Streptococcus pneumoniae (the pneumococcus) is a globally significant bacterial pathogen and the leading cause of pneumonia mortality worldwide. Antibiotics remain critical in the treatment of active pneumococcal disease. However, the rise in multidrug resistant (MDR) isolates compromises our ability to effectively treat infections, necessitating alternative therapeutic approaches.
Metal intoxication is a well-established host-mediated antimicrobial strategy against invading pathogens. Previous studies have shown that, in S. pneumoniae and other human pathogens, metal intoxication can result in increased susceptibility to current antibiotics. Here, we investigate the use of ionophores, a class of compound that facilitates unregulated shuttling of metal ions across the bacterial cell membrane, to rescue the efficacy of frontline antibiotics, tetracycline (TET) and azithromycin (AZI), against MDR S. pneumoniae isolates.
Using in vitro minimal inhibitory concentration (MIC) assays, we show breakage (i.e. restoration of clinically-sensitive MIC values) of AZI and TET resistance in MDR S. pneumoniae clinical isolates upon co-administration with ionophores. Furthermore, bactericidal efficacy (³3-log reduction in CFU compared to controls) was achieved within 4 hours of treatment at clinically relevant levels of antibiotic. Further analysis of these cultures revealed highly dysregulated cellular metal accumulation, with antibiotic-ionophore treatment significantly potentiating hyper-accumulation of the metal ion.
To understand the physiological impacts of this hyper-accumulation and the mechanisms mediating antibiotic re-sensitisation, scanning electron microscopy (SEM) and metabolomics were conducted. SEM revealed visible disruptions to the cell membrane and cellular aggregation under individual and combined antibiotic-ionophore treatment. Metabolomics revealed a severe decrease in acetyl CoA and glutathione abundance, suggesting perturbation to cellular energy production and redox homeostatic mechanisms in response to combined antibiotic-ionophore treatment. Disruption to these essential cellular pathways provides insight into the mechanisms that may contribute to the rapid bactericidal activity of antibiotic-ionophore treatment in vitro.
In vivo analyses using a pneumococcal disease murine model are currently underway to elucidate the utility of this therapeutic approach to overcome antibiotic resistance and aid in reducing the global pneumococcal disease burden.