ArticleScientific reports2020
DNA extraction of microbial DNA directly from infected tissue: an optimized protocol for use in nanopore sequencing.
Article in Scientific reports, 2020. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 18 papers.
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Who cites it
18 citing papers in PubMed, 30 citations in OpenAlex.
- Two novel real-time PCR assays forMicrobiology spectrum · 2026Article
- Development of a multiplex ddPCR assay for simultaneous absolute quantification of bacterial, fungal, and human DNA.PloS one · 2026Article
- Cancer liquid biopsies by Oxford Nanopore Technologies sequencing of cell-free DNA: from basic research to clinical applications.Molecular cancer · 2024Review
- Next Generation Sequencing in orthopaedic infections - Where is the road headed?Journal of clinical orthopaedics and trauma · 2024Article
- Rapid metagenomic sequencing for diagnosis and antimicrobial sensitivity prediction of canine bacterial infections.Microbial genomics · 2023Article
- Assembling Quality Genomes of Flax Fungal Pathogens from Oxford Nanopore Technologies Data.Journal of fungi (Basel, Switzerland) · 2023Article
- Development of an Amplicon Nanopore Sequencing Strategy for Detection of Mutations Conferring Intermediate Resistance to Vancomycin in Staphylococcus aureus Strains.Microbiology spectrum · 2023Article
- An Efficient and Easy- to- Use Method for Extraction ofIranian journal of pathology · 2023Article
- Effect of bacterial DNA enrichment on detection and quantification of bacteria in an infected tissue model by metagenomic next-generation sequencing.ISME communications · 2022Article
- Nanopore sequencing technology, bioinformatics and applications.Nature biotechnology · 2021Review
- Comprehensive Wet-Bench and Bioinformatics Workflow for Complex Microbiota Using Oxford Nanopore Technologies.mSystems · 2021Article
- A graphical, interactive and GPU-enabled workflow to process long-read sequencing data.BMC genomics · 2021Article
- Culture-Independent Genotyping, Virulence and Antimicrobial Resistance Gene Identification ofMicroorganisms · 2021Article
- High-Throughput Metagenomics for Identification of Pathogens in the Clinical Settings.Small methods · 2021Review
- Rapid Diagnostics of Orthopaedic-Implant-Associated Infections Using Nanopore Shotgun Metagenomic Sequencing on Tissue Biopsies.Microorganisms · 2021Article
- DoesVeterinary medicine international · 2021Review
- Long-Reads-Based Metagenomics in Clinical Diagnosis With a Special Focus on Fungal Infections.Frontiers in microbiology · 2021Review
- Characterization of Fecal Microbiota with Clinical Specimen Using Long-Read and Short-Read Sequencing Platform.International journal of molecular sciences · 2020Article
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Authors and funding
2 authors at 1 institution in 1 country.
Funding
No grant is acknowledged in the PubMed record.
Abstract
Identification of bacteria causing tissue infections can be comprehensive and, in the cases of non- or slow-growing bacteria, near impossible with conventional methods. Performing shotgun metagenomic sequencing on bacterial DNA extracted directly from the infected tissue may improve time to diagnosis and targeted treatment considerably. However, infected tissue consists mainly of human DNA (hDNA) which hampers bacterial identification. In this proof of concept study, we present a modified version of the Ultra-Deep Microbiome Prep kit for DNA extraction procedure, removing additional human DNA. Tissue biopsies from 3 patients with orthopedic implant-related infections containing varying degrees of Staphylococcus aureus were included. Subsequent DNA shotgun metagenomic sequencing using Oxford Nanopore Technologies' (ONT) MinION platform and ONTs EPI2ME WIMP and ARMA bioinformatic workflows for microbe and antibiotic resistance genes identification, respectively. The modified DNA extraction protocol led to an additional ~10-fold reduction of human DNA while preserving S. aureus DNA. Including the DNA sequencing and bioinformatics analyses, the presented protocol has the potential of identifying the infection-causing pathogen in infected tissue within 7 hours after biopsy. However, due to low number of S. aureus reads, positive identification of antibiotic resistance genes was not possible.
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