Avaliação da virulência de isolados dos genótipos T3, T4 e T5 de Acanthamoeba provenientes de amostras clínicas e ambientais
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Data
2018-03-23
Autores
Fernandes, Débora da Vitória de Melo
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Universidade Federal do Espírito Santo
Resumo
The genus Acanthamoeba comprises protozoa that are widely distributed in diverse environments and across all continents and that are capable of causing infections in humans, such as keratitis and granulomatous encephalitis. The pathogenesis of Acanthamoeba constitutes a multifactorial process, with factors that involve both ameba and host, but this mechanism of pathogenicity is not yet fully elucidated. There is little standardization in the methodologies applied here, with variable protocols in the literature. The objective of this work was to identify the virulence variations of six isolates of Acanthamoeba, from clinical and environmental backgrounds, with three different genotypes, T3, T4 and T5 - representing the 20 types of findings, and with two doses of amoebae on three types different from mammalian cell lines, MDCK, VERO and CHO, using cytotoxic effect and cytotoxic effect methodologies. Clinical samples are obtained from cultured corneal scrapings of patients diagnosed with amoebic keratitis, and as environmental samples are obtained from faucet outlet, flood water and dust, collected and axenized between the years 2014 and 2017. Considering that prolonged cultivation of Acanthamoeba isolates may cause a decrease or loss of virulence of the amoeba, we use the passage of the amebae in MDCK cell line to reactivate the virulence of the isolates in prolonged culture. Our data demonstrated that a cell line more susceptible to conditioned medium (cytotoxic effect) was MDCK, followed by VERO and CHO. However, for the cytopathogenicity assay, CHO was one more susceptible to exposure to Acanthamoeba trophozoites, followed VERO and MDCK. In the cytopathic effect, the results varied according to the isolate used, the dose used of trophozoites and the passage in cell line. The increase in the number of amoeba trophozoites in incubation with the cell lines was able to provoke an increase in the virulence of the isolates of the three genotypes in the cytopathic effect test, especially the VERO and CHO lines. In the cytopathic effect test, the most virulent genotype was T5, followed by T4 and T3, the most virulent isolate was A3P4 (T5) of environmental origin. Thus, it is concluded that the results obtained with the cytopathic and cytotoxic effect tests with the different mammalian cell lines have variations related to the type of mammalian cell line used, as well as the inherent characteristics of each isolate. Therefore, the results obtained here may be useful for the planning of future research related to the pathogenicity studies of Acanthamoeba.
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Clinical isolate , Environmental isolate , Cytotoxic effect , Cytopathic effect , Acanthamoeba , Isolado clínico , Isolado ambiental , Efeito citotóxico , Efeito citopático , MDCK , VERO , CHO