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. 2023 Aug 1;12(8):10.
doi: 10.1167/tvst.12.8.10.

Experimental Induction of Acute Acanthamoeba castellanii Keratitis in Cats

Eric C Ledbetter  1 , Erotides Capistrano da Silva  1 , Longying Dong  2 , Sean P McDonough  2
Affiliations

Experimental Induction of Acute Acanthamoeba castellanii Keratitis in Cats

Eric C Ledbetter et al. Transl Vis Sci Technol. .

Abstract

Purpose: To develop a feline model of acute Acanthamoeba keratitis using methods that replicate natural routes of infection transmission.

Methods: Corneal Acanthamoeba castellanii inoculation was performed by three methods: topical inoculation with Acanthamoeba solution following corneal abrasion, placement of a contaminated contact lens for 7 days, and placement of a contaminated contact lens for 7 days following corneal abrasion. Sham inoculations with parasite-free medium and sterile contact lenses were also performed. Cats were monitored by ocular examination and in vivo corneal confocal microscopy for 21 days post-inoculation. Corneal samples were collected at intervals for microbiologic assessment, histopathology, and immunohistochemistry.

Results: All cats in the corneal abrasion groups developed clinical keratitis. Clinical ocular disease was inconsistently detected in cats from the contaminated contact lens only group. Initial corneal lesions were characterized by multifocal epithelial leukocyte infiltrates. Ocular lesions progressed to corneal epithelial ulceration and diffuse stromal inflammation. After 14 days, corneal ulcerations resolved, and stromal inflammation consolidated into multifocal subepithelial and stromal infiltrates. Corneal amoebae were detected by culture, in vivo confocal microscopy, histopathology, and immunohistochemistry in cats with keratitis. Neutrophilic and lymphocytic keratoconjunctivitis with lymphoplasmacytic anterior uveitis were identified by histopathology. Coinfection with aerobic bacteria was detected in some, but not all, cats with keratitis. Ocular disease was not detected in the sham inoculation groups.

Conclusions: Feline Acanthamoeba keratitis is experimentally transmissible by contaminated contact lenses and topical inoculation following corneal epithelial trauma.

Translational relevance: Experimentally induced acute Acanthamoeba keratitis in cats is clinically and histopathologically similar to its human counterpart.

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Conflict of interest statement

Disclosure: E.C. Ledbetter, (P); E. Capistrano da Silva, None; L. Dong, None; S.P. McDonough, None

Figures

Figure 1.
Figure 1.
Clinical photographs of cats with experimentally induced Acanthamoeba castellanii keratitis. (A, B) Post-inoculation day 3, corneal lesions were characterized by an irregular epithelium and multifocal epithelial leukocyte infiltrates, (C, D) Post-inoculation day 6, geographic regions of superficial corneal ulceration developed (corneas stained with fluorescein in the photographs), and epithelial and subepithelial leukocyte infiltrates had progressed. (E, F) Post-inoculation day 10, large regions of corneal epithelial ulceration (corneas stained with fluorescein in the photographs), diffuse stromal inflammation, diffuse corneal edema, peripheral stromal vascularization, and anterior uveitis were present. (G, H) Post-inoculation day 14 (G) and post-inoculation day 18 (H), the regions of corneal ulceration epithelialized, and the diffuse stromal inflammation consolidated into multifocal subepithelial and mid-stromal infiltrates associated with progressive peripheral corneal stromal vascularization. In A, C, and H, the cats were topically inoculated following light corneal abrasion. In B, D, and E, the cats were inoculated by placement of a contaminated contact lens following corneal abrasion. In F and G, the cats were inoculated by placement of a contaminated contact lens only.
Figure 2.
Figure 2.
Sequential clinical photographs of two cats with experimentally induced Acanthamoeba castellanii keratitis demonstrating progression of the keratitis over time. (A–D) A cat inoculated topically following corneal abrasion: (A) Post-inoculation day 3, ocular lesions were characterized by mucopurulent discharge, chemosis, and multifocal corneal epithelial leukocyte infiltrates (a temporary tarsorrhaphy suture is present). (B) Post-inoculation day 7, superficial corneal ulceration, progressive epithelial and subepithelial leukocyte infiltrates, and peripheral corneal vascularization developed. (C) Post-inoculation day 14, the diffuse stromal inflammation consolidated into multifocal subepithelial and mid-stromal infiltrates. (D) Post-inoculation day 21, continued consolidation and reduction of the subepithelial and mid-stromal infiltrates associated with progressive corneal vascularization were seen. (E–H) A cat inoculated by placement of a contaminated contact lens following corneal abrasion: (E) Post-inoculation day 3, ocular lesions were characterized by mucopurulent discharge and multifocal corneal epithelial leukocyte infiltrates (a temporary tarsorrhaphy suture and contact lens are present). (F) Post-inoculation day 7, superficial corneal ulceration, progressive epithelial and subepithelial leukocyte infiltrates, and central corneal edema developed. (G) Post-inoculation day 14, the diffuse stromal inflammation consolidated into multifocal subepithelial and mid-stromal infiltrates. (H) Post-inoculation day 21, continued consolidation and reduction of the subepithelial and mid-stromal infiltrates associated with progressive corneal vascularization were seen.
Figure 3.
Figure 3.
Mean keratitis clinical scores (±SDs) calculated every 2 days for cats with experimentally induced Acanthamoeba castellanii keratitis: group 1, topical inoculation with the amoeba suspension following corneal abrasion; group 2, placement of an amoeba contaminated contact lens only; group 3, placement of an amoeba-contaminated contact lens following corneal abrasion; group 4, topical inoculation of parasite-free PYG medium following light corneal abrasion; and group 5, placement of a contact lens incubated with sterile PYG medium following corneal abrasion.
Figure 4.
Figure 4.
In vivo corneal confocal photomicrographs of cats with experimentally induced Acanthamoeba castellanii keratitis. (A, B) At 7 days post-inoculation, leukocytes (arrowheads) and amoebae (arrows) were present within the corneal epithelium (A) and subepithelial stroma (B). (C, D) At 14 and 21 days post-inoculation, small clusters of leukocytes (arrowheads) and amoebae (arrows) were present in the anterior and mid-corneal stroma. In A and C, the cats were topically inoculated following light corneal abrasion. In B and D, the cats were inoculated by placement of a contaminated contact lens following corneal abrasion. Scale bars: 50 µm.
Figure 5.
Figure 5.
Photomicrographs of cats with experimentally induced Acanthamoeba castellanii keratitis. (A) Low-magnification view of neutrophilic and lymphocytic keratitis in a cat on post-inoculation day 21 (H&E stain). Scale bar: 50 µm. (B) Acanthamoeba trophozoite (arrow) in the anterior corneal stroma on post-inoculation day 7 (H&E stain; original magnification, 200×). (C) Acanthamoeba trophozoite in the anterior corneal stroma on post-inoculation day 7; amoeba were visualized using an immunoperoxidase method (appears dark brown) with hematoxylin counterstain (original magnification, 200×). (D) Acanthamoeba cyst (arrow) in the deep corneal stroma on post-inoculation day 21 (H&E stain; original magnification, 200×). In A and B, the cats were inoculated by placement of a contaminated contact lens following corneal abrasion. In C, the cat was topically inoculated following light corneal abrasion. In D, the cat was inoculated by placement of a contaminated contact lens only.
Figure 6.
Figure 6.
Phase-contrast photomicrographs of corneal amoeba cultures from cats with experimentally induced Acanthamoeba castellanii keratitis. (A, B) Acanthamoeba trophozoites and cysts are present. Scale bars: 50 µm.
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