Occurrence and molecular characterisation of
Acanthamoeba isolated from recreational hot springs
in Malaysia: evidence of pathogenic potential
Rosnani Hanim Mohd Hussain, Ahmad Razali Ishak,
Mohamed Kamel Abdul Ghani, Naveed Ahmed Khan,
Ruqaiyyah Siddiqui and Tengku Shahrul Anuar
ABSTRACT
This study aimed to identify the Acanthamoeba genotypes and their pathogenic potential in five
recreational hot springs in Peninsular Malaysia. Fifty water samples were collected between April
and September 2018. Physical parameters of water quality were measured in situ while chemical and
microbiological analyses were performed in the laboratory. All samples were filtered through the
nitrocellulose membrane and tested for Acanthamoeba using both cultivation and polymerase chain
reaction (PCR) by targeting the 18S ribosomal RNA gene. The pathogenic potential of all positive
isolates was identified using thermo- and osmotolerance tests. Thirty-eight (76.0%) samples were
positive for Acanthamoeba. Water temperature (P ¼ 0.035), chemical oxygen demand (P ¼ 0.026),
sulphate (P ¼ 0.002) and Escherichia coli (P < 0.001) were found to be significantly correlated with
the presence of Acanthamoeba. Phylogenetic analysis revealed that 24 samples belonged to
genotype T4, nine (T15), two (T3) and one from each genotype T5, T11 and T17. Thermo- and
osmotolerance tests showed that 6 (15.79%) of the Acanthamoeba strains were highly pathogenic.
The existence of Acanthamoeba in recreational hot springs should be considered as a health threat
among the public especially for high-risk people. Periodic surveillance of hot spring waters and
posting warning signs by health authorities is recommended to prevent disease related to
pathogenic Acanthamoeba.
Rosnani Hanim Mohd Hussain
Tengku Shahrul Anuar (corresponding author)
Centre of Medical Laboratory Technology, Faculty
of Health Sciences,
Universiti Teknologi MARA,
Puncak Alam Campus, Selangor,
Malaysia
and
Integrative Pharmacogenomics Institute,
Universiti Teknologi MARA,
Puncak Alam Campus, Selangor,
Malaysia
E-mail: tengku9235@uitm.edu.my
Ahmad Razali Ishak
Centre of Environmental Health and Safety, Faculty
of Health Sciences,
Universiti Teknologi MARA,
Puncak Alam Campus, Selangor,
Malaysia
Mohamed Kamel Abdul Ghani
Programme of Biomedical Sciences, School of
Diagnostic and Applied Health Sciences,
Universiti Kebangsaan Malaysia,
Jalan Raja Muda Abdul Aziz, Kuala Lumpur,
Malaysia
Naveed Ahmed Khan
Ruqaiyyah Siddiqui
Department of Biological Sciences, School of
Science and Technology,
Sunway University,
Subang Jaya, Selangor,
Malaysia
Key words | Acanthamoeba, culture, genotype, hot spring, Malaysia, PCR
INTRODUCTION
The genus Acanthamoeba is free-living amoebae, which is
ubiquitous in natural and human-made aquatic environ-
ments, including hot spring water, tap water, lakes, ponds,
freshwater sources and swimming pools (Huang & Hsu
). Founded on their cyst characteristics, Acanthamoeba
can be classified into three markedly different groups (I, II
and III) (Pussard & Pons ) and encompassing more
than 25 nominal species (Booton et al. ). Furthermore,
consecutive research investigating molecular and biochemi-
cal techniques have highlighted numerous examples of
incongruity concerning the genotype or Acanthamoeba
grouping (Corsaro & Venditti ). As a result, the genus
classification and taxonomy of Acanthamoeba have pre-
sently been categorised into 21 diverse genotypes that are
813 © IWA Publishing 2019 Journal of Water and Health | 17.5 | 2019
doi: 10.2166/wh.2019.214
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