Treatment of Acanthamoeba keratitis (AK) is difficult because Acanthamoeba cysts are resistant to drugs and as such, successful treatment requires an effective approach that inhibits cyst formation. Histone deacetylase inhibitors (HDACi) are involved in cell proliferation, differentiation, and apoptotic cell death. In this study, the effects of HDACi such as MPK472 and KSK64 on A. castellanii trophozoites and cysts were observed. MPK472 and KSK64 showed at least 60% amoebicidal activity against Acanthamoeba trophozoite at a concentration of 10 μM upon 8 h. Neither of the two HDACi affected mature cysts, but significant amoebicidal activities (36.4% and 33.9%) were observed against encysting Acanthamoeba following treatment with 5 and 10 μM HDACi for 24 h. Light microscopy and transmission electron microscopy results confirmed that the encystation of Acanthamoeba was inhibited by the two HDACi. In addition to this, low cytopathic effects on human corneal epithelial (HCE) cells were observed following MPK472 and KSK64 treatment for 24 h. Our results indicate that the HDACi MPK472 and KSK64 could be used as new candidates for the development of an optimal therapeutic option for AK.

Pathogenic Acanthamoeba spp. cause granulomatous amoebic encephalitis and keratitis. Acanthamoeba keratitis (AK) is a rare but serious ocular infection that can result in permanent visual impairment or blindness. However, pathogenic factors of AK remain unclear and treatment for AK is arduous. Expression levels of proteins secreted into extracellular space were compared between A. castellanii pathogenic (ACP) and non-pathogenic strains. Two-dimensional polyacrylamide gel electrophoresis revealed 123 differentially expressed proteins, including 34 increased proteins, 7 qualitative increased proteins, 65 decreased proteins, and 17 qualitative decreased proteins in ACP strain. Twenty protein spots with greater than 5-fold increase in ACP strain were analyzed by liquid chromatography triple quadrupole mass spectrometry. These proteins showed similarity each to inosine-uridine preferring nucleoside hydrolase, carboxylesterase, oxygen-dependent choline dehydrogenase, periplasmic-binding protein proteinases and hypothetical proteins. These proteins expressed higher in ACP may provide some information to understand pathogenicity of Acanthamoeba.

Acanthamoeba keratitis (AK) is a rare disease but its prevalence throughout the globe continues to grow, primarily due to increased contact lens usage. Since early-stage symptoms associated with AK closely resemble those from other corneal infections, accurate diagnosis is difficult and this often results in delayed treatment and exacerbation of the disease, which can lead to permanent visual impairment. Accordingly, developing a rapid Acanthamoebaspecific diagnostic method is highly desired. In the present study, a rapid and differential method for AK diagnosis was developed using the secretory proteins derived from the pathogenic Acanthamoeba. Among the vast quantities of proteins secreted by the pathogenic Acanthamoeba, an open reading frame of the inosine-uridine preferring nucleoside hydrolase (IPNH) gene was obtained. After expressing and purifying the IPNH protein using the pGEX 4T-3 vector system, mice were immunized with the purified proteins for polyclonal antibody generation. Western blot was performed using protein lysates of the human corneal cell, non-pathogenic amoeba, pathogenic amoeba, and clinical amoeba isolate along with lysates from other causes of keratitis such as Staphylococcus aureus, Pseudomonas aeruginosa, and Fusarium solani to confirm Acanthamoeba-specificity. Western blot using the polyclonal IPNH antibody revealed that IPNH was Acanthamoeba-specific since these proteins were only observed in lysates of Acanthamoeba origin or its culture media. Our findings indicate that the IPNH antibody of Acanthamoeba may serve as a potential agent for rapid and differential AK diagnosis.

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