Unified metagenomic method for rapid detection of bacteria, fungi and viruses in clinical samples

Abstract

Clinical metagenomic sequencing can detect microorganisms causing infection directly from clinical samples. Depletion of host DNA is key to increasing sensitivity and reducing turnaround time (TAT). Several human DNA depletion methods have been previously published for detecting microorganisms with DNA and RNA genomes in clinical samples using metagenomics techniques, however, these methodologies only allow for the detection of either DNA or RNA microbes, but not both simultaneously. Thus, we have developed a mechanical-based human DNA depletion method that allows simultaneous detection of RNA and DNA microorganisms, including viruses, bacteria and fungi, directly from clinical samples using Oxford Nanopore Technology. The method is technically easy and rapid to perform and successfully removes human DNA from the samples, decreasing human DNA detection with a media of eight Ct values. Workflow detects a broad range of organisms: RNA & DNA viruses, bacteria (Gram-negative and Gram-positive and atypical respiratory pathogens (legionella, chlamydia, mycoplasma) and fungi (Candida, Pneumocystis, Aspergillus)​ 2-hour reports have > 90% sensitivity for bacterial and viral detection compared with routine laboratory results. Positive results are first reportable after 30 min sequencing in a 7h end-to-end workflow.​ The whole genome sequence was achieved in 42% of the viruses detected.

Increasing Rate of Fatal Streptococcus pyogenes Bacteriemia—A Challenge for Prompt Diagnosis and Appropriate Therapy in Real Praxis

“…Yet, the most prevalent M types were still emm12 and emm1. The severity of GAS infection was associated with the presence of spea and spej superantigen genes [74]. In the Czech Republic, there was no change in emm types of S. pyogenes before and after the COVID-19 pandemic.…”