Science Papers Challenge Claims that "Alien" Bacteria use Arsenic Instead of Phosphorous
Last December we reported on a controversial paper published in Science which claimed to have discovered bacteria that feed on arsenic instead of phosphorous. According to NASA, this research promised to provide "an astrobiology finding that will impact the search for evidence of extraterrestrial life." At that time the media reported things like:
But soon after the original Science paper was published, credible scientists began critiquing the paper's claims. In the June 3, 2011 issue of Science, several of those scientists have published comments critiquing the original paper. Many of their criticisms focus on the claim that the original paper did not establish or rule out the possibility that the bacteria are not still living off of phosphorous. A review of these criticisms follows:
scientists discovered "a bacteria whose DNA is completely alien to what we know today" (Wired) the "bacteria is made of arsenic" (Wired) the bacteria is "capable of using arsenic to build its DNA, RNA, proteins, and cell membranes" (Gizmodo) the paper had reported "arsenic-based life" which is "very alien in terms of how it's put together" and "NASA has, in a very real sense, discovered a form of alien life" (io9) "you can potentially cross phosphorus off the list of elements required for life" (Nature)
Of course the authors of the original paper, including lead-author Felisa Wolf-Simon, co-authored a reply to the criticisms which should also be read. But critics remain unconvinced. Nature news recently quoted Barry Rosen of Florida International University stating, "I have not found anybody outside of [Wolfe-Simon's] laboratory who supports the work." Likewise, Rosie Redfield observes:
B. Schoepp-Cothenet et al.: This critique argues that "Although their data show that GFAJ-1 [the allegedly arsenophilic bacteria] is an extraordinary extremophile, consideration of arsenate redox chemistry undermines the suggestion that arsenate can replace the physiologic functions of phosphate." The authors argue that the experiments reported in the original paper show that the bacteria still strongly preferred phosphorous and were not preferentially uptaking arsenic:We therefore have to ask whether the case described by Wolfe-Simon et al. differs fundamentally from other extreme forms of life. To us, the answer is straightforwardly provided by the article itself. In the presence of a more than 10,000-fold excess of As over P (the presence of the latter being due to contaminations in the salts used for the culture media), the bacterium integrates only three times (on a molar percentage basis) more arsenic than phosphorus. It is far from proven that As is present in biologically active molecules in the cell. GFAJ-1 appears to do all it can to harvest P atoms from the medium while drowning in As, with cell division occurring once every 2 days (possibly because of the presence of trace P rather than As). This suggests that GFAJ-1 is an extraordinary extremophile but does not support the more exceptional claim that As replaces the functions of P in this organism. Stephen A. Benner: This comment critiques the original paper's claims on the basis that arsenic is chemically incompatible with what we know about the metabolic and biochemical pathways used in life. As the comment explains, "neither the Research Article itself nor the accompanying news article conveyed the extent to which this hypothesis, if true, would require rewriting of many conclusions that we have hitherto accepted based on data from many laboratories." Benner further argues that, "if the arseno-DNA hypothesis proves true, then it will lead to discoveries in metabolism that lie outside metabolic precedent." Finally, Benner cites Carl Sagan noting that "the hypothesis of arseno-DNA would seem to fall into the category of 'exceptional' that, as Carl Sagan remarked, requires exceptional support." Like the previously discussed paper, it explains that the evidence suggests that the bacteria are scavenging phosphate, and have not met Sagan's standard of evidence:The actual numbers reported byWolfe-Simon et al. describing the ratio of arsenic to phosphorus in various subcellular fractions do not allow us confidently to rule out an alternative hypothesis, that the reported microbe aggressively scavenges phosphorus from the environment and uses it where it is most important--in its DNA. Unfortunately, the level of contaminating phosphate in the arsenate that was used to grow these microbes is unknown. Thus, it remains possible that the arsenate added in the growth mixture was itself a source of phosphate that themicrobe might have scavenged. Rosemary J. Redfield: Outspoken critic Rosie Redfield published a comment in Science arguing that "omission of important DNA purification steps, cast doubt on the authors' conclusion that arsenic can substitute for phosphorus in the nucleic acids of this organism." Redfield continues, arguing that "Although the researchers meticulously eliminated contamination of the reagents and equipment used in their elemental analyses, they made much less effort to eliminate contamination in their biological samples." Patricia L. Foster: This critique also suggests that the bacteria were living off of phosphorous rather than arsenic, since "arsenic may have stimulated the bacterium's high-affinity phosphorus assimilation pathway, which is active when phosphate levels are low." James B. Cotner and Edward K. Hall: This criticism observes that while the original paper argued that "the P content in bacteria grown in +As/-P culture medium was far below the quantity needed to support growth" there is evidence from known bacteria groups that "low P content is a common phenotype across a broad range of environmental bacteria that experience P limitation." The paper then reports that "a survey of the elemental content of bacteria from more than 120 freshwater ecosystems ... show that the P content of aquatic bacterial communities sampled in situ and of individual isolates grown in chemostats can be highly depleted in P." The authors themselves have observed bacteria in nature that live off of low phosphorous concentrations:Measurements of individual bacterial cells from one lake indicated that cells contained between 0.01 and 0.1 fmol P per cell with a mean of 0.5%P, with some individuals as low as 0.03% P of dry weight. Additionally, numerous bacterial isolates from temperate lakes grown in chemostats at dilution rates of 1.2 to 2.4 d−1 had P content consistently ranging from 0.2 to 0.4% under Plimiting conditions. We conclude that the P content reported forGFAJ-1, although low, falls within the range we observed for environmental bacteria from a diverse set of aquatic environments. Stefan Oehler: This commenter is skeptical because "straightforward experiments to support this claim [that the bacteria can use arsenic instead of phosphorous], including density gradient centrifugation of DNA assumed to contain arsenic, were either not performed or not presented." The comment further notes that the original paper "presents only preliminary results, not the confirmatory experiments one would expect to find in support of this claim." David W. Borhani: This comment argues that the experimental results reported in the original paper were the "reverse of expectation" if the bacteria had substituted arsenic. It concludes:Does GFAJ-1 incorporate As into its macromolecules? According to the radioactive [73As]- arsenate data shown in table 2 in (1), it apparently does, to some small extent. Perhaps this result is not so surprising for an organism that somehow manages to detoxify high, normally poisonous arsenate concentrations. Have arsenate esters replaced phosphate esters to any appreciable extent, for example, in adenosine triphosphate, DNA, RNA, vitamins, phosphoproteins, or phospholipids? The data presented by Wolfe-Simon et al. seem to suggest not, nor do the authors provide any compelling arguments for how GFAJ-1 avoids deleterious hydrolysis of arsenate esters. In particular, their proposal that critical arsenate ester-containing biomolecules are protected from hydrolysis by sequestration in the observed (putatively poly-bhydroxybutyrate- rich) vacuole-like regions, or in other (unspecified) intracellular regions, implicitly invokes unprecedented mechanisms both for maintaining and for carrying out key metabolic processes in the presence of such a sequestration. István Csabai and Eörs Szathmáry: This comment argues that the original study "lacks crucial experimental evidence to support this claim and suffers from inadequate data and poor presentation and analysis." The critics contend that the original article made incorrect calculations, which if performed correctly, would suggest "much more P than As" in the bacteria. Like other authors, they suggest that the extraordinary claims of the initial paper require more proof: "the data presented by Wolfe-Simon et al. do not show that As is biochemically incorporated into the DNA of GFAJ-1. To support such an extraordinary claim, additional chemical and structural analyses showing the replacement of phosphate by arsenate should have been provided."
"With so many mistakes pointed out, there should be at least some where the authors say, 'you're right, we should have done that but we didn't'," Redfield says. "This as an entirely a 'we were right' response, and that's a bad sign in science."Despite the high levels of skepticism of claims of arsenophilic bacteria, Nature reports that few scientists have taken the initiative to attempt to experimentally reproduce the claims made in the original paper:
However, most labs seem too busy to spend time replicating work that they feel is fundamentally flawed and is not likely to be published in high-impact journals. So principal investigators are reluctant to spend their resources, and their students' time, replicating the work. "If you extended the results to show there is no detectable arsenic, where could you publish that?" asks Simon Silver of the University of Illinois at Chicago, who critiqued the work in FEMS Microbiology Letters in January and on 24 May at the annual meeting of the American Society for Microbiology in New Orleans. "How could the young person who was asked to do that work ever get a job?" Refuting another scientist's work also takes time that scientists could be spending on their own research. For instance, Helmann says he is installing a highly sensitive mass spectrometer that can measure trace amounts of elements. But, he says, "I've got my own science to do."Such admissions do not bode well for those who blindly believe in the perfectly objective, self-correcting nature of science. Indeed, in this case, it seems safe to experimentally critique these claims since so many respected scientists have already expressed vocal skepticism. Yet experiments are apparently not yet forthcoming.