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Background The contemporary Italian wolf (Canis lupus italicus) represents a case of morphological and genetic uniqueness. Today, Italian wolves are also the only documented population to fall exclusively within the mitochondrial haplogroup 2, which was the most diffused across Eurasian and North American wolves during the Late Pleistocene. However, the dynamics leading to such distinctiveness are still debated. Methods In order to shed light on the ancient genetic variability of this wolf population and on the origin of its current diversity, we collected 19 Late Pleistocene-Holocene samples from northern Italy, which we analyzed at a short portion of the hypervariable region 1 of the mitochondrial DNA, highly informative for wolf and dog phylogenetic analyses. Results Four out of the six detected haplotypes matched the ones found in ancient wolves from northern Europe and Beringia, or in modern European and Chinese wolves, and appeared closely related to the two haplotypes currently found in Italian wolves. The haplotype of two Late Pleistocene samples matched with primitive and contemporary dog sequences from the canine mitochondrial clade A. All these haplotypes belonged to haplogroup 2. The only exception was a Holocene sample dated 3,250 years ago, affiliated to haplogroup 1. Discussion In this study we describe the genetic variability of the most ancient wolf specimens from Italy analyzed so far, providing a preliminary overview of the genetic make-up of the population that inhabited this area from the last glacial maximum to the Middle Age period. Our results endorsed that the genetic diversity carried by the Pleistocene wolves here analyzed showed a strong continuity with other northern Eurasian wolf specimens from the same chronological period. Contrarily, the Holocene samples showed a greater similarity only with modern sequences from Europe and Asia, and the occurrence of an haplogroup 1 haplotype allowed to date back previous finding about its presence in this area. Moreover, the unexpected discovery of a 24,700-year-old sample carrying a haplotype that, from the fragment here obtained, falls within the canine clade A, could represent the oldest evidence in Europe of such dog-rich clade. All these findings suggest complex population dynamics that deserve to be further investigated based on mitochondrial or whole genome sequencing.

Summary The evolution of the genera Bos and Bison , and the nature of gene flow between wild and domestic species, is poorly understood, with genomic data of wild species being limited. We generated two genomes from the likely extinct kouprey ( Bos sauveli ) and analyzed them alongside other Bos and Bison genomes. We found that B. sauveli possessed genomic signatures characteristic of an independent species closely related to Bos javanicus and Bos gaurus . We found evidence for extensive incomplete lineage sorting across the three species, consistent with a polytomic diversification of the major ancestry in the group, potentially followed by secondary gene flow. Finally, we detected significant gene flow from an unsampled Asian Bos -like source into East Asian zebu cattle, demonstrating both that the full genomic diversity and evolutionary history of the Bos complex has yet to be elucidated and that museum specimens and ancient DNA are valuable resources to do so.

During historical times many local grey wolf (Canis lupus) populations underwent a substantial reduction of their sizes or became extinct. Among these, the wolf population once living in Sicily, the biggest island of the Mediterranean Sea, was completely eradicated by human persecution in the early decades of the XX century. In order to understand the genetic identity of the Sicilian wolf, we applied ancient DNA techniques to analyse the mitochondrial DNA of six specimens actually stored in Italian museums. We successfully amplified a diagnostic mtDNA fragment of the control region (CR) in four of the samples. Results showed that two samples shared the same haplotype, that differed by two substitutions from the currently most diffused Italian wolf haplotype (W14) and one substitution from the only other Italian haplotype (W16). The third sample showed a wolf-like haplotype never described before and the fourth a haplotype commonly found in dogs. Furthermore, all the wolf haplotypes detected in this study belonged to the mitochondrial haplogroup that includes haplotypes detected in all the known European Pleistocene wolves and in several modern southern European populations. Unfortunately, this endemic island population, bearing unique mtDNA variability, was definitively lost before it was possible to understand its taxonomic uniqueness and conservational value.