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Levels of gonococcal resistance to antimicrobial hydrophobic agents (HAs) are controlled by the mtr (multiple transferrable resistance) system, composed of the mtrRCDE genes. The mtrR gene encodes a transcriptional repressor that appears to regulate expression of the upstream and divergent mtrCDE operon. The mtrCDE genes encode membrane proteins analogous to the MexABOprK proteins of Pseudomonas aeruginosa that mediate export of structurally diverse antimicrobial agents. In this study we found that a single base pair deletion in a 13bp inverted repeat sequence within the mtrR promoter resulted in increased resistance of gonococci to both crystal violet (CV) and erythromycin (ERY) as well as to the more lipophilic non‐ionic detergent Triton X‐100 (TX‐100). However, this cross‐resistance was contingent on the production of a full‐length lipooligosaccharide (LOS) by the recipient strain used in …