Malaria in the Australian military, 2008-2022.

Australian Defence Force (ADF) personnel train and operate in malarious regions that include neighboring countries with high burden and species with latent hepatic parasites.1 We summarized longitudinal malaria case data, following a prior 10-year period review to 2007.2 Malaria case entries within the ADF Malaria and Infectious Diseases Institute (ADFMIDI)-managed Central Malaria Register (CMR) were examined. Data from cases confirmed between January 1, 2008 through December 31, 2022 were analyzed. Sixty ADF members were diagnosed with malaria, including 1 with a mixed Plasmodium falciparum and P. vivax infection. Of 61 malaria infections, 69% (42 of 61) were P. vivax. P. vivax infection resulted in delayed initial case presentation (more than 4 weeks after exposure) in at least 36% (15 of 42) of cases, and 5 personnel experienced further relapse. Most P. vivax infections were acquired in the U.S. Indo-Pacific Command (INDOPACOM) and P. falciparum in the U.S. Africa Command (AFRICOM) regions. The ADF experienced ongoing reduced malaria case incidence following high rates in the early 2000s. Maintenance of prophylactic vigilance, both for eradicating dormant hypnozoites and preventing P. vivax relapse, remains important, however.

Discovery of 1,3,4-oxadiazoles with slow-action activity against Plasmodium falciparum malaria parasites.

To achieve malaria eradication, new preventative agents that act differently to front-line treatment drugs are needed. To identify potential chemoprevention starting points we screened a sub-set of the CSIRO Australia Compound Collection for compounds with slow-action in vitro activity against Plasmodium falciparum. This work identified N,N-dialkyl-5-alkylsulfonyl-1,3,4-oxadiazol-2-amines as a new antiplasmodial chemotype (e.g., 1 96 h IC50 550 nM; 3 96 h IC50 160 nM) with a different action to delayed-death slow-action drugs. A series of analogues were synthesized from thiotetrazoles and carbomoyl derivatives using Huisgen 1,3,4-oxadiazole synthesis followed by oxidation of the resultant thioethers to target sulfones. Structure activity relationship analysis of analogues identified compounds with potent and selective in vitro activity against drug-sensitive and multi-drug resistant Plasmodium parasites (e.g., 31 and 32 96 h IC50 <40 nM; SI > 2500). Subsequent studies in mice with compound 1, which had the best microsomal stability of the compounds assessed (T1/2 >255 min), demonstrated rapid clearance and poor oral in vivo efficacy in a P. berghei murine malaria model. These data indicate that while N,N-dialkyl-5-alkylsulfonyl-1,3,4-oxadiazol-2-amines are a novel class of slow-acting antiplasmodial agents, the further development of this chemotype for malaria chemoprophylaxis will require pharmacokinetic profile improvements.

Feeding Behavior and Plasmodium Detection in Anopheles stephensi, a Malaria Vector in District Khyber, Khyber Pakhtunkhwa, Pakistan.

Background: Anopheles stephensi is a significant malaria vector in Pakistan, and understanding its feeding behavior is necessary to control the spread of malaria. However, limited information is available on the host preferences of A. stephensi in Pakistan. Therefore, we aimed to explore the feeding behavior of A. stephensi, a malaria vector, in the District Khyber, Khyber Pakhtunkhwa, Pakistan. Methods: A total of 7462 mosquitoes were collected between March and September 2021, with 1674 (22.4%) identified as A. stephensi (952 female and 722 male). Among the female A. stephensi, 495 (52%) were blood-fed. DNA was extracted from the blood-fed female A. stephensi mosquitoes using the Ammonium Acetate Precipitation Method followed by PCR analysis, blood meal sources were identified. Nested PCR on 191 pooled samples was used to detect Plasmodium falciparum and Plasmodium vivax. Results: Cattle blood meals were predominant (73%), followed by human (20%) and chicken (7%), with no dog blood meals detected. All individual mosquito samples were negative for Plasmodium falciparum, while two pooled samples (out of 191) tested positive for P. vivax. Conclusion: A. stephensi in Khyber District primarily displayed anthropophagic feeding behavior, with a small portion of the population infected with P. vivax. The results underscore the importance of targeted vector control strategies, environmental management, community engagement and continuous monitoring to suppress malaria transmission.

Analysis of Diagnostic Biomarkers for Malaria: Prospects on Rapid diagnostic test development.

Accurate malaria diagnosis remains a formidable challenge in remote regions of malaria-endemic areas globally. Existing diagnostic methods predominantly rely on microscopy and rapid diagnostic tests (RDTs). While RDTs offer advantages such as rapid results and reduced dependence on highly skilled technicians compared to microscopy, persistent challenges emphasize the critical need to identify novel diagnostic biomarkers to further enhance RDT based malaria diagnosis. This comprehensive review presents a range of promising diagnostic targets. These targets could be useful in developing more robust, accurate, and effective diagnostic tools. Such tools are crucial for the detection of the Plasmodium falciparum malaria parasite. The potential biomarkers discussed here significantly address the challenges posed by HRP2 gene deletion in P.falciparum. Researchers, RDT manufacturers, industrial and other stakeholders involved in malaria diagnosis can harness the crucial information describe in this article, to drive the development of advanced RDTs as viable alternatives. The potential biomarkers discussed here significantly in address the challenges posed by HRP2 gene deletion in P.falciparum. By diversifying the available tools for diagnosis, we can attempt to enhance our ability to knock out malaria effectively and contribute to better health outcomes for peoples residing in malaria-endemic regions. This review serves as a valuable resource for advancing research and development in the field of malaria diagnostics, ultimately aiding to the global fight against this devastating ancient disease.

In vitro antimalarial susceptibility profile of Plasmodium falciparum isolates in the BEI Resources repository.

BEI Resources, a National Institute of Allergy and Infectious Diseases-funded program managed by the American Type Culture Collection, serves researchers worldwide through the provision of a centralized repository for the acquisition, production, characterization, preservation, storage, and distribution of standardized biological resources targeting National Institutes of Health priority pathogens including bacteria, viruses, pathogenic fungi, and parasitic protozoa. These reference materials are critical for the development of diagnostics, vaccines, and therapeutics and are available to qualified registered investigators and institutions worldwide. Bioresources within BEI include well-characterized malaria isolates as part of the Malaria Research and Reference Reagent Resource Center (MR4). These isolates are critical for screening antimalarial compounds, conducting drug resistance studies, and for resistance surveillance and management. In our efforts to enhance the characterization of MR4 P. falciparum isolates, we measured antimalarial susceptibility of >100 isolates against a panel of standard antimalarial compounds. Our results provide valuable information to assist current and prospective users of the BEI Resources repository in making data-driven requests of isolates to meet their research needs.

Simple supplementation of serum-free medium produces gametocytes of Plasmodium falciparum that transmit to mosquitoes.

BACKGROUND: Human serum is a major component of Plasmodium falciparum culture medium, and can be replaced with AlbuMAX™ II, a lipid-rich bovine serum albumin, for asexual cultures. However, gametocytes produced without serum are poorly infective to mosquitoes. Serum suffers from high cost, limited availability, and variability in quality. METHODS: Several commercially-available media supplements were tested for their ability to support parasite growth and production of P. falciparum (3D7) gametocytes in standard RPMI1640 medium containing 0.5% AlbuMAX. The impact on asexual growth and gametocyte production with each supplement was assessed and compared to standard RPMI1640 medium containing 10% human serum, as well as to medium containing 0.5% AlbuMAX alone. The infectivity of gametocytes produced with one supplement to Anopheles gambiae sensu stricto was assessed by standard membrane feeding assay and measuring both prevalence of infection and oocyst intensity. RESULTS: Supplementation of medium containing 0.5% AlbuMAX with five supplements did not affect asexual growth of P. falciparum, and four of the five supplements supported early gametocyte production. The supplement producing the highest number of gametocytes, ITS-X, was further investigated and was found to support the production of mature gametocytes. Infection prevalence and oocyst intensity did not differ significantly between mosquitoes given a membrane feed containing gametocytes grown in medium with 0.5% AlbuMAX + ITS-X and those grown in medium with 10% human serum. Infection prevalence and oocyst intensity was significantly higher in case of ITS-X supplementation when compared to AlbuMAX alone. Infectious gametocytes were also produced from two field clones using ITS-X supplementation. CONCLUSIONS: Serum-free medium supplemented with ITS-X was able to support the growth of gametocytes of P. falciparum that were as infectious to An. gambiae as those grown in medium with 10% serum. This is the first fully serum-free culture system able to produce highly infectious gametocytes, thereby removing the requirement for access to serum for transmission assays.

Circulation of Non-falciparum Species in Niger: Implications for Malaria Diagnosis.

Background: Niger's National Malaria Control Programme and its partners use histidine-rich protein 2-based RDTs, which are specific to Plasmodium falciparum diagnosis. This study aimed to screen for the circulation of non-falciparum species in Zinder, a region of Niger, West Africa. Methods: A cross-sectional study was carried out from July to December 2022 at the district hospital of the Zinder region of Niger. P falciparum histidine-rich protein 2-based rapid diagnostic tests were performed, and dried blood spot samples were collected for further laboratory multiplexed photo-induced electron transfer-polymerase chain reaction (PET-PCR) analysis on positive light microscopy from all patients with fever who attended the Zinder district hospital during the study period. Results: In total, 340 dried blood spots were collected and analyzed by PET-PCR. Overall, 73.2% (95% CI, 68.2%-77.9%; 249/340) were positive for Plasmodium genus and species and represented the study population. Plasmodium species proportions were 89.5% (95% CI, 85.1%-93.1%; 223/249) for P falciparum, 38.5% (95% CI, 32.5%-44.9%; 96/249) for P malariae, 10.8% (95% CI, 7.3%-15.4%; 27/249) for P vivax, and 1.6% (95% CI, .4%-4.1%; 4/249) for P ovale. Single infection with Plasmodium species counted for 61.8% (95% CI, 55.5%-67.9%; 154/249), and the mixed infections rate, with at least 2 Plasmodium species, was 38.1% (95% CI, 32.1%-44.5%; 95/249). Single non-falciparum infections represented a rate of 10.0% (95% CI, 6.6%-14.5%; 25/249). Conclusion: This study confirms the first evidence of Plasmodium vivax by PET-PCR in Niger in addition to the other 3 Plasmodium species. These findings underline the need to adapt malaria diagnostic tools and therapeutic management, as well as the training of microscopists, for recognition of non-falciparum plasmodial species circulating in the country. This will better inform the strategies toward malaria control and elimination, as well as the decision making of the health authorities of Niger.

Mass spectrometry-complemented molecular modeling predicts the interaction interface for a camelid single-domain antibody targeting the Plasmodium falciparum circumsporozoite protein's C-terminal domain.

Background: Bioanalytical methods that enable rapid and high-detail characterization of binding specificities and strengths of protein complexes with low sample consumption are highly desired. The interaction between a camelid single domain antibody (sdAbCSP1) and its target antigen (PfCSP-Cext) was selected as a model system to provide proof-of-principle for the here described methodology. Research design and methods: The structure of the sdAbCSP1 - PfCSP-Cext complex was modeled using AlphaFold2. The recombinantly expressed proteins, sdAbCSP1, PfCSP-Cext, and the sdAbCSP1 - PfCSP-Cext complex, were subjected to limited proteolysis and mass spectrometric peptide analysis. ITEM MS (Intact Transition Epitope Mapping Mass Spectrometry) and ITC (Isothermal Titration Calorimetry) were applied to determine stoichiometry and binding strength. Results: The paratope of sdAbCSP1 mainly consists of its CDR3 (aa100-118). PfCSP-Cext's epitope is assembled from its α-helix (aa40-52) and opposing loop (aa83-90). PfCSP-Cext's GluC cleavage sites E46 and E58 were shielded by complex formation, confirming the predicted epitope. Likewise, sdAbCSP1's tryptic cleavage sites R105 and R108 were shielded by complex formation, confirming the predicted paratope. ITEM MS determined the 1:1 stoichiometry and the high complex binding strength, exemplified by the gas phase dissociation reaction enthalpy of 50.2 kJ/mol. The in-solution complex dissociation constant is 5 × 10-10 M. Conclusions: Combining AlphaFold2 modeling with mass spectrometry/limited proteolysis generated a trustworthy model for the sdAbCSP1 - PfCSP-Cext complex interaction interface.

3-Benzylmenadiones and their Heteroaromatic Analogues Target the Apicoplast of Apicomplexa Parasites: Synthesis and Bioimaging Studies.

The apicoplast is an essential organelle for the viability of apicomplexan parasites Plasmodium falciparum or Toxoplasma gondii, which has been proposed as a suitable drug target for the development of new antiplasmodial drug-candidates. Plasmodione, an antimalarial redox-active lead drug is active at low nM concentrations on several blood stages of Plasmodiumsuch as early rings and gametocytes. Nevertheless, its precise biological targets remain unknown. Here, we described the synthesis and the evaluation of new heteroaromatic analogues of plasmodione, active on asexual blood P. falciparum stages and T. gondii tachyzoites. Using a bioimaging-based analysis, we followed the morphological alterations of T. gondii tachyzoites and revealed a specific loss of the apicoplast upon drug treatment. Lipidomic and fluxomic analyses determined that drug treatment severely impacts apicoplast-hosted FASII activity in T. gondii tachyzoites, further supporting that the apicoplast is a primary target of plasmodione analogues. To follow the drug localization, "clickable" analogues of plasmodione were designed as tools for fluorescence imaging through a Cu(I)-catalyzed azide-alkyne cycloaddition reaction. Short-time incubation of two probes with P. falciparum trophozoites and T. gondii tachyzoites showed that the clicked products localize within, or in the vicinity of, the apicoplast of both Apicomplexa parasites. In P. falciparum, the fluorescence signal was also associated with the mitochondrion, suggesting that bioactivation and activity of plasmodione and related analogues are potentially associated with these two organelles in malaria parasites.

The three Plasmodium falciparum Aurora-related kinases display distinct temporal and spatial associations with mitotic structures in asexual blood stage parasites and gametocytes.

Aurora kinases are crucial regulators of mitotic cell cycle progression in eukaryotes. The protozoan malaria parasite Plasmodium falciparum replicates via schizogony, a specialized mode of cell division characterized by consecutive asynchronous rounds of nuclear division by closed mitosis followed by a single cytokinesis event producing dozens of daughter cells. P. falciparum encodes three Aurora-related kinases (PfARKs) that have been reported essential for parasite proliferation, but their roles in regulating schizogony have not yet been explored in great detail. Here, we engineered transgenic parasite lines expressing GFP-tagged PfARK1-3 to provide a systematic analysis of their expression timing and subcellular localization throughout schizogony as well as in the non-dividing gametocyte stages, which are essential for malaria transmission. We demonstrate that all three PfARKs display distinct and highly specific and exclusive spatiotemporal associations with the mitotic machinery. In gametocytes, PfARK3 is undetectable, and PfARK1 and PfARK2 show male-specific expression in late-stage gametocytes, consistent with their requirement for endomitosis during male gametogenesis in the mosquito vector. Our combined data suggest that PfARK1 and PfARK2 have non-overlapping roles in centriolar plaque maturation, assembly of the mitotic spindle, kinetochore-spindle attachment and chromosome segregation, while PfARK3 seems to be exquisitely involved in daughter cell cytoskeleton assembly and cytokinesis. These important new insights provide a reliable foundation for future research aiming at the functional investigation of these divergent and possibly drug-targetable Aurora-related kinases in mitotic cell division of P. falciparum and related apicomplexan parasites.IMPORTANCEMalaria parasites replicate via non-conventional modes of mitotic cell division, such as schizogony, employed by the disease-causing stages in the human blood or endomitosis during male gametogenesis in the mosquito vector. Understanding the molecular mechanisms regulating cell division in these divergent unicellular eukaryotes is not only of scientific interest but also relevant to identify potential new antimalarial drug targets. Here, we carefully examined the subcellular localization of all three Plasmodium falciparum Aurora-related kinases (ARKs), distantly related homologs of Aurora kinases that coordinate mitosis in model eukaryotes. Detailed fluorescence microscopy-based analyses revealed distinct, specific, and exclusive spatial associations for each parasite ARK with different components of the mitotic machinery and at different phases of the cell cycle during schizogony and gametocytogenesis. This comprehensive set of results closes important gaps in our fragmentary knowledge on this important group of kinases and offers a valuable source of information for future functional studies.

Suppression by RNA Polymerase I Inhibitors Varies Greatly Between Distinct RNA Polymerase I Transcribed Genes in Malaria Parasites.

Transcription of ribosomal RNA (rRNA) by RNA Polymerase I (Pol I) is the rate-limiting step in ribosome biogenesis and a major determinant of cellular growth rates. Unlike virtually every other eukaryote, which express identical rRNA from large tandem arrays of dozens to hundreds of identical rRNA genes in every cell, the genome of the human malaria parasite Plasmodium falciparum contains only a handful single-copy 47S rRNA loci that differ substantially from one another in length, sequence and expression in different cell-types. We found that growth of malaria parasite was acutely sensitive to the Pol I inhibitors 9-hydroxyellipticine and BMH-21 and demonstrate that they greatly reduce the transcription of 47S rRNAs as well as transcription of other non-coding RNA genes. Surprisingly, we found that the various types of Pol I-transcribed genes differed by more than two orders of magnitude in their susceptibility to these inhibitors and explore the implications of these findings for regulation of rRNA in P. falciparum.

A comprehensive review of synthetic strategies and SAR studies for the discovery of PfDHODH inhibitors as antimalarial agents. Part 2: Non-DSM compounds.

Malaria remains a severe global health concern, with 249 million cases reported in 2022, according to the World Health Organization (WHO) [1]. PfDHODH is an essential enzyme in malaria parasites that helps to synthesize certain building blocks for their growth and development. It has been confirmed that targeting Plasmodium falciparum dihydroorotate dehydrogenase (PfDHODH) enzyme could lead to new and effective antimalarial drugs. Inhibitors of PfDHODH have shown potential for slowing down parasite growth during both the blood and liver stages. Over the last two decades, many species selective PfDHODH inhibitors have been designed, including DSM compounds and other non-DSM compounds. In the first chapter [2] of this review, we have reviewed all synthetic schemes and structure-activity relationship (SAR) studies of DSM compounds. In this second chapter, we have compiled all the other non-DSM PfDHODH inhibitors based on dihydrothiophenones, thiazoles, hydroxyazoles, and N-alkyl-thiophene-2-carboxamides. The review not only offers an insightful overview of the synthetic methods employed but also explores into alternative routes and innovative strategies involving different catalysts and chemical reagents. A critical aspect covered in the review is the SAR studies, which provide a comprehensive understanding of how structural modifications impact the efficacy of PfDHODH inhibitors and challenges related to the discovery of PfDHODH inhibitors. This information is invaluable for scientists engaged in the development of new antimalarial drugs, offering insights into the most promising scaffolds and their synthetic techniques.

An adaptable, fit-for-purpose screening approach with high-throughput capability to determine speed of action and stage specificity of anti-malarial compounds.

A revamped in vitro compound identification and activity profiling approach is required to meet the large unmet need for new anti-malarial drugs to combat parasite drug resistance. Although compound hit identification utilizing high-throughput screening of large compound libraries is well established, the ability to rapidly prioritize such large numbers for further development is limited. Determining the speed of action of anti-malarial drug candidates is a vital component of malaria drug discovery, which currently occurs predominantly in lead optimization and development. This is due in part to the capacity of current methods which have low throughput due to the complexity and labor intensity of the approaches. Here, we provide an adaptable screening paradigm utilizing automated high content imaging, including the development of an automated schizont maturation assay, which collectively can identify anti-malarial compounds, classify activity into fast and slow acting, and provide an indication of the parasite stage specificity, with high-throughput capability. By frontloading these critical biological parameters much earlier in the drug discovery pipeline, it has the potential to reduce lead compound attrition rates later in the development process. The capability of the approach in its alternative formats is demonstrated using three Medicines for Malaria Venture open access compound "boxes," namely Pathogen Box (malaria set-125 compounds), Global Health Priority Box [Malaria Box 2 (80 compounds) and zoonotic neglected diseases (80 compounds)], and the Pandemic Response Box (400 compounds). From a total of 685 compounds tested, 79 were identified as having fast ring-stage-specific activity comparable to that of artemisinin and therefore of high priority for further consideration and development.

Performance of rapid diagnostic test, light microscopy, and polymerase chain reaction in pregnant women with asymptomatic malaria in Nigeria.

Objectives: Rapid diagnostic tests (RDTs) offer an attractive tool for diagnosing malaria in pregnancy. This study assessed the effectiveness of a Plasmodium falciparum-specific RDT compared with microscopy and polymerase chain reaction (PCR) in diagnosing asymptomatic malaria in pregnant women in southwest Nigeria. Methods: The study included 406 asymptomatic pregnant women seeking antenatal care. Blood samples were collected and tested using RDT (SD Bioline, Standard Diagnostics Inc. Korea) and light microscopy and confirmed using nested PCR. Results: The study revealed that the malaria parasite positivity rate was 8.9% by RDT, 21% by microscopy, and 32% by nested PCR. RDT had a sensitivity of 51.4% and specificity of 69.5%, whereas microscopy had a sensitivity of 65.3% and specificity of 98.2%. The combined testing of microscopy and RDT had a sensitivity and specificity of 100%. The study also showed a high prevalence of mild anemia among participants. Conclusions: Despite the RDT's low sensitivity, its high negative predictive value suggests it could be useful in combination with microscopy in ruling out asymptomatic malaria in pregnancy. Further study will help identify more suitable RDTs for routine malaria diagnosis in Nigeria and strengthen malaria prevention programs in pregnant women.

Giant membrane rings/loops in the cytosol of P. falciparum-infected erythrocytes and their relation to the parasite.

Striking morphological transformations characterize the invasion of a red blood cell by the malaria parasite. Shortly after the infection, parasite-induced membranes appear in the cytosol of the affected host erythrocyte. One intensely investigated membrane type, commonly called Maurer's clefts, has a slit-like morphology and can be arranged in the form of extended three-dimensional membrane stacks or networks. Here we report the three-dimensional reconstruction of a second membrane type, giant or extended membrane rings/loops, that have only occasionally been described on single ultrathin sections, however that have never been systematically examined so far. Serial ultrathin sectioning of P. falciparum-infected red blood cells, subsequent three-dimensional reconstructions, and in addition examination of Giemsa-stained blood films revealed that intraerythrocytic membrane rings/loops are not isolated structures but are locally in contact with the parasite. They consist either of the parasitophorous vacuolar membrane alone or contain the parasitophorous vacuolar membrane including the plasma membrane of the parasite and small amounts of parasite cytoplasm. We demonstrate that membrane rings/loops represent surface extensions of the parasite that maybe involved in ring stage parasite formation and Maurer's cleft generation at least in a subset of infected red blood cells.

Oleuropein mediated autophagy begets antimalarial drug resistance.

Drug resistance in Plasmodium falciparum presents a formidable challenge to the humanity. And, unavailability of an effective vaccine worsens the situation further. Autophagy is one of the mechanisms employed by parasite to evade drug pressure to survive. Autophagy induced by the P. falciparum in response to the oleuropein pressure may answer many questions related to the parasite survival as well as evolving drug tolerance. The survival/autophagy axis could be an important avenue to explore in order to address certain questions related to the evolution of drug resistance. In addition, humanized mouse model of P. falciparum infection could serve as an important preclinical tool to investigate the oleuropein-induced autophagy, potentially helping to dissect the mechanisms underlying the development of antimalarial drug resistance.

ISCCM Position Statement on the Management of Severe Malaria in Intensive Care Unit.

Malaria is a worldwide health concern, but a great majority of cases occur in tropical countries like India. With almost 95% of Indian population living in malaria endemic regions, India contributes to most of the global malaria cases and deaths, outside of African countries. Despite significant advances towards malaria control and eradication, mortality associated with severe malaria remains particularly high. Changing epidemiology, vulnerable patient population, overlapping symptomatology, and limited availability of parenteral preparations of artemisinin derivatives pose significant challenges in management of severe malaria. Further, the dearth of large-scale randomized trials from the developing countries makes it difficult to establish evidence-based guidelines pertaining to their situation. Thus, this position paper aims to provide guidance to critical care physicians across the country on managing patients with severe malaria in intensive care units (ICUs). How to cite this article: Hegde A, Chhallani AK, Gupta B, Kadapatti K, Karnad D, Maheshwarappa HM, et al. ISCCM Position Statement on the Management of Severe Malaria in Intensive Care Unit. Indian J Crit Care Med 2024;28(S2):S59-S66.

Expression of Plasmodium major facilitator superfamily protein in transporters - Δ Candida identifies a drug transporter.

Aim: To assess the functional relevance of a putative Major Facilitator Superfamily protein (PF3D7_0210300; 'PfMFSDT') as a drug transporter, using Candida glabrata for orthologous protein expression.Methods: Complementary Determining Sequence encoding PfMFSDT was integrated into the genome of genetically engineered C. glabrata strain MSY8 via homologous recombination, followed by assessing its functional relevance as a drug transporter.Results & conclusion: The modified C. glabrata strain exhibited plasma membrane localization of PfMFSDT and characteristics of an Major Facilitator Superfamily transporter, conferring resistance to antifungals, ketoconazole and itraconazole. The nanomolar inhibitory effects of the drugs on the intra-erythrocytic growth of Plasmodium falciparum highlight their antimalarial properties. This study proposes PfMFSDT as a drug transporter, expanding the repertoire of the currently known antimalarial 'resistome'.

[Box: see text].

Raman and resonance Raman spectroscopic study of red blood cells in patients diagnosed with malaria.

Raman spectroscopy was used to study erythrocytes collected from patients diagnosed with malaria at the University Hospital in Kraków and from healthy volunteers. A laser line with a wavelength of 442 nm was used to induce the Raman resonance of haem, while a laser with a wavelength of 785 nm was used for the normal Raman effect. The results were analysed using Principal Component Analysis. For the 442 nm laser line, analysis of the entire spectral range (3200 cm-1 to 300 cm-1) showed satisfactory separation of Raman spectra for healthy cells from infected cells, which was significantly improved in the 1500 cm-1-1200 cm-1 spectral range. For the 785 nm laser line, some separation was observed in each range studied, but the best results were achieved over the full spectral range. Plasmodium-derived nucleic acids and phosphodiester vibrations were observed at excitation lines of 442 nm and 785 nm, respectively.

RH5: rationally-designed malaria vaccine antigen improving efficacy.

Plasmodium falciparum reticulocyte-binding protein homolog 5 (RH5) is a unique asexual blood-stage malaria vaccine candidate because of its high conservation and essential biological function of binding to basigin on the erythrocyte surface. Recent studies by Barrett et al., Wang et al., and King et al., have brought RH5-based vaccine development a step forward based on a rational antigen design strategy.