Background

The skin microbiome has emerged as a promising source for forensic identification. Previous 16S rRNA-based and metagenomic studies have shown that microbial community profiles can differentiate individuals. However, community-level features are inherently dynamic and strongly influenced by environmental exposure, limiting their robustness in forensic applications. In contrast, single nucleotide variation (SNV) markers within dominant skin microbiome species provide higher-resolution genetic variation and may offer more reliable individual-specific signatures.

Methods

We report preliminary results from a large-scale, multi-city cohort of 494 healthy individuals recruited from five geographically distinct cities in China. Skin microbiome samples were collected from the forehead across four seasons, and shotgun metagenomic sequencing was performed. A standardized analytical workflow was established, including taxonomic profiling, reference-based alignment, SNV calling, and stringent quality control filtering. Analyses focused on Cutibacterium acnes, a dominant and consistently detected skin commensal, ensuring sufficient sequencing depth and cross-sample comparability. SNVs were defined under unified criteria across all samples to construct a population-scale genotype dataset.

Results

Using first-season data, we identified 1,401 high-confidence SNV loci in the C. acnes genome, defined by their presence in all samples and a minor allele frequency >0.1. These loci exhibited substantial inter-individual variation. Pairwise comparisons across all 494 samples demonstrated complete discrimination, with no identical SNV profiles observed. This indicates strong individual specificity of microbial SNV patterns at the population scale. To investigate potential drivers of variation, PERMANOVA was conducted incorporating multiple metadata categories, including geographic location, climate conditions, and lifestyle factors. Each factor explained only a very small proportion of the total genetic variation (<0.2%), suggesting limited influence of commonly recorded external variables on SNV profile differentiation.

Conclusions

These results demonstrate that SNV markers derived from dominant skin microbiome species can capture highly individualized genetic variation in a large and geographically diverse cohort. The combination of complete pairwise discrimination and minimal contribution from measured external factors supports the feasibility of microbiome SNV-based individual differentiation in forensic contexts. Ongoing work will extend this framework to longitudinal data, transfer experiments, and likelihood-based evaluation models to further assess its potential for forensic evidence interpretation.

Authors

• Guanju Ma (College of Forensic Medicine, Hebei Medical University, China)
• Lihuan Zhu (College of Forensic Medicine, Hebei Medical University, China)
• Guangping Fu (College of Forensic Medicine, Hebei Medical University, China)
• Shujie Dou (College of Forensic Medicine, Hebei Medical University, China)
• Shujin Li (College of Forensic Medicine, Hebei Medical University, China)