Aedes albopictus is a viable vector for several infectious diseases such as Zika, West Nile, Dengue viruses and others. Originating from Asia, this invasive species is rapidly expanding into North American temperate areas and urbanized places causing major concerns for public health. Previous analyses show that warm temperatures and high humidity during the mosquito season are ideal conditions for A. albopictus development, while its distribution is correlated with population density. To better understand A. albopictus expansion into urban places it is important to consider the role of both environmental and neighborhood factors. The present study aims to assess the relative importance of both environmental variables and neighborhood factors in the prediction of A. albopictus' presence in Southeast Pennsylvania using MaxEnt (version 3.4.1) machine-learning algorithm. Three models are developed that include: (1) exclusively environmental variables, (2) exclusively neighborhood factors, and (3) a combination of environmental variables and neighborhood factors. Outcomes from the three models are compared in terms of variable importance, accuracy, and the spatial distribution of predicted A. albopictus' presence. All three models predicted the presence of A. albopictus in urban centers, however, each to a different spatial extent. The combined model resulted in the highest accuracy (74.7%) compared to the model with only environmental variables (73.5%) and to the model with only neighborhood factors (72.1%) separately. Although the combined model does not essentially increase the accuracy in the prediction, the spatial patterns of mosquito distribution are different when compared to environmental or neighborhood factors alone. Environmental variables help to explain conditions associated with mosquitoes in suburban/rural areas, while neighborhood factors summarize the local conditions that can also impact mosquito habitats in predominantly urban places. Overall, the present study shows that MaxEnt is suitable for integrating neighborhood factors associated with mosquito presence that can complement and improve species distribution modeling.

Plasmodium falciparum malaria is a leading cause of pediatric morbidity and mortality in holoendemic transmission areas. Severe malarial anemia [SMA, hemoglobin (Hb) < 5.0 g/dL in children] is the most common clinical manifestation of severe malaria in such regions. Although innate immune response genes are known to influence the development of SMA, the role of natural killer (NK) cells in malaria pathogenesis remains largely undefined. As such, we examined the impact of genetic variation in the gene encoding a primary NK cell receptor, natural cytotoxicity-triggering receptor 3 (NCR3), on the occurrence of malaria and SMA episodes over time.

Plasmodium species causing malaria in humans are not monophyletic, sharing common ancestors with nonhuman primate parasites. Plasmodium gonderi is one of the few known Plasmodium species infecting African old-world monkeys that are not found in apes. This study reports a de novo assembled P. gonderi genome with complete chromosomes. The P. gonderi genome shares codon usage, syntenic blocks, and other characteristics with the human parasites Plasmodium ovale s.l. and Plasmodium malariae, also of African origin, and the human parasite Plasmodium vivax and species found in nonhuman primates from Southeast Asia. Using phylogenetically aware methods, newly identified syntenic blocks were found enriched with conserved metabolic genes. Regions outside those blocks harbored genes encoding proteins involved in the vertebrate host-Plasmodium relationship undergoing faster evolution. Such genome architecture may have facilitated colonizing vertebrate hosts. Phylogenomic analyses estimated the common ancestor between P. vivax and an African ape parasite P. vivax-like, within the Asian nonhuman primates parasites clade. Time estimates incorporating P. gonderi placed the P. vivax and P. vivax-like common ancestor in the late Pleistocene, a time of active migration of hominids between Africa and Asia. Thus, phylogenomic and time-tree analyses are consistent with an Asian origin for P. vivax and an introduction of P. vivax-like into Africa. Unlike other studies, time estimates for the clade with Plasmodium falciparum, the most lethal human malaria parasite, coincide with their host species radiation, African hominids. Overall, the newly assembled genome presented here has the quality to support comparative genomic investigations in Plasmodium.

The majority of malaria rapid diagnostic tests (RDTs) detect Plasmodium falciparum histidine-rich protein 2 (PfHRP2), encoded by the pfhrp2 gene. Recently, P. falciparum isolates from Peru were found to lack pfhrp2 leading to false-negative RDT results. We hypothesized that pfhrp2-deleted parasites in Peru derived from a single genetic event. We evaluated the parasite population structure and pfhrp2 haplotype of samples collected between 1998 and 2005 using seven neutral and seven chromosome 8 microsatellite markers, respectively. Five distinct pfhrp2 haplotypes, corresponding to five neutral microsatellite-based clonal lineages, were detected in 1998-2001; pfhrp2 deletions occurred within four haplotypes. In 2003-2005, outcrossing among the parasite lineages resulted in eight population clusters that inherited the five pfhrp2 haplotypes seen previously and a new haplotype; pfhrp2 deletions occurred within four of these haplotypes. These findings indicate that the genetic origin of pfhrp2 deletion in Peru was not a single event, but likely occurred multiple times.

Plasmodium vivax is the most prevalent human malaria parasite outside of Africa. Yet, studies aimed to identify genes with signatures consistent with natural selection are rare. Here, we present a comparative analysis of the pattern of genetic variation of five sequenced isolates of P. vivax and its divergence with two closely related species, Plasmodium cynomolgi and Plasmodium knowlesi, using a set of orthologous genes. In contrast to Plasmodium falciparum, the parasite that causes the most lethal form of human malaria, we did not find significant constraints on the evolution of synonymous sites genome wide in P. vivax. The comparative analysis of polymorphism and divergence across loci allowed us to identify 87 genes with patterns consistent with positive selection, including genes involved in the "exportome" of P. vivax, which are potentially involved in evasion of the host immune system. Nevertheless, we have found a pattern of polymorphism genome wide that is consistent with a significant amount of constraint on the replacement changes and prevalent negative selection. Our analyses also show that silent polymorphism tends to be larger toward the ends of the chromosomes, where many genes involved in antigenicity are located, suggesting that natural selection acts not only by shaping the patterns of variation within the genes but it also affects genome organization.

Molecular methods elucidate evolutionary and ecological processes in parasites, where interaction between hosts and parasites enlighten the evolution of parasite lifestyles and host defenses. Population genetics of Plasmodium vivax parasites accurately describe transmission dynamics of the parasites and evaluation of malaria control measures. As a first generation vaccine candidate against malaria, the Circumsporozoite Protein (CSP) has demonstrated significant potential in P. falciparum. Extensive polymorphism hinders the development of a potent malaria vaccine. Hence, the genetic diversity of Pvcsp was investigated for the first time in 60 Sri Lankan clinical isolates by obtaining the nucleotide sequence of the central repeat (CR) domain and examining the polymorphism of the peptide repeat motifs (PRMs), the genetic diversity indices and phylogenetic relationships. PCR amplicons determined size polymorphism of 610, 700 and 710 bp in Pvcsp of Sri Lanka where all amino acid sequences obtained were of the VK210 variant, consisting variable repeats of 4 different PRMs. The two most abundant PRMs of the CR domain, GDRADGQPA and GDRAAGQPA consisted ~2-4 repeats, while GNRAAGQPA was unique to the island. Though, different nucleotide sequences termed repeat allotypes (RATs) were observed for each PRM, these were synonymous contributing to a less polymorphic CR domain. The genetic diversity of Pvcsp in Sri Lanka was due to the number of repetitive peptide repeat motifs, point mutations, and intragenic recombination. The 19 amino acid haplotypes defined were exclusive to Sri Lanka, whereas the 194 Pvcsp sequences of global isolates generated 57 more distinct a.a. haplotypes of the VK210 variant. Strikingly, the CR domain of both VK210 and VK247 variants was under purifying selection interpreting the scarcity of CSP non-synonymous polymorphisms. Insights to the distribution of RATs in the CR region with geographic clustering of the P. vivax VK210 variant were revealed. The cladogram reiterated this unique geographic clustering of local (VK210) and global isolates (VK210 and VK247), which was further validated by the elevated fixation index values of the VK210 variant.

Haemosporidians are a diverse group of vector-borne parasitic protozoa that includes the agents of human malaria; however, most of the described species are found in birds and reptiles. Although our understanding of these parasites' diversity has expanded by analyses of their mitochondrial genes, there is limited information on these genes' evolutionary rates. Here, 114 mitochondrial genomes (mtDNA) were studied from species belonging to four genera: Leucocytozoon, Haemoproteus, Hepatocystis, and Plasmodium. Contrary to previous assertions, the mtDNA is phylogenetically informative. The inferred phylogeny showed that, like the genus Plasmodium, the Leucocytozoon and Haemoproteus genera are not monophyletic groups. Although sensitive to the assumptions of the molecular dating method used, the estimated times indicate that the diversification of the avian haemosporidian subgenera/genera took place after the Cretaceous-Paleogene boundary following the radiation of modern birds. Furthermore, parasite clade differences in mtDNA substitution rates and strength of negative selection were detected. These differences may affect the biological interpretation of mtDNA gene lineages used as a proxy to species in ecological and parasitological investigations. Given that the mitochondria are critically important in the parasite life cycle stages that take place in the vector and that the transmission of parasites belonging to particular clades has been linked to specific insect families/subfamilies, this study suggests that differences in vectors have affected the mode of evolution of haemosporidian mtDNA genes. The observed patterns also suggest that the radiation of haemosporidian parasites may be the result of community-level evolutionary processes between their vertebrate and invertebrate hosts.

The Thailand-Cambodia border is the epicenter for drug-resistant falciparum malaria. Previous studies have shown that chloroquine (CQ) and pyrimethamine resistance originated in this region and eventually spread to other Asian countries and Africa. However, there is a dearth in understanding the origin and evolution of dhps alleles associated with sulfadoxine resistance. The present study was designed to reveal the origin(s) of sulfadoxine resistance in Cambodia and its evolutionary relationship to African and South American dhps alleles. We sequenced 234 Cambodian Plasmodium falciparum isolates for the dhps codons S436A/F, A437G, K540E, A581G and A613S/T implicated in sulfadoxine resistance. We also genotyped 10 microsatellite loci around dhps to determine the genetic backgrounds of various alleles and compared them with the backgrounds of alleles prevalent in Africa and South America. In addition to previously known highly-resistant triple mutant dhps alleles SGEGA and AGEAA (codons 436, 437, 540, 581, 613 are sequentially indicated), a large proportion of the isolates (19.3%) contained a 540N mutation in association with 437G/581G yielding a previously unreported triple mutant allele, SGNGA. Microsatellite data strongly suggest the strength of selection was greater on triple mutant dhps alleles followed by the double and single mutants. We provide evidence for at least three independent origins for the double mutants, one each for the SGKGA, AGKAA and SGEAA alleles. Our data suggest that the triple mutant allele SGEGA and the novel allele SGNGA have common origin on the SGKGA background, whereas the AGEAA triple mutant was derived from AGKAA on multiple, albeit limited, genetic backgrounds. The SGEAA did not share haplotypes with any of the triple mutants. Comparative analysis of the microsatellite haplotypes flanking dhps alleles from Cambodia, Kenya, Cameroon and Venezuela revealed an independent origin of sulfadoxine resistant alleles in each of these regions.

There is an increased interest in potential zoonotic malarias. To date, Plasmodium malariae that infects humans remains indistinguishable from Plasmodium brasilianum, which is widespread among New World primates. Distributed throughout tropical Central and South America, the Callitrichidae are small arboreal primates in which detection of natural Plasmodium infection has been extremely rare. Most prior screening efforts have been limited to small samples, the use of low-probability detection methods, or both. Rarely have screening efforts implemented a longitudinal sampling design. Through an annual mark-recapture program of two sympatric callitrichids, the emperor (Saguinus imperator) and saddleback (Saguinus fuscicollis) tamarins, whole blood samples were screened for Plasmodium by microscopy and nested PCR of the cytochrome b gene across four consecutive years (2012-2015). Following the first field season, approximately 50% of the samples collected each subsequent year were from recaptured individuals. In particular, out of 245 samples from 129 individuals, 11 samples from 6 individuals were positive for Plasmodium, and all but one of these infections was found in S. imperator. Importantly, the cytochrome b sequences were 100% identical to former isolates of P. malariae from humans and P. brasilianum from Saimiri sp. Chronic infections were detected as evidenced by repeated infections (7) from two individuals across the 4-year study period. Furthermore, 4 of the 5 infected emperor tamarins were part of a single group spanning the entire study period. Overall, the low prevalence reported here is consistent with previous findings. This study identifies two new natural hosts for P. brasilianum and provides evidence in support of chronic infections in wildlife populations. Given that callitrichids are often found in mixed-species associations with other primates and can be resilient to human-disturbed environments, they could contribute to the maintenance of P. malariae populations if future work provides entomological and epidemiological evidence indicating human zoonotic infections.

Plasmodium falciparum infections remain among the leading causes of morbidity and mortality in holoendemic transmission areas. Located within region 5q31.1, the colony-stimulating factor 2 gene (CSF2) encodes granulocyte-macrophage colony-stimulating factor (GM-CSF), a hematopoietic growth factor that mediates host immune responses. Since the effect of CSF2 variation on malaria pathogenesis remains unreported, we investigated the impact of two genetic variants in the 5q31.1 gene region flanking CSF2:g-7032 G > A (rs168681:G > A) and CSF2:g.64544T > C (rs246835:T > C) on the rate and timing of malaria and severe malarial anemia (SMA, Hb < 5.0 g/dL) episodes over 36 months of follow-up. Children (n = 1654, aged 2-70 months) were recruited from a holoendemic P. falciparum transmission area of western Kenya. Decreased incidence rate ratio (IRR) for malaria was conferred by inheritance of the CSF2:g.64544 TC genotype (P = 0.0277) and CSF2 AC/GC diplotype (P = 0.0015). Increased IRR for malaria was observed in carriers of the CSF2 AT/GC diplotype (P = 0.0237), while the inheritance of the CSF2 AT haplotype increased the IRR for SMA (P = 0.0166). A model estimating the longitudinal risk of malaria showed decreased hazard rates among CSF2 AC haplotype carriers (P = 0.0045). Investigation of all-cause mortality revealed that inheritance of the GA genotype at CSF2:g-7032 increased the risk of mortality (P = 0.0315). Higher risk of SMA and all-cause mortality were observed in younger children (P < 0.0001 and P = 0.0015), HIV-1(+) individuals (P < 0.0001 and P < 0.0001), and carriers of HbSS (P = 0.0342 and P = 0.0019). Results from this holoendemic P. falciparum area show that variation in gene region 5q31.1 influences susceptibility to malaria, SMA, and mortality, as does age, HIV-1 status, and inheritance of HbSS.

Epidemiological data across the United States of America illustrate health disparities in COVID-19 infection, hospitalization, and mortality by race/ethnicity. However, limited information is available from prospective observational studies in hospitalized patients, particularly for American Indian or Alaska Native (AI/AN) populations. Here, we present risk factors associated with severe COVID-19 and mortality in patients (4/2020-12/2021, n = 475) at the University of New Mexico Hospital. Data were collected on patient demographics, infection duration, laboratory measures, comorbidities, treatment(s), major clinical events, and in-hospital mortality. Severe disease was defined by COVID-related intensive care unit requirements and/or death. The cohort was stratified by self-reported race/ethnicity: AI/AN (30.7%), Hispanic (47.0%), non-Hispanic White (NHW, 18.5%), and Other (4.0%, not included in statistical comparisons). Despite similar timing of infection and comparable comorbidities, admission characteristics for AI/AN patients included younger age (P = 0.02), higher invasive mechanical ventilation requirements (P = 0.0001), and laboratory values indicative of more severe disease. Throughout hospitalization, the AI/AN group also experienced elevated invasive mechanical ventilation (P < 0.0001), shock (P = 0.01), encephalopathy (P = 0.02), and severe COVID-19 (P = 0.0002), consistent with longer hospitalization (P < 0.0001). Self-reported AI/AN race/ethnicity emerged as the highest risk factor for severe COVID-19 (OR = 3.19; 95% CI = 1.70-6.01; P = 0.0003) and was a predictor of in-hospital mortality (OR = 2.35; 95% CI = 1.12-4.92; P = 0.02). Results from this study highlight the disproportionate impact of COVID-19 on hospitalized AI/AN patients, who experienced more severe illness and associated mortality, compared to Hispanic and NHW patients, even when accounting for symptom onset and comorbid conditions. These findings underscore the need for interventions and resources to address health disparities in the COVID-19 pandemic.

Although parasitic organisms are found worldwide, the relative importance of host specificity and geographic isolation for parasite speciation has been explored in only a few systems. Here, we study Plasmodium parasites known to infect Asian nonhuman primates, a monophyletic group that includes the lineage leading to the human parasite Plasmodium vivax and several species used as laboratory models in malaria research. We analyze the available data together with new samples from three sympatric primate species from Borneo: The Bornean orangutan and the long-tailed and the pig-tailed macaques. We find several species of malaria parasites, including three putatively new species in this biodiversity hotspot. Among those newly discovered lineages, we report two sympatric parasites in orangutans. We find no differences in the sets of malaria species infecting each macaque species indicating that these species show no host specificity. Finally, phylogenetic analysis of these data suggests that the malaria parasites infecting Southeast Asian macaques and their relatives are speciating three to four times more rapidly than those with other mammalian hosts such as lemurs and African apes. We estimate that these events took place in approximately a 3-4-Ma period. Based on the genetic and phenotypic diversity of the macaque malarias, we hypothesize that the diversification of this group of parasites has been facilitated by the diversity, geographic distributions, and demographic histories of their primate hosts.

Plasmodium falciparum shares its most recent common ancestor with parasites found in African apes; these species constitute the so-called Laverania clade. In this investigation, the evolutionary history of Plasmodium lineages found in chimpanzees (Pan troglodytes) was explored.

Understanding the origin and spread of mutations associated with drug resistance, especially in the context of combination therapy, will help guide strategies to halt and prevent the emergence of resistance. Unfortunately, studies have assessed these complex processes when resistance is already highly prevalent. Even further, information on the evolutionary dynamics leading to multidrug-resistant parasites is scattered and limited to areas with low or seasonal malaria transmission. This study describes the dynamics of strong selection for mutations conferring resistance against sulphadoxine-pyrimethamine (SP), a combination therapy, in western Kenya between 1992 and 1999, just before SP became first-line therapy (1999). Importantly, the study is based on longitudinal data, which allows for a comprehensive analysis that contrasts with previous cross-sectional studies carried out in other endemic regions.

Timing the origin of human malarias has been a focus of great interest. Previous studies on the mitochondrial genome concluded that Plasmodium in primates, including those parasitic to humans, radiated relatively recently during a process where host switches were common. Those investigations, however, assumed constant rate of evolution and tightly bound (fixed) calibration points based on host fossils or host distribution. We investigate the effect of such assumptions using different molecular dating methods. We include parasites from Lemuroidea since their distribution provides an external validation to time estimates allowing us to disregard scenarios that cannot explain their introduction in Madagascar.

Malaria has reemerged in many regions where once it was nearly eliminated. Yet the source of these parasites, the process of repopulation, their population structure, and dynamics are ill defined. Peru was one of malaria eradication's successes, where Plasmodium falciparum was nearly eliminated for two decades. It reemerged in the 1990s. In the new era of malaria elimination, Peruvian P. falciparum is a model of malaria reinvasion. We investigated its population structure and drug resistance profiles. We hypothesized that only populations adapted to local ecological niches could expand and repopulate and originated as vestigial populations or recent introductions. We investigated the genetic structure (using microsatellites) and drug resistant genotypes of 220 parasites collected from patients immediately after peak epidemic expansion (1999-2000) from seven sites across the country. The majority of parasites could be grouped into five clonal lineages by networks and AMOVA. The distribution of clonal lineages and their drug sensitivity profiles suggested geographic structure. In 2001, artesunate combination therapy was introduced in Peru. We tested 62 parasites collected in 2006-2007 for changes in genetic structure. Clonal lineages had recombined under selection for the fittest parasites. Our findings illustrate that local adaptations in the post-eradication era have contributed to clonal lineage expansion. Within the shifting confluence of drug policy and malaria incidence, populations continue to evolve through genetic outcrossing influenced by antimalarial selection pressure. Understanding the population substructure of P. falciparum has implications for vaccine, drug, and epidemiologic studies, including monitoring malaria during and after the elimination phase.

The origin of Plasmodium falciparum, the etiological agent of the most dangerous forms of human malaria, remains controversial. Although investigations of homologous parasites in African Apes are crucial to resolve this issue, studies have been restricted to a chimpanzee parasite related to P. falciparum, P. reichenowi, for which a single isolate was available until very recently. Using PCR amplification, we detected Plasmodium parasites in blood samples from 18 of 91 individuals of the genus Pan, including six chimpanzees (three Pan troglodytes troglodytes, three Pan t. schweinfurthii) and twelve bonobos (Pan paniscus). We obtained sequences of the parasites' mitochondrial genomes and/or from two nuclear genes from 14 samples. In addition to P. reichenowi, three other hitherto unknown lineages were found in the chimpanzees. One is related to P. vivax and two to P. falciparum that are likely to belong to distinct species. In the bonobos we found P. falciparum parasites whose mitochondrial genomes indicated that they were distinct from those present in humans, and another parasite lineage related to P. malariae. Phylogenetic analyses based on this diverse set of Plasmodium parasites in African Apes shed new light on the evolutionary history of P. falciparum. The data suggested that P. falciparum did not originate from P. reichenowi of chimpanzees (Pan troglodytes), but rather evolved in bonobos (Pan paniscus), from which it subsequently colonized humans by a host-switch. Finally, our data and that of others indicated that chimpanzees and bonobos maintain malaria parasites, to which humans are susceptible, a factor of some relevance to the renewed efforts to eradicate malaria.

Plasmodium vivax is the most prevalent human malaria parasite in the Americas. Previous studies have contrasted the genetic diversity of parasite populations in the Americas with those in Asia and Oceania, concluding that New World populations exhibit low genetic diversity consistent with a recent introduction. Here we used an expanded sample of complete mitochondrial genome sequences to investigate the diversity of P. vivax in the Americas as well as in other continental populations. We show that the diversity of P. vivax in the Americas is comparable to that in Asia and Oceania, and we identify several divergent clades circulating in South America that may have resulted from independent introductions. In particular, we show that several haplotypes sampled in Venezuela and northeastern Brazil belong to a clade that diverged from the other P. vivax lineages at least 30,000 years ago, albeit not necessarily in the Americas. We propose that, unlike in Asia where human migration increases local genetic diversity, the combined effects of the geographical structure and the low incidence of vivax malaria in the Americas has resulted in patterns of low local but high regional genetic diversity. This could explain previous views that P. vivax in the Americas has low genetic diversity because these were based on studies carried out in limited areas. Further elucidation of the complex geographical pattern of P. vivax variation will be important both for diversity assessments of genes encoding candidate vaccine antigens and in the formulation of control and surveillance measures aimed at malaria elimination.

Malaria remains endemic in several countries of South America with low to moderate transmission intensity. Regional human migration through underserved endemic areas may be responsible for significant parasite dispersion making the disease resilient to interventions. Thus, the genetic characterization of malarial parasites is an important tool to assess how endemic areas may connect via the movement of infected individuals. Here, four sites in geographically separated areas reporting 80% of the malaria morbidity in Colombia were studied. The sites are located on an imaginary transect line of 1,500 km from the northwest to the south Pacific Coast of Colombia with a minimal distance of 500 km between populations that display noticeable ethnic, economic, epidemiological, and ecological differences.

Plasmodium vivax is a predominant species of malaria in parts of South America and there is increasing resistance to drugs to treat infections by P. vivax. The existence of latent hypnozoites further complicates the ability to classify recurrent infections as treatment failures due to relapse, recrudescence of hyponozoites or re-infections. Antigen loci are putatively under natural selection and may not be an optimal molecular marker to define parasite haplotypes in paired samples. Putatively neutral microsatellite loci, however, offer an assessment of neutral haplotypes. The objective here was to assess the utility of neutral microsatellite loci to reconcile cases of recurrent parasitaemia in Amazonian P. vivax populations in Peru.