{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,29]],"date-time":"2026-09-29T11:15:15Z","timestamp":1790680515035,"version":"4.1.0"},"reference-count":37,"publisher":"MDPI AG","issue":"11","license":[{"start":{"date-parts":[[2020,11,13]],"date-time":"2020-11-13T00:00:00Z","timestamp":1605225600000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100007180","name":"Faculty of Science, Mahidol University","doi-asserted-by":"publisher","award":["Not Applicable"],"award-info":[{"award-number":["Not Applicable"]}],"id":[{"id":"10.13039\/501100007180","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Diagnostics"],"abstract":"<jats:p>The loop-mediated isothermal amplification coupled with lateral flow dipstick (PfSNP-LAMP-LFD) was recently developed to detect single nucleotide polymorphism (AAT \u2192 ATT), corresponding to substitution of asparagine to isoleucine at amino acid position 51 in the P. falciparumdhfr-ts gene associated with antifolate resistance. In this present study, the PfSNP-LAMP-LFD was validated on 128 clinical malaria samples of broad ranged parasite densities (10 to 87,634 parasites per microliter of blood). The results showed 100% accuracy for the detection of single nucleotide polymorphism for N51I mutation. Indeed, the high prevalence of N51I in the Pfdhfr-ts gene detected in the clinical samples is in line with reports of widespread antifolate resistant P. falciparum in Thailand. The relationship between enzyme choice and reaction time was observed to have an effect on PfSNP-LAMP-LFD specificity; however, the method yielded consistent results once the conditions have been optimized. The results demonstrate that PfSNP-LAMP-LFD is a simple method with sufficient sensitivity and specificity to be deployed in routine surveillance of antifolate resistance molecular marker and inform antimalarial management policy.<\/jats:p>","DOI":"10.3390\/diagnostics10110948","type":"journal-article","created":{"date-parts":[[2020,11,13]],"date-time":"2020-11-13T08:44:02Z","timestamp":1605257042000},"page":"948","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":9,"title":["Validation of PfSNP-LAMP-Lateral Flow Dipstick for Detection of Single Nucleotide Polymorphism Associated with Pyrimethamine Resistance in Plasmodium falciparum"],"prefix":"10.3390","volume":"10","author":[{"given":"Suganya","family":"Yongkiettrakul","sequence":"first","affiliation":[{"name":"National Center for Genetic Engineering and Biotechnology (BIOTEC), National Science and Technology Development Agency (NSTDA), Pathum Thani 12120, Thailand"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Fassou Ren\u00e9","family":"Koli\u00e9","sequence":"additional","affiliation":[{"name":"Department of Microbiology, Faculty of Science, Mahidol University, Bangkok 10400, Thailand"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Darin","family":"Kongkasuriyachai","sequence":"additional","affiliation":[{"name":"National Center for Genetic Engineering and Biotechnology (BIOTEC), National Science and Technology Development Agency (NSTDA), Pathum Thani 12120, Thailand"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-3938-4588","authenticated-orcid":false,"given":"Jetsumon","family":"Sattabongkot","sequence":"additional","affiliation":[{"name":"Mahidol Vivax Research Unit, Faculty of Tropical Medicine, Mahidol University, Bangkok 10400, Thailand"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Wang","family":"Nguitragool","sequence":"additional","affiliation":[{"name":"Department of Molecular Tropical Medicine and Genetics, Faculty of Tropical Medicine, Mahidol University, Bangkok 10400, Thailand"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Namfon","family":"Nawattanapaibool","sequence":"additional","affiliation":[{"name":"Department of Microbiology, Faculty of Science, Mahidol University, Bangkok 10400, Thailand"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Chayanut","family":"Suansomjit","sequence":"additional","affiliation":[{"name":"Mahidol Vivax Research Unit, Faculty of Tropical Medicine, Mahidol University, Bangkok 10400, Thailand"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Saradee","family":"Warit","sequence":"additional","affiliation":[{"name":"National Center for Genetic Engineering and Biotechnology (BIOTEC), National Science and Technology Development Agency (NSTDA), Pathum Thani 12120, Thailand"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Niwat","family":"Kangwanrangsan","sequence":"additional","affiliation":[{"name":"Department of Pathobiology, Faculty of Science, Mahidol University, Bangkok 10400, Thailand"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Sureemas","family":"Buates","sequence":"additional","affiliation":[{"name":"Department of Microbiology, Faculty of Science, Mahidol University, Bangkok 10400, Thailand"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2020,11,13]]},"reference":[{"key":"ref_1","unstructured":"WHO (2019). World Malaria Report 2019, World Health Organization."},{"key":"ref_2","unstructured":"WHO (2013). WHO Seasonal Malaria Chemoprevention with Sulfadoxine\u2013Pyrimethamine plus Amodiaquine in Children: A Field Guide. August 2013, World Health Organization."},{"key":"ref_3","unstructured":"WHO (2014). WHO Policy Brief for the Implementation of Intermittent Preventive Treatment of Malaria in Pregnancy Using Sulfadoxine-Pyrimethamine (IPTp-SP), April 2013 (rev. January 2014), World health Organization."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"8360","DOI":"10.1073\/pnas.84.23.8360","article-title":"Molecular cloning and sequence analysis of the Plasmodium falciparum dihydrofolate reductase-thymidylate synthase gene","volume":"84","author":"Bzik","year":"1987","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"7149","DOI":"10.1073\/pnas.91.15.7149","article-title":"Primary structure and expression of the dihydropteroate synthetase gene of Plasmodium falciparum","volume":"91","author":"Triglia","year":"1994","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"9109","DOI":"10.1073\/pnas.85.23.9109","article-title":"Amino acid changes linked to pyrimethamine resistance in the dihydrofolate reductase-thymidylate synthase gene of Plasmodium falciparum","volume":"85","author":"Cowman","year":"1988","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"9114","DOI":"10.1073\/pnas.85.23.9114","article-title":"Evidence that a point mutation in dihydrofolate reductase-thymidylate synthase confers resistance to pyrimethamine in falciparum malaria","volume":"85","author":"Peterson","year":"1988","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"177","DOI":"10.1016\/0035-9203(90)90246-B","article-title":"The epidemiology of drug-resistant malaria","volume":"84","author":"Bjorkman","year":"1990","journal-title":"Trans. R. Soc. Trop. Med. Hyg."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"3018","DOI":"10.1073\/pnas.87.8.3018","article-title":"Molecular basis of differential resistance to cycloguanil and pyrimethamine in Plasmodium falciparum malaria","volume":"87","author":"Peterson","year":"1990","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"1590","DOI":"10.1086\/514159","article-title":"Mutations in Plasmodium falciparum dihydrofolate reductase and dihydropteroate synthase and epidemiologic patterns of pyrimethamine-sulfadoxine use and resistance","volume":"176","author":"Plowe","year":"1997","journal-title":"J. Infect. Dis."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"2738","DOI":"10.1021\/jm0009181","article-title":"Development of a lead inhibitor for the A16V+S108T mutant of dihydrofolate reductase from the cycloguanil-resistant strain (T9\/94) of Plasmodium falciparum","volume":"43","author":"Yuthavong","year":"2000","journal-title":"J. Med. Chem."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"209","DOI":"10.1016\/S1473-3099(02)00239-6","article-title":"Epidemiology of drug-resistant malaria","volume":"2","author":"Wongsrichanalai","year":"2002","journal-title":"Lancet Infect. Dis."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"1526","DOI":"10.1093\/molbev\/msg162","article-title":"A selective sweep driven by pyrimethamine treatment in southeast asian malaria parasites","volume":"20","author":"Nair","year":"2003","journal-title":"Mol. Biol. Evol."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"1124","DOI":"10.1126\/science.1098876","article-title":"Intercontinental spread of pyrimethamine\u2014Resistant malaria","volume":"305","author":"Roper","year":"2004","journal-title":"Science"},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"389","DOI":"10.1016\/S1368-7646(98)80014-9","article-title":"P. falciparum dihydrofolate reductase and dihydropteroate synthase mutations: Epidemiology and role in clinical resistance to antifolates","volume":"1","author":"Plowe","year":"1998","journal-title":"Drug Resist. Updat."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"E63","DOI":"10.1093\/nar\/28.12.e63","article-title":"Loop-mediated isothermal amplification of DNA","volume":"28","author":"Notomi","year":"2000","journal-title":"Nucleic Acids Res."},{"key":"ref_17","unstructured":"(2020, November 06). Eiken Chemical Co., Ltd.. Available online: https:\/\/www.eiken.co.jp\/en\/products\/lamp\/."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"303","DOI":"10.1373\/clinchem.2005.057901","article-title":"Sensitive and inexpensive molecular test for falciparum malaria: Detecting Plasmodium falciparum DNA directly from heat-treated blood by loop-mediated isothermal amplification","volume":"52","author":"Poon","year":"2006","journal-title":"Clin. Chem."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"2521","DOI":"10.1128\/JCM.02117-06","article-title":"Detection of four Plasmodium species by genus and species-specific loop-mediated isothermal amplification for clinical diagnosis","volume":"45","author":"Han","year":"2007","journal-title":"J. Clin. Microbiol."},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"414","DOI":"10.1016\/j.parint.2010.05.008","article-title":"Development of a reverse transcription-loop-mediated isothermal amplification (RT-LAMP) for clinical detection of Plasmodium falciparum gametocytes","volume":"59","author":"Buates","year":"2010","journal-title":"Parasitol. Int."},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"777","DOI":"10.1016\/j.parint.2014.06.004","article-title":"Application of loop-mediated isothermal amplification assay combined with lateral flow dipstick for detection of Plasmodium falciparum and Plasmodium vivax","volume":"63","author":"Yongkiettrakul","year":"2014","journal-title":"Parasitol. Int."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"3314","DOI":"10.1021\/ac504387c","article-title":"Robust strand exchange reactions for the sequence-specific, real-time detection of nucleic acid amplicons","volume":"87","author":"Jiang","year":"2015","journal-title":"Anal. Chem."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"177","DOI":"10.1016\/j.bios.2018.03.008","article-title":"Polymorphism genotyping based on loop-mediated isothermal amplification and smartphone detection","volume":"109","author":"Yamanaka","year":"2018","journal-title":"Biosens. Bioelectron."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"227","DOI":"10.1186\/1475-2875-11-227","article-title":"Development of an allele-specific, loop-mediated, isothermal amplification method (AS-LAMP) to detect the L1014F kdr-w mutation in Anopheles gambiae sl.","volume":"11","author":"Badolo","year":"2012","journal-title":"Malar. J."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"3723","DOI":"10.7150\/thno.33980","article-title":"One-step colorimetric genotyping of single nucleotide polymorphism using probe-enhanced loop-mediated isothermal amplification (PE-LAMP)","volume":"9","author":"Ding","year":"2019","journal-title":"Theranostics"},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"964","DOI":"10.1016\/j.parint.2016.10.024","article-title":"Simple detection of single nucleotide polymorphism in Plasmodium falciparum by SNP-LAMP assay combined with lateral flow dipstick","volume":"66","author":"Yongkiettrakul","year":"2017","journal-title":"Parasitol. Int."},{"key":"ref_27","doi-asserted-by":"crossref","unstructured":"Itonaga, M., Matsuzaki, I., Warigaya, K., Tamura, T., Shimizu, Y., Fujimoto, M., Fumiyoshi, K., Masao, I., and Murata, S. (2016). Novel methodology for rapid detection of KRAS mutation using PNA-LNA mediated loop-mediated isothermal amplification. PLoS ONE, 11.","DOI":"10.1371\/journal.pone.0151654"},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"98","DOI":"10.1186\/s12936-015-0712-5","article-title":"Microgeography and molecular epidemiology of malaria at the Thailand-Myanmar border in the malaria pre-elimination phase","volume":"14","author":"Parker","year":"2015","journal-title":"Malar. J."},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"91","DOI":"10.1016\/S1383-5769(97)00013-5","article-title":"Identification of the four species of human malaria parasites by nested PCR that targets variant sequences in the small subunit rRNA gene","volume":"46","author":"Kimura","year":"1997","journal-title":"Parasitol. Int."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"737","DOI":"10.1046\/j.1365-3156.2000.00632.x","article-title":"Prevalence of point mutations in the dihydrofolate reductase and dihydropteroate synthetase genes of Plasmodium falciparum isolates from India and Thailand: A molecular epidemiologic study","volume":"5","author":"Biswas","year":"2001","journal-title":"Trop. Med. Int. Health"},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"107","DOI":"10.1186\/s12936-020-03176-x","article-title":"Molecular characterization of Plasmodium falciparum antifolate resistance markers in Thailand between 2008 and 2016","volume":"19","author":"Sugaram","year":"2020","journal-title":"Malar. J."},{"key":"ref_32","doi-asserted-by":"crossref","unstructured":"Breman, J.G., Alilio, M.S., and White, N.J. (2007). A review of malaria diagnostic tools: Microscopy and rapid diagnostic test (RDT). Defining and Defeating the Intolerable Burden of Malaria III: Progress and Perspectives, American Society of Tropical Medicine and Hygiene. Supplement to Volume 77(6) of AJTMH.","DOI":"10.4269\/ajtmh.2007.77.i"},{"key":"ref_33","unstructured":"WHO (2014). Policy Brief on Malaria Diagnostics in Low-Transmission Settings, World Health Organization."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"S86","DOI":"10.1038\/nature16039","article-title":"Comparison of diagnostics for the detection of asymptomatic Plasmodium falciparum infections to inform control and elimination strategies","volume":"528","author":"Wu","year":"2015","journal-title":"Nature"},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"329","DOI":"10.1186\/s12936-018-2476-1","article-title":"Prevalence of asymptomatic Plasmodium infections with sub-microscopic parasite densities in the northwestern border of Thailand: A potential threat to malaria elimination","volume":"17","author":"Sattabongkot","year":"2018","journal-title":"Malar. J."},{"key":"ref_36","doi-asserted-by":"crossref","unstructured":"Chahar, M., Anvikar, A., and Valecha, N. (2019). Development and evaluation of a novel HNB based isothermal amplification assay for fast detection of Pyrimethamine resistance (S108N) in Plasmodium falciparum. Int. J. Environ. Res. Public Health, 16.","DOI":"10.3390\/ijerph16091635"},{"key":"ref_37","doi-asserted-by":"crossref","unstructured":"Mohon, A.N., Menard, D., Alam, M.S., Perera, K., and Pillai, D.R. (2018). A novel single-nucleotide polymorphism Loop Mediated Isothermal Amplification assay for detection of Artemisinin-resistant Plasmodium falciparum malaria. Open Forum Infect. Dis., 5.","DOI":"10.1093\/ofid\/ofy011"}],"container-title":["Diagnostics"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2075-4418\/10\/11\/948\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,11]],"date-time":"2025-10-11T10:33:07Z","timestamp":1760178787000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2075-4418\/10\/11\/948"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2020,11,13]]},"references-count":37,"journal-issue":{"issue":"11","published-online":{"date-parts":[[2020,11]]}},"alternative-id":["diagnostics10110948"],"URL":"https:\/\/doi.org\/10.3390\/diagnostics10110948","relation":{},"ISSN":["2075-4418"],"issn-type":[{"value":"2075-4418","type":"electronic"}],"subject":[],"published":{"date-parts":[[2020,11,13]]}}}