Microbiome Engineering for the Eradication of $\textit{Staphylococcus aureus}$ – Disentangling Bacteria-Bacteria Interactions in the Upper Respiratory Tract

Staphylococcus aureus is a major human pathogen characterised by its ability to asymptomatically colonise approximately 20-30% of the population, either transiently or permanently. This duality in behaviour classifies it as a pathobiont and is, in part, responsible for the frequency of infection, as colonisation has been linked to subsequent infections, which are especially dangerous in a hospital setting. Infections caused by this Gram-positive bacterium range from mild skin infections to life-threatening systemic diseases, such as sepsis. Treatment of these infections is complicated, mostly due to widespread antibiotic resistance, particularly methicillin-resistant S. aureus (MRSA), in addition to the multitude of strategies that the pathogen uses to evade the immune system and persist in the human body. This makes it a significant threat to public health worldwide. In addition to the need for new treatment strategies, there is a need for easy and long-lasting de-colonisation strategies which would lower infection rates. One new strategy is the use of commensal bacteria as live biotherapeutics to alter the upper respiratory tract (URT) microbiome to be resistant to S. aureus colonisation. Chapter #1 investigates the interaction dynamics of two commensals of the URT, D. pigrum and C. pseudodiphtheriticum, with the pathobiont at the macroscopic and microscopic levels. Both commensals robustly inhibited the pathobiont under different conditions. Additionally, we found that pre-conditioning the growth medium with C. pseudodiphtheriticum affected the growth of D. pigrum and S. aureus which in turn influenced the interaction kinetics of the observed inhibition between D. pigrum and S. aureus but did not alter the total observed inhibition at the final time point measurement. This supports the feasibility of using commensals as probiotics against S. aureus and lays the groundwork for future clinical studies. Chapter #2 describes the ongoing work in the lab aimed at identifying the inhibition mechanism employed by D. pigrum. Owing to the genetic intractability of the commensal, we used a transposon mutant library of S. aureus to identify possible targets of inhibition. Additionally, we screened a selection of Gram negative nosocomial pathogens for their susceptibility to inhibition by D. pigrum. Chapter #3 investigates the adaptation of the human URT commensal Corynebacterium accolens, a commensal with potential health benefits due to its antagonistic interactions with another URT pathobiont, Streptococcus pneumoniae, to its preferred habitat, the nose. Together this work investigated the interactions between bacteria of the URT microbiome and how they can be harnessed for human health. $\textit{Full-text embargoed until: 2026-11-13}$