(503) Bacterial characteristics and metabolism of persistent MRSA infection

Background: Chronic infection with methicillin resistant Staphylococcus aureus (MRSA) is associated with worse clinical outcomes compared to MSSA infection. Risk factors among people with CF (pwCF) for developing persistent infection have been shown to include P. aeruginosa co-infection, CF diabetes and pancreatic insufficiency¹. However, bacterial characteristics that are associated with persistence have not been examined in detail. Further, MRSA infection often persists despite treatment with antibiotics that show in vitro susceptibility. We hypothesized that phenotypic and metabolic adaptation during CF airways infection contributes to persistence of MRSA.

Methods: MRSA isolates from incident, and if applicable from persistent infection were collected and examined in vitro. Phenotypic characteristics included minimal inhibitory concentrations (MIC) to commonly used antibiotics, biofilm formation, staphylotaxin, growth rates, lag time of growth, and tolerance. Tolerance a phenomenon associated with bacterial persistence despite lethal doses of antibiotics was tested for vancomycin in these isolates. Metabolism was studied by time-resolved exometabolomics in MRSA isolates cultured from sputum samples of a pwCF collected pre- and post-IV antibiotic therapy.

Results: Between Dec 2017 and Dec 2020, 78 pwCF with incident MRSA were enrolled. Median (SD) age was 14.8 (11.3) with preserved lung function (FEV₁pp 87 (21)). Within the 2-weeks prior to incident MRSA 37% of pwCF were prescribed oral antibiotics, with azithromycin and SMX-TMP being most frequent. Eleven pwCF had only one visit documented; MRSA recurred/persisted in 47 of the remaining 68 (60%) subjects. Frequency of persistence did not differ by age, FEV₁, sample type (sputum vs. OP-swab) or antibiotic use. Antibiotic susceptibility, delta-hemolysis or biofilm formation varied between isolates but no difference was seen for those who remained infected versus those who cleared the infection. Two isolates developed vancomycin resistance per MIC at 8 μg/mL yet 9 isolates showed significant growth in 20 μg/mL of vancomycin indicating tolerance. While lag time was consistent among antibiotic naive isolates, lag times became variable among isolates post vancomycin exposure (Figure). Paired pre- post in vivo antibiotic therapy isolates showed a slower growth rate in the sample collected post-antibiotic therapy. Exometabolomics showed differences in utilization / secretion of several metabolites e.g., pyruvate, acetate pre- vs. post IV antibiotic therapy that jointly indicated changes in TCA cycle activity.

Conclusions: Persistence of incident MRSA is frequent and does not differ by bacterial phenotypes commonly associated with persistence. Yet, despite low rates of resistance as determined per MIC, tolerance occurred in several isolates in early infection. In MRSA isolates collected during chronic infection changes in core energy metabolism may be related to persistence.

Acknowledgements: This work was supported by CFF (MUHLEBAC17A1) and the Lundbeck Foundation.

References: 1. Jennings JT, Dasenbrook LC, Lechtzin N, Boyle MP, MerloCA.Risk Factors for persistent methicillin-resistant Staphylococcus aureus infection in cystic fibrosis. J. Cyst. Fibrosis 2017 doi: 10.1016/j.jcf.2017.04.010.