New discoveries concerning Ötzi's history

Scientists are continually unearthing new facts about Homo sapiens from the mummified remains of Ötzi, the Copper Age man, who was discovered in a glacier in 1991. Five years ago, after Ötzi's genome was completely deciphered, it seemed that the wellspring of spectacular discoveries about the past would soon dry up. An international team of scientists working with paleopathologist Albert Zink and microbiologist Frank Maixner from the European Academy (EURAC) in Bozen/Bolzano have now succeeded in demonstrating the presence of Helicobacter pylori in Ötzi's stomach contents, a bacterium found in half of all humans today. The theory that humans were already infected with this stomach bacterium at the very beginning of their history could well be true. The scientists succeeded in decoding the complete genome of the bacterium.

When EURAC's Zink and Maixner first placed samples from the Iceman's stomach under the microscope in their ancient DNA Lab at EURAC, almost three years ago, they were initially sceptical.

"Evidence for the presence of the bacterium Helicobacter pylori is found in the stomach tissue of patients today, so we thought it was extremely unlikely that we would find anything because Ötzi's stomach mucosa is no longer there," explains Zink. Together with colleagues from the Universities of Kiel, Vienna and Venda in South Africa as well as the Max Planck Institute for the Science of Human History in Jena, the scientists tried to find a new way to proceed. "We were able to solve the problem once we hit upon the idea of extracting the entire DNA of the stomach contents," reports Maixner. "After this was successfully done, we were able to tease out the individual Helicobacter sequences and reconstruct a 5,300 year old Helicobacter pylori genome."

The scientists found a potentially virulent strain of bacteria, to which Ötzi's immune system had already reacted. "We showed the presence of marker proteins which we see today in patients infected with Helicobacter," said the microbiologist.

A tenth of infected people develop further clinical complications, such as gastritis or stomach ulcers, mostly in old age. "Whether Ötzi suffered from stomach problems cannot be said with any degree of certainty," says Zink, "because his stomach tissue has not survived and it is in this tissue that such diseases can be discerned first. Nonetheless, the preconditions for such a disease did in fact exist in Ötzi."

After completing their stomach biopsy, the two EURAC scientists transferred the genome data for analysis by their colleague Thomas Rattei from the University of Vienna. Rattei, in collaboration with geneticists from the USA, South Africa and Germany, came to a surprising conclusion: "We had assumed that we would find the same strain of Helicobacter in Ötzi as is found in Europeans today," explains the computational biologist. "It turned out to be a strain that is mainly observed in Central and South Asia today."

The scientists assume that there were originally two strain types of the bacterium, an African and an Asian one, which at some point recombined into today's European version. Since bacteria are usually transmitted within the family, the history of the world's population is closely linked to the history of bacteria. Up till now, it had been assumed that Neolithic humans were already carrying this European strain by the time they stopped their nomadic life and took up agriculture. Research on Ötzi, however, demonstrates that this was not the case.

"The recombination of the two types of Helicobacter may have only occurred at some point after Ötzi's era, and this shows that the history of settlements in Europe is much more complex than previously assumed," says Maixner.

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New research at the European Academy of Bolzano/Bozen (EURAC) further clarifies the genetic history of man who lived in the Eastern Alps over 5,300 years ago. In 2012 a complete analysis of the Y chromosome (transmitted from fathers to their sons) showed that Ötzi's paternal genetic line is still present in modern-day populations. In contrast, studies of mitochondrial DNA (transmitted solely via the mother to her offspring) left many questions still open. To clarify whether the genetic maternal line of the Iceman, who lived in the eastern Alps over 5,300 years ago, has left its mark in current populations, researchers at the European Academy of Bolzano/Bozen (EURAC) have now compared his mitochondrial DNA with 1,077 modern samples. The study concluded that the Iceman's maternal line -- named K1f -- is now extinct. A second part of the study, a comparison of genetic data of the mummy with data from other European Neolithic samples, provided information regarding the origin of K1f: researchers postulate that the mitochondrial lineage of the Iceman originated locally in the Alps, in a population that did not grow demographically. The study, which also clarifies Ötzi's genetic history in the context of European demographic changes from Neolithic times onwards, was published in Scientific Reports, an open access journal of the Nature group.

"The mummy's mitochondrial DNA was the first to be analysed, in 1994." says Valentina Coia, a biologist at EURAC and first author of the study. "It was relatively easy to analyse and -- along with the Y chromosome -- allows us to go back in time, telling us about the genetic history of an individual. Despite this, the genetic relationship between the Iceman's maternal lineage and lineages found in modern populations was not yet clear." The most recent study regarding the analysis of Ötzi's complete mitochondrial DNA, conducted in 2008 by other research teams showed that the Iceman's maternal lineage -- named K1f -- was no longer traceable in modern populations. The study did not make clear, however, whether this was due to an insufficient number of comparison samples or whether K1f was indeed extinct.

Valentina Coia explains further: "The first hypothesis could not be ruled out given that the study considered only 85 modern comparison samples from the K1 lineage -- the genetic lineage that also includes that of Ötzi -- which comprised few samples from Europe and especially none from the eastern Alps, which are home to populations that presumably have a genetic continuity with the Iceman. To test the two hypotheses, we needed to compare Ötzi's mitochondrial DNA with a larger number of modern samples." The EURAC research team, in collaboration with the Sapienza University of Rome and the University of Santiago de Compostela, thus compared the mitochondrial DNA of the Iceman with that from 1,077 individuals belonging to the K1 lineage, of which 42 samples originated from the eastern Alps and were for the first time analysed in this study. The new comparison showed that neither the Iceman's lineage nor any other evolutionarily close lineages are present in modern populations: the researchers therefore lean towards the hypothesis that Ötzi's maternal genetic branch has died out.

It remains to be explained why Ötzi's maternal lineage has disappeared, while his paternal lineage-- named G2a--still exists in Europe. To clarify this point, researchers at EURAC compared Ötzi's mitochondrial DNA and Y chromosome with available data from numerous ancient samples found at 14 different archaeological sites throughout Europe. The results showed that the paternal lineage of Ötzi was very common in different regions in Europe during the Neolithic age, while his maternal lineage probably existed only in the Alps.

Putting together the genetic data on the ancient and modern samples, namely those already present in the literature and those analysed in this study, researchers have now proposed the following scenario to explain the Iceman's genetic history: Ötzi's paternal lineage, G2a, is part of an ancient genetic substrate that arrived in Europe from the Near East with the migrations of the first Neolithic peoples some 8,000 years ago. Additional migrations and other demographic events occurring after the Neolithic Age in Europe then partially replaced G2a with other lineages, except in geographically isolated areas such as Sardinia. In contrast, the Iceman's maternal branch originated locally in the eastern Alps at least 5,300 years ago. The same migrations that have replaced only in part his paternal lineage caused the extinction of his maternal lineage that was inherited in a small and demographic stationary population. The groups from the eastern Alps in fact significantly increased in size only from the Bronze Age onwards, as evidenced by archaeological studies conducted in the territory inhabited by the Iceman.