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We examined the mitogenomes of a large global collection of human malaria parasites to explore how and when Plasmodium falciparum and P. vivax entered the Americas. We found evidence of a significant contribution of African and South Asian lineages to present-day New World malaria parasites with additional P. vivax lineages appearing to originate from Melanesia that were putatively carried by the Australasian peoples who contributed genes to Native Americans. Importantly, mitochondrial lineages of the P. vivax-like species P. simium are shared by platyrrhine monkeys and humans in the Atlantic Forest ecosystem, but not across the Amazon, which most likely resulted from one or a few recent human-to-monkey transfers. While enslaved Africans were likely the main carriers of P. falciparum mitochondrial lineages into the Americas after the conquest, additional parasites carried by Australasian peoples in pre-Columbian times may have contributed to the extensive diversity of extant local populations of P. vivax.

We evaluated the clinical efficacy of artesunate-mefloquine (ASMQ) fixed-dose combination to treat uncomplicated malaria in Juruá Valley, the main Plasmodium falciparum transmission hotspot in Brazil. Between November 2010 and February 2013, we enrolled 162 patients aged 4-73 years, with fever or a history of fever, and a single-species P. falciparum infection confirmed by microscopy and polymerase chain reaction (PCR). All 154 patients who completed the 42-day follow-up presented an adequate clinical and parasitologic response. ASMQ was well tolerated and no adverse event caused treatment interruption. Gametocytes were detected in 46.3% patients; 35.2% had gametocytes at enrollment, whereas others developed patent gametocytemia 1-14 days after starting ASMQ. By day 3 of treatment, all subjects had cleared asexual parasitemia, but parasite DNA remained PCR detectable in 37.6% of them. Day-3 PCR positivity was associated with prolonged gametocyte carriage. We found no molecular evidence of resistance to either MQ (pfmdr1 gene amplification) or AS (mutations in selected kelch13 gene domains known to be associated with AS resistance) in the local P. falciparum population. These results strongly support the use of ASMQ as a first-line regimen to treat uncomplicated P. falciparum malaria in northwestern Brazil, but underscore the need for gametocytocidal drugs to reduce the transmission potential of ASMQ-treated patients (ClinicalTrials.gov number NCT01144702)., (© The American Society of Tropical Medicine and Hygiene.)

The Plasmodium falciparum var gene family encodes large variant antigens, which are important virulence factors, and also targets of the humoral host response. The frequently observed mild outcomes of falciparum malaria in many places of the Amazon area prompted us to ask whether a globally restricted variant (var) gene repertoire is present in currently circulating and older isolates of this area. By exhaustive analysis of var gene tags from 89 isolates and clones taken during many years from all over the Brazilian Amazon, we estimate that there are probably no more than 350-430 distinct sequence types, less than for any similar sized area studied so far. Detailed analysis of the var tags from genetically distinct clones obtained from single isolates revealed restricted and redundant repertoires suggesting either a low incidence of infective bites or restricted variant gene diversity in inoculated parasites. Additionally, we found a structuring of var gene repertoires observed as a higher pairwise typing sharing in isolates from the same microregion compared to isolates from different regions. Fine analysis of translated var tags revealed that certain Distinct Sequence Identifiers (DSIDs) were differently represented in Brazilian/South American isolates when compared to datasets from other continents. By global alignment of worldwide var DBLalpha sequences and sorting in groups with more than 76% identity, 125 clusters were formed and more than half of all genes were found in nine clusters with 50 or more sequences. While Brazilian/South American sequences were represented only in 64 groups, African sequences were found in the majority of clusters. DSID type 1 related sequences accumulated almost completely in one single cluster, indicating that limited recombination occurs in these specific var gene types. These data demonstrate the so far highest pairwise type sharing values for the var gene family in isolates from all over an entire subcontinent. The apparent lack of specific sequences types suggests that the P. falciparum transmission dynamics in the whole Amazon are probably different from any other endemic region studied and possibly interfere with the parasite's ability to efficiently diversify its variant gene repertoires., (2009 Elsevier B.V. All rights reserved.)

To infer recent patterns of malaria transmission, we measured naturally acquired IgG antibodies to the conserved 19-kDa C-terminal region of the merozoite surface protein (MSP)-1 of both Plasmodium vivax (PvMSP-1(19)) and Plasmodium falciparum (PfMSP-1(19)) in remote malaria-exposed populations of the Amazon Basin. Community-based cross-sectional surveys were carried out between 2002 and 2003 in subjects of all age groups living along the margins of the Unini and Jaú rivers, Northwestern Brazil. We found high prevalence rates of IgG antibodies to PvMSP-1(19) (64.0 - 69.6%) and PfMSP-1(19) (51.6 - 52.0%), with significant differences in the proportion of subjects with antibodies to PvMSP-1(19) according to age, place of residence and habitual involvement in high-risk activities, defining some groups of highly exposed people who might be preferential targets of malaria control measures. In contrast, no risk factor other than age was significantly associated with seropositivity to PfMSP-1(19). Only 14.1% and 19.3% of the subjects tested for antibodies to PvMSP-1(19) and PfMSP-1(19) in consecutive surveys (142 - 203 days apart) seroconverted or had a three fold or higher increase in the levels of antibodies to these antigens. We discuss the extent to which serological data correlated with the classical malariometric indices and morbidity indicators measured in the studied population at the time of the seroprevalence surveys and highlight some limitations of serological data for epidemiological inference.

Most rapid diagnostic tests for Plasmodium falciparum malaria target the Histidine-Rich Proteins 2 and 3 (HRP2 and HRP3). Deletions of the hrp2 and hrp3 genes result in false-negative tests and are a threat for malaria control. A novel assay for molecular surveillance of hrp2/hrp3 deletions was developed based on droplet digital PCR (ddPCR). The assay quantifies hrp2, hrp3, and a control gene with very high accuracy. The theoretical limit of detection was 0.33 parasites/µl. The deletion was reliably detected in mixed infections with wild-type and hrp2-deleted parasites at a density of >100 parasites/reaction. For a side-by-side comparison with the conventional nested PCR (nPCR) assay, 248 samples were screened in triplicate by ddPCR and nPCR. No deletions were observed by ddPCR, while by nPCR hrp2 deletion was observed in 8% of samples. The ddPCR assay was applied to screen 830 samples from Kenya, Zanzibar/Tanzania, Ghana, Ethiopia, Brazil, and Ecuador. Pronounced differences in the prevalence of deletions were observed among sites, with more hrp3 than hrp2 deletions. In conclusion, the novel ddPCR assay minimizes the risk of false-negative results (i.e., hrp2 deletion observed when the sample is wild type), increases sensitivity, and greatly reduces the number of reactions that need to be run.

Most rapid diagnostic tests for Plasmodium falciparum malaria target the Histidine-Rich Proteins 2 and 3 (HRP2 and HRP3). Deletions of the hrp2 and hrp3 genes result in false-negative tests and are a threat for malaria control. A novel assay for molecular surveillance of hrp2/hrp3 deletions was developed based on droplet digital PCR (ddPCR). The assay quantifies hrp2, hrp3, and a control gene with very high accuracy. The theoretical limit of detection was 0.33 parasites/µl. The deletion was reliably detected in mixed infections with wild-type and hrp2-deleted parasites at a density of >100 parasites/reaction. For a side-by-side comparison with the conventional nested PCR (nPCR) assay, 248 samples were screened in triplicate by ddPCR and nPCR. No deletions were observed by ddPCR, while by nPCR hrp2 deletion was observed in 8% of samples. The ddPCR assay was applied to screen 830 samples from Kenya, Zanzibar/Tanzania, Ghana, Ethiopia, Brazil, and Ecuador. Pronounced differences in the prevalence of deletions were observed among sites, with more hrp3 than hrp2 deletions. In conclusion, the novel ddPCR assay minimizes the risk of false-negative results (i.e., hrp2 deletion observed when the sample is wild type), increases sensitivity, and greatly reduces the number of reactions that need to be run. [ABSTRACT FROM AUTHOR]