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Data files for the above publication. ABSTRACT Natural environments harbor a vast and largely unexplored microbial biodiversity. Among these, the Mar Menor, Europe's largest hypersaline coastal lagoon located in southeastern Spain, represents a unique ecosystem whose microbial communities remain poorly characterized. Given the increasing need for CRISPR–Cas systems optimized for diverse biotechnological applications, we sought to identify and classify previously uncharacterized CRISPR–Cas systems, with particular emphasis on novel Cas9 variants. The investigation was conducted through metagenomic analyses of the Mar Menor microbiome, with samples collected between 2019 and 2022. A total of 18 metagenomes were analyzed, generating over 600,000 contigs larger than 2 kb. Among these, 24 complete CRISPR–Cas systems were identified, comprising 58% Class 1 and 42% Class 2 systems. The distribution of subtypes varied based on the plankton size fraction (0.22–3 μm and 3–200 μm) and reflected seasonal dynamics in the microbial communities. All identified CRISPR–Cas systems were of bacterial origin, despite Archaea typically having a higher prevalence of CRISPR–Cas systems. Further phylogenetic analysis of 142 Cas9 proteins revealed a monophyletic clade consisting of 89 sequences from the lagoon, suggesting the existence of previously uncharacterized evolutionary lineages adapted to its unique environmental conditions. This study significantly expands the known diversity of CRISPR–Cas systems, highlighting novel lineages that could have applications in industrial biotechnology. The findings open new opportunities for developing CRISPR-based gene editing tools optimized for extreme environments.