Contribution of the gonococcal <i>NEIS1446-ispD</i> gene conversion to the pathobiology of the <i>Neisseria meningitidis</i> urethritis clade, <i>Nm</i> UC

Over the last decade, a <i>Neisseria meningitidis</i> (<i>Nm</i>) urethritis-causing clade (<i>Nm</i>UC) has emerged to cause clusters of meningococcal urethritis in the United States and globally. One genomic signature of <i>Nm</i>UC is the integration of <i>Neisseria gonorrhoeae</i> (<i>Ng</i>) DNA in an operon, <i>NEIS1446-NEIS1438</i>, which partially replaced the <i>Nm ispD</i> gene. IspD is the 2-C-methyl-d-erythritol 4-phosphate cytidylyltransferase of the terpenoid precursor synthesis pathway, required for the production of ubiquinones of the electron transfer chain. IspD is essential in several gram-negative bacteria. The biological importance of the <i>NEIS1446-ispD</i> gene conversion event for <i>Nm</i>UC was investigated. The <i>ispD</i> gene was found to be essential in <i>Nm</i>UC (CNM3) and non-clade <i>Nm</i> (MC58), and a mutation at the native locus can only be made with the insertion of a second <i>ispD</i> copy in the genome. The IspD_MC58 variant was more efficient at promoting aerobic growth at a low level than IspD_CNM3; the two proteins differ by 15 residues. Maximal aerobic growth densities of strains with an <i>Nm</i>UC background resembled <i>Ng</i> (FA19), and both were significantly lower than <i>Nm</i>. In contrast to non-clade <i>Nm</i>, all <i>Nm</i>UC strains survived well anaerobically. Increasing <i>ispD</i> expression by titrating IPTG in non-clade <i>Nm</i> enhanced anaerobic survival. Translational reporters of the <i>Nm</i>UC and <i>Ng</i> promoters demonstrated similar expression levels, and both were significantly higher than non-clade <i>Nm</i>, under aerobic and microaerobic conditions. Our findings suggest that the integration of gonococcal DNA into the <i>NEIS1446-NEIS1438</i> operon of <i>Nm</i>UC has increased <i>ispD</i> expression<i>,</i> contributing to <i>Nm</i>UC's adaptation to the oxygen-limited environment of the human urogenital tract.