The ocular surface microbiome of mouse models

Abstract

Purpose : The ocular surface microbiome (OSM) consists of all microbes and their genetic material found on the cornea, conjunctiva and eyelid margin. Due to mechanical (eyelid motion) and chemical barriers (e.g. antimicrobial peptides in the tear fluid), the microbial abundance is low on the ocular surface. The introduction of modern sequencing technologies allows the characterization of such low abundance microbiomes. Although mouse models are essential to explore the role of the microbiome in health and disease, very little is known about the murine OSM and its connection to the local immune system.

Methods : Ocular surface swabs were collected from 18 eight-week old mice with an equal male (n=3) to female (n=3) ratio in each group. Germ free (GF) mice (n=6) were used as negative controls and compared to GF mice stably colonized with 12 defined bacteria (sDMDMm; n=6) and to specific pathogen free (SPF) mice (n=6). Host DNA depletion and DNA extraction were performed with the QIAamp DNA Microbiome Kit (Qiagen). Microbial DNA was sequenced by whole-metagenome shotgun sequencing in collaboration with the Next Generation Sequencing Platform of the University of Bern. Filtered reads were matched to the ChocoPhlAn 3 database by the Metagenomic Phylogenetic Analysis tool (MetaPhlAn 3) for taxonomic profiling and to the HMP Unified Metabolic Analysis Network (HUMAnN 3) for functional profiling. Associations between the OSM and the local immune system were established with the R package MaAsLin2, using bulk RNA sequencing data of conjunctival tissue and relative microbial abundance data of OSM swabs.

Results : The results of this study will be presented at ARVO 2023. Preliminary data showed that the relative microbial composition of the OSM of SPF mice is comparable to the human OSM at phylum level. The three predominant phyla in SPF mice were Firmicutes (60.20%, SD = 15.85), Proteobacteria (36.39%, SD = 16.76) and Actinobacteria (2.60%, SD = 2.88).

Conclusions : Applying whole-metagenome shotgun sequencing allows the analysis of eukaryotic and viral DNA in addition to bacterial DNA as well as the description of the functional profile of the microbes. The characterization of the OSM and its associations with local conjunctival immune parameters in mouse models is crucial for future studies about the role of the OSM in homeostasis and disease of the ocular surface.

This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.