To Örebro University

Purpose: To describe the corneal microbiome in infectious keratitis in relation to contact lens wear or not, and culture outcome. To explore if targeted sequencing may provide information concerning: i) culture negative episodes, ii) variables influencing culture outcome of corneal samples dispensed in transport medium.

Methods: Prospective inclusion of patients fulfilling predefined criteria of infectious keratitis. Corneal samples were dispensed in liquid Amies medium, from which both culture and targeted sequencing of the V3-V4 region of the 16S rRNA gene were carried out. Additional standard corneal culture was also performed. Main outcome measures were bacterial findings by targeted sequencing in relation to contact lens wear and culture outcome, and identification of variables influencing corneal culture outcome of indirectly inoculated corneal samples, using quantitative data from the PCR.

Results: In all, 94 episodes of infectious keratitis were included, of which 70 (74%) had bacterial growth on corneal culture. In median, 15 (range 8-30) different bacterial genera per episode of infectious keratitis were detected by targeted sequencing. The contact lens wearers (69/94; 73%) displayed significant (p=0.01) differences in the bacterial community composition of the corneal lesion compared to non-wearers, with higher abundance of Staphylococcus spp. Corynebacterium spp. and Stenotrophomonas maltophilia. Among the culture negative episodes (n=24) Brevundimonas was found to be significantly (adjusted p<0.05) enriched. Sequencing detected a potential corneal pathogen such as Clostridium, Staphylococcus, Brevundimonas, Pseudomonas and Veillonella, with a relative abundance of at least 20% in more than half of the culture negative episodes (14/24; 58%). Bacterial density in the sample had the highest impact on culture outcome (OR 6.3; p=0.009) but also age increased the odds for a positive culture outcome (OR 1.04; p=0.034), while prior antibiotic treatment significantly reduced the odds of a positive corneal culture to a fifth (OR 0.2; p=0.031).

Conclusions: Targeted sequencing can provide a potential corneal pathogen in case of a negative culture outcome in patients with infectious keratitis, as well as providing insights on the corneal microbiome of infectious keratitis and factors influencing its composition.