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Original Article

Establishing a Cre/loxP-based genetic manipulation system for Acanthamoeba: Targeted genome editing and stable reporter expression
Ja Moon Aung, So-Young Joo, Byoung-Kuk Na, Seunghyeok Bang, Minsang Shin, Youn-Kyoung Goo, Yeonchul Hong
Parasites Hosts Dis 2025;63(1):25-36.
Published online February 25, 2025
DOI: https://doi.org/10.3347/PHD.24078
Acanthamoeba is an opportunistic pathogen responsible for granulomatous amoebic encephalitis and amoebic keratitis. Despite its clinical significance, effective treatments remain challenging due to a limited understanding of its pathogenic mechanism. This study developed a genetic manipulation system in Acanthamoeba to facilitate gene function and drug screening studies. We applied the Cre/loxP system to integrate the gene encoding the tdTomato fluorescent protein into the genome of Acanthamoeba castellanii via homologous recombination. The polyubiquitin gene and its untranslated regions were identified and verified, after which the tdTomato gene was cloned between the untranslated regions of the polyubiquitin gene. The construct was then introduced into the Acanthamoeba genome using a modified pLPBLP vector containing loxP sites. Cre recombinase was utilized to remove the neomycin resistance cassette flanked by loxP sites, and genetically modified cells were selected by clonal dilution. The integration of the tdTomato gene, confirmed through PCR and fluorescence microscopy, showed stable expression in both trophozoites and cysts without the need for antibiotic selection. We demonstrated the feasibility of antibiotic-free reporter gene expression in Acanthamoeba. The system provides a valuable tool for functional genomics, allowing us to explore gene functions in Acanthamoeba and develop reliable drug screening models. Furthermore, the ability to express genes without the continuous use of selection markers opens up new possibilities for studying the pathobiology of this pathogen and advancing the development of novel therapeutic strategies against Acanthamoeba infections.

Citations

Citations to this article as recorded by  Crossref logo
  • The effect of Legionella pneumophila infection on the encystation of Acanthamoeba castellanii
    Hye-Jeong Jo, Hae-Ahm Lee, Fu-Shi Quan, Hyun-Hee Kong, Eun-Kyung Moon
    Parasites, Hosts and Diseases.2026; 64(1): 37.     CrossRef
  • 2,449 View
  • 297 Download
  • Crossref

Brief Communication

Appropriate Time for Primaquine Treatment to Reduce Plasmodium falciparum Transmission in Hypoendemic Areas
Polrat Wilairatana, Srivicha Krudsood, Noppadon Tangpukdee
Korean J Parasitol 2010;48(2):179-182.
Published online June 17, 2010
DOI: https://doi.org/10.3347/kjp.2010.48.2.179

Artemesinin-combination therapies (ACT) for falciparum malaria reduce gametocyte carriage, and therefore reduce transmission. Artemisinin derivatives will act against only young gametocytes whereas primaquine acts on mature gametocytes which are present usually in the circulation at the time when the patient presents for treatment. Both artemisinin derivatives and primaquine have short half-lives, less than 1 hr and 7 hr, respectively. Therefore, asexual parasites or young gametocytes remain after completed ACT. A single dose of primaquine (0.50-0.75 mg base/kg) at the end of ACT can kill only mature gametocytes but cannot kill young gametocytes (if present). Remaining asexual forms after completion of ACT course, e.g., artesunate-mefloquine for 3 days, may develop to mature gametocytes 7-15 days later. Thus, an additional dose of primaquine (0.50-0.75 mg base/kg) given 2 weeks after ACT completion may be beneficial for killing remaining mature gametocytes and contribute to more interruption of Plasmodium falciparum transmission than giving only 1 single dose of primaquine just after completing ACT.

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  • Extended-spectrum antiprotozoal bumped kinase inhibitors: A review
    Wesley C. Van Voorhis, J. Stone Doggett, Marilyn Parsons, Matthew A. Hulverson, Ryan Choi, Samuel L.M. Arnold, Michael W. Riggs, Andrew Hemphill, Daniel K. Howe, Robert H. Mealey, Audrey O.T. Lau, Ethan A. Merritt, Dustin J. Maly, Erkang Fan, Kayode K. Oj
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    Daniel Gonçalves, Patrick Hunziker
    Malaria Journal.2016;[Epub]     CrossRef
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    Solomon M Abay, Leonardo Lucantoni, Nisha Dahiya, Geme Dori, Edson G Dembo, Fulvio Esposito, Guilio Lupidi, Sonny Ogboi, Robert K Ouédraogo, Annamaria Sinisi, Orazio Taglialatela-Scafati, R Serge Yerbanga, Massimo Bramucci, Luana Quassinti, Jean Bosco Oué
    Malaria Journal.2015;[Epub]     CrossRef
  • Development of potent and selective Plasmodium falciparum calcium-dependent protein kinase 4 (PfCDPK4) inhibitors that block the transmission of malaria to mosquitoes
    Rama Subba Rao Vidadala, Kayode K. Ojo, Steven M. Johnson, Zhongsheng Zhang, Stephen E. Leonard, Arinjay Mitra, Ryan Choi, Molly C. Reid, Katelyn R. Keyloun, Anna M.W. Fox, Mark Kennedy, Tiffany Silver-Brace, Jen C.C. Hume, Stefan Kappe, Christophe L.M.J.
    European Journal of Medicinal Chemistry.2014; 74: 562.     CrossRef
  • Strategic use of antimalarial drugs that block falciparum malaria parasite transmission to mosquitoes to achieve local malaria elimination
    Rashad Abdul-Ghani, John C. Beier
    Parasitology Research.2014; 113(10): 3535.     CrossRef
  • Blocking malaria transmission to Anopheles mosquitoes using artemisinin derivatives and primaquine: a systematic review and meta-analysis
    Solomon Mequanente Abay
    Parasites & Vectors.2013;[Epub]     CrossRef
  • Plasmodium Cell Biology Should Inform Strategies Used In The Development Of Antimalarial Transmission-Blocking Drugs
    Michael J Delves
    Future Medicinal Chemistry.2012; 4(18): 2251.     CrossRef
  • Efficacy of different primaquine-based antimalarial regimens against Plasmodium falciparum gametocytemia
    Eliana M. Arango, Yulieth A. Upegui, Jaime Carmona-Fonseca
    Acta Tropica.2012; 122(2): 177.     CrossRef
  • Primaquina, gametocitemia de Plasmodium falciparum y bloqueo de transmisión: ineficacia del actual régimen de dosificación
    Jaime Carmona-Fonseca, Eliana María Arango Flórez
    MedUNAB.2012; 15(1): 14.     CrossRef
  • Methodology and application of flow cytometry for investigation of human malaria parasites
    Brian T. Grimberg
    Journal of Immunological Methods.2011; 367(1-2): 1.     CrossRef
  • Blocking Plasmodium falciparum Malaria Transmission with Drugs: The Gametocytocidal and Sporontocidal Properties of Current and Prospective Antimalarials
    Anthony E. Kiszewski
    Pharmaceuticals.2010; 4(1): 44.     CrossRef
  • 9,191 View
  • 129 Download
  • Crossref

Original Articles

Gametocyte Clearance in Uncomplicated and Severe Plasmodium falciparum Malaria after Artesunate-Mefloquine Treatment in Thailand
Noppadon Tangpukdee, Srivicha Krudsood, Sriripun Srivilairit, Nanthaporn Phophak, Putza Chonsawat, Wimon Yanpanich, Shigeyuki Kano, Polrat Wilairatana
Korean J Parasitol 2008;46(2):65-70.
Published online June 20, 2008
DOI: https://doi.org/10.3347/kjp.2008.46.2.65

Artemisinin-based combination therapy (ACT) is currently promoted as a strategy for treating both uncomplicated and severe falciparum malaria, targeting asexual blood-stage Plasmodium falciparum parasites. However, the effect of ACT on sexual-stage parasites remains controversial. To determine the clearance of sexual-stage P. falciparum parasites from 342 uncomplicated, and 217 severe, adult malaria cases, we reviewed and followed peripheral blood sexual-stage parasites for 4 wk after starting ACT. All patients presented with both asexual and sexual stage parasites on admission, and were treated with artesunate-mefloquine as the standard regimen. The results showed that all patients were asymptomatic and negative for asexual forms before discharge from hospital. The percentages of uncomplicated malaria patients positive for gametocytes on days 3, 7, 14, 21, and 28 were 41.5, 13.1, 3.8, 2.0, and 2.0%, while the percentages of gametocyte positive severe malaria patients on days 3, 7, 14, 21, and 28 were 33.6, 8.2, 2.7, 0.9, and 0.9%, respectively. Although all patients were negative for asexual parasites by day 7 after completion of the artesunate-mefloquine course, gametocytemia persisted in some patients. Thus, a gametocytocidal drug, e.g., primaquine, may be useful in combination with an artesunate-mefloquine regimen to clear gametocytes, so blocking transmission more effectively than artesunate alone, in malaria transmission areas.

Citations

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  • Knowledge about Asymptomatic Malaria and Acceptability of Using Artemisia afra Tea among Health Care Workers (HCWs) in Yaoundé, Cameroon: A Cross-Sectional Survey
    Abenwie Suh Nchang, Lahngong Shinyuy, Sandra Noukimi, Sylvia Njong, Sylvie Bambara, Edgar Kalimba, Joseph Kamga, Stephen Ghogomu, Michel Frederich, Jean Talom, Jacob Souopgui, Annie Robert
    International Journal of Environmental Research and Public Health.2023; 20(13): 6309.     CrossRef
  • Parasite clearance dynamics in children hospitalised with severe malaria in the Ho Teaching Hospital, Volta Region, Ghana
    Laura Paris, Richmond G. Tackie, Khalid B. Beshir, John Tampuori, Gordon A. Awandare, Fred N. Binka, Britta C. Urban, Bismarck Dinko, Colin J. Sutherland
    Parasite Epidemiology and Control.2022; 19: e00276.     CrossRef
  • NPC1161B, an 8-Aminoquinoline Analog, Is Metabolized in the Mosquito and Inhibits Plasmodium falciparum Oocyst Maturation
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    Frontiers in Pharmacology.2019;[Epub]     CrossRef
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    Mitali Mishra, Vikash K. Mishra, Varsha Kashaw, Arun K. Iyer, Sushil Kumar Kashaw
    European Journal of Medicinal Chemistry.2017; 125: 1300.     CrossRef
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    Jared N. Balaich, Derrick K. Mathias, Baldwyn Torto, Bryan T. Jackson, Dingyin Tao, Babak Ebrahimi, Brian B. Tarimo, Xavier Cheseto, Woodbridge A. Foster, Rhoel R. Dinglasan
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  • Gametocyte Clearance Kinetics Determined by Quantitative Magnetic Fractionation in Melanesian Children with Uncomplicated Malaria Treated with Artemisinin Combination Therapy
    Stephan Karl, Moses Laman, Brioni R. Moore, John Benjamin, Tamarah Koleala, Clemencia Ibam, Bernadine Kasian, Peter M. Siba, Andreea Waltmann, Ivo Mueller, Robert C. Woodward, Timothy G. St. Pierre, Timothy M. E. Davis
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    Rashad Abdul-Ghani, John C. Beier
    Parasitology Research.2014; 113(10): 3535.     CrossRef
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    Acta Tropica.2012; 122(2): 177.     CrossRef
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    International Journal for Parasitology.2012; 42(11): 999.     CrossRef
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    Michael J Delves
    Future Medicinal Chemistry.2012; 4(18): 2251.     CrossRef
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    Jaime Carmona-Fonseca, Eliana María Arango Flórez
    MedUNAB.2012; 15(1): 14.     CrossRef
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    Polrat Wilairatana, Srivicha Krudsood, Noppadon Tangpukdee
    The Korean Journal of Parasitology.2010; 48(2): 179.     CrossRef
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  • 110 Download
  • Crossref
A survey on head lice infestation in Korea (2001) and the therapeutic efficacy of oral trimethoprim/sulfamethoxazole adding to lindane shampoo
Seobo Sim, In-Yong Lee, Kyu-Jae Lee, Jang-Hoon Seo, Kyung-Il Im, Myeong Heon Shin, Tai-Soon Yong
Korean J Parasitol 2003;41(1):57-61.
Published online March 20, 2003
DOI: https://doi.org/10.3347/kjp.2003.41.1.57

Total of 7,495 children including 3,908 boys and 3,587 girls from a kindergarten and 15 primary schools were examined for head lice infestation (HLI). The overall prevalence of HLI in this study was found to be 5.8%. Head lice were much more commonly detected in girls than in boys with prevalence of 11.2% and 0.9%, respectively. Sixty-nine children with HLI were treated with 1% lindane shampoo alone (group 1), and 45 children with HLI were treated with 1% lindane shampoo and oral trimethoprim/sulfamethoxazole (group 2), and follow-up visits were conducted 2 and 4 weeks later. The children who still had HLI 2 weeks after the primary treatment were treated again. At the 2-week follow-up visit, the treatment success rates of groups 1 and 2 were 76.8% and 86.7%, respectively, and at the 4-week follow-up visit, the rates were 91.3% and 97.8%, respectively. No statistically significant synergistic effect was observed for the combination of a 1% lindane shampoo and oral trimethoprim/sulfamethoxazole.

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Mini Review
Management of malaria in Thailand
Udomsak Silachamroon, Srivicha Krudsood, Nanthaphorn Phophak, Sornchai Looareesuwan
Korean J Parasitol 2002;40(1):1-7.
Published online March 31, 2002
DOI: https://doi.org/10.3347/kjp.2002.40.1.1

The purpose of treatment for uncomplicated malaria is to produce a radical cure using the combination of: artesunate (4 mg/kg/day) plus mefloquine (8 mg/kg day) for 3 days; a fixed dose of artemether and lumefantrine (20/120 mg tablet) named Coartem® (4 tablets twice a day for three days for adults weighing more than 35 kg); quinine 10 mg/kg 8-hourly plus tetracycline 250 mg 6-hourly for 7 days (or doxycycline 200 mg as an alternative to tetracycline once a day for 7 days) in patients aged 8 years and over; Malarone® (in adult 4 tablets daily for 3 days). In treating severe malaria, early diagnosis and treatment with a potent antimalarial drug is recommended to save the patient's life. The antimalarial drugs of choice are: intravenous quinine or a parenteral form of an artemisinin derivative (artesunate i.v./i.m. for 2.4 mg/kg followed by 1.2 mg/kg injection at 12 and 24 hr and then daily for 5 days; artemether i.m. 3.2 mg/kg injection followed by 1.6 mg/kg at 12 and 24 hrs and then daily for 5 days; arteether i.m. (Artemotil®) with the same dose of artemether or artesunate suppository (5 mg/kg) given rectally 12 hourly for 3 days. Oral artemisinin derivatives (artesunate, artemether, and dihydroartemisinin with 4 mg/kg/day) could replace parenteral forms when patients can tolerate oral medication. Oral mefloquine (25 mg/kg divided into two doses 8 hrs apart) should be given at the end of the artemisinin treatment course to reduce recrudescence.

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