Deciphering molecular mechanisms of bacterial cell death and persistence using antibiotic combinations

• Applicant: Professorin Ana Rita Gontao Brochado, Ph.D.
• Subject Area: Medical Microbiology and Mycology, Hygiene, Molecular Infection Biology
• Term: since 2019
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 417574657

Widespread antibiotic resistance demands new strategies against infectious diseases, and antibiotic combinations are an attractive alternative for overcoming the current antibacterial crisis. Yet, we do not fully understand bacterial response to antibiotics, essential for predicting emergent properties of their combinations: synergy and antagonism. This knowledge gap is particularly large for bactericidal antibiotics, which trigger complex responses that result in cell death. The focus of my project is to identify molecular mechanisms of drug interactions leading to cell death. My recent work revealed that the drug interactions largely depend on cellular responses beyond the antibiotic target. My findings convene to the concept that antibiotics elicit complex cellular responses, and more importantly, they establish drug combinations as a prime tool for elucidating such complex traits. Notably, we observed that synergy and antagonism are highly species-specific, suggesting that the mechanisms of drug interactions are mostly based on non-conserved antibiotic traits, beyond their conserved targets. Which traits determine species-specificity of the drug interactions remains a widely open question. Also unknown is whether the antibiotic actions shaping drug interactions based on growth inhibition – as in my previous work – will equally occur when antibiotics are killing. As the mechanisms of cell death by antibiotics differ from those of growth inhibition, the mechanisms of antibiotic interactions may also differ. In the future, I aim at elucidating molecular mechanisms underlying species-specificity of drug interactions, how do antibiotic interactions lead to bacterial cell death, and how do bacteria avoid antibiotic killing through tolerance? First, I propose to investigate the species-specific bactericidal synergy between aminoglycosides and β-lactams, particularly potent against the pathogen Pseudomonas aeruginosa. This will deepen our knowledge on drug action against P. aeruginosa and improve the design of narrow-spectra treatments. Second, we will establish a cross-species approach to decipher how drug combinations impact bacterial cell death and tolerance in Gram–negative pathogens, by measuring cell death upon combinatorial treatment using high-throughput approaches. This will reveal general principles of cell death-based synergy and antagonism, and how they compare to non-killing interactions. Last, we will mine the newly acquired dataset for highly bactericidal synergies (in vitro) and test their efficacy against the intracellular pathogen Salmonella Typhimurium, during infection in macrophages. We will use the S. Typhimurium genome-wide knockout library to probe which bacterial pathways underlie the treatment success. I believe the findings from my Emmy Noether project will contribute to a better understanding of bactericidal effects of drugs and their combinations, in particular those active against Gram-negative pathogens.

• DFG Programme: Independent Junior Research Groups
• Major Instrumentation: High-density Colony Replicator Robot
• Instrumentation Group: 1060 Dilutoren, Pipettiergeräte, Probennehmer