Isolation of Free-living amoeba and Molecular Characterization of Acanthamoeba From Stagnant Water, Kashan, Iran

Abstract

Background and Aims: Acanthamoeba, the causative agent of granulomatous amebic encephalitis)GAE), is among the most prevalent free-living amoebas (FLA) existing in water, soil and dust. This study was conducted to determine FLA and identify Acanthamoeba genotypes isolated from stagnant water in Kashan, Iran. Materials and Methods: In this descriptive study, 138 stagnant water samples were collected from Kashan mosques and public parks. The samples were filtered (0.45µm) and cultured onto non-nutrient agar for the presence of FLA. Acanthamoeba spp. was identified by polymerase chain reaction (PCR) using specific primers, which amplified a 490 bp fragment. Among ten sequenced isolates of Acanthamoeba, different genotypes were determined by sequence analysis. The parameters such as pH, temperature, sampling season and related results were recorded and analyzed using SPSS16. Results: The rate of FLA was 88.4 %, 59.4% of which were confirmed as Acanthamoeba spp. using PCR method. The rate of Acanthamoeba T4 and T2 genotypes were 80% and 20%, respectively. There was a significant relation between FLA rate and sampling season (P= 0.01). The highest rate of FLA and Acanthamoeba was observed at pH 7. There was no significant relationship between FLA and Acanthamoeba spp. with pH and temperature. Conclusions: The rate of FLA and Acanthamoeba in stagnant water were high in Kashan. The dominant Acanthamoeba genotype (T4) is pathogen. Due to serious amoeba-induced complications, hygienic education is recommended to increase the public awareness on transmission and health/preventive measurements.