Background. Quinolone-resistant Neisseria gonorrhoeae (QRNG) arise from mutations in gyrA (intermediate resistance) or gyrA and parC (resistance). Here we tested the consequence of commonly isolated gyrA 91/95 and parC 86 mutations on gonococcal fitness.Methods. Mutant gyrA 91/95 and parC 86 alleles were introduced into wild-type gonococci or an isogenic mutant that is resistant to macrolides due to an mtrR −79 mutation. Wild-type and mutant bacteria were compared for growth in vitro and in competitive murine infection.Results. In vitro growth was reduced with increasing numbers of mutations. Interestingly, the gyrA 91/95 mutation conferred an in vivo fitness benefit to wild-type and mtrR −79 mutant gonococci. The gyrA 91/95 , parC 86 mutant, in contrast, showed a slight fitness defect in vivo, and the gyrA 91/95 , parC 86 , mtrR −79 mutant was markedly less fit relative to the parent strains. A ciprofloxacin-resistant (Cip R ) mutant was selected during infection with the gyrA 91/95 , parC 86 , mtrR −79 mutant in which the mtrR −79 mutation was repaired and the gyrA 91 mutation was altered. This in vivo-selected mutant grew as well as the wild-type strain in vitro.Conclusions. gyrA 91/95 mutations may contribute to the spread of QRNG. Further acquisition of a parC 86 mutation abrogates this fitness advantage; however, compensatory mutations can occur that restore in vivo fitness and maintain Cip R .Neisseria gonorrhoeae is a Gram-negative diplococcus that plays a major role in urogenital tract and perinatal infections [1]. Gonorrhea is the second most frequently reported bacterial sexually transmitted infection (STI) in the United States, with an estimated 700 000 new cases each year. The rate of gonorrhea is

One of the established activities of the nerve agent VX is inhibition of the enzyme acetylcholinesterase (AChE). This inhibition affects the cholinergic nervous system by decreasing the activity of the neurotransmitter-hydrolyzing enzyme cholinesterase (ChE). In an effort to gain a more comprehensive understanding of the molecular pathways affected by low-level exposure to VX, an expression profiling approach was used to identify genes with altered RNA expression patterns after exposure.Specifically, mice were exposed to 0.1, 0.2, 0.4, and 0.6 LD50 VX for a period of 2 weeks. At 2 h, 72 h, and 2 weeks after the final exposure, RNA was isolated from both the hippocampus and the cortex. Changes in gene expression levels were assessed by DNA microarray technology and grouped according to their expression patterns. Data presented here demonstrate that 2 weeks postexposure all up-regulated gene expression has returned to pre-exposure levels, including genes related to the central nervous system. Additionally, this investigation has revealed non-AChE pathway genes involved in other neuronal functions that display altered expression profiles after VX exposure.

With no vaccine available, control of gonorrhea is seriously threatened by the evolution of heightened antibiotic resistance. The fluoroquinolone class of antibiotics served as a first-line treatment option for gonorrhea for only a limited period of time due to the proliferation of resistance mutations. Resistance to fluoroquinolones in Neisseria gonorrhoeae is achieved in a two-step process in which mutations in the quinolone resistance-determining regions of the genes encoding GyrA and ParC confer intermediate and full resistance, respectively. A clinically relevant gyrA 91/95 mutation was previously demonstrated by our laboratory to confer increased in vivo fitness to N. gonorrhoeae strain FA19, leading us to hypothesize that the resistance-conferring gyrA 91/95 mutations are sufficient to alter the in vivo fitness of other N. gonorrhoeae strains. We found that the presence of this allele in two additional strains of N. gonorrhoeae (FA1090 and vi MS11) not only enhanced in vivo fitness, but did so at a level that was 10-fold higher than that reported in strain FA19. We further speculated that the enhanced in vivo fitness phenotype arose through the altered enzymatic behavior of mutant DNA gyrase, leading to a transcriptional profile better suited for bacterial survival or growth within the host.To test this hypothesis, transcriptome signatures were captured and compared between mutant and parent bacteria using total RNA sequencing as a means of identifying potential mechanisms of gyrA 91/95 -driven enhanced in vivo fitness. In vitro assays were also conducted to compare wild-type and gyrA mutant bacterial phenotypes that might explain the enhanced fitness phenotype. We showed that genome-wide expression patterns differed across three sets of mutant and parent strains. Additionally, the mutant that showed the most robust enhancement in fitness relative to its parent strain was also more resistant to human and bacterial antimicrobial peptides, including the cathelicidins and H 2 O 2 in vitro, and the fitness advantage of this mutant was not observed in cathelicidin-deficient mice. Collectively, these results suggest that a fluoroquinolone resistance-conferring mutations in GyrA condition the N. gonorrhoeae genome for increased survival or growth in vivo, and in the case of one strain background, these mutations increase resistance to host assault, allowing enhanced survivability over susceptible wild type bacteria. vii

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