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Quantitative PCR (qPCR) may provide greater sensitivity and permits estimation of the initial template DNA concentration. This study examined the hypothesis that qPCR assays using templates with higher copy numbers may be more sensitive for <jats:italic>P. falciparum<\/jats:italic> than assays based on templates with lower copy numbers.<\/jats:p>\n<\/jats:sec><jats:sec>\n<jats:title>Methods<\/jats:title>\n<jats:p>To test this hypothesis, ten qPCR assays for DNA sequences with template copy numbers from 1 to 160 were compared using parasite DNA standards (<jats:italic>n\u2009<\/jats:italic>=\u20092) and smear-positive filter paper blots from asymptomatic smear-positive subjects (<jats:italic>n\u2009<\/jats:italic>=\u200996).<\/jats:p>\n<\/jats:sec><jats:sec>\n<jats:title>Results<\/jats:title>\n<jats:p>Based on the testing of <jats:italic>P. falciparum<\/jats:italic> parasite DNA standards and filter paper blots, cycle threshold values decreased as the concentrations of template DNA and template copy numbers increased (<jats:italic>p\u2009<\/jats:italic>&lt;\u20090.001). Likewise, the analytical and clinical sensitivities of qPCR assays for <jats:italic>P. falciparum<\/jats:italic> DNA (based on DNA standards and filter paper blots, respectively) increased with template copy number. Despite the gains in clinical sensitivity from increased template copy numbers, qPCR assays failed to detect more than half of the filter paper blots with low parasite densities (\u2264\u2009200 asexual parasites per \u03bcl).<\/jats:p>\n<\/jats:sec><jats:sec>\n<jats:title>Conclusions<\/jats:title>\n<jats:p>These results confirm the hypothesis that the sensitivity of qPCR for <jats:italic>P. falciparum<\/jats:italic> in the blood of individuals with asymptomatic infection increases with template copy number. However, because even the most sensitive qPCR assays (with template copy numbers from 32 to 160) detected fewer than 50% of infections with\u2009\u2264\u2009200 asexual parasites per \u03bcl, the sensitivity of qPCR must be increased further to identify all smear-positive, asymptomatic individuals in order to interrupt transmission.<\/jats:p>\n<\/jats:sec>","DOI":"10.1186\/s12936-020-03365-8","type":"journal-article","created":{"date-parts":[[2020,8,18]],"date-time":"2020-08-18T12:03:21Z","timestamp":1597752201000},"update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":5,"title":["Template copy number and the sensitivity of quantitative PCR for Plasmodium falciparum in asymptomatic individuals"],"prefix":"10.1186","volume":"19","author":[{"given":"Trevor A.","family":"Thompson","sequence":"first","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Mahamoudou B.","family":"Tour\u00e9","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Daouda","family":"Sanogo","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Jeffrey G.","family":"Shaffer","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Seydou O.","family":"Doumbia","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-7972-9667","authenticated-orcid":false,"given":"Donald J.","family":"Krogstad","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"297","published-online":{"date-parts":[[2020,8,18]]},"reference":[{"key":"3365_CR1","doi-asserted-by":"crossref","first-page":"353","DOI":"10.1186\/1475-2875-11-353","volume":"11","author":"G Stresman","year":"2012","unstructured":"Stresman G, Kobayashi T, Kamanga A, Thuma PE, Mharakurwa S, Moss WJ, et al. 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J Infect Dis. 2017;216:713\u201322.","journal-title":"J Infect Dis"}],"container-title":["Malaria Journal"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/link.springer.com\/content\/pdf\/10.1186\/s12936-020-03365-8.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/link.springer.com\/article\/10.1186\/s12936-020-03365-8\/fulltext.html","content-type":"text\/html","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/link.springer.com\/content\/pdf\/10.1186\/s12936-020-03365-8.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2021,8,17]],"date-time":"2021-08-17T23:40:51Z","timestamp":1629243651000},"score":1,"resource":{"primary":{"URL":"https:\/\/malariajournal.biomedcentral.com\/articles\/10.1186\/s12936-020-03365-8"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2020,8,18]]},"references-count":44,"journal-issue":{"issue":"1","published-print":{"date-parts":[[2020,12]]}},"alternative-id":["3365"],"URL":"https:\/\/doi.org\/10.1186\/s12936-020-03365-8","relation":{},"ISSN":["1475-2875"],"issn-type":[{"value":"1475-2875","type":"electronic"}],"subject":[],"published":{"date-parts":[[2020,8,18]]},"assertion":[{"value":"25 March 2020","order":1,"name":"received","label":"Received","group":{"name":"ArticleHistory","label":"Article History"}},{"value":"10 August 2020","order":2,"name":"accepted","label":"Accepted","group":{"name":"ArticleHistory","label":"Article History"}},{"value":"18 August 2020","order":3,"name":"first_online","label":"First Online","group":{"name":"ArticleHistory","label":"Article History"}},{"value":"Subjects were enrolled with their informed assent and their parents\u2019 written informed consent after the study protocol had been reviewed and approved by the NIH (NIAID), the Mali and Tulane Institutional Review Boards (FWA000001769, FWA000002055) and the chief and elders of the village.","order":1,"name":"Ethics","group":{"name":"EthicsHeading","label":"Ethics approval and consent to participate"}},{"value":"Not applicable.","order":2,"name":"Ethics","group":{"name":"EthicsHeading","label":"Consent for publication"}},{"value":"The authors declare that they have no competing interests.","order":3,"name":"Ethics","group":{"name":"EthicsHeading","label":"Competing interests"}}],"article-number":"295"}}