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2018 Agenda and Abstracts | < Previous Next >

2018 OMIG Abstract

Impact of PDAT on S. aureus Inflammatory Response in an EpiCorneal Model

Alejandro Arboleda1, Heather Durkee1, Mariela C. Aguilar1, Jorge Maestre-Mesa2, Guillermo Amescua1,3, Jean-Marie Parel1,3, Darlene Miller2,3

1Ophthalmic Biophysics Center,  2Ocular Microbiology Laboratory, 3Anne Bates Leach Eye Clinic, Department of Ophthalmology, Bascom Palmer Eye Institute, University of Miami Miller School of Medicine

Purpose: Methicillin-resistant Staphylococcus aureus (MRSA) keratitis is a serious, sight-threatening disease which impacts patients worldwide. One of the MRSA mechanisms of resistance includesthe use of virulence factors which protect the organism and promote host tissue destruction. The purpose of this study was to use a 3-D tissue model to help decipher the cornea’s immunological response following MRSA infection and rose bengal photodynamic antimicrobial therapy (PDAT).

Methods: Microarray analysis (Alere) was used to determine genotype and virulence profiles of two MRSA keratitis isolates. One organismwas a USA300, community-associated MRSA (CA-MRSA) and one was a USA100, healthcare-associated MRSA (HA-MRSA). A standard inoculum (105cfu/ml) was prepared for each isolate and a 100ul aliquot was applied to anEpiCorneal tissue model (MatTek).Three groups were tested per strain: 1) Control (EpiCorneal only), 2) MRSA infection (EpiCorneal+ isolate) and 3) MRSA infection-PDAT (EpiCorneal+ isolate + PDAT). For the MRSA infection-PDAT group, after MRSA inoculation, 0.1% rose bengal solution was applied to the wells and irradiated with a custom-made green LED light source. After 18 hours, the fluid from the tissue wells was collected and pro-inflammatory cytokines IL-1b, IL-6, and IL-8 were quantified using ELISA kits (Thermo-Fisher Scientific).

Results: Microarray analysis confirmed the presence of Panton-Valentine Leukotoxin and higher expression of superantigen-like toxins in the CA-MRSAcompared to the HA-MRSA isolate. Following MRSA infections, higher levels of IL-8 were documented, followed by IL-6 and IL-1b.  The cytokine response following PDAT varied between the two MRSA strains but demonstrated a 1-2 fold decrease in expression for both isolates.

Conclusion: Both CA-MRSA and HA-MRSA use virulence factors to invade and damage corneal cells. The body responds by mounting a strong inflammatory response. This study revealed that pro-inflammatory cytokines IL-8 and IL-6 increased following inoculation of MRSA in corneal tissue. However, both markers showed reduced expression following rose bengal PDAT exposure in infected models. These observed effects may be due to the bactericidal and/or toxin-neutralizing properties of rose bengal PDAT against MRSA. Further studies are required to investigate these phenomena.

Disclosure: N

Grant support:  University of Miami Scientific Awards Committee Interdisciplinary Team Science Pilot Award (UM SAC 2016-17), Edward D and Janet K Robson Fund, Florida Lions Eye Bank and Beauty of Sight Foundation, NIH Center Grant P30EY14801, Research to Prevent Blindness, Henri and Flore Lesieur Foundation (JMP)


2018 Agenda and Abstracts | < Previous Next >