Progressive heterogeneity of enlarged and irregularly shaped apicoplasts in persister blood stages after drug treatment.

Morphological modifications and shifts in organelle relationships are hallmarks of dormancy in eukaryotic cells. Communications between altered mitochondria and nuclei are associated with metabolic quiescence of cancer cells that can survive chemotherapy. In plants, changes in the pathways between nuclei, mitochondria, and chloroplasts are associated with cold stress and bud dormancy.

Infectivity of Plasmodium parasites to Aedes aegypti and Anopheles stephensi mosquitoes maintained on blood-free meals of SkitoSnack.

Background: Aedes and Anopheles mosquitoes are responsible for tremendous global health burdens from their transmission of pathogens causing malaria, lymphatic filariasis, dengue, and yellow fever. Innovative vector control strategies will help to reduce the prevalence of these diseases. Mass rearing of mosquitoes for research and support of these strategies presently depends on meals of vertebrate blood, which is subject to acquisition, handling, and storage issues.

Progressive heterogeneity of enlarged and irregularly shaped apicoplasts in persister blood stages after drug treatment.

Morphological modifications and shifts in organelle relationships are hallmarks of dormancy in eukaryotic cells. Communications between altered mitochondria and nuclei are associated with metabolic quiescence of cancer cells that can survive chemotherapy. In plants, changes in the pathways between nuclei, mitochondria, and chloroplasts are associated with cold stress and bud dormancy.

Structure of complex III with bound antimalarial agent CK-2-68 provides insights into selective inhibition of Plasmodium cytochrome bc complexes.

Among the various components of the protozoan Plasmodium mitochondrial respiratory chain, only Complex III is a validated cellular target for antimalarial drugs. The compound CK-2-68 was developed to specifically target the alternate NADH dehydrogenase of the malaria parasite respiratory chain, but the true target for its antimalarial activity has been controversial. Here, we report the cryo-EM structure of mammalian mitochondrial Complex III bound with CK-2-68 and examine the structure-function relationships of the inhibitor's selective action on Plasmodium.

Malaria Control by Mass Drug Administration With Artemisinin Plus Piperaquine on Grande Comore Island, Union of Comoros.

Background: Mass drug administration (MDA) is a powerful tool for malaria control, but the medicines to use, dosing, number of rounds, and potential selection of drug resistance remain open questions.

Methods: Two monthly rounds of artemisinin-piperaquine (AP), each comprising 2 daily doses, were administered across the 7 districts of Grande Comore Island. In 3 districts, low-dose primaquine (PMQ) was also given on the first day of each monthly round.

Pf7: an open dataset of genome variation in 20,000 worldwide samples.

We describe the MalariaGEN Pf7 data resource, the seventh release of genome variation data from the MalariaGEN network.  It comprises over 20,000 samples from 82 partner studies in 33 countries, including several malaria endemic regions that were previously underrepresented.  For the first time we include dried blood spot samples that were sequenced after selective whole genome amplification, necessitating new methods to genotype copy number variations.

A Plasmodium falciparum RING Finger E3 Ubiquitin Ligase Modifies the Roles of PfMDR1 and PfCRT in Parasite Drug Responses.

Protein ubiquitination is an important posttranslational regulation mechanism that mediates development and modifies parasite responses to antimalarial drugs. Although mutations in several parasite ubiquitination enzymes have been linked to increased drug tolerance, the molecular mechanisms by which ubiquitination pathways mediate these parasite responses remain largely unknown. Here, we investigate the roles of a Plasmodium falciparum ring finger ubiquitin ligase (PfRFUL) in parasite development and in responses to antimalarial drugs.

Long read single cell RNA sequencing reveals the isoform diversity of Plasmodium vivax transcripts.

Plasmodium vivax infections often consist of heterogenous populations of parasites at different developmental stages and with distinct transcriptional profiles, which complicates gene expression analyses. The advent of single cell RNA sequencing (scRNA-seq) enabled disentangling this complexity and has provided robust and stage-specific characterization of Plasmodium gene expression. However, scRNA-seq information is typically derived from the end of each mRNA molecule (usually the 3'-end) and therefore fails to capture the diversity in transcript isoforms documented in bulk RNA-seq data.

An open dataset of genome variation in 1,895 worldwide samples.

This report describes the MalariaGEN Pv4 dataset, a new release of curated genome variation data on 1,895 samples of collected at 88 worldwide locations between 2001 and 2017. It includes 1,370 new samples contributed by MalariaGEN and VivaxGEN partner studies in addition to previously published samples from these and other sources. We provide genotype calls at over 4.

Activity of Plasmodium vivax promoter elements in Plasmodium knowlesi, and a centromere-containing plasmid that expresses NanoLuc throughout the parasite life cycle.

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.

Restructured Mitochondrial-Nuclear Interaction in Plasmodium falciparum Dormancy and Persister Survival after Artemisinin Exposure.

Artemisinin and its semisynthetic derivatives (ART) are fast acting, potent antimalarials; however, their use in malaria treatment is frequently confounded by recrudescences from bloodstream parasites that enter into and later reactivate from a dormant persister state. Here, we provide evidence that the mitochondria of dihydroartemisinin (DHA)-exposed persisters are dramatically altered and enlarged relative to the mitochondria of young, actively replicating ring forms. Restructured mitochondrial-nuclear associations and an altered metabolic state are consistent with stress from reactive oxygen species.

Structure-switching aptamer sensors for the specific detection of piperaquine and mefloquine.

Tracking antimalarial drug use and efficacy is essential for monitoring the current spread of antimalarial drug resistance. However, available methods for determining tablet quality and patient drug use are often inaccessible, requiring well-equipped laboratories capable of performing liquid chromatography-mass spectrometry (LC-MS). Here, we report the development of aptamer-based fluorescent sensors for the rapid, specific detection of the antimalarial compounds piperaquine and mefloquine-two slow-clearing partner drugs in current first-line artemisinin-based combination therapies (ACTs).

Transcriptional heterogeneity and tightly regulated changes in gene expression during sporozoite development.

Despite the critical role of sporozoites in malaria transmission, we still know little about the mechanisms underlying their development in mosquitoes. Here, we use single-cell RNA sequencing to characterize the gene expression profiles of 16,038 sporozoites isolated throughout their development from midgut oocysts to salivary glands, and from forced salivation experiments. Our results reveal a succession of tightly regulated changes in gene expression occurring during the maturation of sporozoites and highlight candidate genes that could play important roles in oocyst egress, sporozoite motility, and the mechanisms underlying the invasion of mosquito salivary glands and mammalian hepatocytes.

Sensitive Immunoassay Detection of Lactate Dehydrogenase by Inductively Coupled Plasma Mass Spectrometry.

Rapid, reliable, and sensitive detection of infection is central to malaria control and elimination. Many Malaria Rapid Diagnostic Tests (RDTs) developed for this purpose depend upon immunoassays that can be improved by advances in bound antibody sensor technology. In a previous study, immuno-polymerase chain reaction (PCR) was shown to provide highly sensitive detection of lactate dehydrogenase (PfLDH) in monoclonal antibody (mAb) sandwich assays.

Modulation of Triple Artemisinin-Based Combination Therapy Pharmacodynamics by Genotype.

The first-line treatments for uncomplicated malaria are artemisinin-based combination therapies (ACTs), consisting of an artemisinin derivative combined with a longer acting partner drug. However, the spread of with decreased susceptibility to artemisinin and partner drugs presents a significant challenge to malaria control efforts. To stem the spread of drug resistant parasites, novel chemotherapeutic strategies are being evaluated, including the implementation of triple artemisinin-based combination therapies (TACTs).

Regioisomerization of Antimalarial Drug WR99210 Explains the Inactivity of a Commercial Stock.

WR99210, a former antimalarial drug candidate now widely used for the selection of transfectants, selectively targets the parasite's dihydrofolate reductase thymidine synthase bifunctional enzyme (DHFR-TS) but not human DHFR, which is not fused with TS. Accordingly, WR99210 and plasmids expressing the human gene have become valued tools for the genetic modification of parasites in the laboratory. Concerns over the ineffectiveness of WR99210 from some sources encouraged us to investigate the biological and chemical differences of supplies from two different companies (compounds 1 and 2).

Detection of simple and complex de novo mutations with multiple reference sequences.

The characterization of de novo mutations in regions of high sequence and structural diversity from whole-genome sequencing data remains highly challenging. Complex structural variants tend to arise in regions of high repetitiveness and low complexity, challenging both de novo assembly, in which short reads do not capture the long-range context required for resolution, and mapping approaches, in which improper alignment of reads to a reference genome that is highly diverged from that of the sample can lead to false or partial calls. Long-read technologies can potentially solve such problems but are currently unfeasible to use at scale.

'Artemisinin Resistance': Something New or Old? Something of a Misnomer?

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.

Development of copy number assays for detection and surveillance of piperaquine resistance associated plasmepsin 2/3 copy number variation in Plasmodium falciparum.

Background: Long regarded as an epicenter of drug-resistant malaria, Southeast Asia continues to provide new challenges to the control of Plasmodium falciparum malaria. Recently, resistance to the artemisinin combination therapy partner drug piperaquine has been observed in multiple locations across Southeast Asia. Genetic studies have identified single nucleotide polymorphisms as well as copy number variations in the plasmepsin 2 and plasmepsin 3 genes, which encode haemoglobin-degrading proteases that associate with clinical and in vitro piperaquine resistance.