Next generation sequencing (NGS) has greatly expanded our ability to analyze the human microbiomes, improving our understanding of microbial communities in health and disease. However, that came with the drawback that genome remainders of long vanished microorganisms were included in the analysis, as microbiome sequencing cannot per se differentiate between living and dead microorganisms. In environments like the skin, host defense mechanisms including antimicrobial peptides or low pH result in a high microbial turnover, likely resulting in considerable amounts of DNA from dead bacteria. NGS analyses may thus lead to inaccurate estimations of microbiome structures and consequently functional capacities. To overcome this weakness, we investigated if the approach to digest unprotected DNA by Benzonase before microbial lysis and subsequent DNA preparation (BDA) results in a more accurate assessment of the living microbiome. A skin mock community as well as skin microbiome samples were analyzed using 16S rRNA gene sequencing and metagenomics after DNA extraction with or without a Benzonase digest. The BDA method resulted in less reads from dead bacteria both in the skin mock community and skin swabs spiked with either heat inactivated bacteria or bacterial free DNA. This approach also efficiently depleted host DNA in samples with high human DNA content, with no obvious impact on the microbiome profile. In conclusion, the BDA approach enables both a better assessment of the living microbiota and effective depletion of host DNA reads.

ESDR140 - Improved representation of the living skin microbiome and efficient depletion of host reads by Benzonase pre-digest of unprotected DNA

50th Annual Meeting of the European Society for Dermatological Research. VIRTUAL MEETING