Evaluating the Diagnostic Potential of Chorismate Mutase Poly-Clonal Peptide Antibody for the Acanthamoeba Keratitis in an Animal Model

Abstract

Acanthamoeba spp. is the causative agent of Acanthamoeba keratitis (AK), a vision-threatening parasitic disease whose primary risk factor has been attributed to poor contact lens hygiene. Unfortunately, differential diagnosis of AK is challenging as the clinical manifestations for AK are similar to those of bacterial, fungal, or even viral keratitis. Since delayed AK diagnosis can incur permanent vision impairment, a rapid and sensitive diagnostic method is urgently needed. Here, the diagnostic potential of polyclonal antibodies targeting the chorismate mutase (CM) of Acanthamoeba spp. was evaluated in AK animal models. CM antibody specificity against Acanthamoeba trophozoites and cysts was confirmed by immunocytochemistry after co-culturing Acanthamoeba with Fusarium solani, Pseudomonas aeruginosa, and Staphylococcus aureus, and human corneal epithelial (HCE) cells. Enzyme-linked immunosorbent assay (ELISA) was performed using CM-specific immune sera raised in rabbits, which demonstrated that the antibodies specifically interacted with the Acanthamoeba trophozoites and cysts in a dose-dependent manner. To evaluate the diagnostic potential of the CM antibody, AK animal models were established by incubating contact lenses with an inoculum containing A. castellanii trophozoites and subsequently overlaying these lenses onto the corneas of BALB/c mice for 7 and 21 days. The CM antibody specifically detected Acanthamoeba antigens in the murine lacrimal and eyeball tissue lysates at both time points. Our findings underscore the importance of antibody-based AK diagnosis, which could enable early and differential AK diagnosis in clinical settings.

Development of an Ex Vivo Porcine Eye Model for Exploring the Pathogenicity of Acanthamoeba

“…The amoebae would normally be removed through the blinking action of the eyes [ 27 ]. Therefore, in previous AK animal infection models, to prevent this mechanism from causing failure in establishing the infection model, the eyelids of the infected animals were surgically sutured after administering the infection [ 28 ]. However, this additional suturing to prevent model induction failure due to blinking likely induces stress and physiological pressure on the infected animals, which could further alter their immune response and affect the infection rate of the inoculated Acanthamoeba .…”

New Frontiers in Acanthamoeba Keratitis Diagnosis and Management

“…These potential targets for antibodies may aid in diagnosis in the future: inosine-uridine preferring nucleoside hydrolase (IPNH), chorismite mutase (CM), carboxylesterase, adenylyl cyclaseassociated protein (ACAP), and periplasmic binding protein (PBP) [65][66][67][68][69]. Additional studies have demonstrated the utility of CM and ACAP/PBP antibodies in AK diagnosis in mouse and rabbit animal models, respectively [69,70]. Antibody-based tests hold potential in the diagnosis of AK but still require further investigation and development prior to their widespread use.…”