-
Nature Communications Sep 2023Tryptophan Rich Antigens (TRAgs) are encoded by a multi-gene family found in all Plasmodium species, but are significantly expanded in P. vivax and closely related...
Tryptophan Rich Antigens (TRAgs) are encoded by a multi-gene family found in all Plasmodium species, but are significantly expanded in P. vivax and closely related parasites. We show that multiple P. vivax TRAgs are expressed on the merozoite surface and that one, PVP01_0000100 binds red blood cells with a strong preference for reticulocytes. Using X-ray crystallography, we solved the structure of the PVP01_0000100 C-terminal tryptophan rich domain, which defines the TRAg family, revealing a three-helical bundle that is conserved across Plasmodium and has structural homology with lipid-binding BAR domains involved in membrane remodelling. Biochemical assays confirm that the PVP01_0000100 C-terminal domain has lipid binding activity with preference for sulfatide, a glycosphingolipid present in the outer leaflet of plasma membranes. Deletion of the putative orthologue in P. knowlesi, PKNH_1300500, impacts invasion in reticulocytes, suggesting a role during this essential process. Together, this work defines an emerging molecular function for the Plasmodium TRAg family.
Topics: Humans; Plasmodium vivax; Tryptophan; Plasmodium; Antigens, Protozoan; Malaria, Vivax; Sulfoglycosphingolipids
PubMed: 37709739
DOI: 10.1038/s41467-023-40885-8 -
Tropical Medicine and Infectious Disease Jul 2023The initial and vital stage in the diagnosis of malaria involves extracting DNA. The efficiency of malaria testing is restricted by the multiple steps involved in...
The initial and vital stage in the diagnosis of malaria involves extracting DNA. The efficiency of malaria testing is restricted by the multiple steps involved in commercial DNA extraction kits. We attempted to improve an existing loop-mediated isothermal amplification (LAMP) for the detection of by using a simple DNA extraction approach, making it a feasible option for mass screening. We utilized a simple nucleic acid extraction method directly from whole blood for the detection of , taking only 5 min to complete. The extracted DNA was evaluated by two fluorescent-based LAMP and one colorimetric-based LAMP assay. The detection limit for both SYTO-LAMP and SYBR green-LAMP was 0.00001% and 0.0001% parasitemia, respectively. Meanwhile, neutral red-LAMP had a detection limit of 0.01% parasitemia. Combining this simple and inexpensive DNA extraction method, SYTO-LAMP could serve as an alternative molecular diagnosis for the detection of and other human spp.
PubMed: 37624327
DOI: 10.3390/tropicalmed8080389 -
International Journal of Molecular... May 2024is the only that causes zoonotic disease among the that cause infection in humans. It is fatal due to its short asexual growth cycle within 24 h. Lactate...
is the only that causes zoonotic disease among the that cause infection in humans. It is fatal due to its short asexual growth cycle within 24 h. Lactate dehydrogenase (LDH), an enzyme that catalyzes the final step of glycolysis, is a biomarker for diagnosing infection by spp. parasite. Therefore, this study aimed to efficiently produce the soluble form of LDH (PkLDH) using a bacterial expression system for studying malaria caused by . Recombinant pET-21a(+)- plasmid was constructed by inserting the gene into a pET-21a(+) expression vector. Subsequently, the recombinant plasmid was inserted into the protein-expressing Rosetta(DE3) strain, and the optimal conditions for overexpression of the PkLDH protein were established using this strain. We obtained a yield of 52.0 mg/L PkLDH from the Rosetta(DE3) strain and confirmed an activity of 483.9 U/mg through experiments. This methodology for high-efficiency PkLDH production can be utilized for the development of diagnostic methods and drug candidates for distinguishing malaria caused by .
Topics: Plasmodium knowlesi; L-Lactate Dehydrogenase; Cloning, Molecular; Malaria; Recombinant Proteins; Escherichia coli; Animals; Humans; Gene Expression; Protozoan Proteins
PubMed: 38891805
DOI: 10.3390/ijms25115615 -
PLoS Neglected Tropical Diseases Jan 2024Plasmodium knowlesi is a zoonotic parasite that causes malaria in humans. The pathogen has a natural host reservoir in certain macaque species and is transmitted to... (Review)
Review
BACKGROUND
Plasmodium knowlesi is a zoonotic parasite that causes malaria in humans. The pathogen has a natural host reservoir in certain macaque species and is transmitted to humans via mosquitoes of the Anopheles Leucosphyrus Group. The risk of human P. knowlesi infection varies across Southeast Asia and is dependent upon environmental factors. Understanding this geographic variation in risk is important both for enabling appropriate diagnosis and treatment of the disease and for improving the planning and evaluation of malaria elimination. However, the data available on P. knowlesi occurrence are biased towards regions with greater surveillance and sampling effort. Predicting the spatial variation in risk of P. knowlesi malaria requires methods that can both incorporate environmental risk factors and account for spatial bias in detection.
METHODS & RESULTS
We extend and apply an environmental niche modelling framework as implemented by a previous mapping study of P. knowlesi transmission risk which included data up to 2015. We reviewed the literature from October 2015 through to March 2020 and identified 264 new records of P. knowlesi, with a total of 524 occurrences included in the current study following consolidation with the 2015 study. The modelling framework used in the 2015 study was extended, with changes including the addition of new covariates to capture the effect of deforestation and urbanisation on P. knowlesi transmission.
DISCUSSION
Our map of P. knowlesi relative transmission suitability estimates that the risk posed by the pathogen is highest in Malaysia and Indonesia, with localised areas of high risk also predicted in the Greater Mekong Subregion, The Philippines and Northeast India. These results highlight areas of priority for P. knowlesi surveillance and prospective sampling to address the challenge the disease poses to malaria elimination planning.
Topics: Animals; Humans; Plasmodium knowlesi; Prospective Studies; Asia, Southeastern; Malaria; Malaysia; Macaca; Anopheles
PubMed: 38252650
DOI: 10.1371/journal.pntd.0011570 -
Scientific Reports Jul 2023Molecular detection methods have revealed higher sensitivity and specificity than conventional microscopy or rapid diagnostic tests for malaria diagnosis. In this study,...
Molecular detection methods have revealed higher sensitivity and specificity than conventional microscopy or rapid diagnostic tests for malaria diagnosis. In this study, we implemented, evaluated and validated according to the ISO 15,189 requirements, a multiplex real-time PCR assay to detect and identify the five human malaria parasites. DNA samples were extracted from whole blood or dried blood spots drawn from patients. Based on the External Quality Assessment (whole blood), this method shows 100% sensitivity and specificity. This PCR detected P. vivax up to 0.25 p/µl, P. falciparum and P. knowlesi up to 0.5 p/µl, P. ovale up to 1 p/µl and P. malariae up to 5 p/µl of blood. From blood spots (extraction from four punches), it detected P. vivax at 5 p/µl, P. falciparum, P. ovale and P. knowlesi at 20 p/µl and P. malariae at 125 p/µl. In conclusion, this quantitative PCR shows excellent performance, is easy to use and DNA saver. It is especially useful to actively screen large population groups and identify the five human malaria parasites in a context of low malaria transmission.
Topics: Humans; Real-Time Polymerase Chain Reaction; Plasmodium; Malaria; Malaria, Vivax; Malaria, Falciparum; Sensitivity and Specificity; Plasmodium vivax; Plasmodium falciparum
PubMed: 37452123
DOI: 10.1038/s41598-023-38621-9 -
Tropical Biomedicine Sep 2023In Malaysia presently, the main cause of human malaria is by the zoonotic monkey parasite Plasmodium knowlesi. A previous study has suggested that the P. knowlesi...
In Malaysia presently, the main cause of human malaria is by the zoonotic monkey parasite Plasmodium knowlesi. A previous study has suggested that the P. knowlesi merozoite surface protein 1 (Pkmsp-1) block IV to be a suitable multiplicity of infection (MOI) genotyping marker for knowlesimalaria. This study therefore aimed to investigate the usefulness of Pkmsp-1 block IV in assessing the MOI of P. knowlesi in clinical isolates from Malaysia. Two allele-specific PCR primer pairs targeting the two allelic families of block IV (T1 and T2) were designed, and used to genotype P. knowlesi in 200 blood samples (100 from Peninsular Malaysia and 100 from Malaysian Borneo). Results showed that the mean MOI in Malaysian Borneo was slightly higher as compared to Peninsular Malaysia (1.58 and 1.40, respectively). Almost half of the total blood samples from Malaysian Borneo (52%) had polyclonal infections (i.e., more than one allele of any family type) as compared to Peninsular Malaysia (33%) samples. The T1 allelic family was more prevalent in Peninsular Malaysia (n=75) than in Malaysian Borneo (n=60). The T2 allelic family, however, was more prevalent in the Malaysian Borneo (n=87 vs n=53 respectively). This study shows that the single locus Pkmsp-1 block IV can serve as a simple alternative genetic marker for estimating knowlesi malaria MOI in a population. Future MOI studies should focus on macaque populations as macaques are the natural host of P. knowlesi.
Topics: Humans; Genetic Variation; Plasmodium knowlesi; Malaysia; Genotype; Malaria
PubMed: 37897161
DOI: 10.47665/tb.40.3.004 -
The American Journal of Tropical... Nov 2023Malaysia has maintained zero cases of indigenous human malaria since 2018. However, zoonotic malaria is still prevalent in underdeveloped areas and hard-to-reach...
Malaysia has maintained zero cases of indigenous human malaria since 2018. However, zoonotic malaria is still prevalent in underdeveloped areas and hard-to-reach populations. This study aimed to determine the prevalence of malaria among remote indigenous communities in Peninsular Malaysia. A cross-sectional survey was conducted in six settlements in Kelantan state, from June to October 2019. Blood samples were tested for malaria using microscopy and nested polymerase chain reaction (nPCR) targeting the Plasmodium cytochrome c oxidase subunit III (cox3) gene. Of the 1,954 individuals who appeared healthy, no malaria parasites were found using microscopy. However, nPCR revealed seven cases of Plasmodium knowlesi mono-infection (0.4%), and six out of seven infections were in the group of 19 to 40 years old (P = 0.026). No human malaria species were detected by nPCR. Analysis of the DNA sequences also showed high similarity that reflects common ancestry to other P. knowlesi isolates. These findings indicate low submicroscopic P. knowlesi infections among indigenous communities in Malaysia, requiring PCR-based surveillance to support malaria control activities in the country.
Topics: Humans; Young Adult; Adult; Plasmodium knowlesi; Malaysia; Cross-Sectional Studies; Malaria; Polymerase Chain Reaction
PubMed: 37748768
DOI: 10.4269/ajtmh.23-0184 -
Malaria Journal Oct 2023
PubMed: 37880730
DOI: 10.1186/s12936-023-04733-w -
ChemMedChem Nov 2023Malaria continues to be a major public health challenge worldwide and, as part of the global effort toward malaria eradication, plasmodium carbonic anhydrases (CAs) have...
Malaria continues to be a major public health challenge worldwide and, as part of the global effort toward malaria eradication, plasmodium carbonic anhydrases (CAs) have recently been proposed as potential targets for malaria treatment. In this study, a series of eight hybrid compounds combining the Artesunate core with a sulfonamide moiety were synthesized and evaluated for their inhibition potency against the widely expressed human (h) CAs I, II and the isoform from P. falciparum (PfCA). All derivatives demonstrated high inhibition potency against PfCA, achieving a K value in the sub-nanomolar range (0.35 nM). Two Compounds showed a selectivity index of 4.1 and 3.1, respectively, against this protozoan isoform compared to hCA II. Three Derivatives showed no cytotoxic effects on human gingival fibroblasts at 50 μM with a high killing rate against both P. falciparum and P. knowlesi strains with IC in the sub-nanomolar range, providing a wide therapeutic window. Our findings suggest that these compounds may serve as promising leads for developing new antimalarial drugs and warrant further investigation, including activity against antimalarial-resistant strains, mode of action studies, and in vivo efficacy assessment in preclinical mouse models of malaria.
Topics: Animals; Humans; Mice; Antimalarials; Artesunate; Plasmodium falciparum; Carbonic Anhydrases; Carbonic Anhydrase Inhibitors; Malaria, Falciparum; Malaria; Protein Isoforms
PubMed: 37697903
DOI: 10.1002/cmdc.202300267 -
Parasites & Vectors Aug 2023Indonesia is home to many species of non-human primates (NHPs). Deforestation, which is still ongoing in Indonesia, has substantially reduced the habitat of NHPs in the...
BACKGROUND
Indonesia is home to many species of non-human primates (NHPs). Deforestation, which is still ongoing in Indonesia, has substantially reduced the habitat of NHPs in the republic. This has led to an intensification of interactions between NHPs and humans, which opens up the possibility of pathogen spillover. The aim of the present study was to determine the prevalence of malarial parasite infections in NHPs in five provinces of Indonesia in 2022. Species of the genus Anopheles that can potentially transmit malarial pathogens to humans were also investigated.
METHODS
An epidemiological survey was conducted by capturing NHPs in traps installed in several localities in the five provinces, including in the surroundings of a wildlife sanctuary. Blood samples were drawn aseptically after the NHPs had been anesthetized; the animals were released after examination. Blood smears were prepared on glass slides, and dried blood spot tests on filter paper. Infections with Plasmodium spp. were determined morphologically from the blood smears, which were stained with Giemsa solution, and molecularly through polymerase chain reaction and DNA sequencing using rplU oligonucleotides. The NHPs were identified to species level by using the mitochondrial cytochrome c oxidase subunit I gene and the internal transcribed spacer 2 gene as barcoding DNA markers. Mosquito surveillance included the collection of larvae from breeding sites and that of adults through the human landing catch (HLC) method together with light traps.
RESULTS
Analysis of the DNA extracted from the dried blood spot tests of the 110 captured NHPs revealed that 50% were positive for Plasmodium, namely Plasmodium cynomolgi, Plasmodium coatneyi, Plasmodium inui, Plasmodium knowlesi and Plasmodium sp. Prevalence determined by microscopic examination of the blood smears was 42%. Species of the primate genus Macaca and family Hylobatidae were identified by molecular analysis. The most common mosquito breeding sites were ditches, puddles and natural ponds. Some of the Anopheles letifer captured through HLC carried sporozoites of malaria parasites that can cause the disease in primates.
CONCLUSIONS
The prevalence of malaria in the NHPs was high. Anopheles letifer, a potential vector of zoonotic malaria, was identified following its collection in Central Kalimantan by the HLC method. In sum, the potential for the transmission of zoonotic malaria in several regions of Indonesia is immense.
Topics: Animals; Humans; Indonesia; Mosquito Vectors; Malaria; Plasmodium knowlesi; Primates; Macaca; Anopheles
PubMed: 37550692
DOI: 10.1186/s13071-023-05880-4