RelA Mutant Enterococcus faecium with Multiantibiotic Tolerance Arising in an Immunocompromised Host.

Erin S Honsa,Elaine I Tuomanen,Matthew Frank,Randall T Hayden,Vaughn S Cooper,Joshua Wolf,Amy Iverson,Jeffrey Rubnitz,Jessica Shaker,Mohammed N Mhaissen,Jason W Rosch,Richard E Lee,Charles O Rock,Michael S Gilmore

doi:10.1128/mbio.02124-16

Abstract

ABSTRACTSerious bacterial infections in immunocompromised patients require highly effective antibacterial therapy for cure, and thus, this setting may reveal novel mechanisms by which bacteria circumvent antibiotics in the absence of immune pressure. Here, an infant with leukemia developed vancomycin-resistant Enterococcus faecium (VRE) bacteremia that persisted for 26 days despite appropriate antibiotic therapy. Sequencing of 22 consecutive VRE isolates identified the emergence of a single missense mutation (L152F) in relA, which constitutively activated the stringent response, resulting in elevated baseline levels of the alarmone guanosine tetraphosphate (ppGpp). Although the mutant remained susceptible to both linezolid and daptomycin in clinical MIC testing and during planktonic growth, it demonstrated tolerance to high doses of both antibiotics when growing in a biofilm. This biofilm-specific gain in resistance was reflected in the broad shift in transcript levels caused by the mutation. Only an experimental biofilm-targeting ClpP-activating antibiotic was able to kill the mutant strain in an established biofilm. The relA mutation was associated with a fitness trade-off, forming smaller and less-well-populated biofilms on biological surfaces. We conclude that clinically relevant relA mutations can emerge during prolonged VRE infection, causing baseline activation of the stringent response, subsequent antibiotic tolerance, and delayed eradication in an immunocompromised state.

Highlights

Serious bacterial infections in immunocompromised patients require highly effective antibacterial therapy for cure, and this setting may reveal novel mechanisms by which bacteria circumvent antibiotics in the absence of immune pressure
For Enterococcus species, the stringent response pathway is well characterized and antibiotic resistance is widespread, we report the first example of mutation in the stringent response pathway causing increased baseline alarmone levels, which was responsible for antibiotic tolerance within a biofilm
After blood cultures grew vancomycinresistant Enterococcus faecium, the antibiotic regimen was changed to linezolid, which was eventually supplemented with daptomycin, gentamicin, and quinupristin-dalfopristin (Table 1)

Summary

Introduction

Serious bacterial infections in immunocompromised patients require highly effective antibacterial therapy for cure, and this setting may reveal novel mechanisms by which bacteria circumvent antibiotics in the absence of immune pressure. The mutant remained susceptible to both linezolid and daptomycin in clinical MIC testing and during planktonic growth, it demonstrated tolerance to high doses of both antibiotics when growing in a biofilm This biofilm-specific gain in resistance was reflected in the broad shift in transcript levels caused by the mutation. We conclude that clinically relevant relA mutations can emerge during prolonged VRE infection, causing baseline activation of the stringent response, subsequent antibiotic tolerance, and delayed eradication in an immunocompromised state. Using a new class of compounds that modulate ClpP activity, the biofilms were successfully eradicated These data represent the first clinical emergence of mutations in the stringent response in vancomycin-resistant entereococci. In the absence of host defenses, bacterial killing by antibiotics is required for cure This is ensured by administering drugs with the aim of ensuring that concentrations at the site of infection are above the MIC as reported by the clinical laboratory. For Enterococcus species, the stringent response pathway is well characterized and antibiotic resistance is widespread, we report the first example of mutation in the stringent response pathway causing increased baseline alarmone levels, which was responsible for antibiotic tolerance within a biofilm

Methods

Results

Conclusion