Abstract
Fungi are a large, complex group, increasingly recognized as emerging threats. Their roles as modifiers of health mandate accurate portrayals of fungal communities in humans. As an entry point into the airways and gastrointestinal tract, fungi in the mouth are relevant to several biocompartments. We have revised current practices in sequence-based taxonomy assignments and employed the improvements to address the question of the fungal genera present in the healthy human mouth. The human oral mycobiome was surveyed using massively parallel, high throughput sequencing of internal transcribed spacer 1 (ITS1) amplicons from saliva following robust extraction methods. Taxonomy was assigned by comparison to a curated reference dataset, followed by filtering with an empirically determined BLAST E-value match statistic (10−42). Nomenclature corrections further refined results by conjoining redundant names for a single fungal genus. Following these curation steps, about two-thirds of the initially identified genera were eliminated. In comparison with the one similar metagenomic study and several earlier culture-based ones, our findings change the current conception of the oral mycobiome, especially with the discovery of the high prevalence and abundance of the genus Malassezia. Previously identified as an important pathogen of the skin, and recently reported as the predominant fungal genus at the nostril and backs of the head and ear, this is the first account of Malassezia in the human mouth. Findings from this study were in good agreement with others on the existence of many consensus members of the core mycobiome, and on unique patterns for individual subjects. This research offered a cautionary note about unconditional acceptance of lengthy lists of community members produced by automated assignments, provided a roadmap for enhancing the likely biological relevance of sequence-based fungal surveys, and built the foundation for understanding the role of fungi in health and disease of the oral cavity.
Highlights
The fungi are among the most environmentally abundant and diverse eukaryotes, with estimates ranging from the more conservative approximation of 1.5 million species [1,2] to as many as 5.1 million based on high-throughput sequencing methods [3]
As a first step toward establishing the fungal baseline for this goal, we developed a robust workflow for DNA extraction, 18S internal transcribed spacer 1 (ITS1) amplification, pyrosequencing, and curation, and applied it to analyze the fungal community composition in saliva samples from six healthy individuals
Sequences were submitted to the Fungal Metagenomics Project (FMP [22]) and taxonomic assignments used in analysis
Summary
The fungi are among the most environmentally abundant and diverse eukaryotes, with estimates ranging from the more conservative approximation of 1.5 million species [1,2] to as many as 5.1 million based on high-throughput sequencing methods [3]. The emergence of fungi as a prevalent class of human pathogens has been a relatively recent occurrence, mapping temporally to the past several decades and thought to be facilitated by an increase in the number of immunocompromised persons, due to both widespread viral epidemics and medical interventions (discussed by Casadevall in [10]). Understanding their direct and indirect effects on human health requires a full characterization of fungal commensals and pathogens in both healthy and diseased populations
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