Abstract The bacteria-phage arm race drives the evolution of diverse bacterial defenses. This study identifies and characterizes the defense hotspots in Acinetobacter baumannii using a reference-free approach. Among 4383 high-quality genomes, we found a total of 17,430 phage defense systems and with 54.54% concentrated in 21 hotspots. These hotspots exhibit distinct preferences for different defense systems, and co-occurrence