Parasites, Hosts and Diseases

"18S rDNA"

Brief Communication

Molecular characterization of tick genera Haemaphysalis, Ixodes, and Amblyomma in Korea: Subin Lee, Badriah Alkathiri, Kyu-Sung Ahn, Jiwon Kim, So Youn Youn, Mi-Sun Yoo, Hyang-Sim Lee, Jae-Myung Kim, Dongmi Kwak, Sung Shik Shin, Seung-Hun Lee; Parasites Hosts Dis 2025;63(3):272-277.
Published online August 20, 2025; DOI: https://doi.org/10.3347/PHD.25035

Ticks are important ectoparasites that serve as key vectors of various pathogens, posing significant risks to both human and animal health. In this study, 3 genera and 5 species of ticks, including Haemaphysalis longicornis, H. flava, H. doenitzi, Ixodes nipponensis, and Amblyomma testudinarium, were analyzed both morphologically and molecularly. Morphological identification was conducted using established taxonomic keys, followed by molecular characterization through analysis of complete cytochrome c oxidase subunit 1 (cox1, 1,539 bp) and mitochondrial 16S rDNA (1,191–1,215 bp), and nuclear 18S rDNA (1,812–1,816 bp). Phylogenetic and pairwise distance analyses demonstrated that all 3 genes were effective for genus identification; cox1 and mitochondrial 16S rDNA were more effective than 18S rDNA in species identification. Additionally, this study is the first to identify H. doenitzi in the Korean mainland via molecular characterization. These results may serve as reference data for the molecular identification of tick species.

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Original Article

Loop-Mediated Isothermal Amplification Targeting 18S Ribosomal DNA for Rapid Detection of Acanthamoeba: Hye-Won Yang, Yu-Ran Lee, Noboru Inoue, Bijay Kumar Jha, Dinzouna-Boutamba Sylvatrie Danne, Hong-Kyun Kim, Junhun Lee, Youn-Kyoung Goo, Hyun-Hee Kong, Dong-Il Chung, Yeonchul Hong; Korean J Parasitol 2013;51(3):269-277.
Published online June 30, 2013; DOI: https://doi.org/10.3347/kjp.2013.51.3.269

Abstract
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Amoebic keratitis (AK) caused by Acanthamoeba is one of the most serious corneal infections. AK is frequently misdiagnosed initially as viral, bacterial, or fungal keratitis, thus ensuring treatment delays. Accordingly, the early detection of Acanthamoeba would contribute significantly to disease management and selection of an appropriate anti-amoebic therapy. Recently, the loop-mediated isothermal amplification (LAMP) method has been applied to the clinical diagnosis of a range of infectious diseases. Here, we describe a rapid and efficient LAMP-based method targeting Acanthamoeba 18S rDNA gene for the detection of Acanthamoeba using clinical ocular specimens in the diagnosis of AK. Acanthamoeba LAMP assays detected 11 different strains including all AK-associated species. The copy number detection limit for a positive signal was 10 DNA copies of 18S rDNA per reaction. No cross-reactivity with the DNA of fungi or other protozoa was observed. The sensitivity of LAMP assay was higher than those of Nelson primer PCR and JDP primer PCR. In the present study, LAMP assay based on directly heat-treated samples was found to be as efficient at detecting Acanthamoeba as DNA extracted using a commercial kit, whereas PCR was only effective when commercial kit-extracted DNA was used. This study showed that the devised Acanthamoeba LAMP assay could be used to diagnose AK in a simple, sensitive, and specific manner.

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Mini Review

Molecular Phylogeny of Acanthamoeba: Hyun Hee Kong; Korean J Parasitol 2009;47(Suppl):S21.
Published online October 26, 2009; DOI: https://doi.org/10.3347/kjp.2009.47.S.S21

Abstract
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After morphological grouping of Acanthamoeba by Pussard and Pons, phylogeny of the genus has been always a big topic to the researchers. Because of the variability of morphological characteristics, unchangeable and stable characters have been investigated for phylogenic criteria. Isoenzyme and mitochondrial DNA RFLP (Mt DNA RFLP) analyses revealed different patterns among strains assigned to a same species. Therefore, these characteristics would be considered as tools for strain discrimination than species identification. The most recently developed and the most promising method is the sequence analysis of 18s ribosomal RNA coding DNA (18s rDNA). The phylogenic tree based on comparison of 18s rDNA sequences distinguished the 3 morphological groups of Acanthamoeba and divided them into 12 unique sequence types (T1-T12 genotypes). Most clinical and environmental isolates belonged to the morphological group II and the genotype T4. In the Republic of Korea, 2 strains of Acanthamoeba, YM-2 and YM-3, were first isolated from the environment in 1974. However, phylogenic identification of Korean Acanthamoeba isolates from human infections or the environment were tried from the late 1990s. By RFLP analysis or total sequence analysis of 18s rDNA revealed that almost all clinical isolates including the one from a suspicious granulomatous amebic encephalitis patient belonged to the genotype T4. A large number of environmental isolates from contact lens storage cases, tapped water, and ocean sediments also belonged to the genotype T4. Almost identical strain characteristics, such as Mt DNA RFLP pattern of environmental isolates, with the clinical isolates could make a simple conclusion that most environmental isolates might be a potential keratopathogen.

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Original Articles

Molecular characterization of bacterial endosymbionts of Acanthamoeba isolates from infected corneas of Korean patients: Ying-Hua Xuan, Hak Sun Yu, Hae Jin Jeong, Sung-Yong Seol, Dong-Il Chung, Hyun-Hee Kong; Korean J Parasitol 2007;45(1):1-9.
Published online March 20, 2007; DOI: https://doi.org/10.3347/kjp.2007.45.1.1

Abstract
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The endosymbionts of 4 strains of Acanthamoeba (KA/E9, KA/E21, KA/E22, and KA/E23) isolated from the infected corneas of Korean patients were characterized via orcein stain, transmission electron microscopic examination, and 16S rDNA sequence analysis. Double membrane-bound, rod-shaped endosymbionts were distributed randomly throughout both the trophozoites and cysts of each of Acanthamoeba isolates. The endosymbionts of KA/E9, KA/E22, and KA/E23 were surrounded by electron-translucent areas. No lacunae-like structures were observed in the endosymbionts of KA/E21, the bacterial cell walls of which were studded with host ribosomes. Comparative analyses of the 16S rDNA sequences showed that the endosymbionts of KA/E9, KA/E22 and KA/E23 were closely related to Caedibacter caryophilus, whereas the KA/E21 endosymbiont was assigned to the Cytophaga-Flavobacterium-Bacteroides (CFB) phylum. In the 4 strains of Acanthamoeba, the hosts of the endosymbionts were identified as belonging to the Acanthamoeba castellanii complex, which corresponds to the T4 genotype. Acanthamoeba KA/E21 evidenced characteristics almost identical to those of KA/E6, with the exception of the existence of endosymbionts. The discovery of these endosymbionts from Acanthamoeba may prove essential to future studies focusing on interactions between the endosymbionts and the amoebic hosts.

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Eun Chul Kim, Man Soo Kim
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Molecular characterization of Acanthamoeba isolated from amebic keratitis related to orthokeratology lens overnight wear: Sun Joo Lee, Hae Jin Jeong, Ji Eun Lee, Jong Soo Lee, Ying Hua Xuan, Hyun-Hee Kong, Dong-Il Chung, Mee-Sun Ock, Hak Sun Yu; Korean J Parasitol 2006;44(4):313-320.
Published online December 20, 2006; DOI: https://doi.org/10.3347/kjp.2006.44.4.313

Abstract
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In an effort to characterize, on the molecular scale, the Acanthamoeba initially isolated from the cornea of an amoebic keratitis patient associated with overnight-wear orthokeratology lens in Korea, we conducted mitochondrial DNA restriction fragment length polymorphism, 18S rDNA sequencing, and drug sensitivity analyses on the isolate (KA/PE1). The patient was treated with polyhexamethylene biguanide, chlorhexidine and oral itraconazole, which resulted in resolution of the patient's ocular inflammation. The majority of the molecular characteristics of the KA/PE1 were determined to be identical, or quite similar, to those of A. castellanii Ma strain, which had been isolated also from amoebic keratitis. The risk of Acanthamoeba keratitis as a potential complication of overnight orthokeratology is briefly discussed.

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Genetic diversity of Acanthamoeba isolates from ocean sediments: Hua Liu, Young-Ran Ha, Sung-Tae Lee, Yean-Chul Hong, Hyun-Hee Kong, Dong-Il Chung; Korean J Parasitol 2006;44(2):117-125.
Published online June 20, 2006; DOI: https://doi.org/10.3347/kjp.2006.44.2.117

Abstract
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Genetic diversity of 18 Acanthamoeba isolates from ocean sediments was evaluated by comparing mitochondrial (mt) DNA RFLP, 18S rDNA sequences and by examining their cytopathic effects on human corneal epithelial cells versus reference strains. All isolates belonged to morphologic group II. Total of 16 restriction phenotypes of mtDNA from 18 isolates demonstrated the genetic diversity of Acanthamoeba in ocean sediments. Phylogenetic analysis using 18s rDNA sequences revealed that the 18 isolates were distinct from morphological groups I and III. Fifteen isolates showed close relatedness with 17 clinical isolates and A. castellanii Castellani and formed a lineage equivalent to T4 genotype of Byers' group. Two reference strains from ocean sediment, A. hatchetti BH-2 and A. griffini S-7 clustered unequivocally with these 15 isolates. Diversity among isolates was also evident from their cytopathic effects on human corneal cells. This is the first time describing Acanthamoeba diversity in ocean sediments in Korea.

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Evaluation of taxonomic validity of four species of Acanthamoeba: A. divionensis, A. paradivionensis, A. mauritaniensis, and A. rhysodes, inferred from molecular analyses: Hua Liu, Eun-Kyung Moon, Hak-Sun Yu, Hae-Jin Jeong, Yeon-Chul Hong, Hyun-Hee Kong, Dong-Il Chung; Korean J Parasitol 2005;43(1):7-13.
Published online March 20, 2005; DOI: https://doi.org/10.3347/kjp.2005.43.1.7

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The taxonomy of Acanthamoeba spp., an amphizoic amoeba which causes granulomatous amoebic encephalitis and chronic amoebic keratitis, has been revised many times. The taxonomic validity of some species has yet to be assessed. In this paper, we analyzed the morphological characteristics, nuclear 18s rDNA and mitochondrial 16s rDNA sequences and the Mt DNA RFLP of the type strains of four Acanthamoeba species, which had been previously designated as A. divionensis, A. parasidionensis, A. mauritaniensis, and A. rhysodes. The four isolates revealed characteristic group II morphology. They exhibited 18S rDNA sequence differences of 0.2-1.1% with each other, but more than 2% difference from the other compared reference strains. Four isolates formed a different clade from that of A. castellanii Castellani and the other strains in morphological group II on the phylogenetic tree. In light of these results, A. paradivionensis, A. divionensis, and A. mauritaniensis should be regarded as synonyms for A. rhysodes.

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