Can We Trust Very Old Claims

There are many examples where scientists have trusted their results at first but later the results have turned out to be wrong. Historically, ancient DNA studies have suffered much criticism since they began about 20 years ago. Unfortunatel,y the field is still recovering from the effects of early spectacular and erroneous claims, such as that of DNA being preserved in plant fossils, dinosaur bones, and amber for many millions of years (Hebsgaard et al. 2005; Willerslev and Cooper 2005). Unfortunately, unreplicated results of surprising age continue to be published,

including those from old human remains (Adcock et al. 2001), microorganisms (Cano and Borucki 1995; Vreeland et al. 2000; Fish et al. 2002), and plant fossils (Kim et al. 2004). These studies have routinely underestimated the extent to which ancient DNA research is confounded by contamination with modern DNA, and are widely thought to result from such contamination (Willerslev et al. 2004a; Hebsgaard et al. 2005; Willerslev and Hebsgaard 2005).

In recent years, a greater understanding of postmortem damage and contamination has provided a more robust foundation for the field, although the authentication of studies of human remains and microbes is still highly problematic (Willerslev et al. 2004b; Gilbert et al. 2005b; Hebsgaard et al. 2005; Willerslev and Cooper 2005).

The first report of putative Neanderthal (Homo neanderthalsensis) mitochondrial DNA (mtDNA) was a rare example of a remarkable ancient DNA (aDNA) result obtained using very strict criteria for authenticity, including the independent replication of results and tests of biochemical preservation (Krings et al. 1997; Cooper and Poinar 2001; Hofreiter et al. 2001a; Paabo et al. 2004; Willerslev and Cooper 2005; Hebsgaard et al. 2007). The result is convincing, as the Neanderthal sequence differs from any known modern human (Homo sapiens) and chimpanzee (Pan troglodytes) sequences but is clearly human-like. Furthermore, subsequent independent retrieval of similar, but not identical, mtDNA from other Neanderthal specimens strongly supports the sequence's authenticity (Krings et al. 1999, 2000; Ovchinnikov et al. 2000; Schmitz et al. 2002; Serre et al. 2004; Lalueza-Fox et al. 2005; Hebsgaard et al. 2007). Although the result is convincing it has been shown that the first published Neanderthal sequence may include errors due to postmortem damage in the template molecules for PCR (Hebsgaard et al. 2007). In contrast, inadequate experimental design and a high percentage of chimeric sequences misled Pusch and Bachmann (2004) to suggest the Neanderthal sequences were products of PCR artefacts, a conclusion that later turned out to be wrong (Hebsgaard et al. 2007).

Two recent ice core studies have investigated the long-term survival of DNA in ice from Greenland (Willerslev et al. 2007) and Antarctica (Bidle et al. 2007). The first study showed that DNA can be extracted from ice core samples dated 450,000800,000 years old from the centre of Greenland (Willerslev et al. 2007). Following strict criteria (Willerslev et al. 2004b; Hebsgaard et al. 2005; Willerslev and Cooper 2005), PCR techniques yielded short sequences (less than 120 bp) of plant and insect DNA, which were independently replicated in three different laboratories (Willerslev et al. 2007).

In the study by Bidle and colleagues (2007), ancient DNA and viable cells were isolated from up to 8 million-year-old samples from Antarctica. This is indeed remarkable, and if authentic this study is the first to amplify DNA and viable cells from ice cores as old as 8 million years. Unfortunately, as with many other results of geological ancient DNA, the study did not follow the strict criteria for ancient DNA studies and therefore suffers from inadequate experimental setup and insufficient authentication. Hebsgaard et al. (2007) showed how important it is to follow the strict criteria when working with very old DNA or geological ancient DNA. The results are also interesting compared to results from an 8 million-year-old permafrost sample from Antarctica where not even small fragments of DNA could be amplified (Willerslev et al. 2007). In contrast to Bidle et al. (2007), this study applied strict criteria for ancient DNA work and used dedicated facilities. The results are also interesting because both the DNA and the viable cells isolated from the ice are much older that expected, and are also far reaching compared to the record of long-term DNA survival from permafrost sediment.

Studies of old permafrost samples have been in progress since Willerslev et al. (2003) showed under strict conditions that DNA from extinct animals and plants can be recovered by independent laboratories using strict criteria from samples dated to be 300,000-400,000 years old. Additionally, the study showed that it is not possible to extract DNA from sediment samples dated to be 1.5-2 million years old (Willerslev et al. 2003). A more recent study showed that bacterial DNA can be amplified from 400,000- to 600,000-year-old permafrost samples from Siberia, but not from 8.1 million-year-old samples from Antarctica (Willerslev et al. 2004a). Both of these studies used strict criteria including replication in an independent laboratory, which excluded the possibility that the results were due to laboratory contaminations. A problem with these studies is the risk associated with vertical migration of DNA across different strata. However, it has been shown that for organisms that do not produce copious amounts of liquid urine, the DNA is strati-graphically localized in the sediments, which is true for bacteria, plants and most animals (Haile et al. 2007). Additionally, the results are within the range of what many groups currently accept as maximum ages for DNA survival (Hofreiter et al. 2001b; Smith et al. 2001; Willerslev et al. 2004a, b; Paabo et al. 2004; Willerslev and Cooper 2005).

Since the isolation of 250 million-year-old bacteria from salt crystals (Vreeland et al. 2000), the long-term survival of bacteria has been questioned in several publications (Graur and Pupko 2001; Nickle et al. 2002) and has been found very problematic (Hebsgaard et al. 2005). A recent study, however, investigated the long-term survival of bacteria in sealed permafrost samples up to 1 million years old (Johnson et al. 2007). The study followed strict criteria and showed that bacteria can survive in permafrost up to 500,000 years, which make this the first independently authenticated evidence for viable cells surviving that long.

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