Two Novel Point Mutations in Clinical Staphylococcus aureus Reduce Linezolid Susceptibility and Switch on the Stringent Response to Promote Persistent Infection
Male
0301 basic medicine
METHICILLIN-RESISTANT
LONG-TERM
QH301-705.5
Blotting, Western
Molecular Sequence Data
610
VANCOMYCIN RESISTANCE
UNITED-STATES
Microbial Sensitivity Tests
HEMB MUTANTS
ALARMONE (P)PPGPP
Ligases
03 medical and health sciences
616
Acetamides
Drug Resistance, Bacterial
Humans
Point Mutation
Amino Acid Sequence
RNA, Messenger
MULTIDRUG-RESISTANCE
Biology (General)
23S RIBOSOMAL-RNA
Oxazolidinones
Aged
Oligonucleotide Array Sequence Analysis
0303 health sciences
Gene Expression Profiling
Linezolid
Methyltransferases
RC581-607
ENDOTHELIAL-CELLS
Anti-Bacterial Agents
3. Good health
SMALL-COLONY VARIANTS
RNA, Ribosomal, 23S
Immunologic diseases. Allergy
Biomarkers
Research Article
DOI:
10.1371/journal.ppat.1000944
Publication Date:
2010-06-10T20:31:13Z
AUTHORS (12)
ABSTRACT
Staphylococcus aureus frequently invades the human bloodstream, leading to life threatening bacteremia and often secondary foci of infection. Failure of antibiotic therapy to eradicate infection is frequently described; in some cases associated with altered S. aureus antimicrobial resistance or the small colony variant (SCV) phenotype. Newer antimicrobials, such as linezolid, remain the last available therapy for some patients with multi-resistant S. aureus infections. Using comparative and functional genomics we investigated the molecular determinants of resistance and SCV formation in sequential S. aureus isolates from a patient who had a persistent and recurrent S. aureus infection, after failed therapy with multiple antimicrobials, including linezolid. Two point mutations in key staphylococcal genes dramatically affected clinical behaviour of the bacterium, altering virulence and antimicrobial resistance. Most strikingly, a single nucleotide substitution in relA (SACOL1689) reduced RelA hydrolase activity and caused accumulation of the intracellular signalling molecule guanosine 3', 5'-bis(diphosphate) (ppGpp) and permanent activation of the stringent response, which has not previously been reported in S. aureus. Using the clinical isolate and a defined mutant with an identical relA mutation, we demonstrate for the first time the impact of an active stringent response in S. aureus, which was associated with reduced growth, and attenuated virulence in the Galleria mellonella model. In addition, a mutation in rlmN (SACOL1230), encoding a ribosomal methyltransferase that methylates 23S rRNA at position A2503, caused a reduction in linezolid susceptibility. These results reinforce the exquisite adaptability of S. aureus and show how subtle molecular changes cause major alterations in bacterial behaviour, as well as highlighting potential weaknesses of current antibiotic treatment regimens.
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