Examination of factors that affect the composition and function of the microbiota

The microbiota is an important contributor to host health and fitness, impacting all aspects of life, from development and metabolism to immunity and behavior. Substantial work has been done to characterize the factors that shape the microbiota, and have demonstrated the importance of both environmental and host factors. However, much of the multifaceted relationship between the environment, the host, and the microbiota remains to be elucidated. My dissertation attempts to demonstrate the ways in which hosts can shape their microbiota through a taxonomic and functional evaluation of microbiotas within two experimental systems./nIn chapter 1, I examine the relative contributions of the host and the environment to microbiota composition. Substantial work has been done to characterize the factors that shape the gut microbiota. Studies of the human gut microbiota have shown effects of geographical location, diet, and host genetics. However, the relative contribution of such factors in shaping the gut microbiota remains unclear, probably due to the large inter- individual variation. The nematode Caenorhabditis elegans offers a convenient model to characterize the contributions of the environment and the host to microbiota composition. While C. elegans is a well-characterized model organism, surprisingly little is known about its natural history, especially its interactions with microbes. To characterize the gut microbiota of C. elegans, I used 16S rDNA-targeted sequencing to measure the gut bacteria of worms grown in natural-like environments. By taking advantage of the availability of genetically-homogenous worm populations to reduce noise and average out inter-individual variation, and thus better discern shared features of the C. elegans microbiota, I demonstrate that the worm gut microbiota assembly is a deterministic process. My results suggest a dominant contribution of the host to microbiota composition, and further suggest a role for negative interactions between microbiota members./nIn chapter 2, I further examined the contribution of host genetics to microbiota composition by identifying differences in microbiotas assembled in worms of different genotypes spanning 200-300 million years of nematode evolution. Using 16S rDNA- targeted sequencing, I demonstrate a significant contribution of host genetics to microbiota composition. However, experimental variables affected worm microbiota composition more than the worm genotype, and hindered my identification of host-specific taxa. In an attempt to overcome this, I isolated members of the Enterobacteriaceae core microbiota family from C. elegans and C. briggsae. This, however, did not identify phylogenetic distinctions between commensals of the two species. The functional evaluation of these gut isolates did reveal host-adaptation in the form of host-specific contributions to development, infection resistance, and lifespan. My results support the role of host genetics in shaping microbiota composition, and suggest that the extent of this contribution may surpass what could be deduced based on the commonly available phylogenetic resolution./nIn chapter 3, I surveyed the roles of specific host factors in shaping the C. elegans gut microbiota by examining the gut microbiota of C. elegans mutants deficient in feeding and immune function. When selecting mutants, I considered selection during feeding and selection within the intestine as two general ways a host could shape its microbiota. Altered feeding could affect colonization by changes in food choice, food uptake, and food grinding. Immune responses can significantly affect the colonization of pathogens, and it is likely that these immunity and stress response pathways could affect the colonization of non-pathogenic bacteria. From my previous work, I amassed a collection of C. elegans gut isolates with representatives from almost all core gut microbiota families. Because my previous work did show that the environment does influence the microbiota composition to a degree, I created a synthetic community as a food source to control for environmental variation. My results suggest that changes within the intestinal environment can affect microbiota composition, and demonstrate a role of the TGF-β/DBL-1 pathway in shaping the gut colonization of Enterobacter. This specificity suggests that hosts can shape their microbiota through specific host-microbe interactions./nIn chapter 4, I utilized the tomato plant and P. syringae system to examine the ability of the phyllosphere microbiome to confer resistance to P. syringae infection of tomato leaves. In plants, the phyllosphere (above ground) microbiome is likely to be important for resistance to pathogens that infect the aerial portions of plants; however, relative to the below-ground microbiome, these communities are understudied and we do not yet know how the protective effects of this microbiome might vary as a function of bacterial density or resource availability. My results suggest that the presence of a phyllosphere microbiome does decrease pathogen colonization success, as plants sprayed with a microbiome had significantly less pathogen growth than plants sprayed with sterile buffer. I additionally show that the dose of microbiome spray has an effect on protection, and that this effect is dependent on the composition of the microbial community, but not microbial diversity. Finally, my results suggest that microbe-microbe competition (resource community dynamics) may play an important role in protection, as the addition of fertilizer abolishes the observed microbiome-mediated protection. Together, my results may alter our understanding of microbiome-mediated protection within agricultural settings and the use of plant probiotics.

Advisor: Shapira, Michael