CRISPR spacers acquired from plasmids primarily target backbone genes, making them valuable for predicting potential hosts and host range

In recent years, there has been a surge in metagenomic studies focused on identifying plasmids in environmental samples. Although these studies have unearthed numerous novel plasmids, enriching our understanding of their environmental roles, a significantgap remains: the scarcity of information regarding the bacterial hosts of these newly discovered plasmids. Furthermore, even when plasmids are identifiedwithin bacterial isolates, the reported host is typically limited to the original isolate, with no insights into alternative hosts or the plasmid's potential host range. Given that plasmids depend on hosts for their existence, investigating plasmids without the knowledge of potential hosts offersonly a partial perspective. This study introduces a method for identifying potential hosts and host ranges for plasmids through alignment with CRISPR spacers. To validate the method, we compared the PLSDB plasmids database with the CRISPR spacers database, yielding host predictions for 46% of the plasmids. When compared with reported hosts, our predictions achieved 84% concordance at the family level and 99% concordance at the phylum level. Moreover, the method frequently identifiedmultiple potential hosts for a plasmid, thereby enabling predictions of alternative hosts and the host range. Notably, we found that CRISPR spacers predominantly target plasmid backbone genes while sparing functional genes, such as those linked to antibiotic resistance, aligning with our hypothesis that CRISPR spacers are acquired from plasmid-specificregions rather than insertion elements from diverse sources. Finally, we illustrate the network of connections among differentbacterial taxa through plasmids, revealing potential pathways for horizontal gene transfer.