The DNA is a nucleic acid that contains the genetic information necessary for RNA and protein biosynthesis. DNA extracted from past samples (teeth, bones, faeces, etc.) is defined ancient DNA (aDNA) and needs, to preserve, low level of oxygen, fast decrease in water content and, above all, according to the Arrhenius equation, low temperature. From 1985 (first aDNA extracted from a mummy) with the introduction of the Polymerase Chain Reaction (PCR) several studies of paleogenomic were born, but it is essential to identify some possible errors such as fragmentations, contaminations and post-mortal mutations. The applications of the paleogenomic are: evolutionary biology, population studies, studies of the pathogens and microorganism. We report 3 cases studied by the Division of Paleopathology of Pisa: a) Ferrante I, king of Naples (1431-1494). The natural mummy showed round white formations of the pelvis infiltrating the abdominal wall. b) Maria of Aragon (1503-1568). The well-preserved artificial mummy had a small peduncolate arborescence neoformation in the right inguinal region. An Andean female mummy (so-called ‘Fi9’) dated 10th–11th century A.D by radiocarbonium analyses. The natural young mummy presented a marked megavisceral syndrome characterized by megacolon, megaoesophagus and cardiomegaly. It was possible to perform complete autopsies and collect tissue samples utilized for histological analyses and DNA extraction. a) Histology performed on the round formations confirmed the diagnosis of colorectal adenocarcinoma. Amplification of aDNA highlighted a point mutation of the codon 12 in K-Ras oncogene responsible for the cancer. b) Macroscopic and histological aspects seemed peculiar of condyloma acuminatum, a papillomavirus-induced squamous lesion also called “venereal wart”. Molecular study revealed the presence of HPV 18, a virus with high oncogenic potential. Automated sequencing of several clones revealed 100% similarity sequences of both HPV 18 and JC9813 DNA, a putative novel HPV with low oncogenic potential c) Analysis of the gut microbiome (paleofeces, descending, transverse and ascending colon) underlined the massive presence of Clostridiceae. Sequences homologous to HPVs in the mummified gut (descending colon) was particularly surprising. It was detected also the Tripanosoma cruzi; by comparing a partial sequence homologous to the large ribosomal subunit alpha of the presumptive ancient T. cruzi with modern strains, we suggest that this pathogen may have a more remote origin than previously expected. We also found sequences associated with putative beta-lactamases, penicillin-binding proteins, resistance to fosfomycin, chloramphenicol, aminoglycosides, macrolides, sulfa, quinolones, tetracycline and vancomycin, and multi-drug transporters. Conclusion a) The alimentary “environment” of the Neapolitan court of the XV century, with its abundance of natural alimentary alkylating agents (red smoked meat), well explains the acquired mutation of K-Ras. b) This represented the first molecular diagnosis of HPV in mummies. HPV is a very old virus that evolved together with man. c) Streptococcus, Staphylococcus, Bacillus and Pseudomonas sequences were identified in the mummified gut, opening the opportunity to investigate possible mechanisms by which these bacteria are preserved. The detection of sequences homologous to those of pathogens such as T. cruzi and HPV indicate their presence in the Americas prior to European colonization. The presence of antibiotic-resistance genes in an 11th century pre-Columbian Andean mummy is intriguing as antibiotics were introduced recently. The presence of beta-lactam antibiotic resistance is certainly not unexpected in any culture, as would be in the case of resistance to any natural rather than a semi- or completely synthetic antibiotic as a result of exposure to natural antibiotic-producing microbiota originating from the environment (e.g. soil); however, vancomycin, particularly, was discovered more than 50 years ago, and vancomycin-resistance genes have been mainly implicated with the increased use of this antibiotic. The presence of antibiotic-resistance genes in the ancient human gut microbiome clearly indicates that these genes pre-date therapeutical use of these compounds and that they are not necessarily associated to a selective pressure of antibiotics use. Identification of pathogens and antibiotic-resistance genes in ancient human specimens will aid in the understanding of the evolution of pathogens as a way to treat and prevent diseases caused by bacteria, microbial eukaryotes and viruses. References Willerslev E, Cooper A. Review paper. Ancient dna. Proceedings of the Royal Society of London B: Biological Sciences. 2005; 272(1558):3– 16. Marchetti A, Pellegrini S, Bevilacqua G, Fornaciari G. K-RAS mutation in the tumour of Ferrante I of Aragon, King of Naples. The Lancet; 1996, May 4;347(9010):1272. Fornaciari G, Zavaglia K, Ciranni R. Human papillomavirus in a 16th century mummy. The Lancet; 2003, Oct 3, vol 362. Santiago-Rodriguez TM, Fornaciari G, Luciani S, Dowd SE, Toranzos GA, Marota I, Cano RJ. Gut Microbiome of an 11th Century A.D. PreColumbian Andean Mummy. PLoS One, 2015 Sep 30;10(9).
|Titolo:||Paleoleogenomics and ancient DNA|
GAETA, RAFFAELE (Primo)
FORNACIARI, GINO (Ultimo)
|Anno del prodotto:||2016|
|Appare nelle tipologie:||4.1 Contributo in Atti di convegno|