The pilus antigenic variation (Av) system of Neisseria gonorrhoeae is one of several high-frequency variation systems that utilize gene conversion to switch between numerous forms of an antigen on the cell surface. We have tested three predictions of the first models that explain the movement of DNA during pilin Av: (i) Av requires two recombinations at short regions of identity, (ii) circular intermediates exist that carry pilE/pilS hybrid loci and (iii) these pilE/pilS hybrid loci target the pilS sequences to a recipient pilE gene. We confirm that normal pilin Av utilizes recombination at very short regions of DNA sequence identity and that these recombination events can occur independent of homologous recombination functions. We have isolated covalently closed circular DNA molecules carrying hybrid pilin loci, but propose that an alternative hybrid molecule is the intermediate of pilin Av. Our most striking finding is that transformation of isolated pilE/pilS hybrid loci targets the pilS sequences of the hybrid to a recipient pilE at frequencies much higher than normal recombination frequencies. These results show that the different steps of a model that explains pilin Av can be separately tested to support the validity of these novel models that account for the high-frequency gene conversions that mediate pilin Av.