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Background  The hypotheses put forward to explain the malaria transmission cycle in extra-Amazonian Brazil, an area of very low malaria incidence, are based on either a zoonotic scenario involving simian malaria, or a scenario in which asymptomatic carriers play an important role.    Objectives  To determine the incidence of asymptomatic infection by detecting  Plasmodium  spp. DNA and its role in residual malaria transmission in a non-Amazonian region of Brazil.    Methods  Upon the report of the first malaria case in 2010 in the Atlantic Forest region of the state of Espírito Santo, inhabitants within a 2 km radius were invited to participate in a follow-up study. After providing signed informed consent forms, inhabitants filled out a questionnaire and gave blood samples for PCR, and thick and thin smears. Follow-up visits were performed every 3 months over a 21 month period, when new samples were collected and information was updated.    Results  Ninety-two individuals were initially included for follow-up. At the first collection, all of them were clearly asymptomatic. One individual was positive for  Plasmodium vivax , one for  Plasmodium malariae  and one for both  P. vivax  and  P. malariae , corresponding to a prevalence of 3.4% (2.3% for each species). During follow-up, four new PCR-positive cases (two for each species) were recorded, corresponding to an incidence of 2.5 infections per 100 person-years or 1.25 infections per 100 person-years for each species. A mathematical transmission model was applied, using a low frequency of human carriers and the vector density in the region, and calculated based on previous studies in the same locality whose results were subjected to a linear regression. This analysis suggests that the transmission chain is unlikely to be based solely on human carriers, regardless of whether they are symptomatic or not.    Conclusion  The low incidence of cases and the low frequency of asymptomatic malaria carriers investigated make it unlikely that the transmission chain in the region is based solely on human hosts, as cases are isolated one from another by hundreds of kilometers and frequently by long periods of time, reinforcing instead the hypothesis of zoonotic transmission.    Electronic supplementary material  The online version of this article (10.1186/s12936-018-2263-z) contains supplementary material, which is available to authorized users.

Assessment of asymptomatic Plasmodium spp. infection by detection of parasite DNA in residents of an extra-Amazonian region of Brazil