Identification of pathogens in culture-negative infective endocarditis cases by metagenomic analysis

Abstract

BackgroundPathogens identification is critical for the proper diagnosis and precise treatment of infective endocarditis (IE). Although blood and valve cultures are the gold standard for IE pathogens detection, many cases are culture-negative, especially in patients who had received long-term antibiotic treatment, and precise diagnosis has therefore become a major challenge in the clinic. Metagenomic sequencing can provide both information on the pathogenic strain and the antibiotic susceptibility profile of patient samples without culturing, offering a powerful method to deal with culture-negative cases.MethodsTo assess the feasibility of a metagenomic approach to detect the causative pathogens in resected valves from IE patients, we employed both next-generation sequencing and Oxford Nanopore Technologies MinION nanopore sequencing for pathogens and antimicrobial resistance detection in seven culture-negative IE patients. Using our in-house developed bioinformatics pipeline, we analyzed the sequencing results generated from both platforms for the direct identification of pathogens from the resected valves of seven clinically culture-negative IE patients according to the modified Duke criteria.ResultsOur results showed both metagenomics methods can be applied for the causative pathogen detection in all IE samples. Moreover, we were able to simultaneously characterize respective antimicrobial resistance features.ConclusionMetagenomic methods for IE detection can provide clinicians with valuable information to diagnose and treat IE patients after valve replacement surgery. However, more efforts should be made to optimize protocols for sample processing, sequencing and bioinformatics analysis.Electronic supplementary materialThe online version of this article (10.1186/s12941-018-0294-5) contains supplementary material, which is available to authorized users.

Third-Generation Sequencing in the Clinical Laboratory: Exploring the Advantages and Challenges of Nanopore Sequencing

“…Researchers were able to obtain 99% accuracy of the 16S rRNA and ITS genes using the 9.5 version flow cells, making the option of targeted sequencing feasible for clinical laboratories on the MinION. Nanopore sequencing has also been used to successfully identify pathogens in clinical cases where identification of the infectious agent can be difficult (51,52). mWGS with nanopore sequencing was carried out on seven DNA isolates from resected heart valve tissue obtained from patients diagnosed with infective endocarditis (51).…”

“…Nanopore sequencing has also been used to successfully identify pathogens in clinical cases where identification of the infectious agent can be difficult (51,52). mWGS with nanopore sequencing was carried out on seven DNA isolates from resected heart valve tissue obtained from patients diagnosed with infective endocarditis (51). Although all seven samples were determined to be culture negative by traditional microbiology testing, species identification of pathogens, which included Streptococcus, Coxiella, and Bartonella spp., was attained in all cases.…”

Actinoporins: From the Structure and Function to the Generation of Biotechnological and Therapeutic Tools

“…Oxford Nanopore Technologies (ONT) was the first to launch a DNA sequencer based on protein nanopores constructed from a mutant of the Curli production assembly/transport component (CsgG) protein from Escherichia coli [112,113]. This portable device is known as MinION and has been used in the analysis of nucleic acids to determine the small metagenomics of bacterial communities and to sequence viral genomes and genes from human cell lines [114][115][116][117]. Despite still possessing a higher error rate compared to that of other sequencing technologies, MinION is advantageous in that it can analyze data in real-time and in situ, ultimately making this type of technology ideal for use in the field and hospital diagnoses for the identification of pathogens [113,118,119].…”

Direct Metatranscriptome RNA-seq and Multiplex RT-PCR Amplicon Sequencing on Nanopore MinION – Promising Strategies for Multiplex Identification of Viable Pathogens in Food

“…Nanopore sequencing is a third-generation sequencing platform that can produce long reads on DNA and RNA molecules and perform real-time metagenomic and metatranscriptomic sequence analysis on the pocket-sized Nanopore MinION device (Garalde et al, 2018). This technology can be used to identify viral pathogens, as well as microorganisms such bacteria and fungi (Greninger et al, 2015;Juul et al, 2015;Cheng et al, 2018). A few studies have demonstrated Nanopore's potential for food safety application using metagenomic sequencing in clinical and food samples (Quick et al, 2015), however, like other genomic approaches, stable DNA molecules can cause false-positive identification, and the studies did not include a validation of whether the nanopore metagenomic sequencing data only correlates with viable pathogens.…”