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Table 1 Metagenomic approaches for pathogen detection and their findings and applications

From: Metagenomics for pathogen detection in public health

  Method Applications Recent examples Advantages Limitations
Deep amplicon sequencing • rRNA • Prokaryotic and eukaryotic identification* • Characterization of the healthy human gut microbiome (HMP) [28] • Potentially higher sensitivity • Targeted gene may not be truly universal
   • Determination of taxonomic relationships • Ancient gut microbiomes found to be more similar to modern rural than modern cosmopolitan microbiomes [29] • Less expensive as fewer reads are required for taxonomic classification • Primer bias may alter population structure
  • rpoB • Archaeal and bacterial identification* • Used to divide the species Gardnerella vaginalis into subgroups [30] • rpoB and cpn-60 offer enhanced taxonomic resolution compared to rRNA [31, 32] • Possibility of variable gene copy numbers amongst targeted species
  • cpn-60 • Determination of taxonomic relationships    
  • Viral RNA polymerase (RdRP) • Novel virus discovery • Identified novel families of picornaviruses off the coast of British Columbia [33]   
Metagenomics • Shotgun sequencing • Functional and taxonomic characterization • Detection of African swine fever virus-like sequences representing new members of the family Asfariviridae [9] • Recovery of sequences from all microorganisms • Broad specificity might decrease sensitivity
    • Detection of unexpected microbes from stool samples [12] • No a priori knowledge of microorganisms required • Library preparation is relatively labor intensive
  • Subtraction • Functional and taxonomic characterization • Identified divergent regions in non-coding RNAs in Listeria monocytogenes[34] • Random primers reduce potential for bias • Bioinformatics analysis is more challenging
    • Association of Fusobacterium nucleatum with colorectal carcinoma [35]   • Relatively expensive as more reads are required than for DAS
  • Virus concentration • Novel virus discovery • Detection of the novel H1N1 influenza from nasopharyngeal swabs [13]   • Approximately 50% of sequences generally have no significant homology to known proteins in databases (dark matter) [36]
    • Detection of a novel rhabdovirus from serum [37]   
  • Hybridization capture • Investigation of sequences with very low copy number • Metagenomic analysis of tuberculosis from a mummy [38]   • Increased granularity in population structure determination [39]
    • Investigation of Yersinia pestis from ancient teeth [40]   
  1. *Specific primers need to be made to discriminate between each group. RdRP, RNA-dependent RNA polymerase.