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Abstract

Bacterial pathogenesis requires an invading bacterium to sense and adapt to the harsh environments encountered inside the human host. This is especially true for opportunistic pathogens, such as Acinetobacter baumannii, an organism that is rapidly emerging as a serious source of hospital acquired infections, in part due to the high levels of antibiotic resistance harbored by contemporary clinical isolates. A key limitation to combatting A. baumannii is that a thorough understanding of the molecular mechanisms that underlie A. baumannii pathogenesis is lacking. ,In order to identify potential virulence determinants harbored by A. baumannii, I employed a large-scale forward genetic screen known as transposon insertion sequencing, or TnSeq, with A. baumannii strain AB5075, a contemporary isolate that exhibits both enhanced virulence and resistance to multiple classes of antibiotics. The TnSeq screen identified 300 genes that are specifically required for the survival and/or growth of A. baumannii inside Galleria mellonella larvae, an established insect infection model for studying microbial pathogenesis. Importantly, the genes identified in the screen encompassed many of the previously characterized virulence determinants of A. baumannii and, more importantly, also uncovered several novel genes that are critical for A. baumannii pathogenesis. ,My follow-up studies focus on genes involved with transcriptional regulation and reveal that several of the transcription factors required for growth inside the Galleria larvae, which are given the name ‘gig’ (for Growth in Galleria), are also required for resistance to antibiotics and environmental stress. Using a combination of genetic and biochemical approaches, I demonstrate that GigA and GigB comprise a previously undiscovered signal transduction pathway required for not only virulence in the G. mellonella model, but also for resistance to environmental stress and antibiotic exposure. Additionally, I provide evidence that GigA/GigB are required for establishing an appropriate transcriptional response to antibiotic exposure. My studies also uncover a direct link between GigA/GigB and the nitrogen phosphotransferase system (PTSNtr), a well-conserved metabolic sensing pathway. These findings suggest that GigA/GigB are master regulators of a global stress response and the coupling of the GigA/GigB signaling axis to the PTSNtr allows A. baumannii to integrate cellular metabolic status with environmental cues.

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