Thermotolerant Acanthamoeba spp. isolated from recreational water in Gorgan City, north of Iran

  • Somayeh Maghsoodloorad
  • Elham Maghsoodloorad
  • Amir Tavakoli Kareshk
  • Mohammad Hossein Motazedian
  • Mustapha Ahmed Yusuf
  • Rahmat SolgiEmail author
Original Article


Acanthamoeba as free-living parasites are scattered ubiquitously, throughout the world. This study was aimed to evaluate the presence of Acanthamoeba spp. genotypes in the recreational water sources in Gorgan County, the capital of Golestan Province using both morphological and molecular approaches. Thirty water samples were collected from different recreational waters in Gorgan, the capital of Golestan Province, northern Iran during 2015–2016. Samples were filtered and followed by culture in non-nutrient agar. Acanthamoeba were identified both by morphological and molecular analysis. The pathogenical potential of positive cloned samples were also determined using tolerance test. Twenty-six percent of recreational water were identified as Acanthamoeba spp. based on the morphological analysis and from these positive samples, five samples were successfully sequenced after molecular studies. Phylogenetic analysis showed the clustering of four samples in T4 genotype group and only one sample as T15 genotype. Thermotolerance test revealed that all cloned samples were highly positive. Since the attractiveness of recreational places for people is increasing, the potential risk of this water should be monitored routinely in each region. More studies are needed to better evaluate the risk of this ubiquitous parasite for the human.


Acanthamoeba Genotyping techniques Morphological Molecular Iran 



The authors would like to express their gratitude to Mrs. Shahrbanoo Naderi for her useful collaboration.

Authors’ contributions

All authors contributed to the study design. RS was leader of the research. SM, EM, AT, MHM and MAY carried out experimental tests and prepared the Manuscript. All authors read and approved the final version of the manuscript.


Funding was provided by Golestan University of Medical Sciences.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.


  1. Altschul SF, Madden TL, Schaffer AA, Zhang J, Zhang Z, Miller W, Lipman DJ (1997) Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucleic Acids Res 25(17):3389–3402Google Scholar
  2. Booton GC, Kelly DJ, Chu YW, Seal DV, Houang E, Lam DS, Byers TJ, Fuerst PA (2002) 18S ribosomal DNA typing and tracking of Acanthamoeba species isolates from corneal scrape specimens, contact lenses, lens cases, and home water supplies of Acanthamoeba keratitis patients in Hong Kong. J Clin Microbiol 40(5):1621–1625Google Scholar
  3. Forrest G (2004) Gastrointestinal infections in immunocompromised hosts. Curr Opin Gastroenterol 20(1):16–21Google Scholar
  4. Kimura M (1980) A simple method for estimating evolutionary rates of base substitutions through comparative studies of nucleotide sequences. J Mol Evol 16(2):111–120Google Scholar
  5. Lorenzo-Morales J, Ortega-Rivas A, Martínez E, Khoubbane M, Artigas P, Periago MV, Foronda P, Abreu-Acosta N, Valladares B, Mas-Coma S (2006) Acanthamoeba isolates belonging to T1, T2, T3, T4 and T7 genotypes from environmental freshwater samples in the Nile Delta region, Egypt. Acta Trop 100(1):63–69Google Scholar
  6. Lorenzo-Morales J, Martin-Navarro CM, Lopez-Arencibia A, Arnalich-Montiel F, Pinero JE, Valladares B (2013) Acanthamoeba keratitis: an emerging disease gathering importance worldwide? Trends Parasitol 29(4):181–187Google Scholar
  7. Lorenzo-Morales J, Khan NA, Walochnik J (2015) An update on Acanthamoeba keratitis: diagnosis, pathogenesis and treatment. Parasite 22:10Google Scholar
  8. Marciano-Cabral F, Cabral G (2003) Acanthamoeba spp. as agents of disease in humans. Clin Microbiol Rev 16(2):273–307Google Scholar
  9. Mollalo A, Alimohammadi A, Shirzadi M, Malek M (2015) Geographic information system-based analysis of the spatial and spatio-temporal distribution of zoonotic cutaneous leishmaniasis in Golestan Province, north-east of Iran. Zoonoses Public Health 62(1):18–28Google Scholar
  10. Montalbano Di Filippo M, Santoro M, Lovreglio P, Monno R, Capolongo C, Calia C, Fumarola L, D’Alfonso R, Berrilli F, Di Cave D (2015) Isolation and molecular characterization of free-living amoebae from different water sources in Italy. Int J Environ Res Public Health 12(4):3417–3427Google Scholar
  11. Nazar M, Haghighi A, Niyyati M, Eftekhar M, Tahvildar-Biderouni F, Taghipour N, Abadi A, Nazemalhosseini Mojarad E, Athari A (2011) Genotyping of Acanthamoeba isolated from water in recreational areas of Tehran, Iran. J Water Health 9(3):603–608Google Scholar
  12. Niyyati M, Rezaeian M (2015) Current status of Acanthamoeba in Iran: a narrative review article. Iran J Parasitol 10(2):157Google Scholar
  13. Niyyati M, Lasjerdi Z, Nazar M, Haghighi A, Nazemalhosseini Mojarad E (2012) Screening of recreational areas of rivers for potentially pathogenic free-living amoebae in the suburbs of Tehran, Iran. J Water Health 10(1):140–146Google Scholar
  14. Niyyati M, Lasgerdi Z, Lorenzo-Morales J (2015) Detection and molecular characterization of potentially pathogenic free-living amoebae from water sources in Kish Island, Southern Iran. Microbiol Insights 8(Suppl 1):1–6Google Scholar
  15. Niyyati M, Saberi R, Latifi A, Lasjerdi Z (2016a) Distribution of Acanthamoeba genotypes isolated from recreational and therapeutic geothermal water sources in Southwestern Iran. Environ Health Insights 10:69–74Google Scholar
  16. Niyyati M, Saberi R, Lorenzo-Morales J, Salehi R (2016b) High occurrence of potentially-pathogenic free-living amoebae in tap water and recreational water sources in South-West Iran. Trop Biomed 33(1):95–101Google Scholar
  17. Page FC (1988) A new key to freshwater and soil gymnamoebae. Freshwater Biological Association, AmblesideGoogle Scholar
  18. Pagnier I, Raoult D, La Scola B (2008) Isolation and identification of amoeba-resisting bacteria from water in human environment by using an Acanthamoeba polyphaga co-culture procedure. Environ Microbiol 10(5):1135–1144Google Scholar
  19. Radford CF, Bacon AS, Dart JK, Minassian DC (1995) Risk factors for Acanthamoeba keratitis in contact lens users: a case-control study. BMJ 310(6994):1567–1570Google Scholar
  20. Radford C, Minassian D, Dart J (2002) Acanthamoeba keratitis in England and Wales: incidence, outcome, and risk factors. Br J Ophthalmol 86(5):536–542Google Scholar
  21. Saitou N, Nei M (1987) The neighbor-joining method: a new method for reconstructing phylogenetic trees. Mol Biol Evol 4(4):406–425Google Scholar
  22. Schroeder JM, Booton GC, Hay J, Niszl IA, Seal DV, Markus MB, Fuerst PA, Byers TJ (2001) Use of subgenic 18S ribosomal DNA PCR and sequencing for genus and genotype identification of Acanthamoebae from humans with keratitis and from sewage sludge. J Clin Microbiol 39(5):1903–1911Google Scholar
  23. Schuster FL, Visvesvara GS (2004) Free-living amoebae as opportunistic and non-opportunistic pathogens of humans and animals. Int J Parasitol 34(9):1001–1027Google Scholar
  24. Solgi R, Niyyati M, Haghighi A, Mojarad EN (2012a) Occurrence of thermotolerant Hartmannella vermiformis and Naegleria spp. in Hot Springs of Ardebil Province, Northwest Iran. Iran J Parasitol 7(2):47–52Google Scholar
  25. Solgi R, Niyyati M, Haghighi A, Taghipour N, Tabaei SJ, Eftekhar M, Nazemalhosseini Mojarad E (2012b) Thermotolerant Acanthamoeba spp. isolated from therapeutic hot springs in Northwestern Iran. J Water Health 10(4):650–656Google Scholar
  26. Tamura K, Stecher G, Peterson D, Filipski A, Kumar S (2013) MEGA6: molecular evolutionary genetics analysis version 6.0. Mol Biol Evol 30(12):2725–2729Google Scholar
  27. Thomas V, McDonnell G, Denyer SP, Maillard J-Y (2010) Free-living amoebae and their intracellular pathogenic microorganisms: risks for water quality. FEMS Microbiol Rev 34(3):231–259Google Scholar
  28. Todd CD, Reyes-Batlle M, Pinero JE, Martinez-Carretero E, Valladares B, Streete D, Lorenzo-Morales J, Lindo JF (2015) Isolation and molecular characterization of Acanthamoeba genotypes in recreational and domestic water sources from Jamaica, West Indies. J Water Health 13(3):909–919Google Scholar
  29. Visvesvara GS, Moura H, Schuster FL (2007) Pathogenic and opportunistic free-living amoebae: Acanthamoeba spp., Balamuthia mandrillaris, Naegleria fowleri, and Sappinia diploidea. FEMS Immunol Med Microbiol 50(1):1–26Google Scholar
  30. Wagner C, Reyes-Batlle M, Hernan A, Rojas E, Perez G, Lopez-Arencibia A, Sifaoui I, Martinez-Carretero E, Pinero JE, Valladares B, Lorenzo-Morales J (2016) High occurrence of Acanthamoeba genotype T4 in soil sources from Bolivar State, Venezuela. Acta Parasitol 61(3):466–470Google Scholar

Copyright information

© Indian Society for Parasitology 2019

Authors and Affiliations

  • Somayeh Maghsoodloorad
    • 1
  • Elham Maghsoodloorad
    • 2
  • Amir Tavakoli Kareshk
    • 3
  • Mohammad Hossein Motazedian
    • 4
  • Mustapha Ahmed Yusuf
    • 5
  • Rahmat Solgi
    • 3
    Email author
  1. 1.Department of Parasitology and Mycology, School of MedicineGolestan University of Medical ScienceGorgānIran
  2. 2.Sayyad Shirazi HospitalGolestan University of Medical ScienceGorgānIran
  3. 3.Infectious Disease Research CenterBirjand University of Medical SciencesBirjandIran
  4. 4.Department of Parasitology and Mycology, School of MedicineShiraz University of Medical SciencesShirazIran
  5. 5.Department of Medical Entomology and Vector Control, School of Public Health, International CampusTehran University of Medical ScienceTehranIran

Personalised recommendations