Concordance in the identification of pathogens by next-generation sequencing and conventional microbiology methods among neonates with culture-positive sepsis

IntroductionEarly and accurate diagnosis of sepsis through identification of the causative pathogen is vital for the appropriate treatment of infection and avoiding misuse of antibiotics. Due to low microbial load and the presence of host DNA, earlier studies have shown poor concordance between microbiology and metagenomics results in the case of blood samples. In the present study, we standardized the protocol to obtain the concordance between pathogen(s) identified through conventional microbiological culture and those obtained through 16S metagenomic analyses for the blood and blood culture samples obtained from preterm neonates. MethodologyPaired blood samples (in EDTA) and liquid blood culture samples were collected from neonates suspected of sepsis for microbial DNA extraction and conventional blood culture, respectively. Liquid blood culture samples were also used for microbial DNA extraction. DNA was extracted using Qiagen kit with additional steps of pre-extraction host DNA depletion and post extraction cleanup steps with magnetic beads. The extracted microbial DNA was further subjected to 16S rRNA sequencing using the MinION Mk1C platform. ResultsBy using modified protocol, the concentration and quality of DNA improved. Sixteen paired samples were processed. Results obtained through metagenomics and conventional microbiology showed 100% and 87.5% concordance (at genus level) in case of liquid blood culture and blood samples respectively. ConclusionOur study compared and showed high concordance between pathogen identification in paired blood and liquid blood culture samples obtained from preterm neonates with sepsis using microbiological and molecular methods.