PROJECT SUMMARY Microbial interactions with the neonatal immune system influence the trajectory of an individual's health over the course of a lifetime. The long-term objective of this project is to identify maternal factors, such as microbiota, that educate the neonatal immune system and direct the balance between tolerance and defense. Recently, Lactobacillus reuteri, a maternally transferred probiotic species of bacteria, has been implicated in both exacerbating autoimmunity and raising early neonatal defense by inducing immunoglobulin A (IgA) production. Immunomodulatory characteristics of L. reuteri vary by strain, and it is unknown whether the phenomena of exacerbated autoimmunity and induction of early endogenous IgA are limited to those strains that invoke a pro-inflammatory response. This project aims to clarify the mechanism by which different strains of L. reuteri induce IgA production, focusing on the contributions of surface antigen and metabolites. Aim 1: Determine the mechanism of early endogenous IgA induction by L. reuteri. L. reuteri, a gram- positive organism, has the antigenic potential to stimulate TLR2. TLR stimulation activates B cells, leading to proliferation of antibody secreting cells. Metabolites produced by L. reuteri stimulate AhR, a transcription factor associated with enhanced B cell proliferation. A combination of TLR2 and AhR stimulation may optimize conditions for proliferation of antibody-secreting cells and therefore antibody production. Aim 2: Translate mechanistic findings with models of the human immune system. B cell physiology varies by organism. Similarly, L. reuteri strains derived from different species of hosts have unique characteristics. Therefore, human-derived strains of L. reuteri will be applied to models of the human immune system in vitro and in vivo. Aim3: Determine genesis of auto-IgA antibodies by L. reuteri. Strain-dependent induction of early endogenous IgA will be assessed for autoimmune specificity. Linking the use of L. reuteri as a probiotic on the maternal-neonatal interface to the inception of autoimmunity would warrant investigation of its safety. The proposed research will demonstrate the mechanism of early auto-IgA induction by L. reuteri as strain dependent using a translational model. Training will include specialized techniques for the development of a humanized mouse model and for the generation of mutant L. reuteri. The majority of research and training will will occur at the laboratory of Dr. Xin Luo at Virginia Tech. Bacterial mutagenesis training will take place at the laboratory of Dr. Jan-Peter van Pijkeren at University of Wisconsin-Madison.