Acne-prone skin is associated with dysbiosis involving () and () causing increased seborrhea in sebaceous glands (SG) and inflammation. Human primary sebocytes were cultivated using 1.10 UFC/mL Type IA (facial acne, ATCC6919) and/or 1.10 UFC/mL (unknown origin, ATCC12228) for 48 h in our SEB4GLN-optimized media without antibiotics. Bacteria and sebocytes were enumerated and assessed to determine their viability. Lipid production was imaged and quantified via Nile Red staining. SG with hair follicles were microdissected from healthy skin and cultured using 1.10 UFC/mL Type 1A and/or 1.10 UFC/mL (wild-type facial skin strain) through prior fixation and immunostaining for MC5R, and nuclei (DAPI) via Z-stack confocal microscopy bioimaging (Leica SP5X & FIJI software, Version 2.9.0). growth was not impacted when co-cultivated with sebocytes (2D) or SG (3D) models. Phylotype IA stimulated sebocyte lipid production, which had no impact on viability. The reference strain overproliferated, inducing sebocyte mortality. For 3D SG model, culture conditions were optimized using a wild-type facial skin strain at a lower concentration, 1:10 ratio to reduced contact time, sequential inoculation and rinsing step. Bioimaging revealed strong labeling in the active areas of the pilosebaceous unit. formed biofilm, which was distributed across the SG via non-specific fluorescence imaging. We developed an innovative model of a sebaceous gland that mimics acne-prone skin with lipid overproduction and virulent phylotype IA inoculation.