Next-generation sequencing in neuropathological diagnosis of infections of the nervous system

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Authors: Steven L. Salzberg, Florian P. Breitwieser, Anupama Kumar, Haiping Hao, Peter Burger, Fausto J. Rodriguez, Michael Lim, Alfredo Quiñones-Hinojosa, Gary L. Gallia, Jeffrey A Tornheim, Michael T. Melia, Cynthia L. Sears and Carlos A. Pardo

Preprint posted in bioRxiv online Feb. 9, 2016

COMMENT: In this work NGS is used for differential diagnosis of nervous system pathologies whose diagnosis with conventional methods is problematic. 

The authors studied the presence of microbes in brain or spinal cord biopsies from ten patients with possible infection but for whom conventional clinical and microbiology studies yielded negative or inconclusive results.

This is a relevant work in which NGS demonstrates its usefulness in important clinical decisions for which conventional methodologies are inconclusive. These were the conclusions of this work:

Conclusions: NGS-guided metagenomic studies of brain, spinal cord or meningeal biopsies offer the possibility for dramatic improvements in our ability to detect (or rule out) a wide range of CNS pathogens, with potential benefits in speed, sensitivity, and cost. NGS-based microbiome approaches present a major new opportunity to investigate the potential role of infectious pathogens in the pathogenesis of neuroinflammatory disorders.

The program used to detect microbial reads from pathogens (eukaryotic, bacterial and viral) was Kraken system. The genomes included as reference were

human genome (version GRCh38.p2)

  • 2,817 bacterial genomes representing 891 distinct species
  • 4,383 viral genomes from 2,963 species
  • 14 genomes of eukaryotic pathogens

I wonder if to include more organisms in the reference could improve the efficacy of the methodology for diagnosis.

It is important to note that in this work the reads matching potential causative agents (bacteria or viruses) were re-aligned using BLASTN against the NCBI nucleotide database nt:

Only reads whose BLAST matches hit the same species were considered further. Reads that hit different species were considered uninformative and were excluded from further analysis. For all of the positive diagnoses reported here, the BLAST results confirmed the species identified by Kraken, although the best-matching strain was sometimes a different (draft) genome.

It provides more reliable conclusions.

In my humild opinion in shotgun taxonomic profiling it is fundamental to do a final step of checking that reads are specific of the organisms putatively detected based on the matches detected (in this case with Kraken).

Each patient case is clinically described doing the work more close and interesting for clinical audience. I think that more works like this are needed to advance in the clinical application of NGS technologies.


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Raquel Tobes