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

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]