Abstract
Purpose: :
Proteases represent a class of enzymes which are related with pathogenicity and cytolysis of Acanthamoeba species and genotypes. The proteases of secreted and crude extracts of Acanthamoeba pathogenic strains were studied and compared them with proteases from non-pathogenic A. castellanii strain.
Methods: :
Clinical isolates of Acanthamoeba were obtained from corneal scraping of different patients, while non-pathogenic A. castellanii strain was obtained from American Type Culture Collection (ATCC 30011). All amoebae were grown without shaking, at room temperature, in 5 ml of Neff medium for 72 hours. After three days, the supernatant fluid was collected, filtered and dialyzed for 48 hours against Sodium Chloride buffer. Protein concentration was quantified by the method of Bradford. Protease activities were analyzed after performing electrophoresis of crude extract protein and conditioned medium in 10% polyacrylamide gels, containing gelatin as copolymerized substrate.
Results: :
Our results showed that Acanthamoeba trophozoites produce distinct extracellular proteases. Protease activities showed to be different among Acanthamoeba strains sourced from keratitis patients. Some straisn secreted very weak proteases while others revealed higher proteolitix activity.
Conclusions: :
Heterogeneous extracellular proteases are produced by different Acanthamoeba isolates and corroborated with other studies which showed that this class of enzymes could be used as a virulence marker in the differentiation of Acanthamoeba species and genotypes
Keywords: Acanthamoeba • enzymes/enzyme inhibitors • keratitis