Abstract
ObjectivesThe human oral microbiome is potentially related to diverse health conditions and high-throughput technology provides the possibility of surveying microbial community structure at high resolution. We compared two oral microbiome survey methods: broad-based microbiome identification by 16S rRNA gene sequencing and targeted characterization of microbes by custom DNA microarray.MethodsOral wash samples were collected from 20 individuals at Memorial Sloan-Kettering Cancer Center. 16S rRNA gene survey was performed by 454 pyrosequencing of the V3–V5 region (450 bp). Targeted identification by DNA microarray was carried out with the Human Oral Microbe Identification Microarray (HOMIM). Correlations and relative abundance were compared at phylum and genus level, between 16S rRNA sequence read ratio and HOMIM hybridization intensity.ResultsThe major phyla, Firmicutes, Proteobacteria, Bacteroidetes, Actinobacteria, and Fusobacteria were identified with high correlation by the two methods (r = 0.70∼0.86). 16S rRNA gene pyrosequencing identified 77 genera and HOMIM identified 49, with 37 genera detected by both methods; more than 98% of classified bacteria were assigned in these 37 genera. Concordance by the two assays (presence/absence) and correlations were high for common genera (Streptococcus, Veillonella, Leptotrichia, Prevotella, and Haemophilus; Correlation = 0.70–0.84).ConclusionMicrobiome community profiles assessed by 16S rRNA pyrosequencing and HOMIM were highly correlated at the phylum level and, when comparing the more commonly detected taxa, also at the genus level. Both methods are currently suitable for high-throughput epidemiologic investigations relating identified and more common oral microbial taxa to disease risk; yet, pyrosequencing may provide a broader spectrum of taxa identification, a distinct sequence-read record, and greater detection sensitivity.
Highlights
The NIH Human Microbiome Project, launched as part of the NIH Common Fund’s Roadmap for Medical Research, pointed to the need to accelerate our understanding of how our bodies and microorganisms interact to influence health and disease [1]
With 16S ribosomal RNA (rRNA) gene pyrosequencing, we recovered,79,000 sequences from the 20 oral wash samples (Table S1), with 11 bacterial phyla detected (Table 1), including Firmicutes, Proteobacteria, Bacteroidetes, Actinobacteria, and Fusobacteria as the major phyla accounting for 99.83% of the distribution
Firmicutes, Proteobacteria, Bacteroidetes, Actinobacteria, and Fusobacteria were the major phyla identified by Human Oral Microbe Identification Microarray (HOMIM), accounting for 96.5% of the distribution
Summary
The NIH Human Microbiome Project, launched as part of the NIH Common Fund’s Roadmap for Medical Research, pointed to the need to accelerate our understanding of how our bodies and microorganisms interact to influence health and disease [1]. With the characterization of microbial genetic profiles, molecular technologies can elucidate microbial community population-based studies of human microbiome, providing insight into the diversity and community structure of the human microbiome in relation to health and disease. Oral Microbe Identification Microarray (HOMIM) hybridization assay [7], two well-validated methods for microbiome profiling by assessment of microbial 16S rRNA gene diversity in human samples, with pyrosequencing selected as a broad-based approach applicable generally to the microbiome and HOMIM focused on the oral microbiome. 16S rRNA gene pyrosequencing has been applied in a wide range of human microbiome studies. We (LY and ZP) have designed and validated a 16S rRNA pyrosequencing assay for the V3–V5 region of the gene and reported that 347F/803R is the most suitable pair of primers for classification of the foregut microbiome [6]
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