Cristina Olivieri

Complete Mitochondrial Genome Sequence of the Tyrolean Iceman

The Tyrolean Iceman was a witness to the Neolithic-Copper Age transition in Central Europe 5350-5100 years ago, and his mummified corpse was recovered from an Alpine glacier on the Austro-Italian border in 1991 [1]. Using a mixed sequencing procedure based on PCR amplification and 454 sequencing of pooled amplification products, we have retrieved the first complete mitochondrial-genome sequence of a prehistoric European. We have then compared it with 115 related extant lineages from mitochondrial haplogroup K. We found that the Iceman belonged to a branch of mitochondrial haplogroup K1 that has not yet been identified in modern European populations. This is the oldest complete Homo sapiens mtDNA genome generated to date. The results point to the potential significance of complete-ancient-mtDNA studies in addressing questions concerning the genetic history of human populations that the phylogeography of modern lineages is unable to tackle.

Tracking Plant, Fungal, and Bacterial DNA in Honey Specimens*

Abstract:  Consuming honey can result in adverse effects owing to poisoning by bacterial (botulism) or plant toxins. We have devised a method to extract polymerase chain reaction (PCR) amplifiable DNA of up to c. 400 bp in length based on dialysis of a 15‐mL honey sample for 18 h against deionized water followed by sequential extraction using phenol, phenol/chloroform/isoamyl alcohol, chloroform/isoamyl alcohol, and ether. Sequence analysis of PCR products obtained using “universal” plant, fungal, and bacterial primers targeted to the ribosomal RNA genes has allowed us to identify six different orders of plants (Apiales, Fabales, Asterales, Solanales, Brassicales, and Sapindales), two orders of fungi (Entylomatales and Saccharomycetales), and six orders of bacteria (Sphingomonadales, Burkholderiales, Pseudomonadales, Enterobacteriales, Actinomycetales, and Bifidobacteriales) in a single honey specimen.

Characterization of Nucleotide Misincorporation Patterns in the Iceman's Mitochondrial DNA

Background The degradation of DNA represents one of the main issues in the genetic analysis of archeological specimens. In the recent years, a particular kind of post-mortem DNA modification giving rise to nucleotide misincorporation (“miscoding lesions”) has been the object of extensive investigations. Methodology/Principal Findings To improve our knowledge regarding the nature and incidence of ancient DNA nucleotide misincorporations, we have utilized 6,859 (629,975 bp) mitochondrial (mt) DNA sequences obtained from the 5,350–5,100-years-old, freeze-desiccated human mummy popularly known as the Tyrolean Iceman or Ötzi. To generate the sequences, we have applied a mixed PCR/pyrosequencing procedure allowing one to obtain a particularly high sequence coverage. As a control, we have produced further 8,982 (805,155 bp) mtDNA sequences from a contemporary specimen using the same system and starting from the same template copy number of the ancient sample. From the analysis of the nucleotide misincorporation rate in ancient, modern, and putative contaminant sequences, we observed that the rate of misincorporation is significantly lower in modern and putative contaminant sequence datasets than in ancient sequences. In contrast, type 2 transitions represent the vast majority (85%) of the observed nucleotide misincorporations in ancient sequences. Conclusions/Significance This study provides a further contribution to the knowledge of nucleotide misincorporation patterns in DNA sequences obtained from freeze-preserved archeological specimens. In the Iceman system, ancient sequences can be clearly distinguished from contaminants on the basis of nucleotide misincorporation rates. This observation confirms a previous identification of the ancient mummy sequences made on a purely phylogenetical basis. The present investigation provides further indication that the majority of ancient DNA damage is reflected by type 2 (cytosine→thymine/guanine→adenine) transitions and that type 1 transitions are essentially PCR artifacts.

Phylogenetic Position of a Copper Age Sheep (Ovis aries) Mitochondrial DNA

Background Sheep (Ovis aries) were domesticated in the Fertile Crescent region about 9,000-8,000 years ago. Currently, few mitochondrial (mt) DNA studies are available on archaeological sheep. In particular, no data on archaeological European sheep are available. Methodology/Principal Findings Here we describe the first portion of mtDNA sequence of a Copper Age European sheep. DNA was extracted from hair shafts which were part of the clothes of the so-called Tyrolean Iceman or Ötzi (5,350 - 5,100 years before present). Mitochondrial DNA (a total of 2,429 base pairs, encompassing a portion of the control region, tRNAPhe, a portion of the 12S rRNA gene, and the whole cytochrome B gene) was sequenced using a mixed sequencing procedure based on PCR amplification and 454 sequencing of pooled amplification products. We have compared the sequence with the corresponding sequence of 334 extant lineages. Conclusions/Significance A phylogenetic network based on a new cladistic notation for the mitochondrial diversity of domestic sheep shows that the Ötzis sheep falls within haplogroup B, thus demonstrating that sheep belonging to this haplogroup were already present in the Alps more than 5,000 years ago. On the other hand, the lineage of the Ötzis sheep is defined by two transitions (16147, and 16440) which, assembled together, define a motif that has not yet been identified in modern sheep populations.

Positioning the Red Deer (Cervus elaphus) Hunted by the Tyrolean Iceman into a Mitochondrial DNA Phylogeny

In the last years several phylogeographic studies of both extant and extinct red deer populations have been conducted. Three distinct mitochondrial lineages (western, eastern and North-African/Sardinian) have been identified reflecting different glacial refugia and postglacial recolonisation processes. However, little is known about the genetics of the Alpine populations and no mitochondrial DNA sequences from Alpine archaeological specimens are available. Here we provide the first mitochondrial sequences of an Alpine Copper Age Cervus elaphus. DNA was extracted from hair shafts which were part of the remains of the clothes of the glacier mummy known as the Tyrolean Iceman or Ötzi (5,350–5,100 years before present). A 2,297 base pairs long fragment was sequenced using a mixed sequencing procedure based on PCR amplifications and 454 sequencing of pooled amplification products. We analyzed the phylogenetic relationships of the Alpine Copper Age red deers haplotype with haplotypes of modern and ancient European red deer. The phylogenetic analyses showed that the haplotype of the Alpine Copper Age red deer falls within the western European mitochondrial lineage in contrast with the current populations from the Italian Alps belonging to the eastern lineage. We also discussed the phylogenetic relationships of the Alpine Copper Age red deer with the populations from Mesola Wood (northern Italy) and Sardinia.

Craniofacial superimposition studies on a set of cosimo i de’ medici’s (1519-1574) and eleonora di toledo’s (1522-1562) portraits

A set of portraits of the Florentine grand duke Cosimo I de’ Medici (1519-1574) and of his wife Eleonora di Toledo (1522–1562) painted by two major representatives of Florentine Mannerism (Italian Late Renaissance), Pontormo and Bronzino was compared with the skulls of the two subjects utilizing the forensic technique of craniofacial superimposition. The results show that, in the case of Cosimo I, both a painting (Pontormo, workshop of, Cosimo I de’ Medici, 1537) and a drawing (Pontormo, Cosimo I de’ Medici in Profile, 1537) show lack of fit with the skull in four points (the glabellar outline, the depth of the nasal bridge, the bony lateral wall of the orbit, and the outline of the frontal process of the zygomatic bone). The drawing, therefore, seems to contradict the well established idea that Renaissance painters prepared lifelike sketches of their sitters which were then modified when transferred to the painting. In the case of Eleonora di Toledo, on the other hand, craniofacial superimposition analysis reveals that Bronzino, possibly as the result of a desperate search for the “best angle” of the sitter, adopted an unusual perspective to portray the duchess (Bronzino, Eleonora di Toledo, c. 1539) possibly looking down the sitter, by standing, while the sitter was seated in front of him. The face of the sitter was subsequently, in another painting (Bronzino, Eleonora di Toledo and Her Son Giovanni, c. 1544-45) “transplanted” onto the rest of the body given the impression that the duchess “looks” at the viewer from above.