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Scientific Reports Feb 2023The zoonotic Plasmodium knowlesi parasite is a growing public health concern in Southeast Asia, especially in Malaysia, where elimination of P. falciparum and P. vivax...
The zoonotic Plasmodium knowlesi parasite is a growing public health concern in Southeast Asia, especially in Malaysia, where elimination of P. falciparum and P. vivax malaria has been the focus of control efforts. Understanding of the genetic diversity of P. knowlesi parasites can provide insights into its evolution, population structure, diagnostics, transmission dynamics, and the emergence of drug resistance. Previous work has revealed that P. knowlesi fall into three main sub-populations distinguished by a combination of geographical location and macaque host (Macaca fascicularis and M. nemestrina). It has been shown that Malaysian Borneo groups display profound heterogeneity with long regions of high or low divergence resulting in mosaic patterns between sub-populations, with some evidence of chromosomal-segment exchanges. However, the genetic structure of non-Borneo sub-populations is less clear. By gathering one of the largest collections of P. knowlesi whole-genome sequencing data, we studied structural genomic changes across sub-populations, with the analysis revealing differences in Borneo clusters linked to mosquito-related stages of the parasite cycle, in contrast to differences in host-related stages for the Peninsular group. Our work identifies new genetic exchange events, including introgressions between Malaysian Peninsular and M. nemestrina-associated clusters on various chromosomes, including in parasite invasion genes (DBP[Formula: see text], NBPX[Formula: see text] and NBPX[Formula: see text]), and important proteins expressed in the vertebrate parasite stages. Recombination events appear to have occurred between the Peninsular and M. fascicularis-associated groups, including in the DBP[Formula: see text] and DBP[Formula: see text] invasion associated genes. Overall, our work finds that genetic exchange events have occurred among the recognised contemporary groups of P. knowlesi parasites during their evolutionary history, leading to apparent mosaicism between these sub-populations. These findings generate new hypotheses relevant to parasite evolutionary biology and P. knowlesi epidemiology, which can inform malaria control approaches to containing the impact of zoonotic malaria on human communities.
Topics: Animals; Humans; Genetic Variation; Plasmodium knowlesi; Macaca fascicularis; Malaria; Malaysia; Malaria, Vivax; Malaria, Falciparum; Genetics, Population; Selection, Genetic
PubMed: 36750737
DOI: 10.1038/s41598-023-29368-4 -
Briefings in Functional Genomics Sep 2019Two simian malaria parasite species, Plasmodium knowlesi and Plasmodium cynomolgi, cause zoonotic infections in Southeast Asia, and they have therefore gained... (Review)
Review
Two simian malaria parasite species, Plasmodium knowlesi and Plasmodium cynomolgi, cause zoonotic infections in Southeast Asia, and they have therefore gained recognition among scientists and public health officials. Notwithstanding, these species and others including Plasmodium coatneyi have served for decades as sources of knowledge on the biology, genetics and evolution of Plasmodium, and the diverse ramifications and outcomes of malaria in their monkey hosts. Experimental analysis of these species can help to fill gaps in knowledge beyond what may be possible studying the human malaria parasites or rodent parasite species. The genome sequences for these simian malaria parasite species were reported during the last decade, and functional genomics research has since been pursued. Here research on the functional genomics analysis involving these species is summarized and their importance is stressed, particularly for understanding host-parasite interactions, and potentially testing novel interventions. Importantly, while Plasmodium falciparum and Plasmodium vivax can be studied in small New World monkeys, the simian malaria parasites can be studied more effectively in the larger Old World monkey macaque hosts, which are more closely related to humans. In addition to ex vivo analyses, experimental scenarios can include passage through Anopheline mosquito hosts and longitudinal infections in monkeys to study acute and chronic infections, as well as relapses, all in the context of the in vivo host environment. Such experiments provide opportunities for understanding functional genomic elements that govern host-parasite interactions, immunity and pathogenesis in-depth, addressing hypotheses not possible from in vitro cultures or cross-sectional clinical studies with humans.
Topics: Animals; Genomics; Host-Parasite Interactions; Humans; Plasmodium; Plasmodium cynomolgi; Plasmodium falciparum; Plasmodium knowlesi; Plasmodium vivax; Primates; Systems Biology
PubMed: 31241151
DOI: 10.1093/bfgp/elz013 -
Parasitology Jan 2018Plasmodium knowlesi is increasingly recognized as a major cause of malaria in Southeast Asia. Anopheles leucosphyrous group mosquitoes transmit the parasite and natural...
Plasmodium knowlesi is increasingly recognized as a major cause of malaria in Southeast Asia. Anopheles leucosphyrous group mosquitoes transmit the parasite and natural hosts include long-tailed and pig-tailed macaques. Despite early laboratory experiments demonstrating successful passage of infection between humans, the true role that humans play in P. knowlesi epidemiology remains unclear. The threat posed by its introduction into immunologically naïve populations is unknown despite being a public health priority for this region. A two-host species mathematical model was constructed to analyse this threat. Global sensitivity analysis using Monte Carlo methods highlighted the biological processes of greatest influence to transmission. These included parameters known to be influential in classic mosquito-borne disease models (e.g. vector longevity); however, interesting ecological components that are specific to this system were also highlighted: while local vectors likely have intrinsic preferences for certain host species, how plastic these preferences are, and how this is shaped by local conditions, are key determinants of parasite transmission potential. Invasion analysis demonstrates that this behavioural plasticity can qualitatively impact the probability of an epidemic sparked by imported infection. Identifying key vector sub/species and studying their biting behaviours constitute important next steps before models can better assist in strategizing disease control.
Topics: Animals; Anopheles; Host-Parasite Interactions; Humans; Macaca; Malaria; Models, Biological; Monkey Diseases; Monte Carlo Method; Mosquito Vectors; Plasmodium knowlesi
PubMed: 28345507
DOI: 10.1017/S0031182016002456 -
Malaria Journal Jun 2021Plasmodium knowlesi is now the major cause of human malaria in Malaysia, complicating malaria control efforts that must attend to the elimination of multiple Plasmodium...
BACKGROUND
Plasmodium knowlesi is now the major cause of human malaria in Malaysia, complicating malaria control efforts that must attend to the elimination of multiple Plasmodium species. Recent advances in the cultivation of P. knowlesi erythrocytic-stage parasites in vitro, transformation with exogenous DNA, and infection of mosquitoes with gametocytes from culture have opened up studies of this pathogen without the need for resource-intensive and costly non-human primate (NHP) models. For further understanding and development of methods for parasite transformation in malaria research, this study examined the activity of various trans-species transcriptional control sequences and the influence of Plasmodium vivax centromeric (pvcen) repeats in plasmid-transfected P. knowlesi parasites.
METHODS
In vitro cultivated P. knowlesi parasites were transfected with plasmid constructs that incorporated Plasmodium vivax or Plasmodium falciparum 5' UTRs driving the expression of bioluminescence markers (firefly luciferase or Nanoluc). Promoter activities were assessed by bioluminescence, and parasites transformed with human resistant allele dihydrofolate reductase-expressing plasmids were selected using antifolates. The stability of transformants carrying pvcen-stabilized episomes was assessed by bioluminescence over a complete parasite life cycle through a rhesus macaque monkey, mosquitoes, and a second rhesus monkey.
RESULTS
Luciferase expression assessments show that certain P. vivax promoter regions, not functional in the more evolutionarily-distant P. falciparum, can drive transgene expression in P. knowlesi. Further, pvcen repeats may improve the stability of episomal plasmids in P. knowlesi and support detection of NanoLuc-expressing elements over the full parasite life cycle from rhesus macaque monkeys to Anopheles dirus mosquitoes and back again to monkeys. In assays of drug responses to chloroquine, G418 and WR9910, anti-malarial half-inhibitory concentration (IC) values of blood stages measured by NanoLuc activity proved comparable to IC values measured by the standard SYBR Green method.
CONCLUSION
All three P. vivax promoters tested in this study functioned in P. knowlesi, whereas two of the three were inactive in P. falciparum. NanoLuc-expressing, centromere-stabilized plasmids may support high-throughput screenings of P. knowlesi for new anti-malarial agents, including compounds that can block the development of mosquito- and/or liver-stage parasites.
Topics: Centromere; Luciferases; Microorganisms, Genetically-Modified; Plasmids; Plasmodium knowlesi; Plasmodium vivax; Promoter Regions, Genetic
PubMed: 34090438
DOI: 10.1186/s12936-021-03773-4 -
Scientific Reports Apr 2018The Plasmodium falciparum apical asparagine (Asn)-rich protein (AARP) is one of malarial proteins, and it has been studied as a candidate of malaria subunit vaccine....
The Plasmodium falciparum apical asparagine (Asn)-rich protein (AARP) is one of malarial proteins, and it has been studied as a candidate of malaria subunit vaccine. Basic characterization of PvAARP has been performed with a focus on its immunogenicity and localization. In this study, we further analyzed the immunogenicity of PvAARP, focusing on the longevity of the antibody response, cross-species immunity and invasion inhibitory activity by using the primate malaria parasite Plasmodium knowlesi. We found that vivax malaria patient sera retained anti-PvAARP antibodies for at least one year without re-infection. Recombinant PvAARP protein was strongly recognized by knowlesi malaria patients. Antibody raised against the P. vivax and P. knowlesi AARP N-termini reacted with the apical side of the P. knowlesi merozoites and inhibited erythrocyte invasion by P. knowlesi in a concentration-dependent manner, thereby suggesting a cross-species nature of anti-PvAARP antibody against PkAARP. These results can be explained by B cell epitopes predicted in conserved surface-exposed regions of the AARP N-terminus in both species. The long-lived anti-PvAARP antibody response, cross-reactivity, and invasion inhibitory activity of anti-PvAARP support a critical role of AARP during the erythrocyte invasion and suggest that PvAARP induces long-lived cross-species protective immunity against P. vivax and P. knowlesi.
Topics: Animals; Antibodies, Protozoan; Antigens, Protozoan; Cross Reactions; Female; Humans; Malaria; Male; Mice; Plasmodium knowlesi; Plasmodium vivax; Sequence Analysis, Protein
PubMed: 29636493
DOI: 10.1038/s41598-018-23728-1 -
PLoS Neglected Tropical Diseases Dec 2020Plasmodium knowlesi, a simian malaria parasite, has been in the limelight since a large focus of human P. knowlesi infection was reported from Sarawak (Malaysian Borneo)... (Review)
Review
Plasmodium knowlesi, a simian malaria parasite, has been in the limelight since a large focus of human P. knowlesi infection was reported from Sarawak (Malaysian Borneo) in 2004. Although this infection is transmitted across Southeast Asia, the largest number of cases has been reported from Malaysia. The increasing number of knowlesi malaria cases has been attributed to the use of molecular tools for detection, but environmental changes including deforestation likely play a major role by increasing human exposure to vector mosquitoes, which coexist with the macaque host. In addition, with the reduction in human malaria transmission in Southeast Asia, it is possible that human populations are at a greater risk of P. knowlesi infection due to diminishing cross-species immunity. Furthermore, the possibility of increasing exposure of humans to other simian Plasmodium parasites such as Plasmodium cynomolgi and Plasmodium inui should not be ignored. We here review the current status of these parasites in humans, macaques, and mosquitoes to support necessary reorientation of malaria control and elimination in the affected areas.
Topics: Animals; Asia, Southeastern; Humans; Macaca; Malaria; Monkey Diseases; Mosquito Vectors; Plasmodium knowlesi
PubMed: 33382697
DOI: 10.1371/journal.pntd.0008900 -
Cell Reports. Medicine Jun 2022Serological markers are a promising tool for surveillance and targeted interventions for Plasmodium vivax malaria. P. vivax is closely related to the zoonotic parasite...
Serological markers are a promising tool for surveillance and targeted interventions for Plasmodium vivax malaria. P. vivax is closely related to the zoonotic parasite P. knowlesi, which also infects humans. P. vivax and P. knowlesi are co-endemic across much of South East Asia, making it important to design serological markers that minimize cross-reactivity in this region. To determine the degree of IgG cross-reactivity against a panel of P. vivax serological markers, we assayed samples from human patients with P. knowlesi malaria. IgG antibody reactivity is high against P. vivax proteins with high sequence identity with their P. knowlesi ortholog. IgG reactivity peaks at 7 days post-P. knowlesi infection and is short-lived, with minimal responses 1 year post-infection. We designed a panel of eight P. vivax proteins with low levels of cross-reactivity with P. knowlesi. This panel can accurately classify recent P. vivax infections while reducing misclassification of recent P. knowlesi infections.
Topics: Humans; Immunoglobulin G; Malaria; Malaria, Vivax; Plasmodium knowlesi; Plasmodium vivax
PubMed: 35732155
DOI: 10.1016/j.xcrm.2022.100662 -
Parasites & Vectors Dec 2018In recent years, human infection by the simian malaria parasite Plasmodium knowlesi has increased in Southeast Asia, leading to growing concerns regarding the...
BACKGROUND
In recent years, human infection by the simian malaria parasite Plasmodium knowlesi has increased in Southeast Asia, leading to growing concerns regarding the cross-species spread of the parasite. Consequently, a deeper understanding of the biology of P. knowlesi is necessary in order to develop tools for control of the emerging disease. TatD-like DNase expressed at the surface of P. falciparum has recently been shown to counteract host innate immunity and is thus a potential malaria vaccine candidate.
METHODS
The expression of the TatD DNase of P. knowlesi (PkTatD) was confirmed by both Western-blot and immunofluorescent assay. The DNA catalytic function of the PkTatD was confirmed by digestion of DNA with the recombinant PkTatD protein in the presence of various irons.
RESULTS
In the present study, we investigated the expression of the homologous DNase in P. knowlesi. The expression of TatD-like DNase in P. knowslesi (PkTatD) was verified by Western blot and indirect immunofluorescence assays. Like that of the P. falciparum parasite, PkTatD was also found to be located on the surface of erythrocytes infected by the parasites. Biochemical analysis indicated that PkTatD can hydrolyze DNA and this activity is magnesium-dependent.
CONCLUSIONS
We identified that PkTatD expressed on the surface of P. knowlesi-infected RBCs is a Mg-dependent DNase and exhibits a stronger hydrolytic capacity than TatD from P. falciparum. The data support our previous findings that TatD-like DNase is a unanimously expressed virulence factor of Plasmodium parasites.
Topics: Amino Acid Sequence; DNA, Protozoan; Deoxyribonucleases; Erythrocytes; Humans; Malaria, Falciparum; Plasmodium knowlesi; Protozoan Proteins; Sequence Alignment
PubMed: 30541605
DOI: 10.1186/s13071-018-3251-4 -
Genes Oct 2022The simian malaria parasite causes a high number of zoonotic infections in Malaysia. The thrombospondin-related apical merozoite protein (TRAMP) is an essential ligand...
The simian malaria parasite causes a high number of zoonotic infections in Malaysia. The thrombospondin-related apical merozoite protein (TRAMP) is an essential ligand for binding to the erythrocyte cell surface, whereby it facilitates the invasion. This study is the first attempt to determine the genetic diversity, phylogeography, natural selection and population structure from 97 full-length gene sequences originating from Malaysia. We found low levels of nucleotide diversity (π~0.0065) for the full-length gene despite samples originating from geographically separated regions (i.e., Peninsular Malaysia and Malaysian Borneo). The rate of synonymous substitutions was significantly higher than that of non-synonymous substitutions, indicating a purifying selection for the full-length gene within the clinical samples. The population genetic analysis revealed that the parasite population is undergoing a significant population expansion. The analysis of the amino acid sequence alignment of 97 sequences identified 15 haplotypes, of which a major shared haplotype was noted Hap 1 ( = 68, Sarawak; = 34, Sabah; = 12, Peninsular Malaysia; = 22). The phylogenetic analysis using DNA sequences identified two clusters that separated due to geographical distance and three mixed clusters with samples from both Peninsular Malaysia and Malaysian Borneo. Population structure analyses indicated two distinct sub-populations (K = 2). Our findings point to the potential for independent parasite evolution, which could make zoonotic malaria control and elimination even more challenging.
Topics: Animals; Humans; Plasmodium knowlesi; Merozoites; Phylogeny; Thrombospondins; Protozoan Proteins; Genetic Variation; Sequence Analysis, DNA; Malaria; Genetics, Population
PubMed: 36360181
DOI: 10.3390/genes13111944 -
Malaria Journal Feb 2021Plasmodium falciparum malaria increases plasma levels of the cytokine Fms-like tyrosine kinase 3 ligand (Flt3L), a haematopoietic factor associated with dendritic cell...
BACKGROUND
Plasmodium falciparum malaria increases plasma levels of the cytokine Fms-like tyrosine kinase 3 ligand (Flt3L), a haematopoietic factor associated with dendritic cell (DC) expansion. It is unknown if the zoonotic parasite Plasmodium knowlesi impacts Flt3L or DC in human malaria. This study investigated circulating DC and Flt3L associations in adult malaria and in submicroscopic experimental infection.
METHODS
Plasma Flt3L concentration and blood CD141 DC, CD1c DC and plasmacytoid DC (pDC) numbers were assessed in (i) volunteers experimentally infected with P. falciparum and in Malaysian patients with uncomplicated (ii) P. falciparum or (iii) P. knowlesi malaria.
RESULTS
Plasmodium knowlesi caused a decline in all circulating DC subsets in adults with malaria. Plasma Flt3L was elevated in acute P. falciparum and P. knowlesi malaria with no increase in a subclinical experimental infection. Circulating CD141 DCs, CD1c DCs and pDCs declined in all adults tested, for the first time extending the finding of DC subset decline in acute malaria to the zoonotic parasite P. knowlesi.
CONCLUSIONS
In adults, submicroscopic Plasmodium infection causes no change in plasma Flt3L but does reduce circulating DCs. Plasma Flt3L concentrations increase in acute malaria, yet this increase is insufficient to restore or expand circulating CD141 DCs, CD1c DCs or pDCs. These data imply that haematopoietic factors, yet to be identified and not Flt3L, involved in the sensing/maintenance of circulating DC are impacted by malaria and a submicroscopic infection. The zoonotic P. knowlesi is similar to other Plasmodium spp in compromising DC in adult malaria.
Topics: Acute Disease; Adult; Dendritic Cells; Female; Humans; Malaria; Malaria, Falciparum; Male; Membrane Proteins; Middle Aged; Plasma; Plasmodium falciparum; Plasmodium knowlesi; Young Adult
PubMed: 33593383
DOI: 10.1186/s12936-021-03642-0