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      Isolation and identification of Acanthamoeba from pond water of parks in a tropical and subtropical region in the Middle East, and its relation with physicochemical parameters

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          Abstract

          Background

          Free-living amoeba (FLA) are wide-spread protozoa that are found in different environmental sources including water, soil, dust, hospital units and ventilation areas. These amoebas can act as opportunistic or non-opportunistic pathogens. Among FLAs, some genera such as Acanthamoeba are important because of their potential pathogenic ability in humans . The purpose of this study is to identify of Acanthamoeba isolated from pond water of parks in a tropical and subtropical region in the Middle East, and its relation with physicochemical parameters.From August to December 2015, 90 samples were collected from pond water of parks of 13 regions of Mashhad City. Physicochemical parameters were measured in situ. After filtering, the samples were cultured on Bacto-agar enriched with Escherichia coli. PCR analysis was conducted on the culture-positive samples, and then the PCR products were sequenced. Statistical analysis was performed by SPSS software and Fisher’s Exact and Mann-Whitney test.

          Results

          Among the samples that were examined, 19 samples (21.1%) were positive for Acanthamoeba. The sequencing revealed that Acanthamoeba isolates belonged to T4 genotype of Acanthamoeba. There was no significant relationship between physicochemical parameters and the presence of Acanthamoeba.

          Conclusion

          The prevalence of Acanthamoeba in pond water of parks was relatively high, but there was no significant relationship between physicochemical parameters and the presence of Acanthamoeba.

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          Most cited references13

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          Microbiome of free-living amoebae isolated from drinking water.

          Free-living amoebae (FLA) are protozoa that can be found in water networks where they prey on bacteria within biofilms. Most bacteria are digested rapidly by phagocytosis, however some are able to survive within amoebae and some are even able to multiply, as it is the case for Legionella pneumophila. These resisting bacteria are a potential health problem as they could also resist to macrophage phagocytosis. Several publications already reported intra-amoebal bacteria but the methods of identification did not allow metagenomic analysis and are partly based on co-culture with one selected amoebal strain. The aim of our study was to conduct a rRNA-targeted metagenomic analysis on amoebae and intra-amoebal bacteria found in drinking water network, to provide the first FLA microbiome in environmental strains. Three sites of a water network were sampled during four months. Culturable FLA were isolated and total DNA was prepared, allowing purification of both amoebal and bacterial DNA. Metagenomic studies were then conducted through 18S or 16S amplicons sequencing. Hartmannella was by far the most represented genus of FLA. Regarding intra-amoebal bacteria, 54 genera were identified, among which 21 were newly described intra-amoebal bacteria, underlying the power of our approach. There were high differences in bacterial diversity between the three sites. Several genera were highly represented and/or found at least in two sites, underlying that these bacteria could be able to multiply within FLA. Our method is therefore useful to identify FLA microbiome and could be applied to other networks to have a more comprehensive view of intra-amoebal diversity. Copyright © 2013 Elsevier Ltd. All rights reserved.
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            Detection of bacterial endosymbionts in clinical acanthamoeba isolates.

            To determine the presence of 4 clinically relevant bacterial endosymbionts in Acanthamoeba isolates obtained from patients with Acanthamoeba keratitis (AK) and the possible contribution of endosymbionts to the pathogenesis of AK. Experimental study. Acanthamoeba isolates (N = 37) recovered from the cornea and contact lens paraphernalia of 23 patients with culture-proven AK and 1 environmental isolate. Acanthamoeba isolates were evaluated for the presence of microbial endosymbionts belonging to the bacterial genera Legionella, Pseudomonas, Mycobacterium, and Chlamydia using molecular techniques (polymerase chain reaction and sequence analysis, fluorescence in situ hybridization) and transmission electron microscopy. Corneal toxicity and virulence of Acanthamoeba isolates with and without endosymbionts were compared using a cytopathic effect (CPE) assay on human corneal epithelial cells in vitro. Initial visual acuity, location and characteristics of the infiltrate, time to detection of the infection, and symptom duration at presentation were evaluated in all patients. Prevalence and potential pathobiology of bacterial endosymbionts detected in Acanthamoeba isolates recovered from AK. Twenty-two (59.4%) of the 38 cultures examined contained at least 1 bacterial endosymbiont. One isolate contained 2 endosymbionts, Legionella and Chlamydia, confirmed by fluorescence in situ hybridization. Corneal toxicity (CPE) was significantly higher for Acanthamoeba-hosting endosymbionts compared with isolates without endosymbionts (P 0.05), central location (P 0.05), and longer symptom duration at presentation (P>0.05). Most Acanthamoeba isolates responsible for AK harbor 1 or more bacterial endosymbionts. The presence of endosymbionts enhances the corneal pathogenicity of Acanthamoeba isolates and may impact detection time and clinical features of AK. Copyright 2010 American Academy of Ophthalmology. Published by Elsevier Inc. All rights reserved.
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              Cooccurrence of free-living amoebae and nontuberculous Mycobacteria in hospital water networks, and preferential growth of Mycobacterium avium in Acanthamoeba lenticulata.

              The incidence of lung and other diseases due to nontuberculous mycobacteria (NTM) is increasing. NTM sources include potable water, especially in households where NTM populate pipes, taps, and showerheads. NTM share habitats with free-living amoebae (FLA) and can grow in FLA as parasites or as endosymbionts. FLA containing NTM may form cysts that protect mycobacteria from disinfectants and antibiotics. We first assessed the presence of FLA and NTM in water and biofilm samples collected from a hospital, confirming the high prevalence of NTM and FLA in potable water systems, particularly in biofilms. Acanthamoeba spp. (genotype T4) were mainly recovered (8/17), followed by Hartmannella vermiformis (7/17) as well as one isolate closely related to the genus Flamella and one isolate only distantly related to previously described species. Concerning mycobacteria, Mycobacterium gordonae was the most frequently found isolate (9/17), followed by Mycobacterium peregrinum (4/17), Mycobacterium chelonae (2/17), Mycobacterium mucogenicum (1/17), and Mycobacterium avium (1/17). The propensity of Mycobacterium avium hospital isolate H87 and M. avium collection strain 104 to survive and replicate within various FLA was also evaluated, demonstrating survival of both strains in all amoebal species tested but high replication rates only in Acanthamoeba lenticulata. As A. lenticulata was frequently recovered from environmental samples, including drinking water samples, these results could have important consequences for the ecology of M. avium in drinking water networks and the epidemiology of disease due to this species.
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                Author and article information

                Contributors
                najafpooraa@mums.ac.ir
                Journal
                BMC Microbiol
                BMC Microbiol
                BMC Microbiology
                BioMed Central (London )
                1471-2180
                19 October 2018
                19 October 2018
                2018
                : 18
                : 139
                Affiliations
                [1 ]ISNI 0000 0001 2198 6209, GRID grid.411583.a, Student Research Committee, , Mashhad University of Medical Sciences, ; Mashhad, Iran
                [2 ]ISNI 0000 0001 2198 6209, GRID grid.411583.a, Social Determinants of Health Research Center, , Mashhad University of Medical Sciences, ; Mashhad, Iran
                [3 ]Global Center for Environmental Remediation, University of Newcastle, Callaghan, NSW 2308 Iran
                [4 ]Allergy Research Center Mashhad, University of Medical Sciences, Mashhad, Iran
                [5 ]Department of Epidemiology and Biostatistics, University of Medical Sciences, Mashhad, Iran
                [6 ]ISNI 0000 0004 0405 433X, GRID grid.412606.7, Department of Medical Parasitology and Mycology, School of Medicine, , Qazvin University of Medical Sciences, ; Qazvin, Iran
                Author information
                http://orcid.org/0000-0003-0774-4734
                Article
                1301
                10.1186/s12866-018-1301-x
                6195741
                30340522
                d76bfb43-50e0-456f-9e45-8a7929388214
                © The Author(s). 2018

                Open Access This article is distributed under the terms of the Creative Commons Attribution 4.0 International License ( http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver ( http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.

                History
                : 24 May 2017
                : 4 October 2018
                Funding
                Funded by: The current study is an approved proposal in Mashhad University of Medical Sciences (MUMS) with the code 940310.
                Award ID: Authors would like to thanks from MUMS for the financial support.
                Award Recipient :
                Categories
                Research Article
                Custom metadata
                © The Author(s) 2018

                Microbiology & Virology
                ponds water,parks,acanthamoeba,tropical and subtropical region
                Microbiology & Virology
                ponds water, parks, acanthamoeba, tropical and subtropical region

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