20th International Cave Bear Symposium (ICBS 2014) – September 10th-13th 2014, Corvara/San Cassiano (South Tyrol), Italy “URSUS PROJECT”: A MULTIDISCIPLINARY STUDY OF THE NORTHERN ITALY CAVE BEAR Ermanno Rizzi1, Mario Rossi2, David Caramelli3, Maria Gabriella Fornasiero4, Alex Fontana5, Alessandro Garassino6, Alessandra Modi3, Paolo Schirolli7, Roberto Zorzin2 & Giuseppe Santi8. 1Institute for Biomedical Technologies (ITB), National Research Council (CNR), Segrate (MI) ; 2Museo Civico di Storia Naturale, Verona ; 3Laboratory of Anthropology, Department of Biology, University of Florence, Florence; 4Geological and Paleontological Museum, Department of Geoscience, Padova; 5MuSe, Museo delle Scienze, Trento; 6Museo Civico di Storia Naturale, Milano; 7Museo Civico di Storia Naturale, Brescia; 8Dipartimento di Scienze della Terra e dell’Ambiente, Pavia. Background. Evolutionary studies of the cave bear (Ursus speleaus) are possible thanks to a large number of available retrievals due to its wide diffusion in Europe and Asia during Pleistocene. The high morphological variability detected in European samples, suggested that palaeontologists consider the presence of local evolutionary trends. Recently, the morphological and morphometric analyses of cave bear bones and teeth, were complemented by genetic data performed on the mitochondrial genome. As a consequence, new taxa have been proposed. S. Donà di Lamon Buco dell’Orso (Laglio) Covoli di Velo a Fig. 1 - Position of some selected cave in Northern Italy Tab. 1. Data from Santi & Rossi (Annales de Paléontologie, 100, 237-256 2014) b Fig. 2a - Entries of the Buco dell’Orso in Laglio and 2b of Covoli di Velo (Verona). (kindly provided by: www.comune.di laglio.co.it and www.fabiobona.it). Procedures. The “Ursus project” is based on a multidisciplinary approach to study the cave bear variability of samples recovered from Northern Italian sites. Two sets of data will be considered: 1) morphometric and morphological measures, 2) complete mitochondrial DNA genetic sequence (mtDNA). As an example of morphometric measures in Table 1, the morphodynamic indices (IM) of P4/4 of the cave bears from the most important caves are shown. The samples recovered from different sites of Northern Italy (Fig. 1 and 2a and 2b), will be classified (Fig. 3) and studied under the ancient DNA (aDNA) perspective. After DNA isolation in dedicated “clean rooms” (Fig. 4), the samples will be analysed by the combined approach of genomic capture and next generation sequencing (NGS) (Fig. 5). The genomic capture will be performed using specific oligonucleotides probes, called “molecular baits”, that selectively isolate the endogenous bear aDNA, excluding all possible contaminant sequences and preventing typical problems in aDNA sequencing. The study of the cave bear population will be based on the identification of the haplogroups by a phylogenetic clustering. The cave bear genetic population results will be compared to the available extant bear mtDNA sequences in order to establish a hypothetical gene flow. Fig. 3 – Classified bones and teeth samples from excavation and museum collections. Fig. 4 – A “clean room” for the isolation and managing of aDNA. Goals of the project. The topics of this project are: 1) to integrate the knowledge of the Italian populations using the genetic data and, if possible, the chronological ones, 2) to individuate eventual schemes of distribution allowing to correlate both genetic and morphological-morphometrical data. Fig. 5 – Flowchart of data generation. Once the aDNA is converted to NGS library, a capture step will be performed to enrich the sample in cave bear mitochondrial DNA. NGS runs will be performed using the Illumina (MiSeq) platform, generating millions of reads that will be assembled to obtain the complete mtDNA for the further phylogenetic analysis. References and contacts. Rizzi E. et al. (2012) – Genetics Selection Evolution, 44(21), 1-19. [email protected] Santi G. & Rossi M. (2014) - Annales de Paléontologie, 100, 237-256. [email protected] and [email protected]