A rapid and reliable method for the clonal isolation of Acanthamoeba from environmental samples

Abstract

Acanthamoeba are abundant in a wide range of environments, and some species are responsible for cutaneous infections, keratitis, and granulomatous amoebic encephalitis (GAE). The conventional detection and isolation of amoeba from clinical and environmental samples involves sampling and culture on non-nutrient Ágar medium. Although efficient, this system requires several transfers in order to eliminate contaminants, and is not appropriate for the isolation of individual amoeba from samples with a biodiverse community. In this study we propose an alternative method for the isolation of monocystic clones of Acanthamoeba. The propose method involves sampling, enrichment, encystment induction, and direct cysts micromanipulation and culture on Ágar plates.

A history of over 40 years of potentially pathogenic free-living amoeba studies in Brazil - a systematic review

“…Environmental reports and the assessment of FLA as a water quality marker in Brazil -Acanthamoeba, Naegleria, Vermamoeba, and Balamuthia were detected in Brazilian habitats, including university buildings, hospitals, swimming pools, tap and freshwater, dust, soil samples, and inside insects (118)(119)(120)(121)(122)(123)(124)(125)(126)(127)(128)(129)(130)(131) (Table II).…”

“… M,D 120 2010 Hospital localities Acanthamoeba T4, T5, T3 M 121 2010 Public hospitals Acanthamoeba spp. M,D 126 2011 Tap water Acanthamoeba T2,T6, T4 M,D 127 2012 Soil and water samples Acanthamoeba T4, T5, T2-T6 M 128 2012 Aedes aegypti larvae Acanthamoeba T4, T3, T5 M,D 129 2012 Soil and water samples Acanthamoeba T7, T8, T9, T17 M,D 130 2012 Public hospitals A castellanii , A. polyphaga , A. lenticulata M,D 131 2013 Bromeliads leaves Acanthamoeba T4, T2/T6, T16 M,D …”

Diversity of free-living amoebas in the Monjolinho River in São Paulo state – morphological and molecular approaches

“…(175) On this concern, current Brazilian studies have used conserved regions of PCR amplified 18S rDNA to perform FLA identification. Regarding literature encountered between 2011 to 2020, mots of the FLA environmental and clinical investigations conducted a molecular based assay (127,141,143,150,(176)(177)(178)(179)(180)(181)(182) in contrast with few studies based on amoeba immunodetection tools. (183)(184) Classified as environmental isolated groups, the first one is dated from 1986 by pointing out N. fowleri presence in an artificial lake.…”

“…(150) Brazilian river water evaluated through amplicon based metagenomics ranked positive to the presence of Amoebozoa, however without describing the class or genus that they belong. (180) Reports have amplified Acanthamoeba genes in tap water samples, (176)(177)185) cooling towers (186) and swimming pools. (141,(143)(144) Not only freshwater but also estuarine systems were sampled in Brazilian habitats in which Acanthamoeba T3 and T4 genotypes were encountered.…”

“…into three major groups (I, II, III). (23,177,230) Presently, studies concerned with resolving differences within Acanthamoeba isolates have been using modern non-morphological based taxonomy, mainly based on 18S rDNA sequencing. 23) in which our data is compliant with, due to dimensions varying from about 12-22 µm (Table 3.6, Lines 9 to 11).…”