Isolation and Molecular Characterization of Acanthamoeba Strains from Dental Units in Costa Rica

Abstract

Free-living amoebae are protozoa widely distributed in nature, which can be found in a variety of environments. Four genera are recognized as causal agents of infections in humans and animals: Acanthamoeba, Naegleria, Balamuthia, and Sappinia. In this study, the presence of Acanthamoeba in dental units was determined and the isolates obtained were molecularly characterized; osmotolerance and thermotolerance assays were also performed to evaluate multiplication under these conditions, frequently associated with pathogenicity. The morphological analysis and partial sequencing of the 18S rDNA gene revealed the presence of Acanthamoeba genotype T4 in 14% of the units sampled. Osmotolerance and thermotolerance tests were positive for more than 80% of the isolates. Up to date, this is the first study that reports the detection, identification, and genotyping of Acanthamoeba isolated from dental units in Costa Rica and even in Latin-America. Further assays to determine the potential pathogenicity of these Acanthamoeba isolates are underway.

Isolation of Acanthamoeba T5 from Water: Characterization of Its Pathogenic Potential, Including the Production of Extracellular Vesicles

“…A monolayer of MDCK without amoebae was employed as the negative control. Nonpathogenic Acanthamoeba genotype T4 (GenBank accession number: KP677460), isolated, cultured and physiologically characterized in our laboratory was also employed in this assay as another negative control [19]. Each assay was performed in triplicate.…”

“…The wide distribution of Acanthamoeba, as well as the characteristic of being amphizoic and the severity of the disease that it causes, has motivated researchers to focus not only in the isolation of these organisms, but in demonstrating the presence of direct and indirect factors that could indicate a possible pathogenic potential [15][16][17][18][19]. Most of the literature refers to osmotolerance and thermotolerance, the production of proteases and cytopathic effects in vitro as the main factors related to pathogenicity.…”

Potentially pathogenic Acanthamoeba genotype T4 isolated from dental units and emergency combination showers

“…Acanthamoeba is the genus of FLA most frequently found in nature; nevertheless, not every species or genotype is able to produce damage to a host ( Schuster & Visvesvara 2004 ). In previous studies performed in our laboratory, we isolated and axenically grew eight Acanthamoeba genotype T4 from dental units and combined emergency units ( Retana-Moreira et al 2014 , 2015 ). Considering the location from which they were isolated and, since this genotype is the most frequently associated with clinical cases ( Khan 2006 ), we considered it relevant to physiologically and biologically characterise the isolates through a series of laboratory assays to determine characteristics associated with pathogenic potential.…”

“…Samples - A total of eight axenic isolates of Acanthamoeba genotype T4 were employed in this study. Samples were obtained from dental units (DU3, DU4, DU5, DU6, DU8, DU12) ( Retana-Moreira et al 2015 ) and emergency combination showers (CSU7 and CSUT7) ( Retana-Moreira et al 2014 ). Each axenic isolate was grown on 25 cm 2 tissue culture flasks (Cellstar, Greiner Bio-One GmbH, Frickenhausen, Germany) with 5 mL of PYG medium (0.75% proteose peptone, 0.75% yeast extract and 1.5% glucose) containing 10 μg/mL gentamicin (Sigma Aldrich Co., St. Louis, USA).…”

“…To date, several studies report Acanthamoeba in soil (especially genotype T4), swimming pools, water networks from hospitals, and air conditioning units ( Thomas et al 2006 , Chan et al 2011 , Fabres et al 2016 , Reyes-Batlle et al 2016 ), as well as equipment and devices for medical treatments or emergency situations. This equipment includes dialysis and dental units ( Dendana et al 2008 , Trabelsi et al 2010 , Retana-Moreira et al 2015 ), as well as combination shower units ( Retana-Moreira et al 2014 ).…”