A wide diversity of parasites infects ungulates with varying consequences on health and fitness of hosts depending on abundance and pathogenicity of parasites. Therefore, it is of great interest to deepen the identification of parasites within major taxonomic groups in order to understand how the parasite community changes with individual's traits and with the environment, to understand the consequences of parasites on host population dynamic. Among parasites, gastrointestinal nematodes are ubiquitous and a major concern for both domestic and wild ungulates. Using DNA metabarcoding of ITS-2 rDNA from larvae isolated from fecal samples, we analyzed the gastrointestinal nematodes communities (termed the “nemabiome”) of roe deer of different sex and age classes in two isolated populations. We detected 10 different genera, including parasites primarily found in cervids (e.g., Ostertagia leptospicularis, Spiculopteragia spp.) and also some species more commonly found in domestic ruminants, including pathogenic species (Haemonchus contortus, Bunostomum trigonocephalum, Cooperia punctata, Teladorsagia circumcincta) in one of the population. It probably reflects contact with livestock but the time at which contamination occurred is still to be determined. Among roe deer, young and male roe deer hosted the highest diversity of gastrointestinal nematodes. The effects of age and sex on the nemabiome were more pronounced in the population with the highest diversity of gastrointestinal nematodes and the better body condition. In addition, the effect of age and sex were dependent on gastrointestinal nematodes species, e.g. few variations of Ostertagia leptospicularis, but high variation of Trichostrongylus spp. Overall, our results suggest complex interactions between hosts, environments and parasites and confirm the efficiency of the metabarcoding approach to unravel the nemabiome of wild animals