Our understanding of the microbiome and the role of Propionibacterium acnes in skin homeostasis and acne pathogenesis is evolving. Multiple methods for sampling and identifying the skin’s microbiome exist, and understanding the differences between the abilities of various methods to characterize the microbial landscape is warranted. This study compared the microbial diversity of samples obtained from the cheeks of 20 volunteers, collected by surface swab, pore strips, and cyanoacrylate glue follicular biopsy, all sequenced with 16S rRNA sequencing (V1–V3) and whole-genome metagenomic sequencing. The sequencing method of choice influenced the detection of microbial profiles as whole-genome sequencing captured more species diversity, including viruses, compared with 16S sequencing. The relative abundance of bacterial or fungal species and overall diversity did not differ between sampling methods. However, the viral composition of the skin’s surface is unique compared with the follicle, suggesting distinct viral niches within the skin. P. acnes bacteria, ribotypes, and bacteriophages were identified equally by all sampling methods indicating that the sampling method, whether for the skin’s surface or follicle, does not impact P. acnes-related characterization and that all may be equally useful for acne-related research studies.