Most metazoans engage in mutualistic interactions with their intestinal microbiota. Despite recent progress the molecular mechanisms through which microbiota exerts its beneficial influences on host physiology are still largely uncharacterized. Here we use axenic Drosophila melanogaster adults associated with a standardized microbiota composed of a defined set of commensal bacterial strains to study the impact of microbiota association on its host transcriptome. Our results demonstrate that Drosophila microbiota has a marked impact on the midgut transcriptome and promotes the expression of genes involved in host digestive functions and primary metabolism. We identify the IMD/Relish signaling pathway as a central regulator of this microbiota-mediated transcriptional response and we reveal a marked transcriptional trade-off between the midgut response to its beneficial microbiota and to bacterial pathogens. Taken together our results indicate that microbiota association potentiates host nutrition and host metabolic state, two key physiological parameters influencing host fitness. Our work paves the way to subsequent mechanistic studies to reveal how these microbiota-dependent transcriptional signatures translate into host physiological benefits. Citation: Combe BE, Defaye A, Bozonnet N, Puthier D, Royet J, et al. (2014) Drosophila Microbiota Modulates Host Metabolic Gene Expression via IMD/NF-kB Signaling. PLoS ONE 9(4): e94729.