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Background: To understand how Plasmodium falciparum malaria is controlled, it is essential to elucidate the transcriptomic responses of the human host in naturally-exposed populations. Various individual studies of the human transcriptomic responses to naturally transmitted P. falciparum infections have been reported with varying results. Multicohort gene expression analysis by aggregating data from diverse populations into a single analysis will increase the reproducibility and reliability of the results. Methods: In this study, discovery cohorts GSE1124-GPL96, GSE34404, GSE117613, and validation cohort GSE35858 were obtained from the Gene Expression Omnibus. A meta-analysis using data from the multicohort studies was performed to identify the differentially expressed genes (DEGs) between malaria-infected and noninfected individuals using the MetaIntegrator R package. Subsequently, the protein–protein interaction (PPI) networks of the DEGs were constructed using Cytoscape software. Significant modules were selected, and the hub genes were identified using the CytoHubba and MCODE plug-ins. Multicohort WGCNA was conducted to find a correlation between modules and malaria infection. Furthermore, the immune cell profile of the peripheral blood in different groups was identified using ssGSEA. Results: These analyses reveal that neutrophil activation, neutrophil-mediated immunity, and neutrophil degranulation are involved in the human response to natural malaria infection. However, neutrophil cell enrichment and activation were not significantly different between mild malaria and severe malaria groups. Malaria infection also downregulates host genes in ribosome synthesis and protein translation and upregulates host cell division-related genes. Furthermore, immune cell profiling analysis shows that activated dendritic cells and type 2 T helper cells are upregulated, while activated B cells, immature B cells, and monocytes are downregulated in the malaria-infected patients relative to the noninfected individuals. Significantly higher enrichment of activated dendritic cell-related genes and significantly lower enrichment of monocyte-related genes are also observed in the peripheral blood of the severe malaria group than in the mild malaria group. Conclusion: These results reveal important molecular signatures of host responses to malaria infections, providing some bases for developing malaria control strategies and protective vaccines. [ABSTRACT FROM AUTHOR]

Genetic mapping has been widely employed to search for genes linked to phenotypes/traits of interest. Because of the ease of maintaining rodent malaria parasites in laboratory mice, many genetic crosses of rodent malaria parasites have been performed to map the parasite genes contributing to malaria parasite development, drug resistance, host immune response, and disease pathogenesis. Drs. Richard Carter, David Walliker, and colleagues at the University of Edinburgh, UK, were the pioneers in developing the systems for genetic mapping of malaria parasite traits, including characterization of genetic markers to follow the inheritance and recombination of parasite chromosomes and performing the first genetic cross using rodent malaria parasites. Additionally, many genetic crosses of inbred mice have been performed to link mouse chromosomal loci to the susceptibility to malaria parasite infections. In this chapter, we review and discuss past and recent advances in genetic marker development, performing genetic crosses, and genetic mapping of both parasite and host genes. Genetic mappings using models of rodent malaria parasites and inbred mice have contributed greatly to our understanding of malaria, including parasite development within their hosts, mechanism of drug resistance, and host-parasite interaction. [ABSTRACT FROM AUTHOR]

Drug resistance in malaria parasites is a serious public health burden, and resistance to most of the antimalarial drugs currently in use has been reported. A better understanding of the molecular mechanisms of drug resistance is urgently needed to slow or circumvent the spread of resistance, to allow local treatments to be deployed more effectively to prolong the life span of the current drugs, and to develop new drugs. Although mutations in genes determining resistance to drugs such as chloroquine and the antifolates have been identified, we still do not have a full understanding of the resistance mechanisms, and genes that contribute to resistance to many other drugs remain to be discovered. Genetic mapping is a powerful tool for the identification of mutations conferring drug resistance in malaria parasites because most drug-resistant phenotypes were selected within the past 60 years. High-throughput methods for genotyping large numbers of single nucleotide polymorphisms (SNPs) and microsatellites (MSs) are now available or are being developed, and genome-wide association studies for malaria traits will soon become a reality. Here we discuss strategies and issues related to mapping genes contributing to drug resistance in the human malaria parasite Plasmodium falciparum. [ABSTRACT FROM AUTHOR]

The debate over whether the human malaria parasite Plasmodium falciparum underwent a recent severe population bottleneck (“Malaria''s Eve” hypothesis) has attracted great attention recently. Understanding the genetic diversity and evolutionary history of the parasite has practical implications for developing disease control measures. [Copyright &y& Elsevier]

Abstract Microsatellite loci are generally assumed to evolve via a stepwise mutational process and a battery of statistical techniques has been developed in recent years based on this or related mutation models. It is therefore important to investigate the appropriateness of these models in a wide variety of taxa. We used two approaches to examine mutation patterns in the malaria parasite Plasmodium falciparum: (i) we examined sequence variation at 12 tri-nucleotide repeat loci; and (ii) we analysed patterns of repeat structure and heterozygosity at 114 loci using data from 12 laboratory parasite lines. The sequencing study revealed complex patterns of mutation in five of the 12 loci studied. Alleles at two loci contain indels of 24 bp and 57 bp in flanking regions, while in the other three loci, blocks of imperfect microsatellites appear to be duplicated or inserted; these loci essentially consist of minisatellite repeats, with each repeat unit containing four to eight microsatellites. The survey of heterozygosity revealed a positive relationship between repeat number and microsatellite variability for both di- and trinucleotides, indicating a higher mutation rate in loci with longer repeat arrays. Comparisons of levels of variation in different repeat types indicate that the mutation rate of dinucleotide-bearing loci is 1.6–2.1 times faster than trinucleotides, consistent with the lower mean number of repeats in trinucleotide-bearing loci. However, despite the evidence that microsatellite arrays themselves are evolving in a manner consistent with stepwise mutation model in P. falciparum, the high frequency of complex mutations precludes the use of analytical tools based on this mutation model for many microsatellite-bearing loci in this protozoan. The results call into question the generality of models based on stepwise mutation for analysing microsatellite data, but also demonstrate the ease with which loci that violate model assumptions can be detected using minimal sequencing effort. [ABSTRACT FROM AUTHOR]

Examines the findings of a ...330 kDa protein with complex polymorphisms in chloroquine resistant parasites from Asia and Africa. What chloroquine was used for; What chloroquine resistance can be partially reversed by; Results of the study on the resistance mechanism of chloroquine.

Describes a large and extremely diverse family of Plasmodium falciparum genes (var) that encode 200-350 kDa proteins having the expected properties of antigenically variant adhesion molecules. Predicted amino acid sequences of var genes; 50-150 var genes on multiple parasite chromosomes; Two classes of transcripts in steady-state RNA.

Background: Eukaryotic genes contain introns that are removed by the spliceosomal machinery during mRNA maturation. Introns impose a huge energetic burden on a cell; therefore, they must play an essential role in maintaining genome stability and/or regulating gene expression. Many genes (> 50%) in Plasmodium parasites contain predicted introns, including introns in 5′ and 3′ untranslated regions (UTR). However, the roles of UTR introns in the gene expression of malaria parasites remain unknown. Methods: In this study, an episomal dual-luciferase assay was developed to evaluate gene expression driven by promoters with or without a 5′UTR intron from four Plasmodium yoelii genes. To investigate the effect of the 5′UTR intron on endogenous gene expression, the pytctp gene was tagged with 3xHA at the N-terminal of the coding region, and parasites with or without the 5′UTR intron were generated using the CRISPR/Cas9 system. Results: We showed that promoters with 5′UTR introns had higher activities in driving gene expression than those without 5′UTR introns. The results were confirmed in recombinant parasites expressing an HA-tagged gene (pytctp) driven by promoter with or without 5′UTR intron. The enhancement of gene expression was intron size dependent, but not the DNA sequence, e.g. the longer the intron, the higher levels of expression. Similar results were observed when a promoter from one strain of P. yoelii was introduced into different parasite strains. Finally, the 5′UTR introns were alternatively spliced in different parasite development stages, suggesting an active mechanism employed by the parasites to regulate gene expression in various developmental stages. Conclusions: Plasmodium 5′UTR introns enhance gene expression in a size-dependent manner; the presence of alternatively spliced mRNAs in different parasite developmental stages suggests that alternative slicing of 5′UTR introns is one of the key mechanisms in regulating parasite gene expression and differentiation. [ABSTRACT FROM AUTHOR]

Examines the genetic map and recombination parameters of the human malaria parasite, Plasmodium falciparum. Inheritance patterns within the linkage groups; Analysis of the haplotypes; Details on the meiotic crossover activity in the genome.

The author talks about how his laboratory at the U.S. National Institutes of Health (NIH) is developing disease-control measures and medicines to help cure malaria. Topics discussed include the global incidence of malaria, the laboratory's partnership with National Center for Advancing Translational Sciences (NCATS) scientists in identifying potential new medicines to treat the disease, and the development of vaccines to stop malaria infection.

Artemisinin and its derivatives (ART) are crucial first-line antimalarial drugs that rapidly clear parasitemia, but recrudescences of the infection frequently follow ART monotherapy. For this reason, ART must be used in combination with one or more partner drugs that ensure complete cure. The ability of malaria parasites to survive ART monotherapy may relate to an innate growth bistability phenomenon whereby a fraction of the drug-exposed population enters into metabolic quiescence (dormancy) as persister forms. Characterization of the events that underlie entry and waking from persistence may lead to lasting breakthroughs in malaria chemotherapy that can prevent recrudescences and protect the future of ART-based combination therapies. Artemisinin treatment clears malaria parasitemia rapidly but is frequently followed by recrudescent infection unless the artemisinin (or artemisinin derivative) is combined with an effective partner drug. Recrudescence was identified as RI type artemisinin resistance in 1984, when artemisinin-based combination therapy (ACT) was also proposed. RI type artemisinin resistance is innate to Plasmodium falciparum and has not progressed to higher levels (RII or RIII), despite decades of ACT distributions and partner drug failures. Recrudescences from persisters are not associated with parasite clearance times after artemisinin treatment. Persistence as a survival mechanism is distinct from drug resistance acquired by genetic mutations, a distinction that is at risk of being lost when persisters are incorrectly distinguished as 'the drug-resistant parasites'. Understanding and targeting persisters may suggest strategies for lasting breakthroughs in malaria chemotherapy. [ABSTRACT FROM AUTHOR]

The World Health Organization estimated that more than 1.5 billion people are infected with soil-transmitted helminths globally, and foodborne trematodiases in humans cause ∼2 million life-years lost to disability and death worldwide every year. Investment in prevention, treatment, and awareness of helminth infections and discovery of new, safe, effective, and affordable anti-helminth drugs are urgently needed. Artemisinin (ART) and its derivatives have been widely used to treat malaria and other protozoan infections; they also possess activities against helminths. So far, many papers on ART and its derivatives against schistosomal infections have been reported and reviewed. This review attempts to summarize recent advances in the uses of ART and its derivatives to treat infections of helminth parasites other than Schistosoma spp. in both humans and animals, including nematodes ( Toxocara canis , Trichinella spiralis, Haemonchus contortus , Meloidogyne spp., Globodera rostochiensis , and Xiphinema index ), cestodes ( Echinococcus spp. and Taenia crassiceps ), trematodes ( Echinostoma spp., Fasciola spp., Clonorchis sinensis, Opisthorchis viverrini , Paragonimus westermani , Heterophyes heterophyes , and Paramphistomum microbothrium ), and monogenea parasites ( Dactylogyrus and Gyrodactylus ). We concluded that ART and its derivatives are potentially effective drugs for treating various helminthic diseases of public health significance. [ABSTRACT FROM AUTHOR]

Galectins is a family of multifunctional lectins. Fifteen galectins have been identified from a variety of cells and tissues of vertebrates and invertebrates. Galectins have been shown to play pivotal roles in host–pathogen interaction such as adhesion of pathogens to host cells and activation of host innate and adaptive immunity. In recent years, the roles of galectins during parasite infections have gained increasing attention. Galectins produced by different hosts can act as pattern recognition receptors detecting conserved pathogen-associated molecular patterns of parasites, while galectins produced by parasites can modulate host responses. This review summarizes some recent studies on the roles of galectins produced by parasitic protozoa, nematodes, and trematodes and their hosts. Understanding the roles of galectins in host–parasite interactions may provide targets for immune intervention and therapies of parasitic infections. [ABSTRACT FROM AUTHOR]

Cyclic GMP‐AMP synthase (cGAS) and stimulator of interferon genes (STING) play critical roles in the innate immunity against infectious diseases and are required to link pathogen DNA sensing to immune responses. However, the mechanisms by which cGAS‐STING‐induced cytokines suppress the adaptive immune response against malaria infections remain poorly understood. Here, cGAS‐STING signaling is identified to play a detrimental role in regulating anti‐malaria immunity. cGAS or STING deficiency in mice markedly prolongs mouse survival during lethal malaria Plasmodium yoelii nigeriensis N67C infections by reducing late interleukin (IL)‐6 production. Mechanistically, cGAS/STING recruits myeloid differentiation factor 88 (MyD88) and specifically induces the p38‐dependent signaling pathway for late IL‐6 production, which, in turn, expands CD11b+Ly6Chi proinflammatory monocytes to inhibit immunity. Moreover, the blockage or ablation of the cGAS‐STING‐MyD88‐p38‐IL‐6 signaling axis or the depletion of CD11b+Ly6Chi proinflammatory monocytes provides mice a significant survival benefit during N67C and other lethal malaria‐strain infections. Taken together, these findings identify a previously unrecognized detrimental role of cGAS‐STING‐MyD88‐p38 axis in infectious diseases through triggering the late IL‐6 production and proinflammatory monocyte expansion and provide insight into how targeting the DNA sensing pathway, dysregulated cytokines, and proinflammatory monocytes enhances immunity against infection. [ABSTRACT FROM AUTHOR]

Cyclic GMP‐AMP synthase (cGAS) and stimulator of interferon genes (STING) play critical roles in the innate immunity against infectious diseases and are required to link pathogen DNA sensing to immune responses. However, the mechanisms by which cGAS‐STING‐induced cytokines suppress the adaptive immune response against malaria infections remain poorly understood. Here, cGAS‐STING signaling is identified to play a detrimental role in regulating anti‐malaria immunity. cGAS or STING deficiency in mice markedly prolongs mouse survival during lethal malaria Plasmodium yoelii nigeriensis N67C infections by reducing late interleukin (IL)‐6 production. Mechanistically, cGAS/STING recruits myeloid differentiation factor 88 (MyD88) and specifically induces the p38‐dependent signaling pathway for late IL‐6 production, which, in turn, expands CD11b+Ly6Chi proinflammatory monocytes to inhibit immunity. Moreover, the blockage or ablation of the cGAS‐STING‐MyD88‐p38‐IL‐6 signaling axis or the depletion of CD11b+Ly6Chi proinflammatory monocytes provides mice a significant survival benefit during N67C and other lethal malaria‐strain infections. Taken together, these findings identify a previously unrecognized detrimental role of cGAS‐STING‐MyD88‐p38 axis in infectious diseases through triggering the late IL‐6 production and proinflammatory monocyte expansion and provide insight into how targeting the DNA sensing pathway, dysregulated cytokines, and proinflammatory monocytes enhances immunity against infection. [ABSTRACT FROM AUTHOR]

Highlights: [•] This is the first study that identified gene-related microsatellite of Biomphalaria glabrata using data mining. [•] Genetic linkage map construction based on F2 progenies between resistant and susceptible snails coding B. glabrata markers. [Copyright &y& Elsevier]

Plasmodium falciparum malaria, an infectious disease caused by a parasitic protozoan, claims the lives of nearly a million children each year in Africa alone and is a top public health concern. Evidence is accumulating that resistance to artemisinin derivatives, the frontline therapy for the asexual blood stage of the infection, is developing in southeast Asia. Renewed initiatives to eliminate malaria will benefit from an expanded repertoire of antimalarials, including new drugs that kill circulating P. falciparum gametocytes, thereby preventing transmission. Our current understanding of the biology of asexual blood-stage parasites and gametocytes and the ability to culture them in vitro lends optimism that high-throughput screenings of large chemical libraries will produce a new generation of antimalarial drugs. There is also a need for new therapies to reduce the high mortality of severe malaria. An understanding of the pathophysiology of severe disease may identify rational targets for drugs that improve survival. [ABSTRACT FROM AUTHOR]

Background: Rodent malaria parasites are important models for studying host-malaria parasite interactions such as host immune response, mechanisms of parasite evasion of host killing, and vaccine development. One of the rodent malaria parasites is Plasmodium yoelii, and multiple P. yoelii strains or subspecies that cause different disease phenotypes have been widely employed in various studies. The genomes and transcriptomes of several P. yoelii strains have been analyzed and annotated, including the lethal strains of P. y. yoelii YM (or 17XL) and non-lethal strains of P. y. yoelii 17XNL/17X. Genomic DNA sequences and cDNA reads from another subspecies P. y. nigeriensis N67 have been reported for studies of genetic polymorphisms and parasite response to drugs, but its genome has not been assembled and annotated. Results: We performed genome sequencing of the N67 parasite using the PacBio long-read sequencing technology, de novo assembled its genome and transcriptome, and predicted 5383 genes with high overall annotation quality. Comparison of the annotated genome of the N67 parasite with those of YM and 17X parasites revealed a set of genes with N67-specific orthology, expansion of gene families, particularly the homologs of the Plasmodium chabaudi erythrocyte membrane antigen, large numbers of SNPs and indels, and proteins predicted to interact with host immune responses based on their functional domains. Conclusions: The genomes of N67 and 17X parasites are highly diverse, having approximately one polymorphic site per 50 base pairs of DNA. The annotated N67 genome and transcriptome provide searchable databases for fast retrieval of genes and proteins, which will greatly facilitate our efforts in studying the parasite biology and gene function and in developing effective control measures against malaria. [ABSTRACT FROM AUTHOR]

• Australian tea tree oil display a range of biological and pharmacological properties. • Tea tree oil and its 15 major constituents have effects on parasitic infections. • The parasites include protozoa, nematodes, cestodes, and monogenean parasites. • Prolonged survival or curative effect on protozoan infections in treated animals. • Helminthic burden was reduced or controlled in treated animals. Australian tea tree (Melaleuca alternifolia) oil (TTO) and its monoterpene constituents such as terpinen-4-ol (T4O), 1,8-cineole, limonene, p-cymene, and α-terpinene have been shown to be effective in controlling a wide range of parasitic infections. The anti-parasitic effects of these compounds are mainly due to their anti-histamine and anti-acetylcholinesterase activities as well as their ability to modulate host inflammatory responses. This review attempts to summarize recent advances in the uses of TTO and its 15 major monoterpene constituents in treating parasitic infections in both humans and animals. Activities against parasitic protozoans (Plasmodium falciparum, Leishmania spp., Trypanosoma spp., Acanthamoeba castellanii, Trichomonas vaginalis, Eimeria, and Ichthyophthirius multifiliis), nematodes (Haemonchus contortus and Anisakis simplex), cestode (Echinococcus ortleppi), and monogeneans (Gasterosteus spp. and Dactylogyrus minutus) have been reported, showing good potentials in treating parasitic infections. Further studies are necessary for developing anti-parasite therapies using TTO or its monoterpenes constituents. [ABSTRACT FROM AUTHOR]

Autochthonous Plasmodium falciparum malaria (PFM) in Central Europe has been reported repeatedly, transmission of the parasite being attributed to blood transfusion or imported P. falciparum-infected vectors. We report two cases of PFM in German children without travel history to malaria-endemic areas. Both infections occurred during a stay in a hospital where a child from Angola with chronic P. falciparum infection was hospitalized at the time. Known routes of transmission, such as imported mosquitoes or blood transfusion, were very unlikely or could be excluded, whereas evidence was obtained for transmission by the indigenous mosquito species Anopheles plumbeus. [ABSTRACT FROM AUTHOR]

• Plasmodium yoelii can repair CRISPR/Cas9-generated DNA breaks by microhomology-mediated end joining. • Size mutants of the central repeat in circumsporozoite surface protein (CSP) were successfully generated. • A minimum size of the CSP central repeat region is required for mosquito stage development. CRISPR/Cas9 is a powerful genome editing method that has greatly facilitated functional studies in many eukaryotic organisms including malaria parasites. Due to the lack of genes encoding enzymes necessary for the non-homologous end joining DNA repair pathway, genetic manipulation of malaria parasite genomes is generally accomplished through homologous recombination requiring the presence of DNA templates. Recently, an alternative double-strand break repair pathway, microhomology-mediated end joining, was found in the Plasmodium falciparum parasite. Taking advantage of the MMEJ pathway, we developed a MMEJ-based CRISPR/Cas9 (mCRISPR) strategy to efficiently generate multiple mutant parasites simultaneously in genes with repetitive sequences. As a proof of principle, we successfully produced various size mutants in the central repeat region of the Plasmodium yoelii circumsporozoite surface protein without the use of template DNA. Monitoring mixed parasite populations and individual parasites with different sizes of CSP-CRR showed that the CSP-CRR plays a role in the development of mosquito stages, with severe developmental defects in parasites with large deletions in the repeat region. However, the majority of the csp mutant parasite clones grew similarly to the wild type P. yoelii 17XL parasite in mice. This study develops a useful technique to efficiently generate mutant parasites with deletions or insertions, and shows that the CSP-CRR plays a role in parasite development in mosquito. [ABSTRACT FROM AUTHOR]

The RNA-guided endonuclease Cas9 has applied as an efficient gene-editing method in malaria parasite Plasmodium . However, the size (4.2 kb) of the commonly used Cas9 from Streptococcus pyogenes (SpCas9) limits its utility for genome editing in the parasites only introduced with cas9 plasmid. To establish the endogenous and constitutive expression of Cas9 protein in the rodent malaria parasite P. yoelii , we replaced the coding region of an endogenous gene sera1 with the intact SpCas9 coding sequence using the CRISPR/Cas9-mediated genome editing method, generating the cas9-knockin parasite ( PyCas9ki ) of the rodent malaria parasite P. yoelii . The resulted PyCas9ki parasite displays normal progression during the whole life cycle and possesses the Cas9 protein expression in asexual blood stage. By introducing the plasmid (pYCs) containing only sgRNA and homologous template elements, we successfully achieved both deletion and tagging modifications for different endogenous genes in the genome of PyCas9ki parasite. This cas9-knockin PyCas9ki parasite provides a new platform facilitating gene functions study in the rodent malaria parasite P. yoelii . [ABSTRACT FROM AUTHOR]

Malaria parasites invade red blood cells (RBCs), consume copious amounts of hemoglobin, and severely disrupt iron regulation in humans. Anemia often accompanies malaria disease; however, iron supplementation therapy inexplicably exacerbates malarial infections. Here we found that the iron exporter ferroportin (FPN) was highly abundant in RBCs, and iron supplementation suppressed its activity. Conditional deletion of the Fpn gene in erythroid cells resulted in accumulation of excess intracellular iron, cellular damage, hemolysis, and increased fatality in malaria-infected mice. In humans, a prevalent FPN mutation, Q248H (glutamine to histidine at position 248), prevented hepcidin-induced degradation of FPN and protected against severe malaria disease. FPN Q248H appears to have been positively selected in African populations in response to the impact of malaria disease. Thus, FPN protects RBCs against oxidative stress and malaria infection. [ABSTRACT FROM AUTHOR]

Parasitic protozoan diseases continue to rank among the world’s greatest global health problems, which are also common among poor populations. Currently available drugs for treatment present drawbacks, urging the need for more effective, safer, and cheaper drugs. Artemisinin (ART) and its derivatives are some of the most important classes of antimalarial agents originally derived from Artemisia annua L. However, besides the outstanding antimalarial and antischistosomal activities, ART and its derivatives also possess activities against other parasitic protozoa. In this paper we review the activities of ART and its derivatives against protozoan parasites in vitro and in vivo, including Leishmania spp., Trypanosoma spp., Toxoplasma gondii , Neospora caninum , Eimeria tenella , Acanthamoeba castellanii , Naegleria fowleri , Cryptosporidium parvum , Giardia lamblia , and Babesia spp. We conclude that ART and its derivatives may be good alternatives for treating other non-malarial protozoan infections in developing countries, although more studies are necessary before they can be applied clinically. [ABSTRACT FROM AUTHOR]

CRISPR/Cas9 has been successfully adapted for gene editing in malaria parasites including Plasmodium falciparum and Plasmodium yoelii . However, the reported methods were limited to editing one gene at a time. In practice, it is often desired to modify multiple genetic loci in a parasite genome. Here we describe a CRISPR/Cas9 mediated genome editing method that allows successive modification of more than one gene in the genome of P. yoelii using an improved single-vector system (pYCm) we developed previously. Drug resistant genes encoding human dihydrofolate reductase (hDHFR) and a yeast bifunctional protein (yFCU), with cytosine deaminase (CD) and uridyl phosphoribosyl transferase (UPRT) activities in the plasmid, allowed sequential positive (pyrimethamine, Pyr) and negative (5-fluorocytosine, 5FC) selections and generation of transgenic parasites free of the episomal plasmid after genetic modification. Using this system, we were able to efficiently tag a gene of interest ( Pyp28 ) and subsequently disrupted two genes ( Pyctrp and Pycdpk3 ) that are individually critical for ookinete motility. Disruption of the genes either eliminated ( Pyctrp ) or greatly reduced ( Pycdpk3 ) ookinete forward motility in matrigel in vitro and completely blocked oocyst development in mosquito midgut. The method will greatly facilitate studies of parasite gene function, development, and disease pathogenesis. [ABSTRACT FROM AUTHOR]

Summary Type I interferon (IFN) is critical for controlling pathogen infection; however, its regulatory mechanisms in plasmacytoid cells (pDCs) still remain unclear. Here, we have shown that nucleic acid sensors cGAS-, STING-, MDA5-, MAVS-, or transcription factor IRF3-deficient mice produced high amounts of type I IFN-α and IFN-β (IFN-α/β) in the serum and were resistant to lethal plasmodium yoelii YM infection. Robust IFN-α/β production was abolished when gene encoding nucleic acid sensor TLR7, signaling adaptor MyD88, or transcription factor IRF7 was ablated or pDCs were depleted. Further, we identified SOCS1 as a key negative regulator to inhibit MyD88-dependent type I IFN signaling in pDCs. Finally, we have demonstrated that pDCs, cDCs, and macrophages were required for generating IFN-α/β-induced subsequent protective immunity. Thus, our findings have identified a critical regulatory mechanism of type I IFN signaling in pDCs and stage-specific function of immune cells in generating potent immunity against lethal YM infection. [ABSTRACT FROM AUTHOR]

Both type I interferon (IFN-I) and CD40 play a significant role in various infectious diseases, including malaria and autoimmune disorders. CD40 is mostly known to function in adaptive immunity, but previous observations of elevated CD40 levels early after malaria infection of mice led us to investigate its roles in innate IFN-I responses and disease control. Using a Plasmodium yoelii nigeriensis N67 and C57BL/6 mouse model, we showed that infected CD40-/- mice had reduced STING and serum IFN-β levels day-2 post infection, higher day-4 parasitemia, and earlier deaths. CD40 could greatly enhance STING-stimulated luciferase signals driven by the IFN-β promoter in vitro, which was mediated by increased STING protein levels. The ability of CD40 to influence STING expression was confirmed in CD40-/- mice after malaria infection. Substitutions at CD40 TRAF binding domains significantly decreased the IFN-β signals and STING protein level, which was likely mediated by changes in STING ubiquitination and degradation. Increased levels of CD40, STING, and ISRE driven luciferase signal in RAW Lucia were observed after phagocytosis of N67-infected red blood cells (iRBCs), stimulation with parasite DNA/RNA, or with selected TLR ligands [LPS, poly(I:C), and Pam3CSK4]. The results suggest stimulation of CD40 expression by parasite materials through TLR signaling pathways, which was further confirmed in bone marrow derived dendritic cells/macrophages (BMDCs/BMDMs) and splenic DCs from CD40-/-, TLR3-/- TLR4-/-, TRIF-/-, and MyD88-/- mice after iRBC stimulation or parasite infection. Our data connect several signaling pathways consisting of phagocytosis of iRBCs, recognition of parasite DNA/RNA (possibly GPI) by TLRs, elevated levels of CD40 and STING proteins, increased IFN-I production, and longer host survival time. This study reveals previously unrecognized CD40 function in innate IFN-I responses and protective pathways in infections with malaria strains that induce a strong IFN-I response, which may provide important information for better understanding and management of malaria. [ABSTRACT FROM AUTHOR]

Rodent malaria parasites have been widely used in all aspects of malaria research to study parasite development within rodent and insect hosts, drug resistance, disease pathogenesis, host immune response, and vaccine efficacy. Rodent malaria parasites were isolated from African thicket rats and initially characterized by scientists at the University of Edinburgh, UK, particularly by Drs. Richard Carter, David Walliker, and colleagues. Through their efforts and elegant work, many rodent malaria parasite species, subspecies, and strains are now available. Because of the ease of maintaining these parasites in laboratory mice, genetic crosses can be performed to map the parasite and host genes contributing to parasite growth and disease severity. Recombinant DNA technologies are now available to manipulate the parasite genomes and to study gene functions efficiently. In this chapter, we provide a brief history of the isolation and species identification of rodent malaria parasites. We also discuss some recent studies to further characterize the different developing stages of the parasites including parasite genomes and chromosomes. Although there are differences between rodent and human malaria parasite infections, the knowledge gained from studies of rodent malaria parasites has contributed greatly to our understanding of and the fight against human malaria. [ABSTRACT FROM AUTHOR]

Leucocytozoon parasites infect many species of avian hosts, including domestic chicken, and can inflict heavy economic loss on the poultry industry. Two major species of Leucocytozoon parasites have been reported in China, L. sabrazesi and L. caulleryi, although L. sabrazesi appears to be more widespread than L. caulleryi in southern China. The traditional method for detecting Leucocytozoon infection is microscopic examination of blood smears for the presence of mature gametocytes in circulation, which may miss infections with low parasitemia (gametocytemia) or immature gametocytes. Here we developed a PCR-based method to monitor L. sabrazesi infections at seven sites in four provinces of China after testing two PCR primer pairs based on parasite mitochondrial cytochrome b (cytb) and cytochrome c oxidase III (coxIII) genes. We compared the results of PCR detection with those of microscopic observation. As expected, the PCR assays were more sensitive than microscope examination in detecting L. sabrazesi infection and were able to detect parasite DNA after gametocytes disappeared in the blood stream. Using these methods, we investigated monthly dynamics of L. sabrazesi in chickens from a free-range farm in Xiamen, Fujian province of China, over one year. Our results showed that chickens were infected with L. sabrazesi year-round in southern China. Finally, we tested several compounds for potential treatment of Leucocytozoon infections, including primaquine, ketotifen, clomipramine hydrochloride, desipramine hydrochloride, sulfaquinoxaline, and pyrimethamine. Only primaquine had activity against L. sabrazesi gametocytes. Our results provide important information for controlling parasite transmission in southern China and disease management. [ABSTRACT FROM AUTHOR]

Leucocytozoon parasites infect a large number of avian hosts, including domestic chicken, and cause significant economical loss to the poultry industry. Although the transmission stages of the parasites were observed in avian blood cells more than a century ago, the specific host cell type(s) that the gametocytes infect remain uncertain. Because all the avian blood cells, including red blood cells (RBCs), are nucleated, and the developing parasites dramatically change the morphology of the infected host cells, it has been difficult to identify Leucocytozoon infected host cell(s). Here we use cell-type specific antibodies to investigate the identities of the host cells infected by Leucocytozoon sabrazesi gametocytes. Anti-RBC antibodies stained RBCs membrane strongly, but not the parasite-infected cells, ruling out the possibility of RBCs being the infected host cells. Antibodies recognizing various leukocytes including heterophils, monocytes, lymphocytes, and macrophages did not stain the infected cells either. Antisera raised against a peptide of the parasite cytochrome B (CYTB) stained parasite-infected cells and some leukocytes, particularly cells with a single round nucleus as well as clear/pale cytoplasm suggestive of thrombocytes. Finally, a monoclonal antibody known to specifically bind chicken thrombocytes also stained the infected cells, confirming that L. sabrazesi gametocytes develop within chicken thrombocytes. The identification of L. sabrazesi infected host cell solves a long unresolved puzzle and provides important information for studying parasite invasion of host cells and for developing reagents to interrupt parasite transmission. [ABSTRACT FROM AUTHOR]

Summary Tissue-infiltrating Ly6C hi monocytes play diverse roles in immunity, ranging from pathogen killing to immune regulation. How and where this diversity of function is imposed remains poorly understood. Here we show that during acute gastrointestinal infection, priming of monocytes for regulatory function preceded systemic inflammation and was initiated prior to bone marrow egress. Notably, natural killer (NK) cell-derived IFN-γ promoted a regulatory program in monocyte progenitors during development. Early bone marrow NK cell activation was controlled by systemic interleukin-12 (IL-12) produced by Batf3 -dependent dendritic cells (DCs) in the mucosal-associated lymphoid tissue (MALT). This work challenges the paradigm that monocyte function is dominantly imposed by local signals after tissue recruitment, and instead proposes a sequential model of differentiation in which monocytes are pre-emptively educated during development in the bone marrow to promote their tissue-specific function. [ABSTRACT FROM AUTHOR]

Leucocytozoon parasites infect many species of avian hosts, including domestic chicken, and can inflict heavy economic loss to the poultry industry. Although the prevalence and distribution of two Leucocytozoon species (L. sabrazesi and L. caulleryi) have been reported in China previously, there are many questions related to the parasite infection that remain unanswered, including population diversity and transmission dynamics in domestic chickens. Here we surveyed chicken blood samples from seven sites in four provinces of China to identify Leucocytozoon infection, characterized parasite diversity within individual infected hosts and between sampling sites, and investigated the dynamics of gametocytemia in chickens over time. We found high infection rates in three of the seven sites. Clustering parasite sequences of the mitochondrial cytochrome oxidase III (coxIII) and cytochrome b (cytb) genes showed lack of grouping according to geographic origins and individual hosts carrying large numbers of L. sabrazesi strains. Monitoring gametocytemia in blood samples from infected chickens over time showed ‘relapse’ or persistence of low-level gametocytemia for 4–5 months, which could be explored as an in vivo model for testing drugs against liver stages of Apicomplexan parasites. This study provides important information on population diversity and transmission dynamics of L. sabrazesi and for disease control. [ABSTRACT FROM AUTHOR]

Highlights: [•] Genome sequencing of Plasmodium yoelii parasite strains. [•] Identification of large numbers of single nucleotide polymorphism (SNP). [•] Development of a high throughput and reliable genotyping microarray. [•] Detection of SNPs among P. yoelli strains and genetic cross progeny. [Copyright &y& Elsevier]

Malaria parasites are transmitted by mosquitoes, and blocking parasite transmission is critical in reducing or eliminating malaria in endemic regions. Here, we report the pharmacological characterization of a new class of malaria transmission-blocking compounds that acts via the inhibition of Plasmodia CDPK4 enzyme. We demonstrate that these compounds achieved selectivity over mammalian kinases by capitalizing on a small serine gatekeeper residue in the active site of the Plasmodium CDPK4 enzyme. To directly confirm the mechanism of action of these compounds, we generated P. falciparum parasites that express a drug-resistant methionine gatekeeper (S147M) CDPK4 mutant. Mutant parasites showed a shift in exflagellation EC50 relative to the wild-type strains in the presence of compound 1294, providing chemical-genetic evidence that CDPK4 is the target of the compound. Pharmacokinetic analyses suggest that coformulation of this transmission-blocking agent with asexual stage antimalarials such as artemisinin combination therapy (ACT) is a promising option for drug delivery that may reduce transmission of malaria including drug-resistant strains. Ongoing studies include refining the compounds to improve efficacy and toxicological properties for efficient blocking of malaria transmission. [ABSTRACT FROM AUTHOR]

Highlights: [•] Quantitative high-throughput screen (qHTS) of small molecules (SMs). [•] SMs as a tool for studying phenotypes and gene function. [•] Combination of SM screening and genome-wide approaches. [•] Target identification and pathway/network building. [ABSTRACT FROM AUTHOR]

Spatial relationships within the eukaryotic nucleus are essential for proper nuclear function. In Plasmodium falciparum, the repositioning of chromosomes has been implicated in the regulation of the expression of genes responsible for antigenic variation, and the formation of a single, peri-nuclear nucleolus results in the clustering of rDNA. Nevertheless, the precise spatial relationships between chromosomes remain poorly understood, because, until recently, techniques with sufficient resolution have been lacking. Here we have used chromosome conformation capture and second-generation sequencing to study changes in chromosome folding and spatial positioning that occur during switches in var gene expression. We have generated maps of chromosomal spatial affinities within the P. falciparum nucleus at 25 Kb resolution, revealing a structured nucleolus, an absence of chromosome territories, and confirming previously identified clustering of heterochromatin foci. We show that switches in var gene expression do not appear to involve interaction with a distant enhancer, but do result in local changes at the active locus. These maps reveal the folding properties of malaria chromosomes, validate known physical associations, and characterize the global landscape of spatial interactions. Collectively, our data provide critical information for a better understanding of gene expression regulation and antigenic variation in malaria parasites. [ABSTRACT FROM AUTHOR]

Cerebral malaria (CM) is a serious and often fatal complication of Plasmodium falciparum infections; however, the precise mechanisms leading to CM is poorly understood. Mouse malaria models have provided insight into the key events in pathogenesis of CM. T-cell immune response is known to play an important role in malaria infection, and members of the T-cell immunoglobulin- and mucin-domain-containing molecule (Tim) family have roles in T-cell-mediated immune responses. Tim-1 and Tim-3 are expressed on terminally differentiated Th2 and Th1 cells, respectively, and participate in the regulation of Th immune response. Until now, the role of Tim family proteins in Plasmodium infection remains unclear. In the present study, the mRNA levels of Tim-1, Tim-3, and some key Th1 and Th2 cytokines in the spleen of Kunming outbred mice infected with Plasmodium berghei ANKA ( PbANKA) were determined using real-time polymerase chain reaction (qRT-PCR). Compared with uninfected controls, Tim-1 expression was significantly decreased in infected mice with CM at day 10 postinfection (p.i.) but significantly increased in infected mice with non-CM at day 22 p.i.; in contrast, Tim-3 expression was significantly increased in infected mice both with CM at day 10 p.i. and with non-CM at day 22 p.i. The expressions of IFN-γ, TNF-α, IL-10, and IL-12 were significantly increased but IL-4 was significantly decreased in infected mice with CM at days 10 p.i., whereas the expressions of IFN-γ, TNF-α, IL-4, IL-10, and TGF-β were significantly increased but IL-12 was significantly decreased in infected mice with non-CM at days 22 p.i. Furthermore, the expression of Tim-1 and Tim-3 could reflect Th2 and Th1 immune response in the spleen of PbANKA-infected mice, respectively. Our data suggest that PbANKA infection could inhibit the differentiation of T lymphocytes toward Th2 cells, promote the Th1 cell differentiation, and induce Th1-biased immune response in the early infective stage, whereas the infection could promote Th2 cell differentiation and induce Th2-biased immune response in the late infective stage. Our data indicate that both Tim-1 and Tim-3 may play a role in the process of PbANKA infection, which may represent a potential therapeutic target. [ABSTRACT FROM AUTHOR]

The recent detection of clinical Artemisinin (ART) resistance manifested as delayed parasite clearance in the Cambodia-Thailand border area raises a serious concern. The mechanism of ART resistance is not clear; but the P. falciparum sarco/endoplasmic reticulum Ca2+-ATPase (PfSERCA or PfATP6) has been speculated to be the target of ARTs and thus a potential marker for ART resistance. Here we amplified and sequenced pfatp6 gene (∼3.6 Kb) in 213 samples collected after 2005 from the Greater Mekong Subregion, where ART drugs have been used extensively in the past. A total of 24 single nucleotide polymorphisms (SNPs), including 8 newly found in this study and 13 nonsynonymous, were identified. However, these mutations were either uncommon or also present in other geographical regions with limited ART use. None of the mutations were suggestive of directional selection by ARTs. We further analyzed pfatp6 from a worldwide collection of 862 P. falciparum isolates in 19 populations from Asia, Africa, South America and Oceania, which include samples from regions prior to and after deployments ART drugs. A total of 71 SNPs were identified, resulting in 106 nucleotide haplotypes. Similarly, many of the mutations were continent-specific and present at frequencies below 5%. The most predominant and perhaps the ancestral haplotype occurred in 441 samples and was present in 16 populations from Asia, Africa, and Oceania. The 3D7 haplotype found in 54 samples was the second most common haplotype and present in nine populations from all four continents. Assessment of the selection strength on pfatp6 in the 19 parasite populations found that pfatp6 in most of these populations was under purifying selection with an average dN/dS ratio of 0.333. Molecular evolution analyses did not detect significant departures from neutrality in pfatp6 for most populations, challenging the suitability of this gene as a marker for monitoring ART resistance. [ABSTRACT FROM AUTHOR]

Abstract: Genetic cross is a powerful tool for studying malaria genes contributing to drug resistance, parasite development, and pathogenesis. Cloning and identification of recombinant progeny (RP) is laborious and expensive, especially when a large proportion of progeny derived from self-fertilization are present in the uncloned progeny of a genetic cross. Since the frequency of cross-fertilization affects the number of recombinant progeny in a genetic cross, it is important to optimize the procedure of a genetic cross to maximize the cross-fertilization. Here we investigated the factors that might influence the chances of obtaining RP from a genetic cross and showed that different Plasmodium yoelii strains/subspecies/clones had unique abilities in producing oocysts in a mosquito midgut. When a genetic cross is performed between two parents producing different numbers of functional gametocytes, the ratio of parental parasites must be adjusted to improve the chance of obtaining RP. An optimized parental ratio could be established based on oocyst counts from single infection of each parent before crossing experiments, which may reflect the efficiency of gametocyte production and/or fertilization. The timing of progeny cloning is also important; cloning of genetic cross progeny from mice directly infected with sporozoites (vs. frozen blood after needle passage) at a time when parasitemia is low (usually <1%) could improve the chance of obtaining RP. This study provides an optimized protocol for efficiently cloning RPs from a genetic cross of malaria parasites. [Copyright &y& Elsevier]

BACKGROUND: Although transmission of Plasmodium falciparum (Pf) infection during red blood cell (RBC) transfusion from an infected donor has been well documented, malaria parasites are not known to infect hematopoietic stem cells. We report a case of Pf infection in a patient 11 days after peripheral blood stem cell transplant for sickle cell disease. STUDY DESIGN AND METHODS: Malaria parasites were detected in thick blood smears by Giemsa staining. Pf HRP2 antigen was measured by enzyme-linked immunosorbent assay on whole blood and plasma. Pf DNA was detected in whole blood and stem cell retention samples by real-time polymerase chain reaction using Pf species-specific primers and probes. Genotyping of eight Pf microsatellites was performed on genomic DNA extracted from whole blood. RESULTS: Pf was not detected by molecular, serologic, or parasitologic means in samples from the recipient until Day 11 posttransplant, coincident with the onset of symptoms. In contrast, Pf antigen was retrospectively detected in stored plasma collected 3 months before transplant from the asymptomatic donor. Pf DNA was detected in whole blood from both the donor and the recipient after transplant, and genotyping confirmed shared markers between donor and recipient Pf strains. Lookback analysis of RBC donors was negative for Pf infection. CONCLUSIONS: These findings are consistent with transmission by the stem cell product and have profound implications with respect to the screening of potential stem cell donors and recipients from malaria-endemic regions. [ABSTRACT FROM AUTHOR]

Summary: Background: Artemisinin-resistant Plasmodium falciparum has been reported in Pailin, western Cambodia, detected as a slow parasite clearance rate in vivo. Emergence of this phenotype in western Thailand and possibly elsewhere threatens to compromise the effectiveness of all artemisinin-based combination therapies. Parasite genetics is associated with parasite clearance rate but does not account for all variation. We investigated contributions of both parasite genetics and host factors to the artemisinin-resistance phenotype in Pursat, western Cambodia. Methods: Between June 19 and Nov 28, 2009, and June 26 and Dec 6, 2010, we enrolled patients aged 10 years or older with uncomplicated falciparum malaria, a density of asexual parasites of at least 10 000 per μL of whole blood, no symptoms or signs of severe malaria, no other cause of febrile illness, and no chronic illness. We gave participants 4 mg/kg artesunate at 0, 24, and 48 h, 15 mg/kg mefloquine at 72 h, and 10 mg/kg mefloquine at 96 h. We assessed parasite density on thick blood films every 6 h until undetectable. The parasite clearance half-life was calculated from the parasite clearance curve. We genotyped parasites with 18 microsatellite markers and patients for haemoglobin E, α-thalassaemia, and a mutation of G6PD, which encodes glucose-6-phosphate dehydrogenase. To account for the possible effects of acquired immunity on half-life, we used three surrogates for increased likelihood of exposure to P falciparum: age, sex, and place of residence. This study is registered with ClinicalTrials.gov, number NCT00341003. Findings: We assessed 3504 individuals from all six districts of Pursat province seeking treatment for malaria symptoms. We enrolled 168 patients with falciparum malaria who met inclusion criteria. The geometric mean half-life was 5·85 h (95% CI 5·54–6·18) in Pursat, similar to that reported in Pailin (p=0·109). We identified two genetically different parasite clone groups: parasite group 1 (PG1) and parasite group 2 (PG2). Non-significant increases in parasite clearance half-life were seen in patients with haemoglobin E (0·55 h; p=0·078), those of male sex (0·96 h; p=0·064), and in 2010 (0·68 h; p=0·068); PG1 was associated with a significant increase (0·79 h; p=0·033). The mean parasite heritability of half-life was 0·40 (SD 0·17). Interpretation: Heritable artemisinin resistance is established in a second Cambodian province. To accurately identify parasites that are intrinsically susceptible or resistant to artemisinins, future studies should explore the effect of erythrocyte polymorphisms and specific immune responses on half-life variation. Funding: Division of Intramural Research, National Institute of Allergy and Infectious Diseases, National Institutes of Health. [Copyright &y& Elsevier]

The recent reports of artemisinin (ART) resistance in the Thai-Cambodian border area raise a serious concern on the long-term efficacy of ARTs. To elucidate the resistance mechanisms, we performed in vitro selection with dihydroartemisinin (DHA) and obtained two parasite clones from Dd2 with more than 25-fold decrease in susceptibility to DHA. The DHA-resistant clones were more tolerant of stressful growth conditions and more resistant to several commonly used antimalarial drugs than Dd2. The result is worrisome as many of the drugs are currently used as ART partners in malaria control. This study showed that the DHA resistance is not limited to ring stage, but also occurred in trophozoites and schizonts. Microarray and biochemical analyses revealed pfmdr1 amplification, elevation of the antioxidant defence network, and increased expression of many chaperones in the DHA-resistant parasites. Without drug pressure, the DHA-resistant parasites reverted to sensitivity in approximately 8 weeks, accompanied by de-amplification of pfmdr1 and reduced antioxidant activities. The parallel decrease and increase in pfmdr1 copy number and antioxidant activity and the up and down of DHA sensitivity strongly suggest that pfmdr1 and antioxidant defence play a role in in vitro resistance to DHA, providing potential molecular markers for ART resistance. [ABSTRACT FROM AUTHOR]

Malaria elimination strategies require surveillance of the parasite population for genetic changes that demand a public health response, such as new forms of drug resistance. Here we describe methods for the large-scale analysis of genetic variation in Plasmodium falciparum by deep sequencing of parasite DNA obtained from the blood of patients with malaria, either directly or after short-term culture. Analysis of 86,158 exonic single nucleotide polymorphisms that passed genotyping quality control in 227 samples from Africa, Asia and Oceania provides genome-wide estimates of allele frequency distribution, population structure and linkage disequilibrium. By comparing the genetic diversity of individual infections with that of the local parasite population, we derive a metric of within-host diversity that is related to the level of inbreeding in the population. An open-access web application has been established for the exploration of regional differences in allele frequency and of highly differentiated loci in the P.?falciparum genome. [ABSTRACT FROM AUTHOR]

Macrophage migration inhibitory factor (MIF) has been shown to be involved in the pathogenesis of severe malaria. Malaria parasites express an MIF homolog that may play a role in regulating host immune responses, and a recent study showed that overexpression of MIF reduced parasitemia in a mouse malaria model. Another recent study showed migration of monocytes to the spleen contributed to the control of blood stage infection. However, there are few papers describing the effect of MIF on monocyte recruitment/activation during the infection. We generated recombinant Plasmodium yoelii MIF (rPyMIF) and investigated its function on purified mouse CD11b cells in vitro and monocyte responses in vivo. The result shows that rPyMIF protein bound to mouse CD11b cells and inhibited their random migration in vitro. On the other hand, rPyMIF did not induce cytokine release from the cells directly or modulate lipopolysaccharide-induced cytokine release. Mice immunized with rPyMIF showed transient but significantly lower parasitemia than the control mice at day 3 after lethal Py17XL challenge. The total number of CD11b cells in the spleens was significantly higher in rPyMIF-immunized group. Further investigation revealed that there were significantly higher numbers of recruited and activated monocytes in the spleens of rPyMIF immunization group on day 3. These results indicate that PyMIF potentially modulates monocyte recruitment and activation during infection of P. yoelii erythrocytic stages. [ABSTRACT FROM AUTHOR]

Of all the parasitic diseases, malaria is the number one killer. Despite tremendous efforts in disease control and research, nearly a million people, primarily children, still die from the disease each year, partly due to drug resistance and the lack of an effective vaccine. Many parasite antigens have been identified and evaluated for vaccine development; however, none has been approved for human use. Antigenic variation, complex life cycle, and inadequate understanding of the mechanisms of parasite-host interaction and of host immune response all contribute to the lack of an effective vaccine for malaria control. In a recent search of genome-wide polymorphism in Plasmodium falciparum, several molecules were found to be recognized by sera from patients infected with the P. falciparum parasite. Here, we have expressed a 350-amino acid N terminus from one of the homologous candidate antigen genes from the rodent malaria parasite Plasmodium yoelii (Py01157, a putative dentin phosphorin) in bacteria and evaluated the immune response and protection generated after immunization with the recombinant protein. We showed that the recombinant protein was recognized by sera from both mice and humans infected with malaria parasites. Partial protection was observed after challenge with non-lethal P. yoelii 17XNL but not with the lethal P. yoelii 17XL parasite. Further tests using a full-length protein or the conserved C terminus may provide additional information on whether this protein has the potential for being a malaria vaccine. [ABSTRACT FROM AUTHOR]

Abstract: Despite significant improvement in the malaria situation of the Greater Mekong Subregion (GMS), malaria control for the region continues to face a multitude of challenges. The extremely patchy malaria distribution, especially along international borders, makes disease surveillance and targeted control difficult. The vector systems are also diverse with dramatic differences in habitat ecology, biting behavior, and vectorial capacity, and there is a lack of effective transmission surveillance and control tools. Finally, in an era of heavy deployment of artemisinin-based combination therapies, the region acts as an epicenter of drug resistance, with the emergence of artemisinin resistant Plasmodium falciparum posing a threat to both regional and global malaria elimination campaigns. This problem is further exacerbated by the circulation of counterfeit and substandard artemisinin drugs. Accordingly, this Southeast Asian Malaria Research Center, consisting of a consortium of US and regional research institutions, has proposed four interlinked projects to address these most urgent problems in malaria control. The aims of these projects will help to substantially improve our understanding of malaria epidemiology, vector systems and their roles in malaria transmission, as well as the mechanisms of drug resistance in parasites. Through the training of next-generation scientists in malaria research, this program will help build up and strengthen regional research infrastructure and capacities, which are essential for sustained malaria control in this region. [Copyright &y& Elsevier]

Abstract: The Greater Mekong Subregion (GMS), comprised of six countries including Cambodia, China''s Yunnan Province, Lao PDR, Myanmar (Burma), Thailand and Vietnam, is one of the most threatening foci of malaria. Since the initiation of the WHO''s Mekong Malaria Program a decade ago, malaria situation in the GMS has greatly improved, reflected in the continuous decline in annual malaria incidence and deaths. However, as many nations are moving towards malaria elimination, the GMS nations still face great challenges. Malaria epidemiology in this region exhibits enormous geographical heterogeneity with Myanmar and Cambodia remaining high-burden countries. Within each country, malaria distribution is also patchy, exemplified by ‘border malaria’ and ‘forest malaria’ with high transmission occurring along international borders and in forests or forest fringes, respectively. ‘Border malaria’ is extremely difficult to monitor, and frequent malaria introductions by migratory human populations constitute a major threat to neighboring, malaria-eliminating countries. Therefore, coordination between neighboring countries is essential for malaria elimination from the entire region. In addition to these operational difficulties, malaria control in the GMS also encounters several technological challenges. Contemporary malaria control measures rely heavily on effective chemotherapy and insecticide control of vector mosquitoes. However, the spread of multidrug resistance and potential emergence of artemisinin resistance in Plasmodium falciparum make resistance management a high priority in the GMS. This situation is further worsened by the circulation of counterfeit and substandard artemisinin-related drugs. In most endemic areas of the GMS, P. falciparum and Plasmodium vivax coexist, and in recent malaria control history, P. vivax has demonstrated remarkable resilience to control measures. Deployment of the only registered drug (primaquine) for the radical cure of vivax malaria is severely undermined due to high prevalence of glucose-6-phosphate dehydrogenase deficiency in target human populations. In the GMS, the dramatically different ecologies, diverse vector systems, and insecticide resistance render traditional mosquito control less efficient. Here we attempt to review the changing malaria epidemiology in the GMS, analyze the vector systems and patterns of malaria transmission, and identify the major challenges the malaria control community faces on its way to malaria elimination. [Copyright &y& Elsevier]