Branched broomrape (Phelipanche ramosa (L.) Pomel) is an achlorophyllous root parasitic plant with a wide host range. Its complex management is leading to the abandonment of tobacco or oilseed rape cultivation in the most affected regions in France. Among broomrape regulation factors, soil microorganisms such as fungi seem to be a relevant biocontrol lever. The aim of this work was to detect potential mycoherbicides among fungal endophytic colonizers of P. ramosa parasitizing tobacco. Our hypothesis was that both the inhibitory of broomrape seed germination and the necrotic activities are characteristic of the fungal isolates whatever their taxonomic position. To test this hypothesis, we analysed the taxonomic and functional diversity of fungal isolates of symptomatic P. ramosa collected from infested tobacco-growing regions in France in order to identify one or more fungal strains for future biocontrol. The fungal isolates were characterized using morphological and molecular identification tools and tested for their ability to inhibit the germination of P. ramosa seeds, their necrotic activity on the stems of the pest and their non-pathogenicity to the host plant. We highlighted the specific richness of fungal colonizers associated with symptomatic P. ramosa. Among the 374 collected isolates, nearly 80% belonged to 19 Fusarium species. Eighty-seven isolates representative of this diversity also showed functional diversity by inhibiting seed germination of the parasite. The 20 best-performing isolates showed differences in germination inhibition of P. ramosa at the intraspecific level. Among these 20 fungal isolates, a set of 15 randomly selected isolates was tested for their necrotic activity on the parasite stems. Fusarium venenatum isolates showed dual competence, i.e. germination inhibition and necrotic activity, and were non-pathogenic to tobacco. This led us to discuss the potential mycoherbicidal effect of this fungal species on P. ramosa.