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Of late a lot of hubbub has brewed over Richard Lenski’s latest paper on the laboratory evolution of citrate-eating E. coli (PNAS 105(23):7899-7906 (2008)). Lenski figures very prominently in Carl Zimmer’s overview of E. coli research entitled Microcosm, and for good reason. For those that don’t know of the content of the current paper, for the last 20 years Lenski and his students have been growing bacteria (starting from a single bacterium and creating 12 lines) and periodically freezing samples (every 500 generations or so) to see if these bacteria evolve in the lab. At the time this paper was published, over 44,000 generations had passed.

Periodically, the frozen ‘fossil’ bacteria can be thawed and DNA compared over time. Since the bacteria are cloned and reproduce asexually, only natural selection and genetic drift can act on the gene populations, greatly simplifying analysis. Previous to this paper, changes in growth rates, reduced lag phases when the bacteria were transfered to a fresh culture medium, reduced peak population densities and larger cell sizes relative to their ancestors were reported. Most populations evolved increased DNA supercoiling.

So, as expected, things changed over time. But something new happened that surprised the researchers. Under normal circumstances, E. coli does not take up citrate and use it as an energy source (rare cases have been reported where populations of E. coli have acquired a plasmid from other citrate-using bacteria allowing them to do so, but this could not have happened in this experiment), but they do secrete it into the medium. Thus, one would expect that bacterial populations which could take up citrate would have an advantage over the wild type variant which can not. Lo-and-behold, after 31,500 generations one of the populations suddenly developed the ability to uptake citrate and the gene which allowed this very rapidly spread throughout the population. But this Cit+ variant did not make the Cit- one extinct within the population, but coexisted with it.

The Cit+ variant could not be the result of a single mutation. Why not? Because the number of generations passed coupled with the rate of DNA copying errors would result in more mutations than the number of base pairs in the entire genome (4.6 million bp’s) of this strain of E. coli. Thus, if a single mutation imparted this kind of advantage, it would have manifested itself long since.

Two competing hypotheses were tested: the first – that Cit+ arose through a rare mutation, one that does not scale with typical mutation rates; the second – the mutation is ordinary, but the expression is contingent on prior mutations in the population. It is tempting to speculate that this is exactly what occurred with the development of a nylon-eating strain of Flavobacteria. In this case it was a duplicated gene coupled with a frameshift mutation.

The long-term evolution experiment design allowed the researchers to attempt something that Stephen Gould suggested couldn’t happen – to rewind the tape of life and watch the same course of evolution happen. Thawing out samples of the ‘fossil’ bacteria, they repeated the same experiment with that bacterial line. Samples from the 20,000 gen mark became Cit+, whereas samples from prior to this point did not become Cit+! Thus, some mutation happened that prepared the way for becoming a Cit+ variant. It did not in and of itself cause the bacteria to become Cit+, but was necessary for it to happen and greatly increased the chances that the bacteria in that line would go on to become Cit+. Gould was (at least to this extent) wrong

The results of their experiments suggest that it was not one, not two, but three mutations which occurred to produce the Cit+ variant. The first, as noted, occurred around 20,000 generations. The second at around 31,000 generations (which, in fact, slowed citrate uptake). The final one was at 33,000 generations.

Michael Behe is on record in his book The Edge of Evolution as saying that when more than one mutation needed to produce an evolutionary advantage, the likelihood of occurrence is vanishingly small. (Sidebar: I have yet to see Behe ever mention ‘genetic drift’. The first of the two mutations need only be benign or even mildly disadvantageous – genetic drift can spread such a gene through a small population like wildfire. Natural selection is not the only mechanism through which allele frequencies can be increased.) Of course, Behe never does any experimental work to back up his claims. He has a long history of this. In Darwin’s Black Box he claims that the blood clotting cascade is irreducibly complex, yet performs no experiments to show that this is so. And coming up with an experimental design is rather easy for this one. Just knock out a gene, two, three, whatever, encoding the proteins in the pathway in mice. And this has been done. Guess what? Blood still clots. Not as well, but that is to be expected. What is not expected by Behe is that it clots at all. Of course, there are already animals which have incomplete (or reduced, I would say) cascades: dolphins and whales.

Why doesn’t Behe do any research before writing such nonsense? Here’s his answer:

‘I myself would prefer to spend time in what I would consider to be more fruitful endeavors.’

Mind-boggling.

Anyway, here is what Behe has to say about the paper by Blout et al:

‘I think the results fit a lot more easily into the viewpoint of The Edge of Evolution. One of the major points of the book was that if only one mutation is needed to confer some ability, then Darwinian evolution has little problem finding it. But if more than one is needed, the probability of getting all the right ones grows exponentially worse. ‘If two mutations have to occur before there is a net beneficial effect — if an intermediate state is harmful, or less fit than the starting state — then there is already a big evolutionary problem.’ And what if more than two are needed? The task quickly gets out of reach of random mutation.’

In the immortal words of Jon Stewart, ‘Waaaaaaa?’ As PZ Myer rhetorically asked, did he even read the paper? There were three mutations! And again, either Behe is unaware of the concept of genetic drift (unlikely, though possible) or he is being intellectually dishonest like all the other IDiots. You pick. Neither is other than contemptible. It’s no wonder Behe’s colleagues at Lehigh have officially disowned him! They want nothing to do with the guy. How the heck did he get tenure?

Enter the YEC lawyer. The latest bruhaha involves Conservapedia creator Andy Schlafly (you already know from that intro what a nutjob he is) in which he demands to Lenski to pass over his data to him and his ‘experts’! Schlafly cites PNAS’s objective to transparency in the peer review process, which Schlafly seems to think means he can have access to the lab notebooks!!! The data, as Lenski was kind enough to point out, is spelled out in the paper, as is their methodology. Everything which is necessary to reproduce the experiments and compare data between the two is there! What does Schlafly think he will do with the original data even in the event that he understands even a tenth of it? This is unlikely from a guy that I would expect to consider feces throwing the height of intellectual discourse. His demand to ‘make the key underlying data available for independent review’ is farcical. I wouldn’t trust that guy to sit the right way on a toilet seat!

It is quite clear from Schlafly’s communication that he is calling Lenski’s research fraudulent but hasn’t the balls to actually come out and make the accusation. Does Schlafly see something in the methodology that he thinks is odd? Is there something in Schlafly’s own experiments which show that these results could not possilby be correct? Oh, wait… Schlafly’s only experience with E. coli is intimately involved with feces-flinging, isn’t it?

No, Schlafly’s problem is that he doesn’t like the implications of the research and so it must be wrong. Truth be damned – for Schlafly, it’s all about Schlafly! What a maroon! Like Behe, it is quite obvious from this correspondence that he hasn’t read the paper either. He figures that because he is a ‘tax payer’ and that he in part funded the work that he has the right to the actual laboratory notebooks. I don’t think so. Besides, it would be like a monkey trying to understand Shakespeare.

Undaunted, Schlafly wrote directly to PNAS in September and in his whining tone pleaded with the editors to publish it. After pointing out that only letters with content “of wide interest to the readership of PNAS or should illuminate some obscure or subtle point” (in other words, something useful), the editor followed with one hell of a zinger:

‘The issues raised by Mr. Schlafly are neither obscure nor subtle, but are part of everyday statistical analysis at a level too elementary to need rehearsal in the pages of PNAS.’

(Sorry for the links to such an idiotic website as Conservapedia. Feel free to apply disinfectant to your computer and eyes to get rid of any contamination of the stupids….) Ouch! Everyone has a right to be heard on an issue, but with Schlafly’s complete lack of expertise in any area of this research he has no expectation of being taken seriously and the tone of the response (which was in the same vein as flicking a fly off the arm) correctly reflects this.

I suggest going to that last link I provide periodically. The correspondence seems to be piling up, though I would truly be surprised if Lenski continues the conversation. I sure wouldn’t.

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One Trackback/Pingback

  1. […] An Instance of Historical Contingency Observed in the Laboratory I’ve written on my own blog (Shamelessly Atheist – perhaps I should change it to ‘Shameless Self-promoter’) on the observation of the ability of E. coli to suddenly utilize citrate as an energy source by Richard Lenski. Published in the Proceedings of the National Academy of Sciences, the work has received more attention from the anti-evolution crowd than I have ever witnessed outside of The Origin itself. The infamous creationist lawyer Andy Schlafly, creator of the somewhat less-than-objective version of Wikipedia, Conservapedia, objected strenuously to the conclusions of this experiment and in a comical farce has tried to obtain the original data for the experiment I will be describing presently. But you can read about that in my blog on the matter. […]

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