Moxifloxacin Activates the SOS Response in Mycobacterium tuberculosis in a Dose- and Time-Dependent Manner.

Angelo Iacobino,Giovanni Piccaro,Lanfranco Fattorini,Manuela Pardini,Federico Giannoni

doi:10.3390/microorganisms9020255

Abstract

Previous studies on Escherichia coli demonstrated that sub-minimum inhibitory concentration (MIC) of fluoroquinolones induced the SOS response, increasing drug tolerance. We characterized the transcriptional response to moxifloxacin in Mycobacterium tuberculosis. Reference strain H37Rv was treated with moxifloxacin and gene expression studied by qRT-PCR. Five SOS regulon genes, recA, lexA, dnaE2, Rv3074 and Rv3776, were induced in a dose- and time-dependent manner. A range of moxifloxacin concentrations induced recA, with a peak observed at 2 × MIC (0.25 μg/mL) after 16 h. Another seven SOS responses and three DNA repair genes were significantly induced by moxifloxacin. Induction of recA by moxifloxacin was higher in log-phase than in early- and stationary-phase cells, and absent in dormant bacilli. Furthermore, in an H37Rv fluoroquinolone-resistant mutant carrying the D94G mutation in the gyrA gene, the SOS response was induced at drug concentrations higher than the mutant MIC value. The 2 × MIC of moxifloxacin determined no significant changes in gene expression in a panel of 32 genes, except for up-regulation of the relK toxin and of Rv3290c and Rv2517c, two persistence-related genes. Overall, our data show that activation of the SOS response by moxifloxacin, a likely link to increased mutation rate and persister formation, is time, dose, physiological state and, possibly, MIC dependent.

Highlights

Mycobacterium tuberculosis (Mtb) is a major human pathogen, with 10 million new cases and 1.5 million deaths reported in 2018 [1]
We treated mid-log phase Mtb H37Rv cultures with MX concentrations ranging from 32 × minimum inhibitory concentration (MIC) (4 μg/mL) to 1/50 × MIC (0.0025 μg/mL), and measured the expression of three SOS response genes: recA, lexA and dnaE2
All three genes were up-regulated in a dose dependent manner (Figure 1A), though following a different pattern of induction, since recA and lexA peaked at 2 × MIC, whereas dnaE2 showed a maximal activation at 32 × MIC

Summary

Introduction

Mycobacterium tuberculosis (Mtb) is a major human pathogen, with 10 million new cases and 1.5 million deaths reported in 2018 [1]. The insurgence of drug resistance depends on various causes, including poor compliance with medication regimens by the TB patients, a condition that may select genotypically-resistant mutant strains [2] Such a long therapy is related to other factors including the difficulty for the drugs to penetrate inside the cellular and caseous granulomas, where mycobacteria are confined by the host immune response [3,4]. For this reason, inside the granulomas drug concentrations may not be optimal to kill Mtb, in particular since physical and biological conditions such as low oxygen concentration may cause a growth arrest termed dormancy [5,6]. Dormant Mtb is a form of resistance of the microorganism to adverse conditions enabling it to develop phenotypic resistance (drug tolerance) to several drugs [7,8], a condition termed persistence

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Conclusion