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Classification of ancient mammal individuals using dental pulp MALDI-TOF MS peptide profiling.

Tran TN, Aboudharam G, Gardeisen A, Davoust B, Bocquet-Appel JP, Flaudrops C, Belghazi M, Raoult D, Drancourt M - PLoS ONE (2011)

Bottom Line: Our work was organized into three successive steps.The resulting peptide profiles accurately classified every individual at the species level in agreement with parallel cytochrome b gene sequencing gold standard.The dental pulp is a new source for collagen and other proteins for the species classification of modern and ancient mammal individuals.

View Article: PubMed Central - PubMed

Affiliation: Unité de Recherche sur les Maladies Infectieuses et Tropicales Émergentes, UMR CNRS 6236 IRD 198, IFR48, Faculté de Médecine, Université de la Méditerranée, Marseille, France.

ABSTRACT

Background: The classification of ancient animal corpses at the species level remains a challenging task for forensic scientists and anthropologists. Severe damage and mixed, tiny pieces originating from several skeletons may render morphological classification virtually impossible. Standard approaches are based on sequencing mitochondrial and nuclear targets.

Methodology/principal findings: We present a method that can accurately classify mammalian species using dental pulp and mass spectrometry peptide profiling. Our work was organized into three successive steps. First, after extracting proteins from the dental pulp collected from 37 modern individuals representing 13 mammalian species, trypsin-digested peptides were used for matrix-assisted laser desorption/ionization time-of-flight mass spectrometry analysis. The resulting peptide profiles accurately classified every individual at the species level in agreement with parallel cytochrome b gene sequencing gold standard. Second, using a 279-modern spectrum database, we blindly classified 33 of 37 teeth collected in 37 modern individuals (89.1%). Third, we classified 10 of 18 teeth (56%) collected in 15 ancient individuals representing five mammal species including human, from five burial sites dating back 8,500 years. Further comparison with an upgraded database comprising ancient specimen profiles yielded 100% classification in ancient teeth. Peptide sequencing yield 4 and 16 different non-keratin proteins including collagen (alpha-1 type I and alpha-2 type I) in human ancient and modern dental pulp, respectively.

Conclusions/significance: Mass spectrometry peptide profiling of the dental pulp is a new approach that can be added to the arsenal of species classification tools for forensics and anthropology as a complementary method to DNA sequencing. The dental pulp is a new source for collagen and other proteins for the species classification of modern and ancient mammal individuals.

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The dendrogram obtained by the software Maldi Biotyper 2.0 (Bruker Daltonics) after peptide spectral analysis is congruent with the one derived from cytochrome b sequencing by the software Tree View (Free down load from the Internet site:http://darwin.zoology.gla.ac.uk/~rpage/treeviewx/download.html). The images indicate the mammalian species (left) from which the ancient teeth were identified (right).
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pone-0017319-g001: The dendrogram obtained by the software Maldi Biotyper 2.0 (Bruker Daltonics) after peptide spectral analysis is congruent with the one derived from cytochrome b sequencing by the software Tree View (Free down load from the Internet site:http://darwin.zoology.gla.ac.uk/~rpage/treeviewx/download.html). The images indicate the mammalian species (left) from which the ancient teeth were identified (right).

Mentions: We applied the same approach to ancient teeth. By using the software MALDI Biotyper 2.0 and comparing with our modern database, we could classify 10/18 (56%) teeth collected in 8/15 (53.3%) individuals from five burial sites dating between 700 and 8,500 years ago (Table S6). In these specimens, protein concentrations of 0.28±0.12 g/L yielded reproducible MALDI-TOF MS profiles that differed by 1–17 USSPs (Table S7). We did not observe contamination from one species to another one in this study. Peptide sequencing confirmed these data by giving 4 different proteins including collagen (alpha-1 type I and alpha-2 type I) in ancient human dental pulp (Table S4). Two isolated teeth collected from a first- or second-century site were identified as cow, four other teeth as dog and one additional tooth as cat. After these profiles of ancient individuals were incorporated into ancient mammal database, further comparison of teeth that had not been identified by first-round comparison with modern profiles yielded 100% identification with cow, cat, pig and human, with identification scores of≥1.8. All MALDI-TOF MS results were in agreement with the observed morphological characteristics and were confirmed by cytochrome b sequencing (Figure 1, Table S8). We therefore successfully identified the remains of individuals representing four domestic mammalian species frequently encountered in excavations. Two isolated teeth collected from an 18th-century site were identified as human, as were teeth collected from 300-year-old burial site in France. This approach also proved effective in classifying 8,500-year-old human dental pulp specimens collected in Syria from the deciduous teeth of two Neolithic children (Figure 2).


Classification of ancient mammal individuals using dental pulp MALDI-TOF MS peptide profiling.

Tran TN, Aboudharam G, Gardeisen A, Davoust B, Bocquet-Appel JP, Flaudrops C, Belghazi M, Raoult D, Drancourt M - PLoS ONE (2011)

The dendrogram obtained by the software Maldi Biotyper 2.0 (Bruker Daltonics) after peptide spectral analysis is congruent with the one derived from cytochrome b sequencing by the software Tree View (Free down load from the Internet site:http://darwin.zoology.gla.ac.uk/~rpage/treeviewx/download.html). The images indicate the mammalian species (left) from which the ancient teeth were identified (right).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3045434&req=5

pone-0017319-g001: The dendrogram obtained by the software Maldi Biotyper 2.0 (Bruker Daltonics) after peptide spectral analysis is congruent with the one derived from cytochrome b sequencing by the software Tree View (Free down load from the Internet site:http://darwin.zoology.gla.ac.uk/~rpage/treeviewx/download.html). The images indicate the mammalian species (left) from which the ancient teeth were identified (right).
Mentions: We applied the same approach to ancient teeth. By using the software MALDI Biotyper 2.0 and comparing with our modern database, we could classify 10/18 (56%) teeth collected in 8/15 (53.3%) individuals from five burial sites dating between 700 and 8,500 years ago (Table S6). In these specimens, protein concentrations of 0.28±0.12 g/L yielded reproducible MALDI-TOF MS profiles that differed by 1–17 USSPs (Table S7). We did not observe contamination from one species to another one in this study. Peptide sequencing confirmed these data by giving 4 different proteins including collagen (alpha-1 type I and alpha-2 type I) in ancient human dental pulp (Table S4). Two isolated teeth collected from a first- or second-century site were identified as cow, four other teeth as dog and one additional tooth as cat. After these profiles of ancient individuals were incorporated into ancient mammal database, further comparison of teeth that had not been identified by first-round comparison with modern profiles yielded 100% identification with cow, cat, pig and human, with identification scores of≥1.8. All MALDI-TOF MS results were in agreement with the observed morphological characteristics and were confirmed by cytochrome b sequencing (Figure 1, Table S8). We therefore successfully identified the remains of individuals representing four domestic mammalian species frequently encountered in excavations. Two isolated teeth collected from an 18th-century site were identified as human, as were teeth collected from 300-year-old burial site in France. This approach also proved effective in classifying 8,500-year-old human dental pulp specimens collected in Syria from the deciduous teeth of two Neolithic children (Figure 2).

Bottom Line: Our work was organized into three successive steps.The resulting peptide profiles accurately classified every individual at the species level in agreement with parallel cytochrome b gene sequencing gold standard.The dental pulp is a new source for collagen and other proteins for the species classification of modern and ancient mammal individuals.

View Article: PubMed Central - PubMed

Affiliation: Unité de Recherche sur les Maladies Infectieuses et Tropicales Émergentes, UMR CNRS 6236 IRD 198, IFR48, Faculté de Médecine, Université de la Méditerranée, Marseille, France.

ABSTRACT

Background: The classification of ancient animal corpses at the species level remains a challenging task for forensic scientists and anthropologists. Severe damage and mixed, tiny pieces originating from several skeletons may render morphological classification virtually impossible. Standard approaches are based on sequencing mitochondrial and nuclear targets.

Methodology/principal findings: We present a method that can accurately classify mammalian species using dental pulp and mass spectrometry peptide profiling. Our work was organized into three successive steps. First, after extracting proteins from the dental pulp collected from 37 modern individuals representing 13 mammalian species, trypsin-digested peptides were used for matrix-assisted laser desorption/ionization time-of-flight mass spectrometry analysis. The resulting peptide profiles accurately classified every individual at the species level in agreement with parallel cytochrome b gene sequencing gold standard. Second, using a 279-modern spectrum database, we blindly classified 33 of 37 teeth collected in 37 modern individuals (89.1%). Third, we classified 10 of 18 teeth (56%) collected in 15 ancient individuals representing five mammal species including human, from five burial sites dating back 8,500 years. Further comparison with an upgraded database comprising ancient specimen profiles yielded 100% classification in ancient teeth. Peptide sequencing yield 4 and 16 different non-keratin proteins including collagen (alpha-1 type I and alpha-2 type I) in human ancient and modern dental pulp, respectively.

Conclusions/significance: Mass spectrometry peptide profiling of the dental pulp is a new approach that can be added to the arsenal of species classification tools for forensics and anthropology as a complementary method to DNA sequencing. The dental pulp is a new source for collagen and other proteins for the species classification of modern and ancient mammal individuals.

Show MeSH
Related in: MedlinePlus