PROJECT SUMMARY/ABSTRACT Enteric viruses encounter a vast array of microbes in the mammalian intestine, and microbiota influence their infection efficiency. Previous work has shown that most enteric viruses benefit from the microbiota, and microbiota depletion reduces infection with enteric viruses including poliovirus, coxsackievirus, noroviruses, and one strain of reovirus called T3SA+. However, recent studies indicate that members of the Reoviridae family are outliers in microbiota effects. For example, rotavirus infection is inhibited by bacteria, and most tested strains of reovirus have enhanced replication upon microbiota depletion. Interestingly, a pair of reovirus strains differ by a single amino acid but have opposing effects from microbiota depletion. Microbiota depletion decreases replication of strain T3SA+ but increases replication of strain T3SA-. These viruses differ by a proline-leucine polymorphism in the s1 attachment protein, which confers sialic acid binding to the T3SA+ strain but not the T3SA- strain. Recent studies indicate that these two reovirus strains also differ in intestinal cell tropism and sensitivity to host innate immune responses in mice. Thus, these isogenic viruses provide an unprecedented opportunity to define how microbiota influence viral infection. In this work we will 1) examine the specificity of reovirus-glycan interactions including interactions with microbial glycans, 2) determine how microbiota facilitate infection with reovirus strain T3SA+, and 3) elucidate mechanisms of microbiota inhibition of reovirus strain T3SA-. These studies will provide mechanistic insight into how microbiota influence enteric virus infection in the complex environment of the intestine.