Human Conjunctival Transcriptome in Acanthamoeba Keratitis: An Exploratory Study

Abstract

Purpose: The purpose of this study was to identify conjunctival transcriptome differences in patients with Acanthamoeba keratitis compared with keratitis with no known associated pathogen.

Methods: The host conjunctival transcriptome of 9 patients with Acanthamoeba keratitis (AK) is compared with the host conjunctival transcriptome of 13 patients with pathogen-free keratitis. Culture and/or confocal confirmed Acanthamoeba in 8 of 9 participants with AK who underwent metagenomic RNA sequencing as the likely pathogen. Cultures were negative in all 13 cases where metagenomic RNA sequencing did not identify a pathogen.

Results: Transcriptome analysis identified 36 genes differently expressed between patients with AK and patients with presumed sterile, or pathogen-free, keratitis. Gene enrichment analysis revealed that some of these genes participate in several biologic pathways important for cellular signaling, ion transport and homeostasis, glucose transport, and mitochondrial metabolism. Notable relatively differentially expressed genes with potential relevance to Acanthamoeba infection included CPS1 , SLC35B4 , STEAP2 , ATP2B2 , NMNAT3 , and AKAP12 .

Conclusions: This research suggests that the local transcriptome in Acanthamoeba keratitis may be sufficiently robust to be detected in the conjunctiva and that corneas infected with Acanthamoeba may be distinguished from the inflamed cornea where no pathogen was identified. Given the low sensitivity for corneal cultures, identification of differentially expressed genes may serve as a suggestive transcriptional signature allowing for a complementary diagnostic technique to identify this blinding parasite. Knowledge of differentially expressed genes may also direct investigation of disease pathophysiology and suggest novel pathways for therapeutic targets.

Figure 1.

Heatmap of the 36 differentially expressed transcripts between patients with Acanthamoeba keratitis (AK) compared to keratitis associated with no known pathogen (pathogen-free keratitis). Normalized expression levels, arranged by unsupervised hierarchical clustering, demonstrates under expression (blue) or over expression (red) for each conjunctival sample. The closeness of the patients’ samples (x-axis) indicated similar gene expression patterns. Here, patients with Acanthamoeba keratitis algorithmically clustered separately from patients without a pathogen identified. Patient’s expression profiles do not cluster similarly based upon steroid use at time of diagnosis, contact lens uses, sex or age.