myself, I was teasing him about his inevitable losses, when he suddenly hit a jackpot... gave me a dirty look at walked away. Right then I began giving some thought to the actual numerology of slot machines; in doing so, it dawned on me that slot machines and bacterial mutations have something to teach each other." (From Luria's autobiography: A Slot Machine, A Broken Test Tube).

Luria returned to the lab and set up a large number of bacterial cultures containing just a small number of bacteria in each, to which he added some bacteriophage. Luria reasoned that if mutations were spontaneous, then their distribution would resemble jackpots. Here the number of surviving bacteria would be small in most cultures, but large in a handful. On the other hand, if mutations were directed as the Lamarckists supposed, then their payoffs would be evenly distributed. Each culture would contain a small number of mutants, as the figure here shows:

Luria and Delbruck's experiments showed unequivocally that mutations were spontaneous and emphasized the role of chance and historicity in evolutionary biology, thus putting the final nail in the coffin of Lamarckism. See Fig. 2 from L&D's paper where the number of jackpots (>9 resistant bacteria) far exceeds that expected by chance. The reason I selected the Fluctuation Test as this week's citation classic is because of a recent exchange between Rich Lenski and I, of which I reprint portions of here:

"I've always been fascinated by the tension between chance and necessity, between randomness and repeatability. As a kid, for example, I especially liked games with dice and cards that involved both luck and skill.

Then, when I was at Oberlin College, I took a wonderful course in which we used Gunther Stent's "Molecular Genetics: an Introductory Narrative" as a text. Unlike most science textbooks, it emphasized the history of who did what experiments and why. I remember reading about the "fluctuation test" performed by Salvador Luria and Max Delbruck, and trying to make sense of it, and then having that eureka moment when the whole point of the experiment hit me. It's my all-time favorite experiment and to this day, whenever I think about it, I'm struck not only by its elegance, but also by the subtlety of the interpretation and by an appreciation of why the problem had been so difficult until they did their experiment.

As you know, a main focus of the long-term evolution experiment with E. coli has been to better understand the repeatability of evolution that arises from the tension between random mutation, on the one hand, and the systematic process of natural selection, on the other hand, that pushes populations toward greater fitness in the environments in which they live. So in a way, you might think of my long-term evolution experiment as a descendant of the fluctuation test, one that examines the role of random mutation in producing statistically quantifiable variation between replicate lineages, not in overnight cultures but across, now, more than 40,000 generations of evolution."

It is precisely this randomness of evolution that led to Lenski's and colleagues latest discovery that, after 33127 generations, a strain of E. coli evolved the ability to digest citrate. Carl Zimmer does a bang up job of covering that story.

Lenski also had a recent dustup with the IDiots, and his tolerant response is covered here.

Update:P Jonathan Eisen of Tree of Life also wrote about this paper here.

Photo: Max Delbruck, Salvador Luria, and Frank Exner at Cold Spring Harbor Laboratory from the National Library of Medicine.