Elevated Levels of Interleukins, Leukocyte Protein and Cathelicidin Antimicrobial Peptide are Strongly Associated with Early to Mid-Stage of <i>Pythium insidiosum</i> Infection in Rabbit Corneas

Corneal infection in humans caused by <i>Pythium insidiosum</i> can lead to blindness and the host ocular immune response to it is less studied. Herein, we investigate the expression of mediators of innate and adaptive immune responses in a rabbit model. <i>P. insidiosum</i> zoospores were injected intracorneally in right eye of the nine <i>New Zealand White</i> rabbits while left eye was injected with 1XPBS. RT-qPCR and multiplex ELISA (mELISA) were used to study the expression of antimicrobial peptides (AMPs) and immune mediators in infected cornea on 3rd, 7th and 9th day of post-infection(PI). STRING-11.0 analysis was used to predict the interactions of immune mediators. mRNA expressions of pathogen recognition receptors (PRRs) were determined in human corneal epithelial cells (HCECs) stimulated with <i>P. insidiosum</i> zoospores. Data was analyzed using one-way ANOVA with Post-Hoc Tukey HSD test and <i>p</i>-value &lt;0.05 was considered significant. mRNA expression assay for IL-1β, IL-6, IL-8 and Cathelicidin antimicrobial peptide (CAP)-18 showed significant upregulation (<i>p</i>-value &lt; 0.05) on 7th day post-infection (PI) compared to 3rd and 9th day while Leukocyte protein (LeukoP) was elevated significantly on 3rd day followed by 7th and 9th day PI . Only IL-17A among other adaptive immune cytokines showed significant upregulation on 7th day compared to 9th day PI. Expressed in pg/mL, mELISA showed significant higher levels (<i>p</i>-value &lt; 0.05) of IL-1β, IL-8 in infected tissue in each of the time points compared to control. STRING analysis revealed co-expression of IL-1β, IL-8 and IL-6. Among PRRs, Dectin 1 and TLR4 showed significant upregulation in HCECs at 12 hrs compared to 6 hrs. In the rabbit <i>P. insidiosum</i> keratitis model, innate immune mediators: IL-1β, IL-8, IL-6, AMPs: LeukoP and CAP-18 are strongly associated in early to mid-stage of corneal infection. Dectin 1 and TLR4 were observed to be associated with recognition of <i>P. insidiosum.</i>