Interaction between Streptococcus pneumoniae and Staphylococcus aureus Generates (·)OH Radicals That Rapidly Kill Staphylococcus aureus Strains.

Streptococcus pneumoniae rapidly kills Staphylococcus aureus by producing membrane-permeable hydrogen peroxide (H(2)O(2)). The mechanism by which S. pneumoniae-produced H(2)O(2) mediates S. aureus killing was investigated. An in vitro model that mimicked S. pneumoniae-S. aureus contact during colonization of the nasopharynx demonstrated that S. aureus killing required outcompeting densities of S. pneumoniae Compared to the wild-type strain, isogenic S. pneumoniae ΔlctO and S. pneumoniae ΔspxB, both deficient in production of H(2)O(2), required increased density to kill S. aureus While residual H(2)O(2) activity produced by single mutants was sufficient to eradicate S. aureus, an S. pneumoniae ΔspxB ΔlctO double mutant was unable to kill S. aureus A collection of 20 diverse methicillin-resistant S. aureus (MRSA) and methicillin-susceptible S. aureus (MSSA) strains showed linear sensitivity (R(2) = 0.95) for S. pneumoniae killing, but the same strains had different susceptibilities when challenged with pure H(2)O(2) (5 mM). There was no association between the S. aureus clonal complex and sensitivity to either S. pneumoniae or H(2)O(2) To kill S. aureus, S. pneumoniae produced ∼180 μM H(2)O(2) within 4 h of incubation, while the killing-defective S. pneumoniae ΔspxB and S. pneumoniae ΔspxB ΔlctO mutants produced undetectable levels. Remarkably, a sublethal dose (1 mM) of pure H(2)O(2) incubated with S. pneumoniae ΔspxB eradicated diverse S. aureus strains, suggesting that S. pneumoniae bacteria may facilitate conversion of H(2)O(2) to a hydroxyl radical ((·)OH). Accordingly, S. aureus killing was completely blocked by incubation with scavengers of (·)OH radicals, dimethyl sulfoxide (Me(2)SO), thiourea, or sodium salicylate. The (·)OH was detected in S. pneumoniae cells by spin trapping and electron paramagnetic resonance. Therefore, S. pneumoniae produces H(2)O(2), which is rapidly converted to a more potent oxidant, hydroxyl radicals, to rapidly intoxicate S. aureus strains.IMPORTANCEStreptococcus pneumoniae strains produce hydrogen peroxide (H(2)O(2)) to kill bacteria in the upper airways, including pathogenic Staphylococcus aureus strains. The targets of S. pneumoniae-produced H(2)O(2) have not been discovered, in part because of a lack of knowledge about the underlying molecular mechanism. We demonstrated that an increased density of S. pneumoniae kills S. aureus by means of H(2)O(2) produced by two enzymes, SpxB and LctO. We discovered that SpxB/LctO-produced H(2)O(2) is converted into a hydroxyl radical ((·)OH) that rapidly intoxicates and kills S. aureus We successfully inhibited the toxicity of (·)OH with three different scavengers and detected (·)OH in the supernatant. The target(s) of the hydroxyl radicals represents a new alternative for the development of antimicrobials against S. aureus infections.