AbstractQuality and quantity of food items consumed has a crucial effect on phenotypes. In addition to direct effects mediated by nutrient resources, an individual’s diet can also affect the phenotype indirectly by altering its gut microbiota, a potent modulator of physiological, immunity and cognitive functions. However, most of our knowledge of diet-microbiota interactions is based on mammalian species, whereas little is still known about these effects in other vertebrates. We developed a metabarcoding procedure based on cytochrome c oxidase I high-throughput amplicon sequencing and applied it to describe diet composition in breeding colonies of an insectivorous bird, the barn swallow (Hirundo rustica). To identify putative diet-microbiota associations, we integrated the resulting diet profiles with an existing dataset for faecal microbiota in the same individual. Consistent with previous studies based on macroscopic analysis of diet composition, we found that Diptera, Hemiptera, Coleoptera and Hymenoptera were the dominant dietary components in our population. We revealed pronounced variation in diet consumed during the breeding season, along with significant differences between nearby breeding colonies. In addition, we found no difference in diet composition between adults and juveniles. Finally, our data revealed a correlation between diet and faecal microbiota composition, even after statistical control for environmental factors affecting both diet and microbiota variation. Our study suggests that variation in diet induce slight but significant microbiota changes in a non-mammalian host relying on a narrow spectrum of items consumed.