In bacteria, the FtsK/Xer/dif (chromosome dimer resolution site) system is essential for faithful vertical genetic transmission, ensuring the resolution of chromosome dimers during their segregation to daughter cells. This system is also targeted by mobile genetic elements that integrate into chromosomal dif sites. A central question is thus how Xer/dif recombination is tuned to both act in chromosome segregation and stably maintain mobile elements. To explore this question, we focused on pathogenic Neisseria species harboring a genomic island in their dif sites. We show that the FtsK DNA translocase acts differentially at the recombination sites flanking the genomic island. It stops at one Xer/dif complex, activating recombination, but it does not stop on the other site, thus dismantling it. FtsK translocation thus permits cis discrimination between an endogenous and an imported Xer/dif recombination complex.