Methods for the analysis of skin microbiomes: a comparison of sampling processes and 16S rRNA hypervariable regions.

Background The skin microbiome is increasingly recognised as a critical component to the pathogenesis of many inflammatory skin disorders including atopic dermatitis. However, few studies have looked in detail at the impact of changing both sampling methodology and 16S rRNA hypervariable region sequencing in tandem for skin microbiome analysis. Methods We set out to undertake a detailed analysis comparing microbiota population diversity and composition using swab, tape, scrape and scrape then swab sampling assayed by sequencing of two 16S rRNA hypervariable regions. Comparisons were done using triplicate samples from the antecubital fossa taken from healthy volunteers. Results Alpha diversity was the greatest with tape sampling using the V1-3 region, whereas for V4, tape and swab were equivalent. Scrape sampling showed lower diversity with both V1-3 and V4. All measures of beta diversity showed the scrape methodology yielded a lower diversity. Phyla composition was similar across all sampling methodologies. Minor differences in composition were noted between V1-3 and V4 sequencing, but V1-3 was optimal for identification of firmicutes (including staphylococci). Conclusions In the methodological planning of skin microbiome analysis one needs to consider the microbes of interest to select the optimal 16S rRNA hypervariable region to sequence, and harmonisation of methodological approaches would be beneficial for the field. For detection of staphylococci on flexural skin, we recommend tape sampling with analysis of V1-3.