title: Comparison of microbial DNA enrichment tools for metagenomic whole genome sequencing.
abstract: Metagenomic whole genome sequencing for detection of pathogens in clinical samples is an exciting new area for discovery and clinical testing. A major barrier to this approach is the overwhelming ratio of human to pathogen DNA in samples with low pathogen abundance, which is typical of most clinical specimens. Microbial DNA enrichment methods offer the potential to relieve this limitation by improving this ratio. Two commercially available enrichment kits, the NEBNext Microbiome DNA Enrichment Kit and the Molzym MolYsis Basic kit, were tested for their ability to enrich for microbial DNA from resected arthroplasty component sonicate fluids from prosthetic joint infections or uninfected sonicate fluids spiked with Staphylococcus aureus. Using spiked uninfected sonicate fluid there was a 6-fold enrichment of bacterial DNA with the NEBNext kit and 76-fold enrichment with the MolYsis kit. Metagenomic whole genome sequencing of sonicate fluid revealed 13- to 85-fold enrichment of bacterial DNA using the NEBNext enrichment kit. The MolYsis approach achieved 481- to 9580-fold enrichment, resulting in 7 to 59% of sequencing reads being from the pathogens known to be present in the samples. These results demonstrate the usefulness of these tools when testing clinical samples with low microbial burden using next generation sequencing.
authors.affiliation: Division of Infectious Diseases, Department of Medicine, Mayo Clinic, Rochester, MN, USA.
authors.forename: Matthew
authors.initials: M
authors.lastname: Thoendel
authors.affiliation: Center for Individualized Medicine, Mayo Clinic, Rochester, MN, USA; Department of Surgery, Mayo Clinic, Rochester, MN, USA.
authors.forename: Patricio R
authors.initials: PR
authors.lastname: Jeraldo
authors.affiliation: Division of Clinical Microbiology, Department of Laboratory Medicine and Pathology, Mayo Clinic, Rochester, MN, USA.
authors.forename: Kerryl E
authors.initials: KE
authors.lastname: Greenwood-Quaintance
authors.affiliation: Center for Individualized Medicine, Mayo Clinic, Rochester, MN, USA.
authors.forename: Janet Z
authors.initials: JZ
authors.lastname: Yao
authors.affiliation: Center for Individualized Medicine, Mayo Clinic, Rochester, MN, USA; Department of Surgery, Mayo Clinic, Rochester, MN, USA.
authors.forename: Nicholas
authors.initials: N
authors.lastname: Chia
authors.affiliation: Department of Orthopedic Surgery, Mayo Clinic, Rochester, MN, USA.
authors.forename: Arlen D
authors.initials: AD
authors.lastname: Hanssen
authors.affiliation: Department of Orthopedic Surgery, Mayo Clinic, Rochester, MN, USA.
authors.forename: Matthew P
authors.initials: MP
authors.lastname: Abdel
authors.affiliation: Division of Infectious Diseases, Department of Medicine, Mayo Clinic, Rochester, MN, USA; Division of Clinical Microbiology, Department of Laboratory Medicine and Pathology, Mayo Clinic, Rochester, MN, USA. Electronic address: patel.robin@mayo.edu.
authors.forename: Robin
authors.initials: R
authors.lastname: Patel
mesh_terms: D004269:DNA, Bacterial / Q000235:genetics / Q000302:isolation & purification*; D016680:Genome, Bacterial; D059014:High-Throughput Nucleotide Sequencing; D006801:Humans; D007202:Indicators and Reagents; D054892:Metagenome; D056186:Metagenomics / Q000379:methods; D064307:Microbiota; D013203:Staphylococcal Infections / Q000382:microbiology; D013211:Staphylococcus aureus / Q000235:genetics*
chemical_list: D004269:DNA, Bacterial; D007202:Indicators and Reagents
keywords: Clinical samples; Enrichment; Metagenomics; Pathogen detection; Whole genome sequencing