Role of Clinicogenomics in Infectious Disease Diagnostics and Public Health Microbiology

Abstract

Clinicogenomics is the exploitation of genome sequence data for diagnostic, therapeutic, and public health purposes. Central to this field is the high-throughput DNA sequencing of genomes and metagenomes. The role of clinicogenomics in infectious disease diagnostics and public health microbiology was the topic of discussion during a recent symposium (session 161) presented at the 115th general meeting of the American Society for Microbiology that was held in New Orleans, LA. What follows is a collection of the most salient and promising aspects from each presentation at the symposium. The explosion of microbiome research is driven by highthroughput DNA sequencing, so-called next-generation sequencing (NGS), technologies that allow the genomic content of entire microbial communities (bacterial, viral, and eukaryotic organisms) to be described. Although much of this work is aimed at describing the structure of "commensal" communities, the methodology works equally well to identify pathogens in clinical samples. The key concept in using NGS methodology is that detection of microbes is independent of culture and is not limited to targets used for PCR assays. Rather, it is a process of generating large-scale sequence data sets that adequately sample a specimen for microbial content and then of applying computational methods to resolve the sequences into individual species, genes, pathways, or other features.Most microbiome analyses have focused on describing bacterial content, and this is usually performed by sequencing the 16S rRNA gene. PCR primers with degenerative sequences are used to amplify all or part of the 16S rRNA gene from a broad range of species in the sample. The mix of amplicons generated from different organisms in the community is then sequenced, and the abundance of each species is determined by the number of sequences found for its respective 16S rRNA gene. Although this is useful for defining communities, it also affords the identification of pathogens with unique 16S rRNA sequences.The sensitivity and specificity of this method are determined in large part by the NGS technology. Before NGS, the full-length 16S rRNA gene was sequenced with high-quality, 700-base-long reads of Sanger, or chain termination, sequencing (sometimes referred to as "first-generation" sequencing technology). This was laborious and expensive, and deep sampling was not possible. When NGS became available, most work was done on the FLX sequencing instrument (a second-generation sequencing technology) from 454 Life Sciences (Roche Diagnostics, Indianapolis, IN, USA). This only permitted 400-base-long sequencing reads, and only a portion of the 16S rRNA gene was sequenced. The 16S rRNA gene has nine hypervariable regions that provide much of the specificity in species identification. With 454 sequencing, typically only three of these regions can be sequenced. Nevertheless, this allowed detection to the genus level of most taxa. This methodology can correctly identify pathogens in stool samples from patients with diarrhea comp...

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