In a previous article, Wadsworth and colleagues (mBio 9:e01419-18, 2018, https://doi.org/10.1128/mBio.01419-18) described isolates that express low levels of azithromycin (Azi) resistance. Whole-genome sequencing and bioinformatic analysis suggested that the isolates had acquired DNA from commensal spp. that caused numerous nucleotide changes in the locus, which contains genes for a transcriptional repressor (MtrR) and three proteins (MtrC-MtrD-MtrE) that form a multidrug efflux pump known to export macrolides. Strong regions of linkage disequilibrium mapped to the overlapping and promoters and Genetic analyses revealed that these mosaic-like sequences increased transcription of and MtrD function, respectively. These changes also had strong epistatic effects that collectively were responsible for decreased susceptibility to MtrCDE substrates, including Azi. The report emphasizes the importance of gene exchange among neisserial species and development of antibiotic resistance in gonococci, both of which have ramifications for detection of resistance markers and efficacy of antibiotic treatment regimens for gonorrhea.