Background Antimicrobial resistance is one of the major challenges affecting public health . It is mostly due to the continuous emergence of extended-spectrum β-lactamase from various environments followed by their rapid dissemination and selection in clinical settings . The warming of Earth ’ s climate is the other global threat facing human society, in particular with the Arctic regions experiencing a twice faster warming than the global average and permafrost affected by widespread thawing . A potentially dreadful combination of these two threats would be the release and dispersion of harmful microbes that have remained confined to largely uninhabited Arctic regions, or are stored dormant in permafrost . Methods Environmental DNA was isolated from 12 soil samples from various Arctic and subarctic pristine regions in Siberia (Yakutia and Kamchatka), including nine permafrost samples collected at various depths . The large datasets obtained from high throughput sequencing was assembled in contigs and their protein-gene contents predicted . We used exhaustive similarity searches to perform taxonomical assignments of bacterial, archaeal, and eukaryotic organisms, as well as DNA viruses . In addition, we specifically identified β-lactamase genes and their prevalence per bacterial genome estimated through the detection of two universal single copy genes . Findings A total of 9.217 1011 bp were exploited, leading to a total of 525,313 contigs at least 5kb in size . The DNA content of the various samples was found to be highly variable, not strictly correlated with the depth or radio-carbon-based deposit age, and most likely linked to the global density of microbes trapped in the corresponding permafrost layers . Bacteria account for more than 90% of the contigs in most samples, followed by Eukaryotes and Archaea (always lower than 10 %). Viruses represented less than 2% of all contigs in all samples . The taxonomic profiles of surface cryosoils and deep permafrost samples exhibited a high diversity, including between permafrost samples originating from various depths in the same borehole . In all samples, bacterial contigs carrying different β-lactamases from class A to D were identified . Interpretation No clear common taxonomic feature could be found shared by surface cryosoils or ancient permafrost layers . However, most samples (9/12) exhibited a high frequency of β-lactamase genes, with an estimated average close to 1 copy/bacterial genome . In addition to the well-documented reactivation of infectious ancient pathogens (bacteria, viruses, protozoa), we show now that global warming could contribute to the emergence of new antibiotic resistances through the mobilization by contemporary bacteria of ancient DNA released from thawing permafrost . Funding CNRS PRC research grant (PRC1484-2018) to C.A . E C-F was supported by a PhD grant (DGA/DS/MRIS) #2017 60 0004 . GG and JS were funded by ERC PETA-CARB (# 338335) and the HGF Impulse and Networking Fund (ERC-0013).