Defining Fluoroquinolone Resistance-Mediating Mutations from Non-Resistance Polymorphisms in Mycoplasma hominis Topoisomerases

Abstract

Often dismissed as a commensal, Mycoplasma hominis is an increasingly prominent target of research due to its role in septic arthritis and organ transplant failure in immunosuppressed patients, particularly lung transplantation. As a mollicute, its highly reductive genome and structure render it refractile to most forms of treatment and growing levels of resistance to the few sources of treatment left, such as fluoroquinolones. We examined antimicrobial susceptibility (AST) to fluoroquinolones on 72 isolates and observed resistance in three (4.1%), with corresponding mutations in the quinolone resistance-determining region (QRDR) of S83L or E87G in gyrA and S81I or E85V in parC. However, there were high levels of polymorphism identified between all isolates outside of the QRDR, indicating caution for a genomics-led approach for resistance screening, particularly as we observed a further two quinolone-susceptible isolates solely containing gyrA mutation S83L. However, both isolates spontaneously developed a second spontaneous E85K parC mutation and resistance following prolonged incubation in 4 mg/L levofloxacin for an extra 24-48 h. Continued AST surveillance and investigation is required to understand how gyrA QRDR mutations predispose M. hominis to rapid spontaneous mutation and fluoroquinolone resistance, absent from other susceptible isolates. The unusually high prevalence of polymorphisms in M. hominis also warrants increased genomics' surveillance.

Keywords: Mycoplasma hominis; United Kingdom; antibiotic resistance; epidemiology; fluoroquinolone; genome analysis; genomics; topoisomerase.

Figure 1

The single nucleotide polymorphisms (SNPs) of gyrA and parC genes found in the QRDR regions of all strains analyzed. Orange lines represent functional mutations, while blue lines represent synonymous mutations. No functional mutations were found in the QRDR regions of gyrB or parE.

Figure 2

Minimum inhibitory concentration for M. hominis isolates for levofloxacin and moxifloxacin. Internationally agreed thresholds for resistance are shown as dotted lines and isolates above the line are resistant to the respective fluoroquinolones. Isolates with QRDR gyrA-only mutations are shown (S019M as red circles and DF28 as blue circles), and MICs for these isolates following one step induction of resistance, resulting in an additional QRDR parC mutation, are shown as colored squares (S019M2R as red squares and DF282R as blue squares).

Figure 3

A phylogenetic tree of 72 isolates analyzed (and two induced resistant strains linked by arrows), constructed via concatenations of all four topoisomerase genes (gyrA, gyrB, parC and parE). The outgroup used was Mycoplasma pneumoniae M129, listed on the tree under its accession number (NC000912). Naturally occurring resistant strains are identified by arrowheads.