Project Summary/Abstract Acute lower respiratory tract infections, including pneumococcal pneumonia, are a leading cause of morbidity and mortality worldwide. Among children, pneumonia is the top cause of death globally and of hospitalization in the U.S. After early childhood, rates of severe pneumonia fall rapidly and do not increase again until the 4th or 5th decade of life. Very little is known about the natural immune mechanisms that provide protection against pneumonia in young to middle-aged healthy adults. Streptococcus pneumoniae, or the pneumococcus, is the leading cause of community-acquired pneumonia. Over 90 serotypes of this pathogen exist, each with a unique polysaccharide capsule. Nearly all children are colonized by or infected with pneumococcus multiple times by the age of 2. We hypothesize that these repeated exposure events result in naturally acquired heterotypic immunity in the lung that is protective against all serotypes during young adulthood but wanes with advancing age. We define heterotypic immunity as adaptive immune responses to related but not identical organisms, and believe this is the mechanism behind natural lung defenses against pneumococcus. We have developed a mouse model of pneumococcus infection-elicited heterotypic immunity against virulent pneumococcal pneumonia, which enables mechanistic studies into these lung immune defenses. This model has been used to identify a population of lung resident memory CD4+ T cells that are important for the heterotypic immune protection observed in our mice. However, these T cells are not sufficient on their own for maximum protection, and preliminary data show that a lung-confined population of B cells bearing the memory B cell marker PD-L2 are generated in mouse lung lobes that received prior pneumococcal infections. To test our central hypothesis that lung memory B cells contribute to naturally acquired heterotypic immunity against bacterial pneumonia, we will pursue 2 specific aims: (1) we will determine whether multiple pneumococcal infections in mice generate a population of resident memory B cells, a never-before demonstrated cell population in the lung; and (2) we will test whether reactivated lung memory B cells, including the observed PD-L2+ B cell population, secrete heterotypic antibodies and contribute to heterotypic lung protection against pneumococcal pneumonia in our model. Elucidating the importance of B cells in protecting the lung from heterotypic infection will be critical to future efforts to improve pneumococcal vaccine design and develop therapies based on stimulating appropriate antibacterial activities from these cells. This study would also be the first to demonstrate the importance of lung memory B cells in defense against any bacterial pathogen.