Histological evaluation of lungs of infected mice pre-treated with Ly-6G antibody revealed a dramatic decrease in the magnitude and overall area of inflammatory foci compared to that seen in untreated animals (Figures 4B, S5B). per treatment condition ATP1A1 are shown. Asterisks indicate a significant difference in area between sample conditions (*?=? .001, **?=? .0001 by two-way ANOVA).(TIFF) ppat.1003679.s005.tiff (773K) GUID:?46E97037-5CBF-4490-8E5E-0C7CEEDA92DE Abstract Inhalation of causes primary pneumonic plague, a highly lethal syndrome with mortality rates approaching 100%. PHT-7.3 Pneumonic plague progression is usually biphasic, with an initial pre-inflammatory phase facilitating bacterial growth in the absence of host inflammation, followed by a pro-inflammatory phase marked by extensive neutrophil influx, an inflammatory cytokine storm, and severe tissue destruction. Using a FRET-based probe to quantitate injection of effector proteins by the type III secretion system, we show that these bacteria target alveolar PHT-7.3 macrophages early during contamination of mice, followed by a switch in host cell preference to neutrophils. We also demonstrate that neutrophil influx is unable to limit bacterial growth in the lung and is ultimately responsible for the severe inflammation during the lethal pro-inflammatory phase. Author Summary Inhalation of the bacterium results in primary pneumonic plague, a severe necrotizing pneumonia with mortality rates approaching 100% in the absence of timely antibiotic administration. Despite the notoriety of as a potential biological weapon and its well-established pandemic potential, very little is known regarding early host-pathogen interactions that lead to the progression of pulmonary contamination. harbors a type III secretion system (T3SS) for delivery of outer protein (Yop) effectors into host cells, an early and essential step in pathogenesis. In the work presented here, we identify the host cell targets of Yop secretion in the lung. We show that initially targets alveolar macrophages, followed by a shift in host cell preference to neutrophils. Through cellular depletion studies, we demonstrate that is highly resistant to macrophage- and neutrophil-mediated clearance, and that the accumulation of neutrophils in the lung is responsible for the severe necrotizing pneumonia that develops during the pro-inflammatory phase of pneumonic plague. Introduction The historical impact of on humanity cannot be understated, as three major pandemics including the Black Death of the Middle Ages have been attributed to contamination [1], [2], [3]. In the event of respiratory exposure in humans, mortality rates are nearly 100% with a time to death of typically between four and seven days [1]. Its extreme lethality and history of weaponization have led to the assignment of as a Tier 1 Select Agent and compound fears of its intentional release as a weapon of PHT-7.3 bioterrorism [1]. Using a murine intranasal contamination model, our laboratory demonstrated that the primary pneumonic plague syndrome progresses in two distinct phases [4]: an initial pre-inflammatory phase characterized by rapid bacterial replication in the lung in the absence of host innate immune responses, followed by a pro-inflammatory phase marked by extensive neutrophil influx, a massive pro-inflammatory cytokine storm, and considerable tissue destruction within the lung. In mice and humans, progression into this pro-inflammatory phase invariably results in death without immediate treatment [5], [6]. Recently, our laboratory showed that creates a unique protective environment in the lungs of mice that allows for the growth of typically avirulent organisms [7], suggesting that suppresses host innate immune responses in the lung early during contamination. The mechanism and host cell types involved in this phenomenon remain unknown. utilizes a plasmid-encoded type III secretion system (T3SS) to deliver effector proteins (Yops) directly to the cytosol of target cells. Injection of Yop effectors is essential for virulence and is known to have anti-inflammatory and anti-phagocytic effects on mammalian cells [2], [8], [9]. The pulmonary cells targeted by during primary pneumonic plague have yet to be identified. In the work presented here, we use fully virulent CO92 expressing a YopE-TEM -lactamase hybrid protein to identify the host cells targeted in the lung, and to evaluate the effect of depletion of these cell types around the progression of pneumonic plague. We show that this T3SS primarily targets macrophages and neutrophils early during the pre-inflammatory phase of disease. We also monitor the host cell dynamics in the lung in response to challenge and show that neutrophils are ultimately responsible for the severe necrotizing pneumonia during the pro-inflammatory disease phase. This work is the first identification/evaluation of.