Ancient bacteria of the Ötzi's microbiome: a genomic tale from the Copper Age

Abstract

Background: Ancient microbiota information represents an important resource to evaluate bacterial evolution and to explore the biological spread of infectious diseases in history. The soft tissue of frozen mummified humans, such as the Tyrolean Iceman, has been shown to contain bacterial DNA that is suitable for population profiling of the prehistoric bacteria that colonized such ancient human hosts.

Results: Here, we performed a microbial cataloging of the distal gut microbiota of the Tyrolean Iceman, which highlights a predominant abundance of Clostridium and Pseudomonas species. Furthermore, in silico analyses allowed the reconstruction of the genome sequences of five ancient bacterial genomes, including apparent pathogenic ancestor strains of Clostridium perfringens and Pseudomonas veronii species present in the gut of the Tyrolean Iceman.

Conclusions: Genomic analyses of the reconstructed C. perfringens chromosome clearly support the occurrence of a pathogenic profile consisting of virulence genes already existing in the ancient strain, thereby reinforcing the notion of a very early speciation of this taxon towards a pathogenic phenotype. In contrast, the evolutionary development of P. veronii appears to be characterized by the acquisition of antibiotic resistance genes in more recent times as well as an evolution towards an ecological niche outside of the (human) gastrointestinal tract.

Keywords: Genomic evolution; Genomics; Gut bacteria; Metagenomics.

Fig. 1

Bacterial abundance in the Ötzi’s gut. Panel a displays a bar plot with the abundance of the major species identified in the Tyrolean Iceman gut using CoCla script. The x axis represents the identified bacterial species, while the y axis represents the number of nucleotides assembled in contigs. Each color reflects a specific sample, i.e., B0625 (lower part of the large intestine), C1824 and C1825 (upper part of the large intestine), and B0621 (small intestine). Panel b shows the visual abundance of the identified species

Fig. 2

Phylogenetic diversity of the reconstructed ancient genomes. Panel a depicts a phylogenetic supertree based on the sequences of identified core proteins shared by the analyzed C. perfringens genomes. Panel b shows the same supertree based on the analyzed P. fluorescent genomes, while panel c displays the P. veronii supertee

Fig. 3

Comparative genomic analysis of P. veronii PVER with other fully sequenced P. veronii strains. Circular genome atlas of P. veronii PVER (red circle) with mapped orthologues (defined as reciprocal best BLASTp hits with more than 50% identity over at least 50% of both protein lengths) in four publicly available P. veronii genomes (orange through green circle). Internal circles illustrate P. veronii PVER GC% deviation and GC skew (G − C/G + C), while the external maps exhibit the sequence identity between the unique loci of P. veronii PVER compared to other bacteria retrieved from the database. Each arrow indicates an ORF, whereas the length of the arrow is proportional to the length of the predicted ORF. Red arrows correspond to the P. veronii PVER genes, while orange arrows display orthologous genes

Fig. 4

Carbohydrate-active enzymes of the ancient genomes. Panel a exhibits a bar plot with the abundance in Glycosyl hydrolase (GH) or Glycosyltransferase (GT) families encoded by the genomes of Clostridium sp. CADE and Clostridium sp. Ade.TY. Panel b shows a similar bar plot for C. algidicarnis CALG and C. algidicarnis B3, while panels c, d, and e display the GH and GT average between the publicly available genomes of C. perfringens, P. fluorescens and P. veronii, respectively