Gene

Biology

DNA extraction

Operating system

Computer science

DNA sequencing

Computational biology

DNA profiling

Genetics

Microbiome

Genome

Deep sequencing

Polymerase chain reaction

Host (biology)

Metagenomics

Profiling (computer programming)

Frontiers in Microbiology

The amount of host DNA, poses a major challenge to metagenome analysis. However, there is no guidance on the levels of host DNA, nor on the depth of sequencing needed to acquire meaningful information from whole metagenome sequencing (WMS). Here, we evaluated the impact of a wide range of amounts of host DNA and sequencing depths on microbiome taxonomic profiling using WMS. Synthetic samples with increasing levels of host DNA were created by spiking DNA of a mock bacterial community, with DNA from a mouse-derived cell line. Taxonomic analysis revealed that increasing proportions of host DNA led to decreased sensitivity in detecting very low and low abundant species. Reduction of sequencing depth had major impact on the sensitivity of WMS for profiling samples with 90% host DNA, increasing the number of undetected species. Finally, analysis of simulated datasets with fixed depth of 10 million reads confirmed that microbiome profiling becomes more inaccurate as the level of host DNA increases in a sample. In conclusion, samples with high amounts of host DNA coupled with reduced sequencing depths, decrease WMS coverage for characterization of the microbiome. This study highlights the importance of carefully considering these aspects in the design of WMS experiments to maximize microbiome analyses.

Impact of Host DNA and Sequencing Depth on the Taxonomic Resolution of Whole Metagenome Sequencing for Microbiome Analysis