{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,30]],"date-time":"2026-09-30T22:57:45Z","timestamp":1790809065781,"version":"4.1.0"},"update-to":[{"DOI":"10.1371\/journal.pntd.0010760","type":"new_version","label":"New version","source":"publisher","updated":{"date-parts":[[2022,9,23]],"date-time":"2022-09-23T00:00:00Z","timestamp":1663891200000}}],"reference-count":32,"publisher":"Public Library of Science (PLoS)","issue":"9","license":[{"start":{"date-parts":[[2022,9,13]],"date-time":"2022-09-13T00:00:00Z","timestamp":1663027200000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100001711","name":"Swiss National Science Foundation","doi-asserted-by":"crossref","award":["310030-134889"],"award-info":[{"award-number":["310030-134889"]}],"id":[{"id":"10.13039\/501100001711","id-type":"DOI","asserted-by":"crossref"}]},{"DOI":"10.13039\/501100001711","name":"Swiss National Science Foundation","doi-asserted-by":"crossref","award":["310030-159580"],"award-info":[{"award-number":["310030-159580"]}],"id":[{"id":"10.13039\/501100001711","id-type":"DOI","asserted-by":"crossref"}]},{"name":"NIH NIAID","award":["U19 AI089686"],"award-info":[{"award-number":["U19 AI089686"]}]},{"name":"NIH NIAID","award":["U19 AI129392-01"],"award-info":[{"award-number":["U19 AI129392-01"]}]},{"DOI":"10.13039\/100000865","name":"Bill and Melinda Gates Foundation","doi-asserted-by":"publisher","award":["TransEpi Consortium"],"award-info":[{"award-number":["TransEpi Consortium"]}],"id":[{"id":"10.13039\/100000865","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100000925","name":"National Health and Medical Research Council","doi-asserted-by":"publisher","award":["GNT11155075"],"award-info":[{"award-number":["GNT11155075"]}],"id":[{"id":"10.13039\/501100000925","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100000925","name":"National Health and Medical Research Council","doi-asserted-by":"publisher","award":["GNT1161627"],"award-info":[{"award-number":["GNT1161627"]}],"id":[{"id":"10.13039\/501100000925","id-type":"DOI","asserted-by":"publisher"}]},{"name":"Agence Nationale pour le Recherche","award":["ANR-10-LABX-62-IBEID"],"award-info":[{"award-number":["ANR-10-LABX-62-IBEID"]}]},{"name":"Agence Nationale pour la Recherche","award":["ANR-16-CONV-0005"],"award-info":[{"award-number":["ANR-16-CONV-0005"]}]}],"content-domain":{"domain":["www.plosntds.org"],"crossmark-restriction":false},"short-container-title":["PLoS Negl Trop Dis"],"abstract":"<jats:sec id=\"sec001\">\n<jats:title>Background<\/jats:title>\n<jats:p>Co-infection of the four major species of human malaria parasite <jats:italic>Plasmodium falciparum<\/jats:italic> (<jats:italic>Pf<\/jats:italic>), <jats:italic>P<\/jats:italic>. <jats:italic>vivax<\/jats:italic> (<jats:italic>Pv<\/jats:italic>), <jats:italic>P<\/jats:italic>. <jats:italic>malariae<\/jats:italic> (<jats:italic>Pm<\/jats:italic>), and <jats:italic>P<\/jats:italic>. <jats:italic>ovale<\/jats:italic> sp. (<jats:italic>Po<\/jats:italic>) is regularly observed, but there is limited understanding of between-species interactions. In particular, little is known about the effects of multiple <jats:italic>Plasmodium<\/jats:italic> species co-infections on gametocyte production.<\/jats:p>\n<\/jats:sec>\n<jats:sec id=\"sec002\">\n<jats:title>Methods<\/jats:title>\n<jats:p>We developed molecular assays for detecting asexual and gametocyte stages of <jats:italic>Pf<\/jats:italic>, <jats:italic>Pv<\/jats:italic>, <jats:italic>Pm<\/jats:italic>, and <jats:italic>Po<\/jats:italic>. This is the first description of molecular diagnostics for <jats:italic>Pm<\/jats:italic> and <jats:italic>Po<\/jats:italic> gametocytes. These assays were implemented in a unique epidemiological setting in Papua New Guinea with sympatric transmission of all four <jats:italic>Plasmodium<\/jats:italic> species permitting a comprehensive investigation of species interactions.<\/jats:p>\n<\/jats:sec>\n<jats:sec id=\"sec003\">\n<jats:title>Findings<\/jats:title>\n<jats:p>The observed frequency of <jats:italic>Pf<\/jats:italic>-<jats:italic>Pv<\/jats:italic> co-infection for asexual parasites (14.7%) was higher than expected from individual prevalence rates (23.8%<jats:italic>Pf<\/jats:italic> x 47.4%<jats:italic>Pv<\/jats:italic> = 11.3%). The observed frequency of co-infection with <jats:italic>Pf<\/jats:italic> and <jats:italic>Pv<\/jats:italic> gametocytes (4.6%) was higher than expected from individual prevalence rates (13.1%<jats:italic>Pf<\/jats:italic> x 28.2%<jats:italic>Pv<\/jats:italic> = 3.7%). The excess risk of co-infection was 1.38 (95% confidence interval (CI): 1.09, 1.67) for all parasites and 1.37 (95% CI: 0.95, 1.79) for gametocytes. This excess co-infection risk was partially attributable to malaria infections clustering in some villages. <jats:italic>Pf-Pv-Pm<\/jats:italic> triple infections were four times more frequent than expected by chance alone, which could not be fully explained by infections clustering in highly exposed individuals. The effect of co-infection on parasite density was analyzed by systematic comparison of all pairwise interactions. This revealed a significant 6.57-fold increase of <jats:italic>Pm<\/jats:italic> density when co-infected with <jats:italic>Pf<\/jats:italic>. <jats:italic>Pm<\/jats:italic> gametocytemia also increased with <jats:italic>Pf<\/jats:italic> co-infection.<\/jats:p>\n<\/jats:sec>\n<jats:sec id=\"sec004\">\n<jats:title>Conclusions<\/jats:title>\n<jats:p>Heterogeneity in exposure to mosquitoes is a key epidemiological driver of <jats:italic>Plasmodium<\/jats:italic> co-infection. Among the four co-circulating parasites, <jats:italic>Pm<\/jats:italic> benefitted most from co-infection with other species. Beyond this, no general prevailing pattern of suppression or facilitation was identified in pairwise analysis of gametocytemia and parasitemia of the four species.<\/jats:p>\n<\/jats:sec>\n<jats:sec id=\"sec005\">\n<jats:title>Trial registration<\/jats:title>\n<jats:p>This trial is registered with ClinicalTrials.gov, Trial ID: <jats:ext-link xmlns:xlink=\"http:\/\/www.w3.org\/1999\/xlink\" ext-link-type=\"uri\" xlink:href=\"https:\/\/clinicaltrials.gov\/ct2\/show\/NCT02143934\" xlink:type=\"simple\">NCT02143934<\/jats:ext-link>.<\/jats:p>\n<\/jats:sec>","DOI":"10.1371\/journal.pntd.0010760","type":"journal-article","created":{"date-parts":[[2022,9,13]],"date-time":"2022-09-13T17:41:35Z","timestamp":1663090895000},"page":"e0010760","update-policy":"https:\/\/doi.org\/10.1371\/journal.pntd.corrections_policy","source":"Crossref","is-referenced-by-count":27,"title":["Co-infection of the four major Plasmodium species: Effects on densities and gametocyte carriage"],"prefix":"10.1371","volume":"16","author":[{"given":"Aurel","family":"Holzschuh","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Maria","family":"Gruenberg","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Natalie E.","family":"Hofmann","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Rahel","family":"Wampfler","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Benson","family":"Kiniboro","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Leanne J.","family":"Robinson","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ivo","family":"Mueller","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ingrid","family":"Felger","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-7472-4138","authenticated-orcid":true,"given":"Michael T.","family":"White","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"340","published-online":{"date-parts":[[2022,9,13]]},"reference":[{"key":"pntd.0010760.ref001","doi-asserted-by":"crossref","first-page":"855","DOI":"10.4269\/ajtmh.1937.s1-17.855","article-title":"Simultaneous inoculation with Plasmodium vivax and Plasmodium falciparum","volume":"17","author":"MF Boyd","year":"1937","journal-title":"Am. J. Trop. Med. Hyg"},{"key":"pntd.0010760.ref002","doi-asserted-by":"crossref","first-page":"1289","DOI":"10.2307\/4582611","article-title":"Antagonism between species of malaria parasites in induced mixed infections","volume":"53","author":"B Mayne","year":"1938","journal-title":"Public Health Reports (1896\u20131970)."},{"key":"pntd.0010760.ref003","doi-asserted-by":"crossref","first-page":"725","DOI":"10.4269\/ajtmh.1980.29.725","article-title":"A longitudinal study of human malaria in the West African Savanna in the absence of control measures: relationships between different Plasmodium species, in particular P. falciparum and P. malariae","volume":"29","author":"L Molineaux","year":"1980","journal-title":"Am. J. Trop. Med. Hyg"},{"key":"pntd.0010760.ref004","doi-asserted-by":"crossref","first-page":"12","DOI":"10.2307\/3285692","article-title":"Multispecies Plasmodium infections of humans","volume":"85","author":"FE McKenzie","year":"1999","journal-title":"J. Parasitol"},{"key":"pntd.0010760.ref005","doi-asserted-by":"crossref","first-page":"233","DOI":"10.1017\/S0031182001007272","article-title":"Methods for estimation of associations between multiple species parasite infections","volume":"122","author":"SC Howard","year":"2001","journal-title":"Parasitology"},{"key":"pntd.0010760.ref006","first-page":"33","article-title":"Systematic review and meta-analysis of the interaction between Plasmodium falciparum and Plasmodium vivax in humans.","volume":"44","author":"AA Haghdoost","year":"2007","journal-title":"J. Vector Borne Dis"},{"key":"pntd.0010760.ref007","doi-asserted-by":"crossref","first-page":"E105","DOI":"10.1086\/651587","article-title":"Mixed infections and the evolution of virulence: effects of resource competition, parasite plasticity, and impaired host immunity","volume":"175","author":"M Choisy","year":"2010","journal-title":"Am. Nat"},{"key":"pntd.0010760.ref008","doi-asserted-by":"crossref","first-page":"278","DOI":"10.1016\/j.pt.2007.04.009","article-title":"Plasmodium malariae and Plasmodium ovale\u2013the \u2018bashful\u2019 malaria parasites.","volume":"23","author":"I Mueller","year":"2007","journal-title":"Trends Parasitol"},{"key":"pntd.0010760.ref009","first-page":"45","article-title":"Persistent detection of Plasmodium falciparum, P. malariae, P. ovale curtisi and P. ovale wallikeri after ACT treatment of asymptomatic Ghanaian school-children","volume":"3","author":"B Dinko","year":"2013","journal-title":"Int. J. Parasitol.: Drugs and Drug Resistance."},{"key":"pntd.0010760.ref010","doi-asserted-by":"crossref","first-page":"350","DOI":"10.1186\/s12936-016-1409-0","article-title":"Low prevalence of Plasmodium malariae and Plasmodium ovale mono-infections among children in the Democratic Republic of the Congo: a population-based, cross-sectional study.","volume":"15","author":"SM Doctor","year":"2016","journal-title":"Malar. J"},{"key":"pntd.0010760.ref011","doi-asserted-by":"crossref","first-page":"9784","DOI":"10.1038\/s41598-019-46194-9","article-title":"Characterization of Plasmodium infections among inhabitants of rural areas in Gabon.","volume":"9","author":"TG Woldearegai","year":"2019","journal-title":"Sci Rep."},{"key":"pntd.0010760.ref012","doi-asserted-by":"crossref","first-page":"S23","DOI":"10.1017\/S003118200001698X","article-title":"Concomitant infections, parasites and immune responses","volume":"122","author":"FEG Cox","year":"2001","journal-title":"Parasitology"},{"key":"pntd.0010760.ref013","doi-asserted-by":"crossref","first-page":"522","DOI":"10.1038\/383522a0","article-title":"High incidence of malaria in \u03b1-thalassaemic children","volume":"383","author":"TN Williams","year":"1996","journal-title":"Nature"},{"key":"pntd.0010760.ref014","doi-asserted-by":"crossref","first-page":"1095","DOI":"10.1016\/S0140-6736(94)90203-8","article-title":"Mixed infections with Plasmodium falciparum and P. malariae and fever in malaria","volume":"343","author":"J Black","year":"1994","journal-title":"Lancet"},{"key":"pntd.0010760.ref015","doi-asserted-by":"crossref","first-page":"4","DOI":"10.1016\/S2666-5247(21)00009-4","article-title":"Plasmodium interspecies interactions during a period of increasing prevalence of Plasmodium ovale in symptomatic individuals seeking treatment: an observational study","volume":"2","author":"HM Akala","year":"2021","journal-title":"Lancet Microbe"},{"key":"pntd.0010760.ref016","doi-asserted-by":"crossref","first-page":"1713","DOI":"10.1086\/504330","article-title":"Fever in patients with mixed-species malaria","volume":"42","author":"FE McKenzie","year":"2006","journal-title":"Clin. Infect. Dis"},{"key":"pntd.0010760.ref017","doi-asserted-by":"crossref","first-page":"e127","DOI":"10.1371\/journal.pmed.0050127","article-title":"Plasmodium vivax and mixed infections are associated with severe malaria in children: A prospective cohort study from Papua New Guinea.","volume":"5","author":"B Genton","year":"2008","journal-title":"PLOS Med."},{"key":"pntd.0010760.ref018","doi-asserted-by":"crossref","first-page":"442","DOI":"10.4269\/ajtmh.2008.78.442","article-title":"Increased Plasmodium falciparum gametocyte production in mixed infections with P. malariae","volume":"78","author":"JT Bousema","year":"2008","journal-title":"Am, J. Trop. Med. Hyg"},{"key":"pntd.0010760.ref019","doi-asserted-by":"crossref","first-page":"e76316","DOI":"10.1371\/journal.pone.0076316","article-title":"Strategies for detection of Plasmodium species gametocytes.","volume":"8","author":"R Wampfler","year":"2013","journal-title":"PLOS One."},{"key":"pntd.0010760.ref020","doi-asserted-by":"crossref","first-page":"e23708","DOI":"10.7554\/eLife.23708","article-title":"The complex relationship of exposure to new Plasmodium infections and incidence of clinical malaria in Papua New Guinea.","volume":"6","author":"NE Hofmann","year":"2017","journal-title":"eLife"},{"key":"pntd.0010760.ref021","doi-asserted-by":"crossref","first-page":"e1001891","DOI":"10.1371\/journal.pmed.1001891","article-title":"Strategies for understanding and reducing the Plasmodium vivax and Plasmodium ovale hypnozoite reservoir in Papua New Guinean children: A randomised placebo-controlled trial and mathematical model.","volume":"12","author":"LJ Robinson","year":"2015","journal-title":"PLOS Med."},{"key":"pntd.0010760.ref022","doi-asserted-by":"crossref","first-page":"253","DOI":"10.1016\/S1471-4922(03)00091-6","article-title":"The epidemiology of malaria in Papua New Guinea","volume":"19","author":"I M\u00fcller","year":"2003","journal-title":"Trends Parasitol"},{"key":"pntd.0010760.ref023","doi-asserted-by":"crossref","first-page":"1745","DOI":"10.1111\/tmi.12616","article-title":"Prevalence of malaria across Papua New Guinea after initial roll-out of insecticide-treated mosquito nets","volume":"20","author":"MW Hetzel","year":"2015","journal-title":"Trop. Med. Int. Health"},{"key":"pntd.0010760.ref024","doi-asserted-by":"crossref","first-page":"340","DOI":"10.1186\/s13071-016-1635-x","article-title":"Changes in malaria burden and transmission in sentinel sites after the roll-out of long-lasting insecticidal nets in Papua New Guinea.","volume":"9","author":"MW Hetzel","year":"2016","journal-title":"Parasit. Vectors"},{"key":"pntd.0010760.ref025","doi-asserted-by":"crossref","first-page":"e0126747","DOI":"10.1371\/journal.pone.0126747","article-title":"Blood-stage parasitaemia and age determine Plasmodium falciparum and P. vivax gametocytaemia in Papua New Guinea.","volume":"10","author":"C Koepfli","year":"2015","journal-title":"PLOS One"},{"key":"pntd.0010760.ref026","doi-asserted-by":"crossref","first-page":"e0005753","DOI":"10.1371\/journal.pntd.0005753","article-title":"Effects of liver-stage clearance by primaquine on gametocyte carriage of Plasmodium vivax and P. falciparum.","volume":"11","author":"R Wampfler","year":"2017","journal-title":"PLOS Negl. Trop. Dis"},{"key":"pntd.0010760.ref027","doi-asserted-by":"crossref","first-page":"361","DOI":"10.1186\/1475-2875-9-361","article-title":"Comparison of diagnostic methods for the detection and quantification of the four sympatric Plasmodium species in field samples from Papua New Guinea.","volume":"9","author":"A Rosanas-Urgell","year":"2010","journal-title":"Malar. J."},{"issue":"6\u201310","key":"pntd.0010760.ref028","doi-asserted-by":"crossref","first-page":"677","DOI":"10.1016\/j.ijpara.2011.01.004","article-title":"Plasmodium ovale curtisi and Plasmodium ovale wallikeri circulate simultaneously in African communities","volume":"41","author":"MC Oguike","year":"2011","journal-title":"Int J Parasitol"},{"key":"pntd.0010760.ref029","doi-asserted-by":"crossref","first-page":"440","DOI":"10.1016\/j.pt.2004.07.004","article-title":"Why do we need to know more about mixed Plasmodium species infections in humans?","volume":"20","author":"PA Zimmerman","year":"2004","journal-title":"Trends Parasitol"},{"key":"pntd.0010760.ref030","doi-asserted-by":"crossref","first-page":"41","DOI":"10.1186\/1475-2875-8-41","article-title":"High sensitivity detection of Plasmodium species reveals positive correlations between infections of different species, shifts in age distribution and reduced local variation in Papua New Guinea.","volume":"8","author":"I Mueller","year":"2009","journal-title":"Malar. J"},{"key":"pntd.0010760.ref031","doi-asserted-by":"crossref","first-page":"1532","DOI":"10.1016\/j.cell.2017.10.020","article-title":"Lysophosphatidylcholine regulates sexual stage differentiation in the human malaria parasite Plasmodium falciparum","volume":"171","author":"NMB Brancucci","year":"2017","journal-title":"Cell"},{"key":"pntd.0010760.ref032","doi-asserted-by":"crossref","first-page":"36","DOI":"10.4269\/tropmed.1999.61-036","article-title":"A retrospective examination of sporozoite- and trophozoite-induced infections with Plasmodium falciparum in patients previously infected with heterologous species of Plasmodium: effect on development of parasitologic and clinical immunity","volume":"61","author":"WE Collins","year":"1999","journal-title":"Am. J. Trop. Med. Hyg"}],"updated-by":[{"DOI":"10.1371\/journal.pntd.0010760","type":"new_version","label":"New version","source":"publisher","updated":{"date-parts":[[2022,9,23]],"date-time":"2022-09-23T00:00:00Z","timestamp":1663891200000}}],"container-title":["PLOS Neglected Tropical Diseases"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/dx.plos.org\/10.1371\/journal.pntd.0010760","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2022,9,23]],"date-time":"2022-09-23T18:36:00Z","timestamp":1663958160000},"score":1,"resource":{"primary":{"URL":"https:\/\/dx.plos.org\/10.1371\/journal.pntd.0010760"}},"subtitle":[],"editor":[{"given":"Hans-Peter","family":"Fuehrer","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"editor"}]}],"short-title":[],"issued":{"date-parts":[[2022,9,13]]},"references-count":32,"journal-issue":{"issue":"9","published-online":{"date-parts":[[2022,9,13]]}},"URL":"https:\/\/doi.org\/10.1371\/journal.pntd.0010760","relation":{"new_version":[{"id-type":"doi","id":"10.1371\/journal.pntd.0010760","asserted-by":"object"}]},"ISSN":["1935-2735"],"issn-type":[{"value":"1935-2735","type":"electronic"}],"subject":[],"published":{"date-parts":[[2022,9,13]]}}}