Since Alexander Fleming's discovery of penicillin, antibiotics have been arguably the single medical intervention that has saved more lives than any other. However, this life saving intervention is now being threatened by the problem of antibiotic resistance that is outpacing the discovery of new antibiotics, resulting in the WHO and CDC declaring antibiotic resistance as one of the greatest threats to human health. Projections include the possibility of 10 million deaths per year by 2050 with tremendous impact on the global economy in the absence of a significant shift in the current antibiotic landscape. Despite important renewed calls for investment in antibiotic discovery and an encouraging increase of activity and investment in this space, the pipeline of new antibiotics remains alarmingly sparse, particularly for agents with new mechanisms of action and that target Gram-negative pathogens in serious infection. Clearly new agents are needed; however, equally important is the need for novel strategies and antibiotic discovery platforms that can overcome these barriers and create a pipeline both now and into the future. Herein, we propose an interdisciplinary center (Center for Innovation to Transform Antibiotic Discovery; CITADel) that will take on the challenges of antibiotic discovery against the important Gram negative pathogens Pseudomonas aeruginosa, Acinetobacter baumannii, and Klebsiella pneumoniae (carbapenem-resistant Enterobactericeae in general) both with the goal of producing new antimicrobial candidates with new mechanisms of action that can progress into the clinics, and developing innovative platforms to create a sustainable pipeline of new antibiotic candidates. CITADel will consist of 4 projects and an administrative core that will focus on innovation and synergy. Novel concepts to be championed include the development of narrow spectrum, combination, or biological agents, multiplexed target-based whole cell screening using next generation sequencing, massively high-throughput combinatorial screening using droplet microfluidic technologies, the targeting of outer membrane essential proteins (to circumvent the need for small molecule intracellular accumulation), leveraging recent synthetic biology technologies to create small molecule libraries consisting of natural products whose syntheses are encoded within microbial genomes but to date are untapped, novel pathogen targeting-drug conjugates, and novel macrocyclization chemistries and protein engineering strategies, with each of these tackling many of the critical barriers that hinder antibiotic discovery efforts. Importantly, CITADel will uniquely bring together (1) the complementary expertises of investigators with experience in the development of small molecules and biological agents to advance the development novel therapeutics; (2) the innovation arising in academia with the expertise in therapeutic development in biotechnology and pharmaceutical companies; and (3) clinical infectious disease, regulatory, and commercialization expertise with basic science expertise.