Acanthamoeba castellanii as an alternative interaction model for the dermatophyte Trichophyton rubrum.

BACKGROUND: Trichophyton rubrum (Tr) is the main aetiological agent of human dermatophytosis, being isolated from the environment and keratinised tissues. In the environment, Tr can interact with other organisms, such as free-living amoebas (FLA), which can act as an alternative host system to study the interaction between microbes and phagocytic cells. OBJECTIVES: To characterise the Acanthamoeba castellanii (ALX)-Tr interaction. METHODS: Interaction was characterised in three conditions: trophozoites (PYG), late (PYG/NES) and early (NES) encystation stimulus, evaluating encystation kinetics, phagocytosis, exocytosis and fungicidal activity dynamics. RESULTS: Tr was able to induce ALX encystation and be internalised by ALX. The number of internalised conidia was high at 1 hour, and ALX presented fungicidal activity with increased intracellular ROS production and exocytosis. In PYG/NES, phagocytosis and ROS production were reduced, with decreased ALX's fungicidal activity. However, in NES there was an increased fungal engulfment, and a reduced ROS production and higher fungal burden. Furthermore, exogenous mannose decreased phagocytosis of Tr conidia, and divalent cations induced ROS production and increased ALX's fungicidal activity. Interestingly, phagocytosis was reduced in the presence of cytoskeleton inhibitor, but exocytosis was increased, suggesting that Tr conidia may have alternative pathways to escape ALX's cells. CONCLUSION: A castellanii is a proper model for studying Tr-FLA interaction, since ALX can engulf, produce ROS and kill Tr, and all these parameters are influenced by an encystation stimulus and divalent cations. Moreover, this interaction is likely to occur in the environment implicating in the adaptation to environmental stressful conditions in both organisms.

Morphological, genotypic, and physiological characterization of Acanthamoeba isolates from keratitis patients and the domestic environment in Vitoria, Espírito Santo, Brazil.

Amoebae of the genus Acanthamoeba are free-living protozoa that can cause granulomatous encephalitis and keratitis in humans. In this study, four clinical and three household dust isolates obtained in Vitória, Espírito Santo, Brazil were characterized by their morphological, genotypic, and physiological properties. All isolates belonged to group II according to Pussard and Pons' cyst morphology. Analysis of their 18S rDNA sequence identified one isolate from household dust as genotype T11 and the others six samples as genotype T4. Five T4 isolates presented a highly variable region (DF3) in 18S rDNA identical to those previously described. Physiological assays carried out with trophozoites in co-culture with bacteria or in axenic conditions showed all samples tolerated temperatures up to 37°C, regardless of culture method. One keratitis isolate grew at 42°C in co-culture with bacteria. Most isolates in co-culture survived at 1.0M, except a T11 isolate, which tolerated up to 0.5M. The isolates did not grow at 42°C and did not tolerate 0.5M and 1.0M under axenic condition. This is the first report of 18S rRNA gene genotyping applied to Acanthamoeba isolated from keratitis patients in Brazil. The results also indicated that osmo-tolerance is dependent on the culture system.