Epithelial surfaces of most animals are colonized by diverse microbial communities. Although it is generally agreed that commensal bacteria can serve beneficial functions, the processes involved are poorly understood. Here we report that in the basal metazoan Hydra, ectodermal epithelial cells are covered with a multilayered glycocalyx that provides a habitat for a distinctive microbial community. Removing this epithelial microbiota results in lethal infection by the filamentous fungus Fusarium sp. Restoring the complex microbiota in gnotobiotic polyps prevents pathogen infection. Although mono-associations with distinct members of the microbiota fail to provide full protection, additive and synergistic interactions of commensal bacteria are contributing to full fungal resistance. Our results highlight the importance of resident microbiota diversity as a protective factor against pathogen infections. Besides revealing insights into the in vivo function of commensal microbes in Hydra, our findings indicate that interactions among commensal bacteria are essential to inhibit pathogen infection.

The intestinal immune system is tailored to fight pathogens effectively while tolerating the indigenous microbiota. Impairments of this homeostatic interaction may contribute to the etiology of various diseases including inflammatory bowel diseases. However, the molecular architecture underlying this complex regulatory interaction is not well understood. Here, we show that the fruit fly Drosophila melanogaster has a multilayered intestinal immune system that ensures strictly localized antimicrobial responses. Enterocytes, a major cell population of the intestine, produced antimicrobial peptides (AMPs) in a FoxO- but not NF-κB-dependent manner. Consequently, animals impaired in FoxO-mediated signaling had a significantly lowered resistance to intestinal infections; they were unable to increase the expression of AMP genes and males showed an increased bacterial load in response to an infection. Conventional innate immune signaling converging onto NF-κB activation was operative in only a few regions of the intestine, comprising the proventriculus, copper cells, and intestinal stem cells. Taken together, our results imply that danger-mediated as well as conventional innate immune signaling constitute modules that contribute to the fruit fly's intestinal immune system. We propose that this special architecture ensures localized and efficient antimicrobial responses against invasive pathogens while preserving the microbiota.

Outer membrane vesicles (OMVs) are continuously produced by Gram-negative bacteria and are increasingly recognized as ubiquitous mediators of bacterial physiology. In particular, OMVs are powerful effectors in interorganismal interactions, driven largely by their molecular contents. These impacts have been studied extensively in bacterial pathogenesis but have not been well documented within the context of mutualism. Here, we examined the proteomic composition of OMVs from the marine bacterium Vibrio fischeri, which forms a specific mutualism with the Hawaiian bobtail squid, Euprymna scolopes. We found that V. fischeri upregulates transcription of its major outer membrane protein, OmpU, during growth at an acidic pH, which V. fischeri experiences when it transitions from its environmental reservoir to host tissues. We used comparative genomics and DNA pulldown analyses to search for regulators of ompU and found that differential expression of ompU is governed by the OmpR, H-NS, and ToxR proteins. This transcriptional control combines with nutritional conditions to govern OmpU levels in OMVs. Under a host-encountered acidic pH, V. fischeri OMVs become more potent stimulators of symbiotic host development in an OmpU-dependent manner. Finally, we found that symbiotic development could be stimulated by OMVs containing a homolog of OmpU from the pathogenic species Vibrio cholerae, connecting the role of a well-described virulence factor with a mutualistic element. This work explores the symbiotic effects of OMV variation, identifies regulatory machinery shared between pathogenic and mutualistic bacteria, and provides evidence of the role that OMVs play in animal-bacterium mutualism. IMPORTANCE Beneficial bacteria communicate with their hosts through a variety of means. These communications are often carried out by a combination of molecules that stimulate responses from the host and are necessary for development of the relationship between these organisms. Naturally produced bacterial outer membrane vesicles (OMVs) contain many of those molecules and can stimulate a wide range of responses from recipient organisms. Here, we describe how a marine bacterium, Vibrio fischeri, changes the makeup of its OMVs under conditions that it experiences as it goes from its free-living lifestyle to associating with its natural host, the Hawaiian bobtail squid. This work improves our understanding of how bacteria change their signaling profile as they begin to associate with their beneficial partner animals.

Highlights d Chronic epithelial immune activation leads to structural changes in the airways d Activation of JNK signaling via TAK1 mediates this airway remodeling d FoxO acts downstream of JNK signaling in inducing this airway remodeling d NF-kB factors are of minor relevance for this response

Nutritional interventions such as caloric and dietary restriction increase lifespan in various animal models. To identify alternative and less demanding nutritional interventions that extend lifespan, we subjected fruit flies ( Drosophila melanogaster) to weekly nutritional regimens that involved alternating a conventional diet with dietary restriction. Short periods of dietary restriction (up to 2 d) followed by longer periods of a conventional diet yielded minimal increases in lifespan. We found that 3 or more days of contiguous dietary restriction (DR) was necessary to yield a lifespan extension similar to that observed with persistent DR. Female flies were more responsive to these interventions than males. Physiologic changes known to be associated with prolonged DR, such as reduced metabolic rates, showed the same time course as lifespan extension. Moreover, concurrent transcriptional changes indicative of reduced insulin signaling were identified with DR. These physiologic and transcriptional changes were sustained, as they were detectable several days after switching to conventional diets. Taken together, diets with longer periods of DR extended lifespan concurrently with physiologic and transcriptional changes that may underlie this increase in lifespan.-Romey-Glüsing, R., Li, Y., Hoffmann, J., von Frieling, J., Knop, M., Pfefferkorn, R., Bruchhaus, I., Fink, C., Roeder, T. Nutritional regimens with periodically recurring phases of dietary restriction extend lifespan in Drosophila.