IntroductionAcne vulgaris is a prevalent chronic inflammatory skin disease, and Cutibacterium overproliferation is a core pathogenic factor, yet findings remain controversial, partly due to single anatomical area sampling bias.MethodsWe conducted a crosssectional study with anatomically paired sampling across multiple facial anatomical areas in 85 subjects (30 healthy subjects, 55 acne subjects), analyzing 763 samples from 6 facial anatomical areas for 16S rRNA V4 region sequencing and microbial community analysis.ResultsAt the whole-face level, acne subjects had a significantly higher relative abundance of Cutibacterium and Staphylococcus compared with healthy subjects. Critically, Cutibacterium enrichment was highly dependent on facial anatomical areas: only the forehead and glabella showed significant intergroup differences, with the forehead showing a 127% higher median relative abundance (adjusted q = 0.043) and the glabella showing a 53% higher median relative abundance (adjusted q = 0.043), while Staphylococcus enrichment was widespread (present in 5 of 6 areas, except the jaw), and Corynebacterium showed no intergroup differences in any facial area, although LEfSe classified it as a health-associated taxon. Tax4Fun-based functional prediction suggested upregulated antimicrobial resistance and adaptation pathways, and downregulated core metabolic pathways, in acne-associated microbiota.DiscussionThis study demonstrates the facial anatomical area specificity of Cutibacterium enrichment, identifies the forehead and glabella as the areas with pronounced relative abundance differences between acne and healthy groups.