Bacterial DNA on the skin surface overrepresents the viable skin microbiome

Abstract

The skin microbiome provides vital contributions to human health. However, the spatial organization and viability of its bacterial components remain unclear. Here we apply culturing, imaging, and molecular approaches to human and mouse skin samples, and find that the skin surface is colonized by fewer viable bacteria than predicted by bacterial DNA levels. Instead, viable skin-associated bacteria are predominantly located in hair follicles and other cutaneous invaginations. Furthermore, we show that the skin microbiome has a uniquely low fraction of viable bacteria as compared to other human microbiome sites, indicating that most bacterial DNA on the skin surface is not associated with viable cells Additionally, a small number of bacterial families dominate each skin site and traditional sequencing methods overestimate both the richness and diversity of the skin microbiome. Finally, we performed an in-vivo skin microbiome perturbation-recovery study using human volunteers. Bacterial 16S rRNA gene sequencing revealed that, while the skin microbiome is remarkably stable even in the wake of aggressive perturbation, repopulation of the skin surface is driven by the underlying viable population. Our findings help explain the dynamics of skin microbiome perturbation, as bacterial DNA on the skin surface can be transiently perturbed but is replenished by a stable underlying viable population. These results address multiple outstanding questions in skin microbiome biology with significant implications for future efforts to study and manipulate it. This study provides a crucial update to the skin microbiome paradigm by showing that viable bacteria of the skin microbiome are primarily localized to hair follicles and other sub-cutaneous structures rather than the skin surface. The native distribution of skin-associated bacteria has not been previously evaluated, and here we show that the skin surface has few intact bacteria while deeper structures are replete with bacterial contents. We used orthogonal approaches to evaluate the source of bacterial DNA and the dynamics of bacterial repopulation on the skin surface to develop an updated model of the skin microbiome. By adjusting the current understanding of the skin microbiome to match this model, we will be able to address outstanding questions in the field.

Highly-resolved within-species dynamics in the human facial skin microbiome

“…The predominance of complex topologies in C. acnes is consistent with our previous work indicating that each sebaceous follicle is independently colonized, creating an archipelago-like community structure which maintains coexisting genotypes by preventing direct competition ( 13 ). It is unclear if there is an anatomical basis for the coexistence of multiple transmitted S. epidermidis genotypes, given the unknown fine-scale spatial structure of S. epidermidis populations on human skin ( 27 , 31 ). C. acnes and S. epidermidis have significantly different within-lineage topologies, with C. acnes having a more comb-like structure than S. epidermidis , which could indicate that populations of S. epidermidis are more efficiently mixed (Fig.…”

“…To limit collection of surface contaminants, each subject's face was washed using a gentle cleansing cloth, rinsed with a saline wipe, and air dried. This process enabled repopulation of the surface from sebaceous secretions during the drying period (1,27).…”

“…To limit collection of surface contaminants, each subject’s face was washed using a gentle cleansing cloth, rinsed with a saline wipe, and air dried. This process enabled repopulation of the surface from sebaceous secretions during the drying period ( 1 , 27 ). From each sample, we profiled the entire community using metagenomic sequencing (mean non-human reads/sample = 4.5×10 6 ), enabling study of both species-level and within-species diversity.…”

Commensal skin bacteria exacerbate inflammation and delay skin healing

“…Intact skin is composed of two layers: the outer epidermis, composed of proliferating keratinocytes and melanocytes protected by an outermost layer of cornified dead cells; and the inner dermis, composed of hair follicles, blood vessels, nerves and immune cells. The human skin microbiome is thought to colonize both the epidermal skin surface and hair follicles within the dermis (Conwill et al 2022; Acosta et al 2023).…”