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Location: Los Angeles, California, United States

My biggest motivation for creating my own blogs was to avoid the arbitrary censorship practiced by other blogs and various other Internet forums. Censorship will be avoided in my blogs -- there will be no deletion of comments, no closing of comment threads, no holding up of comments for moderation, and no commenter registration hassles. Comments containing nothing but insults and/or ad hominem attacks are discouraged. My non-response to a particular comment should not be interpreted as agreement, approval, or inability to answer.

Monday, June 30, 2008

A reason for glucose-cycling in Lenski's E. coli experiment

I asked Zachary Blount, the lead author of the paper on the evolution of Cit+ (citrate-eating) E. coli bacteria, if favoring Cit+ evolution was a purpose or one of the purposes of the glucose-cycling (giving the bacteria an insufficient supply of glucose so as to cause alternating glucose feeding and starvation), and he did not answer. What is he trying to hide? Maybe he is trying to hide the fact that the glucose cycling favored Cit+ evolution, because some people might regard that as cheating. Aha -- that must be the reason! Also, he did not answer my question about whether there were other purposes of the glucose-cycling. Now I have found another purpose on my own, and I deserve sole credit for the answer.

Each daily population of the 12 lines of bacteria was raised in 10 ml of medium, and one-percent -- i.e., 0.1 ml -- of each old population is transferred to start the next population. That ratio of the medium transfer allows a 100:1 growth in population to maintain the same final population density in the populations. Assuming a doubling of population with each generation, a 100:1 population growth would be reached in 6-7 (6.64, to be exact) generations. 6-7 generations occur in just a few hours (the populations are started in the morning and the glucose is exhausted by the afternoon). If more generations were allowed, the population density would become too high, which would limit capacity for population growth when the next population is started, so glucose supply is limited to limit the number of generations of glucose-eating-only bacteria. I presume that the 10 ml and 0.1 ml quantities are limited by the physical limitations of the lab equipment, e.g., the incubator size and the means for transferring the medium. If desired, the glucose starvation period could be eliminated by restarting the populations more frequently, but that would make the experiment even more labor intensive and I think it was desirable to have a glucose starvation period, anyway. The medium recipe says that the glucose supply could be varied, and the supply might have been varied to control population growth -- I don't know. The experimental procedure is described here.
.
Also, as I noted before, Blount did not give straight answers to my question about whether Cit+ evolution was a goal of the experiment. If, say, Cit+ evolution was a secondary goal or a longshot goal, he would be misleading people by going around flatly telling them that Cit+ evolution was not a goal of the experiment. In a sane world, Blount's failure to answer my simple, basic questions would be widely condemned -- but this is not a sane world.

Another somewhat minor concern I have is the potential loss of mutations during transfer, but that loss is inevitable. Mutations occurring in one of the last generations in a daily population might exist in only one, two, or just a few bacteria and so would have only a small chance of being in the one-percent of the medium that is transferred to start the next generation.

The glucose supply was unbelievably small -- only 25 mg per liter or just 0.25 mg for the 10 ml medium for each population!

Anyway, what is the likelihood that anything similar to the carefully controlled conditions in this experiment could occur in nature? Even with these carefully controlled conditions that favored Cit+ evolution, the Cit+ evolution was a rare occurrence, occurring in only one of 12 lines of bacteria in 20 years.

Anyway, unlike Andy Schlafly over at Conservapedia, I see no reason to suspect error or fraud in the claims of Cit+ evolution. Lenski et al. claim that they have the Cit+ E. coli bacteria -- which are very unusual -- and also claim that they have E. coli bacteria that have a strong tendency to evolve the Cit+ trait (bacteria of 20,000 generations or later). It would be easy to verify or disprove the claims about Cit+ evolution.
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49 Comments:

Blogger Phae said...

You are an idiot savant, only your skill is being an idiot. So I am not sure what the correct terminology is there. You're unusually gifted at being stupid, I guess is the gist of it, so phrase it however you want.

I asked Zachary Blount, the lead author of the paper on the evolution of Cit+ (citrate-eating) E. coli bacteria, if favoring Cit+ evolution was a purpose or one of the purposes of the glucose-cycling (giving the bacteria an insufficient supply of glucose so as to cause alternating glucose feeding and starvation), and he did not answer.

Blount said that to the best of his knowledge, Lenski intended twenty years ago (when he started the experiment) that the citrate present in DM0 and DM25 would provide a potential avenue for adaptive evolution of the bacteria. He then elaborated by saying that was not the intent - not the goal. He was really clear and obvious, and you even quote those very comments yourself in your asinine first blog post about it. So WHAT PART of that do you NOT UNDERSTAND?! It was understood as a POSSIBILITY but was not the INTENT.

I don't know what you need. Do you only understand things written in crayon?

Maybe he is trying to hide the fact that the glucose cycling favored Cit+ evolution, because some people might regard that as cheating. Aha -- that must be the reason!

It was a POSSIBLE ROUTE FOR ADAPTIVE EVOLUTION, but not the intention. He said that. You read it. You even quoted it, many times.

Now I have found another purpose on my own, and I deserve sole credit for the answer.

Oh, I wouldn't worry about that.

Also, as I noted before, Blount did not give straight answers to my question about whether Cit+ evolution was a goal of the experiment. If, say, Cit+ evolution was a secondary goal or a longshot goal, he would be misleading people by going around flatly telling them that Cit+ evolution was not a goal of the experiment. In a sane world, Blount's failure to answer my simple, basic questions would be widely condemned -- but this is not a sane world.

...you QUOTED HIM. "The evolution of a citrate-utilizing variant E. coli was seen from the beginning as a possible occurrence, and one that would be pretty neat should it occur (and indeed as it has proven to be now that it has happened), but not a goal."

Seriously, what aspect is confusing to you? Is it the dizzying array of letters? The punctuation? Which vocabulary word gives you trouble?

Anyway, what is the likelihood that anything similar to the carefully controlled conditions in this experiment could occur in nature? Even with these carefully controlled conditions that favored Cit+ evolution, the Cit+ evolution was a rare occurrence, occurring in only one of 12 lines of bacteria in 20 years.

The level of ignorance displayed in this comment has been equaled seldom in history. Once, in 1256 when an English peasant transplanted from France named Regineau Monegre said, "They do not zeem to be very fond of us here." And again in 1954, when a young white woman wondered why black people loved the back of the bus so much. But at no other time has anyone displayed such a baldfaced amount of ignorance. You are almost peerless, good sir. I congratulate you.

Okay, a little hyperbole. But seriously, this reveals an enormous amount you don't understand.

First of all, evolution takes a very, very long time. For most complex creatures, adaptations on this order (the ability to primarily utilize a previously useless nutrient) would take millions of years. Even in a bacterium subjected to stress to spur on its Poly IV inclination to mutate, it took twenty years to run this experiment. So it is an incredibly obvious and idiotic statement to note blithely that it was a rare evolution.

Secondly, are you seriously asking what are the chances of a bacterium naturally finding itself in a series of petri dishes with precisely varying quantities of food and no exterior competition or variation? Do you understand what an experiment is? How on earth could it have otherwise been run?

The point of the experiment was to use evolutionary theory and our almost complete knowledge of E coli (and it is almost wholly complete; we have mapped out its whole structure, genes, and so on) and isolate the survival pressure which spurs on evolution to cause it to occur in a controlled and observable environment. An organism was put in a consistent circumstance where it had to adapt if it wanted to thrive, and it did so immediately. Eventually, it adapted so remarkably as to change into virtually another organism altogether. It made an adaptation - a leap forward - that was theoretically known to be possible, but was still brought about by evolutionary pressure.

The experiment was a dizzying, astonishingly complete success. Evolution was observed, and its process was recorded along every step.

Now I have read this whole post. You recount laboriously but without innovation a passable summary of the experiment. What exactly do you claim to have discovered? I want you to state it precisely and simply, please, so that I can link you to exactly where in the papers or in dialog this was already pointed out to you.

Monday, June 30, 2008 7:57:00 PM  
Anonymous Good Commenter said...

"I deserve sole credit for the answer."

Yes, YESSS, hooray!

"... this is not a sane world."

I've noticed that also. How about that!

Tuesday, July 01, 2008 12:06:00 AM  
Anonymous Anonymous said...

Jeff, Man, please resume taking your meds. Now- It is already too late.


I think it would be interesting to watch what would happen if all the fundies, jeffies and their ilk were just given their own state, and had to make everything work all by themselves. No help from rational, open to 'secular knowlege' engineers, doctors or scientists.....

My guess is cannibalism within 6 months.

Tuesday, July 01, 2008 12:33:00 AM  
Anonymous Hector said...

> he did not answer. What is he trying to hide? <

Perhaps he doesn't feel a need to answer every jackass who brays at him?

Tuesday, July 01, 2008 4:39:00 AM  
Blogger Larry Fafarman said...

Phae,
>>>>> Blount said that to the best of his knowledge, Lenski intended twenty years ago (when he started the experiment) that the citrate present in DM0 and DM25 would provide a potential avenue for adaptive evolution of the bacteria. <<<<<

What? Blount said nothing about DM0 -- I presume that means the medium (Davis Minimum broth) without glucose. Your statement is essentially correct about DM25 (with 25 mg/l glucose) -- Blount actually said,

When Dr. Lenski started, he figured the citrate would provide an opportunity that the populations might or might not figure out a way to exploit, thereby presenting a potential point of divergence between the populations (this is my understanding -- I will need to check with him to make certain I understand this properly). (comment #115 on Carl Zimmer's blog)

>>>>> He then elaborated by saying that was not the intent - not the goal. <<<<<<

Can we please stop using the word "intent" here? "Intent" is what one plans to do, but one cannot plan on having an uncertain result.

My printed dictionary defines "goal" as "an object or end that one strives to attain." Basically, a goal is something foreseen as a possible and desirable result of an effort. In a search for the Lost Dutchman Mine, finding it is a "goal." If Cit+ evolution was one of several goals, or a secondary goal, or a longshot goal, or whatever kind of goal of the experiment, then saying that it was "not a goal" confuses and misleads people. Blount actually said,

The evolution of a citrate-utilizing variant E. coli was seen from the beginning as a possible occurrence, and one that would be pretty neat should it occur (and indeed as it has proven to be now that it has happened), but not a goal. (comment #122 on Zimmer's blog)

Someday Blount may really mislead people in a big way by merely saying "it was not a goal" without the clarification, which is likely because he doesn't understand what the word "goal" means. Or someone could quote him out of context as follows: It was "not a goal." I deserve thanks -- not abuse -- for pointing out that the word "goal" is being misused here.

Cit+ evolution might have even been a major goal because of the following factors:

(1) It had been observed once before.

(2) This was a long-term experiment with 12 lines of bacteria.

(3) The glucose-cycling favored Cit+ evolution. Lenski might not have considered this factor -- I don't know. Also, I don't know if this was a factor when Cit+ evolution was first observed.

Finally, as I have already said umpteen times, Blount gave no answer at all to my questions about glucose cycling.

>>>>> An organism was put in a consistent circumstance where it had to adapt if it wanted to thrive, and it did so immediately. <<<<<<

What? The bacteria did not have to adapt -- they already ate glucose, which was regularly fed to them. The term "thrive" is vague -- the bacteria were surviving quite well before developing the ability to eat citrate. And the bacteria did not adapt "immediately" to the citrate -- it took one line of bacteria about 14 years to adapt to the citrate and the eleven other lines of bacteria have not adapted to the citrate in 20 years. I thought that you read Blount et al.'s paper but you know practically nothing about the experiment.

>>>>>> So it is an incredibly obvious and idiotic statement to note blithely that it was a rare evolution. <<<<<<

One of the problems, dunghill, is that a lot of Darwinists don't recognize that it was a rare evolution.

>>>>> Secondly, are you seriously asking what are the chances of a bacterium naturally finding itself in a series of petri dishes with precisely varying quantities of food and no exterior competition or variation? <<<<<

The quantities of food (glucose) and potential food (citrate) were not "precisely varying." The citrate quantity was fixed and the glucose supply was specified as 25 mg/l but it was stated that the glucose supply could be varied.

>>>>> Do you understand what an experiment is? <<<<<<

Some experiments are accurate simulations of what happens in nature -- this one was not.

>>>>> The experiment was a dizzying, astonishingly complete success. <<<<<

Yes, I know -- it proves Darwin was right and Behe is wrong.

>>>>>> What exactly do you claim to have discovered? I want you to state it precisely and simply, please, so that I can link you to exactly where in the papers or in dialog this was already pointed out to you.<<<<<<

I discovered that a purpose of the glucose-cycling was to limit the bacteria's population densities so that there would be big capacities for population growth when the populations are started by transferring one-percent of the previous populations. As I said umpteen times plus one, Blount gave no answer at all to my questions about glucose cycling, so you would be wasting your time trying to find where this purpose was pointed out to me.

Your constant findings of fault are just meaningless knee-jerk reactions to everything I say.

Tuesday, July 01, 2008 4:43:00 AM  
Blogger Larry Fafarman said...

Hectoring Hector barfed,

>>>>> Perhaps he doesn't feel a need to answer every jackass who brays at him? <<<<<

As the saying goes, dunghill, "if you can't stand the heat, stay out of the kitchen."

Tuesday, July 01, 2008 5:01:00 AM  
Blogger Phae said...

What? Blount said nothing about DM0 -- I presume that means the medium (Davis Minimum broth) without glucose. Your statement is essentially correct about DM25 (with 25 mg/l glucose) -- Blount actually said,

I was paraphrasing. Yes, DM0 is the Davis Minimal Broth without glucose. See how much you're learning!? But I see someone's craaaaanky... naptime for you!

Can we please stop using the word "intent" here? "Intent" is what one plans to do, but one cannot plan on having an uncertain result.

You can actually use the word in that way. Just stop and think about it, you moron. "I intend to make the sandwich" is a proper sentence. So is "I intend to make some sort of lunch, maybe a sandwich" even though there is an element of uncertainty. In the same way, "I intend to apply evolutionary pressure, which may result in the bacteria using citrate." See how that works?

My printed dictionary defines "goal" as "an object or end that one strives to attain." Basically, a goal is something foreseen as a possible and desirable result of an effort. In a search for the Lost Dutchman Mine, finding it is a "goal." If Cit+ evolution was one of several goals, or a secondary goal, or a longshot goal, or whatever kind of goal of the experiment, then saying that it was "not a goal" confuses and misleads people. Blount actually said,

All of your above statements are factually true. You're putting together the perfectly correct and sound sides of a cube and trying to make a sphere.

Someday Blount may really mislead people in a big way by merely saying "it was not a goal" without the clarification, which is likely because he doesn't understand what the word "goal" means. Or someone could quote him out of context as follows: It was "not a goal." I deserve thanks -- not abuse -- for pointing out that the word "goal" is being misused here.

Someday he might mislead people? He stated with perfect accuracy what the facts were! It WAS NOT a goal, it was an admitted POSSIBILITY. Saying "evolving cit+ bacteria was not a goal" would be an entirely valid and true statement. The "goal" was to observe a phenomenon, adaptation and evolution, and see what forms it might take. Cit+ could have been and was among the possible adaptations taken by a bacteria strain, along with enlargement.

Cit+ evolution might have even been a major goal

It wasn't. As Blount has told you. And I have told you. Many, many times.

because of the following factors:
(1) It had been observed once before.


That is not a reason it might have been a goal this time. I have observed people playing pianos but I don't have to play one just because I have seen it. Explain yourself.

(2) This was a long-term experiment with 12 lines of bacteria.

What does that have to do with aside from the opportunity to watch twelve parallel sets of specimens evolve? Explain yourself.

(3) The glucose-cycling favored Cit+ evolution. Lenski might not have considered this factor -- I don't know. Also, I don't know if this was a factor when Cit+ evolution was first observed.

Why did cycling "favor" cit+? You could just as easily say it favored cell enlargement, yet you're not saying that was a goal. In fact, you could also say it favored the bacterium learning to utilize glass, since that would have been even more advantageous. Explain yourself and the relevant literature that leads you to believe a limited supply of food in a famine-and-plenty environment would cause a bacterium to favor cit+ in particular.

Finally, as I have already said umpteen times, Blount gave no answer at all to my questions about glucose cycling.

He directed you to read the paper, which you apparently still have not done. If some random asshole asks me what happened at the first civilian court Gitmo detainee review, I am to be forgiven for not explaining the intricacies of biology to him and instead pointing him to the newspaper.

Do I honestly have to explain to you AGAIN about Polyamerase IV, mutations, and glucose-cycling? Is it conceivable that you might instead go and read the papers?

What? The bacteria did not have to adapt -- they already ate glucose, which was regularly fed to them. The term "thrive" is vague -- the bacteria were surviving quite well before developing the ability to eat citrate. And the bacteria did not adapt "immediately" to the citrate -- it took one line of bacteria about 14 years to adapt to the citrate and the eleven other lines of bacteria have not adapted to the citrate in 20 years. I thought that you read Blount et al.'s paper but you know practically nothing about the experiment.

I have actually read MOST of the papers they wrote about it. You will note I said ADAPT, not cit+ adapt, you raging assclown. Read the words I type, not the random ones in your brain. You are the Da Vinci of morons... years from now, people will discover your notebooks and wonder how you could be so magnificently wrong. The cells, as I told you previously, IMMEDIATELY ADAPTED by getting larger (among other adaptations), as discussed in the first paper published about the experiment, three years after it began.

Your other assertions are bizarre in the extreme. So are you saying that they could have all just continued to eat glucose and not adapted to the much more advantageous ability of cit+? Do you not understand the concept of "thrive" as it refers to doing BETTER?

To make it simple for you: was cit+ advantageous for the bacterium that developed it?

If it was, then it was a GOOD thing that it happened, yes?

One of the problems, dunghill, is that a lot of Darwinists don't recognize that it was a rare evolution.

Oh, then I was mistaken. Please point me to where a biologist (that's what we call them in the current era, Yar; Darwin lived some time ago) is indicating that this mutation, found previously only once, is common. Show me.

The quantities of food (glucose) and potential food (citrate) were not "precisely varying." The citrate quantity was fixed and the glucose supply was specified as 25 mg/l but it was stated that the glucose supply could be varied.

No, the supply found WITHIN the standard DM medium CAN be varied. In the experiment, if you READ THE FUCKING PROTOCOL, they used DM25 until it became DM0. They didn't add varying amounts of glucose; it was precisely calibrated.

Some experiments are accurate simulations of what happens in nature -- this one was not.

What was inaccurate? Please, bother to explain some of your wild accusations for once.

You do understand that experiments must happen in controlled and observable conditions, right? What exactly should he have done differently to meet the requirements I outline for you below?

Yes, I know -- it proves Darwin was right and Behe is wrong.

Only in the broadest strokes. Behe can still claim it is meaningless, I am sure; ID proponents go through elaborate mental contortions to justify creationism. And Darwin was "right" only in his primary principle of natural selection. They didn't even have the concept of DNA in Darwin's time, so he did not have the opportunity to weigh in on genetic mutation except in the loosest way.

I discovered that a purpose of the glucose-cycling was to limit the bacteria's population densities so that there would be big capacities for population growth when the populations are started by transferring one-percent of the previous populations.

No. You are incorrect. The purpose of it was to provide an inadequate food supply for the bacteria, causing it environmental stress and increasing its levels of Polyamerase IV, a well-understood mechanism in many cells such as this which increase the chances of data error during DNA transcription, and accordingly mutation. This increased mutation and environmental stress would provide an opportunity for the bacteria to adapt into a form better-suited for the specific environment in which it consistently found itself. This made it more likely that dramatic evolution would occur and be observable in the relatively short time frame afforded by the career of a scientist.

Once again, this was in the first paper. Once again, you just don't understand what's going on. Once again, you are glaringly wrong and wonder why everyone laughs at you.

As I said umpteen times plus one, Blount gave no answer at all to my questions about glucose cycling, so you would be wasting your time trying to find where this purpose was pointed out to me.

Gee, did you make that lie before? I hadn't noticed.

Your constant findings of fault are just meaningless knee-jerk reactions to everything I say.

Actually, it would be more accurate to say that my constant findings of fault are just knee-jerk reactions to constantly finding faults. Quit fucking up, asshole.

Tuesday, July 01, 2008 6:53:00 AM  
Anonymous brossa said...

I posted comments at Conservapedia in response to yours, and I'll continue to do so here, because I'll take at face value your comment there that "Right now I am just trying to understand the experiment."

First of all, please just drop the 'Blount won't answer my questions' line of argument. Blount is under no obligation to respond to your questions. He is not obligated to respond to the questions of the President of the United States or the questions of the Nobel Prize committee. His lack of response to you can be construed, at worst, as insufficient politeness.

"Now I have found another purpose on my own, and I deserve sole credit for the answer"

You have reached a partially correct understanding of what was explicitly stated in the paper and its supporting materials.

"Each daily population of the 12 lines of bacteria was raised in 10 ml of medium, and one-percent -- i.e., 0.1 ml -- of each old population is transferred to start the next population. That ratio of the medium transfer allows a 100:1 growth in population to maintain the same final population density in the populations. Assuming a doubling of population with each generation, a 100:1 population growth would be reached in 6-7 (6.64, to be exact) generations. 6-7 generations occur in just a few hours (the populations are started in the morning and the glucose is exhausted by the afternoon)"

This is correct, and is stated in the paper and the references.

"If more generations were allowed, the population density would become too high, which would limit capacity for population growth when the next population is started, so glucose supply is limited to limit the number of generations of glucose-eating-only bacteria. I presume that the 10 ml and 0.1 ml quantities are limited by the physical limitations of the lab equipment, e.g., the incubator size and the means for transferring the medium."

Your assumptions in the last sentence are incorrect. The number of bacteria transferred from one flask to the next is easily controlled by varying the volume and/or concentration that is transferred. The initial paper describing the LTEE states that the DM25 medium allows a "stationary phase bacterial density of about 5 x 10^7 cells per ml." That is to say, that the cells will grow near-exponentially until they exhaust the glucose supply, at which point there will be 50 million bacteria per milliliter. Under different glucose concentrations, that number would be lower or higher, but regardless, the number of bacteria transferred to the next flask can easily be manipulated by transferring a larger or smaller volume of mature culture. The experiment could be run with 10:1, 100:1, or 1000:1 growth between transfers. It's arbitrary. Lenski is not restricted to transferring 0.1 ml each time; even if he was, he could dilute that 0.1 ml into 9.9 ml of media, then take 0.1 ml of that solution and transfer it to the next growth flask.

"If desired, the glucose starvation period could be eliminated by restarting the populations more frequently, but that would make the experiment even more labor intensive and I think it was desirable to have a glucose starvation period, anyway."

The glucose starvation period could be eliminated entirely by adding 10000x more glucose, thereby allowing for exponential growth of the bacteria right up to the point at which the next sample is drawn, with no increase in labor. You are correct in that it is desirable to have a glucose starvation period, because that will tend to favor the reproduction of those mutants that are better able to compete for the limited resource. The harsher environment makes the favorable mutants stand out more.

"The medium recipe says that the glucose supply could be varied, and the supply might have been varied to control population growth -- I don't know."

No doubt Lenski ran trials of differnt DM formulas before he started the LTEE. Once it started, however, the DM25 formula remained fixed.

"What is he trying to hide? Maybe he is trying to hide the fact that the glucose cycling favored Cit+ evolution, because some people might regard that as cheating. Aha -- that must be the reason!"

If Lenski had set up a long-term E. coli breeding program with the explicit purpose of developing a strain of Cit+ E. coli, and then succeeded after 20 years and merely published it as a fait accompli with no other explanation, nobody would care. The Cit+ trait is a novelty. It's not inherently valuable. The value arises from the supporting research that shows the manner in which this novelty arose, and provides the opportunity for others to repeat the process by preserving ancestral strains. Lenski could have published a paper about how a strain of E. coli arose during the LTEE that make blue colonies rather than white ones. The particular trait that arose is not important. Merely possessing a Cit+ E. coli strain will not win Lenski any prizes.

Glucose cycling is not 'cheating'. The Cit+ trait was a possible development right from the beginning of the LTEE, but so were cannabalism, toxin production, rapid division, and an unknowable number of other possible traits. Glucose starvation is a fixed part of the environment and favors ALL adaptive traits. Please remember that 'success' from the POV of the bacteria means having your clones make up a larger proportion of the transferred cells than all the other clones. A Cit+ mutant can be outcompeted if it is killed by a toxin-secreting mutant, even if the toxin mutant can't grow on citrate.

Tuesday, July 01, 2008 8:30:00 AM  
Anonymous W. Kevin Vicklund said...

Phae-

>>>Blount said that to the best of his knowledge, Lenski intended twenty years ago (when he started the experiment) that the citrate present in DM0 and DM25 would provide a potential avenue for adaptive evolution of the bacteria.<<<

Your use of "intended" is erroneous - you end up contradicting yourself in the next sentence. "Realized" is a much better word. Subsequent use of "intent" and cognates are appropriate. I should also note that while the becteria ate all the glucose in the DM25, the stationary phase medium is not technically DM0 because other ingredients have also been metabolized (though it is probably functionally equivalent for the experimental setup)

Larry-

>>>Can we please stop using the word "intent" here? "Intent" is what one plans to do, but one cannot plan on having an uncertain result.<<<

We can't stop using "intent" until you stop trying to make claims about what was and wasn't said. Zachary used "intent" and the whole disagreement is over what he meant when he used that word. Also, you are using "plan" in an incorrect and inconsistent manner. "Plan to" and "plan on" have different meanings. "Plan to" is the mental part of an attempt to make something happen, while "plan on" is the additional planning that takes place after an event has occured. It's a "before and after" difference. Furthermore, while it may be unwise to plan on an uncertain outcome, it is not impossible to do so.

And look at one of the most common sports phrases: "[quarterback]'s pass intended for [receiver], broken up by [defender]" There is no definitional requirement that planning require a certain outcome, and the way "plan" and "intent" is used in daily language, it is clear that people use it when talking about trying to make uncertain outcomes take place.

So what does "intent" really mean? According to Webster, it is "the act or fact of intending" - not the most helpful definition. So what does "intend" mean? Again according to Webster, "to have in mind as a purpose or goal" - and I should note that this definition dates back to at least 1971, before Zachary or I were born.

So substituting the definition of "intend" in what Zachary said, here is what he meant:

despite some misunderstanding on the internet, [what they had in mind as the purpose or goal] of the experiment was never to evolve a Cit+ E. coli variant - comment #115

[what they] original[ly had inmind as the purpose or goal] of the experiment was not evolve a citrate-utilizing variant of E. coli, but to, among other things, study the repeatability of evolution and adaptive dynamics.

Zachary then tells us where to look to find out what the original goals (plural) of the experiment were, and unequivocally states that Cit+ evolution was not one of them.

The logical conclusion, therefore, is that while Lenski realized that the way the experiment was set up it could promote Cit+ evolution, he did not design the experiment with Cit+ evolution in mind as a purpose or goal. Whether or not something can be a purpose or goal without having it in mind as a purpose or a goal is a matter best left to Larry's shrink.

As to what the actual purpose of glucose cycling, I'll address that in a separate comment. I will say that Larry at least has made some strides towards understanding the purposes.

Tuesday, July 01, 2008 9:24:00 AM  
Blogger Larry Fafarman said...

Phae barfed,
>>>>>> "I intend to make the sandwich" is a proper sentence. So is "I intend to make some sort of lunch, maybe a sandwich" even though there is an element of uncertainty. In the same way, "I intend to apply evolutionary pressure, which may result in the bacteria using citrate." <<<<<<

But "I intend to have Cit+ evolution" is not a proper sentence, because I have no control over whether there will be Cit+ evolution. But that is the way you and other idiots have been using the word "intend" and its cognates. You think that "intention" is a substitute for "goal" here.

>>>>>> Someday he might mislead people? He stated with perfect accuracy what the facts were! It WAS NOT a goal, it was an admitted POSSIBILITY. <<<<<<

You stupid fathead, as I said, a "goal" is anything foreseen as a possible and desirable result of an effort. In searches for the Lost Dutchman Mine, finding it is a "goal."

>>>>>Cit+ evolution might have even been a major goal

It wasn't.<<<<<<

You can't speak for Lenski, dunghill.

>>>>>(1) It had been observed once before.

That is not a reason it might have been a goal this time <<<<<<

It is a very good reason. When someone finds gold in an area, others are going to come seeking gold in that area. It's called a "gold rush."

>>>>>>(2) This was a long-term experiment with 12 lines of bacteria.

What does that have to do with aside from the opportunity to watch twelve parallel sets of specimens evolve? <<<<<<

Idiot, it means that the bacteria would have a lot of opportunity for Cit+ evolution. Duh

>>>>> Why did cycling "favor" cit+? <<<<<<

I explained it numerous times, idiot, the Cit+ bacteria had an advantage because they had something to eat after the glucose was exhausted. And this doesn't even need explanation -- it is self-evident.

>>>>> You could just as easily say it favored cell enlargement, yet you're not saying that was a goal. <<<<<

I didn't say that wasn't a goal.

>>>>>> He directed you to read the paper, which you apparently still have not done. <<<<<

I browsed through the paper -- it is a long paper, 8 pages of fine print. As I said, presumably most people who have commented about the paper have not seen it at all. The Internet has hundreds of comments about the paper.

>>>>> Do I honestly have to explain to you AGAIN about Polyamerase IV, mutations, and glucose-cycling? Is it conceivable that you might instead go and read the papers? <<<<<<

That is just sending me on a wild goose chase -- most of the material in the papers does not concern my questions. It is appropriate to direct me to specific pages of the papers. Anyway, Blount did not give any answer at all to my questions about glucose-cycling -- he did not even use "bibliography bluffing."

>>>>>> You will note I said ADAPT, not cit+ adapt, you raging assclown. <<<<<

You stupid dunghill, we -- or at least I -- was talking specifically about the Cit+ adaptation, which was by far the most important adaptation.

>>>>>> they used DM25 until it became DM0 <<<<<

Of course it became DM0 -- the bacteria used up the glucose!

>>>>>>They didn't add varying amounts of glucose<<<<<

You said before that the amount of glucose was "precisely varying" -- now you are saying that they didn't add varying amounts of glucose.

>>>>>I discovered that a purpose of the glucose-cycling was to limit the bacteria's population densities so that there would be big capacities for population growth when the populations are started by transferring one-percent of the previous populations.

No. You are incorrect. <<<<<<<

Wrong -- my original post explains this in greater detail. And I didn't say that this was necessarily the only purpose of the glucose-cycling.

>>>>>> The purpose of it was to provide an inadequate food supply for the bacteria, causing it environmental stress and increasing its levels of Polyamerase IV, a well-understood mechanism in many cells such as this which increase the chances of data error during DNA transcription, and accordingly mutation . . . .This made it more likely that dramatic evolution would occur and be observable in the relatively short time frame afforded by the career of a scientist. <<<<<

So maybe that was another way that the experiment was unnaturally rigged to favor Cit+ evolution (as well as other kinds of evolution).

>>>>>> Quit fucking up, asshole. <<<<<<

Your abuse is increasing as you become more desperate because of the hopelessness of your position.

It is time to stop feeding the trolls.

Tuesday, July 01, 2008 9:25:00 AM  
Anonymous Hector said...

> "if you can't stand the heat, stay out of the kitchen." <

Larry's policy is that if he can't stand the heat, he censors.

Tuesday, July 01, 2008 9:28:00 AM  
Blogger Phae said...

Kevin:
Good catch, thanks. You are right, my mistake on that one.

Larry:
But "I intend to have Cit+ evolution" is not a proper sentence, because I have no control over whether there will be Cit+ evolution. But that is the way you and other idiots have been using the word "intend" and its cognates. You think that "intention" is a substitute for "goal" here.

Indeed! There you have it! Thank goodness no one ever said he "intended to have Cit+ evolution," and that it was just one of the possibilities for adaptation and evolution!

"Intention" actually is a serviceable synonym for "goal" in this context, but fortunately for you cit+ was neither a goal nor their intention ;)

You stupid fathead, as I said, a "goal" is anything foreseen as a possible and desirable result of an effort. In searches for the Lost Dutchman Mine, finding it is a "goal."

No. A goal is not anything foreseen as possible, nor just a desirable result of an effort. It is the desired result of an effort.

You can't speak for Lenski, dunghill.

To my great good fortune and in contrast to you, I am fucking literate and so I could read his stated intentions, moron.

It is a very good reason. When someone finds gold in an area, others are going to come seeking gold in that area. It's called a "gold rush."

But when someone finds gold in Alaska, and someone else starts mining in Montana, it does not follow that the Montanan is looking for gold. Instead in our analogy, he is mining looking for whatever might happen to prove his thesis about strata, and gold happened to be one of the results to prove it.

While it is possible they are lying about Lenski's intentions, it seems unlikely and more than your unfounded assertions would be needed.

Idiot, it means that the bacteria would have a lot of opportunity for Cit+ evolution. Duh

They would also have a lot of opportunity to grow larger, or learn to eat glass. Or a lot of other things. Duh.

I explained it numerous times, idiot, the Cit+ bacteria had an advantage because they had something to eat after the glucose was exhausted. And this doesn't even need explanation -- it is self-evident.

Not at all. You are stating that cit+ is an advantage, but it is one among MANY possible advantages. Why did the cycling favor that one in particular, if that is your assertion?

I didn't say that wasn't a goal.

Then kindly please list all of the many "goals" there were for the experiment in your fevered imagination, aside from the ones the experimenters stated.

I browsed through the paper -- it is a long paper, 8 pages of fine print. As I said, presumably most people who have commented about the paper have not seen it at all. The Internet has hundreds of comments about the paper.

Jesus Christ... eight pages of small print?! That's like... that's almost one tenth of a novella! Or a hundredth of a typical novel! GOOD LORD!

Of course no one else casting aspersions on the authors or defending them could be bothered to read that! Never mind, I was seriously mistaken. No wonder it took me that agonizing thirty minutes to read a couple of times... they had to bring me food several times, and I passed out during the ordeal.

Whew, thanks for cleaning that up, and you are a saint for even managing to browse through five pages!

That is just sending me on a wild goose chase -- most of the material in the papers does not concern my questions. It is appropriate to direct me to specific pages of the papers. Anyway, Blount did not give any answer at all to my questions about glucose-cycling -- he did not even use "bibliography bluffing."

It might also be appropriate to read a paper thoroughly before you criticize it, you raging moron.

Blount is not bound to explain elementary biology to a moron, and he was well within his rights to ignore you.

You stupid dunghill, we -- or at least I -- was talking specifically about the Cit+ adaptation, which was by far the most important adaptation.

YOU were. In my careful summation that was trying to teach you, I was in fact talking about the many mutations that occurred.

Of course it became DM0 -- the bacteria used up the glucose!

Very good. You're coming right along. Tomorrow we move past primary colors during fingerpainting class, congrats!

You said before that the amount of glucose was "precisely varying" -- now you are saying that they didn't add varying amounts of glucose.

To be more precise, they added it daily and the amount of glucose in the medium varied according to that schedule.

Wrong -- my original post explains this in greater detail. And I didn't say that this was necessarily the only purpose of the glucose-cycling.

It was not a purpose at all, as explained to you here. Your summation of your "discovery" was entirely inaccurate and mistaken. You fail. Massively and publicly. Again.

So maybe that was another way that the experiment was unnaturally rigged to favor Cit+ evolution (as well as other kinds of evolution).

You here assert again that this setup favors cit+. Please EXPLAIN how it favors that over other adaptations.

It did "favor" evolution in the sense of making it observable and measurable. That's the point of an experiment.

Your abuse is increasing as you become more desperate because of the hopelessness of your position.

Not really. The way I treat you depends on the current level of my amusement with you and charity. When you dance sufficiently to entertain me, I generally spend more time mocking you for my enjoyment. When you are just repeating yourself as you ram headfirst into the wall, I find it more succinct to just call you a stupid prick.

It is time to stop feeding the trolls.

This is code for "Oh shit three people pointing out how wrong I am, gotta run."

Cowardice becomes you.

Tuesday, July 01, 2008 9:53:00 AM  
Anonymous Good Commenter said...

"GOOD LORD!"

Shall we try to roust Him?

("Don't make me come down there.")

:-}

Tuesday, July 01, 2008 11:34:00 AM  
Blogger Larry Fafarman said...

brossa said...
>>>>I posted comments at Conservapedia in response to yours, <<<<<

I know -- and I am not allowed to answer them because of the 90/10 rule -- that is very annoying.

>>>>> First of all, please just drop the 'Blount won't answer my questions' line of argument. Blount is under no obligation to respond to your questions. <<<<<<

I will drop it when hell freezes over. Would Blount dodge simple, basic questions from peer-reviewers and questioners at paper presentations? As I said, if he can't stand the heat, he should get out of the kitchen. Which do you prefer -- my approach of asking simple questions or Conservapedia's Andy Schlafly's approach of requesting the raw data? Anyway, a lot of the discussions here are just wild speculation when we don't have any answers from Lenski, Blount, et al.

>>>>> His lack of response to you can be construed, at worst, as insufficient politeness. <<<<<

Thank you for that generous concession.

>>>>>Lenski is not restricted to transferring 0.1 ml each time; even if he was, he could dilute that 0.1 ml into 9.9 ml of media, then take 0.1 ml of that solution and transfer it to the next growth flask. <<<<<

Good point -- that's true. But for one thing, that would greatly increase the daily labor required, a big concern because this experiment is very long in duration. Also, as I said, mutations occurring in the last generations of a population would be possessed by very few bacteria, and the chances of these late mutations not being included in the transfer would increase if the transfer ratio is increased. Anyway, I think that my original post here has a reasonable explanation of the glucose-cycling's purpose and effect regarding the experiment as actually conducted.

>>>>>> The glucose starvation period could be eliminated entirely by adding 10000x more glucose, thereby allowing for exponential growth of the bacteria right up to the point at which the next sample is drawn, with no increase in labor. <<<<<<

I assume that there is about one generation of bacteria per hour (the populations started growing in the morning, the glucose was exhausted by the afternoon, and there were about 6-7 generations per population). So if the bacteria were fed glucose for the full 24-hour period, then the population theoretically would grow by a factor of 2 to the 24th power, or close to 17 million times. Of course, the maximum carrying capacity of the medium would presumably be reached long before then, so there could be no "exponential growth of the bacteria right up to the point at which the next sample is drawn."

Also, there would be a big increase in labor if the sample had to be diluted prior to transfer (in the manner that you described above). This dilution would have to be done for 12 lines of bacteria.

>>>>> No doubt Lenski ran trials of differnt DM formulas before he started the LTEE. Once it started, however, the DM25 formula remained fixed. <<<<<

I see no evidence that Lenski ran trials of different DM formulas before starting the experiment. The basic medium he used was a standard one -- "Davis minimal broth." And the recipe for the liquid medium says that the glucose concentrations could be varied:

One can, of course, add different concentrations of glucose or other carbon sources as so desired.

>>>>>> The Cit+ trait is a novelty. <<<<<

It is quite a novelty -- its evolution was observed only once before, according to Blount.

>>>>>> The value arises from the supporting research that shows the manner in which this novelty arose, <<<<<

Well, I think that the Cit+ evolution alone was a significant result. But I agree that the "historical contingency" (the high tendency of Cit+ evolution in populations descended from unfrozen populations of 20,000 generations or later) is an important result.

>>>>> Glucose cycling is not 'cheating'. <<<<<

I was half-joking when I called it "cheating." It could be "cheating" in a sense if it is a condition that is unlikely to occur in nature.

>>>>> Glucose starvation is a fixed part of the environment and favors ALL adaptive traits. <<<<<

Not all adaptive traits are necessarily favored by the glucose starvation. For example, glucose starvation would hurt the advantage of Cit- (glucose-eating-only) bacteria with shorter generational times by reducing the time available for exploiting that advantage. Glucose starvation gives an especially big advantage to the Cit+ bacteria because they have something to eat after the glucose is exhausted.

Tuesday, July 01, 2008 12:07:00 PM  
Anonymous 'Nonymous said...

Which do you prefer -- my approach of asking simple questions or Conservapedia's Andy Schlafly's approach of requesting the raw data?

Perhaps both could be sent unpasteurized data?

Tuesday, July 01, 2008 12:11:00 PM  
Anonymous W. Kevin Vicklund said...

So what is the purpose of glucose cycling?

Well, the answer to that requires a lengthy and detailed response. Which is why Zachary decided to answer it in the long post that he announced he was going to write - a post he announced he was going to write before Larry started demanding immediate answers.

In any evolutionary experiment of sufficient timespan (length is relative to the experimental conditions), size of environment must logically be limited. As things multiply, they take up more space, and a laboratory is limited in physical space. So there are two options: either let the population grow until it runs out of physical space, or remove excess population. Since the population in question is basically immortal and has no predators, growth and evolution will stop once the physical limitations are reached. Therefore, we must choose the second option – remove excess population. That’s purpose number 1: size.

There are two ways to remove excess population. One is continuous removal, which is accomplished by chemostat. The advantage of a chemostat is that, once the population reaches steady-state, the environment is constant and only changes as a result of evolution (or outside contamination, which will henceforth be ignored for the sake of argument except as noted). Also, the flow rate determines the growth rate, so we can give accurate estimates of generation number. The disadvantage is that when you take samples, growth continues, making it difficult to compare samples taken at different times. The second way is to perform serial transfer. Assuming that the time elapsed between transfers is greater than the time needed to replenish the original population density (if not, the population will go extinct) the average number of generations will eventually be determined by the binary log of the dilution ratio. The disadvantages are that the environment is cyclical and that not all populations can survive these conditions. The advantage is that generation number is easy to estimate and samples can be saved that (nearly) perfectly record the state of the population and environment at the time of sampling, allowing a myriad of possibilities for experimentation. This includes having a backup in case of contamination, being able to compare ancestral states for relative fitness, being able to go back to look for when mutations were acquired, and being able to rerun a line from an intermediate state to see if the same events take place – you can even halt the experiment for holidays or when tyou need to move the lab cross-country. Carl, Zachary, and the recommended papers listed these as the reasons the second option was chosen.

But why limit a resource other than space? Because it is impractical, and perhaps impossible, to include all of the necessary resources in aqueous solution to support the physically limited population size. It also creates a problem for making transfers and estimating population size, as the population itself would no longer be suspended in water. Keeping things well stirred is also an issue at that point, creating micro-environments and impacting whether the population is randomly distributed throughout the medium. All of these factors contribute to the decision to limit based on resource other than space.

Before going into why limit glucose, let’s visit why they chose the resources they did. About 40 years before the experiment started, a guy named Davis determined the chemical ingredients for a broth which would support E. Coligrowth, but only when a carbon source that a particular strain could metabolize was added. If that strain could not metabolize the carbon source (or if no carbon source was added), the bacteria simply went into a stationary phase – kind of like suspended animation – and wouldn’t grow. They could be frozen, stored for decades, thawed out, and resume growing as soon as a suitable carbon source was added. As decades passed, the response numerous carbon sources was examined and catalogued, including glucose. The advantage of using glucose plus Davis minimal broth is that the environment is easily controlled and we already know how the ancestral train responded to the environment, eliminating the need to perform preliminary experiments on the ancestral response. Additionally, using glucose as the sole (or so they thought!) carbon source enabled them to test whether adapting to glucose-only environment had trade-offs in the ability to metabolize other carbon sources. Yet another reason for glucose only is that the Ara- mutation in 6 of the lines is neutral wrt Ara+ in a glucose only environment, allowing them to be able to compete ancestral and evolved populations in the same flask (and check for cross-contamination). All of these reasons are covered in the recommended papers, and a number of them are covered by Carl and Zachary.

So why limit glucose, rather than another resource? One reason is that because of the reasons why Davis first formulated his broth, the previous experiments focused on the response to carbon-limiting the media. The ancestral population’s response is therefore well-known. Which ingredient in Davis minimal broth is the limiting resource if the carbon source is plentiful? That question is undertemined. Another thing to consider is that Davis minimal broth without glucose is a standard mixture. You can order it direct from several chemical supply companies. All you have to do is add glucose (and thiamine) to get the DM25, or you can make it from scratch if you want. If you altered anything but the glucose, you wouldn’t have Davis minimal broth anymore. Also, as indicated above, one of the purposes was to see whether there would be trade-offs in the ability to metabolize other carbon sources. If the carbon source was not the limiting factor, the ability to metabolize other carbon sources would be almost totally confined to drift, rather than to evolutionary trade-offs.

Now comes the question, why limit glucose to the degree they did? The answer is simple. They wanted to be able to control the population density, and they deliberately set it low. Lenski didn’t explain exactly why they did so in the recommended papers, but he made it clear that it was a deliberate decision to limit the size (and tying back to the previous point, they knew from previous experiments what the population density of the ancestral strain would be). My best guess? One of the metrics they used in determining how the strains changed was density of bacteria in stationary phase. By reducing the density, it made it easier to count. Knowing the population size allows them to perform a number of useful calculations beyond simple numerical analysis, such as mutation rate, fixation rate, and relative fitness of beneficial and harmful mutations.

One final issue: why the 100:1 dilution ratio, and why wait a full day? Because, as Lenski said in paper 147 (the seventh page), waiting a full day is convenient, and diluting any further reduces the effective population size (he later shows that the effective population size is not the same thing as the stationary phase population). Additionally, a 100:1 ratio has some interesting properties. It works out well for our base 10 numbering system, and 500 generations equals 75 days, an easy metric for when to save a sample.

Tuesday, July 01, 2008 12:26:00 PM  
Anonymous W. Kevin Vicklund said...

>>>But "I intend to have Cit+ evolution" is not a proper sentence,<<<

It's an awkward sentence. The proper construction is "I intend to evolve Cit+ E. coli"

>>>because I have no control over whether there will be Cit+ evolution.<<<

Sure you do. You have partial control in the form of the design of the experiment. Of course, if you design the experiment for purposes other than and not including Cit+ evolution, then you would not be intending to evolve Cit+ - just as Zachary said.

>>>But that is the way you and other idiots have been using the word "intend" and its cognates.<<<

That's because, unlike you, we use words to mean what they actually mean, rather than what we want them to mean.

>>>You think that "intention" is a substitute for "goal" here.<<<

And with good reason - my print dictionary says that it is, in fact, a synonym. Which is a high-falutin' term for "a word that you can substitute"

Tuesday, July 01, 2008 1:17:00 PM  
Blogger Phae said...

...such massive ownage ...can't speak ...should have sent a poet...

Tuesday, July 01, 2008 1:43:00 PM  
Blogger Larry Fafarman said...

Heavens -- one of my worst nightmares -- nitpicking pettifogger and cyberbully Kevin Vicklund is back.

I am responding to three different comments here.

Kevin said (Tuesday, July 01, 2008 9:24:00 AM) --

>>>>>> Zachary used "intent" and the whole disagreement is over what he meant when he used that word.<<<<<

Actually, most of the controversy was over "goal." I said that if the Cit+ evolution was one of many goals, or a secondary goal, or a longshot goal, or an incidental goal, or whatever, then people should call it what it is rather than falsely stating that it was not a goal.

>>>>>The logical conclusion, therefore, is that while Lenski realized that the way the experiment was set up it could promote Cit+ evolution, he did not design the experiment with Cit+ evolution in mind as a purpose or goal. <<<<<<

You stupid fathead, how do you know "he did not design the experiment with Cit+ evolution in mind as a purpose or goal"? Sure, there were other purposes of the glucose-cycling and including citrate in the medium, but how do you know that Lenski didn't have Cit+ evolution in mind when he designed the experiment? You people just don't understand that there are different kinds of goals and purposes. And Cit+ evolution could still have been a goal even if nothing specific was done to favor it.

>>>>>> Whether or not something can be a purpose or goal without having it in mind as a purpose or a goal is a matter best left to Larry's shrink. <<<<<<

You're the one who needs a shrink, you stupid fathead.


Kevin said (Tuesday, July 01, 2008 12:26:00 PM) --
>>>>>> So what is the purpose of glucose cycling?

Well, the answer to that requires a lengthy and detailed response. Which is why Zachary decided to answer it in the long post that he announced he was going to write - a post he announced he was going to write before Larry started demanding immediate answers. <<<<<<

Then why didn't he just say that he was going to answer the questions about glucose cycling later? And he never admitted that he was misusing the word "goal."

>>>>> Now comes the question, why limit glucose to the degree they did? The answer is simple. They wanted to be able to control the population density, and they deliberately set it low. <<<<<<

Isn't that something I discovered on my own?

>>>>> One final issue: why the 100:1 dilution ratio, and why wait a full day? Because, as Lenski said in paper 147 (the seventh page), waiting a full day is convenient, and diluting any further reduces the effective population size <<<<<

So why didn't Blount refer me to that specific page rather than use bibliography bluffing? Also, that still does not answer my questions about the intended purposes of the glucose cycling.

Pettifogger Kevin said (Tuesday, July 01, 2008 1:17:00 PM) --
>>>>> Sure you do. You have partial control in the form of the design of the experiment. Of course, if you design the experiment for purposes other than and not including Cit+ evolution, then you would not be intending to evolve Cit+ - just as Zachary said. <<<<<

You are making this much too complicated. "Intend" and its cognates are too ambiguous to be used here.

I am not going to answer Phae's crap at all.

Tuesday, July 01, 2008 2:28:00 PM  
Anonymous Saas-Phae said...

"I am not going to answer Phae's crap at all."

He didn't say anything -- what's to answer?
-- Saas-Phae.

Tuesday, July 01, 2008 3:00:00 PM  
Anonymous brossa said...

I will drop it when hell freezes over. Would Blount dodge simple, basic questions from peer-reviewers and questioners at paper presentations?

The Internet is not a scientific meeting. You are not a scientific peer of Blount. Blount's peers would not ask the questions that you are asking, because they would have a)read the papers and b)already understood the points that seem to confuse you. But this line of argument is a dead end.

But for one thing, that would greatly increase the daily labor required, a big concern because this experiment is very long in duration.

My point was only that the glucose was not limited because it 'had' to be so that there were not 'too many' bacteria transferred to the next flask. The number of bacteria transferred can be controlled by other means, some of which are no more labor-intensive than the current LTEE setup.

-I went on here a bit about the glucose cycling, but on preview I see that w. kevin vicklund covered it in much more detail and clarity than I could.-

Not all adaptive traits are necessarily favored by the glucose starvation. For example, glucose starvation would hurt the advantage of Cit- (glucose-eating-only) bacteria with shorter generational times by reducing the time available for exploiting that advantage. Glucose starvation gives an especially big advantage to the Cit+ bacteria because they have something to eat after the glucose is exhausted.

The entire basis of evolutionary theory is that not all traits are equally favored by the environment. I admit my wording was awkward. The development of the Cit+ trait is not a game-ending proposition. You seem to think that the Cit+ mutants are highly efficient utilizers of citrate right from the get-go, and that this highly effective trait, in this setting, would allow them to outcompete any other possible mutation. It is true that the Cit+ mutants appear to have a significant advantage at this time. But the Cit- bacteria still coexist with the Cit+ ones in about a 1:9 ratio which has remained stable for generations - so the Cit+ advantage is clearly not overwhelming, and could be reversed tomorrow by further mutation in the Cit- strain (although to be fair the mutation that comes to dominate completely will probably emerge from the Cit+ population - call it Cit+++, just for giggles- because that's where 90% of the action is coming from). Furthermore, the early,weakly Cit+ mutants were nearly driven extinct by the Cit- once already.

Tell me, Larry, does the glucose cycling favor the development of the Cit+ trait over the development of a toxin? Over the development of an even faster Cit- reproducer? Over predatory E. coli? Over a Cit- mutant that can jam the machinery of the Cit+ pathway? When the next big mutation shows up, will it have been inevitable that the experimental design favored it?

Tuesday, July 01, 2008 3:11:00 PM  
Blogger Larry Fafarman said...

Brossa said,
>>>>> The Internet is not a scientific meeting. <<<<<

Wrong. The Internet is a huge scientific meeting. Laypeople are always participating in Internet discussions on topics in professional and technical fields. IANAL (I hate that expression), but I frequently participate in discussions on law blogs.

>>>>> You are not a scientific peer of Blount. <<<<<

Wrong -- he is not a scientific peer of me. He refuses to answer my simple, basic questions. Scientists treat laypeople with disdain and then moan that they are not understood by the public.

>>>>> Blount's peers would not ask the questions that you are asking, because they would have a)read the papers and b)already understood the points that seem to confuse you. <<<<<

There is no reason why attendees at a presentation of the paper at a scientific conference would not ask the same questions I asked. Maybe an attendee is unfamiliar with Blount's area of research, or asks a question about something not covered in the paper, or wants a clarification of something in the paper, or disagrees with something in the paper, or didn't read the paper thoroughly or didn't read it at all (often a lot of papers are presented at conferences and it is not practical to read them all), or didn't have a chance to read the paper, or whatever.

>>>>> But this line of argument is a dead end. <<<<<<

It's a dead end for you but not for me. I -- like a lot of laypeople -- am fed up with being snubbed by the so-called "experts."

>>>>>>But for one thing, that would greatly increase the daily labor required, a big concern because this experiment is very long in duration.

My point was only that the glucose was not limited because it 'had' to be so that there were not 'too many' bacteria transferred to the next flask. <<<<<<

That is contrary to the following point that Kevin Vicklund made (Tuesday, July 01, 2008 12:26:00 PM) --

Now comes the question, why limit glucose to the degree they did? The answer is simple. They wanted to be able to control the population density, and they deliberately set it low. Lenski didn’t explain exactly why they did so in the recommended papers, but he made it clear that it was a deliberate decision to limit the size.

I thought you said that Kevin explained the glucose cycling better than you could have. Also, note that Kevin said that Lenski did not explain exactly why the population density was deliberately set low.

>>>>> The number of bacteria transferred can be controlled by other means, some of which are no more labor-intensive than the current LTEE setup. <<<<<<

The only means you suggested for very high transfer ratios is to dilute the sample before the final transfer, and that involves more labor.

>>>>> I went on here a bit about the glucose cycling, but on preview I see that w. kevin vicklund covered it in much more detail and clarity than I could.-<<<<<<

There is a lot of detail -- most of it unnecessary -- but not much clarity. Like Phae, Kevin is a charlatan who tries to impress people with his bullshit.

>>>>> You seem to think that the Cit+ mutants are highly efficient utilizers of citrate right from the get-go, and that this highly effective trait, in this setting, would allow them to outcompete any other possible mutation. <<<<<

I never said that. According to this article on Carl Zimmer's "The Loom" blog, it took about 1000 generations for the first Cit+ bacteria to increase from 0.5% to 19% of the population.

>>>>> It is true that the Cit+ mutants appear to have a significant advantage at this time. But the Cit- bacteria still coexist with the Cit+ ones in about a 1:9 ratio which has remained stable for generations <<<<<

Maybe the Cit+ bacteria would do better if they got more citrate -- I hear that the citrate supply runs out before the end of the 24-hour period.

>>>>> does the glucose cycling favor the development of the Cit+ trait over the development of a toxin? Over the development of an even faster Cit- reproducer? Over predatory E. coli? Over a Cit- mutant that can jam the machinery of the Cit+ pathway? When the next big mutation shows up, will it have been inevitable that the experimental design favored it? <<<<<

There is no question that Cit+ bacteria have an advantage because of the glucose cycling, and the Cit+ mutation is probably more likely than most of those other mutations because the bacteria were close to having the ability to eat citrate. They already had the ability to metabolize citrate -- all they needed was the ability to pass citrate through the cell walls.

Tuesday, July 01, 2008 6:22:00 PM  
Anonymous Voice in the Urbanness said...

> nitpicking pettifogger and cyberbully Kevin Vicklund is back. <

You call him a cyberbully because he always shows the absurdity of your positions?

> I am not going to answer Phae's crap at all. <

You are getting better. You have finally realized that you are at the bottom of a large hole and you have announced that you will stop digging.

Tuesday, July 01, 2008 6:53:00 PM  
Blogger Larry Fafarman said...

>>>>> You call him a cyberbully because he always shows the absurdity of your positions? <<<<<

No, dunghill, I call him a cyberbully because he is always trying to get me kicked off of other blogs. Like you, he doesn't deserve to be allowed to post comments here but my no-censorship policy prevents me from kicking you two off.

Tuesday, July 01, 2008 7:40:00 PM  
Blogger Larry Fafarman said...

Brossa said (Tuesday, July 01, 2008 3:11:00 PM) --
>>>>> Blount's peers would not ask the questions that you are asking, because they would have a)read the papers and b)already understood the points that seem to confuse you. <<<<<<

BTW, I neglected to ask -- exactly where in the paper are my questions answered?

Tuesday, July 01, 2008 9:33:00 PM  
Anonymous brossa said...

My point was only that the glucose was not limited because it 'had' to be so that there were not 'too many' bacteria transferred to the next flask.

That is contrary to the following point that Kevin Vicklund made


The protocol that determines the final population density is independent of the protocol for transferring cells to the next flask. I was addressing your initial statement:

If more generations were allowed, the population density would become too high, which would limit capacity for population growth when the next population is started, so glucose supply is limited to limit the number of generations of glucose-eating-only bacteria.

You can have high final population density AND allow for growth in the next flask by transferring a smaller volume.

I presume that the 10 ml and 0.1 ml quantities are limited by the physical limitations of the lab equipment, e.g., the incubator size and the means for transferring the medium

The 0.1 ml quantity is not fixed; it can be 0.02 ml, 0.002 ml, or 0.0002ml. You can have dense final cultures and still transfer small numbers of cells to the next generation, if you so desire.

The only means you suggested for very high transfer ratios is to dilute the sample before the final transfer

I said in my first post here that Lenski was not limited to transfers of 0.1 ml. The Pipetman (www.pipetman.com) is ubiquitous in bio labs and allows for transfers of as little as 0.2 microliters.

Also, note that Kevin said that Lenski did not explain exactly why the population density was deliberately set low.

This is addressed in the first paper on the LTEE. Basically, the rate at which a favorable mutant comes to dominate a population is proportional on how favorable it is and inversely proportional to how many individuals there are in the population. Assuming that the mutant is 10% more fit than its neighbors, it will reach 50% of the population more quickly in a small population than a large one, in a nonlinear fashion. So smaller populations allow small differences in fitness to establish themselves more easily, at the expense of taking longer to run through all possible mutations. In the LTEE, duration of the experiment is not a problem, so the best way to encourage the fixation of a useful mutation in the population is to keep the overall population size small.

Maybe the Cit+ bacteria would do better if they got more citrate -- I hear that the citrate supply runs out before the end of the 24-hour period.

No doubt they would - and the Cit- bacteria would do better if the transfers occurred at six hours rather than 24 - but that's not the LTEE protocol. As an aside, if Cit+ mutation was meant to be favored over all other pathways, wouldn't it have made sense to include an unlimited supply of citrate, to really kick things off?

There is no question that Cit+ bacteria have an advantage because of the glucose cycling, and the Cit+ mutation is probably more likely than most of those other mutations because the bacteria were close to having the ability to eat citrate. They already had the ability to metabolize citrate -- all they needed was the ability to pass citrate through the cell walls

I could eat paper pulp like mashed potatoes if I could only figure out how to break that darn beta-glycosidic bond. After all, cellulose is just made of sugar, and I can digest that just fine.

Tuesday, July 01, 2008 9:46:00 PM  
Blogger Phae said...

Larry, how do you even look in the mirror after getting humiliated so badly? Time and time again, you make specious claims and repeat them. And time and time again, you are shown why you are foolish nigh-unto-imbecility.

Tuesday, July 01, 2008 10:02:00 PM  
Blogger Larry Fafarman said...

Brossa said,
>>>>>> The protocol that determines the final population density is independent of the protocol for transferring cells to the next flask. <<<<<

No, that's not true. If there is a high final population density and a large fraction of the old population is transferred, then the large number of transferred cells would consume the glucose supply too quickly, not allowing a reasonable number of generations. Of course, it might be possible to fix that by increasing the glucose supply, but all of these things have to be coordinated. It is evident that the following procedure was used for this experiment: 100:1 transfer ratio coordinated with enough glucose for 100:1 population growth (6-7 generations with a doubling of the population in each generation).

>>>>>> You can have high final population density AND allow for growth in the next flask by transferring a smaller volume. <<<<<

Yes, but as I pointed out, mutations occurring in the last generations of a population will be carried by just a few bacteria. If the transfer ratio is extremely high, there would be a high likelihood that none of these mutant bacteria would be transferred to the next population.

>>>>>The only means you suggested for very high transfer ratios is to dilute the sample before the final transfer

I said in my first post here that Lenski was not limited to transfers of 0.1 ml. The Pipetman (www.pipetman.com) is ubiquitous in bio labs and allows for transfers of as little as 0.2 microliters. <<<<<<

OK, but you did not mention these Pipetman (or similar) pipettes before. But that's OK -- you mentioned them now (if I had made an omission like that, I would never hear the end of it).

Also, as I said, assuming one generation per hour and a doubling in population per generation, the population growth in a day would be 2 to the 24th power or about 17 million, and presumably the carrying capacity of the medium would be reached long before then, so it appears that a period of glucose starvation would be unavoidable in a 24-hour population. This glucose starvation has been called an "environmental stressor," but I am wondering if the bacteria are harmed or even stressed by just a few hours of glucose starvation.

>>>>>>Also, note that Kevin said that Lenski did not explain exactly why the population density was deliberately set low.

This is addressed in the first paper on the LTEE. <<<<<<

I counted 41 papers on the LTEE! Now tell me why I should have to wade through all that stuff in a possibly futile search for answers to my questions when Zachary Blount -- the lead author of the last paper -- was right there to answer questions on Carl Zimmer's blog?

>>>>> So smaller populations allow small differences in fitness to establish themselves more easily, at the expense of taking longer to run through all possible mutations. In the LTEE, duration of the experiment is not a problem <<<<<<

But the duration of the experiment was a problem so far as the Cit+ evolution was concerned (though the Cit+ evolution was evidently a secondary or longshot goal of the experiment) -- the Cit+ trait appeared in only one of twelve lines of bacteria in 20 years and took about 14 years to appear in that one line and that was a very lucky break. So as you say, there are tradeoffs.

>>>>>> As an aside, if Cit+ mutation was meant to be favored over all other pathways, wouldn't it have made sense to include an unlimited supply of citrate, to really kick things off? <<<<<

According to the medium recipe, there is 0.25 g of sodium citrate in 500 ml or 500 mg/l compared to a standard supply of 25 mg/l of glucose, so there is 20 times more sodium citrate by weight and the glucose might be more efficient as a food source than the sodium citrate. So there is enough glucose in comparison to the citrate to maintain the ratio you mentioned of Cit+ bacteria to Cit- (glucose-only) bacteria of 9:1. Also, the generational times of the different bacteria could be a factor in that ratio.

>>>>>>They already had the ability to metabolize citrate -- all they needed was the ability to pass citrate through the cell walls

I could eat paper pulp like mashed potatoes if I could only figure out how to break that darn beta-glycosidic bond. After all, cellulose is just made of sugar, and I can digest that just fine. <<<<<<<<

Why do you think that the bacteria are more likely to become cannibals -- or develop the ability to produce toxins that harm their neighbors while not being affected themselves -- than they are to develop the ability to pass citrate through the cell walls?

Phae driveled,
>>>>> Larry, how do you even look in the mirror after getting humiliated so badly? <<<<<

How do you even look in the mirror when you are such a lousy troll?

Wednesday, July 02, 2008 12:22:00 AM  
Anonymous 'Nonymous said...

"I could eat paper pulp like mashed potatoes ..."

You may have my share; thanks! :-)

Stuff might taste better to us if we could digest it.

(Good post BTW.)

Wednesday, July 02, 2008 12:45:00 AM  
Anonymous E. Coli said...

"I am wondering if the bacteria are harmed or even stressed by just a few hours of glucose starvation."

Yes we are! And how come so little attention is paid to our civil rights in all this?

Wednesday, July 02, 2008 12:59:00 AM  
Anonymous Troll said...

The mirror broke when I butted my head against it.

Wednesday, July 02, 2008 1:08:00 AM  
Blogger Larry Fafarman said...

Well, Brossa? Phae? Kevin? Others? Exactly where in the papers are all the answers to my questions, the answers that you said were there? I am still waiting for you to show me where.

I should have asked this question a long time ago -- it would have shut them up sooner.

Wednesday, July 02, 2008 2:54:00 AM  
Anonymous Zen Master said...

"Exactly where in the papers are all the answers to my questions ... ?"

With a little work, you might have the basis for a koan there.

Wednesday, July 02, 2008 8:05:00 AM  
Blogger Phae said...

Well, Brossa? Phae? Kevin? Others? Exactly where in the papers are all the answers to my questions, the answers that you said were there? I am still waiting for you to show me where.

I should have asked this question a long time ago -- it would have shut them up sooner.


How fucking spoonfed do you need this to be? How about you read the paper and find out, as you have been directed to? Do you think it is so unreasonable for you to read something before you criticize its content?

Wednesday, July 02, 2008 9:28:00 AM  
Anonymous Voice in the Urbanness said...

> No, dunghill, I call him a cyberbully because he is always trying to get me kicked off of other blogs. <

Your own actions, like that lie about Kevin, have gotten you kicked off an ever increasing number of blogs.

> my no-censorship policy <

Which you practice on and off arbitrarily.

Wednesday, July 02, 2008 9:44:00 AM  
Blogger Larry Fafarman said...

Phae barfed,
>>>>>How fucking spoonfed do you need this to be? How about you read the paper and find out, <<<<<<

Aha, I knew it! It was just bibliography bluffing.

Voice in the Urbanness barfed,
>>>>> Your own actions, like that lie about Kevin, have gotten you kicked off an ever increasing number of blogs. <<<<<<

Wrong, dunghill, Kevin has tried to get me kicked off of blogs even when I was not violating any rules.

>>>>>> my no-censorship policy <

Which you practice on and off arbitrarily. <<<<<<<<

You deserve to be kicked off this blog for lying but I can't do it because of my no-censorship policy.

Wednesday, July 02, 2008 11:28:00 AM  
Anonymous W. Kevin Vicklund said...

>>>Wrong, dunghill, Kevin has tried to get me kicked off of blogs even when I was not violating any rules.<<<

This is a flat-out lie. I have never tried to get Larry kicked off any blog. In fact, I tried to prevent Larry from getting himself kicked off Pandas Thumb, going so far as to offer him information that he had been requesting - information that would prove him right and me wrong. Larry refused my offer, and was later kicked off PT just as I warned him that he would.

I have pointed out when Larry was violating a ban, but he had already been kicked off. Never have I tried to get Larry kicked off of a blog.

Wednesday, July 02, 2008 12:59:00 PM  
Anonymous Super Troll said...

"The mirror broke when I butted my head against it."

That's nothin'. I just looked at mine.

Wednesday, July 02, 2008 2:20:00 PM  
Anonymous Anonymous said...

> Wrong, dunghill, Kevin has tried to get me kicked off of blogs even when I was not violating any rules. <

This has been shown to be a lie. You are not doing your already limited credibility any good by repeating it.

Wednesday, July 02, 2008 5:02:00 PM  
Blogger Phae said...

Aha, I knew it! It was just bibliography bluffing.

It was unwillingness to pander to a moron. But fine, I will take you by the hand and show you.

p7905 of the newest paper notes methodology, and directs those interested in the details of the overall experiment to the first paper, as I have said to you. Note the reference to the footnote about it, after the first sentence in ''Materials and Methods.'' Following the reference to the '91 paper now on JSTOR, which should have been incredibly easy for you since I have told you where it was previously and linked you there (not that you were willing to read it) - even had it not been in the quite obvious publication list to which I also linked you - we come to the '91 paper, which immediately summarizes the whole of the experiment's intent. On page 1316 is a very clear summary of the intent of the experiment, expressed "metaphorically" rather than technically for the convenience of idiots (hint: you). Notice that citrate is not mentioned.

Now, is there anything else you don't understand? This is the only opportunity I am going to give you to be spoonfed this sort of information, and otherwise will consider it very justifiable to ask that you read the papers before criticizing them.

Wednesday, July 02, 2008 6:48:00 PM  
Anonymous Jim said...

"It's a dead end for you but not for me. I -- like a lot of laypeople -- am fed up with being snubbed by the so-called "experts."

Are you really arrogant enough to believe that you are personally owed an explanatin? That someone who has to worry about REAL questions from other scientists in the field should waste their time explaining background info that can be learned by reading a few papers? A scientist has no obligation to answer your questions and in the process tutor you on basic biology and microbiology. Scientists don't "snub" laypeople, they simply spend their time answering more pointed, well informed questions from their collegues and, more than likely, their competitors.

Wednesday, July 02, 2008 10:38:00 PM  
Blogger Larry Fafarman said...

Jim driveled,
>>>>> Are you really arrogant enough to believe that you are personally owed an explanatin? <<<<<

Yes, I was personally owed an explanation, and no, I am not arrogant. Zachary Blount, lead author of the Cit+ evolution paper, was answering questions on Carl Zimmer's "The Loom" blog. My questions were simple, well-informed, and basic. The questions mostly had nothing to do with basic biology and microbiology. There was more than a few papers to read -- there was 41, and reading them all would not necessarily answer my questions. Compare me to Conservapedia's Andy Schlafly, who thinks that he is owed all of the raw data from 20 years of the experiment.

Thursday, July 03, 2008 12:23:00 AM  
Anonymous Voice in the Urbanness said...

Perhaps if you showed more willingness to actually read the references that were given to you, you might be taken seriously.

Thursday, July 03, 2008 8:36:00 AM  
Blogger Larry Fafarman said...

>>>>> Perhaps if you showed more willingness to actually read the references that were given to you, you might be taken seriously. <<<<<<

Dunghill, answering simple, specific questions -- e.g., was Cit+ evolution a goal, what was the purpose of the glucose cycling -- by giving people big references that consist mostly or entirely of unrelated material is wrong. And you lousy trolls can't identify the exact places (reference and page number(s)) where the questions are answered.

Thursday, July 03, 2008 9:08:00 AM  
Anonymous 'Nonymous said...

"Compare me to Conservapedia's Andy Schlafly"

Why do you wish to be compared to a rude and ignorant person? Lowering the bar?

Thursday, July 03, 2008 9:23:00 AM  
Anonymous "Esch" Coli said...

"Dunghill, answering simple, specific questions ... by giving people big references ... is wrong."

At least you got answers. Yesterday I posted the following message:

"Yes we are! And how come so little attention is paid to our civil rights in all this?"

And got zero responses! I'm hurt!

Do you really favor this sort of rampant discrimination on the basis of phylum?

Do you even know how many of us there are in each dunghill?

-- "Esch" Coli

Thursday, July 03, 2008 10:02:00 AM  
Anonymous Zmidponk said...

Larry bilously snorted...

>>>>>>And you lousy trolls can't identify the exact places (reference and page number(s)) where the questions are answered.<<<<<<

I didn't give you a page number - I just pointed out that, on the paper that Zachary Blount directed to you, there was a large section right at the very start which answered your questions in great detail. Phae DID give you precise page numbers on precise papers. So that is a complete, total and utter, grade 1 lie.

Thursday, July 03, 2008 2:00:00 PM  
Blogger Phae said...

Dunghill, answering simple, specific questions -- e.g., was Cit+ evolution a goal, what was the purpose of the glucose cycling -- by giving people big references that consist mostly or entirely of unrelated material is wrong. And you lousy trolls can't identify the exact places (reference and page number(s)) where the questions are answered.

I did. I actually took the trouble to write out the chain of words your eyes should follow in order to obtain answers to your questions. Even though I knew I shouldn't since you weren't going to actually read the pages I instructed, I took you by the hand and guided you gently through it.

Blount told you cit+ wasn't a requirement and told you to read the paper to find out your other answers, since he didn't feel it was his job to have to explain some of these elementary concepts to you. When you refused to read the latest paper, and its referent, the first paper, which contained all of your answers (as I have shown), then you lost whatever credibility you had in the argument.

You fail.

Thursday, July 03, 2008 8:31:00 PM  
Anonymous Zachary Blunt said...

My non-response to a particular comment should not be interpreted as agreement, approval, or inability to answer.

Thursday, July 10, 2008 7:23:00 AM  

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