Metagenome Quantitation and Assembly of Complex Microbial Samples Using High Definition Mapping
The role of the human microbiome in disease is increasingly well established. The characterization of microbe populations has historically been limited to known genetic markers of strains such as 16S rRNA genes. More recently, shotgun sequencing has been employed in an attempt to provide a more accurate measure of biodiversity. While whole metagenome sequencing is more comprehensive than single marker typing, short read lengths prevent accurate quantitation of related strains within a mixture, as well as consistent characterization of large-scale structural variation that can significantly impact pathogenicity.
To address these issues, we have applied the Nabsys HD-Mapping™ platform to the identification and quantitation of microbial strains in the context of defined complex samples. HD-Mapping employs fully electronic detection of tagged single DNA molecules, hundreds of kilobases in length, at a resolution superior to existing optical mapping approaches. On the length scales of the HD-Mapping reads, there are frequently differences, even between closely related strains of the same species. This means that individual reads tend to be much more specific to the genomes from which they derive than do NGS reads. This specificity allows for strain identification in complex mixtures with minimal bioinformatics work.
Nabsys single-molecule reads derived from a complex mixture were mapped to a database of microbial references resulting in identification of strains within the sample. Titrating the relative abundance of a single bacterial genome in a complex sample resulted in quantitation that was linear over three orders of magnitude. In addition, assembly of the mixed reads into map-level contigs allowed for the identification and structural comparison of distinct but related strains contained within the same sample.
