Molecular characterization of the ocular microbiome in healthy adults

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SUMMARY The ocular surface is constantly exposed to the environment and is inhabited by several microbes. The most commonly used methods for ocular surface microbiome characterization consist of traditional microbiological culturing and, more recently, next-generation sequencing. Culture-based methods provide information on absolute abundances but are limited in identifying only fast-growing microorganisms on culture media and a large number of nonculturable microbes. Recent 16S rRNA marker gene sequencing studies provide the most extensive and diverse characterization of the ocular microbiome. This data suggests the existence of the resident ocular microbiome; however, little is known about the core microbiome constituents of the anterior segment in a healthy population. Furthermore, the optimal method of sample collection that will yield the most representative results remains unclear. This study aims to implement whole-metagenome sequencing (WMS) and metatranscriptomics sequencing methods optimized for low-biomass samples to characterize the ocular microbiome inhabiting the anterior segment of the eye and their interaction with the immune system in healthy adults. In this project, we will establish a large human cohort (n=500) and will prospectively collect serial samples from inferior and superior conjunctival fornix from healthy adults for a period of 1 year. In addition to characterizing the core ocular microbiome in healthy adults, we will examine seasonal changes in the diversity and enrichment of specific microbial taxon (bacteria, fungi, and viruses); ocular microbiome differences between sexes and different age groups (young adults, middle age, and older adults). The metatranscriptomics data will enable us to profile active microbial pathways (bacteria, fungi, and virus) and host epithelial (conjunctival) immune response for underpinning associations between the microbiome and ocular immune modulation. The outcomes of this study will contribute towards understanding the core microbial constituents in the anterior segment and their role in immune interactions to maintain homeostasis.