Background The cause of folliculitis decalvans (FD) remains unknown. We hypothesized that a bacterial biofilm could be involved in its pathogenesis. Objective To assess the presence or not of a bacterial biofilm in the hair roots of the scalp in FD. Patients and methods Hairs plucked from four patients and three controls were examined by field emission scanning electron microscopy (FESEM) and confocal laser scanning microscopy (CLSM). Results Bacterial communities organized as biofilms were observed both by FESEM and CLSM in the under infundibular part of hair follicles in all patients and in two of the three controls. In patients and controls, these biofilms were formed exclusively of bacilli of comparable shapes. Conclusion This pilot study provides the first evidence of the presence of bacterial biofilms in the infra infundibular part of human scalp hair follicles. These biofilms were detected both in FD patients and controls, suggesting their ubiquity as a commensal biofilm with a possible pathogenic shift in FD.