Plasmodium vivax is the most prevalent malarial species in South America and exerts a substantial burden on the populations it affects. The control and eventual elimination of P. vivax are global health priorities. Genomic research contributes to this objective by improving our understanding of the biology of P. vivax and through the development of new genetic markers that can be used to monitor efforts to reduce malaria transmission. Here we analyze whole-genome data from eight field samples from a region in Cordóba, Colombia where malaria is endemic. We find considerable genetic diversity within this population, a result that contrasts with earlier studies suggesting that P. vivax had limited diversity in the Americas. We also identify a selective sweep around a substitution known to confer resistance to sulphadoxine-pyrimethamine (SP). This is the first observation of a selective sweep for SP resistance in this species. These results indicate that P. vivax has been exposed to SP pressure even when the drug is not in use as a first line treatment for patients afflicted by this parasite. We identify multiple non-synonymous substitutions in three other genes known to be involved with drug resistance in Plasmodium species. Finally, we found extensive microsatellite polymorphisms. Using this information we developed 18 polymorphic and easy to score microsatellite loci that can be used in epidemiological investigations in South America.

Multiplicity of infection (MOI) refers to the average number of distinct parasite genotypes concurrently infecting a patient. Although several studies have reported on MOI and the frequency of multiclonal infections in Plasmodium falciparum, there is limited data on Plasmodium vivax. Here, MOI and the frequency of multiclonal infections were studied in areas from South America where P. vivax and P. falciparum can be compared.

Even though malaria incidence has decreased substantially in Guatemala since 2000, Guatemala remains one of the countries with the highest malaria transmission in Mesoamerica. Guatemala is committed to eliminating malaria as part of the initiative 'Elimination of Malaria in Mesoamerica and the Island of Hispaniola' (EMMIE); however, it is still in the control phase. During the past decade, the government strengthened malaria control activities including mass distribution of long-lasting insecticide-impregnated bed nets, early diagnosis and prompt treatment. This study aimed to determine the prevalence of malaria, including gametocytes, in three areas of Guatemala using active case detection (ACD) and quantitative polymerase chain reaction (qPCR).

Plasmodium vivax 48/45 protein is expressed on the surface of gametocytes/gametes and plays a key role in gamete fusion during fertilization. This protein was recently expressed in Escherichia coli host as a recombinant product that was highly immunogenic in mice and monkeys and induced antibodies with high transmission-blocking activity, suggesting its potential as a P. vivax transmission-blocking vaccine candidate. To determine sequence polymorphism of natural parasite isolates and its potential influence on the protein structure, all pvs48/45 sequences reported in databases from around the world as well as those from low-transmission settings of Latin America were compared.

We sequenced and annotated the genomes of four P. vivax strains collected from disparate geographic locations, tripling the number of genome sequences available for this understudied parasite and providing the first genome-wide perspective of global variability in this species. We observe approximately twice as much SNP diversity among these isolates as we do among a comparable collection of isolates of P. falciparum, a malaria-causing parasite that results in higher mortality. This indicates a distinct history of global colonization and/or a more stable demographic history for P. vivax relative to P. falciparum, which is thought to have undergone a recent population bottleneck. The SNP diversity, as well as additional microsatellite and gene family variability, suggests a capacity for greater functional variation in the global population of P. vivax. These findings warrant a deeper survey of variation in P. vivax to equip disease interventions targeting the distinctive biology of this neglected but major pathogen.

The study of the three protagonists in malaria-the Plasmodium parasite, the Anopheles mosquito, and the human host-is key to developing methods to control and eventually eliminate the disease. Genomic technologies, including the recent development of next-generation sequencing, enable interrogation of this triangle to an unprecedented level of scrutiny, and promise exciting progress toward real-time epidemiology studies and the study of evolutionary adaptation. We discuss the use of genomics by the International Centers of Excellence for Malaria Research, a network of field sites and laboratories in malaria-endemic countries that undertake cutting-edge research, training, and technology transfer in malarious countries of the world.

Transmission of malaria parasites from humans to Anopheles mosquitoes can be inhibited by specific antibodies elicited during malaria infection, which target surface Plasmodium gametocyte/gamete proteins. Some of these proteins may have potential for vaccine development. Pvs48/45 is a P. vivax gametocyte surface antigen orthologous to Pfs48/45, which may play a role during parasite fertilization and thus has potential for transmission blocking (TB) activity. Here we describe the expression of a recombinant Pvs48/45 protein expressed in Escherichia coli as a ∼60kDa construct which we tested for antigenicity using human sera and for its immunogenicity and transmission blocking activity of specific anti-mouse and anti-monkey Pvs48/45 antibodies. The protein reacted with sera of individuals from malaria-endemic areas and in addition induced specific IgG antibody responses in BALB/c mice and Aotus l. griseimembra monkeys. Sera from both immunized animal species recognized native P. vivax protein in Western blot (WB) and immunofluorescence assays. Moreover, sera from immunized mice and monkeys produced significant inhibition of parasite transmission to An. Albimanus mosquitoes as shown by membrane feeding assays. Results indicate the presence of reactive epitopes in the Pvs48/45 recombinant product that induce antibodies with TB activity. Further testing of this protein is ongoing to determine its vaccine potential.

Reported urban malaria cases are increasing in Latin America, however, evidence of such trend remains insufficient. Here, we propose an integrated approach that allows characterizing malaria transmission at the rural-to-urban interface by combining epidemiological, entomological, and parasite genotyping methods.

Previous work suggests that Brazilian Plasmodium falciparum has limited genetic diversity and a history of bottlenecks, multiple reintroductions due to human migration, and clonal expansions. We hypothesized that Brazilian P. falciparum would exhibit clonal structure. We examined isolates collected across two decades from Amapá, Rondônia, and Pará state (n = 190). By examining more microsatellites markers on more chromosomes than previous studies, we hoped to define the extent of low diversity, linkage disequilibrium, bottlenecks, population structure, and parasite migration within Brazil. We used retrospective genotyping of samples from the 1980s and 1990s to explore the population genetics of SP resistant dhfr and dhps alleles. We tested an existing hypothesis that the triple mutant dhfr mutations 50R/51I/108N and 51I/108N/164L developed in southern Amazon from a single origin of common or similar parasites. We found that Brazilian P. falciparum had limited genetic diversity and isolation by distance was rejected, which suggests it underwent bottlenecks followed by migration between sites. Unlike Peru, there appeared to be gene flow across the Brazilian Amazon basin. We were unable to divide parasite populations by clonal lineages and pairwise FST were common. Most parasite diversity was found within sites in the Brazilian Amazon, according to AMOVA. Our results challenge the hypothesis that triple mutant alleles arose from a single lineage in the Southern Amazon. SP resistance, at both the double and triple mutant stages, developed twice and potentially in different regions of the Brazilian Amazon. We would have required samples from before the 1980s to describe how SP resistance spread across the basin or describe the complex internal migration of Brazilian parasites after the colonization efforts of past decades. The Brazilian Amazon basin may have sufficient internal migration for drug resistance reported in any particular region to rapidly spread to other parts of basin under similar drug pressure.

Several of the intended Plasmodium falciparum vaccine candidate antigens are highly polymorphic and could render a vaccine ineffective if their antigenic sites were not represented in the vaccine. In this study, characterization of genetic variability was performed in major B and T-cell epitopes within vaccine candidate antigens in isolates of P. falciparum from Peru.

Plasmodium vivax malaria is a neglected disease, particularly during pregnancy. Severe vivax malaria is associated with inflammatory responses but in pregnancy immune alterations make it uncertain as to what cytokine signatures predominate, and how the type and quantity of blood immune mediators influence delivery outcomes. We measured the plasma concentrations of a set of thirty-one biomarkers, comprising cytokines, chemokines and growth factors, in 987 plasma samples from a cohort of 572 pregnant women from five malaria-endemic tropical countries and related these concentrations to delivery outcomes (birth weight and hemoglobin levels) and malaria infection. Samples were collected at recruitment (first antenatal visit) and at delivery (periphery, cord and placenta). At recruitment, we found that P. vivax-infected pregnant women had higher plasma concentrations of proinflammatory (IL-6, IL-1β, CCL4, CCL2, CXCL10) and TH1-related cytokines (mainly IL-12) than uninfected women. This biomarker signature was essentially lost at delivery and was not associated with birth weight nor hemoglobin levels. Antiinflammatory cytokines (IL-10) were positively associated with infection and poor delivery outcomes. CCL11 was the only biomarker to show a negative association with P. vivax infection and its concentration at recruitment was positively associated with hemoglobin levels at delivery. Birth weight was negatively associated with peripheral IL-4 levels at delivery. Our multi-biomarker multicenter study is the first comprehensive one to characterize the immunological signature of P. vivax infection in pregnancy thus far. In conclusion, data show that while TH1 and pro-inflammatory responses are dominant during P. vivax infection in pregnancy, antiinflammatory cytokines may compensate excessive inflammation avoiding poor delivery outcomes, and skewness toward a TH2 response may trigger worse delivery outcomes. CCL11, a chemokine largely neglected in the field of malaria, emerges as an important marker of exposure or mediator in this condition.

Emperor Geese (Anser canagicus) are iconic waterfowl endemic to Alaska and adjacent areas of northeastern Russia that are considered to be near threatened by the International Union for Conservation. This species has been identified as harboring diverse viruses and parasites which have, at times, been associated with disease in other avian taxa. To better assess if disease represents a vulnerability for Emperor Geese breeding on the Yukon-Kuskokwim Delta, Alaska, we evaluated if haemosporidian parasites were associated with decreased mass or survival among adult female nesting birds captured during 2006-2016. Through molecular analyses, we detected genetically diverse Leucocytozoon, Haemoproteus, and Plasmodium parasites in 28%, 1%, and 1% of 607 blood samples screened in triplicate, respectively. Using regression analysis, we found evidence for a small effect of Leucocytozoon infection on the mass of incubating adult female Emperor Geese. The estimated mass of infected individuals was approximately 43 g (95% CI: 20-67 g), or approximately 2%, less than uninfected birds when captured during the second half of incubation (days 11-25). We did not, however, find support for an effect of Leucocytozoon infection on survival of adult female nesting Emperor Geese using a multi-state hidden Markov framework to analyze mark-resight and recapture data. Using parasite mitochondrial DNA cytochrome b sequences, we identified 23 haplotypes among infected Emperor Geese. Leucocytozoon haplotypes clustered into three phylogenetically supported clades designated as 'L. simondi clade A', 'L. simondi clade B', and 'other Leucocytozoon'. We did not find evidence that parasites assigned to any of these clades were associated with differential mass measures among nesting adult female Emperor Geese. Collectively, our results provide negligible evidence for Leucocytozoon parasites as causing detrimental effects to adult female Emperor Geese breeding on the Yukon-Kuskokwim Delta.

Vector control for malaria prevention relies most often on the use of insecticide-treated bed net (ITNs) and indoor residual spraying. Little is known about the longevity of long-lasting insecticidal nets (LLINs) in the Americas. The physical integrity and insecticide retention of LLINs over time were monitored after a bed net distribution campaign to assess community practices around LLIN care and use in Waspam, northeastern Nicaragua.

Malaria incidence has reached staggering numbers in Venezuela. Commonly, Bolívar State accounted for approximately 70% of the country cases every year. Most cases cluster in the Sifontes municipality, a region characterized by an extractive economy, including gold mining. An increase in migration to Sifontes, driven by gold mining, fueled a malaria spillover to the rest of the country and the region. Here samples collected in 2018 were compared with a previous study of 2003/2004 to describe changes in the parasites population structures and the frequency of point mutations linked to anti-malarial drugs.

Colombia aims to eliminate malaria by 2030 but remains one of the highest burden countries in the Americas. Plasmodium vivax contributes half of all malaria cases, with its control challenged by relapsing parasitaemia, drug resistance and cross-border spread. Using 64 Colombian P. vivax genomes collected between 2013 and 2017, we explored diversity and selection in two major foci of transmission: Chocó and Córdoba. Open-access data from other countries were used for comparative assessment of drug resistance candidates and to assess cross-border spread. Across Colombia, polyclonal infections were infrequent (12%), and infection connectivity was relatively high (median IBD = 5%), consistent with low endemicity. Chocó exhibited a higher frequency of polyclonal infections (23%) than Córdoba (7%), although the difference was not significant (P = 0.300). Most Colombian infections carried double pvdhfr (95%) and single pvdhps (71%) mutants, but other drug resistance mutations were less prevalent (< 10%). There was no evidence of selection at the pvaat1 gene, whose P. falciparum orthologue has recently been implicated in chloroquine resistance. Global population comparisons identified other putative adaptations. Within the Americas, low-level connectivity was observed between Colombia and Peru, highlighting potential for cross-border spread. Our findings demonstrate the potential of molecular data to inform on infection spread and adaptation.

Resistance against artemisinin-based combination therapy is one of the challenges to malaria control and elimination globally. Mutations in different genes (Pfdhfr, Pfdhps, Pfk-13 and Pfmdr1) confer resistance to artesunate and sulfadoxine-pyrimethamine (AS + SP) were analysed from Mandla district, Madhya Pradesh, to assess the effectiveness of the current treatment regimen against uncomplicated Plasmodium falciparum.

Malaria sterile immunity has been reproducibly induced by immunization with Plasmodium radiation-attenuated sporozoites (RAS). Analyses of sera from RAS-immunized individuals allowed the identification of P. falciparum antigens, such as the circumsporozoite protein (CSP), the basis for the RTS, S and R21Matrix-M vaccines. Similar advances in P. vivax (Pv) vaccination have been elusive. We previously reported 42% (5/12) of sterile protection in malaria-unexposed, Duffy-positive (Fy +) volunteers immunized with PvRAS followed by a controlled human malaria infection (CHMI). Using a custom protein microarray displaying 515 Pv antigens, we found a significantly higher reactivity to PvCSP and one hypothetical protein (PVX_089630) in volunteers protected against P. vivax infection. In mock-vaccinated Fy + volunteers, a strong antibody response to CHMI was also observed. Although the Fy- volunteers immunized with non-irradiated Pv-infected mosquitoes (live sporozoites) did not develop malaria after CHMI, they recognized a high number of antigens, indicating the temporary presence of asexual parasites in peripheral blood. Together, our findings contribute to the understanding of the antibody response to P. vivax infection and allow the identification of novel parasite antigens as vaccine candidates.Trial registration: ClinicalTrials.gov number: NCT01082341.

Foamy viruses are complex retroviruses that have been shown to be transmitted from nonhuman primates to humans. In Bangladesh, infection with simian foamy virus (SFV) is ubiquitous among rhesus macaques, which come into contact with humans in diverse locations and contexts throughout the country. We analyzed microsatellite DNA from 126 macaques at six sites in Bangladesh in order to characterize geographic patterns of macaque population structure. We also included in this study 38 macaques owned by nomadic people who train them to perform for audiences. PCR was used to analyze a portion of the proviral gag gene from all SFV-positive macaques, and multiple clones were sequenced. Phylogenetic analysis was used to infer long-term patterns of viral transmission. Analyses of SFV gag gene sequences indicated that macaque populations from different areas harbor genetically distinct strains of SFV, suggesting that geographic features such as forest cover play a role in determining the dispersal of macaques and SFV. We also found evidence suggesting that humans traveling the region with performing macaques likely play a role in the translocation of macaques and SFV. Our studies found that individual animals can harbor more than one strain of SFV and that presence of more than one SFV strain is more common among older animals. Some macaques are infected with SFV that appears to be recombinant. These findings paint a more detailed picture of how geographic and sociocultural factors influence the spectrum of simian-borne retroviruses.

Individuals exposed to malaria infections for a long time develop immune responses capable of blocking Plasmodium transmission to mosquito vectors, potentially limiting parasite spreading in nature. Development of a malaria TB vaccine requires a better understanding of the mechanisms and main effectors responsible for transmission blocking (TB) responses. The lack of an in vitro culture system for Plasmodium vivax has been an important drawback for development of a standardized method to assess TB responses to this parasite. This study evaluated host, vector, and parasite factors that may influence Anopheles mosquito infection in order to develop an efficient and reliable assay to assess the TB immunity.

Malaria control programmes rely on confirmation of parasite presence in patients' blood prior to treatment administration. Plasmodium parasites are detected mostly by microscopy or rapid diagnostic test (RDT). Although these methods contribute significantly to malaria control/elimination, they are not suitable for detecting the significant proportion of asymptomatic subjects harbouring low levels of parasitaemia, which endure untreated as potential reservoirs for transmission. Malaria prevalence was assessed in endemic regions of Colombia over a 4-year follow-up.