PROJECT SUMMARY Within hospitals, infected patients are sources of bacterial transmission, often through the colonization of high- touch surfaces and equipment. Acinetobacter baumannii (Ab) is the Gram-negative bacterium with the highest rate of multidrug resistance, and is a common cause of hospital-acquired pneumonia, bloodstream, soft tissue, and urinary tract infections (UTIs). Although infected patients are sources of bacterial transmission within hospitals, no common reservoirs in the human body have been established for Ab. Hospitals often institute active surveillance strategies to mitigate the spread of bacteria between patients. However, outbreaks of Ab are still common. Moreover, there are increasing reports of community-acquired Ab infections across the globe, suggesting the existence of extra-hospital reservoirs. Two questions remain unanswered: how does the first patient get infected, and how are new Ab strains introduced into hospitals? We have recently demonstrated that Ab can hide undetected in murine bladder cells and then reactivate when stimulated by medical intervention. This lead to the hypothesis that Ab can be asymptomatically carried and introduced into hospitals by patients in intracellular reservoirs before hospitalization, and that subsequent medical interventions, such as the use of catheters, could trigger a resurgence of Ab infection from such reservoirs. Here we will employ murine models to explore this hypothesis. We recently developed catheter- and non-catheter-associated murine Ab UTI models by employing UPAB1, a recent MDR community-acquired urinary clinical isolate. In the non-catheter-associated UTI (ncUTI) model, although immunocompetent mice were not susceptible to long-term ncUTI, immunocompromised mice exhibited high bacterial burdens in urine, bladders, and kidneys, taking up to 8 weeks to completely resolve the infection. Notably, we found that introduction of a catheter into resolved mice led to the development of Ab CAUTI in ~50% of the mice in just 24 hours. Genetic characterization of the recovered bacteria confirmed that the same strain re-colonized the urinary tract. Moreover, intracellular bacteria were detected in the bladders of resolved mice up to two months after resolution of the initial infection. We named these small groups of intracellular bacteria ABIRs (Ab intracellular reservoirs). Furthermore, we also detected intracellular bacteria in alveolar macrophages during a respiratory infection. More than 300,000 patients receive mechanical ventilation in the US every year. Several complications often arise from the use of ventilators, such as ventilator-associated pneumonia (VAP). We have developed a murine model for VAP. Here we propose to investigate the relationship between ABIR formation and resurgent urinary and respiratory infections Understanding how Ab strains reside in the host and the events causing resurgence may lead, in the future, to modified surveillance strategies and/or prev...