Antibiotic Resistance Revisited
[Note: This response is co-authored with Explore Evolution co-author Ralph Seelke, Professor of Biology at University of Wisconsin-Superior.]
In its rebuttal to Explore Evolution (EE) on antibiotic resistance, the National Center for Science Education (NCSE) calls EE “incoherent,” “deeply confused,” and asserts that it “significantly misrepresents” the data.[1] But this appears to be little more than harsh rhetoric: the NCSE cites papers that demonstrate trivial degrees of evolution and when read carefully, actually validate EE’s arguments about fitness costs associated with antibiotic resistance. The NCSE’s rebuttal to EE with regards to antibiotic resistance contains many misstatements about EE and the data, and the entire rebuttal makes only one valid point—a point which when properly understood actually strengthens the case against macroevolution in Explore Evolution. Moreover, the NCSE makes multiple assertions about the text of EE that are simply not true, some of which are self-contradictory.
A recurrent error in the NCSE’s response to EE on antibiotic resistance is that it wrongly accuses EE of committing a logical fallacy called the fallacy of the undistributed middle. This fallacy is committed when someone argues like this: “My dog is red, therefore all dogs are red.” Obviously, the fact that I observe that one dog is red, or even that many dogs are red, does not mean that all dogs are red. And it would be patently unfair for someone who merely says “My neighbor’s dog is red” to then be accused of arguing that “All dogs are red.” Yet the NCSE consistently misrepresents EE as making such false absolutist statements when in fact EE’s arguments are far more nuanced and careful than the NCSE acknowledges.
If anything, the nature and tone of the NCSE’s weak rebuttal to EE with regards to antibiotic resistance should inspire confidence in the accuracy and balance of EE’s treatment of the subject.
A. The NCSE Claims that According to EE, “Antibiotic resistance is just selection of pre-existing variability” without a Requirement for Mutation, but in fact the NCSE Misrepresents EE on this Point.
The NCSE alleges that according to EE, “[a]ntibiotic resistance is just selection of pre-existing variability.” The NCSE further alleges that according to EE, “a ‘resistance gene’ does not develop through mutation.” This misrepresents EE: not only does the phrase “resistance gene” (which the NCSE directly attributes to EE) exist nowhere in the textbook, but EE also nowhere implies that antibiotic resistance “does not develop through mutation.” This is a prime example of the NCSE wrongly accusing EE of committing the fallacy of the undistributed middle: EE does not make this generalized absolute argument, and in fact the textbook explicitly states that mutations generate variability:
For mutations to provide the raw materials on which natural selection can operate, two things need to happen. First, the ‘mutants’ must be viable (that is, able to survive, and capable of reproducing). Second, the mutation must be heritable. (EE, pg. 101)
In another section on antibiotic resistance where EE describes “The neo-Darwinian Mutation Scenario,” the textbook explains that “mutations in the DNA sometimes modify this program. Thus, descendants may possess modified structures that are similar—but not identical—to those of their parents,” explicitly stating that, “The second way that bacteria become resistant to some antibiotics is through mutation … In a few generations, an antibiotic-resistant strain arises. And mutations are the key.” (pg. 100) EE clearly states (as it should) that mutations are a vital component of the process of antibiotic resistance, for they provide the “raw materials” upon which selection can act.
The NCSE later admits that “Explore Evolution then says mutations do confer resistance but with a ‘fitness cost.'” So which is it? Either EE implies that mutations play a role in antibiotic resistance, or it doesn’t. We already know the answer, but it’s worth noting that the NCSE’s response is self-contradictory, and the NCSE’s admissions defeat its own arguments, misrepresenting EE by claiming that the textbook says that antibiotic resistance is “just selection.” Somehow the NCSE managed to miss EE’s extensive discussion of the importance of mutations.
The NCSE then launches into a lengthy discussion of the history and mechanisms of antibiotic resistance. The NCSE is to be complimented for its understanding of this process, however EE’s coverage of antibiotic resistance is in full agreement with their discussion, except for the fact that EE, because of its target audience, only covers certain types of mechanisms of antibiotic resistance. However, this does not mean that EE is committing the fallacy of the undistributed middle and arguing that those are the only mechanisms of resistance.
The data cited by the NCSE is consistent with EE’s observation that the genes for bacterial resistance to penicillin can pre-exist in the populations of bacteria prior to the introduction of the antibiotic drug. We would hope that the NCSE would see the fallacy of their claim: The fact that EE observes that these genes can pre-exist in a population does not imply that EE is saying that antibiotic resistance “does not develop through mutation” or that EE says that “[a]ntibiotic resistance is just selection of pre-existing variability.” The NCSE is putting words into the mouth of EE that the textbook never stated.
B. When Attacking EE on Antibiotic Resistance, the NCSE Cites Examples of Evolution that have Nothing to Do with Antibiotic Resistance—and those Papers in fact Support EE’s Arguments with Respect to the Trivial Nature of Viable Mutations.
In a section attempting to rebut EE with respect to antibiotic resistance, the NCSE complains that EEdoes not talk about mutations in cis-regulatory elements (CREs), which are stretches of DNA near genes which help control gene-expression. The NCSE relies heavily upon a paper by Prud’homme et al. (2007)[2] which has nothing to do with antibiotic resistance, and in fact gives examples of trivial types of evolution that bolster EE’s observation that viable or advantageous mutations tend to have trivial or minuscule morphological effects. Additionally, the arguments of Prud’homme et al. (2007) have been sharply critiqued and rebutted by leading evolutionary scientists.
The NCSE claims that Prud’homme et al. (2007) shows that “mutations in cis-regulatory elements (CREs)… have minimal fitness costs and are considered by many evolutionary biologists to have the greatest potential for generating evolutionary change.” Aside from the fact that there are very good reasons to expect fitness costs from mutations that control gene expression (this is discussed further in Section F below), the NCSE’s implication is that fitness costs are not important factors in evolution. As is often the case with Darwinists, the devil is the footnotes: A fair read of Prud’homme et al. (2007) shows that the paper confirms the arguments of EE.
Prud’homme et al. (2007) did find that CREs can influence evolution without fitness costs, but the type of evolution reported was trivial: all it found was changing coloration patterns change on the wings of fruit flies. That’s right, the NCSE’s prime example of genetic evolution is changes in wing-coloration patterns on fruit flies. If this is the best example of evolution that Darwinists can produce, it’s no wonder so many scientists are growing skeptical of the claims of neo-Darwinian evolution.
None of the evolution claimed to have been observed in Prud’homme et al. (2007) was in response to insecticides or any pest-control factor, and in fact the paper cited no reason why the different coloration patterns should provide any evolutionary advantage. Thus, despite this example’s lack of any linkage to adaptive selection, few Darwin-skeptics would be surprised by findings that small-scale features like wing coloration patterns on insects can evolve; diverse coloration patterns among obviously closely-related species are common in nature.
Few who want to understand how a car works trouble themselves with investigating the color of paint on the car. But that’s what these wing-spot studies look at. What would have been far more impressive to those who are thinking critically about neo-Darwinism would be if the NCSE had produced a paper that explained how wings could evolve in the first place, not how mutations can change mere coloration patterns on insect wings. EE investigates this far more important question, observing that mutations face an “either/or” problem where “Major Mutations not Viable; Viable Mutations not Major.” As EE states:
Critics of the mutation argument say these textbook examples point to a kind of Catch-22. Small, limited mutations (like those that produce antibiotic resistance) can be beneficial in certain environments, but they don’t produce enough change to produce fundamentally new forms of life. Major mutations can fundamentally alter an animal’s anatomy and structure, but these mutations are always harmful or outright lethal. (pg. 106)
Changes in wing-color are hardly sufficient to explain how mutations can produce “fundamentally new forms of life.” Thus, if anything, the paper confirms the arguments of EE by showing that viable mutations tend to be minor and “don’t produce enough change to produce fundamentally new forms of life.” Tellingly, no evidence that such mutations can produce major changes was produced by the NCSE in its citations.
In fact, Prud’homme et al. (2007) confirms that mutations that change the amino acid sequence in proteins often have a high fitness cost, just as EE argues:
In theory, the loss of a particular pigmentation pattern could occur by the loss of pigmentation gene expression or the disruption of pigmentation protein functions through mutations in their coding sequences. However, the latter kinds of genetic changes would have substantial collateral effects, affecting all pigmentation patterns and other processes in which these proteins are involved. Many fly pigmentation proteins are also involved in cuticle formation and the metabolism of dopamine, an essential neurotransmitter, and D. melanogaster yellow mutants are notorious for their poor mating success (37, 40-42). Hence, losses of pigmentation through changes in the coding sequences of pigmentation genes are unlikely to be tolerated by natural selection, because their fitness cost is too high.
While mutations in protein-coding sequences tend to result in a fitness cost, other evolutionary biologists have observed that mutations in CREs do not produce significant evolutionary change. Hopi E. Hoekstra and Jerry Coyne’s review article, “The Locus of Evolution: Evo Devo and the Genetics of Adaptation,” published in the journal Evolution in 2007, offers a potent rebuttal to arguments from “evo-devo” based upon changes in CREs:
An important tenet of evolutionary developmental biology (“evo devo”) is that adaptive mutations affecting morphology are more likely to occur in the cis-regulatory regions than in the protein-coding regions of genes. This argument rests on two claims: (1) the modular nature of cis-regulatory elements largely frees them from deleterious pleiotropic effects, and (2) a growing body of empirical evidence appears to support the predominant role of gene regulatory change in adaptation, especially morphological adaptation. Here we discuss and critique these assertions. We first show that there is no theoretical or empirical basis for the evo devo contention that adaptations involving morphology evolve by genetic mechanisms different from those involving physiology and other traits. … Genomic studies lend little support to the cis-regulatory theory…[3]
Significantly, the authors argue that the case for cis-regulatory evolution is presently weak because “Supporting the evo devo claim that cis-regulatory changes are responsible for morphological innovations requires showing that promoters are important in the evolution of new traits, not just the losses of old ones.” The article concludes, “evo devo’s enthusiasm for cis-regulatory changes is unfounded and premature. There is no evidence at present that cis-regulatory changes play a major role—much less a pre-eminent one—in adaptive evolution.”
Hoekstra and Coyne of course instead suggest that genetic evolution takes place primarily within protein coding sequences, not regulatory sequences. But Prud’Homme et al. (2007) have made it clear that mutations in protein coding sequences face, in their own words, a “fitness cost.” Taken together, these papers make precisely the same points as EE: mutations that do cause significant phenotypic effects (like those in protein coding sequences) tend to be harmful, and mutations that aren’t harmful (like these mutations in regulatory regions) don’t tend to cause significant phenotypic effects. (Nevertheless, mutations in regulatory regions likely do cause significant fitness costs, as discussed in Section F.) The NCSE has provided no evidence of, as EE puts it, mutations that are both “major” and “viable.”
Prud’Homme et al. (2007) makes one final an observation that corroborates with the text of EE, as the article states: “If a functional CRE were to evolve from naïve DNA, the evolutionary path to acquire all of the necessary transcription factor-binding sites, in a functional arrangement, would be relatively long, and it is difficult to see how selection might favor the intermediates.” This corroborates with EE’s claim that functionally intermediate genetic sequences can be difficult to imagine. To avoid this pitfall, Prud’Homme et al. (2007) suggests that CREs evolve from pre-existing CREs that are already functional. This of course begs the all-important question: From where do functional CREs evolve in the first place? By acknowledging the difficulty in de novo CRE generation, this paper again corroborates a fundamental point of EE: “where does new biological information come from? Critics of neo-Darwinism contend that contemporary evolutionary theory doesn’t have an adequate answer for this question.” (pg. 94)
The NCSE complains that “Explore Evolution does not even discuss mutations in cis-regulatory elements,” but as we have seen, EE does discuss the very category of small-scale-change producing mutations that mutations in CREs represent. Moreover, EE should not be expected to discuss proposed mutational mechanisms for evolution about which leading evolutionary biologists are saying: “There is no evidence at present that cis-regulatory changes play a major role—much less a pre-eminent one—in adaptive evolution.” If the NCSE thinks CREs provide a magic genetic bullet for macroevolutionary change, it may need to keep looking.
C. The NCSE’s Rebuttal Wrongly Implies that Fitness Costs in Resistant Bacteria and Other Organisms are Unimportant to Microbiologists.
The evolution of antibiotic resistance is typically the result of small changes allowing for survival in a microbe or other organism under special circumstances where the organism faces extremely strong selection pressure due to the presence of some antibiotic drug. In other cases, it is the result of the transfer of pre-existing antibiotic resistance genes from one microbe to another, and the selection of such microbes in an environment containing antibiotics. Even in the first example, evolution does not produce a truly new function. In fact the change produced often makes the microbe less fit when the antibiotic is removed—it reproduces slower than it did before it was changed. This effect is widely recognized, and is called the fitness cost of antibiotic resistance. It is the existence of these costs and other examples of the limits of evolution that call into question the neo-Darwinian story of macroevolution.
Fitness costs are real, and biological realities like fitness cost and other limits to evolution play a vital role in shaping strategies used to combat antibiotic resistance, antiviral resistance, and pesticide resistance. In fact, were it not for the existence of fitness cost, in many cases antibiotic resistant bacteria would proliferate and resistant strains would soon replace non-resistant strains. Because of fitness costs, resistant strains are outcompeted by non-resistant bacteria once selection pressure is relaxed, allowing doctors to combat antibiotic resistance through various drug usage strategies.
Yet under the approach adopted by the National Center for Science Education (NCSE) in its critique of EE, organisms are treated as if they are nearly infinitely-plastic; evolution is viewed as if it can do anything. If the NCSE were right—which thankfully it isn’t—then medical researchers would have little hope in the fight against antibiotic resistant microbes.
Not only is the NCSE’s mindset challenged by the evidence[4], but if it were true, the implications for medicine would be drastic: If biological realities like fitness cost and limits-to-evolution did not exist, it would be pointless for medical doctors to try to combat antibiotic resistance or antiviral drug resistance, because evolution could always produce an adaptation such that bacteria would become resistant without incurring a fitness cost. Thankfully, Explore Evolution informs students about the realities of limits to bacterial evolution that give doctors and scientists empirically-based hope in the fight against antibiotic resistance.
The NCSE wrongly implies that fitness costs are a minor issue for those trying to fight antibiotic resistance and other forms of resistance, stating, “Mutations do not necessarily impair a protein’s normal functioning nor impose a fitness cost.” After complaining that “Explore Evolution … says mutations do confer resistance but with a ‘fitness cost,'” the NCSE then claims that “Explore Evolution significantly misrepresents how antibiotic resistance arises in this description.” Unfortunately, it appears that the NCSE misunderstands both EE and the importance of fitness costs to evolutionary biologists.
Many scientific papers discuss the stark reality of fitness costs, supporting the emphasis that EEplaces on this topic. In fact, one paper cited by the NCSE acknowledges that the reality of fitness costs is vital to help scientists predict whether resistance will spread: “biological cost of resistance might be a more relevant predictor of the risk for resistance development.”[5] Another paper published in Environmental Toxicology and Chemistry found that “[t]he topic of fitness costs is a central theme in evolutionary biology” because “fitness costs constrain the evolution of resistance to environmental stress.”[6] Yet another paper observed that “[i]t is generally established that drug resistance mutations reduce viral fitness.”[7] Regarding the specific case of antibiotic resistance, one study in the Journal of Antimicrobial Chemotherapy observed that “[t]he biological fitness cost of antibiotic resistance is a key parameter in determining the rate of appearance and spread of antibiotic-resistant bacteria.”[8] Indeed, science journals are replete with documented examples of fitness costs, as the following selections amply demonstrate:
- An article published in the journal Genetics in 2007 by Marciano et al. titled “A Fitness Cost Associated With the Antibiotic Resistance Enzyme SME-1 β-Lactamase” found that blaSME-1 β-lactamase gene, which confers antibiotic resistance to the use of carbapenems, has a fitness cost associated with mutations in its signal sequence. Only by artificially swapping the gene’s signal sequence with the signal sequence from a different gene could this fitness cost be alleviated; there was no natural evolutionary elimination of this fitness cost. The article found that identifying this fitness cost barrier to evolution helped them prevent the spread of antibiotic resistant bacteria: “The identification of a SME-1-mediated fitness cost allows the direct application of genetic techniques that have been utilized to understand structural features of β-lactamase function and evolution.” See David C. Marciano, Omid Y. Karkouti and Timothy Palzkill, “A Fitness Cost Associated With the Antibiotic Resistance Enzyme SME-1 β-Lactamase,” Genetics, Vol. 176: 2381-2392 (August, 2007).
- A paper in BiomedCentral’s journal Evolutionary Biology titled “Acetylcholinesterase alterations reveal the fitness cost of mutations conferring insecticide resistance” found that some insects exposed to insecticides which target acetylcholinesterase, an important enzyme involved in the nervous system of insects, evolve resistance that comes only at a fitness cost. According to the article, “Our findings suggest that the alteration of activity and stability of acetylcholinesterase are at the origin of the fitness cost associated with mutations providing resistance.” As the paper put it, “higher the number of [resistance-conferring] mutations, the lower the stability of the mutant” enzyme. When seeking mutations that compensated for loss of stability in the mutant enzymes, the study found that “no mutation increased the stability of the enzyme, all combinations resulted in proteins still less stable.” In other words, there was a clear fitness cost faced by insecticide-resistant mutant insects. See David C. Marciano, Omid Y. Karkouti and Timothy Palzkill, “A Fitness Cost Associated With the Antibiotic Resistance Enzyme SME-1 β-Lactamase,” Genetics, Vol. 176: 2381-2392 (August, 2007).
- A paper in the Journal of Antimicrobial Chemotherapy, titled “Nitrofurantoin resistance mechanism and fitness cost in Escherichia coli,” observes the reality of fitness cost, stating: “The biological fitness cost of antibiotic resistance is a key parameter in determining the rate of appearance and spread of antibiotic-resistant bacteria.” The paper found that because of the fitness cost associated with E. coli that are resistant to Nitrofurantoin, “even though resistant mutants will appear in the bacterial population in the bladder, they will be unable to become enriched and establish an infection because of their impaired growth at these therapeutic antibiotic concentrations.” The article further observes that, “Resistance to antibiotics is most often accompanied by a biological cost, observed as a decrease in fitness, i.e. a reduced growth rate or virulence.” Ironically, the paper cited by this study to bolster this claim—a claim that corroborates EE’s statements about fitness cost—is Andersson (2006) [see below], the same paper that the NCSE cites to back its claim that “not all mutations produce fitness costs!” It seems that research scientists have interpreted Andersson (2006) differently than the NCSE. See Linus Sandegren, Anton Lindqvist, Gunnar Kahlmeter, and Dan I. Andersson, “Nitrofurantoin resistance mechanism and fitness cost in Escherichia coli,” Journal of Antimicrobial Chemotherapy, Vol. 62, 495-503 (2008).
- Andersson (2006) explicitly observes that fitness cost is important to understanding whether resistant populations will persist after selection is relaxed:
A key parameter influencing the rate and trajectory of the evolution of antibiotic resistance is the fitness cost of resistance. Recent studies have demonstrated that antibiotic resistance, whether caused by target alteration or by other mechanisms, generally confers a reduction in fitness expressed as reduced growth, virulence or transmission. These findings imply that resistance might be reversible, provided antibiotic use is reduced. However, several processes act to stabilize resistance, including compensatory evolution where the fitness cost is ameliorated by additional mutation without loss of resistance, the rare occurrence of cost-free resistance mechanisms and genetic linkage or co-selection between the resistance markers and other selected markers. Conceivably we can use this knowledge to rationally choose and design targets and drugs where the costs of resistance are the highest, and where the likelihood of compensation is the lowest.
Thus, Andersson (2006) observes that “cost-free resistance mechanisms” are “rare” and that fitness cost is a very common phenomenon, stating that antibiotic resistance “generally confers a reduction in fitness.” EE thus properly discusses this common phenomenon, and Andersson (2006) actually bolsters the points of EE. We find it unfortunate that the NCSE has misused this paper in its attempt to downplay the importance and reality of fitness costs. Additionally, Andersson (2006) states that “A rational antibiotic design strategy is therefore to identify targets for which the resistance mechanism has the most negative effect on fitness.” This is a good strategy, but it would be pointless if bacteria didn’t face evolutionary limits and could essentially always evolve to avoid fitness costs, as the NCSE implies. Again, we see that fitness cost is a real phenomenon and is vitally important to understand as microbiologists seek to slow the spread of antibiotic resistant bacteria. EE is justified in discussing it. See Dan I Andersson, “The biological cost of mutational antibiotic resistance: any practical conclusions?,” Current Opinion in Microbiology, Vol. 9:461-465 (2006).
Many similar examples could be cited. Given the scientific literature, how can the NCSE seriously maintain that fitness cost is not an important issue in microbiology or that EE is mistaken by highlighting its importance to evolutionary processes? The NCSE asserts that EE “significantly misrepresents how antibiotic resistance arises” when EE states that “[e]xperiments show that once antibiotics are removed from the environment, the original (non-resistant) strain ‘out-competes’ the resistant strain, which dies off within a few generations” But studies like those discussed here directly corroborate this claim of EE.
D. The NCSE Wrongly Claims EE Mis-describes how Antibiotic Resistance Can Evolve.
The NCSE further claims that EE “significantly misrepresents” antibiotic resistance by claiming that “[a] mutation [changes] the shape of the active site on the ‘target’ protein so that the antibiotic no longer recognizes the site.” The NCSE’s response is to discuss other mechanisms of bacterial antibiotic resistance, mechanisms which while true, do not contradict EE’s accurate description of the major way that antibiotic resistance develops.
The mechanism of antibiotic resistance highlighted by EE is indeed an extremely important mechanism of resistance. For example, an authoritative review of antibiotic resistance mechanisms in Nature by Walsh (2000) found that one of three major mechanisms of antibiotic resistance is to “[r]eprogramme the target structure” where bacteria resistant to erythromycin “have learned to mono- or dimethylate a specific adenine residue A2058, in the peptidyl transferase loop of the 23S RNA component of the ribosome” which prevent the antibiotic from binding to its target.[9]
The NCSE asserts that EE is wrong to state that these changes that confer resistance can have negative effects upon the organism, but clearly EE was not wrong to make this claim. As an article titled “The origins and molecular basis of antibiotic resistance,” in British Medical Journal states, “Alterations in the primary site of action may mean that the antibiotic penetrates the cell and reaches the target site but is unable to inhibit the activity of the target because of structural changes in the molecule.”[10] This article explains that “[m]utants of Streptococcus pyogenes that are resistant to penicillin and express altered penicillin binding proteins can be selected in the laboratory,” but notes that these resistant mutants “have not been seen in patients, possibly because the cell wall can no longer bind the anti-phagocytic M protein” (emphasis added). Thus, the alteration of the enzyme enables resistance but prevents it from binding an important protein that protects the cell from the immunoresponse of the host patient, a severe fitness cost. As Marciano (2007) observes in the journal Genetics, “Antibiotic resistance that occurs via mutation of an antibiotic target often results in a fitness cost to the bacteria under permissive conditions.”[11] How can the NCSE attack EE’s statements about fitness cost as being anything but the mainstream scientific view?
E. The NCSE Misrepresents EE by Insinuating It Does Not Discuss How Compensatory Mutations Can Overcome Fitness Costs.
The NCSE claims that EE is wrong to observe that “Mutations impair normal protein function resulting in a fitness cost” and implies that EE teaches that mutations “necessarily impair a protein’s normal functioning [and/or] impose a fitness cost.” (emphasis added to NCSE’s statement) Once again, the NCSE overstates EE’s arguments and wrongly represents EE as making absolute statements: The NCSE claims that EE argues that there is always a fitness cost, but EE does discuss compensatory mutations and explains how they can allow some resistance-conferring mutations to overcome the fitness cost. As EE plainly states:
Researchers have noticed that this fitness cost is sometimes offset by additional mutations. Because these mutations make up for—or compensate for—the damage caused by the first mutation, biologists call them ‘compensatory mutations.’ … Compensatory mutations act on the “companion” protein components, and this helps restore some of the original machine function that was lost due to the first mutation. In this way, the compensatory mutations allow the bacterium to keep its resistance to antibiotics, while recovering its lost fitness. This suggests that mutations can cause new varieties to arise without loss of function to vital cellular systems. (EE, pg. 108)
Sadly, the NCSE never gives its readers any hint that EE contains a balanced discussion of this topic which explains that compensatory mutations can take place in some cases to alleviate fitness cost. The NCSE misleads its readers by insinuating that EE does not mention the importance of compensatory mutations.
The NCSE further claims that EE is wrong to state that “[t]he cell cannot endure an unlimited number of mutation-induced changes at these critical active sites” and claims that EE is “deeply confused about the role of mutations and adaptations.” The NCSE’s extrapolations from the evidence of compensatory mutations, however, is unwarranted by the data, as it makes the sweeping assertion that “[t]herefore a large number of mutations can be tolerated.” But the evidence from compensatory mutations does not necessarily support that claim. In fact, in many cases of resistance, mutations which confer resistance have great fitness costs because they impair survival and reproduction, whereas others have lesser fitness costs. While those costs are lesser, they are still costs. Obviously the mutations with lesser fitness costs become more strongly fixed into the population than those with more severe fitness costs, but this does not justify the NCSE’s claim that “[t]herefore, a large number of mutations can be tolerated.” Quite the opposite, it implies that only particular mutations can be tolerated. For example, one paper that studied compensatory mutations cited earlier found that only certain mutations were able to restore fitness, whereas others resulted in severe fitness costs:
Spcr mutations in the 16S rRNA gene at positions 1191 and 1193 were associated with a marked impairment of C. psittaci biological fitness, and the bacteria were severely outcompeted by the wild-type parent. In contrast, mutations at position 1192 had minor effects on the bacterial life cycle, allowing the resistant isolates to compete more efficiently with the wild-type strain. Thus, mutations with a wide range of fitness costs can be selected in the plaque assay, providing a new strategy for prediction and monitoring of the emergence of antibiotic resistance in chlamydiae.[12]
The NCSE protests that the “loss of fitness” isn’t an absolute loss of fitness because it is “always in relation to the wild-type organism in the original environment.” However, in many cases the fitness cost is an absolute one, particularly where resistance creates fundamental impairments of an organism’s ability to perform basic life-processes like protein-production, cell-wall maintenance, or replication. In these common cases, we see vital life-processes become less efficient, causing organisms to lose function rather than gain function in some new environment. The biological reality of fitness costs is a far cry from the NCSE’s hypothetical example of penguins evolving the ability to use their wings to swim.
The NCSE further complains about EE’s treatment of compensatory mutations, asserting that “many compensatory mutations” do not have “hidden fitness costs.” Here, the NCSE is attempting to do a difficult thing by proving a negative, namely stating that hidden fitness costs absolutely do not exist. This is difficult to prove because there are many potential environmental changes that a bacterium could face, and fitness costs might only be apparent in some of them. Hidden fitness costs are, of course, hidden, and remain so until one finds an environment in which the cost is shown. Further research often discovers these hidden costs which the NCSE would have researchers believe are non-existent.
For example, in a study titled “Fitness Cost and Impaired Survival in Penicillin-Resistant Streptococcus gordonzii Isolates Selected in the Laboratory,” Haenni and Moreillon reported mutations that increased the resistance of Streptococcus gordonii to penicillin as well as compensatory mutations that restored the loss of fitness caused by the resistant mutations. The fitness cost did not show up either in the rate of growth or in the rate of death during the stationary phase. However, the strain with the compensatory mutations died off much faster when grown in the presence of the unmutated strain. This was a cost that was hidden until further research revealed it.[13]
The NCSE goes on to suggest that EE’s example of a compensatory mutation with hidden fitness costs may be an “imaginary example.” In fact, EE’s example is not imaginary, for the example given by EE—changes in temperature and salinity—commonly influence bacterial behavior, and these are likely changes that a bacterium could face. In particular, for bacteria grown near their maximum growth temperature, mutant proteins often show instability that the original protein lacks.
Finally, it must be noted that the NCSE at one points admits that there are cases “where it is true” that compensatory mutations have “hidden fitness costs.” For all of the NCSE’s protestations, this effectively concedes EE’s point. Compensatory mutations and hidden fitness costs indeed represent areas where further research is needed, which is precisely how EE frames the issues, noting that “[t]his disagreement is far from over, and the field is wide open to more research.” (pg. 109) In contrast to the NCSE’s sweeping insinuations that hidden fitness costs are unimportant and “a large number of mutations can be tolerated,” EE acknowledges that this is a field of active research, and EE makes its claims about compensatory mutations in a tentative, balanced, and scientific fashion.
F. The NCSE’s One Valid Point Actually Strengthens the Case Against Macroevolution in EE, Demonstrating that Antibiotic Resistance Does Not Produce New Features.
The NCSE makes one valid point: The first edition of EE incorrectly states that “[i]n every case where mutations lead to antibiotic resistance, resistance results from small changes to a single protein molecule.” This is an overstatement, as the aforementioned review by Walsh (2000) found that in addition to small changes in single protein molecules, other mechanisms of antibiotic resistance include overproduction of pre-existing “efflux pumps” that remove the antibiotic from the cell interior, or overproducing pre-existing enzymes that can neutralize the antibiotic. Another mechanism is to overproduce the target protein. The NCSE calls these mutations that “change the expression of drug transporters,” but in each of these mechanisms, nothing new has evolved—the only change is the level of production of pre-existing parts. (The NCSE also gives an example of mutations in a cell-wall enzyme, but this too represents “small changes to a single protein molecule,” the very mechanism cited by EE, and thus does not contradict EE’s text in any way.)
EE focuses on the major mechanism of antibiotic resistance that seems to provide the best opportunity for antibiotic resistance to evolve something new. But as EE explains, even in this case, antibiotic resistance still falls short, because it is “doubt[ful] that the kind of mutations that produce antibiotic resistance can ever produce fundamentally new forms of life—no matter how many times the same molecule is altered.” (pg. 104) The NCSE responds by citing other mechanisms of antibiotic resistance that don’t even entail the evolution of anything new, mechanisms that seem even less able to produce “fundamentally new forms of life” than do mutations in proteins. In fact, the NCSE’s example of “[c]hanges in gene expression” are known to have severe fitness costs: Mortlock (1984) discusses classic experiments where bacteria evolve the ability to metabolize the sugar alcohol, xylitol (you may have eaten some in sugar-free gum). However, this newfound metabolic ability is due to the simple overproduction of a pre-existing enzyme, i.e. a change in gene expression. Removing xylitol reveals the fitness cost to the microbe of this mutation. Once the enzyme is no longer needed, overproduction of this now unneeded enzyme makes the microbe less fit. The population quickly becomes dominated by revertants that have lost the ability to use xylitol.[14] From a genetics standpoint, it seems likely that the same kind of fitness cost would be felt by bacteria that become resistant to antibiotics due to overproduction of particular proteins or efflux pumps, and then in the absence of the antibiotics, incur a fitness cost because they’re overproducing proteins they no longer need. Thus, the NCSE’s examples would probably entail great fitness costs. Far from harming EE’s case against macroevolution, the NCSE’s clarifications bolster EE’s arguments.
In this regard, we appreciate the NCSE making this note about other mechanisms of antibiotic resistance and thereby strengthening the arguments of EE: Not only do the NCSE’s examples bolster EE’s point that antibiotic resistance is an insufficient mechanism to explain, via great extrapolation, the evolution of complex new biological features, but the NCSE’s examples also strengthen EE’s explanatory discussion of the severity of fitness costs experienced by resistant microbes.
G. The NCSE Overstates EE’s Claims Regarding Bacterial Speciation and Fails to Provide Evidence Sufficient to Rebut EE on this Point.
The NCSE claims that EE says “No species of bacteria has ever changed into another” but again, the NCSE is overstating the EE’s arguments. In reality, Explore Evolution quotes an authority about the state of the evidence, and makes no dogmatic statements akin to the NCSE’s false characterization of EE; the textbook simply observes that “British bacteriologist Alan Linton has noted, ‘Throughout 150 years of the science of bacteriology, there is no evidence that one species of bacteria has changed into another.'” (pgs. 104-105) The NCSE apparently takes issue with EE for quoting Linton, disregarding the fact that Linton himself is an esteemed bacteriologist who is an expert in this field. In this regard, the NCSE attempts to respond to Linton by adopting a much weaker standard of “evidence” for speciation than that used by Linton.
Linton seems to be talking about direct observation of bacterial speciation. The papers cited by the NCSE to rebut Linton[15] do not provide any direct observational evidence of bacterial speciation, and thus do not rebut Linton’s claim. Instead, the NCSE’s citations infer relatedness of bacterial species based upon the observation that two bacteria genera—Escherichia and Salmonella—share some genetic similarities. In fact, the paper estimates that their speciation event took place over the course of 70 million years. Based upon these genetic similarities, it was inferred that the two groups evolved from a common ancestor—but an alleged divergence that started over 70 million years ago was obviously not observed; at best, it was inferred from mere genetic similarity.
The NCSE thinks that mere genetic similarity is sufficient “evidence” to show that two species evolved from a common ancestor. But the assumption that similarity implies inheritance from a common ancestor is just that—an assumption. In fact, EE documents that numerous instances where this assumption and standard methodologies for inferring common descent and homology break down (see especially the Anatomical Homology and Molecular Homology chapters of EE). There are many instances where similarities among living species contradict expectations of common descent, including examples given in EE like identical genes being used to construct eyes in widely diverse types of animals or examples of extreme convergent evolution in morphology. Skeptics have much empirical justification for questioning the NCSE’s assumption about similarity necessarily implying homology and inheritance from a common ancestor.
The fact that these bacteria share genetic similarities does NOT dictate that we have “evidence” of an unguided speciation event that began some 70 million years ago. If we go off of the empirical observations we do have, Linton’s claim remains justified. Moreover, if the NCSE argues that it takes 70 million years for two relatively similar species of bacteria to diverge from one-another, how can evolutionary biologists ever hope to account for the explosive appearance of many new animal body plans during the Cambrian period in just a few million years?
References Cited
[1] National Center for Science Education. 2008. Section on “Bacteria” in the NCSE critique of Explore Evolution. Available at https://ncse.com/creationism/analysis/bacteria as of January 16, 2009.
[2] Benjamin Prud’homme, Nicolas Gompel, and Sean B. Carroll, “Emerging principles of regulatory evolution,” Proceedings of the National Academy of Sciences, USA, Vol. 104:8605-8612 (May 15, 2007).
[3] Hopi E. Hoekstra and Jerry A. Coyne, “The Locus of Evolution: Evo Devo and the Genetics of Adaptation,” Evolution, Vol. 61-5: 995-1016 (2007).
[4] See R. Seelke and S. Ebnet. “An unexpectedly low evolutionary potential for a trpA 49V,D60N double mutant In Escherichia coli.,” Presented at the 107th Annual Meeting, Abstract R-055, American Society for Microbiology, Toronto, Canada, May 21-25, 2007; R. P. Mortlock (ed.), Microorganisms as Model Systems for Studying Evolution (Plenum Press, New York, 1984). Note: This book contains seven examples of situations in which evolution fails to produce a new function.
[5] Dan I Andersson, “The biological cost of mutational antibiotic resistance: any practical conclusions?,” Current Opinion in Microbiology, Vol. 9:461-465 (2006).
[6] Lingtian Xie and Paul L. Klerks, “Fitness costs constrain the evolution of resistance to environmental stress in populations,” Environmental Toxicology and Chemistry, Vol. 23(6):1499-1503 (2004).
[7] M. Cong, D.E. Bennett, W, Heneine and J.G. GarcÃa-Lerma, “Fitness Cost of Drug Resistance Mutations is Relative and is Modulated by Other Resistance Mutations: Implications for Persistance of Transmitted Resistance,” Antiviral Therapy, Vol. 10, Suppl 1:S169 (June 7-11, 2005).
[8] Linus Sandegren, Anton Lindqvist, Gunnar Kahlmeter, and Dan I. Andersson, “Nitrofurantoin resistance mechanism and fitness cost in Escherichia coli,” Journal of Antimicrobial Chemotherapy, Vol. 62, 495-503 (2008).
[9] Christopher Walsh, “Molecular mechanisms that confer antibacterial drug resistance,” Nature, Vol. 406:775-781 (August 17, 2000).
[10] Peter M Hawkey, “The origins and molecular basis of antibiotic resistance,” British Medical Journal, Vol. 317(7159):657-660 (September 5, 1998) (emboldened emphasis added).
[11] David C. Marciano, Omid Y. Karkouti and Timothy Palzkill, “A Fitness Cost Associated With the Antibiotic Resistance Enzyme SME-1 β-Lactamase,” Genetics, Vol. 176: 2381-2392 (August, 2007) (emphasis added).
[12] Rachel Binet and Anthony T. Maurelli, “Fitness Cost Due to Mutations in the 16S rRNA Associated with Spectinomycin Resistance in Chlamydia psittaci 6BC,” Antimicrobial Agents and Chemotherapy, Vol. 49(11):4455-4464 (November, 2005).
[13] Marisa Haenni and Philippe Moreillon, “Fitness Cost and Impaired Survival in Penicillin-Resistant Streptococcus gordonzii Isolates Selected in the Laboratory,” Antimicrobial Agents and Chemotherapy, Vol. 52(1): 337-339 (January, 2008).
[14] R. P. Mortlock (ed.), Microorganisms as Model Systems for Studying Evolution (Plenum Press, New York, 1984).
[15] Adam C. Retchless and Jeffrey G. Lawrence, “Temporal Fragmentation of Speciation in Bacteria,” Science, Vol. 317:1093-1096 (August 24, 2007) and Christophe Fraser, William P. Hanage, Brian G. Spratt, “Recombination and the Nature of Bacterial Speciation,” Science, Vol. 315:476-480 (January 26, 2007).