Heteroresistance to the model antimicrobial peptide polymyxin B in the emerging Neisseria meningitidis lineage 11.2 urethritis clade: mutations in the pilMNOPQ operon.

Clusters of Neisseria meningitidis (Nm) urethritis among primarily heterosexual males in multiple US cities have been attributed to a unique non-encapsulated meningococcal clade (the US Nm urethritis clade, US_NmUC) within the hypervirulent clonal complex 11. Resistance to antimicrobial peptides (AMPs) is a key feature of urogenital pathogenesis of the closely related species, Neisseria gonorrhoeae. The US_NmUC isolates were found to be highly resistant to the model AMP, polymyxin B (PmB, MICs 64-256 µg ml-1 ). The isolates also demonstrated stable subpopulations of heteroresistant colonies that showed near total resistant to PmB (MICs 384-1024 µg ml-1 ) and colistin (MIC 256 µg ml-1 ) as well as enhanced LL-37 resistance. This is the first observation of heteroresistance in N. meningitidis. Consistent with previous findings, overall PmB resistance in US_NmUC isolates was due to active Mtr efflux and LptA-mediated lipid A modification. However, whole genome sequencing, variant analyses and directed mutagenesis revealed that the heteroresistance phenotypes and very high-level AMP resistance were the result of point mutations and IS1655 element movement in the pilMNOPQ operon, encoding the type IV pilin biogenesis apparatus. Cross-resistance to other classes of antibiotics was also observed in the heteroresistant colonies. High-level resistance to AMPs may contribute to the pathogenesis of US_NmUC.