{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,1,16]],"date-time":"2026-01-16T18:42:04Z","timestamp":1768588924077,"version":"3.49.0"},"reference-count":29,"publisher":"MDPI AG","issue":"11","license":[{"start":{"date-parts":[[2022,10,25]],"date-time":"2022-10-25T00:00:00Z","timestamp":1666656000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"Department of Biotechnology, Govt. of India","award":["BT\/RLF\/Re-entry\/09\/2017"],"award-info":[{"award-number":["BT\/RLF\/Re-entry\/09\/2017"]}]},{"name":"Department of Biotechnology, Govt. of India","award":["NU\/IFC\/ENT\/01\/007"],"award-info":[{"award-number":["NU\/IFC\/ENT\/01\/007"]}]},{"name":"Department of Biotechnology, Govt. of India","award":["2018R1A6A1A03025124"],"award-info":[{"award-number":["2018R1A6A1A03025124"]}]},{"name":"Department of Biotechnology, Govt. of India","award":["HV20C0142"],"award-info":[{"award-number":["HV20C0142"]}]},{"name":"Ministry Agency for Research and Innovation, Ministry of Education, Saudi Arabia and Najran University, Najran, Saudi Arabia","award":["BT\/RLF\/Re-entry\/09\/2017"],"award-info":[{"award-number":["BT\/RLF\/Re-entry\/09\/2017"]}]},{"name":"Ministry Agency for Research and Innovation, Ministry of Education, Saudi Arabia and Najran University, Najran, Saudi Arabia","award":["NU\/IFC\/ENT\/01\/007"],"award-info":[{"award-number":["NU\/IFC\/ENT\/01\/007"]}]},{"name":"Ministry Agency for Research and Innovation, Ministry of Education, Saudi Arabia and Najran University, Najran, Saudi Arabia","award":["2018R1A6A1A03025124"],"award-info":[{"award-number":["2018R1A6A1A03025124"]}]},{"name":"Ministry Agency for Research and Innovation, Ministry of Education, Saudi Arabia and Najran University, Najran, Saudi Arabia","award":["HV20C0142"],"award-info":[{"award-number":["HV20C0142"]}]},{"name":"National Research Foundation of Korea (NRF)","award":["BT\/RLF\/Re-entry\/09\/2017"],"award-info":[{"award-number":["BT\/RLF\/Re-entry\/09\/2017"]}]},{"name":"National Research Foundation of Korea (NRF)","award":["NU\/IFC\/ENT\/01\/007"],"award-info":[{"award-number":["NU\/IFC\/ENT\/01\/007"]}]},{"name":"National Research Foundation of Korea (NRF)","award":["2018R1A6A1A03025124"],"award-info":[{"award-number":["2018R1A6A1A03025124"]}]},{"name":"National Research Foundation of Korea (NRF)","award":["HV20C0142"],"award-info":[{"award-number":["HV20C0142"]}]},{"name":"Ministry of Health &amp; Welfare, Republic of Korea","award":["BT\/RLF\/Re-entry\/09\/2017"],"award-info":[{"award-number":["BT\/RLF\/Re-entry\/09\/2017"]}]},{"name":"Ministry of Health &amp; Welfare, Republic of Korea","award":["NU\/IFC\/ENT\/01\/007"],"award-info":[{"award-number":["NU\/IFC\/ENT\/01\/007"]}]},{"name":"Ministry of Health &amp; Welfare, Republic of Korea","award":["2018R1A6A1A03025124"],"award-info":[{"award-number":["2018R1A6A1A03025124"]}]},{"name":"Ministry of Health &amp; Welfare, Republic of Korea","award":["HV20C0142"],"award-info":[{"award-number":["HV20C0142"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Genes"],"abstract":"<jats:p>The simian malaria parasite Plasmodium knowlesi causes a high number of zoonotic infections in Malaysia. The thrombospondin-related apical merozoite protein (TRAMP) is an essential ligand for binding to the erythrocyte cell surface, whereby it facilitates the invasion. This study is the first attempt to determine the genetic diversity, phylogeography, natural selection and population structure from 97 full-length PkTRAMP gene sequences originating from Malaysia. We found low levels of nucleotide diversity (\u03c0~0.0065) for the full-length gene despite samples originating from geographically separated regions (i.e., Peninsular Malaysia and Malaysian Borneo). The rate of synonymous substitutions was significantly higher than that of non-synonymous substitutions, indicating a purifying selection for the full-length gene within the clinical samples. The population genetic analysis revealed that the parasite population is undergoing a significant population expansion. The analysis of the amino acid sequence alignment of 97 PkTRAMP sequences identified 15 haplotypes, of which a major shared haplotype was noted Hap 1 (n = 68, Sarawak; n = 34, Sabah; n = 12, Peninsular Malaysia; n = 22). The phylogenetic analysis using DNA sequences identified two clusters that separated due to geographical distance and three mixed clusters with samples from both Peninsular Malaysia and Malaysian Borneo. Population structure analyses indicated two distinct sub-populations (K = 2). Our findings point to the potential for independent parasite evolution, which could make zoonotic malaria control and elimination even more challenging.<\/jats:p>","DOI":"10.3390\/genes13111944","type":"journal-article","created":{"date-parts":[[2022,10,25]],"date-time":"2022-10-25T22:00:27Z","timestamp":1666735227000},"page":"1944","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":4,"title":["Genetic Diversity and Population Genetic Structure Analysis of Plasmodium knowlesi Thrombospondin-Related Apical Merozoite Protein (TRAMP) in Clinical Samples"],"prefix":"10.3390","volume":"13","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-3407-7220","authenticated-orcid":false,"given":"Md Atique","family":"Ahmed","sequence":"first","affiliation":[{"name":"ICMR-Regional Medical Research Centre, NER, Dibrugarh, Assam, Bhubaneswar 786010, India"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-7993-3174","authenticated-orcid":false,"given":"Rehan Haider","family":"Zaidi","sequence":"additional","affiliation":[{"name":"Department of Biotechnology and Microbiology, National College, Tiruchirapalli 620001, India"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Gauspasha Yusuf","family":"Deshmukh","sequence":"additional","affiliation":[{"name":"Department of Biotechnology and Microbiology, National College, Tiruchirapalli 620001, India"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-9614-3238","authenticated-orcid":false,"given":"Ahmed","family":"Saif","sequence":"additional","affiliation":[{"name":"Department of Clinical Laboratory Sciences, Faculty of Applied Medical Sciences, Najran University, Narjan 55461, Saudi Arabia"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Mohammed Abdulrahman","family":"Alshahrani","sequence":"additional","affiliation":[{"name":"Department of Clinical Laboratory Sciences, Faculty of Applied Medical Sciences, Najran University, Narjan 55461, Saudi Arabia"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Syeda Sabiha","family":"Salam","sequence":"additional","affiliation":[{"name":"Department of Life Sciences, Dibrugarh University, Assam, Dibrugarh 786004, India"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-8231-2902","authenticated-orcid":false,"given":"Mohammed Mohieldien Abbas","family":"Elfaki","sequence":"additional","affiliation":[{"name":"Department of Microbiology and Parasitology, Faculty of Medicine, Jazan University, Jizan 45142, Saudi Arabia"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Jin-Hee","family":"Han","sequence":"additional","affiliation":[{"name":"Department of Medical Environmental Biology and Tropical Medicine, School of Medicine, Kangwon National University, Chuncheon 24341, Korea"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-0651-2739","authenticated-orcid":false,"given":"Saurav Jyoti","family":"Patgiri","sequence":"additional","affiliation":[{"name":"ICMR-Regional Medical Research Centre, NER, Dibrugarh, Assam, Bhubaneswar 786010, India"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-0419-9339","authenticated-orcid":false,"given":"Fu-Shi","family":"Quan","sequence":"additional","affiliation":[{"name":"Medical Research Center for Bioreaction to Reactive Oxygen Species and Biomedical Science Institute, School of Medicine, Graduate School, Kyung Hee University, Seoul 02447, Korea"},{"name":"Department of Medical Zoology, School of Medicine, Kyung Hee University, Seoul 02447, Korea"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2022,10,25]]},"reference":[{"key":"ref_1","unstructured":"World Health Organization (2022, June 23). World Malaria Report 2020: 20 Years of Global Progress and Challenges. Available online: www.who.int\/teams\/global-malaria-programme\/reports\/world-malaria-report-2020."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"134","DOI":"10.1111\/voxs.12115","article-title":"Plasmodium knowlesi\u2014An emerging pathogen","volume":"10","author":"Ahmed","year":"2015","journal-title":"ISBT Sci. Ser."},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"13506","DOI":"10.1074\/jbc.272.21.13506","article-title":"Characterization of simian malarial parasite (Plasmodium knowlesi)-induced putrescine transport in rhesus monkey erythrocytes. A novel putrescine conjugate arrests in vitro growth of simian malarial parasite (Plasmodium knowlesi) and cures multidrug resistant murine malaria (Plasmodium yoelii) infection in vivo","volume":"272","author":"Singh","year":"1997","journal-title":"J. Biol. Chem."},{"key":"ref_4","doi-asserted-by":"crossref","unstructured":"Ahmed, M.A., Saif, A., and Quan, F.S. (2019). Diversity pattern of Plasmodium knowlesi merozoite surface protein 4 (MSP4) in natural population of Malaysia. PLoS ONE, 14.","DOI":"10.1371\/journal.pone.0224743"},{"key":"ref_5","first-page":"1","article-title":"Diversity and natural selection on the thrombospondin-related adhesive protein (TRAP) gene of Plasmodium knowlesi in Malaysia","volume":"17","author":"Ahmed","year":"2018","journal-title":"Malar. J."},{"key":"ref_6","doi-asserted-by":"crossref","unstructured":"Pinheiro, M.M., Ahmed, M.A., Millar, S.B., Sanderson, T., Otto, T.D., Lu, W.C., Krishna, S., Rayner, J.C., and Cox-Singh, J. (2015). Plasmodium knowlesi genome sequences from clinical isolates reveal extensive genomic dimorphism. PLoS ONE, 10.","DOI":"10.1371\/journal.pone.0121303"},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"531","DOI":"10.1073\/pnas.1216457110","article-title":"Adaptation of the genetically tractable malaria pathogen Plasmodium knowlesi to continuous culture in human erythrocytes","volume":"110","author":"Moon","year":"2013","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"8","DOI":"10.1186\/1475-2875-12-8","article-title":"Limitations of microscopy to differentiate Plasmodium species in a region co-endemic for Plasmodium falciparum, Plasmodium vivax and Plasmodium knowlesi","volume":"12","author":"Barber","year":"2013","journal-title":"Malar. J."},{"key":"ref_9","first-page":"31","article-title":"Severe Plasmodium knowlesi infection with multiorgan involvement in north east peninsular Malaysia","volume":"31","author":"Azidah","year":"2014","journal-title":"Trop. Biomed."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"502","DOI":"10.1128\/iai.28.2.502-507.1980","article-title":"Alterations in the distribution and proliferative responses of rhesus monkey peripheral blood and spleen cells during malaria (Plasmodium knowlesi) infection","volume":"28","author":"Taylor","year":"1980","journal-title":"Infect. Immun."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"6","DOI":"10.1017\/S0031182016001888","article-title":"The role of Plasmodium knowlesi in the history of malaria research","volume":"145","author":"Butcher","year":"2018","journal-title":"Parasitology"},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"225","DOI":"10.1016\/j.molbiopara.2003.12.003","article-title":"PTRAMP; a conserved Plasmodium thrombospondin-related apical merozoite protein","volume":"134","author":"Thompson","year":"2004","journal-title":"Mol. Biochem. Parasitol."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"5195","DOI":"10.1093\/emboj\/18.19.5195","article-title":"The A-domain and the thrombospondin-related motif of Plasmodium falciparum TRAP are implicated in the invasion process of mosquito salivary glands","volume":"18","author":"Wengelnik","year":"1999","journal-title":"EMBO J."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"39","DOI":"10.1016\/j.meegid.2017.06.019","article-title":"Genetic diversity in the C-terminus of merozoite surface protein 1 among Plasmodium knowlesi isolates from Selangor and Sabah Borneo, Malaysia","volume":"54","author":"Yap","year":"2017","journal-title":"Infect. Genet. Evol."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"283","DOI":"10.1186\/1475-2875-9-283","article-title":"Identification and characterization of the Plasmodium vivax thrombospondin-related apical merozoite protein","volume":"9","author":"Mongui","year":"2010","journal-title":"Malar. J."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"1341","DOI":"10.1111\/cmi.12118","article-title":"A thrombospondin structural repeat containing rhoptry protein from Plasmodium falciparum mediates erythrocyte invasion","volume":"15","author":"Siddiqui","year":"2013","journal-title":"Cell Microbiol."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"114","DOI":"10.1016\/j.molbiopara.2006.06.010","article-title":"Plasmodium thrombospondin related apical merozoite protein (PTRAMP) is shed from the surface of merozoites by PfSUB2 upon invasion of erythrocytes","volume":"150","author":"Green","year":"2006","journal-title":"Mol. Biochem. Parasitol."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"785","DOI":"10.1038\/nmeth.1701","article-title":"SignalP 4.0: Discriminating signal peptides from transmembrane regions","volume":"8","author":"Petersen","year":"2011","journal-title":"Nat. Methods"},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"2731","DOI":"10.1093\/molbev\/msr121","article-title":"MEGA5: Molecular evolutionary genetics analysis using maximum likelihood, evolutionary distance, and maximum parsimony methods","volume":"28","author":"Tamura","year":"2011","journal-title":"Mol. Biol. Evol."},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"1451","DOI":"10.1093\/bioinformatics\/btp187","article-title":"DnaSP v5: A software for comprehensive analysis of DNA polymorphism data","volume":"25","author":"Librado","year":"2009","journal-title":"Bioinformatics"},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"945","DOI":"10.1093\/genetics\/155.2.945","article-title":"Inference of population structure using multilocus genotype data","volume":"155","author":"Pritchard","year":"2000","journal-title":"Genetics"},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"359","DOI":"10.1007\/s12686-011-9548-7","article-title":"STRUCTURE HARVESTER: A website and program for visualizing STRUCTURE output and implementing the Evanno method","volume":"4","author":"Earl","year":"2012","journal-title":"Conserv. Genet. Resour."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"564","DOI":"10.1111\/j.1755-0998.2010.02847.x","article-title":"Arlequin suite ver 3.5: A new series of programs to perform population genetics analyses under Linux and Windows","volume":"10","author":"Excoffier","year":"2010","journal-title":"Mol. Ecol. Resour."},{"key":"ref_24","doi-asserted-by":"crossref","unstructured":"Ahmed, A.M., Pinheiro, M.M., Divis, P.C., Siner, A., Zainudin, R., Wong, I.T., Lu, C.W., Singh-Khaira, S.K., Millar, S.B., and Lynch, S. (2014). Disease progression in Plasmodium knowlesi malaria is linked to variation in invasion gene family members. PLoS Negl. Trop. Dis., 8.","DOI":"10.1371\/journal.pntd.0003086"},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"e6141","DOI":"10.7717\/peerj.6141","article-title":"The Plasmodium knowlesi Pk41 surface protein diversity, natural selection, sub population and geographical clustering: A 6-cysteine protein family member","volume":"6","author":"Ahmed","year":"2018","journal-title":"Peer J."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"442","DOI":"10.1186\/s12936-018-2583-z","article-title":"Within-population genetic diversity and population structure of Plasmodium knowlesi merozoite surface protein 1 gene from geographically distinct regions of Malaysia and Thailand","volume":"17","author":"Ahmed","year":"2018","journal-title":"Malar. J."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"241","DOI":"10.1186\/s12936-016-1294-6","article-title":"Clustering and genetic differentiation of the normocyte binding protein (nbpxa) of Plasmodium knowlesi clinical isolates from Peninsular Malaysia and Malaysia Borneo","volume":"15","author":"Ahmed","year":"2016","journal-title":"Malar. J."},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"5781","DOI":"10.1038\/s41598-018-23728-1","article-title":"Cross-species analysis of apical asparagine-rich protein of Plasmodium vivax and Plasmodium knowlesi","volume":"8","author":"Muh","year":"2018","journal-title":"Sci. Rep."},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"1371","DOI":"10.3201\/eid2208.151885","article-title":"Phylogeographic Evidence for 2 Genetically Distinct Zoonotic Plasmodium knowlesi Parasites, Malaysia","volume":"22","author":"Yusof","year":"2016","journal-title":"Emerg. Infect. Dis."}],"container-title":["Genes"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2073-4425\/13\/11\/1944\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,11]],"date-time":"2025-10-11T01:02:35Z","timestamp":1760144555000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2073-4425\/13\/11\/1944"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2022,10,25]]},"references-count":29,"journal-issue":{"issue":"11","published-online":{"date-parts":[[2022,11]]}},"alternative-id":["genes13111944"],"URL":"https:\/\/doi.org\/10.3390\/genes13111944","relation":{},"ISSN":["2073-4425"],"issn-type":[{"value":"2073-4425","type":"electronic"}],"subject":[],"published":{"date-parts":[[2022,10,25]]}}}